Specimen Collection Urinalysis - clean catch used if culture is needed. Reflex testing - microscopic or culture if UA abnormal Proper collection of cultures
Urinalysis Specimen type (clean catch, catheter, suprapubic aspiration, U-bag (nonsterile) Gross analysis - color, clarity Dipstick Microscopic - cellular elements Culture if indicated: if infection is possible from dipstick or microscopic results
Urinalysis Color: Normal = straw, light yellow. –Amber/orange = bilirubin, urobilinogen –Red = blood –Other colors Clarity: Normal = clear –Hazy –Cloudy Artifact or cellular elements
Urine Dipstick Protein: Usually in form of albumin or globulins. –Trace amounts in DM associated with increased mortality due to diabetic nephropathy. –Globulins (Bence-Jones) associated with multiple myeloma –Large amounts in nephrotic syndrome
Urine Dipstick pH: Normal 5-9, usually around 6. Acidotic or alkalotic can be due to diet, medication, disease or metabolic changes. Some bacteria incr. pH Specific Gravity: Normal Weight of particles in solution, correlates with osmolality.
Urine Dipstick Bilirubin: Increased in obstructive biliary disease, hepatocellular injury. Not incr in hemolytic jaundice. Urobilinogen: Formed by bacterial conversion of conj. Bilirubin in intestine. Incr in hepatocellular injury and jaundice, not obstructive biliary disease. Also increased in CHF with liver congestion, cirrhosis, hepatitis.
Urine Dipstick Blood: Detects blood & hemoglobin. Can cross react with myoglobin. Increased in hemolysis, GU tract cancer, UTI, calculi, coagulopathies, glomerulonephritis.
Urine Dipstick Glucose: Present if serum glucose is > 180 mg/dL. Increased in DM. Ketones: Screening for ketoacidosis in diabetics. Increased in starvation, fever, pregnancy.
Urine Dipstick Leukocyte Esterase: Enzyme released by WBCs. Marker of infection or inflammation Nitrite: Urine nitrates are converted to nitrite by some bacteria (E. coli, Klebsiella, Proteus, etc.)
Microscopic Done if dipstick abnormal Detects cellular elements –WBC –RBC –Bacteria –Epithelial cells – if contaminated sample, or tubules sloughing –Casts, crystals
Urine culture Semi-quantitative >100,000 colonies/ml indicative of infection >10,000 colonies/ml in symptomatic,immunosuppressed or abx treated patients Lower numbers suprapubic (>150)
Serology Testing serum to determine antibody levels. Used for many viruses and other infectious agents IgM - early infection IgG - lifelong, immunity
Hepatitis - Causes Drugs: antihypertensives, statins, antibiotics, others. Toxic agents: acetaminophen, alcohol, others. Viruses: Hepatitis A (HAV), B (HBV), C (HCV) commonly. Uncommon: EBV, CMV, measles, rubella, etc.
Liver Function Tests Serum Aminotransferases (ALT and AST) Serum and urine Bilirubin Serum Alkaline Phosphatase Additionally: LDH, GGTP, Albumin, Prothrombin Time
Liver Function Tests ALT – Alanine Aminotransferase AST – Aspartate Aminotransferase –Inflammation and cell necrosis –Most sensitive marker of liver injury (from infections, toxins, autoimmunity, etc)
Bilirubin Hemoglobin breakdown product Conjugated by liver, excreted in bile, eliminated in urine Bilirubin increased in: –Biliary tract obstruction (tumor, stone, pancreatitis) –Inflammation (hepatitis) –Hemolysis (Gilberts syndrome)
Bilirubin Bilirubinuria occurs in both inflammation and obstruction (but not hemolysis) Jaundice results when levels exceed 2.5 mg/dl In viral hepatitis, bilirubin not always elevated, therefore: Elevated serum bilirubin is neither sensitive nor specific for viral hepatitis
More Liver Function Tests Serum albumin: decreased in cirrhosis and severe, fulminant disease Prothrombin time: Prolonged in severe liver disease (vitamin K deficiency) LDH (lactate dehydrogenase): non specific, not very useful. ALKP: sensitive marker of biliary tract obstruction, mildly elevated in viral hep.
Lab Findings ALT usually >8x upper limit of normal ALT usually elevated >AST ALKP modestly elevated Bilirubin normal to highly elevated –These are quick tests if you suspect hepatitis. –If elevated, proceed to serology testing
Hepatitis A Serology HAV IgM – rises early in illness, will remain positive for up to six months. HAV IgG – will appear soon after IgM and remain elevated for years. –Most common cause of acute viral hepatitis (AVH), no chronicity, no carrier state
Hepatitis A Serology Testing for Hep A includes HAV Total (IgG and IgM), and HAV IgM. So, –If someone has a positive HAV Total and an positive HAV IgM, they have a current infection. –If someone has a positive Total and a negative IgM, they had a past infection or passive immunity (vaccination).
Hepatitis B Second most common cause of acute viral hepatitis Most complex hepatitis virus -infective particle made up of viral core plus an outer surface coat 5-10% become chronic, lead to cirrhosis, hepatocellular cancer
Hepatitis B Serology HBsAg: First evidence of infection, persists through clinical illness. Anti-HBs: Antibody to HBsAg appears after clearance of HBsAg and after vaccination (titer >=10 mU/mL). –Neg HBsAg and pos Anti-HBs means recovery from HBV infection, noninfectivity and immunity.
Hepatitis B Serology Anti-HBc: –IgM anti-HBc appears shortly after HBsAg. Indicates acute hepatitis. Persists for 3-6 months or longer. May appear during flares of chronic HBV. –IgG anti-HBc appears during acute HBV but persists indefinitely, whether recovery or chronic hepatitis occurs.
Hepatitis B Serology HBeAg: A soluble protein found in HBsAg positive patients. Indicates viral replication and infectivity. Appearance beyond 3 months indicates increased likelihood of chronicity. HBV DNA: Parallels presence of HBeAg, more sensitive and precise
Acute Hepatitis B Course
Hepatitis B Review Acute HBV: HBsAg+, IgM anti-HBc+, Total anti-HBc+, anti-HBs-, HBeAg+. Chronic HBV: HBsAg+, IgM anti-HBc-, Total anti-HBc+, anti-HBs-. Past resolved HBV: HBsAg-, IgM anti- HBc-, Total anti-HBc+, anti-HBs+. Vaccination (immunity): anti-HBs+.
Hepatitis C Chronicity common (>70%) Prolonged viremia Aminotransferases elevated off and on (can have ALT >7x normal) Diagnose with Anti-HCV EIA –False negatives early in disease (low sensitivity) –False positives with elevated gamma glob (low specificity)
Hepatitis C Serology Positive Anti-HCV EIA needs confirmation HCV RIBA (Recombinant Immunoblot Assay) confirms + EIA. Does not distinguish between past/present infection. Being replaced by HCV-RNA Liver Biopsy
Hepatitis C Serology HCV-RNA by RT-PCR. –Most sensitive test –Diagnose acute infection prior to seroconversion –May be intermittent (neg does not mean no disease) –Qualitative and quantitative tests –Response to therapy
Hepatitis C NO immunization No post exposure prophylaxis Chronicity common Different genotypes respond differently to therapy
Other Hepatitis Viruses Hepatitis D (Delta). –Due to ssRNA virus. –Always associated with Hepatitis B. –Acute or chronic. –Often severe, high mortality. Hepatitis E. Due to ssRNA virus. –Rare, occurs in endemic areas.
Chronic Hepatitis HBV – 5-10% of acute infections HCV - >70% of acute infections HDV – with HBV coinfection or superinfection –Elevated aminotransferases for >6 months –My lead to cirrhosis, hepatocellular ca –Liver transplant may be indicated
Review of Hepatitis Serology Excellent website with graphic representation of each type of viral hepatitis, case studies (click on Training Resources, then Viral Hepatitis Online Serology Training)
HIV testing ELISA antibody (screening) Western Blot (confirmation) RNA PCR (viral load) CD4 count and %
LYME Antibodies Lyme ab. IgM and IgG, screening with ELISA testing. Confirm with WB Poor sensitivity/specificity IgM – 2-4 wks post infection, decline by 4-6 months IgG – 4-8 wks post infection, high for months or years Must correlate clinically
Antigen testing Tests for the actual infectious agent Example: Some Hepatitis testing Rapid antigen tests: Rapid strep, Rapid flu, C. diff, etc. Test for a protein or other marker on the bacteria or virus, not a full culture
Stool Testing O&P, will determine if there are parasites present in feces. May need more than one sample, not always shedding. Culture: tests for Salmonella, Shigella, Campylobacter, E.coli 0157 WBC, occult blood (Guiac) C. diff - rapid antigen test
Viral Cultures Viruses are slow to grow in culture medium, may take weeks for a result. Therefore, serology is utilized more often. May see herpes culture ordered to confirm outbreak.
Bacterial Cultures Sterile vs nonsterile site Normal flora Aerobic vs anaerobic Gram stain is routinely performed on cultures from certain sites: sputum, wound, CSF, etc. Urine culture is semiquantiative others isolate bacterial colonies
Common Specimens Abscess, pus, fluid – swab or aspirate Blood– special vacutainer Body fluids – (amniotic, pericardial, peritoneal, pleural, synovial) – needle aspiration CSF – lumbar puncture Cutaneous -skin or nail scrapings, swab of infection Ear - middle ear myringotomy, outer ear swab Eye – swab conjunctiva, corneal scrapings FB – IV cath tips, prosthetic heart valves, IUD, etc. GI – Gastric biopsy for H. pylori, rectal swab, stool for O&P, culture Genital – cervical, urethral, vaginal secretions or swab. Resp – sputum, lavage, nasopharynx/pharynx swab Tissue – biopsy Urine – clean catch, cath, suprapubic aspirate
Bacterial Culture – Gram stain Done quickly Only on certain sites Need to correlate with clinical picture Results will be verified by culture in hrs, but can start empirically on antibiotics Report: Gram +/-, shape, other cellular elements (WBC, epithelials, etc)
Gram Stain: G+
Gram Stain: G-
Bacterial Cultures Most streak for isolation Plated on specific media for site of culture Grown in appropriate environment Will only be grown anaerobically if requested
Streak for isolation
Bacterial Cultures Sensitivities if bacteria is a pathogen Antibiotics tested vary from lab to lab, depending on g+ or g- Certain species do not have sensitivities performed. Ex: Strep, usually sens to penicillins so no sensitivities performed Hospital antibiogram – common bacteria and their susceptibilities
Alpha, Beta Hemolysis
Nosocomial Infections Originate in hospitals Account for tens of thousands of deaths per year 5-10% of hospitalized patients Due to: –Prevalence of pathogens –Compromised hosts –Effective transmission
Nosocomial Infections Primary pathogens: –Enterococcus (VRE) –E. coli –Pseudomonas –Staph Aureus (MRSA) –C. diff Acquire antibiotic resistance Become normal flora for hospital workers Common sites: urinary tract, wounds, respiratory, skin, blood, GI
CA MRSA Community acquired MRSA Athletes, children, military recruits, close living quarters Not hospitalized or in long-term care
WBC overview Viral vs. bacterial infection –Viral: lymphocyte response (T, B or NK). May have slightly elevated or suppressed total WBC count –Bacterial: Neutrophil response with early forms (bands). Often higher total WBC Sepsis –Neutropenia or neutrophilia, immature forms