Laboratory MedicineInfectious Disease

Brenda Beckett, PA-C

Clinical Assessment II

Infectious Disease

Urinalysis Serum serologies (antibody testing)

– Hepatitis, others

Cultures: bacterial, viral Rapid antigen testing O&P Nosocomial infections CBC results with viral vs bacterial infection

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 1.010-1.025. 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 (Gilbert’s 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

Acute Viral Hepatitis Panel

HBsAg IgM anti-HBc Igm anti-HAV Anti-HCV EIA

Post Exposure Testing

Source Patient: – Anti-HIV – HBsAg – Anti-HCV EIA

Injured HCW: – Anti-HIV – Anti-HCV EIA – Anti-HBs

Review of Hepatitis Serology

http://www.cdc.gov/hepatitis/

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

Giardia

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 SpecimensAbscess, 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 24-

48 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

Urine culture

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

Beta Hemolysis

Alpha, Beta Hemolysis

Blood Culture

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