2. protozoan dysentrey & intestinal infections
Dr. M. Shahnazi

3. Parasitic Amoebas Entamoeba histolytica** Entamoeba dispar
Entamoeba hartman Colon
Entamoeba coli
Entamoeba gingivalis
Entamoeba moshkovskii
Entamoeba poleki
Iodomoeba butschlii
Endolimax nana Colon
Dientamoeba fragilis** Colon
(**=pathogenic)  

4. Entamoeba histolytica/ Entamoeba dispar
15 to 20 µm
12 to 15 µm

5. histolytica/E. dispar trophozoites stained with trichrome.

6. Entamoeba histolytica
Trophozoite

7. Entamoeba coli
15 to 25 µm
20 to 25 µm

8. A: E. coli cyst in a concentrated wet mount stained with iodine
A:  E. coli cyst in a concentrated wet mount stained with iodine.  The cyst shows 5 nuclei in this focal plane.
B:  E. coli cyst in a formalin concentrated wet mount stained with iodine.

9. Iodomoeba butschlii 10 to 12 µm (range 5 to 20 µm)

10. Iodamoeba buetschlii 10 to 12 µm (range 5 to 20 µm)
A: Line drawing of an I. buetschlii cyst
I. buetschlii cysts stained with trichrome. Note the well defined glycogen mass in B (black arrow).

11. Endolimax nana 6 to 8 µm (range 5 to 10 µm)

12. A: E. nana cyst stained with trichrome.

13. 8 to 10 µm (range 6 to 12 µm).
A: Line drawing of an E. nana trophozoite. B: E. nana trophozoite stained with trichrome

14. E. nana trophozoites stained with trichrome.

15. E. gingivalis FF EE
EE, FF: Trophozoites of E. gingivalis from culture, stained with trichrome.

16. Dientamoeba fragilis Worldwide.
Despite its name, Dientamoeba fragilis is not an ameba but a flagellate. 
This protozoan parasite produces trophozoites;
cysts have not been identified.
  transmitted by fecal-oral route and transmission via helminth eggs (e.g., Ascaris, Enterobius spp.)
Infection may be either symptomatic or asymptomatic.

17. Dientamoeba fragilis
5 to 15 µm
5 to 15 µm

18. 5 to 15 µm
  Binucleate forms of trophozoites of D. fragilis, stained with trichrome.

20. Entamoeba histolytica/dispar
Entamoeba hartmani
Entamoeba coli
Endolimax nana
Iodamoeba beutschlii
         
                                 
                          
                                             
                                
                        
                      
Trophozoites
Cysts

21. Entamoeba histolytica/E. dispar
Worldwide, with higher incidence of amebiasis in developing countries.
- tropical
- Sub tropical
1% population of world
In industrialized countries, risk groups include
- male homosexuals,
- travelers and recent immigrants
Transmission
- Food&Water
- Insect

22. Metacystic Trophozoit
Entamoeba histolytica
Metacystic Trophozoit

23. E. histolytica: Life Cycle&Clinical Features
A wide spectrum, from asymptomatic infection ("luminal amebiasis"),
Invasive intestinal amebiasis
- Dysentery
- Colitis
- Amebomas
- appendicitis
-Toxic megacolon
- Flask ulcer
Invasive extraintestinal amebiasis
- Peritonitis
- liver absces
- Hepatitis
- cutaneous amebic lesions
- pleuropulmonary abscess
- Brain amebic lesions
- Genital amebic lesions

24. amoebic liver abscess

25. with perforation of abscess through abdominal skin
Patient with amoebiasis liver absess,
with perforation of abscess through abdominal skin

26. افتراق دیسانتری آمیبی و شیگلایی
PH* مدفوع
- اسیدی
- قلیایی
*تب
- بدون تب یا ضعیف
- تب دار
*بلورهای شارکوت لیدن
- در 25% موارد
- وجود ندارد
*سن
- اغلب در ساله
- ” در کودکان
*دوره کمون
- بیش از یک هفته
- کمتر از یک هفته
*درد و پیچش شکم
- موضعی و اطراف ناف
- اغلب منتشر
*همه گیری
- اندمیک گاهی اپیدمیک
- اپیدمیک گاهی اندمیک

27. E. histolytica: Laboratory Diagnosis
  Microscopic identification of cysts and trophozoites in the stool
- Fresh stool
wet mounts, with or without iodine stain
permanently stained preparations (e.g., trichrome)
- Concentrates from fresh stool
*Concentration procedures, however, are not useful for demonstrating trophozoites
- aspirates or biopsy samples obtained during colonoscopy or surgery.
Serology (IFA, IHA, ELISA)
Isoenzymes
Monoclonal antibody (antigen detection)
PCR .

28. Subphylum Mastigophora

29. Giardia lamblia

30. Giardia lamblia: Morphology
Location
Size
Reproduction
Nucleus
Sucking plate
10-20 micrometers
8-19 µm (average µm)

31. Life Cycle of Giardia lamblia

32. Giardia lamblia: Pathogenesis1
Giardiasis
symptomatic only about 30-40%
* چسبیدن به مخاط
- انسداد مکانیکی
- ترشح توکسین
- آتروفی پرزها
* تجزیه و مصرف نمک های صفراوی
- جلوگیری از عمل پانکراس ( کاهش لیپاز )
- اخلال در جذب چربی
▪ آویتامینوزAو
* رقابت با میزبان
diarrhea
steatorrhoea
Malnutrition
Malnutrition

33. Giardia lamblia: Pathogenesis2
Symptoms of infection include
- diarrhea
- vomiting
- nausea
- excessive gas
- malaise
- epigastric pain
- diminished interest in food
- weight loss
* Symptoms in gallbladder
■ Pus, mucus and blood are not commonly present in the stool
■ the condition is usually self-limiting, although the infection can be prolonged in patients who are immunocompromised, or who have decreased gastric acid secretion
■ People with recurring Giardia infections, particularly those with a lack of IgA, may develop chronic disease.
■ Some studies have shown that giardiasis should be considered as a cause of Vitamin B12 deficiency
■ Blood groupA
Infective doseََ ■
nausea vomit vomit weight loss malaise

34. Giardia lamblia: Epidemiology1
Worldwide
About 200 Million in world
1- 25%
more prevalent in
- warm climates
- children (4 – 11 year old )
*In children ( 4 year old )
maybe zoonosis
Resestance of cysts
In Iran
- 16%

35. Giardia lamblia: Epidemiology2
Transmission
- Direct
- Water&Food
- Insects
- Homosexual

36. Giardia lamblia: Diagnosis
Stool Examination ( formed, Loose )
- Wet mount
- Concentration
Repeated samplings may be necessary
samples of duodenal fluid (e.g., Enterotest)
duodenal biopsy may demonstrate trophozoites. 
Differentiation ffrom Bacterial diarrhea
- Pus, mucus and blood are not commonly present in the stool
Serologic Examination
- Antibody response
- Antigen detection
PCR

37. G. intestinalis cysts stained with trichrome.

38. G. intestinalis trophozoites stained with trichrome.

39. Giardia intestinalis: Control&prevention
Health
- Personal
-environment
- Sewage
- Insects
- Water&Food
* Filtration
* Boiling
Education
Control&Treatment of Carriers

40. Balantidum coli

41. Balantidum coli Location large ciliated protozoan Size Reproduction
Nutrition
Vacuole
Nucleus
- macronucleus
- Micronucleus
Other species
- B. suis
90 – 120 × 60 – 80 Mic.
50 – 55 × 45 – 65 Mic.

42. Balantidium coli: Life Cycle

43. Balantidum coli: Pathpgenesis1
Most cases are asymptomatic
Symptomatic
* Mild Symptomatic
* severe Symptomatic
Mechanism
- Physical
- Enzym secretion

44. Balantidum coli: Pathpgenesis2
Symptomatic
* Mild Symptomatic
- Intermittent diarrhea and constipation
- inappetence
- weight loss
- Insomnia
- headache

45. Balantidum coli: Pathpgenesis3
* severe Symptomatic
■ Intestinal Balantidiasis
- persistent diarrhea
- dysentery
- dehydration
- Anemia
- Colitis
- appendicitis
- abdominal pain
- weight loss
- unrest
Symptoms can be severe in debilitated persons.
■ Extraintestinal Balantidiasis
- Peritonitis
- liver Balantidiasis
- Lymph Nodes “
- pleuropulmonary “
- Genital “
■Death For Peritonitis, Anemia, dehydration

46. Balantidum coli: Epidemiology
Worldwide
- special in Trppical, Sub Tropical,Temperate
High Risk In
- debilitated persons
- alcoholic “
- IgA dificiency
- Malnutrition
pigs are an animal reservoir
human infections occur more frequently in areas where pigs are raised.
Other potential animal reservoirs include rodents and nonhuman primates
Resistece OF cysts
Transmission
In Iran
- sporadic

47. Balantidum coli: Diagnosis
Stool Examination ( formed, Loose )
- Wet mount
- Concentration
- permanently stained preparations
Serologic Examination

48. C: B. coli trophozoite in a wet mount, 1000× magnification. 
Note the visible cilia on the cell surface. 
D: B. coli trophozoite in a Mann's hematoxylin stained smear, 500× magnification. 
Note the cytosome (black arrow) and the bean shaped macronucleus.

49. Balantidium coli trophozoites in tissue stained with H&E.

50. Balantidium coli: Control&prevention
Health
- Personal
- environment
- Sewage
- Insects
- Water&Food
Education
Control&Treatment of Carriers

51. Phylum Apicoplexa Phylum Apicoplexa Genus Cryptosporidium
Class Sporozoa
Sub-class coccidia
Order Eucoccidiida
Family
Genus
Genus Cryptosporidium
“ Isospora
“ Cyclospora
“ Sarcocystis

52. Cryptosporidium spp. 4.2 to 5.4 µm
Cryptosporidium has been found worldwide.
Intra cellular – Extra cytoplasmic
Monoxenous
cryptosporidiosis
Outbreaks of cryptosporidiosis have been reported in several countries,
Many species of Cryptosporidium exist that infect humans and a wide range of animals. 
Although Cryptosporidium parvum and Cryptosporidium hominis (formerly known as C. parvum anthroponotic genotype or genotype 1) are the most prevalent species causing disease in humans,
infections by C. felis, C. meleagridis, C. canis, C. baileyi and C. muris have also been reported.
Oocysts Sporolated ( Direct )
4.2 to 5.4 µm

53. Cryptosporidiosis – Life cycle
مرحله عفونت زا: اووسیست های رسیده حاوی 4 اسپوروزوئیت
تکثیر انگل، داخل حاشیه پرز دار سلولهای اپی تلیال
- به روش شیزوگونی: ایجاد شیزونت( با مروزوئیت نوع I) و( مروزوئیت نوع II)
- به روش گامتوگونی: ایجاد میکرو و ما کروگامت، zygot و در نهایت oocyst
- اووسیست ها دارای دیواره ضخیم و نازک
در میزبانهای طبیعی از نظر ایمنی:
- چرخه تولید مروزوئیت های نوع II و اووسیست با دیواره نازک کند، مانع تکثیر بیشتر انگل و در نهایت خاتمه عفونت
در افراد دچار ضعف سیستم ایمنی:
- هر دو مورد به سیر خود ادامه ودر نهایت ایجاد عفونت های شدید و بادوام
Oocyst Excretion 1- several Month
Cryptosporidiosis – Life cycle

54. Cryptosporidium spp.: Clinical Features
Infection with Cryptosporidium sp.  results in a wide range of manifestations,
asymptomatic infections
symptomatic infections Endotoxin & Intestinal Irritation
- incubation period is an average of 7 days (but can range from 2 to 10 days). 
* In immunocompetent persons,
- Watery diarrhea is the most frequent symptom, and can be accompanied by
- dehydration, weight loss, abdominal pain, fever, nausea and vomiting.
- Malabsorption
 - symptoms are usually short lived (1 to 2 weeks);
- Self limited
* immunocompromised patients
- Severe symptom,
- they can be chronic and more severe in, especially those with CD4 counts <200/µl. 
- life-threatening illness
- Cryptosporidium infections have also been found in other organs including other digestive tract organs, the lungs, and possibly conjunctiva.
* Prognosis

55. Cryptosporidium spp.: Epidemiology1
Worlwide
Many Reservoirs
Zoonosis
Infective Dose
Infection in Children
“ Neoborn
“ Adults
“ Homosexual
Rubbish
Trasmission
- Direct
- Water & Food
Day-Care Center & Hospital
Resistance of Oocysts
- environment
- temperature
- Disinfectant

56. Cryptosporidium spp.: Epidemiology2
Prevalence in Countries
- Developing – 32%
- Developed – 20%
- AIDS – 50%
In Iran
- AIDS, Tehran %
- Diarrhoeal Children, Tehran, – 7%
- Eslamshahr Laboratories %
- Dialysis Patients, esfahan hospitals %
- Kidney Transmplant Hospital, Uromia
- Haematopoietic Disorders     22%
- In Hamadan, Tabriz, Ahvaz, Khorram Abad, Uromia

57. Cryptosporidium spp.: Laboratory Diagnosis
Acid-fast staining methods,
- with or without stool concentration, are most frequently used in clinical laboratories.  For greatest sensitivity and specificity,
* Modified zinc sulphate Flotation
* Sheather Flotation
immunofluorescence microscopy is the method of choice (followed closely by enzyme immunoassays).
Molecular methods are mainly a research tool.
Serologic Methods
Safety Oocysts in stool specimens infective
Thus stool specimens should be preserved in
- 10% buffered formalin
- sodium acetate-acetic acid-formalin (SAF) to render oocysts nonviable.
- aqueous potassium dichromate (2.5% w/v, final concentration). 

58.                                               A B
4.2 to 5.4 µm
Cryptosporidium parvum oocysts stained with modified acid-fast.  Against a blue-green background, the oocysts stand out in a bright red stain.
  Sporozoites are visible inside the two oocysts to the right

59.  Cryptosporidium parvum oocysts stained with the fluorescent stain auramine-rhodamine.

60. Agarose gel (2%) analysis of a PCR diagnostic test for detection of Cryptosporidium parvum DNA.  PCR was performed using primers CPBDIAGF and CPBDIAGR.1

61. Isospora Monoxenous Isosporiasis
Worldwide, especially in tropical and subtropical areas.
Isospora In hhman
- I. belli
- I. natalensis
infects the epithelial cells of the small intestine
Oocysts unsporolated
20 – 33 Mic.
Disporic - Tetrazoic

62. Isospora belli: Clinical Features
Infection causes acute, nonbloody diarrhea with crampy abdominal pain,
which can last for weeks and result in malabsorption and weight loss. 
In immunodepressed patients, and in infants and children, the diarrhea can be severe.
- Somewhen Extra Intestinal Infection
* repetitious Schizogony
  Eosinophilia may be present (differently from other protozoan infections).

63. Isospora belli: Laboratory Diagnosis
Microscopic demonstration of the large, typically shaped oocysts, is the basis for diagnosis. 
Because the oocysts may be passed in small amounts and intermittently, repeated stool examinations and concentration procedures are recommended.
If stool examinations are negative, examination of duodenal specimens by biopsy or string test (Enterotest®) may be needed.
The oocysts can be visualized on wet mounts by microscopy with bright-field,
They can also be stained by modified acid-fast stain.

64. 20 – 33 Mic
Isospora belli oocyst with 2 sporoblasts (Imature oocyst) (Direct wet preparation without staining)

65. Isospora belli: Control&prevention
personal hygiene
Public health
- Food & Water hygiene
- Water Filtering
- Environmental Hygiene
- vegetable hygiene

66. Cyclospora Monoxenous Infect mor Animals
human cases are caused by C. cayetanensis
Oocysts unsporolated
8 – 10 Mic.
Disporic - Dizoic

67. Cyclospora:Clinical Features
Infections, especially in disease-endemic settings
can be asymptomatic.
After an average incubation period of 1 week,
symptomatic infections typically manifest as watery diarrhea, which can be severe.
  Other symptoms include anorexia, weight loss, abdominal pain, nausea and vomiting, myalgias, low-grade fever, and fatigue. 
Untreated infections typically last for weeks and may follow a relapsing course ( AIDS ) 

68. Cyclospora: Laboratory Diagnosis
Microscopic examination: - direct microscopy
- concentration procedures
*formalin-ethyl acetate sedimentation
*Sheather’s flotation procedure).
- preserved
- stained smears (using modified acid-fast stain or a modified safranin stain)
- UV fluorescence microscopy
Sporulation assay
- 2.5% potassium dichromate
Molecular methods
- frozen without fixation

69. A, B:  Oocysts of C. cayetanensis viewed under UV microscopy

70. Oocysts of C. cayetanensis Oocysts of Cryptosporidium
µm in diameter
4.2 to 5.4 µm
Oocysts of C. cayetanensis
  Oocysts of Cryptosporidium
modified acid-fast

71. Cyclospora: Epidemiology
Cyclosporiasis has been reported in many countries, but is most common in tropical and subtropical areas.
Traveler's diarrhea
Transmission
- Water
- Fruit
- No Animal To Human & Human To human
  Since 1990, at least 11 foodborne outbreaks of cyclosporiasis, affecting approximately 3600 persons, have been documented in the United States and Canada.
In Iran
- 1375, 1378
- need to study

73. Stool specimens: Specimen Collection1
Collect the stool in a dry, clean, leakproof container.
Make sure no urine, water, soil or other material gets in the container.
* This table demonstrates the distribution of protozoa in relation to stool consistency and should be taken into consideration when specimens are received.