Vibrio Cholerae - Bacteriology - Lecture Slides, Slides of Bacteriology

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Vibrio cholerae

Key Words

• Vibrio cholerae
• Late lactose fermenter
• Oxidase positive
• String test
• Hemodigestion
• Choleragen (cholera toxin)
• Rice water stool
• Fish in Stream appearance
  • Darting motility
  • Cholera red reaction
  • TCBS
  • Transport media
  • Classical Vs El Tor
  • Endemic
  • Severe Dehydration
  • Halophilic vibrios

Cultural characteristics

• Strongly aerobic but can grow anaerobically.

• Grows at 16-40 C optimum 37 C

• pH 6.4 – 9.6 optimum 8.

• NaCl is required for growth at 0.5-1%

• Grows on ordinary culture media

• Nutrient agar – Moist translucent disc like colonies,

1-2 mm diameter with bluish tinge in transmitted

light. Distinct odour present.

• MacConkey’s agar – Initially colourless later become

pink due to lactose fermentation (late lactose

fermenter).

Cultural characteristics

• Blood agar – Initially surrounded by zone of

greening, later the zone is clear due to

hemodigestion.

• Gelatin stab culture – Infundibuliform

liquifaction in 3 days at 22 C.

• Peptone water – In six hours fine surface

pellicle is formed. Turbidity and powdery

deposit later.

Enrichment media

• Alkaline peptone water at pH 8.

• Monsur’s taurocholate tellurite peptone water

at pH 9.

• Both these are good transport as well as

enrichment media

Plating media

• Alkaline Bile Salt Agar (BSA) –

• pH 8.2, widely used. Colonies like on Nutrient agar.

• Monsur’s Gelatin Taurocholate Trypticase Tellurite

Agar (GTTA) –

• Small translucent colonies with greyish black centre and

turbid halo. The colonies become 3-4 mm in 48 hrs.

• Thiosulphate Citrate Bile Salt Sucrose (TCBS) medium

• Widely used. Yellow colonies turning green on further

incubation.

Biochemical reactions

• Catalase & Oxisase tests are positive.
• Glucose, Mannitol, Maltose, Sucrose fermented with acid production.
• Lactose fermented late.
• Indole formed & nitrates reduced to nitrites. These two give cholera red reaction. Few drops of Conc. H 2 SO 4 added to 24 hr peptone water culture. Development of reddish pink colour due to production of nitroso-indole indicates presence of V. cholerae.
• Methyl Red & urease tests negative. - Catalase + - Oxidase + - G A - M A - M A - S A - L Late A - I + - MR _ - U _ - Cholera red +

Resistance

• Susceptible to heat, drying & acids.

• Can survive for 2-3 days on linen in dry

condition.

• Survive in tap water for 30 days, cold storage

food, for 30 days.

• El Tor strains survive better than classical

strains

• Susceptible to disinfectants including chlorine.

Classical Cholera Vs El Tor Vibrio

Test Classical cholera El Tor vibrio

Hemolysis (^) - +

Voges Proskauer (^) - +

Chick erythrocyte

agglutination

Polymixin B sensitivity@ (^) + -

Group IV phage

susceptibility

El Tor phage V

susceptibility

@ 50 iu disc

O serotypes of Cholera vibrios

SEROTYPE O ANTIGENS

Ogawa AB

Inaba AC

Hikojima ABC

Cholera toxin (CT) - Choleragen

• B binds to gangliosides
• provides channel for A
• A catalyses ADP-ribosylation
  • regulator complex
  • activates adenylate cyclase

CT determined by chromosomal gene.

Each molecule has 1 A & 5 B subunits

A – Active B - Binding

B attaches to GM1 ganglioside receptor of eneterocyte

Allowing entry of A subunit in enterocyte

Splits into A1 & A

allows only biologically active A1 to be

linked with B

Prolonged activation of cellular

adenylate cyclate and accumulation of

cAMP

Outpouring of large qty of water + electrolytes + Carbonates + K Diarrhea

A1 – Active A2 - Linking

Laboratory Diagnosis

• Specimen collection –

– Stool collected in acute stage before

administration of antibiotic

– Best collected by No. 26 or 28 rectal catheter

– Rectal swabs may be used particularly in

convalescent patients.

– Grossly – “Rice water stool”

– Not suitable

• Specimen from pans
• Vomitus

Transportation

• Whenever possible, specimen plated at bedside and

inoculated plates sent to the lab.

• Preserve at 4 C or in holding medium whenever delay is more

than few hours

 Cary – Blair medium  VR fluid

 If delay is only few hours, transport medium can be used

 Monsur’s medium  Alkaline peptone water

 If transport media are not available, filter paper strips can be

soaked in stool sample, sealed in plastic envelopes and sent to

lab.