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What are the biochemical reactions used to ID E. coli?
TSI-
KIA-
Gas-
H2S-
MR-
VP-
IND-
CIT-
PDA-
URE-
MOT-
LDC-
ADH-
ODC-
ONPG-
TSI- A/A
KIA- A/A
Gas- P
H2S- N
MR- P
VP- N
IND- P
CIT- N
PDA- N
URE- N
MOT- P
LDC- P
ADH- V
ODC- V
ONPG- P
What are the reactions used to presumptively ID E. coli?
hemolytic, LF with diffuse bile salts, green sheen on EMB, oxidase negative, spot indole positive
What is the difference between an outbreak, epidemic, and pandemic?
Outbreak-- rise in incidence of a disease localized by a common factor, usually contained
Epidemic-- increased incidence and spread compared to an outbreak, usually regional
Pandemic-- increased incidence and spread compared to an epidemic, usually international
What is the difference between sensitivity and specificity?
Sensitivity detects true positives (few false negatives)
Specificity detects true negatives (few false positives)
--these terms describe the test method itself
What are PPV and NPV?
PPV = positive predictive value (positive results correlate to disease)
NPV = negative predictive value (negative results correlate to no disease)
--these terms describe the relationship between test method and disease
What is meant by the term “beta strep”?
these are the strep species classified by Lancefield group, related by beta hemolysis on SBA
--S. pneumo and viridans Strep are alpha streps, S. pyogenes and S. agalactiae are beta streps
What diseases are caused by GAS and GBS beta streps? What disease is caused by viridans alpha strep?
GAS (S. pyogenes)-- strep throat, ear infection, deep soft tissue infection, Childbed Fever, TS-like syndrome, glomerulonephritis
GBS (S. agalactiae)-- meningitis, Childbed Fever, cystitis, pyelonephritis
Alpha strep (viridans)-- endocarditis
What virulence factors are associated with GAS? (there are 10 to know)
M proteins
lipotechoic acid
hyaluronic acid capsule
hyaluronidase
C5a peptidase
pyogenic (erythrogenic) toxins
streptokinase
streptodornase (DNase B)
streptolysin O
streptolysin S
What virulence factors are associated with GBS? (there are 4 to know)
C5 peptidase
beta hemolysin/ cytolysin
capsule
hyaluronidase
What is the principle of the Lancefield ID grouping test?
Nitrous acid is used to extract the carbohydrate Lancefield Ag from the cell wall of the bacteria. Latex particles are coated with strep Ab and mixed with the extraction. Granular agglutination occurs when the Ab and Ag correspond.
How is the Bacitracin disk used to differentiate beta streps?
Group A strep are sensitive to Bacitracin and resistant to SXT. Group B strep are resistant to both Bacitracin and SXT. (Bacitracin resistance = Group B)
How is the CAMP test used to differentiate beta streps?
Group B strep secretes CAMP factor (a protein) that interacts with beta hemolysins from S. aureus producing enhanced/synergistic hemolysis
How are Staph species distinguished? (2 tests)
Catalase positive (strep catalase neg)
Coagulase negative (micrococcus coag pos)
How does S. aureus acquire resistance? (In VRSA, MRSA, Oxacillin Résistant)
VRSA mechanism is acquisition of vanA gene from enterococci, also use it for Vanc ressistance
MRSA mechanism is hyperproduction of beta lactamases via blaZ (inducible beta lac, methicillin specific, modifies drug binding to PBP sites) and mecA (induce PBP2a prod at low levels)
Oxacillin Resistant SA mechanism is like mecA,/PBP2a called mecC
What toxin is produced by coag neg staph? (general)
Staphylokinase-- digests clots, cleaves IgG and C3b, inhibits phagocytosis
What diseases result from staph toxins?
Scalded Skin Syndrome-- mediated by exfoliative (epidermolytic) toxin
Food Intoxication-- mediated by heat-stable enterotoxin, causes cytokine release and vagus nerve stimulation
What tests are used to help ID S. aureus?
Catalase, Coagulase or Rapid ID, Novobiocin resistance, Morphology (yellow)
How is S. pneumo identified?
Catalase negative, *bile soluble, optichin susceptible
What diseases are associated with S. pneumo?
Meningitis, pneumonia, ear/eye infection, necrotizing fasciitis, associated with splenectomies
What are the virulence factors associated with S. pneumo?
M proteins, lipotechoic acid, hyaluronic acid capsule, hyaluronidase, C5a peptidase, pyogenic (erythrogenic) toxins, streptokinase, streptodornase (DNase B), streptolysin O, streptolysin S
How is Pseudomonas aeruginosa identified?
Morphology, smell, oxidase positive, beta hemolytic
What is the biochemical profile for Pseudomonas aeruginosa?
Catalase, nitrate, certamide positive
Non-fermenting
K/NC or K/K on TSI
What is the specific name for the oxidase reagent? What is the principle of this test?
tetramethyl-penyldiamine-dihydrochloride (oxidase acts on this to form indophenol-- ie, the substrate acts as artificial final electron acceptor)
What are the roles of hyaluronidase, collagenase, and DNAse?
Hyluronidase and collagenase aid in tissue breakdown, DNase acts as a “thinner” to release trapped bacteria
What are the general roles of virulence factors? (8/9 to know)
as enzymes, as siderophores, anti-complement activity, anti-phagocytic (prevent, kill, or survive), antibody resistance, immunosuppression, adhesion, as endo or exo toxins
How do superantigens work?
also called type 1 exotoxins
superantigens bind to host cell membrane receptors (not translocated into cell) > produce a signal to act within target cell cytoplasm (contact-dependent mechanism); specifically bind MHC II on APC and T Cell-receptors; activate T Cell response at very high rates (overstimulation);
cellular overstimulation can result in fever/cardiovascular shock as too many cytokines are released and Ag response is uncoordinated; can lead to lysis of MHC II expressing cells and toxicity to host cells; can induce immune tolerance
What are the 3 common superantigens?
associated with toxic shock (usually staph), staph exotoxin (food poisoning), strep exotoxin (scarlet fever)
Which organisms require CO2 for growth?
A(H)ACEK organisms (Agregattibacter (Haemophilus) aphrophilus, Agregattibacter (Actinobacillus) actinomycetemcomitans, Cardiobacterium hominis, Eikenella corrodens, Kingella kingae)
Haemophilus influenzae
What is the difference between antisepsis, disinfection, and sterilization?
Sterilization kills all forms of microbial life in areas or on objects (ex, autoclave)
Antisepsis eliminates microorganisms in living tissues (ex, iodine, alcohol)
Disinfection eliminates microorganisms on inanimate objects (ex, bleach)
When are specimens rejected? (3 cases minimum)
C. diff-- reject on children under 2, be strict (treating carriers without true clinical infection increases MDR pathogens, VRE/recurrent)
Anaerobes-- only accept normally sterile body fluids, blood, pus/aspirates from abscesses and deep wounds, tissue
Sputum-- can reject based on gram stain when more than 10 SECs per lpf (10x) are observed
What are the common CSF pathogens by age group?
Infants 6-12 moz-- N. meningitidis, S. agalactiae, Listeria monocytogenes, E. coli K1, Coronobacter sakasakii, Chryseobacterium meningisepticum
Children-- H. influenzae type B, S. pneumoniae
Young adults-- N. meningitidis
Adults over 30-- S. pneumoniae
Misc-- S. aureus, GNRs
What tests are commonly performed on CSF pathogens?
Rapid IDs available for Neisseria sp, Haemophilus sp, perform beta lactamase testing on both
Haemophilus will not grow on BAP
Coag, Bile solubility, Optichin, Bacitracin, CAMP, Hippurate, Strep typing all options for staph and strep
Hemolysis on BAP used to classify strep
Listeria: hippurate, motility, bile esculin, catalase, CAMP, beta hemolysis
What are the symptoms associated with N. meningitidis meningitis?
flu-like, myalgias, arthralgias, nuchal rigidity (50% cases), rapid development of rash (50-60% cases)-- pink maculopapular eruption becoming petechial, starts on mucous membranes and moves to trunk (indicates bleeding complications)
Which pathogens are associated with wound infections?
Primary infections-- beta strep esp GAS, S. aureus, GNRs, anaerobes, Candida albicans, Corynebacterium
Secondary infections-- P. aeruginosa, S. aureus, Erysipelothrix rhusiopathae
Bite wounds (animal)-- Capnocytophaga, Pasturella multicoda
Bite wounds (human)-- Eikenella corrodens
“Jungle rot”-- Burkholderia cepacia
Anthrax, Tetanus
Gas gangrene-- Clostridium perfringens
What are the virulence factors of S. aureus?
afimbrial adhesion
requires damage to skin for SST infection (not normally invasive)
catalase, hyaluronidase, DNAse, beta lactamase
prevents phagocytosis with capsule, protein A, leukocidins, coagulase
What are the toxins of S. aureus?
exfoliatin superantigen-- induces fever, rash, desquamation in Scalded Skin Syndrome
hemolysins
What diseases are associated with S. aureus? (2 classes)
Toxin-mediated: Scalded Skin, food poisoning, Toxic Shock Syndrome
Not toxin-mediated: abscesses, wound infection, pneumonia, septicemia (endocarditis, osteomyelitis, meningitis), impetigo
What causes Erysipeloid?
Erysipelothrix rhusiopathae
How is Erysipelothrix rhusiopathae IDed?
Regular, short GPRs
Catalase negative
Nonmotile, H2S positive, Hippurate and BE negative
What bacteria causes myonecrosis? What toxin is associated with the disease?
Clostridium perfringens
Alpha Toxin (CPA)-- phospholipase, attacks cell membrane, primary damage occurs from this
Beta Toxin (CPB), Epsilon Toxin (ETX), Iota Toxin (ITX)-- lyse RBCs, digest connective tissue, kill soft tissue
What is hidendritis suppurativa?
a recurrent infection of apocrine sweat gland bs (groin, axillary), multiple infections can lead to tissue fibrosis, sinus tract formation, disfigurement, caused by staph, strep, GNRs, anaerobes
How is anthrax transmitted?
via direct contact with infectious material or close contact with animals-- a small break in the skin can introduce organism or can travel over mucosal surface
How does Bacillus anthraxis survive in the body? (What are its virulence factors?)
capsule (resists phagocytosis), PA (protective Ag)
EF (edema factor), LF (lethal factor); EF+LF increases intracellular cAMP, inhibits phagocytosis, blocks oxidative burst, promotes macrophage apoptosis
What is the colony morphology of Bacillus anthraxis?
Nonhemolytic, ground glass
How is Bacillus anthraxis ruled out in the lab?
gamma hemolytic, appearance, GPR
catalase positive, nonmotile
How does anthrax progress if untreated?
incubation 2-5 days, initial lesion of pruritic papule > develops into hemorrhagic vessel that ruptures > black-bottomed central eschar forms w/surrounding edema > satellite lesions may begin > death occurs from pulmonary obstruction
What is the difference between food poisoning and gastroenteritis?
food poisoning results from ingesting contaminated food or water, gastroenteritis is an infection in the GI tract
Which GI bugs utilize toxins?
S. aureus (heat stabile enterotoxin), Bacillus cereus (heat labile and heat stabile enterotoxins), Botulism (simple AB toxin), Clostridium botulinum, Clostridium perfringens (toxin A), Helicobacter pylorii (vacuolating toxin), Shigella sp (enterotoxin, cytolethal distending toxin, shiga toxin), EHEC (shiga-like toxin), Vibrio cholerae (cholera, ZOT, ACE toxins), Vibrio parahemolyticus, Aeromonas hydrophila, Plesiomonas shigelloides
Why is honey dangerous to infants?
potential for botulism infection via spore ingestion (not a problem for adults, must ingest preformed toxin)
How are toxin-producing GI bugs treated?
vaccines, antibiotics, antitoxins (Ab drug) in some cases
What are the Salmonella and Shigella serotypes?
Salmonella: typhii ID is based on agglutination with Vi antiserum, non-typhii serotypes are based on O and K antigen antisera
Shigella: serotype based on O antigen (Kauffman-White scheme), S. dysentariae (group A), S. flexneri (group B), S. boydii (group C), S. sonnei (group D)
How are pathogens and NF differentiated on GI media?
look for NLF on MAC and/or NLF with H2S on HE, XLD
How are stool screen results interpreted?
What are the common stool pathogens?
Shigella, Salmonella, Aeromonas, Plesiomonas, Yersinia
How does Vibrio infection commonly occur? What disease is associated with this infection?
From eating raw oysters, swimming with new tattoos
Associated with liver disease
How is H. pylori IDed?
can be diagnosed using Urease Breath Test--swallow urea pill, check for CO2 in breath (increased bc urease broken down into CO2 and ammonia), correlates with infection (culture usually overrun with NF)
What are the clinically relevant subtypes of E. coli?
What is EHEC?
enterohemorrhagic E. coli, aka E. coli O157H7, produces Shiga Toxin, also associated with HUS, hemorrhagic colitis– ID uses growth as non-sorbitol fermenter on MAC w/ Sorbitol
What is ETEC?
enterotoxigenic E. coli, causes cholera-like disease
What is EPEC?
enteropathogenic E. coli, serotypes O55 and O111, causes mild/mod diarrhea with high mortality in undev countries
What is EAEC?
enteroaggregative E. coli, causes mild persistant diarrhea in children, produces heat stabile toxin but does not cause HUS
How is Campylobacter IDed?
Skirrow’s Media aka Campylobacter agar used– only Campylobacter, Pseudomonas, Enterococcus, & Staph sp. will grow
Gull-wing GNRs
microaerophillic
oxidase positive
hippurate positive for Jejuni, negative for Coli
How are the different oxidase positive GI pathogens differentiated?
What growing conditions are used for UGT samples?
Media reduces normal vaginal flora growth, incubated in high-humidity CO2, held 72 hours
What organisms are associated with Urethritis, Vaginitis, Cervicitis?
yeast, Group B Strep (S. agalactiae), enteric GNRs, Neiserria gonorrhoeae
What organisms are associated with bacterial vaginosis?
Gardnerella vaginalis, Lactobacillus
What organisms are associated with PID?
Chlamydia trachomatis, Neisseria gonorrhoeae, anaerobic organisms (Peptococcus, Peptostreptococcus, Bacteroides), Mycoplasma and Ureaplasma, intestinal coliforms
What organisms are associated with Purepurial Sepsis?
prev pyogenes (GAS) now caused by S. agalactiae (GBS)
What is the KOH cervical exam?
used to diagnose fungal infection
When does disseminated gonorrhea occur?
in patients with complement deficiency
What is Rheumatic fever?
autoimmune reaction where Ab are produced against M protein of strep are cross-reactive to connective tissue of valves and proteins in synovial fluid
What is the name for the lesions associated with rheumatic fever?
erythema magnum lesions on trunk, arms, legs
What are the HACEK organisms?
Agregattibacter (Haemophilus) aphrophilus, Agregattibacter (Actinobacillus) actinomycetemcomitans, Cardiobacterium hominis, Eikenella corrodens, Kingella kingae
What diseases are associated with the HACEK pathogens?
Aggregatibacter aphrophilus – subacute native valve bacterial endocarditis
Aregattibacter actinomycetemcomitans -- lumpy jaw, endocarditis
Cardiobacterium hominis -- endocarditis
Eikenella corrodens -- endocarditis
Kingella – septic arthritis in kids
What lab tests are used to ID HACEK organisms?
fastidious GNRs, slow growing (48-72 hrs on BAP and CHOC, 2 weeks in blood cultures), require CO2 and enriched media, do not grow on MAC
Which systemic organism forms rosettes in liquid media?
How is significance determined in blood cultures?
What is the pathogenesis for the plague?
organism spreads in lymph to lymph nodes, groin and armpit are most common first sites bc transmission bites are most common on extremities > “buboes” may burst and release organisms into circulation, migrate to liver, spleen, lungs (organisms multiplying in blood, called septicemic plague), high fever, dark purple skin, 100% fatal > may also multiply in lung macrophages, leads to tissue damage and hemorrhage –necrosis caused by DIC, death occurs from overwhelming septicemia and endotoxin shock (weeks if bubonic, days for pneumonic)
Why are leuconostoc and pediococcus important?
they are environmental pathogens that cause endocarditis in immunocompromised patients
What antibiotic resistance do leuconostoc and pediococcus have?
Vancomycin
What is added to blood cultures in order to inactivate antibiotics and prevent clotting?
SPS (sodium polyanetholesulfonate)
What causes catheter related blood infections?
Staphylococcus aureus, Bacillus sp., GNR’s & Corynebacterium (rare, from skin), Acinetobacter, Pseudomonas, Stenotrophomonas, & Candida (from healthcare workers)
What are the parameters for significant growth on a catheter blood culture?
the same organism should be recovered from all sets
if only cath is positive indicates tip infection only
the cath culture may be positive for two organisms, but only that which matches the PB culture and patient symptoms is significant
What are the non-micro tests used for sepsis monitoring and prediction?
lactic acidosis > 4mmol
hypotension, renal failure, altered mental state, acute respiratory failure
nonspecific sepsis indicators– CBC, ESR >100 mm/hr, respiratory alkalosis – pH ↑; PCO2↓, metabolic acidosis – pH ↓; Bicarb decr, HS, CRP, IL-6 & IL-8, TNF-α, Lipopolysaccharide binding protein (LBP)
Ferritin used as an exclusionary test– if elevated not sepsis
procalcitonin should be increased 2-4 hours after bacterial or endotoxin challenge (serum lactate also used to diagnose but rises later)
monocyte distribution width also indicates sepsis–
molecular assay ideal, requires positive blood culture first
What should be done if an organism does not grow on SBA, but it appears not to be an anaerobe?
it might be ? Starts with A
Streak plate with S. aureus, will grow satellite colonies due to NAD production
When do blood cultures become positive with upper resp. tract infections?
What organism class is always considered pathogenic?
GAS
What type of selective media is used for respiratory samples?
BCYE - Buffered Charcoal Yeast Extract Agar for Legionella pneumophil
Chocolate with Bacitracin for Haemophilus
Mannitol Salt Agar for Staphylococcus aureus
What does it mean if columnar epi cells are present on a gram stain?
the sample is good
Diptheria?
What specific media is used for samples from CF patients (sputum)?
FPBL aka Cepacia agar
Which pathogens are uniquely significant in CF patients?
Which organism is associated with the formation of smelly crusts around the nose? What disease is this?
Klebsiella pneumoniae spp. ozaenae
Atrophic Rhinitis
Which organism causes Scarlet Fever? What are the symptoms of this disease?
GAS
flushed face, sore throat, fever, chills, swollen glands in neck, Pastia’s Lines, strawberry tongue
What organisms commonly cause UTIs?
lower UTIs caused by E. coli, S. saprophyticus, S. epidermidis, S. agalactiae
Which organism causes alkaline encrusted cystitis?
urease producing bacteria (Proteus, Corynebacterium)
Which drug classes are inactivated by beta lactamases?
Penicillin, Cephalosporin
Which drug classes are inactivated by ESBLs?
extended spectrum penicillin, 3rd/4rd gen cephalosporin
Which drug classes are inactivated by AmpC beta lactamases?
Cephalosporins only
When is a D-test indicated?
performed on Methicillin Resistant Staphylococcus species that appear resistant to erythromycin but sensitive to clindamycin
Note 
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