PHT313.

PHT313

E. coli Klebsiella Citrobacter Enterobacter Salmonella Shigella Proteus Vibrio Yersinia Bacteroides Pseudomonas Brucella Bordetella Haemophilus Niesseria Spirochetes Rickettsia Chlamydia Mycoplasma

Introduction Host-Parasite Relationship

Introduction Microbiology
The science deals with the study of microorganisms (M.O) Microorganism Living organism that can not see by naked eye Examples: bacteria, fungi, viruses, protozoa Medical microbiology The science deals with the study of M.O. causing infection/disease

ECOLOGICAL RELATIONSHIPS
SYMBIOSIS: neutral, antagonistic or synergistic relationship between two dissimilar organisms (SYMBIOTES, SYMBIONTS) living in close association with each other MUTUALISM (+/+): mutually beneficial relationship between two species COMMENSALISM (+/0): relationship between two species in which one is benefited and the other is not affected, neither negatively nor positively PARASITISM (+/-): relationship between two species in which one benefits (parasite) from the other (host); usually involves detriment to the host

Microorganism Parasite Strict or obligate parasite
Living organism lives in another organism Receives shelter & nourishment from another organism Strict or obligate parasite Organism that cannot live without a host No free-living existence. e.g.: Rickettsiae, Spirochetes, viruses Facultative parasite Has both a free-living & a parasitic existence, e.g. Clostridium, Pseudomonas

Normal flora (Commensals)
Present on the skin and mucous membrane May be non-pathogen or opportunistic pathogen 2 types of Normal flora: Resident flora M.O. present in the region at a given age S. mutans in oral cavity, E. coli in intestine Transient flora Temporarily M.O. is present only for hours to weeks Streptococcus pyogenes in throat

Important normal flora
Less common Common Site None, Sterile Blood, CSF, Internal organs S. aureus, Diphtheroids, streptococci, anaerobes, yeasts Staphylococcus epidermidis Cutaneous surfaces including urethra & outer ears S. epidermidis, diphtheroides, assorted streptococci S. aureus Nose assorted streptococci, non-pathogenic Neisseria, nontypeale H. influenzae Viridans streptococci Oropharynx None Stomach Lactobacillus, Fusobacterium assorted gram-negative rods, E. faecalis Bacterioids E. coli Colon Assorted streptococci, gram-negative rods, diphteroids, yeasts Lactobacillus Vagina

Opportunistic pathogen
Saprophyte Heterotroph Organism that lives in/on dead organic matter Non-pathogen Organism that does not cause disease May be a member of normal flora Pathogen Primary pathogen which can cause disease Opportunistic pathogen Organism (commensal or saprophyte) that can cause disease only in immunocompromised individuals.

Factors affecting diseases
Microbial factors Pathogenicity and Virulence Host resistance factors Natural and acquired immunity (PHT324)

Microbial Factors Pathogenicity Virulence Virulence factors
Ability of a pathogen to produce a disease by overcoming the defenses of the host Virulence Degree/extent/measure of pathogenicity Highly virulent organism  Sever disease Moderate virulent organism  Mild disease Avirulent organism  unable to produce disease Virulence factors Properties of M.O. responsible for its pathogenicity Pathogenesis The mechanism that causes the disease Development of disease

Virulence Factors Bacterial structures Enzymes Toxins

Virulence Factors

Virulence Factors Bacterial structures Flagella Capsules Pili

Virulence Factors Enzymes Coagulase Kinases Hyaluronidase Lecithinase
Necrotizing enzymes

Virulence Factors Toxins Endotoxin Exotoxins Neurotoxins Enterotoxins
Toxins produces by the bacteria

Steps in the Pathogenesis of Infectious Diseases
Contact Attachment Entry Multiplication Invasion or spread Evasion of host defenses Damage to host tissues

Portal of Entry Sites of entry of M.O. in human hosts Digestive tract
Respiratory tract Skin Urogenital tract Conjunctiva

Mechanism of Pathogenicity (Infectivity and Toxicity)
Invasiveness Ability to invade tissues Colonization (Adhesins) Invasion (invasins) Evasion host defense mechanisms Damage and Toxigenesis Ability to produce damage e.g. Exotoxins and endotoxins

Mechanism of Pathogenicity Invasiveness
Colonization First step of microbial infection Important unless organism is traumatically implanted Establishment of the pathogen at portal of entry Organisms infect the regions of portal entry have Developed tissue adherence mechanisms Some ability to overcome host defenses at the surface

Mechanism of Pathogenicity
Colonization includes Adherence to mucosal cell surfaces involves Pilli/fimbriae- primary mechanism in gram - Teichoic acids- primary mechanism in gram + Adhesins IgA protease- Cleaved FC portion may coat bacteria and bind them to cellular Fc receptors Adherence to inert materials, biofilm: S. epidermidis, Streptococcus mutans

Invasion Aided by production of Invasins Bacterial extracellular substances Act locally To damage host cells and/or To facilitate growth and spread of pathogen Spreading Factors Hyaluronidase, Neuraminidase, Streptokinase Enzymes that Cause Hemolysis and/or Leucolysis Lecithinases, hemolysins, leukocidins, Streptolysin Staphylococcal coagulase

Extracellular Digestive Enzymes Proteases, lipases, glycohydrolases, nucleases Not have a direct role in invasion or pathogenesis, but may aid in invasion Toxins With Short-Range Effects Related to Invasion Thought to promote bacterial invasion Toxins which have adenylate cyclase activity  cAMP and disruption of cell permeability e.g. Anthrax toxin (EF) and Bordetella pertussis toxin

Ability to bypass or overcome of host defense Avoiding immediate destruction by host defense Antiphagocytic surface components Capsule /slime layers S. pyogenes M protein N. gonorrhoeae pilli S. aureus A protein IgA protease Destruction of mucosal IgA: Neisseria, Heamophilus, S. pneumoniae

Ability to survive intracellularly Escape from the phagosome Essential for growth & virulence of some intracellular M.O. Rickettsiae produce a phospholipase enzyme that lyses the phagosome membrane within 30 seconds after ingestion Survival inside the phagolysosome Some extracellular pathogens resist killing in phagocytes B. anthracis, M. tuberculosis and S. aureus Inhibition of phagosome-lysosome fusion Salmonella, M. tuberculosis, Legionella, Chlamydiae

Replication cycle of Rickettsia
Phagosome is vesicle formed around particle absorbed by phagocytosis Some pathogens that enter cell inside phagosome either Reproduce inside of phagolysosome e.g. Coxiella spp or Escape phagosome fuses with lysosome e.g. Rickettsia spp. into cytoplasm before

Replication of C. burnetii

Damage and Toxigensis Using the Host's Nutrients Bacteria take Fe from host's iron transport proteins Bacteria secrete siderophores, which bind Fe Direct Damage Host cells can be destroyed when pathogens metabolize and multiply inside the host cell Toxins Exotoxin Endoxins

Toxin Toxin Substances that contribute to pathogenicity Toxigenicity
Ability to produce a toxin Toxemia Presence of toxin the host's blood Toxoid Inactivated toxin used in a vaccine Antitoxin Antibodies against a specific toxin

A. Exotoxins Exotoxins classified according to
A-B toxins (Type III toxins) Consist of 2 polypeptides (A and B subunits) A polypeptide is enzyme & B is a binding component. B binds to the cell, the complete toxin is taken into cell 2 subunits separate & A subunit enzymatically kills cell

A. Exotoxins Membrane disrupting toxins (Type II toxins)
Disturb host cell plasma membranes Leukocidins kill phagocytes Hemolysins destroy erythrocytes Superantigens (Type I toxins) Provoke intense immune responses TSST(Toxic shock syndrome toxin), Erythrogenic toxin

A. Exotoxins Exotoxins classified into Cytotoxins Neurotoxins
Diphtheria toxin (A-B toxin inhibits protein synthesis) Erythrogenic toxins (Superantigens that damage capillaries) Neurotoxins Botulinum toxin (A-B toxin) Tetanus toxin (A-B toxin) Enterotoxins Induce fluid and electrolyte loss from host cell Vibrio entertoxin - A-B toxin stimulates secretion of fluid and electrolytes results in diarrhea. Staphylococcal enterotoxin, a superantigen that produces the same results as Vibrio enterotoxin.

B. Endotoxins (Pyrogen)
Endotoxins are LPS, lipid A component of the outer membrane of gram-negative bacteria Bacterial cell death, antibiotics, and antibodies may cause the release of endotoxins Endotoxins cause fever by inducing the release of interleukin-1 and shock because of TNF-induced decrease in blood pressure TNF causes edema by damaging capillaries. LPS stimulates release of NO from macrophages, which causes vasodilation Endotoxins allow bacteria to cross the blood-brain barrier

INFECTION vs. DISEASE INFECTION
Colonization and/or invasion and multiplication of infectious agent in the host with or without the manifestation of disease DISEASE When infection has detectable clinical consequence

INFECTIOUS VS. NONINFECTIOUS DISEASES
Communicable/Transmissible diseases Transmitted directly or indirectly from one host to another Caused by infectious agents Patient may exhibit symptoms and signs A specific group of symptoms or signs that always accompanies a specific disease is called syndrome A contagious disease is one that is easily spread from one person to another. Non infectious Diseases Commonly called Noncommunicable diseases (NCD) Chronic diseases are not passed from person to person 4 main types of NCD are cardiovascular diseases, cancers, chronic respiratory diseases and diabetes.

INFECTIOUS DISEASES Community acquired infection:
Infection contracted outside of a health care setting or present on admission CA respiratory infection commonly involve H. influenzae, S. pneumoniae, usually more antibiotic sensitive Nosocomial infection: acquired in hospital not present in the host prior to admission generally occurring within 72 hours of admission e.g. Pseudomonas, Hepatitis B Zoonotic infection: transmitted under natural conditions from vertebrate into man e.g. brucellosis, plague,, anthrax and T.B. Occupational infection: associated with certain profession, usually zoonotic e.g. Brucellosis, anthrax

EPIDEMIOLOGY Epidemiology: study of all factors connected to incidence and spread of a disease Sporadic : occurs occasionally (typhoid fever in U.S.) Epidemic: Disease occurred suddenly in a given area in short period (influenza) Endemic:Disease present or usually prevalent in a population or geographic area at all times (Cholera in India) Pandemic: world wide epidemic, occuring widely throughout a region, country, continent, or globally influenza occasionally, AIDS might be considered pandemic

The Severity or Duration of a Disease
Acute Develops rapidly but lasts a short time (influenza) Chronic Develops more slowly, reactions to the disease are less severe, likely to be continual or recurrent for long periods (TB, hepatitis B) Subacute Intermediate between acute and chronic Latent The causative agent remains inactive for a time but then becomes active to produce symptoms

Characterized by appearance of first mild signs & symptoms
Incubation period Time interval between the initial infection and the first appearance of signs and symptoms Prodromal period Characterized by appearance of first mild signs & symptoms Period of illness Disease is at its height and all disease signs and symptoms are apparent. Death may occur during the period of illness. Period of decline The signs and symptoms subside. Period of convalescence Body returns to its pre-diseased state & health is restored.

Infection involves the following :
Source or reservoir of infectious agent Transmission of the infectious agent from the source to the host Susceptible host Portal of entry of the agent in the host, its localization, multiplication and finally host-parasite interactions which result in either Destruction of the agent or Infectious disease

I. Source (Reservoir) Human Source Exogenous source: Patient (case) or carrier Endogenous source: The individual himself Case: Either clinical or sub-clinical form Carrier. One who harbors organisms in their body without manifest symptoms, thus acting as a distributor of infection. Carrier: the most dangerous reservoir for infections Vector: carrier, especially the animal that transfers an infectious agent from one host to another, usually an ARTHROPOD

Self-infection Infection that occurs from patient's own flora Infection of wound of a patient by staphylococci carried by individual in his nose Coliforms and anaerobes released from his bowel during surgery Cross-infection is the infection derived from other patients or healthy carriers by direct spread Cross-infection is common in hospitals

Classification of carriers A) According to type
Healthy carrier: A person who receive the infection from another carrier (very dangerous). Incubatory carrier: A person who shed the M.O. during the incubation period (specially last days of the incubation period) e.g. mumps, measles and diphtheria Convalescent carrier: a person who still shed the M.O. during the convalescent period e.g. typhoid, dysentery and whooping cough.

Classification of carriers B) According to duration
Temporary carrier: Person who shed the m.o. for short period Chronic carrier: Person who shed the m.o. for indefinite periods (very dangerous). C) According to portal of exit Respiratory carrier: e.g. meningococcus Fecal (intestinal) carrier: e.g. typhoid, dysentery, cholera Blood carrier: e.g. hepatitis B and AIDs Urinary or sexual Carrier e.g. gonococcus, AIDs

Source (Reservoir) Foods and Drinks Food: Any contaminated food
Water contaminated with bacteria of typhoid fever, cholera, diarrhoea and dysentery, Milk contaminated with salmonella, M. bovis

Source (Reservoir) : Animals: Zoonoses
Cow: Bovine tuberculosis, brucellosis, Salmonella food poisoning, anthrax Poultry. Salmonella food poisoning by eggs and meat, Dog. Rabies, Weil's disease Horse. Tetanus Goat. Anthrax and brucellosis (by B. melitensis ), Sheep. Anthrax and tetanus, Rat. Plaque, Weil's disease Parrot and pigeons. Psittacosis Soil: Tetanus, gas gangrene.

II. Modes of Transmission
Water-borne e.g. cholera and other diarrhoeal diseases, enteric fever from contaminated water. Food-borne (contaminated food) Enteric fever, salmonella food poisoning (eggs) Milk and milk products- enteric fever, bovine TB. Air-borne e.g. Droplets. During coughing, sneezing & talking: Diphtheria, TB, measles, chickenpox, influenza Dust-borne. e.g. tuberculosis, Soil-Tetanus, gas gangrene.

Modes of Transmission Contaminated fomites like beddings, clothing, utensils: Trachoma. Direct contact- STD like gonorrhea, syphilis, AIDS. Transplacental: Rubella, congenital syphilis. Vector-borne.(Insects) Actively, e.g. Spotted fever, dengue, plague, yellow fever Passively, e.g. diarrhoeal diseases, enteric fever, salmonella food poisoning.

III. Host Host is the harboring organism of a parasite
Definitive host. When the adult stage of animal parasites lives and sexual reproduction takes place Man is the definitive host for all animal parasites Intermediate host (IH). Asexual reproduction takes place, or the larval stages of animal parasites develops When development of larval stage takes place in 2 different hosts they are called 'first' & 'second' IH Man is IH for malaria & hydatid tapeworm Man is both definitive and IH for T. solium Paratenic host. A carrier or transport host

Prevention and control of diseases
Mean the measures to be followed prior to the onset of disease General measures 1)Environment sanitation: Housing, water supply, waste and sewage disposal 2) Health education 3) Health promotion: Nutrition and persons hygiene

Specific measures Control of reservoir and susceptible hosts
Control of reservoir (Cases or carriers) Clinical & lab diagnosis followed by treatment Isolation to protect community by preventing the spread of infection. Notification of health authorities for all communicable diseases except common cold, rubella, influenza, mumps, dysentery, sore throat and WHO for cholera, plague, typhus and yellow fever. Quarantine of contacts for a period longer than that of I.P. e.g. cholera, plague, typhus and yellow fever. Immunization: Active and passive.

A. Gram Positive Cocci 1. Staphylocoocus
> 30 species 3 species of medical importance Staphylococcus aureus Present in the anterior nares (50-75%) Pathogenic Staphylococcus epidermidis Normal flora Opportunistic pathogen Staphylococcus saprophyticus

General characteristics of Staphylococci
Morphology Gram positive cocci Grape-like (Cluster, punches of grapes) Non Motile, Non Spore Forming Capsule is usually present in vivo General physiological characteristics Non Fastidious Halotolerant (Grows in media containing 10% NaCl) Facultative anaerobes and Fermentative (O+/F+) Catalase positive

Virulence Factors of S. aureus 1. Structural components
Protein A Major protein in cell wall and important virulence factor Disable our immune defenses / adhesin Binds to Fc region of IgG Preventing the binding to the complement Disrupts opsonization and phagocytosis Polysaccharide Capsule: Antiphagocytosis Facilitates adherence to foreign bodies

1. Structural components
Peptidoglycan Activates complement by alternative pathway N.B. Complement is part of immune response called innate immune system Means ability of antibodies and phagocytes to clear pathogens Teichoic acid Activates complement and involved in adherence

2. Toxins Disable our immune defenses Hemolysins
Four types (alph, beta, gamma and delta) Destroy RBCs, neutrophils, macrophage and platelets Leukocidins Kills WBC’s (PMNL) and prevents phagocytosis Releases and ruptures lysosomes Lysosomes - contain powerful hydrolytic enzymes which cause more tissue damage Staphyloxanthin (Carotenoid pigment) Responsible for its characteristic golden yellow color Has antioxidant action- help M.O evade death by reactive O2

2. Toxins Exotoxins attack weaponry TSST-1 (Superantigen)
TSST-1 causes TSS associated with tampon use. TSST-1 stimulates the release of IL-1 and IL-2 Exfoliatin ET (A and B) Produced by staphylococci of phage group II ET causes SSSS in young children Enterotoxins Six types (A-F), heat stable and resistant to GIT enzymes Stimulate the release of large amounts of IL-1 and IL-2 Increasing intestinal peristalsis, fluid loss, nausea & vomiting

S. epidermidis & S. saprophyticus
3. Enzymes Coagulase This enzyme can lead to fibrin formation around the bacteria, protecting it from phagocytosis Coagulase converts fibrinogen into fibrin Free coagulase Bound Coagulase (Clumping Factor) Coagulase test Coagulase Positive S. aureus Coagulase-Negative S. epidermidis & S. saprophyticus

S. epidermidis & S. saprophyticus
3. Enzymes Deoxyribonuclease (DNAase) DNase hydrolyze DNA into oligonucleotides S. aureus produces DNase while S. epidermidis and most staphylococci have not DNase DNase test Positive Staphylococus aureus Negative S. epidermidis & S. saprophyticus

C. Catalase Oxidative killing of the organism by phagocyte achieved by reduction of O2 to superoxide and peroxide H2O2 is toxic to the microorganism Catalase catalyzes removal of H2O2 H2O2 Catalase H2O + O2 (gas, ↑) Staphylococci Catalase test Positive Microcococcaceae Staphylococci Negative Streptococcaceae Streptococci

Other enzymes proteins to tunnel through tissue
Hyaluronidase Spreading factor Breakdown hyaluronic acid in connective tissue Staphylokinase (Fibrinolysin) Lyses formed fibrin clot (Like streptokinase) Conversion of plasminogen into plasmin Helps to spread bacteria - Bacteremia

Other enzymes proteins to tunnel through tissue
Lipase Breaks down fats & oils which often accumulate on the surface of our body This facilitates colonization of sebaceous glands Protease Destroy tissue proteins Beta-lactamase

Clinical finding/diseases
Due to direct effect of organism Inflammatory/Suppurative/ القيحي Local lesions of skin (The most common) Deep abscesses (metastatic) Systemic infections Toxin mediated Food poisoning Toxic shock syndrome (TSS) Scalded skin syndrome (SSSS)

1. Skin lesions/superficial infections
Pustules, Boils and Abscesses Furuncles (infection of hair follicle) Carbuncles (infection of several hair follicles) Impetigo: Skin lesion with blisters بثور that break and become covered with crusting تقشر)) exudates Wound infections (Hospital acquired) Progressive appearance of swelling and pain in a surgical wound after about 2 days from the surgery Conjunctivitis (Pink eye) Inflammation of the conjunctiva (the outermost layer of the eye and the inner surface of the eyelids)

2. Deep abscesses Metastic in any organ after Bacteremia
Can be single or multiple Breast abscess can occur in 1-3% of nursing mothers in puerperium (النفاس) Can produce mild to severe disease Other sites - kidney, brain from septic foci in blood

3. Systemic Infections Osteomyelitis, septic arthritis
Either hematogenous or traumatic Acute bacterial endocarditis Bacteremia/Septicaemia (IV drug users) S. aureus is the most common cause of G + bacteremia Commonly in hospital strains UTI Staphylococcal pneumonia (Post viral) Meningitis

B. Toxin mediated diseases
Staphylococcal food poisoning Due to ingestion of heat stable enterotoxins They performed in food (protein)contaminated from hands or nose of handlers & kept non-refrigerated Very short incubation period (1-8 hours) Severe vomiting+++, diarrhea + and No fever Resolves on its own within about 24 hours

2. Toxic shock syndrome (TSS)
Mediated via TSST High fever, diarrhea, shock & erythematous skin rash which goes to desquamate May be hypotension with cardiac and renal failure 10% mortality rate Described in young women using tampons (retained highly absorbent tampons) during menstruation

3. Staphylococcal Scalded skin syndrome (SSSS)
Disease of young children Mediated through minor Staphylococcal infection by ‘epidermolytic/exofoliative toxins’ producing strains Mild erythema and blistering of skin followed by shedding of sheets of epidermis Children are eventually recover

Phage typing S. aureus has a differential ability to spread and cause outbreak of infections Characterization of these strains is important to control the spread of infection in hospitals and the community Typing of these strains is important in epidemiology to Differentiate and evaluate the importance of these strains in spreading the infection.

Treatment Drain Pus if any + Antibiotics
95% of S. aureus resistant to penicillin G (β-lactamase production) If sensitive, Penicillin is the drug of choice If resistant, use β-lactamase stable penicillins Cloxacillin, oxacillin, methicillin If resistant to β-lactamase stable penicillins due to alteration of penicillin binding proteins Are called Methcillin Resistant S. aureus (MRSA) Prevalence of MRSA about 20-30% Vancomycin is the drug of choice for MRSA VISA (vancomycin intermediate resistant S. aureus) VRSA (vancomycin resistant S. aureus) Fucidic Acid- penetrate well in bones

Staphylococcus epidermidis
Skin commensal True opportunistic pathogen Pathogen of nosocomial infections Infections are associated with implanted foreign bodies Intravascular devices (prosthetic heart valves, shunts) Septicemia and endocarditis Prosthetic joints, catheters, and large wounds Catheter infections along with catheter-induced UTIs Treatment Has variable ABS pattern Treatment should be aided with ABST Similar to S. aureus

What makes S. epidermidis virulant?
The organism produces slime layers (Glycocalyx) Glycocalyx forms a hydrophobic biofilm Biofilm is adhesive to hydrophobic polymers of prosthetics Causes resistance to phagocytosis and some antibiotics Creating diseases such as endocarditis Biofilms are sources of infection from which bacteria are flushed into the bloodstream and cause disease

Stapylococcus saprophyticus
GIT is the major reservoir of S. saprophyticus Young adult are more susceptible to UT colonization Cause of UTI 2nd most-frequent causative agent of acute UTI Virulence factors include adherence to urothelial cells; Surface-associated protein, lipoteichoic acid; a hemagglutinin that binds to fibronectin, a hemolysin; and production of extracellular slime Usually sensitive to wide range of antibiotics Resistant to Novobiocin

Laboratory Diagnosis I- Specimen:
Pus, Mid-stream urine, Sputum, Stool, Blood, CSF II- Gram Stain: Gram positive cocci, arranged in cluster, etc III- Culture: A. Blood agar (Non-Selective Media) S.aureus colonies are golden yellow & -hemolytic S. epidermidis are non-pigmented and non-hemolytic

B. Mannitol Salt Agar (MSA)
Selective and differential medium for staphylococci Selective agent: 7.5% NaCl Differential agent: Mannitol pH indicator: Phenol Red Turns yellow in acidic pH & turns red in alkaline pH S. aureus ferment mannitol (yellow colonies) S. epidermidis & S. saprophyticus do not ferment mannitol and appear red on MSA. IV Biochemical Test Coagulase, DNAse, Catalase tests

A. Gram Positive Cocci 2. Streptcocci
Morphology Gram positive cocci arranged in chains or pairs Capsulated Non motile, Non spore forming General physiological characteristics Fastidious Grow on blood agar, chocolate agar Facultative anaerobes, Fermentative Catalase negative

Classification of Streptococci
Hemolysis on Blood Agar α-hemolytic streptococci Partial hemolysis Green discoloration around the colonies e.g. S. pneumoniae, viridans streptococci -hemolytic streptococci Complete hemolysis Clear zone of hemolysis around the colonies e.g. S. pyogenes and S. agalactiae -hemolytic streptococci No lysis Enterococcus faecalis

Classification of Streptococci
Serology Classification (Lanciefield) Based on C- carbohydrate antigen of cell wall 20 groups from A-H & K-V One or more species per group Groupable streptococci A, B and D (more frequent) C, G and F (Less frequent) Non-groupable streptococci S. pneumoniae viridans streptococci

Pathogenesis and Virulence Factors
Structural components Hyaluronic acid capsule Antiphagocytic Non-immunogenic Capsule, M protein, F protein and lipoteichoic acid facilitate attachment (Adhesion) to various host cells F protein & lipotecichoic acid have receptor on fibronectin It is responsible for the adhesion to pharynx and skin

M protein M proteins, major virulence factors, found in fimbriae
Used to determine the type of GAS (>100 types) Responsible for colonization & resistance to phagocytosis It binds fibrinogen and blocks binding of complement Its the weakest point in the organism’s defense Why?? Because B cells generates antibodies binds to M protein, aiding destruction of organism by macrophage Certain M protein types are considered rheumatogenic

Enzymes Streptokinases (Fibrinolysin)
Activates plasminogen to form plasmin Lysis blood clot helping in spreading of infection Used therapeutically to dissolve clot Nicotinamid Adenine Dinucleotidase (NADase) Kills leucocytes

Enzymes Deoxynucleases (streptodornase) Four different DNases
DNases protect bacteria from being trapped in neutrophil extracellular traps (NETs) By digesting NET's web of DNA, to which are bound neutrophil serine proteases that can kill bacteria Hyaluronidase Destroy hyaluronic acid in connective tissue Speeding factor

Toxins Streptococcal Pyrogenic Toxins (SPT)
SPT is also called Erythrogenic toxin 3 types A, B, C Produced by certain strains lysogenized by bacteriophage carrying the gene for toxin Stimulate macrophages & helper T cells cytokines Causes erythematous rash of scarlet fever

Toxins Streptolysins (Hemolysins)
Streptolysin O lyse RBCs, antigenic, unstable to O2, inactive aerobically and active anaerobically Streptolysin S lyse RBCs, non-antigenic, Stable to O2, active aerobically and anaerobically Exotoxin B It has Protease activity Destroy tissues rapidly Produced in large amounts by strains of S. pyogenes that cause necrotizing fasciitis

Disease caused by S. pyogenes
Suppurative Non-Invasive Pharyngitis Skin infection  Impetigo Invasive Erysipeals Necrotizing fasciitis Scarlet fever Puerperal sepsis Non Suppurative (Post-streptococcal ) Rheumatic fever Glomerulonephritits

Pharyngitis and tonsillitis
Represents 90% of cases of bacterial pharyngitis Streptococcal pharyngitis is acquired by droplet Red swollen tonsils and pharynx, a purulent exudates & swollen lymph nodes Fever & sore throat with white spots on tonsil membrane Extend to otitis media, sinusitis, mastoiditis & meningitis 1-3% develop rheumatic fever weeks after infection Diagnosis: Throat swab is taken and inoculated on a plate of blood agar including bacitracin disk S. pyogenes is beta-hemolytic & sensitive to bacitracin

Skin Infections: Impetigo
Involves infection of epidermal layers of skin Young children are the most susceptible Development of superficial blisters (بثور) Blisters break readily and spread by continuity Infected area is covered with honey colored crusts Diagnosis: Swab is taken from the lesion and inoculated on blood agar as before

Skin Infections: Erysipelas
Infection of the dermis Type of cellulitis generally caused GAS(gpA Strept) Manifested by creeping inflammation with vesicular sharply demarcated margin and browny edema Fever and shaking chills are common Most cases involve the legs, followed by the face. Way of infection: contamination of wound Diagnosis: The vesicular content is inoculated on blood agar and examined as before

Necrotizing Fasciitis/Flesh Eating
Rare infections and infect the fascia Certain strains have M proteins that block phagocytosis This allow bacteria to move rapidly through tissues S. pyogenes enters through a break in skin caused by trauma and then follow a path along fascia May proceed rapidly to underlying muscle Destructive effect due to exotoxin B (Protease) Swelling, heat and redness within a day Day latter, changes from red to purple to blue & large blister Later the skin dies & muscle may become infected

Scarlet Fever Streptococcal Pyrogenic Toxins mediated disease
Way of infection: Droplet infection Clinical finding Fever, sore throat, bright red tongue with a "strawberry" appearance and erythematous rash Rash is caused by erythrogenic (pyrogenic) toxins Rash begins on trunk & neck then spreads to extremities

Diagnosis of Scarlet Fever
Schultz-Charlton Reaction In vivo neutralization test I.D. injection of anti-erythrogenic toxin in erythematous area Positive: Disappearance of the rash within 6-12 hours Susceptibility to scarlet fever This is done by the Dick Test 0.1 ml erythrogenic toxin injected ID in one forearm (test) The same dose of heated toxin in the other forearm (control) Dick negative Dick positive No Rash Rash Test Control

Puerperal Sepsis Endogenous-Her throat and flora in the vagina
Clinical Picture Fever following labor or septic abortion accompanied with foul smelling uterine discharge The disease is always accompanied by bacteremia Way of infection: Endogenous-Her throat and flora in the vagina Exogenous-Droplet coming from medical staph, instruments, gloves Diagnosis Uterine swab and Blood culture

Post-streptococcal diseases
Rheumatic fever and Acute glomerularnephritis (AGN) Life threatening inflammatory diseases These diseases are not due to the direct effect of bacteria but due to hypersensitivity response About 2-4 weeks after acute GAS infection Nephritis is commonly preceded by infection of the skin Rheumatic fever is commonly preceded by pharyngitis

A. Rheumatic fever Early termination of throat infections with penicillin decreases the incidence of rheumatic fever Affecting primarily the heart valves and muscle and joints Certain M proteins are considered rheumatogenic Cross-reacts with heart tissue antigens leading to autoimmunity Fever, migratory polyarithritis, carditis Carditits damages Myo- and endo-cardial especially mitral and aortic valves Marked reactivated by recurrent infection Protection from recurrent by prophylaxis penicillin administration

Diagnosis of Rheumatic fever
Antistreptolysis titer (ASO) Principal Antibodies against streptolysin O neutralize streptolysin O Inhibiting its hemolytic activity Procedure Serial dilutions of patient’s serum + standard amount of streptolysin O and Incubate at 370C for 30 min Rabbit RBCs are added to each tube, reincubated for 30 min The titer is the last tube showing no hemolysis It’s expressed as reciprocal of that dilution & given in Tood units In positive cases the titer is usually above 200 Tood C-Reactive protein & Erythrocyte Sedimentation Rate

B. Acute glomerulonephritis (AGN)
Disease is initiated by antigen-antibody complexes on glomerular basement membrane M protein type 49 causes AGN most frequently Hypertension, edema in the face and ankles and smoky urine (Blood and protein in the urine) Few patients die & most patients recover completely Some develops chronic glomerulonephritis Re-infection rarely leads to recurrence of AGN

Treatment Prevention S. pyogenes do not produce β-lactamases
Drug of choice is Penicillin G ,000U/kg/day. In severe cases, such as with necrotizing fasciitis, surgical removal of the infected tissue + Clindamycin to block metabolism of toxin production Prevention No vaccines are currently available Rheumatic fever can be prevented by prompt treatment of pharyngitis Long acting Penicillin is used to prevent recurrence of disease in patient with Rheumatic fever

Group B Streptococcus (GBS)
GBS includes S. agalactiae General characteristics of S. agalactiae is similar to GAS Cause mastitis in cattle Cause pneumonia, septicemia & meningitis in neonates Colonize UT & GIT of adults and genital tract in women 25% of women carry S. agalactiae in their vagina Baby can acquire these bacteria during delivery Transmission rate from mothers to neonates is 50% 1-2% of colonized neonates develop invasive disease Pregnant women should be screened and treated

Streptococcus agalactiae: Invasive Infections
Cause invasive diseases in newborns Early-onset infection Occurs in neonates (< 7 days old neonates) Vertical transmission from the mother Manifests in the form of pneumonia or meningitis with bacteremia (neonatal sepsis) Associated with a high mortality rate S. agalactiae neonatal meningitis doesn't present with hallmark sign of adult meningitis, a stiff neck

Late-onset infection Occurs between 1 week and 3 months after birth Usually occurs in the meningitis form Mortality rate is not as high as early-onset S. agalactiae, E. coli and L. monocytogenes are the most common pathogens causing neonatal meningitis (<3 months) In adults Occurs in immunosuppressed patients Wound & skin infections & endocarditis

Laboratory Diagnosis: Group B β-Hemolytic Streptococcus
Specimen Sputum, CSF, Blood On Blood Agar β-hemolysis Presumptive Identification tests Catalase-negative CAMP test–positive Bacitracin-resistant

Group D -1: Enterococcus
Formerly called Streptococcus E. faecalis & E. faecium are the most prevalent species Members of the gut flora Tolerant to 6.5 % Na CL Hydrolyze esculin in the presence of bile Important clinical infections include: UTI (associated with urinary tract catheterization ) Bacteremia Bacterial endocarditis (5-15% of endocarditis cases) Meningitis (uncommon )

Treatment of Enterococcus
Have high level of intrinsic antibiotic resistance Some are intrinsically resistant to β-lactams & aminoglycoside Sensitive strains treated with ampicillin and vancomycin Emergence of Vancomycin Resistant Enterococcus (VRE) in nosocomial infections VRE may be treated with quinupristin/dalfopristin

Group D 2- Non-Enterococcus
Streptococcus bovis Normally found in the rumen of cattle and the colon of simple stomached animals Hydrolyze esculin in presence of bile But not tolerate 6.5% NaCl Important clinical infections include: Most common infection: Bacteremia with or without Endocarditis Less-common infections Neonatal septicemia, meningitis, UTI, Wound infection

-Hemolytic Streptococci: Viridans Group
Large complex group Strep. mutans, S. oralis, S. salivarus, S. mitis Gram positive cocci,  hemolytic, catalase –ve etc Members of normal oral and nasopharyngeal flora Diseases Subacute bacterial endocarditis (Serious infection) 30-40% of cases due to viridans streptococci Involved in dental caries Bacteremia

Subacute bacterial endocarditis
People with congenital heart problems, heart surgery, previous episodes of endocarditis, and intravenous drug use Dental or surgical procedures facilitate entrance to blood Bacteria settle and grow on heart lining or valves Colonization of heart by forming biofilms Treatment is prolonged course of Penicillin Receive prophylactics before surgery or dental procedures Diagnosis Blood Culture 5-10 ml patient’s blood during fibril ml nutrient broth Subculture on blood agar -hemolytic, optochin R

Dental Caries (tooth decay)
S. mutans is the leading cause of dental caries Plaque is a sticky film on teeth Bacteria in a mouth convert sucrose into acids (lactic acid) These acids may cause demineralization, which is the dissolution of its mineral content If demineralization continues over time, enough mineral content may be lost forming a cavity or hole S. mutans produce extracellular enzyme glucosyltransferase Synthesizes glucans from sucrose Insoluble Glucan (Slime layer) is important for enhancement of attachment of the S. mutans to the tooth enamel

Strept. pneumoniae (Pneumococcus)
General characteristics Gram +ve small, lancet-shaped, arranged in pairs Culture requires blood or chocolate agar Growth improved by 5-10% CO2 Inhabits nasopharyngeal areas of 5-50% healthy individuals Typical opportunist Very delicate, does not survive long outside of its habitat Virulence factors Polysaccharide capsule (antiphagocytic) 90 capsular types (Serotypes) have been identified IgA protease (Colonization factor)

Disease caused by S. pneumoniae
Invasive Pneumococcal Disease (IPD) Meningitis, bacteremia (life-threatening infections) Non-Invasive Pneumococcal Disease NIPD Pneumonia (life-threatening infection) Causes 60-70% of all bacterial pneumonias Young children, elderly, immune compromised, those with lung diseases or viral infections, persons living in close quarters are predisposed to pneumonia Sinusitis/otitis media

CLINICAL PRESENTATION
Pneumococcal disease is common during winter/ spring Characterized by Sudden onset of fever and malaise In pneumonia, fever may precede the usual symptoms of cough, pleuritic chest pain & production of purulent or blood-tinged sputum by 12–24 hours Bacteremia occurs in 15-25% of cases Meningitis with stiff neck, headache, lethargy or seizures MODE OF TRANSMISSION S. pneumoniae is transmitted directly from person to person through close contact via respiratory droplets

Cultivation and Diagnosis
Specimen Sputum, CSF, Blood Isolation from specimen with normal flora provide supportive evidence but not proof of infection Predominant growth of Pneumococci in sputum with numerous pus is likely to be significant & confirmed Specimen is cultured on blood agar then Gram stain Presumptive identification Quellung test or capsular swelling reaction Pneumococci mixed with antiserum on a slide The capsule swells markedly Optochin sensitive and soluble in bile

Treatment and Prevention
Traditionally treated with penicillin G or V Emergence of Penicillin Resistant S. pneumoniae PRSP treated with others such as macrolides Two vaccines are available 23-valent pneumococcal polysaccharide vaccine (PPSV23) Recommended for adults ≥65 years and for people aged 2–64 years with underlying medical conditions Covers 85%-90% of serotypes that cause IPD Because PPSV23 is not effective in children <2 years so conjugate vaccines were developed

Pneumococcal Conjugate Vaccines
7-valent pneumococcal conjugate vaccine (PCV7) Routine infant immunization schedule in US since 2000 Recommended for all children aged <5 years PCV7 covers 80-85% of invasive disease in young 13-valent pneumococcal conjugate vaccine (PCV13) Other pneumococci are becoming increasingly more common in young children so PCV13 replaced PCV7 PCV7 & PCV13 are recommended for children aged 2–59 M as a 4-doses at ages 2, 4, 6, and 12–15 M

General Characteristics of Bacillus
>60 species; Most are saprophytic or normal flora Bacillus are ubiquitous in soil, water and airborne dust 2 medically important; B. anthracis and B. cereus Morphology Large Gram-positive Spore forming (central & oval) bacilli and arranged in chains Spores contaminate soil & remain viable for many years Motile except B. anthracis General physiological characteristics Non-fastidious, facultative anaerobic, Fermentative Catalase positive Clostridium (catalase –ve) Thermophilic (< 75°C) and psychrophilic (≥5-8°C) Can grow at extremes of acidity & alkalinity (pH 2-10)

Bacillus anthracis Pathogenesis “Virulence Factor”
Poly-D-glutamyl Capsule Non-immunogenic and unique Mediates the invasive stage of the infection Antiphagocytic Anthrax Exotoxin (Thermolabile protein) Mediates the toxigenic stage Consists of 3 distinct antigenic components Protective Antigen Edema Factor Lethal Factor

A. Bacillus anthracis Disease: Anthrax
B. anthracis primarily infects herbivores animals zoonotic disease Direct person-to-person spread is unlikely to occur Portal of entry: Skin, GIT, Respiratory tract Mode of transmission Direct contact with infected animals or soils When eating meat of infected animals Inhalation of spores when Handling with infected animal products (e.g. wool) Biological war and Bioterrorism

Types of Anthrax Cutaneous Anthrax (Malignant Pustule)
95% of human anthrax is cutaneous Resolves spontaneously in 80-90% of cases 20% mortality in untreated cutaneous anthrax Portal of entry: Injured skin Spores contaminate injured skin (Face, neck, arms) Spores germinate, multiply then produce exotoxin

1. Cutanoues Anthrax After 1-5 days contact:
Small, pruritic, non-painful erythermatous papule Papule develops into hemorrhagic vesicle and ruptures to form ulcer Slow-healing painless ulcer covered with black scar surrounded by edema Pus & pain are absent (non-pyogenic organism) May progress to toxemia and death Infection spread to lymphatics with local adenopathy

2. Intestinal Anthrax Rare but virtually 100% fatal
Occurs accidentally among butchers and in primitive societies eating meat of infected animals It is analogous to cutaneous anthrax but occurs on intestinal mucosa Vomiting, abdominal pain and bloody diarrhea

3. Pneumonic Anthrax (Woolsorters disease)
High mortality rate (50-100%) Result from inhalation of spore-containing dust Spores germinate in the lungs and secrete toxins Toxin absorbed into the bloodstream Disease progress from malaise with mild fever & non-productive cough to respiratory distress, massive chest edema, cyanosis and death Complication cutaneous & pneumonic anthrax: Meningitis

Prevention and Control
Rapid identification and treatment Penicillin is critical in preventing high mortality of anthrax Ciproflaxacin and many other antimicrobials are effective Control the disease in animals by Vaccination of uninfected animals Burning or burying in lime pits of infected animals Destroy spores on animal hairs

Animal Vaccine Live attenuated vaccine (loss of the capsule) The animal vaccine is dangerous to human Human Vaccine Human vaccine composed of PA Given to people under risk such as military personnel 6 doses, first 3 doses are given at 2-week intervals 3 additional doses, at 6, 12 & 18 M after first dose Annual booster dose are needed for ongoing protection

B. Bacillus cereus B. cereus is a normal inhabitant of soil
Isolated from food such as grains and spices Diseases Food poisoning (self-limiting) usually lasts < 24 hrs after onset Opportunistic pathogen can cause Septicemia, Endocarditis, Meningitis, Pneumonia, Wound & eye infections Virulence factors 2 types of Enterotoxins (HL and HS) Dermonecrotic and lethal toxin Hemolysins, lecithinase, proteases, and nucleases Treatment Clindamycin with or without gentamicin may be used Resist penicillin due to production of β-lactamases

Food poisoning Two types of food poisoning
Emetic form or short incubation (1-6 hrs): HS enterotoxin associated with rice dishes Victim eats preformed toxin (Intoxication form ) Nausea, vomiting and abdominal cramps It resembles S. aureus food poisoning Diarrheal form or long incubation (6-18 hrs): HL enterotoxin associated with meat dishes Victim ingests organism then produce toxin Diarrhea and abdominal cramps It resembles Cl. perfringens food poisoning

Identification of Bacillus Spp.
Specimen Pastular exudates in malignant pustule Sputum in pneumonic anthrax Stool in intestinal anthrax Stool in intestinal anthrax and food poisoning Cultural characteristics Nutrient agar, blood agar Microscopical Gram stain, spore stain, capsule and motility

Key Characteristics to Distinguish between B
Key Characteristics to Distinguish between B. anthracis & Other Species of Bacillus Other Bacillus B. anthracis β No-hemolysis Hemolysis Motile Non-Motile Motility - + Capsule Gelatin hydrolysis Salicin fermentation Resistant Sensitive Penicillin Sensitivity

A. Clostridium tetani General characteristics
Gram positive spore forming bacilli with rounded ends Round terminal spores give drumstick appearance Obligate anaerobe and Fermentative Motile by peritrichous flagella Produces β-hemolysis & thin spreading film on blood agar Habitat Colonizes the intestinal tract in humans and animals The spores are distributed widely in manure-treated soils Disease Cl. tetani is the causative agent of tetanus (LOCK JAW)

A. Clostridium tetani Mode of infection
Contamination of wound with spores of Cl. tetani In developing countries most cases occur in Children secondary to wounds Neonates with contaminated umbilical stump after labor After surgery done with non-sterilized instruments Use of badly sterilized cut gut "surgical ligatures" Virulence factors (Pathogenesis) Cl. tetani produces two types of exotoxins: Tetanolysin, which causes lysis of RBCs Tetanospasmin: neurotoxin

Incubation period How tetanus occur?
Several months but is usually about 8 days How tetanus occur? Spores germinate in wound & elaborate tetanospasmin Tetany occur when toxin is reached at neuromuscular junction and Toxin transported to CNS Binds to inhibitory neurotransmitters (GABA) Blocks their release of inhibitor molecules Lead to send a high frequency of impulses to muscles This lead to sustained tetanic contraction Death is usually from respiratory failure

Types of Tetanus Generalized tetanus Signs and Symptoms
The most common and the most severe form Affect all skeletal muscles Signs and Symptoms Trismus (lockjaw) & facial spasms (risus sardonicus) Followed by stiffness of the neck, difficulty in swallowing and rigidity of muscles  temperature, sweating, blood pressure, heart rate Spasms occur frequently & last for minutes with body shaped into characteristic form (opisthotonos)

Similar to generalized tetanus except affects neonates
Neonatal tetanus Similar to generalized tetanus except affects neonates Rare in developed countries Local tetanus Muscle spasms at or near the infected wound Can progress to generalized tetanus Cephalic tetanus Affect facial muscles:2days after head injury or ear infection Involvement of cranial nerves, especially in facial area Lockjaw and facial spasms Can easily progress to generalized tetanus

Laboratory Diagnosis of Tetanus
It depends primarily upon the clinical manifestation of tetanus including muscle spasm and rigidity Specimen: Wound exudates using capillary tube Culture: On blood agar and incubated anaerobically Growth appears as fine spreading film & β-hemolytic Gram stain is a good method for identifying Gram positive rod, motile with a round terminal spore giving a drumstick appearance

Prevention Prevented by active Immunization with tetanus toxoid
Toxoid is inactivated toxin either by formal or alum DTP give at 2, 4, 6 and 18 months of birth Booster dose is given before school entry at 4-6 years Prophylactic dose is performed once every 10 years Pregnant women must take the vaccine

Treatment Clear the toxin and the toxin producing bacteria
Give Anti-Tetanic Sera (ATS)- Passive immunization To neutralize the circulating toxins ATS is tetanus immune globulins Give booster dose DTP to stimulate immune response Clean the wound, excising any devitalized tissue To remove source of C. tetani Give antibiotics such as penicillin or metronidazole To eradicate the toxin-producing organism Provide intensive support therapy until the toxin is cleared Administration of Muscle relaxants Patients placed on a Ventilator

b. Clostridium botulinum
General Characteristics Strict anaerobic Gram positive bacillus Oval sub-terminal spores Motile with peritrichous flagella Often diffusely beta hemolytic Natural habitats Widely distributed saprophyte occurring in soil, vegetable, fruits Disease Botulism (rare and severe) Food poisoning

Ingestion of Organism, Spores and Neurotoxin
Pathogenesis Botulinal toxins (Exotoxins, Neurotoxins) Among the most poisonous natural substances Divided into four groups (I-IV) based on the type of toxin Seven toxin types (A-G) A, B, E & F most frequently associated with human disease All botulinum toxins produce the same clinical signs Mode of infection Spores widely distributed in soil & contaminate vegetables & meat Ingestion of Organism, Spores and Neurotoxin Wound contamination Inhalation (Biological war) Person-to-person not documented

Types of human botulism
Adult botulism Caused by eating foods that contain toxin Foods usually preserved meat, large sausages, home-preserved meats, vegetables, smoked fish, canned food etc. Infant botulism Caused by consuming the spores of Cl. botulinum Grow and release toxin in the intestine Wound botulism (Rare) Caused by toxin produced from a wound infected with Cl. botulinum

1. Human botulism (Food borne)
Food cooked insufficiently that placed into a jar or can Organism matures and secretes neurotoxin Man ingest canned food containing toxins Toxin is absorbed from GIT & affects cholinergic system It blocks release of acetylcholine (Excitatory neurotransmitters) at points in peripheral nervous system Blocking muscle stimulation Resulting in flaccid paralysis Clinical features The incubation period is usually 1-2 days Nausea, vomiting, diarrhea Double vision and difficulty speaking or swallowing Descending weakness or paralysis Symmetrical flaccid paralysis & respiratory muscle paralysis

2. Infant Botulism Ingestion of food (HONEY) contaminated with the spores Spores germinate and the organism colonized GIT of infants The organism elaborates the toxin Infants < 1 year old- 94% < 6 months old The infant will be constipated for 2-3 days Followed by difficulty swallowing and muscle weakness Lethargy (كسل وفتور ونوم), Poor feeding, Weak cry Bulbar palsies (impairment of function of the cranial nerves IX, X, XI and XII) Failure to thrive (poor physical growth ) The baby must be hospitalized and given supportive therapy Prognosis is excellent, so antitoxin is generally not used

Laboratory diagnosis Clinical signs provide tentative diagnosis for botulism intoxication The diagnosis may be confirmed by demonstration of: Organism (usually not cultured) is detected in Patient's stool and Suspected food Specimens are inoculated on blood agar anaerobically Gram stain of the suspected colonies revealed that the organism is gram positive bacilli, motile and sporulated Toxin is detected in Patient's stool, Suspected food, Patient's serum by toxin-antitoxin neutralization test in mice

Home canning of foodstuffs should be avoided Commercial canning must be strictly controlled Swollen canned food must be discarded Sterilization of all canned and vacuum packed food is essential Prevention germination of spores in foods by Maintaining food in acidic pH and high sugar content Storing the food at 4 0C or colder Destroy toxins by heating of food for 20 min at 80 0C A prophylactic dose of polyvalent antitoxin should be given to all persons who have eaten food suspected of causing botulism Active immunization should be considered for under risk Person handles organism or specimen containing organism/toxin Infants ≤ 1 year should not be given honey or food containing it

Treatment The priorities are To remove unabsorbed toxin from the GIT
To neutralize unfixed toxin by giving polyvalent antitoxin To give relevant intensive care and support Intubation and ventilatory support is critical To give antibiotics such as penicillin or metronidazole Most cases of botulism require immediate intensive care treatment. Due to respiratory paralysis, a mechanical ventilator will be needed if respiratory failure occurs. An intravenous equine-derived botulinum antitoxin is available on a case-by-case basis from the CDC through state and local health departments. Botulism immune globulin was approved for use on October 23, 2003 for the treatment of infant botulism caused by types A and G.

C. Clostridium perfringens
General characteristics Anaerobic Large Gram positive Rod-shaped with stubby (short) ends Oval, sub-terminal and not bulging Spore Spores are seldom to see Non-motile Double-zone hemolysis on blood agar Natural habitats Animal and human excreta Widely distributed in soil and sewage Diseases Gas gangrene, myonecrosis (Histotoxic) Food poisoning (Enterotoxegenic)

Pathogenesis (Virulence factor)
Toxins Five different biotype of Cl. perfringens (A-E) They are differentiated on the basis of production of toxins Four type of toxins (, β, epsilon and iota) The most important is  toxin (lethicinase) All biotypes contain  toxin Splits lecithin in cytoplasmic membrane Necrotizing and hemolytic effect (RBCs rupture ) Responsible for sever toxemia in gas gangrene All biotypes causing gas gangrene Collagenase Hyaluronidase DNase Enterotoxin (heat labile) Biotype A cause food poisoning due to enterotoxin Digest subcutaneous tissue and muscles

i. Histotoxic: Gas Gangrene
Mode of Infection Contamination of wounds with a soil which contain spores Post operative from badly sterilized cat-gut Gas gangrene is likely to occur when: There is extensive damage to tissues Interruption of blood supply Contamination of wound with soil and foreign material Such conditions are particularly associated with War wounds, Accidental fracture or Industrial injuries Uterine infections may occur after septic abortion Incubation period 1-4 day & earliest symptom is severe pain at site of wounds

How Gas gangrene occur? Impairment of blood supply of tissue cause an anaerobic focus Infection initiated in a focus of devitalized anaerobic tissue The organisms multiply and produce a range of toxins Spread and attack adjacent viable tissue (muscle fibers) Lead to kill tissues and render it anaerobic Fermentation of muscle glycogen with acid & gas production The gas separates the fibers from its sheath Cutting of blood supply leading to necrosis This produce the black color and foul smelling of gangrene Organism is further colonized with production of more toxins

Clinical findings Pain, odema, cellulitis occur in the wound area
Crepitation indicates the presence of gas in issues Hemolysis and Jaundice are common Shock and death Mortality rate is high

ii. Food poisoning Food poisoning is caused by biotype A
Food (meat) is often contaminated with the spores The heat resistant spores survive cooking Spores germinate and multiply (especially if unrefrigerated) When consumed, enterotoxin is produced in the GIT After ingestion (8-12 h): Abdominal cramps then diarrhea Fever and vomiting are not typically encountered Symptoms generally subside within a day or two Similar to food poisoning with B. cereus "Diarrheal form”

Prevention and treatment
Wound must be cleaned to prevent gas gangrene Gas gangrene is fatal unless identified & treated very early Removal of necrotic tissue can be life saving Hyberbaric oxygen Antibiotics (such as penicillin G in large doses) Administration of antitoxin No Vaccine available to protect against gas gangrene Food poisoning is prevented by avoid mishandling of foods Type A toxoid vaccine for animals to prevent gastroenteritis

Laboratory Diagnosis Gas gangrene
Specimen: Histological specimen or wound exudates They should be taken from the deeper areas of wound Culture: Anaerobically at 370C On Robertson's cooked meat medium → blackening of meat will observed with the production of H2S and NH3 On blood agar → double zones of β-hemolytic colonies Microscopical examination Gram and spore stains give the characteristics of organism Biochemical test Positive for Stormy Clot and Nagler reaction Food poisoning Lab diagnosis is not usually done Isolation of Cl. perfringens from feces of patients, from those at risk who have eaten the suspected food & from food itself

D. Clostridium difficile
General characteristics Strict anaerobic Gram positive rod Sub-terminal Oval spores and Motile Non-hemolystic Natural habitats Cl. difficile is part of the intestinal flora Quite commonly in the faces of neonates 2% of healthy adults colonized 10-20% of elderly individuals colonized 20-40% of hospitalized patients colonized Isolated from toilets, bedpans, floors

Disease Way of infection Toxins Antibiotic associated diarrhea
Pseudomembranous colitis Way of infection Endogenous route Unrestricted multiplication of organism in GIT after Abdominal or intestinal surgery Certain antibiotics such as penicillin, clindamycin Exogenous route Non colonized patients may become colonized (20-40%) Toxins Proliferation of Cl. difficile with localized production of their toxins in the colon leads to disease Toxin A (Enterotoxin) and Toxin B (Cytotoxic)

Clinical findings Laboratory diagnosis Treatment
Fever, bloody diarrhea and abdominal cramps The most serious complication is toxic megacolon Laboratory diagnosis Observation of colonic pseudomembranes (white exudates on surface of large intestine) by colonoscopy is diagnostic for pseudomembranous colitis Isolation from faces on selective media. Toxin can be detected in the patient's feces by ELISA Culture without demonstration of toxin has little diagnostic value Treatment It is essential to discontinue the antibiotic Vancomycin or metronidazole

1. Corynebacterium C. diphtheriae is the most important pathogen
It inhabits nasopharynx but only on carrier state Not considered part of normal flora Isolation from health human (Carrier) is not common Diphtheroids (e.g. C. xerosis ) Some are saprophytic Some produce disease in animals Colonizes skin, GIT, UT, respiratory and genital tracts

Corynebacterium diphtheriae
Small, pleomorphic gram-positive bacilli Club-shaped (tapered at one end) Arranged in palisades or in V or Y configurations or Clumps resembling “Chinese letters” They are beaded Beads contain metachromatic granules Stained with polychrome methylene blue stain Non motile, Non spore forming, Capsulated (K antigen) Fastidious; Slow growth on enriched medium Facultative anaerobic , Fermentative , Catalase positive C. diphtheriae is the causative agent of DIPHTHERIA Transmitted by respiratory droplet from case or carrier

Pathogenesis (Virulence factors)
C. diphtheriae produce powerful exotoxin Not all C. diphtheriae secretes the toxin Lysogenic bacteriophage encodes tox gene tox responsible for production of potent toxin in virulent strain The toxin inhibits protein synthesis which results in cell death

Diphtheria Diphtheria is childhood and Toxin mediated disease
Incubation period 2-6 days 2 stages of disease: Local infection –upper respiratory tract inflammation Sore throat, pharyngitis, swollen lymph nodes, low-grade fever, Firmly adherent pseudomembrane covered tonsils, uvula, palate The major complication is respiratory obstruction due to Pseudomembrane extends up or down into nasopharynx & larynx Difficulty in breathing due to respiratory obstruction Palate perforation (Localized damage)

Diphtheria Antibody neutralizes toxin by blocking interaction of subunit B Prevent entry the cell Immunity status assessed by SCHICK’S TEST If toxin is reached the blood, the disease proceeds 2nd stage Diptherotoxin production and toxemia Target primarily heart (Myocarditis) and nerves (Neuritis) Recovery or complication and death can occur Most mortality from systemic toxin-mediated heart failure

Laboratory diagnosis Specimen Culture
Throat swap by gentle touch of membrane to avoid bleeding Culture Loeffler's serum medium or Blood Tellurite Agar (BTA) 3 biotypes of C. diphtheriae are characterized on BTA Gravis, mitis and intermedius biotypes The most severe disease is associated with gravis biotype Microscopy Polychrome methylene blue stain shows metachromatic granule Gram stain shows Gram-positive pleomorphic rods Elek’s Test In vitro test is used to detect toxins This done by a precipitation of exotoxin with diphtheria antitoxin

How to test for susceptibility to diphtheria by Schick test
Diphtheria toxin produces marked local reaction when injected ID A small dose of toxin injected ID in one forearm (Test) Similar dose of heated toxin is injected ID in other forearm (Control) Positive Schick test Redness and swelling in test forearm Reaction increases for several days and then slowly disappear Means the person is non-immune and sensitive Susceptible to diphtheria Negative Schick test No reaction appears in both forearms (Test & Control) This means the person is immune and non-sensitive

Immunization Treatment Active immunization (Prophylaxis)
Diphtheria Toxoid (Inactivated diphtheria toxin) DTP vaccine (Containing Diphtheria Toxoid) 3 doses at 2 months intervals Given at 2,4,6 months of birth Booster doses at 18 month and school entry age Booster dose each 10 years Passive immunization (Treatment) Administration of antitoxin Treatment Administration of antitoxin to neutralize the circulating toxin Penicillin or Erythromycin to eradicate organism

Listeria General Characteristics
Gram-positive short coccobacilli (0.5-2 by µm) Singly or in short chains Non-spore formers, motile (tumbling motility at room temp) Facultative anaerobes, Fermentative, Catalase positive Weak beta-hemolytsis Growth range = (-1 to 45°C) optimum 30-35ºC Grows at pH 4.4 and 9.6 optimum pH Facultative intracellular pathogen 10 species; L. monocytogenes is primary human pathogen

Pathogenesis: Virulence factors
Growth at room temperature, Motility Adherence and invasion Attachment and invasion of intestinal mucosa D-galactose receptors on host intestinal epithelial cells by bacterial surface D-galactose residues The bacteria are then taken up by induced phagocytosis Organism can grow in macrophages and epithelial cells Listeriolysin O (Hemolysin) The most significant virulence factor Lysis RBCs (β-hemolysis on blood agar) It breaks phagosome and enter the cytoplasm of the phagocyte Multiplies in the phagocyte and invades other tissues Catalase and superoxide dismutase Neutralize the effects of the phagocytic oxidative burst

Where do we find Listeria?
Widely distributed in nature Environment Soil, water, animal feces, sewage, decaying vegetation Food Fresh food products of animal or plant origin Animal food mammals, fish, birds (especially chickens) Animals (reservoirs ) carry Listeria without appearing ill 5-10% of humans may be asymptomatic intestinal carriers

Mode of Transmission Portal of entry is from GIT to blood and from blood to menings It is a foodborne pathogen and transmitted by Usually by ingestion of contaminated food with Listeria Raw or undercooked food of animal origin (Luncheon, Hot dogs) Soft cheeses and un-pasteurized milk Unwashed raw vegetables Listeria grows in a wide range of pH and in cold temperatures Refrigeration of contaminated foods permit the multiplication of the organism to an infectious dose Colonized mothers may pass organism onto fetus Its presence intracellularly in phagocyte permits access to the brain and transplacental migration to fetus in pregnant women Listeriosis is not contagious from person to person

Clinical Finding Non-perinatal infections Perinatal infections
Listeriosis is general group of disorders It is defined when Listeria is isolated from blood, CSF The manifestations of listeriosis include Non-perinatal infections Septicaemia, Meningitis, Encephalitis Perinatal infections Spread to the fetus Sepsis, Miscarriage, Stillbirth Spread to the newborn baby Meningitis Listeria is 3rd most common cause of neonatal meningitis People under risk: Neonates and Pregnant women Immunocomprised patients with defective in cell-mediated immunity (Renal transplant, Cancer, AIDS, Elderly)

Why L. monocytogenes invades neonates & certain imuno-compromised?
L. monocytogenes is a resistant fellow, able to hide and survive within macrophages and neutrophils Facultative intracellular pathogen It can survive either within or outside the cells In immune person, the immune system release factors that activate macrophage so macrophage destroy Listeria There is defect in cell-mediated immunity in immuno-compromised

Diagnosis of Listeriosis
Listeriosis can only be positively diagnosed by culturing the organism from blood, CSF, or stool (although the latter is difficult and of limited value) CSF contains neutrophils,  protein level,  glucose level Culture: Cold technique can be used to isolate Listeria selectively On 5% blood agar: small, white, smooth, translucent, moist, β-hemolytic colonies Gram stain Gram-positive bacilli, motile at 20 0C etc CAMP Test positive (like Group B Streptococcus)

Prevention and Treatment
L. monocytogenes is killed by cooking or by heating methods, including pasteurization So Avoid consumption of raw milk or milk products Pregnant or immunocompromised should avoid soft cheeses Reheat leftover or ready-to-eat foods hot dogs until steaming hot Cook food to a safe internal temperature Wash raw vegetables Wash hands, knives & cutting boards after handling uncooked foods Treatment Listeriosis is a serious disease requiring hospitalization Parenteral penicillin or ampicillin with or without an aminoglycoside Trimethoprim/sulfamethoxazole (in patients allergic to penicillin) The duration of antibiotic treatment is at least 2 weeks

II- Acid-Fast Bacilli Genus Mycobacterium
Grow slowly Acid-fast staining Resist decolourization with HCl 3% and 95% Ethanol Gram-positive irregular bacilli Difficult to stain by Gram stain High lipid content (40%) in its cell wall Colonies rough and hydrophobic Strict aerobes, non-capsulated, non-motile, non-spore forming Facultative intracellular pathogen Produce catalase, peroxidases and superoxide dismutase Possess mycolic acids and a unique type of peptidoglycan

Mycobacterium tuberculosis
Appendix 12 - Fundamentals of TB Presentation Mycobacterium tuberculosis Cell wall contains several complex antigens Lipoid fraction (Cord factor) Trehalose dimycolate correlates with virulence of TB Virulent strains grow in a characteristic cordlike pattern Avirulent strains do not Loss of the cord factor results in attenuation of TB (vaccine) Lipid fraction Long chain fatty acids (MYCOLIC ACID) Responsible for acid fastness and slow growth Wax D Phospahtides- play role in caseation necrosis Protein fraction (Purified proteins derivatives, PPD) When combined with wax D elicit delayed hypersensitivity They used in the skin test

Appendix 12 - Fundamentals of TB Presentation
Pathogenesis No exotoxins or enzymes Contain complex waxes and cord factor that prevent destruction by lysosomes or macrophages after phagocytosis When phagocyte dies & disintegrates So it multiplies in the cytoplasm of phagocyte Immunity of TB: T.B. stimulates Cellular immunity (Protective) (CD4-positive T cells) Ability to localize TB and retard its multiplication Humoral immunity (non protective) Delayed hypersensitivity (harmful) Related to the pathology of the disease Due to the protein fraction

Disease Tuberculosis: The most important communicable disease
M. tuberculosis (Human type) and M. bovis (Bovine type) WHO estimates that 1/3rd of world population carry latent TB Around 3 million people worldwide die of TB each year The symptoms are due to delayed hypersensitivity or host tissue reaction response that result in tissue damage Predisposing factors include: Inadequate nutrition, debilitation of the immune system (HIV), crowdedness, poor access to medical care Way of infection Human type by airborne respiratory droplets Bovine type by consumption of milk from infected animals

I- Pulmonary tuberculosis a. Primary tuberculosis
Occur by airborne respiratory droplets (10 cells) Alveolar macrophage phagocytosed TB and multiply intracellularly Primary site of infection in the lungs, known as "Ghon focus” Tubercle granuloma is formed after immune system attacks 4 week It consist of central core contain TB surrounded by WBC If center of tubercle breaks down into necrotic caseous lesions Gradually heal spontaneously or become Latent TB infection (LTBI) TB remain in this impasse of dormant infection (LTBI) for years Primary tuberculosis leaves the patient immune

I- Pulmonary tuberculosis b. Secondary TB
Secondary TB occurred by reactivation or re-infection in: Patients have been recently infected LTBI’s patients reactivated to TB disease 10% of LTBI will develop TB disease at some point in their lives 5% within 1-2 years and 5% at some point in their lives This happens to a weakened or suppressed immune system The tubercle is ruptured and T.B. spread through the lungs One or more lung lesions progress to caseation and cavitation Create case of open tuberculosis i.e. A patients cough up T.B. in a sputum and infect others Symptoms: Fever, anorexia, weight loss, fatigue, night sweats, violent coughing, greenish or bloody sputum, chest pain Sometimes in active disease the defensive forces of the body may overcome TB and render it inactive (arrested cases)

LTBI TB Disease Tubercle bacilli in the body Tuberculin Skin Test result usually positive Chest x-ray usually normal Chest x-ray usually abnormal Sputum smears and cultures negative Sputum smears and cultures positive No symptoms Symptoms such as cough, fever, weight loss Not infectious Often infectious before treatment Not a case of TB A case of TB Needs treatment for LTBI to prevent TB disease Needs treatment to treat TB disease

II- Extrapulmonary TB Alternative name: Disseminated TB, Miliary tuberculosis 1.5% of patients are estimated to have miliary tuberculosis Miliary tuberculosis is the widespread dissemination of TB through infected macrophages from lung via hematogenous spread to regional lymph nodes, kidneys, long bones, genital tract, brain and meninges It may occur in an individual organ (very rare, < 5%), in several organs, or throughout the entire body (>90%), including the brain Set up many foci of infection, all appearing as tiny, white tubercles in the tissues It most common in young children and those with HIV

Laboratory Diagnosis Direct Specimen Sputum, blood, urine, or CSF Sputum specimens are essential to confirm TB Collect 3 morning specimens on 3 different days Acid Fast Stain (AFS) Positive in patients with smears containing acid-fast bacilli Only positive when large numbers of T.B. begin to be excreted Culture Culture all specimens, even if smear is negative On Lowenstein-Jensen Medium 4-6 weeks needed for growth

Indirect Tuberculin skin test (PPD skin test, Mantoux test) To determine if someone has developed immune response to TB This response can occur if someone currently has TB, if They were exposed to it in the past, or if They received the BCG vaccine against TB It based on delayed-type hypersensitivity skin reaction to PPD Local ID injection of 0.1 ml of PPD in the forearm Look for red wheal induration to form in h <5 mm is considered negative Negative test doesn’t always mean that a person is free of TB ≥5 mm is considered positive In people exposed to TB infection, with or without disease People have not received vaccine yet A positive test has limited diagnostic significance in older age

PREVENTION BCG Vaccine Eradication of infected animals by slaughtering
Pasteurization of milk Improved social conditions- housing /nutrition Case detection & treatment Treatment of infected / diseased contacts BCG Vaccine Based on attenuated Bacilli Calmet-Guerin strain of M. bovis Its given as single dose (0.1 ml ) within the first month of age Tuberculin test must be done before vaccination in children ≥ 36 months

Treatment of TB Disease
Appendix 12 - Fundamentals of TB Presentation Treatment of TB Disease Without treatment, Mortality is 50% Antitubercular drugs 1st-line drugs in initial regimen (more effective) Isoniazid (INH), Rifampin (REF), Pyrazinamide (PZA), Streptomycin (SM) Second-line medicines include Ethambutal (EMB), para-amino-salicylic acid (PAS), kanamycin, amikacin Why is treatment of T.B required long term therapy? Presence of TB in caseous protective material Bacteria can live by intracellular Drugs penetration is slow

Multi-drug therapy for long duration is recommended
Primary resistance - infection with a strain of TB that is already resistant to one or more drugs Acquired resistance - infection with a strain of TB that becomes drug resistant due to inappropriate or inadequate treatment Multi-drug therapy for long duration is recommended Drug combinations used to reduce emergence of resistance Second drug will inhibit T.B. that become resistant to one drug Routine chemotherapy INH+SM+PAS month Short-term chemotherapy INH+RFP M Directly Observed Therapy (DOT) Health care worker watches patient swallow each dose of drugs Patient goes to the clinic or DOT worker goes to patient’s home DOT helps a patient complete TB treatment Reduces relapse of TB disease and acquired drug resistance

b. Mycobacterium leprae (Hansen’s bacillus)
Acid fast bacilli (Modified ZN stain) Cannot grow in bacterial media/culture Grow on mouse foot pads and in nine banded armadillo Causes leprosy, a chronic disease that begins in the skin and mucous membranes and progresses into nerves The lesions involve cooler tissue of the body Skin, superficial nerve, nose, pharynx, larynx, eyes & testicles Man is the only reservoir of proven significance Patient discharges M. leprae in nasal secretion & from skin lesions Transmitted by prolonged contact with Lepromatous patients

b. Mycobacterium leprae
Leprosy occurs worldwide Commonly in tropical areas of Asia and Africa World prevalence million Incubation from 2-10 years The onset of leprosy is insidious Macrophages phagocytize T.B, but a weakened macrophage or slow T cell response may not kill bacillus If untreated, bacilli grow slowly in the skin macrophages and Schwann cells of peripheral nerves Spectrum of clinical presentations Dependent on host –parasite interactions 2 clinical forms possible: Tuberculoid- and Lepromatous Leprosy

Course of Infection and Disease
Tuberculoid Leprosy (TL) TL is less severe and more superficial than LL The affected person manifests considerable biological resistance to the disease Superficial infection without skin disfigurement which damages nerves and causes loss of pain perception It advances slowly, if there is a little chance of it progressing Few bacilli present in the lesion So called Paucibacillary (PB) Hansen's disease Positive lepromin test Because cell mediated immunity (CMI) is intact and the skin infiltrated with helper T cell

Course of Infection and Disease
Lepromatous Leprosy (LL) A deeply nodular infection that causes severe disfigurement of the face and extremities The affected person manifests no resistance to the disease Abundant bacilli present in the lesion So called multibacillary Hansen's disease Negative lepromin test because the cell mediated immunity is markedly deficient and the skin infiltrated with suppressor T cell

Laboratory Diagnosis Detection of acid-fast bacilli in skin lesions, nasal discharges, and tissue samples Lepromin Skin Test Similar to tuberculin test A test to evaluate leprosy using ID injection of a lepromin Induration is observed 2-4 weeks The test classifies the stage of leprosy based on reaction and differentiates tuberculoid leprosy, in which there is a positive delayed reaction at the injection site, from lepromatous leprosy, in which there is no reaction despite the active infection.

Treatment and Prevention
The three most commonly used antibiotics are: Dapsone (The most important), Rifampin and Clofazimine Treatment by long-term combined therapy (≥2 years) Combination therapy (MDT) is recommended Dapson+Rifampcin for Tuberculoid Leprosy (PB) Dapson+Rifampcin+Clofazimine for Lepromatous Leprosy (MB) Prevention requires constant surveillance of high risk populations WHO sponsoring a trial vaccine

Infections by Non-Tuberculosis Mycobacteria (NTM)
M. avium complex – third most common cause of death in AIDS patients M. kansaii – pulmonary infections in adult white males with emphysema or bronchitis M. marinum – water inhabitant; lesions develop after scraping on swimming pool concrete M. scrofulaceum – infects cervical lymph nodes M. paratuberculosis – raw cow’s milk; recovered from 65% of individuals diagnosed with Crohn’s disease

1. Neisseria species Primary pathogens: Non-pathogenic (Commensals)
N. gonorrhoeae (Gonococcus) ALWAYS pathogenic N. meningitidis (Meningococcus) May be carried as commensal in nasopharynx flora (10-25% carrier) Grow on chocolate agar at 370C under 5-10% CO2 Non-pathogenic (Commensals) Example N. lactamica Grow on ordinary medium at room temperature Habitat of non-pathogenic Upper respiratory, Genitourinary and Alimentary tract

Pathogenic Neisseria General characteristics
Gram-negative diplococci with adjacent sides flattened (like coffee beans) Non motile Oxidase and Catalase positive Aerobic, capnophilic (5% CO2) and oxidative Requires complex media pre-warmed to 370C Susceptible to cool temperatures, drying and fatty acids Soluble starch added to neutralize fatty acid toxicity Intracellular human-specific pathogen

a. Neisseria gonorrhoeae
Virulence factors Fimbrae (pili) To adhere to host cells and to each other >100 serotypes known according to pilus protein Has not polysaccharide Capsule Outer membrane proteins (formerly Proteins I, II, & III) Involved in adherence to host cells IgA protease- cleaves IgA on mucosal surfaces IgA blocks the ability of bacteria to adhere IgA protease involved in successful colonization Lipopolysaccharide Prevents phagocytosis

Pathogenicity Pyogenic infection of columnar and transitional epithelial cells Urethral, endocervix, anal canal, pharynx, conjunctiva Venereal (Sexual Transmitted Disease (STD)) Genital infections Gonorrhea Extragenital infections Pharyngitis and Anorectal infections Non-venereal Ophthalmia neonatorum Vulvo-vaginitis in small girls Contaminated toilet seats and contaminated towels

Gonorrhea Gonorrhea transmitted by sexual intercourse
Gonorrhea is the second most common venereal disease Incubation period: 2 to 7 days Females Males 50% risk of infection after single exposure 20% risk of infection after single exposure Asymptomatic infections frequently not diagnosed Most initially symptomatic (95% acute) Major reservoir is asymptomatic carriage Genital infection primary site is cervix (cervicitis), but vagina, urethra, rectum can be colonized Genital infection generally restricted to urethra (urethritis) with purulent discharge and dysuria Ascending infections in 10-20% including salpingitis, tubo-ovarian abscesses, PID, chronic infections can lead to sterility Rare complications may include epididymitis, prostatitis, and periurethral abscesses Disseminated infections more common, including septicemia, infection of skin and joints (1-3%) Bacteremia and Gonococcal arthritis occurs as a result of disseminated gonococcal bacteremia Disseminated infections are very rare Can infect infant at delivery (opthalmia neonatorum) More common in homosexual/bisexual men than in heterosexual population

Ophthalmia Neonatorum
Contamination of infant’s eye during labour through the birth canal of mother In infancy, an eye infection (ophthalmia neonatorum) may occur during vaginal delivery Conjunctivitis with mucopurulent discharge May cause blindness if not treated Infection is preventable with the application of erythromycin eye drops at birth (chlamydia & Neisseria)

Laboratory Diagnosis Clinical specimens Direct microscopy (Gram stain)
Gonnorhea Female Cervical discharge in acute and chronic Cervical swab may give positive results Male Acute: Urethral purulent discharge after urinating Chronic: The morning drop or prostatic massage Ophthalmia neonatorum Mucopurulent discharge Direct microscopy (Gram stain) Small, gram-negative diplococci in presence of PMN’s

Laboratory Diagnosis Culture Biochemical tests
Inoculate specimen on non selective (chocolate agar) or selective media [Thayer-Martin or Modified Thayer-Martin (MTM)] Incubated at 350 C in 5% CO2 The suspected colonies are Gram-negative diplococci with adjacent sides flattened (like coffee beans) Biochemical tests Oxidase + & produce acid from glucose only Fresh growth must be used for testing, because N. gonorrhoeae produces autolytic enzymes

Prevention and control
Penicillin no longer drug of choice due to: Continuing rise in the MIC Beta-lactamase production (some strains) Ceftriaxone, cefixime or fluoroquinolone combined with doxycycline or azithromycin for dual infections with Chlamydia Chemoprophylaxis against gonorrhea is of little value Chemoprophylaxis against ophthalmia neonatorum with 1% silver nitrate 1% tetracycline or 0.5% erythromycin eye ointment Measures to limit epidemic include education, detection, and follow-up screening of sexual partners, use of condoms No vaccine is available

b. Neisseria meningitidis (Meningcoccus)
Encapsulated small, gram-negative diplococci Similar to Neisseria gonorrhoeae The main differences are N. gonorrhea N. meningitidis Portal of entry Genital tract Respiratory tract Polysaccharide capsule Absent Present Beta-lactamase Some None Acid from maltose Negative Positive Vaccine Not available Available

Pathogenicity: Pili-mediated receptor-specific colonization of cells of nasopharynx Antiphagocytic polysaccharide capsule Allows systemic spread in absence of specific immunity 13 serogroups six of which (A, B, C, W135, X, Y) can cause epidemics Serogroups A, B, C, Y, W135 account for 90% of all infections Serotypes A, B and C are the most common worldwide Serotype A is common in epidemics in Africa Toxic effects mediated by hyperproduction of LPS (endotoxin) Other virulence factors as in N. gonnorhea

Clinical Infections Meningitis (30-50%)
Meningitis with meningococcemia (40%), or Meningococcemia without obvious meningitis (7-10%) Second most common cause of community-acquired meningitis Person-to-person transmission by respiratory droplets Commonly colonize nasopharynx of healthy individuals; highest oral and nasopharyngeal carriage rates in school-age children, young adults and lower socioeconomic groups Requires close contact with infectious person in crowded conditions (e.g. military barracks, prisons, Hajj and other) Requires lack of specific antibody (susceptibility) Incubation period is 2-10 days Symptoms: Fever, headache, stiff neck, nausea, vomiting and purulent meningitis with increased WBCs

Laboratory Diagnosis:
Specimens CSF appears turbid (meningitis) Blood (meningecoocemia) Nasopharyngeal swabs (carrier) Large numbers (e.g. >107cells/ml) of encapsulated, small, gram-negative diplococci and PMN’s can be seen microscopically in CSF Extracellular and intracellular Culture Transparent, non-pigmented nonhemolytic colonies on chocolate, MTM agar with enhanced growth in 5% CO2 Biochemical and Immunologic tests Oxidase-positive Acid production from glucose and maltose Immunologic methods are available for sero-grouping

Prevention and control
Prophylaxis Antibiotic prophylaxis is recommended for certain close contacts Avoid crowdedness Vaccination Several vaccines are available to control the disease They are designed to prevent serogroups A, C, Y, and W-135 They are lacking serogroup B antigen Tetravalent Meningococcal Polysaccharide vaccine (MPSV4) Available since 1981 and used for people >55 years Meningococcal Conjugate Vaccine (MCV) When polysaccharides are conjugated to carrier proteins, the polysaccharide antigens become immunogenic in infants and prime for memory anticapsular antibody responses

Meningococcal A conjugate vaccine
Meningococcal C conjugate vaccines Tetravalent meningococcal conjugate vaccine (MCV4) MCV4 is licensed in 2005 and is available for ages 9 M-55 Y 2 doses of MCV4 are recommended (5 years interval) MPSV4 is taken every 3 years 7–10 days are required for development of immunity No vaccines are available for prevention of group B meningococcal disease, which accounts for>50% of all cases Group B capsule is not a suitable vaccine target Polysaccharide B is poor immunogenic Treatment Penicillin (drug of choice) Other options: rifampin or sulfonamide (Chemoprophylaxis)

Enterobacteriaceae Coliforms (lactose ferementers)
Normal inhabitants of GIT of human and animals Source of noscomial infections Opportunistic or cause secondary infections of wounds, urinary and respiratory tracts and the circulatory system e.g. E. coli, Klebsiella E. coli used as biological indicator in water pollution True pathogens (Lactose non-fermenters) Salmonella spp., Shigella spp., Yersinia spp. Certain strains of E. coli (ETEC, EPEC, EIEC, EHEC)

Media for isolation Selective differential media for enteric pathogens
MacConkey agar EMB agar SS agar Selective by incorporation of dyes and bile salts Differential by incorporation of lactose and/or Fe+3 Fe+3 is incorporated to detect H2S Classified as lactose fermenters & non-lactose fermenters

I- Escherichia coli General characteristics: Gram-negative Motile rods
Non-spore forming,Facultative anaerobic, Oxidase -ve Ferment glucose and lactose Normal flora of intestine Opportunistic pathogens E. coli may be pathogenic inside or outside Intestine Some strains (Pathogenic) cause various forms of gastroenteritis

Virulence factors Adhesions (Colonization factors)
Pili or fimbriae & nonfimbrial factors Host defense Capsule OMPs are involved in helping the organism to invade by helping in attachment and in initiating endocytosis Exotoxin production (Enterotoxin) Heat-Labile (LT) & Heat-Stable Toxin (ST) (ETEC) Shiga-like toxin (Verotoxin) (EHEC) Endotoxin (Pyrogen) Lipid A of LPS causes fever and endotoxic shock

Diseases caused by E. coli
Intestinal: Diarrhea (Toxin and/or adhesion) Enterotoxegenic E. coli (ETEC) Enteropathogenic E. coli (EPEC) Enteroinvasive E. coli (EIEC) Enterohemorrhagic E. coli (EHEC) Enteroaggregative E. coli (EAEC) Extraintestinal: Urinary Tract Infections [UTI] (Pili) Neonatal meningitis (K1 antigen) Sepsis (commonly in debilitated hospitalized patients) Endotoxic shock Due to lipid A (Pyrogen) Fever and sudden hypotension Acquired by ingestion of contaminated food and water

Summary of E. coli gastroenteritis
Symptoms Diseases Invasion Pathogenesis Site M.O. Watery diarrhea, cramps, nausea, low grade fever Traveler's diarrhea Infant diarrhea Non invasive LT + STcAMP + cGMP  Fluid + electrolyte loss Small intestine ETEC dysentery-like diarrhea, severe inflammation, fever Dysentery Invasive nonfimbrial adhesin (NFA): OMP Large intestine EIEC Watery diarrhea With mucous without blood or pus With fever & vomiting Infantile diarrhea Poorly invasive NFA: intimin EPEC adherence factor Some reports of shiga-like toxin EPEC Hemorrhagic colitis with sever abdominal cramps, watery diarrhea followed by blood, no fever HUS Bloody diarrhoea and haemolytic uraemic syndrome Cytotoxic shiga-like toxin (verotoxin) EHEC Vomiting Abdominal pain Without inflammation or fever Aute and persistent diarrhoea in children and adults Production of enterotoxin that similar to ETEC EAEC

Urinary Tract Infection (UTI)
E. coli is the most common organism causing UTI Community acquired 90% Hospital acquired (50%) UTI is the disease of female (Short urethra) Fecal E. coli acquires pili to colonize mucosa of UT Travel up urethra & infect balder (Cystitis)& sometimes move further up to infect kidney (pyelonephritis) Symptoms:urinary frequency, dysuria, hematuria, pyuria

Diagnosis of UTI Specimen MSU (Mid-Stream Urine) Culture
On MacConkey agar Gram negative, Lactose fermented (Pink colonies) Viable count ≥100,000 (105) cfu/ml urine IMViC

Neonatal meningitis E. coli is the second most common cause
S. galactaiae (Group B) is the first Occur during the first month of life Lab diagnosis Specimen: CSF Culture: Pink colonies on MacConkey agar (LF)

II- Salmonella General characteristics:
Gram-negative rods belonging to Enterobacteriaceae Do not ferment lactose and H2S positive Salmonellae live in the intestinal tracts of animals Not considered part of normal intestinal flora in human Always PATHOGENIC to human

Classification of Salmonella
Kaufman-White- Le Minor Classification Based on O and H antigen serotyping 64 O and 114 H variants identified Classified into 9 groups (A-I) according to O antigen Each group can be classified into subgroups according to H Ag Salmonella can be detected by its group O antigen and then by its type specific H antigen >2500 known serovars

Classification of Salmonella
US CDC (Center for Disease Control) S. enterica Subdivided into 6 subspecies enterica, salamae, arizonae, diarizonae, indica, houtanae Of these six subspecies, only subspecies enterica is associated with disease in warm-blooded animals S. enterica subsp. enterica serovar Typhi or S. Typhi S. enterica subsp. enterica ser.Typhimurium or S. Typhimurium S. enterica subsp. enterica ser. Enteritidis or S. Enteritidis Salmonella bongori (Subspecies V)

Species of Salmonella Two important members of Salmonella causing diseases: Salmonella causing enteric fever Salmonella Typhi or Salmonella Paratyphi A,B, and C These organisms penetrate intestinal mucosa Detected in blood, urine and stool Salmonella causing food poisoning Salmonella Enteritidis and Salmonella Typhimurium These organisms do not penetrate intestinal mucosa Detected in stool only

Enteric Fever “Typhoid”
Enteric fevers are severe systemic forms of salmonellosis. Caused by S. typhi whereas a milder form “Paratyphoid” caused by S. paratyphi A, B or C Mode of Transmission Via fecal-oral route through fecally-contaminated food or water from either chronic carrier or case A few individuals continue to harbour Salmonella in their gall-bladders and intermittently excrete organisms in their faces Temporary excretors: Patients who excrete bacteria for <year Chronic carrier: Patients who excrete bacteria for > year Incubation Period 10-14 day during which bacteria multiplies in Peyer’s patches Pass to blood via lymphatics resulting in bacteriaemia in 1st week In 2nd week M.O. passes to different organs including peyer’s patches causing ulcers, gall bladder, liver, kidney & rarely menings

Symptoms of Enteric Fever
Symptoms begin after an incubation period of 2 weeks Enteric fevers may be preceded by gastroenteritis, which usually resolves before the onset of systemic disease The symptoms of enteric fevers are nonspecific and include Fever, headache, delirium (sustained fever), malaise and tender abdomen Complications include intestinal hemorrhage and perforation

Laboratory diagnosis Direct diagnosis:
Specimen: Blood during 1st week, urine during 2nd week and stool during 3rd week Isolation of microorganism: From blood using blood culture Five to 10 ml of blood is taken during the 1st week of infection, Add to ml sterile nutrient broth and incubate at 370C for 24 hrs. Subculture is done on MacConkey's agar which shows colorless colonies in positive case

Laboratory diagnosis From stool
By culture on enrichment medium such as selenite F or tetrathionate broth which inhibits the growth of coliform and allow the growth of Salmonella and Shigella Subculture on MacConkey, SS or DCA agar On MacConkey's agar they give colorless colonies On SS agar they give colorless colonies with black edges due to H2S production Biochemical Reactions: The suspected colonies were subjected to biochemical reactions Oxidase negative, not ferment lactose and sucrose, H2S positive

Laboratory diagnosis Indirect: Serological diagnosis (Widal Test):
In the 2nd week of the disease, antibodies against Salmonella are present in the patient's serum and can be detected serologically by Widal test (agglutination test) Widal test is positive and valid during 2nd week Serial dilutions of patient's serum are added to an equal volume of common O and specific H antigens Agglutination of O- antigen and one only of the H-antigens at a titer 1/80 or above is diagnostic

Salmonella causing food poisoning
The causative agent : S. typhimurium & S. enteritidis Mode of infection: Consumption of contaminated food Food as cakes, pastries and various milk and egg dishes Cattle, sheep, hens, ducks and turkeys are often infected and the organism may contaminate meat and meat products Infective dose 100,000 bacteria Gastric juice is the an important host defense and decreased acidity is a predisposing factor Incubation period is hrs and recovery within 4-7 days It is self limiting disease Manifestations include Nausea, vomiting, and abdominal discomfort, non-bloody diarrhea and slight fever

I- Shigella General Characteristics Gram negative rods Non motile
Non spore-forming Non capsulated Oxidase negative Ferment glucose with acid only Non lactose fermentating organism H2S negative

Shigella species Disease: Bacillary dysentery (Shigellosis)
S. sonnei is the most common, followed by S. flexneri Mode of Transmission Oral-fecal transmission <200 bacilli are needed for infection in health individuals Incubation periods It varies between 1-3 days Symptoms Ranges from asymptomatic to severe bacillary dysentery Watery diarrhea changing to dysentery with frequent small stools with blood, pus and mucus Fever, tenesmus and abdominal cramps

Stages of shigellosis Early stage:
Ingestion of contaminated food or water Noninvasive colonization and cell multiplication Production of the enterotoxin in the small intestine Watery diarrhea attributed to enterotoxic activity of Shiga toxin (similar to LT of ETEC) Fever attributed to neurotoxic activity of toxin Second stage: Adherence to and tissue invasion of large intestine Typical symptoms of dysentery Cytotoxic activity of Shiga toxin increases severity

Laboratory identification
Specimen: Stool (mucous bloody part of stool) or rectal swap Stool culture Specimen is inoculated in Selenite broth at 370C for 24 h Then subculture on MacConkey or SS On MacConkey agar: they give colorless colonies On SS agar: they give colorless colonies Gram stain Gram negative bacilli, NON MOTILE Biochemical reactions Oxidase negative, ferment glucose, non ferment lactose and H2S NEGATIVE

COMMENSAL ENTEROBACTERIAE
Opportunistic pathogens and are common cause of nosocomial infections Klebsiella Enterobacter Serratia Proteus Morganella Providencia Lactose Fermnters Non-Lactose Fermnters

Klebsiella-Enterobacter-Serratia group
General characteristics: These organisms are very similar All are motile except Klebsiella MR negative; VP positive Simmons citrate positive H2S negative Some weakly urease positive Phenylalanine deaminase negative These organisms belonging to Enterobacteriaceae Frequently in large intestine but also found in soil and water Usually opportunistic pathogens Wide variety of infections: primarily pneumonia, wound and UTI

Klebsiella pneumoniae
Virulence factors Polysaccharide capsule Protects against phagocytosis and antibiotics Makes the colonies moist and mucoid Adhesions Clinical significance Major cause of nosocomial infections Nosocomial Pneumonia Important respiratory tract pathogen outside hospitals 3% of bacterial pneumonia Bloody sputum (50%) (Thick, Jelly and Red Sputum) Septicemia, Meningitis and UTI

Proteus-Providencia-Morganella
These organisms belonging to Enterobacteriaceae All are normal intestinal flora but also found in soil and water Opportunistic pathogens Non-lactose fermenters Phenylalanine deaminase and urease positive Urease positive after 2-6 hrs (urea → NH3+ CO2) All are highly motile Proteus sp. swarming on blood agar Swarming characterized by expanding waves (rings) of organisms over the surface of blood agar

Proteus species P. mirabilis and P. vulgaris are widely recognized human pathogens Isolated from urine, wounds and ear and bacteremic infections They found in colon and able to colonize urethra especially in female P. mirabilis causes urinary tract infections (UTI): Proteus sp. produces urease which alkalinizes urine →precipitation of calcium and magnesium salts → stone formation →renal epithelium damage P. vulgaris causes nosocomial infections (pneumonia, bacteremia) and UTIs

Laboratory diagnosis Specimen: Culture:
Urine or Stool. Culture: On MacConkey agar →non lactose fermenters→ colorless colonies On SS agar →non lactose fermenters → colorless colonies with black center On ordinary media, such as nutrient agar, blood agar, show swarming (successive waves on the surface) due to high motility of Proteus Biochemical reactions: Urease , phenyldeaminase and H2S positive

Pseudomonas aeruginosa
Gram-negative bacilli belonging to Pseudomonadaceae Motile by single polar flagellum, non spore Non fastidious- Minimal nutritional requirements Produces grape-like odor and blue-green pus and colonies Two types of soluble pigments: Pyoverdin : Yellow-green pigment & fluorescent Pyocyanin: Blue-green pigment and non-fluorescent Strict aerobic, O+/F-, Oxidase and catalase positive Optimum temperature is 37 0C & able to grow at 42 0C Resistant to dyes, weak antiseptics and many antibiotics

Epidemiology The most important pathogenic is Ps. aeruginosa
Opportunistic pathogen Important agent in causing nosocomial infections Ubiquitous in moist environment, hospital, in soil and water Colonized in the intestine in10% of human Represents 10-20% of hospital-acquired infections Transiently colonize respiratory & GIT of hospitalized patients: Treated with broad-spectrum antibiotics Exposed to respiratory therapy equipment Hospitalized for long periods

Pathogenesis 1. Adhesins Invasins Proteases - Inactivate IFN and TNF
Fimbriae, Pilli, Alginate (mucoid exo-polysaccharide) Alginate slim forms biofilm that protect from antibodies, complement, phagocytosis & antibiotic Invasins Proteases - Inactivate IFN and TNF Elastase Break down of elastin-containing tissues e.g. blood vessels, lung tissue, skin Cleaves collagen, IgG, IgA and complement) Produce hemorrhagic lesions associated with disseminated infection Alkaline protease (lyses fibrin) Hemolysins (phospholipase & lecithinase) & Leukocidin

Pathogenesis Toxins Exotoxin A & S
Inhibit protein synthesis (Like diphtheria toxin) Immunosuppressive Causes dermatonecrosis in burn wounds, corneal damage in ocular infections, and tissue damage in chronic pulmonary infections Pyocyanin – inhibits mitochondrial enzymes, impairs ciliary function, mediates tissue damage LPS (endotoxin)

Clinical Finding UTIs (Catheterized patients)
Wound, burn & other skin & soft tissues infections Echtyma Gangrenosum Pneumonia (Cystic Fibrosis) Eye infection- Contaminated contact lens clearing fluids Chronic otitis media & otitis externa (swimming pool) Meningitis- following lumbar puncture Systemic infections- Septicemia in debilitated patients Pseudomonal Endocarditis

Echtyma Gangrenosum Causative agent: Ps. aeruginosa
Under risk: immunocompromised, burn patients, and other critically ill patients Round or oval (1-15cm) severe invasive cutaneous ulceritic single or multiple lesion with halo of erthryma Pseudomonas exotoxin: Tissue destruction Elastase degrades elastin in blood vessels wall Phospholipase C degrades phospholipids in cell membranes Pyocyanin generates reactive oxygen species

Typing methods Bacteriocin typing
Three types of bacteriocins are produced-R,F,S Pyocin produced by test strain is employed to assess the growth inhibition of 13 indicators strains Depending upon the growth inhibition of 13 indicators strains,105 types are recognized Most popular method used Phage typing Serotyping-based on O & H,17 serotypes are recognized

Laboratory diagnosis Specimen
Urine, pus, sputum, CSF, blood, skin swap Microscopical Examination Gram-negative rods and motile Cultural Characteristics On Nutrient agar: Colonies are surrounded by bluish green coloration On selective media "Cetermide" Pigments are more obvious On Blood agar: -hemolytic colonies On MacConkey agar Pale yellow colonies i.e. non lactose fermenters Ps. aeruginosa able to grow at 42 0C for 3 days Pyocin typing: Identification for epidemiological purpose

Vibrionaceae General characteristics
Gram negative, curved, comma shaped bacilli Motile by single polar flagella Non spore, Non capsulated Facultative anaerobes, Fermentative Oxidase and catalase positive Most vibrios have relatively simple growth factor requirements and grow well in alkaline pH Natural inhabitants : Aquatic environment

Vibrio cholerae >200 serogroups based on O-antigen
V. cholerae O1 (classical and El Tor biotypes) and O139 are primarily responsible for cholera outbreaks V. El Tor biotype causes cholera-like but milder Recently, V. cholerae O75 and O141 strains has been associated with cholera-like diarrhea Some O1 strains do not produce cholera enterotoxin (atypical or nontoxigenic O1 V. cholerae) Cholera is endemic in southern Asia (India, Pakistan and Bangladesh), Latin America

Pathogenesis Mode of Transmission
Consumption of contaminated water or food Incubation period:1-4 days High infectious dose: >108 CFU V. cholerae attach to mucosa of small intestine V. cholerae multiply & secrete enterotoxin (cholargen) Toxin binds to specific receptor on the intestinal mucosal cell Toxin stimulates activity of cAMP, resulting in active secretion of chloride & secondary loss of Na and H2O The patient loss 20 Litre fluids/day V. cholerae cause same disease as ETEC but more severe

Diagnosis of V. cholerae
Specimen: Rice watery stool or rectal swab Culture on Alkaline Peptone Water (APW) then subculture on selective differential medium Thiosulfate Citrate Bile Salts Sucrose agar Selective due to alkalinity pH9 & contains bile salts Differential because contains sucrose Sucrose fermenting V. cholerae yellow colonies Sucrose non fermenting V. parahemolyticus appears as blue to green colonies Sucrose fermentation is gold standard in identification

Diagnosis of V. cholerae
Any sucrose fermenting colonies were subjected to Gram stain and oxidase test Gram stain Gram negative short rods, comma shaped, motile Biochemical reactions: Oxidase positive O+/F+ Cholera red reaction Serology: Diagnosis confirmed & serotyping done by agglutination with specific antisera (O1, O139)

V. parahaemolyticus It is the cause of acute gastroenteritis following ingestion of contaminated sea-food such as raw fish Marine organism which live in high salt conc. So it requires 2% Na Cl in the isolation medium It is invasive affecting the colon V. cholerae is noninvasive, affecting the small intestine through secretion of an enterotoxin Allied Vibrios are large group of organisms; some are saprophytic while others cause disease in animals

Campylobacter Campylobacter, belong to Campylobacteriaceae,
It was formerly grouped with vibrios why? Gram-negative curved, S-shaped or spiral short bacilli Motile by single polar flagella (Cork screw) Non-spore forming ,Microaerophilic, Oxidase and catalase positive 15 species; 6 subspecies 12 associated with human infection Divided into 2 groups on the basis of growth temperature: Thermotolerant species which grow at 420C C. jejuni (poultry; humans) C. coli (porcine; human) C. jejuni accounts for >80% of all Campylobacter infection Non-thermotolerant - grow at 250C (C. fetus; veterinary pathogen) Primarily zoonotic with variety of animals (reservoirs) for infection Also, food animal such as poultry, cattle, sheep and pigs

Campylobacter Campylobacter is the main causative agent of diarrhoea (enteritis ), mainly children, in developing countries C. jejuni, E. coli and Rotavirus are the three most common cause of infantile diarrhoea in the world Infections usually sporadic, occurring in summer Mode of transmission Acquired by the fecal-oral route via ingestion of; Improperly handled or cooked food primarily poultry Also, drinking of un-pasteurized milk Children is the most commonly affected worldwide Incubation period The illness appears to last from 2-7 days

Pathogenesis and Virulence factor
C. jejuni infection vary considerably from asymptomatic to severe bloody diarrhea, high fever, and prostration Enterocolitis begins as watery, foul smelling diarrhoea followed by bloody stools accompanied by fever, headache and sever abdominal cramps Systemic infection: bacteremia (very rare) Rarely C. jejuni spreads systematically (bacteremia) This organism multiply in the small intestine, invade the epithelium and produce gastroenteritis Then C. jejuni secretes Enterotoxin similar to cholera toxin and LT of E. coli Cytotoxin that destroy mucosal cell

Diagnosis of C. jejuni Specimen Stool culture is done on:
Selective medium (Skirrow’s medium, Blood agar + cefoperazone, vancomycin, amphotericin B) Incubation temperature at 420C in microaerophilic condition (5%CO2, 10%O2 & 85%N2) for hrs Gram-stain Gram stain examination of the colony should be performed along with oxidase test Oxidase +ve colony exhibiting characteristic Gram stain appearance can be reported as Campylobacter

Diagnosis & Treatment Special test for C. jejuni
Hippurate hydrolysis is the major test for distinguishing between C. jejuni (positive) and other Campylobacter spp. Treatment Campylobacter jejuni isolates have variable susceptibilities to a variety of antimicrobials Azithromycin and erythromycin (Macrolides) are the drug of choice

4. Helicobacter Helicobacter is closely resemble to Campylobacter
Gram-negative spiral bacilli Non-spore forming Motile by multiple polar flagella Microaerophilic Grow at 370C and slow growing organism (7-10 days) Requires enriched media with blood, hemin Such supplements protects the organism from oxygen free radicals, H2O2 and fatty acids present in the media Oxidase and catalase positive Helicobacter is urease +ve while Campylobacter -ve It survives in acidic environment of stomach & duodenum It hides in mucus & neutralizes acid in its local environment

Diseases Associated with H. pylori
Gastritis (irritation & inflammation of lining of stomach) Symptoms include nausea, vomiting and frequent complaints about pain in the abdomen Peptic and duodenal ulcer Sores that form in the stomach or the duodenum The common symptom is burning pain in abdomen Ulcers that bleed, causing hematemesis (bloody vomit or vomit that looks like coffee grounds) or melena (stool that's black, bloody or looks like tar) Stomach cancer later in life No bacteremia or disseminated diseases occurs

Mode of transmission Scientists suspect that H. pylori infection may be contagious The infection seems to run in families More common in crowded or unsanitary conditions Infection may be passed from person to person?? How spread happens isn't really known Transmission believed to be by fecal-oral route

Pathogenesis Virulence Factors
Most bacteria killed in environment of gastric lumen H. pylori proliferates in mucus layer by aid of flagella Then reach epithelial cells underneath- more neutral pH H. pylori survives because of virulence factors that contribute to gastric inflammation, alter gastric acid production and cause tissue destruction Virulence Factors Initial colonization facilitated by: Acid inhibitory protein - blocks acid secretion from parietal cells during acute infection Urease - neutralizes gastric acids due to NH3 production Urease stimulates monocytes and neutrophil chemotaxis; stimulates production of inflammatory cytokines

Heat shock protein - enhances urease expression; co-expressed with urease on bacterial surface
Flagella - allows penetration into gastric mucous layer Adhesins - mediate binding to host cells Localized tissue damage mediated by: Mucinases and phospholipases - disrupt gastric mucus Vacuolating cytotoxin - induces vacuolation in epithelial cells that results in epithelial cell damage SOD and catalase - prevent from phagocytosis and intracellular killing

Diagnosis H. pylori is diagnosed by non-culture method such as
Blood antibody tests Urea breath tests (UBT) Endoscopic biopsies Stool antigen tests H. pylori isolated by culture of gastric biopsy on non-selective media enriched with blood or serum for 7-10 day Skirrow’s media supplemented with blood or serum

Diagnosis Blood antibody test
Blood tests for the presence of antibodies to H. pylori can be performed easily and rapidly Urea Breath Test (UBT) UBT is a safe, easy & accurate test for detection of H. pylori UBT relies on ability of H. pylori to hydrolyse urea into CO2 CO2 is absorbed from stomach & eliminated in the breath 10-20 minute after swallowing a capsule containing a minute amount of radioactive urea, a breath is collected The presence of radioactive CO2 in the breath (a positive test) means that there is active infection

Test kit showing negative and positive results for Helicobacter
Endoscopic biopsies Endoscopy is an accurate test for diagnosing H. pylori as well as the inflammation and ulcers that it causes For endoscopy, the doctor inserts a flexible viewing tube (endoscope) through the mouth, down the oesophagus, and into the stomach and duodenum During endoscopy, small tissue samples (biopsies) from the stomach lining can be removed A biopsy is placed on a special slide containing urea If the urea is broken down, this means that there is an infection with H. pylori Test kit showing negative and positive results for Helicobacter

Haemophilus General characteristics: Belonging to the Pasteurellaceae
Non intestinal pleomorphic Gram negative coccobacilli Microaerophilic Non motile, Non spore forming Usually capsulated Fastidious Dependent on the growth promoting substances X (haematin) and V (NAD) which present in blood Oxidase and catalase positive

Differentiation of Species
Hemolysis Growth Factor X V H. influenzae - + H. aegyptius H. ducreyi H. parainfluenzae

H. influenzae Opportunistic pathogen Exclusively human pathogen
Not pathogenic for animals Diseases due to H. influenzae considered under 2 groups –invasive and non invasive Nonencapsulated strains (Non invasive) Part of normal flora of the respiratory tract Colonize in nasopharynx of 80% of healthy individuals Causes pneumonia, otitis media and sinusitis

H. influenzae Encapsulated strains (Invasive)
Typing based on capsule polysaccharide a → f Type b is the most pathogenic & colonized in 5% Polyribose-ribitol phosphate (PRP) capsule (type b) Penetrate nasopharynx and invade blood directly Causes meningitis, pneumonia, bacteremia 95% of invasive disease caused by type b Vaccination with Hib conjugate vaccine Given IM at 2,4,6 month & poster dose at month

Virulence factors of H. influenzae
Polysaccharide capsule Antiphagocytic and major pathogenesis factor Fimbriae the adherence to human mucosal cells LPS- lipid A -major role in non capsule strains All virulent strains produce neuraminidase and IgA protease Many respiratory pathogens, including H. influenzae, S. pneumoniae and P. aeruginosa, express neuraminidases Bacterial neuraminidase facilitates mucosal infection by participating in biofilm production No exotoxins PBP- Poly ribitol phospate capsule

H. aegypticus Cause of acute mucopurulent conjunctivitis (Pinkeye)
The disease is common in Egypt It is spread very easily, especially among children Pinkeye is transmitted mechanically by common towels or by flies Diagnosis is made from the conjunctival discharge

H. ducreyi This is the cause of chancroid (Soft chancre) which is venereal disease Chancroid is characterized by painful genital ulcers This organism needs only X factor Diagnosis is made from the discharge of the ulcer

Diagnosis of Haemophilus
Specimen: CSF, blood, sputum Diagnosis is confirmed when the organism is isolated from a sterile body site (Blood & CSF) Diagnosis is not confirmed when it is isolated from nasopharyx or sputum Stain: Gram negative coccobacilli Culture: On chocolate agar: A 24 h colony of H. influenzae is larger than that observed on blood agar On nutrient agar plate with added X & V factors at 37°C in an enriched CO2 incubator

Diagnosis of Haemophilus
Satellitism: Blood agar contains much X factor and little V factor S. aureus produces V factor A lawn of test bacteria is plated onto a blood agar plate S. aureus is placed on plate & the culture is incubated H. influenzae will grow in the hemolytic zone of S. aureus The hemolysis of cells by S. aureus releases nutrients H. influenzae will not grow outside the hemolytic zone of S. aureus due to the lack of nutrients in these areas Capsule swelling: Specific antiserum added to a slide of the organism allows swelling of the bacterial capsule thus permitting rapid diagnosis of H. influenzae in sputum

Bordetella Small capsulated Gram negative coccobacillus
Strict aerobe, oxidative, oxidase variable? Three important species B. pertussis causes WHOOPING COUGH (Pertussis) Strictly human pathogen: infant & young children B. parapertussis–causes mild form of whooping cough B. bronchoseptica Widespread in animals (dog) & causes kennel cough Rarely causes respiratory or wound infection in humans

B. pertussis Pathogenesis
Respiratory Colonization Attaches to ciliated epithelium of URT VIA Filamentous hemagglutinin (FHA) Protein on pili Antibodies against FHA inhibit colonization Fimbriae are NOT involved Pertussis Toxin Do not invade underlying lung tissues Decreased cilia activity & epithelial cell death occur 7-10 days, NO symptoms Positive cultures toward the end of this stage

Toxin-mediated disease
Pertussis toxin Responsible for adherence ↑Adenylate cyclase ↑ intracellular cAMP This causes cellular dysfunction Inhibition of host phagocyte oxidative responses and the inhibition of natural killer cell activity Dermonecrotic toxin (lethal toxin) Released upon cell lysis causing strong vasoconstriction Results in O2  convulsion Tracheal cytotoxin Prevents ciliated epithelial cells from beating LPS –endotoxin-irritation and damage of cell

Clinical significance
Pertussis is Toxin-mediated disease Very contagious & can cause serious illness―especially in babies too young MODE OF TRANSMISSION: droplet inhalation INCUBATION PERIOD:1-2 weeks Organism multiply & start to liberate toxins Pertussis lasts for 6 weeks and appear in 4 stages CATARRHAL STAGE occurs - rhinorrhea, mild cough, sneezing & fever whereby a lot of organisms are spread through respiratory secretions This last ~ 1-2 weeks

PAROXYSMAL STAGE that lasts 2-4 weeks
Patient has rapid, consecutive cough with a rapid intake of air between coughs (has a whooping sound) Ciliary action of RT has been compromised Mucous has accumulated Patient is trying to cough up mucous accumulations Coughs are strong enough to break ribs! Finally there is a CONVALESCENT STAGE during which symptoms gradually subside This can last for months

Diagnosis Specimen is taken during PAROXYSMAL STAGE
Nasopharyngeal swabs Cough plate method Highly fastidious & requires special media+additional nutrients for growth+absorbents to remove toxic substances found in complex media e.g. fatty acids Culture on Bordet-Gengou medium Contains glycerol, potato infusion, albumin (binds fatty acids), and up to 50% defibrinated SRBCs Charcoal agar +10% horse blood ± Cephalexin 3-7 days for growth, hemolytic colonies A characteristic pearl like colonies are observed Direct fluorescent antibody testing, PCR, Slide agglutination . The bacteria is slow growing and an incubation is done for days. After 4 weeks of infection cultures are generally negative. Direct immunofluorescence assay of nasopharyngeal secretions have variable sensitivity and low specificity.

Differentiation of Bordetella species Sheep blood agar and MacConkey
Growth on Urease Oxidase? Motility Sheep blood agar and MacConkey Bordet-Gengou agar B. pertussis - + B. parapertussis B. bronchiseptica

Prevention Pertussis Whole cell vaccine
Merthiolate-killed bacterial cell suspension Part of the DTP vaccine P in DTP for Pertussis Pertussis Acellular vaccines Consist of 1–5 purified, detoxified toxins & adhesins Introduced in developed regions ∼15 years ago to replace Killed vaccines Has fewer side effects than the whole cell vaccine Diphtheria-Tetanus-Pertussis vaccine using acellular pertussis is known as DTaP 2, 4, 6, months and 4-6 years of age.

Legionellae Gram negative rod belonging to Legionellaceae
Stains poorly with gram stain Facultative intracellular pathogen > 57 species, ½ of species implicated in human disease L. pneumophila is the most important human pathogen Recently became opportunistic human pathogen ~ 90% of all cases of legionellosis (Legionnaires' disease) 15 serogroups, 88.6% of cases are caused by serogroup 1 Serogroup 1 seems to be more virulent Not grow on Sheep Blood Agar Requires specialized media to grow Cysteine and iron are essential for growth

L. pneumophila Reservoir principally aquatic
L. pneumophila is not a free-living organism but an intracellular parasite of amoebae Natural environment Lakes and rivers Artificial environment Showers, taps, air conditioning system Normally found in hot water (up to 500C) tanks Multiply in pipes in sediment & live many years Sediment provides shelter and nutrition for other bacteria that can supply cysteine Important to detect L. pneumophila serogroup1 in water

Legionnaires disease The portal of entry is the respiratory tract via inhalation Legionellae are associated with water-based aerosols Air conditioning cooling towers Sauna Person to person spread does not occur Nosocomial infections Due to presence of bacteria in water taps, sinks & showers No person to person spread

Legionnaires disease In general, healthy people rarely get disease
People under Risk People with defect in cell mediated immunity Typically occurred in older man who smokes and consumes substantial amount of alcohol Patients with AIDS, cancers, Renal transplants, patient treated with corticosteriods or elderly No person to person spread

L. pneumophila Initial host response: acute inflammatory response of aveoli & then bronchioles (similar to pneumococcal infection) Neutrophils accumulate followed by macrophage Different from pneumococcal infection Organisms located inside of macrophage Inhibit lysosomal fusion & acidification of phagocyte

Clinical Presentations
Pontiac fever Pontiac, Michigan in 1968 Incubation period 1-2 days Flu-like, Milder (no mortality) and self-limiting Persists for 2-5 days, then spontaneously resolves Legionnaire's disease Incubation period 2-10 days Atypical pneumonia Most cases resolve spontaneously in 7-10 days but In older or immunocomprmised may be fatal 15-75% mortality

Laboratory Diagnosis of Legionella
Specimen: sputum and blood Culture: on Buffered Charcoal Yeast Extract Agar (BCYE) Grow after 3-5 days Appear as small colonies with ground glass appearance Microscopy -Difficult because of Lack of staining Intracellular nature Require large number of organisms to detect Detection of antigen in respiratory secretion or urine Seroconversion: ≥4 fold rise in specific serum antibody titer Direct fluorescent antibody (DFA) staining

Yersinia, Francisella and Brucella
Small Gram-negative rods Zoontic diseases True pathogens: isolation always associated with disease; i.e., always clinically significant NOTE: Previously studied nonfermenters were all opportunistic pathogens Very virulent and able to penetrate any body area they touch Skin through insect bite, animal bite or direct contact with animals Lung after inhalation of infected aerosolized matter Non-fermentative Facultative intracellular pathogens Common treatment: Aminoglycosides and/or Doxycycline which must be given for prolonged period so as to reach the hidden intracellular bacteria

1. Brucella Brucella are small gram-negative coccobacilli, strict aerobic, non-fermentative lacking a capsule, flagella, endospores Oxidase and catalase are positive Some species require 5-10% CO2 (B. abortus ) for primary isolation in Lab. Requires specialized media and prolonged incubation for growth in culture Facultative Intracellular pathogen Brucella spp. is the causative agent of brucellosis Brucellosis is a zoonotic infection transmitted to humans The disease is rarely, if ever, transmitted between humans Brucellosis also called Undulant fever, Malta fever, Mediterranean fever

Human Brucellosis & Associated Species
Severe

Epidemiology Brucellosis occurs worldwide; major endemic areas include countries of the Mediterranean basin, Arabian Gulf, Indian subcontinent and Mexico, Central and South America Major in Saudi Arabia especially Aseer B. melitensis is the species that infects humans most frequently The incubation period ranges from a few days to a few months The disease is manifested as fever accompanied by a wide array of other symptoms

Human brucellosis is mostly found among farmers, Vet and others who work with infected animals
The organism enters the body through Ingestion of contaminated food such as raw milk, cheese made from unpasteurized milk or raw meat Direct inoculation through skin abrasions from handling animal carcass, placenta or contact with animal vaginal secretions Inhalation of infectious aerosols Localize on reticule-endothelia system (Lymph nodes, liver, spleen, bone marrow) Many organisms are killed by macrophage but some survive where they are protected from antibodies

Clinical Signs of Human Brucellosis
Multi-systemic disease with a broad spectrum of symptoms Human brucellosis is manifested by intermittent fever Patient has 3-4 weeks fever and then 3-4 weeks without fever Incubation Period:1-6 weeks Asymptomatic infections are common The disease is extremely variable and the clinical signs may appear abruptly in symptomatic cases After incubation period, an acute febrile illness with non-specific flu-like signs occur Fever, malaise, chills, night sweats, fatigue, weakness, myalgias, weight loss, arthralgias and nonproductive cough Splenomegaly, hepatomegaly and chest pain Anorexia, nausea, vomiting, diarrhea and constipation

Clinical Signs of Human Brucellosis
In many patients, the symptoms last for 2-4 weeks and are followed by spontaneous recovery Others develop an intermittent fever Most with this undulant (Rising and falling) form recover completely in 3-12 Months A few become chronically ill Relapses (10%) can occur months after the initial symptoms Chronic disease and recurrence are common. Why? Because it can survive in phagocytes & multiply to high concentrations Complications are seen occasionally, particularly in the undulant and chronic forms The most common complications are arthritis, spondylitis, epididymo-orchitis and chronic fatigue

Brucellosis in Animals
Brucella cause abortion in animals Brucella infects organs rich in erythritol Breast, uterus, placenta and epididymis Does Brucella cause abortion in humans? Brucella does not cause abortion in Human. Why? Because of absence of erythritol in human placenta and fetus Asymptomatic carriage, sterility or abortions Transmitted between animals in aborted tissues

Laboratory Diagnosis A. Culture
When brucellosis is suspected, Lab. should be informed to maintain cultures for minimum of 4 weeks Blood culture is done into liver infusion broth during febrile attack Inoculate 2 tubes, one of them in 10-20% CO2 and the other in the normal atmospheric conditions Rate of isolation from blood ranges from 15-70% depending on methods used and incubation period Bone marrow or lymph node cultures can be done if the blood culture is negative Bone marrow cultures have higher yield than blood Most Labs. use rapid isolation techniques, (e.g. BACTEC) PCR used for rapid diagnosis of Brucella in blood specimens

Laboratory Diagnosis Serological Tests
Most serological studies for diagnosis of Brucellosis are based on antibody detection Antibodies appear in the patient’s serum 7-10 days after the onset of clinical features These include: Serum agglutination (Standard tube agglutination) ELISA Complement fixation Indirect Coombs

Serum agglutination This test is insensitive and should not be relied on for diagnosis A titer of >1:160 supports the diagnosis of brucellosis A fourfold increase in agglutinating antibodies over 4 to 12 weeks provides even stronger evidence for the diagnosis A titer of 1:100 may be met in normal people Due to previous subclinical infection False-negative reactions Result from prozone phenomenon (High serum concentration) False-positive reactions Result from cross-reaction with antibodies to Yersinia, Cholera and Tularemia These can be avoidable by routinely diluting serum beyond 1:320 Must be done using wide range of dilutions of patient’s serum In certain conditions antibodies are blocked giving no visible agglutination, so Coomb’s test must be done

Brucellosis can be eradicated from animals by testing them and slaughtering the positive Immunization of animals with a live attenuated vaccine Pasteurization of milk and dairy products Use of proper safety techniques in clinical laboratory working with Brucella

Treatment Intracellular localization of brucellae, believed to offer some protection against antimicrobials, thus drugs with good intra-cellular penetration are necessary for cure Tetracyclines among most active drugs for treating brucellosis Relapse rate unacceptably high with single-drug therapy Combination therapy is recommended There are two major regimens: Regimen A: Doxycycline 100 mg orally twice daily for 6 weeks + Streptomycin 1 gm IM once daily for the first days Regimen B: Doxycycline 100 mg orally twice daily plus rifampin mg (15 mg/kg) orally once daily for 6 weeks

Yersinia Species Yersinia formerly classified in the Pasteurellaceae family So Yersinia formerly called Pasteurella Yersinia reclassified in the Enterobacteriaceae family Three species of Yersinia cause disease in humans Y. pseudotuberculosis Y. enterocolitica Both are animal pathogens Y. pseudotuberculosis causes pseudotuberculosis in animals Y. enterocolitica sometimes infects man causing Yersiniosis Fever, diarrhea, abdominal pain and arthritis Yersiniosis is usually self-limiting and does not require treatment Both are enteric food and water borne pathogens Acquired by ingestion of contaminated food Y. pestis

Y. pestis (formerly Pasteurella pestis)
Discovered in 1894 in Hong Kong by Alexandre Yersin Gram negative short bacilli (coccobacilli) Exhibit bipolar staining (special stain e.g. Giemsa stain) Dark blue bioplar and the remaining appears light blue Facultative anaerobes, Fermentative, non-motile, non-spore forming Oxidase negative (Pasteuralla and Brucella are oxidase positive) Have capsule when freshly isolates Loss of their capsule when passed in laboratory Found in low frequency in wild rodent populations (rat) Rat flea is prime transmitter of disease Disease: Plague (Black death) Disease has an extremely important place in human history

Virulence Factors Pathogenic Y. pestis produce two antiphagocytic components; Fraction 1 (F1) capsular antigen and the VW antigens Both are required for virulence They are only produced when the organism grows at 37°C They not expressed at temperatures < 37°C Yersinia is not virulent in fleas Why? Because their body temperature normally levels around 25°C The bacteria are capable of surviving and multiplying within monocytes, but not PMNs, and upon emerging from the monocytic host, the bacteria possess their F1 and VW antigens

Transmission Cycles The transmission cycle of Y. pestis is vector-dependent, and is usually dependent on X. cheopis, the rat flea (Gage and Kosoy, 2005). Bubonic plague is endemic in a number of rodent populations throughout the world, but it can go through epidemic stages as well. If Y. pestis reaches a new, naïve population, it can explode through that population, creating massive die-offs before being extinguished or becoming endemic again. These epidemic stages, when fleas infected with the bacteria are looking for new hosts, are the times when humans are most likely to be infected. Once humans are infected, further human-flea-human transmission may occur, or pneumonic plague may be transmitted via droplet transmission (Gage and Kosoy, Bichat guidelines). This unique set of transmission cycles, both epidemic and endemic have allowed a unique evolutionary pattern within the Y. pestis genome, which is reflected in the genetic data. Initially the ancestral genotype is dominant, spreading quickly and very rapidly, covering a large amount of geographic area with the same strain of bacteria (Girard, et al. 2004). Once the bacteria become endemic in a population, however, the genotypes begin to diverge and to form local lineages. While these local lineage groups generally do not have any impact on the virulence of the genes, it allows the study of the evolution of the Y. pestis genome. While this study was done in Gunnison’s prairie dog populations, it also is an excellent model for the historical epidemics of plague in the 14th-17th C in Europe. A historical characteristic of the plague in Europe was the rapid spread of the disease between centers of commerce and trade followed by a period in which the disease became endemic in the population centers. Occasionally these endemic varieties would become more virulent again, and the rapid spread of the disease would repeat, which is seen in the evolutionary picture of genomic alterations.

Transmission of Y. pestis
Bubonic and Septicemic plague Disease endemic to rat species Flea bites rat and then Y. pestis replicates in flea’s digestive tract A solid mass forms, Obstructs the fleas gut, Transfers to the flea’s mouth and then Introduced to human by flea bits Bubonic and Septicemic can occur directly when rat bites human Bubonic and Septicemic can not be transferred human to human Pneumonic – primary: Contracted when infected droplets are directly inhaled This can be passed from person to person Secondary: develops when bubonic or septicemic goes untreated  moves to lungs and then can be spread to someone else

Types of Plague Incubation period 2-6 days Bubonic Plague Most common
Infection of the lymph system (attacks immune system) Fever, headache, chills, weakness, swollen and tender lymph glands Pneumonic Plague Most serious type of plague Infection of the lungs leading to pneumonia Fever, headache, weakness, rapid onset of pneumonia (accompanied by: shortness of breath, chest pain, cough, bloody or watery sputum) Septicemic Plague Bacteria reproduces in the blood Can be contracted like bubonic plague but is most often seen as a complication of untreated bubonic or pneumonic plague Fever, chills, weakness, abdominal pain, shock, bleeding underneath skin or other organs Pneumonic – primary: contracted when infected droplets are directly inhaled. This can be passed from person to person. Secondary: develops when bubonic or septicemic goes untreated  moves to lungs and then can be spread to someone else.

Stages of Disease Bacteria travel through the blood to the nearest lymph nodes In lymph nodes, Y. pestis is ingested by fixed macrophages Y. pestis grows in macrophages but not PMNs and replicates Elicits an inflammatory response (the bubo) Bubo is swelling of the lymph nodes Bacteria from the bubo leak into blood stream (septicemic plague) Lysis of the bacteria releases LPS, which causes septic shock Eventually bacteria reach the lung, where they parasitize the lung macrophages (pneumonic plague) At the pneumonic stage, the bacteria can be spread to others via aerosols (respiratory droplets) Direct inhalation at this point of the disease, induces more rapid development (than flea) At this stage the bacteria have well developed virulence factors needed to colonize the human body

Lab diagnosis Diagnosis is based primarily on clinical suspicion
Specimen Blood (Septicemic) Sputum or (Pneumonic) Pus from the bubo (Bubonic) Direct smear By Giemsa stain shows the characteristic morphology of Y. pestis Smear is the best diagnostic procedure Culture Blood agar: non-hemolytic or enteric media (MacConkey agar): non-lactose ferment After hrs the colonies were pinpoint grey to greyish white, and slightly mucloid

Treatment Without treatment, fatality rates: up to 90% for bubonic plague, 100% for Septicemic or pneumonic plague Treatment, fatality rate= (5-20%) Rapid treatment is critical to improved survival No major antibiotic resistances have developed Plague is usually treated with Streptomycin (Drug of choice) Other antibiotics : tetracyclines, chloramphenicol, fluoroquinolones Course of treatment: days

Keep rodent population down by proper disposal of garbage Anti-rats, anti fleas measures Eliminate crowded living conditions of substandard housing Strict Isolation of cases (Quarantine) Vaccines Effective but protects for less than a year Heat killed vaccine consists of whole killed Y. pestis cells Requires a series of injections over a 6 month period Live attenuated vaccines: Injection of non-pathogenic mutant, derived from a fully virulent strain Chemoprophylaxis Sulfonamides and tetracyclines during epidemics

Francisella tularensis
Small gram-negative coocobacilli (0.2 by µm) Nonmotile, non-spore forming, strict aerobic, and non-fermentative Fastidious and slow-growing Requires cysteine-supplemented specialized media Blood-glucose-cysteine agar Requires prolonged growth Facultative intracellular bacterium Survive up to weeks at low temperatures in water, soil, and animal carcasses Major target organs are lymph nodes, lungs, pleura, spleen, liver and kidney One of the most infectious pathogenic bacteria known Substantial capacity to cause illness and death

Francisella tularensis
Disease: Tularemia Also Known As… Rabbit fever, Deer-fly fever, Glandular tick fever It resembles bubonic plague Virulence Factors Antiphagocytic capsule Thin lipid capsule present in pathogenic strains Facultative intracellular parasite that can survive in macrophages of the reticuloendothelial system

Clinical Presentation of Tularemia
NOTE: Also Gastrointestinal & Pneumonic forms of disease

Two subspecies (biovars)
Jellison Type B Jellison Type A Characteristics Europe, Asia North America Geographic distribution Contaminated water or rodents Rabbit and ticks Source of infection Relatively virulent (Mild) Highly virulent (Sever) Severity <0.5% in untreated cutaneous disease 5-6% in untreated cutaneous disease Mortality Negative Positive Biochemical properties Glycerol fermentation Citrulline ureidase Divided based on virulence, acid production from glycerol, citrulline ureidase activity (conversion of L-cittruline to ornithine) 10 CFU—50% lethal dose

Infection Incubation period 1-21 days (average=3-5 days)
Infective dose 10-50 organisms Duration of illness 2 weeks Mortality treated: low untreated: moderate Ulceroglandular and glandular tularemia are rarely fatal (mortality rate < 3%) Pulmonic or Septicemic tularemia is more acute form of disease (mortality rate %) Vaccine efficacy good, ~80% Wild animals are carriers – flies are infectious for up to 2 weeks, ticks are infectious for life, in blood 2 weeks, and in lesions 1 month Rarely fatal even without treatment, although it may take significant amount of time to resolve Typhoidal tularemia = septicemia without lymphadenopathy or the appearance of an ulcer, possibly have delirium and shock

Very small number of organisms to become infected (10-50)
Route of Transmission Mode of Transmission Skin or conjunctiva Handling of infected animals Skin Bite of arthropod (vector) which has fed on an infected animal (Reservoir) GI tract Ingestion of improperly cooked meat or contaminated water Respiratory tract Aerosol inhalation Very small number of organisms to become infected (10-50) Humans cannot transmit infection to others Reservoirs Rabbits, Aquatic Rodents, Rats, Squirrels, Lemmings, Mice Vectors Ticks, Mosquitoes, Biting Flies Ulceroglandular Tularemia (Most common) Transmitted through a bite from an arthropod vector which has fed on an infected animal (Reservoir)

Diagnosis of F. tularensis
Because this bacterium so virulent, most labs will not culture it from pus or blood It is not advisable to drain infected lymph nodes Fastidious and slow-growing Requires cysteine-supplemented specialized media Requires prolonged growth Diagnosis rests on Clinical picture Skin test similar to tuberculin and brucellin Measurement of antibodies titer

Prevention Avoidance of reservoirs and vectors
Protective clothing and gloves Laboratory personnel should be made aware of potential for Fransicella in clinical specimens Best Immunity (Permanent) Previous infection with avirulent strain Live Vaccine Strain (LVS) Best prophylactic Only effective vaccine against tularemia Doesn’t provide 100% immunity Possibility of varying immunogenicity between different batches Possibility of a spontaneous return to virulence Foshay’s Vaccine (killed bacteria) Provides lesser immunity towards systemic and fatal aspects of disease than LVS Immunity to reinfection is permanent and is the result of a CMI mechanism in which macrophages activated by MAF released from antigen sensitized effector T cells destroy the phagocytized organisms LVS

Antibiotics to Treat Tularemia
Antibiotics are effective for treatment after exposure Antibiotic treatment must begin several days post-exposure to prevent relapse Streptomycin and gentamicin Effective against tularemia Require intramuscular or intravenous administration High toxicity profile Can be relapses of tularemia on aminoglycosides There exist streptomycin-resistant strains of F. tularensis Tetracyclines and chloramphenicol Can be administered orally Higher relapse rate than aminoglycosides Quinolones (including ciprofloxacin) Generally works well and Low relapse rate Has not been used extensively for treatment

Anaerobic Gram negative bacilli
Gram negative rods, Anaerobic, Non-spore forming Over 24 genera of Gram-negative anaerobic bacilli The most important genera: Bacteroides, Prevotella, Fusibacterium and Porphyromonas B. fragilis, B. melaningenicus (now P. melaningenicus) and B. corrodens are the most important species They predominant components of the florae of mucous membranes B. fragilis group are the predominant in the colon (1011/g of feaces) Anaerobes outnumber aerobes by 1000:1 in the large intestine They play an important role in almost all intra-abdominal infections 60% of female carry it in their vagina P. melaningenicus and B. corrodens occur primarily in oral cavity

Anaerobic Gram negative bacilli
Opportunistic pathogens Cause serious infections Infections are generally polymicrobial Transmission Endogenous spread to normally sterile tissues or fluids Bacteroides infections can develop in all body sites, including the CNS, the head, the neck, the chest, the abdomen, the pelvis, the skin, and the soft tissues Predisposing factors Any condition predisposing to anaerobic infections such as diabetes Surgical/Trauma patients and chronic diseases Human and animal bites Decreased redox potential: Local tissue necrosis, impair blood supply and growth of facultative anaerobes (E. coli) at the site contribute to anaerobic infection E. coli utilize O2 therefore reduce it to level allow anaerobic to grow

Anaerobic Gram-Negative Infections
CNS Infections Brain abscess and meningitis Brain abscesses are commonly caused by adjacent chronic infections in ears, mastoids, sinuses, oropharynx, teeth or lungs Head and neck Infections Chronic otitis media; sinusitis; mastoiditis; tonsillar, peritonsillar, and retropharyngeal abscesses; cervical lymphadenitis Dental infections, include periodontal disease, gingivitis, localized periodontitis, pericoronitis, endodontitis, periapical and dental abscesses and post-extraction infection Pleuropulmonary Infections 90% of patients with community-acquired aspiration pneumonia B. fragilis is found in 25% of lung abscesses Intra-abdominal Infections Secondary peritonitis, Appendicitis and Abdominal abscesses

Anaerobic Gram-Negative Infections
Gynecoloical Infections Bacterial vaginosis; soft tissue perineal, tubo-ovarian, pelvic and vulvar abscesses; endometritis; salpingitis; pelvic cellulitis; intrauterine device–associated infection; septic abortion and postsurgical obstetric and gynecologic infections Skin and Soft Tissues Infections Infected cutaneous ulcers, secondary diaper rash Cutaneous and subcutaneous abscesses, breast abscesses Necrotizing fasciitis, necrotizing synergistic cellulitis, gas gangrene Osteomyelitis and septic arthritis Bacteremia (5-15%)

Bacteroides Virulence Factors
Polysaccharide capsule Antiphagocytic and adhesive Lipopolysaccharide Stimulates leucocytes migration and chemotaxis Agglutinins Enhance adherence Histolytic enzymes Tissues destruction, inactivates Immunogloublins Oxygen tolerance Catalase and superoxide dismutase Beta-lactamases Inactivates penicillins

Diagnosis Direct-needle aspiration is the best method of obtaining a culture Specimens obtained from normally sterile sites, such as blood or spinal, joint, or peritoneal fluids Bacteroides can be isolated anaerobically on blood agar containing kanamycin and vancomycin to inhibit unwanted bacteria and incubated anaerobiacally Treatment Directed at all major aerobic and anaerobic pathogens Clindamycin, cefoxitin and cefotetan Almost sensitive to metronidazole, carbapenems, chloramphenicol and combinations of penicillin/β-lactamase inhibitors

Avoiding conditions that reduce the redox potential of the tissues Preventing the introduction of anaerobes of the normal flora to wounds, closed cavities, or other sites prone to infection Surgical intervention, abscesses drainage and antibiotic treatment Prophylactic therapy are used when medical procedure disrupt the mucosa or if disruption of mucosa causing trauma No vaccines are available

General characteristics Spirochetes
Tiny (0.2 W x 6-20 m L), thin walled, flexible, spirals (helical) that like corkscrew They are coiled into regular helices (6-14/cell) Move in a unique spinning fashion via 6 thin endoflagella called axial filaments Flagella lie between outer membrane & PDG layer Gram negative Too small to be seen using the light microscope

Special procedures are required to view spirochetes
Dark field microscopy Immunofluorescence Silver stain (Fontana’s silver or Giemsa’s stain) Pathogenic species do not grow on artificial media While non-pathogenic species can be grow Can be propagate some strains of spirochetes by inoculating it into the testes of the living rabbit

Classification of Spirochetes
Human pathogens belong to following 3 genera: Treponema Leptospira Borrelia Others (saprophytes) are found in water, sewage & in mouth & genital tracts of humans

Treponema pallidium The causative agent for STD “syphilis”
Syphilis is either Acquired (STD, Venereal) Transmitted from infected skin or mucous membrane (genitalia, mouth, rectum) to other person by intimate contact Congenital Transmitted from pregnant women to their fetus Blood transfusion Rarely reported Men are more susceptible to contract syphilis than women Marked increase in syphilus in homosexual men

Stages of venereal syphilis
Primary syphilis Primary lesion is Hard chancre Develop on genitals in day Chancre heals spontaneously in 3-12 week without scar Enlargement of lymph nodes on genital organs Firm, painless, indurated & ulcerated lesion; covered with a thick exudate rich in spirochetes Highly infectious Spirochetes spread widely in tissue leading to 2ry syphilis

Secondary syphilis Most infectious stage Untreated patient enters bacteremic or 2ry syphilis 1-3 months later, secondary lesions may appear T. pallidium multiply and spread via blood This stage is systematic and characterized by Generalized lymphadenopathy Maculopapular skin rashes on the body & condylom latum (wart) at mucocutaneous junctions Lesions heal spontaneously over 6 week and yet the disease may progress latency & then to tertiary stage

Latent Syphilis Most patients are asymptomatic during this period Early latent period Last for 1-2 years after the 2nd stage 25% of patient develop symptoms of 2ry syphilis Patient can infect other Late latent period Last for many years No symptoms occur and patients are not infectious 30% of untreated patients slowly progress to 3ry syphilis The rest will remain asymptomatic

Tertiary syphilis Develops over 6-40 year with slow inflammatory damage to organs, blood vessels & nerve cells It can be grouped into 3 general categories Gummatous syphilis Occurs 10 year after 1ry stage in 10% of untreated May show localized granulomatous lesions (gumma) which eventually necrosis and become fibrotic Noninfection lesion are found mainly in skin & bones

Cardiovascular syphilis 3-10 year after 1ry stage in 15% of untreated patients Characterized by aortic aneurysm & necrosis in aorta Neurosyphilis (Late tertiary or quaternary syphilis) Occurs year in 8% of untreated patients Asymptomatic neurosyphilis, subacute meningitis, brain infarction, mental deteriorations & psychiatric symptoms In 3ry lesion , T. pallidium are rarely seen

Congenital syphilis Transplacental spread of T. pallidium after the 1st trimester of pregnancy may result in Fetal death Prematurity or Congenital syphilis The early syndrome may not present at birth, only developing during the subsequent 4 weeks They are characterized by Generalized rash resemble 2nd acquired syphilis Enlargement of the liver, spleen and lymph nodes Congenital syphilis leaves residual characteristic sign Hutchinson’s teeth Saddle nose or nose deformity

Immunity of syphilis Immunity of T. pallidium is incomplete
Antibodies do not stop the progression of the disease Treated syphilitic Patient can contract syphilis again Late syphilitic patient is relatively resistant to reinfection The antigens of T. pallidium induce Specific antibodies (non-treopenemal); detected by immuno-fluorescence or hemagglutination tests Nonspecific antibodies (reagin): detected by the flocculation of cardiolipin extracted from beef heart tissues Both specific and non-specific antitreponemal antibodies are used in serological diagnosis of syphilis

Diagnosis of Syphilis Syphilis is confirmed by
Finding T. pallidium in exudates from the lesions Or by detection of antibodies in the serum During t 1ry and 2ry stages spirochetes are demonstrated in the early lesions in wet preparation by dark field or immunfluorescense microscopy Stained with either Fontana or Giemsa stain Not seen on a Gram-stained smear During 2ry & 3ry syphilis can be detected serologically

Serological Tests of Syphilis
Patients develop 3 types of antibodies 1st is known as a lipoidophil antibody called regain Tests depend on the detection of regain Non-specific test/Standard Tests for Syphilis (STS) STS used for routine screening STS involves use of non-treponemal antigens (Cardiolipin) which react with regain in patient’s serum Can be demonstrated by CFT (Wassermann test) or by Flocculation Tests such as Venereal Disease Research Laboratory (VDRL) or Rapid Plasma Regain (RPR) test

2nd antibody called Reiter which react with a protein component found in non-pathogenic T. reiter This test called Group specific test Demonstrated by Reiter Protein CFT This test is more specific than the STS but gives fewer false positive results

3rd type of antibody reacts directly with T. pallidium These tests are specific Using Nichol’s strain It can be demonstrated by T. pallidium Immobilization (TPI) test, Fluorescent Treponemal Antibody (FTA) test and T. pallidium Haemagglutination (TPHA) test This test is the most specific tests

Prevention, Control & Treatment
There is no vaccine against syphilis This disease is controlled by Early diagnosis and appropriate treatment Use of condoms Follow-up of infected individuals & their contacts This disease is treated by Penicillin Tetracycline or Erythromycin can be used in allergic patients to Penicillin

Non-venereal Treponema
There are three subspecies of T. pallidium T. pallidium subspecies endemicum It causes endemic syphilis (Bejel) T. pallidium subspecies pertenue It causes yaws T. pallidium subspecies carateum It causes pinta

Borrelia causing diseases
Borellia Causing Relapsing Fever Endemic (European) Longer febrile period Fewer relapses B. recurrentis Transmitted by Lice Endemic (Africa) Shorter febrile period More relapses B. duttoni Transmitted by Ticks Lyme B. burgdorferei

Main differences between the two types of relapsing fever
European or louse-borne type West African or Tick-borne type Caused by B. recurrentis B. duttoni Transmitted by body louse Tick Often appears in epidemics Endemic Long febrile period and fewer relapses Shorter febrile and more relapses Spirochetes not transmitted to louse progeny Transmitted to tick progeny

Rickettsiae and Chlamydiae
Are Rickettsiae & Chlamydiae Bacteria or Virus? They share a few characteristics of viruses Their size relative to virus nm Obligate intracellular parasites Lack of ATP-generating ability Need to obtain ATP from the host cell Grow in living media (yolk sac of chick embryo) They similar to bacteria They have both DNA and RNA They have ribosomes Sensitive to antibacterial agents

Similarity between Rickettsiae & Chlamydiae Rickettsiae are similar to Chlamydia in that Both are obligate intracellular energy parasites Both are similar in size as large viruses Both grow in living media (yolk sac of chick embryo) They have both DNA and RNA They have ribosomes Both are replicate by binary fission Both are sensitive to antibacterial agents Share with viruses Share with bacteria

Rickettsiae are different to Chlamydia in that Require arthropod vector (except for Q fever) Replicates freely in cytoplasm while Chlamydia replicates in endosomes Ricketsia has tropism for endothelial cell that line blood vessels while Chlamydia like columnar epithelium

Comparison of Chlamydiae & Rickettsiae with Bacteria & Viruses
Chlamydia & Rickettsia Bacteria 15-350 350 Size (nm) Yes No Obligatory intracellular DNA OR RNA DNA & RNA Nucleic acids Synthesis & assembly Complex cycle with fission Fission Reproduction Antibacterial sensitivity Ribosomes Metabolic enzymes Energy production

Rickettsiae Taxonomy of Rickettsia Family: Ricketsiaceae
Genus: Rickettsia, Coxiella, Bartonella Species Rickettsia prowazekii (epidemic typhus) Rickettsia typhi (endemic typhus) Rickettsia rickettsii (spotted fever) Bartonella quintana (trench fever) Bartonella henselae (cat scratch fever) Coxiella burnetii (Q fever)

Rickettsiae Gram-negative, coccobacilli, non-motile
Best stained by Giemsa or Machiavello stains Small ( x 1-2 m) prokaryotic cells Contain both RNA & DNA & Multiply by binary fission Obligate intracellular energy Except Bartonella quintana They grow in tissue culture or yolk sac Except B. quintana: grow on blood agar under 5%CO2 All Rickettsial diseases are zoonotic Except Epidemic typhus and Trench fever

Rickettsiae Transmitted through arthropod vector from reservoir
Arthropods serves as both vector and reservoir Except C. burnetii does not transmitted through vector Cause skin rash, fever, headache & malaise ESCHAR, black ulcer developed at site of inoculation Except C. burnetii cause pneumonia, slow fever & hepatitis Rickettsiae are susceptible to antiseptics, dryness & heat Coxiella resist pasteurization at 600C for 30 m Rickettsiae are sensitive to chloramphenicol & doxycycline

Endothelial cells, micro blood vessels
Pathogenic Mechanism Virulence factors： Endotoxin, Phospholipase A, slime layer Mechanism： Bites or faeces of arthropod Local lymph or micro blood vessels (1st bacteremia) Endothelial cells, micro blood vessels in whole body (2nd bacteremia) Fever, rash, headache, etc Targets: Endothelial cells, micro blood vessels

Rickettsial Diseases: 1-Typhus Group
Epidemic Typhus Endemic Typhus Organism Rickettsia prowazekii Rickettsia typhi Reservoir Human and flying squirrels Rats and small rodent Vector Lice borne Flea borne Transmission Infected louse feces rubbed into broken skin Infected flea feces rubbed into broken skin Inc. period 8 days 7-14 days Distribution Worldwide worldwide Clinical Sudden onset of fever & headache Rashes which spares the palms, soles and face Delirium/ stupor Gangrene of hands or feet Gradual onset of Fever, Headache, Rash

Rickettsial Diseases:2- Spotted fever Group
Rocky Mountain Spotted Fever Mediterranean Sea Spotted Fever Causative agent Rickettsia rickettsii Rickettsia conorii Reservoir Dogs, rabbits, & wild rodents Rodent Vector Tick borne Transmission Wild and dog ticks bite Tick bite Geographic distribution USA Southern Europe, Africa, Middle East Clinical Fever Conjunctival redness Sever headache Rash on wrests, ankles, soles, palms initially and becomes more generalized later Severe Headache Rash

Rickettsial Diseases: 3. Trench Fever
Also called Five-day fever Causative agent: Bartonella quintana Reservoir: Human only Vector: Body human louse Transmission: Via feces of infected body lice being scratched into the skin Geographic distribution: Temperate regions and high elevations in the tropics, including South America Clinical Fever, Headache and Back pain It lasts for 5 days and recurs at 5 day intervals

Rickettsial Diseases: 4- Q Fever
Coxiella burnetii is the causative agent of Q fever. Q from Query Geographic distribution: Worldwide Clinical: Fever, Headache, Viral-like pneumonia, No Rash Complication of Q fever are Hepatitis and Endocarditis It can grow at pH 4.5 within phagolysosomes Coxiella burnetii is unique to the Rickettsiae Why??? Because like Gram-positive spore formers This endospore confers properties to the bacteria that differ from other Rickettsiae: This make the organism resistance to heat and drying Also extracellular existence

Q Fever Q fever is zoonotic and occupational disease
Reservoir: Cattle, sheep and goats Non-arthropod vector transmission due to Coxiella burnetii grows in Ticks and Cattle The spores remain viable in; Dried tick feces deposited in cattle hide and Dried cow placentas following the birthing So Pneumonia occurred via inhalation of Spores via Airborne transmission of spore from hide or dried placenta or Via consumption of spore contaminated unpasteurized milk

Laboratory diagnosis Specimen: Blood and/or autopsy (Tissue) Staining
Gram stain: gram negative coocobacilli (poorly stain) Giemsa stain: purple Isolation: difficult and dangerous The organism can be inoculated into tissue culture and grown over 4-7 days but this is very hazardous to personnel Serodiagnosis Complement Fixation test (CFT) Complement Complement Ag + Ab Ag-Ab No lysis Ag-Ab If the complement is free Indicator system RBCs-anti-RBCs Indicator system RBCs-anti-RBCs Ag-Ab + Lysis

Laboratory diagnosis Indirect Immuno-fluorescence reaction (IIFR)
Rckettsial antigen + Patient’s serumn Ag-Ab complex Add anti-antibody (Fluorescence) Ab-F Result in Ag-Ab-Ab-F which detected under fluorescence microscope The use of immunofluorescent antibodies to examine a biopsy can be diagnostic Weil-Fleix Reaction Some Rickettsia share antigenic characteristics with non-motile Proteus vulgaris strain P. vulgaris strains that share these common antigens are designated OX-2, OX-19, OX-K This is done by mixing patient’s serum (Antibody) and P. vulgaris strains (Antigen) Agglutination means positive test Results are shown in the Table

Weil-Fleix Reaction + Rocky Mountain spotted fever
Disease OX-K OX-2 OX-19 - + Rocky Mountain spotted fever Mediterranean Sea fever Rickettsial pox Epidemic typhus Endemic typhus Trench fever Q fever

Control Sanitary: Arthropod and rodent control Immunological:
No vaccines are currently available Chemotherapeutic: Tetracycline or chloramphenicol are drugs of choice

Chlamydiae Chlamydia is extremely tiny
Gram-negative cocci bacteria, non-motile Have rigid cell wall Have outer membrane & Cytoplasmic membrane Lacking PDG and muramic acid Basophilic because stained blue by Giemsa stain Elementary bodies stain purple Reticulate body stain blue Inclusion body stain dark purple All share common complement fixing antigen

Special Growth developmental Cycle
Elementary body Infects host epithelial cell Taken by endocytosis Reticulate body Formed 1-2 hr latter Larger than EB Divided by Binary fission Inclusion body Aggregates of small particles Formed within hrs Host cell rupture releasing EB Infects again new host cell Chlamydia pass through a series of developmental forms while multiplying by binary fission. This is termed the "developmental cycle." Two morphologically different developmental forms with a continuous gradation of intermediates between them can be recognized. One is a small cell about 0.3 u in diameter, with an electron-dense nucleoid. The other is a large cell, 0.5 to 1.0 u in diameter without a dense center. There appears to be no significant difference in morphology or developmental cycle among the various chlamydia, and a single generalized description applies to all. The development cycle may be regarded as an orderly alternation of the small and large cell type. It is initiated by the highly infectious small cell which is taken into the host cell by phagocytosis. The engulfed small cell retains its morphological integrity in vacuoles bound by membrane derived from the surface of the host cell, and there is no eclipse (period in which the parasite loses the infectious ability). Instead, without loss of individuality, the small cell is reorganized into a large cell which is the vegetative multiplying form of these organisms. Then, still within the membrane-bound vacuole, the large cell grows in size and multiplies by repeated binary fission. The developmental cycle is completed by the reorganization of most of the large cells into small ones which are then available for infection of new host cells. The time required for completion of a cycle varies from hours, depending on the particular host/parasite system involved.

Characteristics of elementary and reticulate bodies of Chlamydia
RETICULATE BODY (RB) ELEMENTARY BODY (EB) Size um ( nm) Size 0.3 um (300 nm) RNA:DNA content = 3.1 RNA:DNA content = 1.1 Not infectious Infectious Adapted for intracellular growth Adapted for extracellular survival Hemagglutinin absent Hemagglutinin present Does not induce endocytosis Induces endocytosis Metabolically active Metabolically inactive

Classification and Differentiation of Chlamydiae
Subgroup B Subgroup A Bird parasites Mammalian parasites Diffuse inclusions Compact inclusions Transmitted by inhalation Transmitted by contact Glycogen not synthesized Glycogen synthesized Folates not synthesized (resistant) Folates synthesized (Sensitive) Resistant to D-cycloserine Sensitive to D-cycloserine Broadening of host range Restricted host range Chlamydia psittaci Chlamydia pneumoniae Both infect LUNG Chlamydia trachomatis Infects non-ciliated columnar epithelial cells (EYE and GENITAL)

SubgroupA: Chlamydia trachomatis
Three biovars (biological variants): Biovar Trachoma (15 Serologic types (A-K) A, B, Ba, C, D, Da, E, F, G, H, I ,Ia, J, Ja, K Serotypes A, B & C cause Trachoma Serotypes D-K cause inclusion conjunctivitis (Newborn), Non-Gonococcal Urethritis, Cervicitis, Infant pneumonia Biovar lymphogranuloma venereum, LGV (4 serologic types) (L1, L2, L3,L4) Biovar mouse Infects human non-ciliated columnar epithelial cells: Eye and Genitals Except Biovar mouse

a. Trachoma Severe form of chronic conjunctivitis
Caused by C. trachomatis biovar Trachoma Serotypes A, B & C Transmission occurs by hand-to-hand transfer of infected secretions to eye by infected articles (Towels) Can be also transmitted by droplets, contaminated clothing, flies and by passage through an infected birth canal Infections occur most commonly in children Incubation period of 5 to 12 days

a. Trachoma Conjunctivitis, pink eye, Eye discharge, Swollen eyelids, Swelling of lymph nodes in front of the ears, corneal ulcer Producing scarring and deformity of the eyelids and corneal vascularization and opacities which may lead to blindness Blindness develops slowly over years Occurs worldwide primarily in areas of poverty & overcrowding 500 million people are infected worldwide 7 - 9 million people are blind as a consequence

b. Inclusion conjunctivitis
Collection of initial bodies in cytoplasm of conjunctival cells STD (Sexual Transmitted Disease) caused by C. trachomatis biovar Trachoma Serotypes D-K Infection derives from mother to neonates during birth Mucopurulent yellow discharge and swelling of eyelids Develops 5-14 days after birth Newborns are given Erythromycin eye drops prophylactically

c. Infant pneumonia Spread of Biovar Trachoma serotypes D-K through nasolacrimal duct Common in babies Develops between 4-11 weeks of life Initially, the baby develops upper respiratory symptoms followed by rapid breathing, cough and respiratory distress Diagnosed by presence of Chlamydial anti-IgM and/or demonstration of C. trachomatis in clinical specimen Treated with oral erythromycin

d. Urethritis (Male/Female)
Caused by C. trachomatis Biovar Trachoma Serotypes D-K Urethritis –One cause of Non-Gonococcal Urethritis (NGU) NGU are most common caused by C. trachomatis & U. urealyticum STD and Majority (>50%) asymptomatic Symptoms : mucoid or clear urethral discharge, dysuria Incubation period unknown (5-10 days in symptomatic infection) Clinically, no difference between NGU and gonococcal urethritis Urethritis usually occurs as mixed infections Empirical therapy for urethritis Single dose of ceftriaxon (against Gonococci) followed by days course of doxycycline or azithromycin (against NGU agents)

e. Cervicitis and Pelvic Inflammatory Disease (PID)
Majority (70%-80%) are asymptomatic Local signs of infection, when present, include: Mucopurulent endocervical yellow discharge Edematous cervical ectopy with erythema and friability Infection can spread upwards to involve uterus, fallopian tubes (Salpingitis) and ovaries PID develops abnormal vaginal discharge or uterine bleeding, pain with sexual intercourse, nausea , vomiting and fever The most common symptom is lower abdominal pain Infection by both N. gonorrhoeae & C. trachomatis is called PID Treated by ceftriaxon followed by 14-days doxycycline

f. Lymphogranuloma Venereum (LGV)
STD Caused by C. trachomatis biovar lymphogranuloma venereum L1, L2, L3 serotypes Starts with painless papule or ulceration on the genitals that heal spontaneously The bacteria migrate to regional lymph nodes which enlarge over the next 2 months These nodes become increasingly tender and may break open and drain pus Fever, skin rash, nausea, and vomiting are often found

Lab diagnosis Specimen:
Sputum, Conjunctival scrapping, urethral discharge or pus in LGV Satin: Giemsa’s stain Culture Cell culture treated with cycloheximide Non-culture tests Nucleic Acid Amplification Tests (NAATs) Non-Nucleic Acid Amplification Tests (Non-NAATs) Serology (Complement Fixation Test)

Subgroup B: C. psittaci It infects > 130 species of birds
It is the causative agent of; Psittacosis (parrots; parrot fever) or Ornithosis (Pigeons, chicken, ducks and turkey) Occupational disease for poultry workers Zoonotic disease (animal diseases) Infection occurred by inhalation of respiratory discharge, feather or contaminated fecal material of birds Occurs after 1-3 weeks after exposure Infection results in atypical pneumonia

Subgroup B: C. pneumoniae
First recognized in 1983 as a respiratory pathogen, after isolation from a student with pharyngitis Pneumonia or bronchitis, gradual onset of cough with little or no fever. Less common presentations are pharyngitis, laryngitis, and sinusitis Person-to-person transmission by respiratory secretions All ages at risk but most common in school-age children

Mycoplasma & Ureplasma
Smallest free-living organisms ( nm) Pass through some bacterial filters Lack of a cell wall Three layer membranes Outer and inner: proteins and saccharide Middle: 1/3 cholesterol Multiple shapes: round, pear shaped & even filamentous Require complex media containing sterol Require sterols for growth and for membrane synthesis Grow slowly (3 weeks) by binary fission and produce "fried egg" or “T strain” (tiny strain) colonies on agar plates

Two genera that infect humans: Mycoplasma and Ureaplasma
They belong to Mycoplasmataceae There are many species of mycoplasmas Only four are recognized as human pathogens; Mycoplasma pneumoniae Upper respiratory tract disease, Tracheobronchitis, atypical pneumonia Mycoplasma hominis Pyelonephritis, pelvic inflammatory disease, postpartum fever Mycoplasma genitalium Nongonococcl urethritis (NGU) Ureaplasma urealyticum NGU and may play a role in male fertility Isolation pathogen cultured in pH 6.0 media Requires 10% urea for growth

Pathogenesis Adherence factors Toxic Metabolic Products
Adherence proteins are one of the major virulence factors Adhesin localizes at tips of the cells and binds to sialic acid residues on host epithelial cells Toxic Metabolic Products Intimate association provides an environment in which toxic metabolic products accumulate and damage host tissues Products of metabolism : hydrogen peroxide and superoxide -- oxidize host lipids Inhibit host cell catalase Immunopathogenesis M. pneumoniae is a superantigen Activate macrophages and stimulate cytokine production and lymphocyte activation Host factors contribute to pathogenesis

M. pneumoniae Need 10-20% Serum to culture in pH 7.8-8.0
Pathogenesis: P1 protein, capsule and saccharide Spread by close contact via aerosolized droplets Incubation period: 1-3 weeks Cause tracheobronchitis, primary atypical pneumonia Fever, headache, sore throat and cough Initially cough is non-productive but occasionally paroxysmal Antibodies play a role in controlling infection, particularly IgA Delayed type hypersensitivity spread in confined populations

Medical Mycology Myco = Fungi Ology = Science
4/27/2017 Medical Mycology Myco = Fungi Ology = Science Mycology is the science deals with fungi Mycoses = Fungal infection Dermatophytosis - "ringworm" disease of the nails, hair, and/or stratum corneum of the skin caused by fungi called dermatophytes Dermatomycosis - more general name for any skin disease caused by a fungus Mohamed Al Agamy Mycology

What is the FUNGUS? Eukaryotic → a true nucleus
4/27/2017 What is the FUNGUS? Eukaryotic → a true nucleus Do not contain chlorophyll Organoheterotrophic Cannot photosynthesize their own food Live either as saprophytes or parasites Have chitin cell walls Produce filamentous structures Reproduce by spores (sexual & asexual) Are aerobic life forms Mohamed Al Agamy Mycology

What are Actinomyces? True Bacteria (Prokaryotic)
Similar to fungi (fungi-like bacteria) WHY? Clinical infection resembles mycoses Grow on mycotic media Grow slowly >48 h Gross colonies resemble fungi (rough, heaped, short aerial filaments) Resemble mycelia microscopically, with branched mycelia in tissue and smears

Fungi vs. Bacteria Fungi Bacteria Nucleus Cell Wall Nutrition Example
Eukaryotes Prokaryotes Nucleus *Chitin Peptidoglycan Cell Wall Heterotrophs Auto- or Heterotrophs Nutrition Sexually & asexually Binary fission Reproduction Aspergillus E. coli Example Chitin is not found in any other microorganisms

Types of fungi Multicellular: Molds- filamentous
Penicillium, Aspergillus Unicellular: Yeasts Candida, Cryptococcus Dimorphic Fungi Dermatophytes, Histoplasma

YEAST Facultative Anaerobes Fermentation=ethanol and CO2
Non-filamentous unicellular fungi Spherical or oval Reproduction by Fission or budding 17

DIMORPHIC FUNGI Growth as a mold or as a yeast
Most pathogenic fungi are dimorphic fungi At 37o C yeast-like At 25o C mold-like Can also occur with changes in CO2 Fungi grow differently in tissue vs nature/culture; often dictated by temp 19

Basic structure of fungi
Hyphae Main body of most fungi is made up of fine, Cylindrical, branching threads called hyphae Tubular cell wall filled with cytoplasm & organelles Most hyphae are 2-10 m diameter Mycelia When formed of many cells, cellular units connect together (intertwining) to form long filamentous Aerial mycelium Part projects above the surface of medium Vegetative mycelium Part penetrate into medium and absorb food

Basic structure of fungi
Septa—regular cross-walls formed in hyphae Septate: Hyphae with septa - or Aseptate or coenocytic: Lacking septa except to delimit reproductive structures & aging hyphae Primary septa: Formed as a process of hyphal extension & generally have a septal pore, which allows for cytoplasmic & organelle movement Secondary or adventitious septa are imperforate, formed to wall off ageing parts of the mycelium

Reproduction Propagate via formation of spores
Sexual and asexual spores The shape and type of spores are Different from one type of fungi to another Important in the classification & identification of different species of fungi

Asexual spore Conidiospore
Multiple (chains) or single spores formed at the end of an aerial hypha Not enclosed by a SAC Aspergillus spp. Penicillium spp Conidiophore: filament that forms Conidospore

ConidioSpores Arthrospores Blastospores
Cells in hyphae develop thick wall & separate by disarticulation Coccidioides- genus of dimorphic Ascomycete Blastospores Thickened wall Bud from the parent cell Candida, Cryptococcus

Chlamydospores Spore contained within hypha Rounded & Thick cell wall
Chlamydophores Aerial hypha with chlamydospores e.g. Candida albicans

Sporangiospore Spore contained in a sporangium at the end of a sporangoiphore Sporangium: A sac or cell containing spores produced asexually Sproangiophore- aerial hypha with sporangium Rhizopus spp

Sexual spores Sexual spores - exhibit fusion of nuclei Ascospore
Formed in sac-like cell (ascus) Often 8 spores formed Ascomycetes Basidiospore Produced on a specialized club-shaped structure (basidium) Basidiomycetes Zygospore Thick-walled spore formed during sexual reproduction Zygomycetes/ Mucor and Rhizopus

Classification of Fungi
Comprised of over 100,000 species Classified into 4 orders according to the presence or absence of sexual reproductive cycle and the nature of sexual spores Orders of Fungi Mycelium Reproduction Zygomycetes Non-septate Sexual Ascomycetes Septate Basidomycetes Deutromycetes (Fungi imperfecti) No sexual

ZYGOMYCETES Phylum Zygomycota/Sexual/Non-septate Lower fungi
< 1000 species Produce zygospore, or sporangiospores Include the common bread molds and other fungi that cause food spoilage Mucor and Rhizopus are most familiar example

Ascomycetes Phylum Ascomycota, Septate/Sexual/Higher fungi
Certain yeasts & some fungi that causes plant disease Contains more than 30,000 species of unicellular and multi-cellular fungi Produce sexual spores (ascospores) Produce asexual exospores (conidia) Unicellular e.g. Saccharomyces, Candida Multicellular e.g. Penicillum, Asperigullus, Claviceps, Dermatophytes, Sporothrix schenckii, Histoplasma

Basidomycetes Phylum Basidiomycota/Septate/Sexual
Higher Fungi-Over 30,000 different species Sexual spores borne on clublike stalks (basidia) Mushrooms (Agaricus bisporus, Agaricus campestris) are the most familiar members Among Basidiomycota, only Malassezia & Cryptococcus are frequent human pathogens

Septate mycelium: Deutromycetes:
Fungi imperfecti Sexual life cycle is either unknown or absent Reproduce by various types of asexual spores including budding Have an abundant mycelium at times while at other times they grow as yeast-like cells Includes majority of pathogenic to man & animal Resemble Ascomycetes in morphology Examples: Trichophyton, Epidermophyton, Microsporon, Candida, Cryptococcus, Histoplasma

Fungal cell wall composition
Consists of  80% of polysaccharides Polysaccharide fibrils provide rigidity/integrity of wall  20% of the wall consists of Proteins Structural components (Fibrous) Chitin microfibrils Polymers of ß (1-4)-linked N-acetylglucosamine Chitosan (in Zygomycota only) De-acetylated chitin ß-linked glucans Polymers of β-1,3-linked glucose residues with short β -1,6-linked side chains

Fungal cell wall composition
Matrix components (Gel-like components) Structural polymers are contained in gel-like matrix Mannoproteins Glycoproteins (form matrix throughout wall)  (1,3) glucan

Structure of cell membrane
Semi-permeable Phospholipid bilayer Involved in uptake of nutrients Anchorage for enzymes/proteins, e.g., chitin synthase, glucan synthase, etc. Signal transduction Differs in that it contains ERGOSTEROL Site of action for certain antifungal drugs Human cell doesn't contain ergosterol Human cell contains cholesterol

Medical Effect of Fungi
I. Mycotoxicoses: Mould produces secondary metabolite (MYCOTOXINS) Highly toxic to humans Ingestion of toxic fungi or their metabolites Ergotism is caused by eating bread prepared from rye infected with Claviceps purpurea Historically, several large scale outbreaks of madness in populations have been attributed to ergotism Ergot are -adrenergic blockers inhibits response to adrenaline vasoconstriction necrosisgangaren Symptoms consisted of inflammation of infected tissue, followed by necrosis and gangrene

Natural occurrence: Food products contaminated with AFLATOXINS include cereal (maize, rice & wheat), oilseeds (groundnut, soybean & cotton), spices (black pepper, coriander & zinger), tree nuts (almonds, and coconut) & milk Physical and chemical properties: Aflatoxins are potent toxic, carcinogenic, mutagenic, immunosuppressive agents, produced as secondary metabolites by Aspergillus flavus and A. parasiticus

II. Hypersensitivity Disease: Fungal spores are inhalated They can be an antigenic  elicit immune response  production of Ig or sensitized lymphocyte Example is hypersensitivity pneumonitis

III. Colonization and resultant disease: They may attack: Outermost layers of Skin, hair and/or mucous membrane  superficial mycoses Epidermis as well as nail and hair  cutaneous mycoses Dermis, subcutaneous tissues, muscle and face  Subcutaneous mycoses The internal organs as the lungs, CNS, bones etc.  systemic mycoses Opportunistic - cause infection only in the immunocompromised

A. Superfacial mycosis Tinea versicolor (Pityriasis versicolor)
4/27/2017 A. Superfacial mycosis Tinea versicolor (Pityriasis versicolor) Tinea nigra They are extremely superficial mycoses Primary Manifestation is pigment change of the skin Both are named for their respective skin manifestation Mohamed Al Agamy Mycology

Tinea versicolor (Pityriasis versicolor)
4/27/2017 Tinea versicolor (Pityriasis versicolor) Causative agent: Malassezia globosa Less common-Malassezia furfur Lipophilic Yeasts belongs to Basidomycota Normal flora of skin and scalp Superficial opportunistic pathogens of the skin Associated with seborrheic dermatitis, dandruff (Pityriasis capitis) and atopic dermatitis Pityriasis versicolor is chronic superficial mycoses Characterized by a blotchy discoloration of skin which may itch With sunlight exposure the skin around patches will tan, but patches remains white Mohamed Al Agamy Mycology

Pityriasis versicolor
Chronic superficial mycoses Characterized by hyperpigmented lesions Well-demarcated white, pink, or brownish lesions, often coalescing and covered with thin furfuraceous scales The colour varies according to; The normal pigmentation of the patient exposure of the area to sunlight the severity of the disease. Lesions occur on the trunk, shoulders and arms, Rarely on the neck and face Fluoresce a pale greenish colour under Wood's ultra-violet light.

B. cutaneous Mycoses Dermatophytes attack keratinous structure of skin, hair and nail and cause the group of disease known as Ringworms or Tinea Candida can attack the skin, the mucous membranes and rarely the internal organs Non-dermatophyte moulds e.g. Hendrsonula toruloidea, Scytalidium hyalium, Scopulariopsis brevicaulis

i. Dermatophytes Confined to the outer layers of skin, hair & nails
Do not invade living tissues Called dermatophytes (keratinophilic fungi) Produce extracellular keratinas  hydrolyze keratin Utilize keratin for their nutrition Keratin is the chief protein in skin, hair and nail They caused dermatophycosis“Ringworm" or “Tinea" Dermatophycosis refers to the characteristic central clearing that often occurs in dermatophyte infections

I. dermatophytes Trichophyton (19 species) Affect hair, skin & nails
Infect both children & adults Epidermophyton Infect skin, nails (rarely hair) Infect adults, rarely in children (ringworm) Epidermophyton floccosum Microsporum (13 species) Affect hair, skin (rarely nails) Frequently in children, rarely in adults M. canis is the most common infect man

Ecology and Mode of transmission
To determine the source of infection Anthrophilic Some Dermatophytes affect man only Person -to-person transmission through contaminated objects (comb, hat, etc.) T. rubrum and T. mentagrophytes Zoophilic Other affect animal mainly Direct transmission to humans by close contact with animals M. canis and T. verrucosum

Ecology and Mode of transmission
Geophilic Other live in soil and can affect man Transmitted to humans by direct exposure M. gypseum Each geographic locality has its own dermatophyton T. violacium is the prevalent causative organism of Tinea capitis in Egypt M. audouini is prevalent cause in England

Tinea corporis Small lesions occurring anywhere on the body
Causative agent: Trichophyton rubrum, T. mentagrophytes, M. canis, and M. audouinii Live on the skin surface (opportunistic) Reservoir: Humans, soil & animals Acquired by person-to-person transfer usually via direct skin contact with infected individual Animal-to-human transmission is common

Tinea pedis "athlete's foot"
Infection of toe webs and soles of feet Causative agent: Trichophyton rubrum, T. mentagrophytes and Epidermophyton floccosum Requires warmth and moisture to survive and grow Causes scaling, flaking & itch of affected areas Reservoirs: Humans Athlete's foot is a communicable disease Transmission: when people who regularly wear shoes go barefoot in a moist environment

Tinea capitis Infection of the scalp Causative agent:
Trichophyton and Microsporum invade the hair shaft Trichophyton infection prevail in Central America to USA & in parts of Western Europe Microsporum infection are in South America, Southern & Central Europe, Africa & Middle East Reservoirs: Humans & animals (dogs, cats and cattle) Transmitted by humans, animals or objects that harbor the fungus

Dermatophytes Tinea cruris - "jock itch" Tinea barbae
Infection of the groin, perineum or perianal area Trichophyton rubrum Some other contributing fungi are Candida albicans, T. mentagrophytes and Epidermophyton floccosum. Tinea barbae Ringworm of the bearded areas of the face and neck Trichophyton mentagrophytes and T. verrucosum

Dermatophytes Tinea unguium (onychomycosis) Infection of nails
Common - Trichophyton rubrum Less common- T. interdigitale, Epidermophyton floccosum, T. violaceum, Microsporum gypseum, T. tonsurans, T. soudanense Reservoirs: Humans and rarely animals or soil

Treatment of Dermatophytes
All are sensitive to grisofulvin Tolfnatate available over the counter – Topical Terbinifine (Lamisil) - oral, topical. Ketoconazole seems to be most effective for tinea versicolor and other dermatophytes Itraconazole - oral Echinocandins (caspofungin)

Lab diagnosis of Dermatophytes
Specimen collection Skin Scrapings, nail scrapings and epilated hairs Direct Microscopy Specimens should be examined using 10% KOH and Parker ink or calcofluor white mounts Characteristic hyphae can be seen

Lab diagnosis of Dermatophytes
Culture Specimens should be inoculated onto Sabouraud's dextrose agar (General purpose) Selective – Mycosel agar Gentamicin: inhibits normal bacterial flora Cycloheximide: inhibits saprophytic fungi containing cycloheximide and incubated at 250C for 4 weeks The growth of any dermatophyte is significant

II. Candida C. albicans, C. tropicalis & C. glabrata
Yeast-like organism that lives as a commensal in oral mucosa, throat, skin, scalp, vagina, fingers, nails, bronchi, lungs, or intestine and vagina Opportunistic organism The causative agent of candidiasis C. albicans, C. tropicalis & C. glabrata Candidiasis encompasses infections that range from superficial such as thrush and vaginitis Rarely become systemic in immunocompromised patients Septicaemia, endocarditis and meningitis

Symptoms of Candida albicans
Thrush appears as creamy-white or bluish-white patches on the tongue - which is inflamed and sometimes-beefy red - and on the lining of the mouth, or in the throat. Diaper rash caused by candida is an inflammation of the skin, usually red and sometimes scaly. Vaginitis is characterized by a white or yellow discharge. Inflammation of the walls of vagina and of the vulva (external genital area) causes burning and itching. Infections of the fingernails and toenails appear as red, painful swelling around the nail. Later, pus may develop.

Laboratory diagnosis of candida
Specimen: A scraping or swab of the affected area or blood (candidemia) is placed on a microscope slide Microscopic examination A drop of 10% KOH solution is added to the specimen. KOH dissolves the skin cells, but leaves Candida intact, permitting visualization of pseudohyphae and budding yeast cells typical of many Candida species. Culture Swab/blood is streaked on a culture SDA medium The culture is incubated at 37°C for several days The characteristics of colonies may allow initial diagnosis of organism causing disease symptom

Laboratory diagnosis of candida
A germ tube test is a diagnostic test in which a sample of fungal spores are suspended in serum and examined by microscopy for the detection of any germ tubes It is particularly indicated for colonies of white or cream color on fungal culture, where a positive germ tube test is strongly indicative of Candida albicans

Treatment of candidacies
Oral drugs: Amphotericin B, fluconazole, and ketoconazole, are the drugs most commonly used to treat candidiasis Topical administration of antifungal drugs such as clotrimazole, miconazole , tioconazole, and nystatin The drug of choice is nystatin

c. Subcutaneous Mycoses
4/27/2017 c. Subcutaneous Mycoses Rare Confined to subcutaneous tissue & rarely spread systemically Confined mainly to tropical regions Include heterogeneous group of soil fungal infections Introduced into the extremities by trauma/wound Tend to be slow in onset and chronic in duration The main subcutaneous fungal infections include Sporotrichosis, chromoblastomycosis, MYCETOMA, lobomycosis, rhinosporidiosis, subcutaneous zygomycosis, & subcutaneous phaeohyphomycosis Mohamed Al Agamy Mycology

C. Subcutaneous Mycoses
Sporotrichosis caused by Sporothrix schenckii The fungus is saprophyte on dead plant material Dimorphic fungi Colonies of media at 250C have delicate radiating forms that appear as white at first but turned black with prolonged incubation Microscopic examination reveals branched hyphae with numerous conidia

Sporotrichosis It was once common in Europe but cases are now rare
Most prevalent in Americas, South Africa & Australia Infection usually follows and insect bite, thorn pricks or scratches from a fish spine. Certain occupation groups appear to have increased risk from infection These include florists, farm workers and others who handle hay and moss The most common symptom is an ulcerative lesion that may develop into lymphangitis Treated by saturated solution of KI and Amphotericin B

2. Chromoblastomycosis Chronic, localized, slowly progressive infection of the subcutaneous tissue caused by several species of dematiaceous fungi Tissue proliferation occurs around the area of inoculation producing crusted, verrucose, wart-like lesions Causative agent Most common: Cladophialophora carrionii & Fonsecaea pedrosoi Less common: Fonsecaea compactum, Phialophora verrucosa

Chromoblastomycosis Mode of Transmission
Traumatic implantation of fungal elements into skin The infection occurs in tropical or subtropical climates, often in rural areas. Traetment Chromoblastomycosis treated by surgical removal and 5-flurocytosine

Mycetoma Also called Maduromycosis or Madura foot
Madura foot referring to first case seen in Madura region of India which was in the foot of patient Syndrome involving cutaneous & subcutaneous tissues, fascia, and bone Infection focused in one area of the body (Foot) Distribution: World-wide but most common in bare-footed populations in tropical sub- or regions Characterized by draining sinuses, granules (vary in size, colour & degree of hardness) & tumefaction

Mycetoma Mode of transmission
Traumatic implantation of causative agent Causative agent (50% bacteria & 50% fungi) Soil-inhabiting bacteria (actinomycotic mycetoma or actinomycosis) Actinomadura madurae, Actinomyces israelii and Nocardia brasiliensis Soil-inhabiting fungi (eumycetoma) Madurella mycetomatis & Madurella grisea

Treatment EUMYCETOMA ACTINOMYCOTIC MYCETOMA
Treatment for eumycetoma is less successful than for actinomycetoma EUMYCETOMA surgical treatment is still usually required Ketoconazole 400mg daily, itraconazole 300 mg daily & IV amphotericin B 50 mg daily Therapy is suggested for 1-2 years Flucytosine, Topical nystatin & potassium iodide ACTINOMYCOTIC MYCETOMA Penicillin, gentamicin & co-trimoxazole for 5 weeks Followed by maintenance therapy with amoxicillin & co-trimoxazole

Systemic Mycoses Invasive infections of the internal organs with the organism gaining entry by; Lungs, GIT or through intravenous lines They may be caused by: Primary (TRUE) pathogenic fungi Opportunistic fungi that are of marginal pathogenicity but can infect the immunocompromised host

I. Primary Pathogenic Fungi
Histoplasma capsulatum (Histoplasmosis ) Blastomyces dermatidis (Blastomycosis ) Coccidioides immitis (Coccidiomycosis) Paracoccidioides brasiliensis (paracoccidioidomycosis) Dimorphic fungi normally found in soil Infection occurs in previously healthy persons Arises through the respiratory route (inhalation)

Histoplasmosis Caused by dimorphic Histoplasma capsulatum
H. capsulatum is endemic in many parts of the world including North and South America It is found in the soil and growth is enhanced by the presence of bird and bat excreta Environments containing such material are often implicated as sources of human infection

Histoplasmosis Lungs are the main site of infection but dissemination to liver, heart & CNS can occur. Pulmonary infection can resemble symptoms seen in tuberculosis Treatment: Amphotericin B Ketoconazole and other new azoles

II. Opportunistic Fungi
Patients usually have some serious immune or metabolic defect, or have undergone surgery The diseases include aspergillosis, systemic Candidiasis, cryptococcosis, Zygomycosis, Pneumocystis carinii Exceptionally, other fungi that are normally not pathogenic, such as Trichosporon, Fusarium or Penicillium, may cause systemic infections.

Cryptococcosis Systemic infection caused by encapsulated yeast - Cryptococcus neoformans Inhabits soil around pigeon roosts Host defense is CMI Affects both healthy & immunosuppressed individuals Common infection of AIDS, cancer or diabetic Primary infection in lungs via inhalation Pulmonary infection leads to cough, fever & lung nodules Polysaccharide capsule resists phagocytosis Cryptococcal meningitis is most common disseminated manifestation Can spread to skin, bone and prostate

Cryptococcosis Lab. Diagnosis
Lumbar puncture and microscopic examination of cerebrospinal fluid is diagnostic India Ink for CSF Culture on SDA White mucoid colonies within 48hours Cryptococcal antigen Serum and CSF are 99% sensitive in AIDS Serum is less sensitive in normal hosts Treatment Amphotericin B and fluconazole

Aspergillosis Aspergillus is found in soil, on plants & in decaying organic matter. 600 species, A. fumigatus is one of the most ubiquitous of the airborne saprophytic fungi A. fumigatus is the main causative agent of Aspergillosis Diseases of the soil fungus called Aspergillus Major portal of entry is the respiratory tract Via inhalation of conidia Conidia are eliminated in immunocompetent host by innate immune mechanism For people with weakened immune systems, breathing in conidia can lead to infection

Aspergillosis Most commonly affects the sinuses or lungs
Symptoms of sinus infections include fever, headache, and sinus pain Lung infections can cause fever and cough In the immunosuppressed host, Aspergillus can disseminate throughout the body. Treatment Amphotericin B and nystatin Voriconazole is currently the first-line treatment for invasive aspergillosis

Candidiasis In severely immunocompromised patients, C. albicans can proliferate and disseminate throughout the body. An infection in the bloodstream can affect the kidneys, heart, lungs, eyes, or other organs causing high fever, chills, anemia, and sometimes a rash or shock

Symptoms of Candida albicans
Candida can cause the following problems depending upon the organ infected: in kidneys can cause blood in the urine in heart can cause murmurs & valve damage in the lungs can cause bloody sputum in the eyes can cause pain and blurred vision in the brain can cause seizures and acute changes in mental function or behavior

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