Module 1 Flashcards by Taylor Horlock

What types of nonselective media are there?

Chocolate agar- RBCs are lysed and release nutrients, supplements are added (supplement B, Iso-Vitatex)

Selective Neisseria media but with out antimicrobials (specific for Neisseria)

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What is the purpose of selective media?

Prevents the overgrowth of contaminating normal flora

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What types of selective media are there?

Modified Thayer Martin (MTM)

New York City (NYC)

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What does MTM agar contain? What does each do?

Vancomycin- inhibits G+

Colistin- inhibits G- and no pathogenic Neisseria

Nystatin- inhibits yeast

Trimethoprim lactate- inhibits Proteus spread

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What does NYC agar contain?

Vancomycin

Colistin

Amphotericin B- inhibits yeast

Trimethoprim lactate

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What type of media is ideal for isolation of vancomycin resistant strains of Neisseria?

Combo of selective and nonselective media

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What types of transport media are there?

Non nutrient holding media- without growth, Amie’s transport medium, up to 6hrs

Growing transport media- used when there’s a delay, container with CO2 generating system, more recovery of N. gonorrhoea, vancomycin resistant strains may not be isolated

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What is the genus test for Neisseria and Moraxella? How does the test work?

Oxidase positive

Amine turns positive when oxidized by enzyme

Reagent- dimethyl or tetramethyl-para-phenylenedianire (Kovac’s oxidase reagent ~1%)

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Why can oxidase reagent only stand for a few hours after prep? How can it be stabilized?

Readily oxidized by oxygen

Ascorbic acid- reduces sensitivity

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What are the methods for performing the oxidase test?

Filter paper method- reagent on paper, rub colonies on

Alternate procedure- drop of reagent on colonies

Positive- purple

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What can cause false positives or negatives in oxidase testing?

False neg- old colonies, inhibitory/selective media, carb utilization media

False pos- nichrome wire, MAC

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What is the QC for the oxidase test?

PC- Neisseria or Moraxella catarrhalis

NC- Staph, Strep, Enterobacteriaceae

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What are the cellular characteristics of N. gonorrhoea?

G- cocci, kidney beans in pairs, 0.8-1.0um

Intracellular G- cocci

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What are the growth requirements for N. gonorrhoea?

Obligate aerobes

Capnophiles (3-7%)

35°C

Moist environment required

Enriched media required (chocolate agar)

72hrs of incubation

Colonies- 0.5-3mm, grey, translucent, entire, gummy, older colonies tend to stick

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What tests are used for Neisseria species ID?

Carb utilization- determines what carbs bacteria use, phenol red detects acid produced by oxidative mech

Chromogenic substrate test- detects specific enzymes, Gonochek II

Ag-Ab tests- confirmatory, coagglutination, direct fluorescent Ab, test for Protein I

Gene probes- detects unique NA sequences, ssDNA probe, positive = high RLU

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What are the ID results for N. gonorrhoea?

Carb utilization- glucose +

Chromogenic substrate test- no colour change, reagent added, red-pink colour (proline aminopeptidase enzyme)

Coagglutination with Protein I antisera (N. gonorrhoea)

What causes false negatives in carb utilization tests?

Extended incubation can cause reversion and loss of acid pH

Utilization of peptone in the presence of O2

What sugars are used for reference in carb utilization testing?

CTA sugars

What rapid carb utilization test kits are there?

QuadFerm+kit

API NH

What colours indicate what antibiotics in chromogenic substrate tests?

Blue- N. lactamica (beta galactosidase)

Yellow- N. meningitidis (glutamylaminopeptidase)

No colour, reagent, red-pink- N. gonorrhoea (proline aminopeptidase enzyme)

What can cause false negatives in Ag-Ab testing?

Some strains lack Protein I

What is the antimicrobial susceptibility of N. gonorrhoea?

Originally all isolates were sensitive to penicillin

B lactamase resistant cephalosporins are effective (ceftizoxime and ciprofloxacin)

How is penicillin resistance caused in N. gonorrhoea?

Plasmid- codes for B lactamase, PPNG

Genetic info on chromosome- changes the structure of PBPs and outer membrane permeability (resistance without B lactamase)

What is the pathogenicity of N. gonorrhoea?

STI- urethral inflammation, dysuria, purulent discharge, prostatitis and epididymitis, salpingitis, endometriosis, peritonitis, infertility

Rectal infection

Purulent conjunctivitis

Occasionally spreads to bloodstream- DGI

What are the growth requirements of N. meningitidis?

Aerobic Capnophilic (3-7%) 50% humidity 35°C Nutrient media not recommended, good growth on MTM and NYC SPS anticoagulant is inhibitory to some strains

What are the colonial characteristics of N. meningitidis?

Moist Smooth May be mucoid No hemolysis Translucent Older colonies tend to autolyse

What are the species ID results for N. meningitidis?

Oxidase + Carb- glucose + and maltose + Coagglutination and latex reagents Polyvalent meningococcal antisera

What is the antimicrobial susceptibility of N. meningitidis?

Most strains are susceptible to penicillin Resistance due to plasmid induced B lactamase

What is the pathogenicity of N. meningitidis?

Nasopharynx of ~5% of healthy individuals Children and young adults are most susceptible Petechial rash Elevated WBC in spinal fluid Fulminate form AKA Waterhouse Friderichsen syndrome Endocarditis, pneumonia, arthritis, conjunctivitis

What are the characteristics of other Neisseria species?

G- cocci in pairs, not usually intracellular O2 required, but not increased CO2 Grows well on nutrient agar without enrichment, don't grow on selective media Dry, hard, wrinkled colonies

What is the species ID for other Neisseria species?

Oxidase + N. lactamica- glucose + maltose + lactose + N. species- may be positive for sucrose, negative for all others

What are the cellular characteristics of Moraxella catarrhalis (M. cat)?

G- cocci in pairs, resist decolourizing Found intra and extracellularly

What are the growth conditions of M. cat?

Aerobe, doesn't require increased CO2 35°C Grows on nutrient agar without enrichment (optimum growth on blood and chocolate agar)

What are the species ID results of M. cat?

Strongly oxidase and catalase positive Carb utilization- doesn't produce any acid Lipase pos DNase pos (Neisseria are neg) Butyrate esterase test- pos (most Neisseria neg)

What is the antimicrobial susceptibility of M. cat?

Most strains are resistant to vancomycin, colistin, trimethoprim and clindamycin Suitable antimicrobials- amoxicillin with clavlanate, cotrimoxazole, erythromycin, tetracycline, second gen cephalosporins ~80% produce B lactamase

What is the pathogenicity of M. cat?

Normal respiratory flora Otitis media and maxillary sinus infection Conjunctivitis, pneumonia, bronchitis, tonsillitis, endocarditis, meningitis, septicaemia

Why generas belong to the family Neisseriaceae?

Acinetobacter- G- rods Kingella- G- coccobacilli Neisseria- G- cocci Moraxella- G- cocci and coccobacilli