Bacillus
Laboratory Diagnosis • Specimen Fluid or pus from local lesion, blood, sputum • Microscopy • Culture In septicemic anthrax, blood culture should be done • Serological test • Animal inoculation
Microscopy • Large aerobic, non motile, Gm+ve bacilli • Arranged singly, in pairs or in short chains, the entire chain is surrounded by a capsule • Capsules are produced in the presence of bicarbonates or 10-25% CO2 • Spores are oval and centrally located, non bulging • Spores are stained by special stains – Sudan black B.
Microscopic features Staining blood films with polychrome methylene blue: - Pink amorphous material around blue bacillus (M’ Fadyean’sreaction): represents capsular material used for the presumptive diagnosis of anthrax in animals.
Cultural Characteristics • Grow on blood or nutrient agar, at 37°C • Irregular, round, raised, dull, opaque, greyish white colonies with a frosted glass appearance. • Low power – edge of the colony is composed of long, interlacing chains of bacilli, resembling locks of matted hair – “Medusa Head Appearance” • Gelatin stab culture – “inverted fir tree” appearance, with slow liquefaction starting from top.
Inverted fir tree • Medusa Head Appearance • wavy colonies with small projections
Cultural Characteristics • “String of Pearls reaction” – • solid medium containing 0.05-0.5 units of Pn/ ml, in 3-6 hrs the cells become large, spherical and occur in chains on agar surface, resembling a string of pearls. differentiates B. anthracis from B. cereus • Selective medium – PLET medium
Smear from colony Morphology in stained smears from cultures • “Bamboo stick appearance” : bacilli arranged end to end in long chains.
Laboratory Diagnosis • Animal inoculation By rubbing contaminated tissues over shaven skin of a guinea pig • Serology • Ascoli’s thermoprecipitation test –to demonstrate anthrax Ag in tissue extracts • EIA(using purified anthrax toxin Ag) • PCRto detect anthrax contamination of animal & agricultural products
Resistance • Bacilli destroyed at 60°C in 30 mins. • Animal carcasses – bacilli remain viable in BM for a wk & in skin for 2 wks. • Spores – highly resistant, survive in soil for 60 yrs • Spores can be destroyed by • 4% KMnO4 in 15 mins • ‘Duckering’ – using formaldehyde solution for animal products imported into non endemic countries
Duckering • For disinfection of wool – 2% soln of formaldehyde at 30- 40°C for 20 mins • Animal hair & bristles – 0.25% at 60°C for 6 hrs
Prophylaxis • General methods of prevention • Improvement of factory hygiene • Proper sterilisation of animal products • Animal carcasses to be buried deep in quicklime or cremated
Prophylaxis • Active immunisation of • Domestic animals with live attenuated spore vaccines • Persons with occupational risk (butchers, farmers, veterinarians) with a cell- free vaccine containing purified protective antigen as immunogen. 3 doses IM with annual booster injections. * Anthrax infection in humans give life long permanent immunity & secondary infections are very rare.
Anthrax vaccines • Original anthrax vaccine – developed by Pasteur – live attenuated bacilli vaccine – strain rendered avirulent by the loss of plasmids which encodes anthrax toxin • Live attenuated anthrax spore vaccine • Sterne vaccine – contains spores of a noncapsulated avirulent mutant strain - loss of plasmid which controls capsule production • Mazucchi vaccine – containsspores of stable attenuated Carbazoo strain
Bacillus cereus • Readily isolated from soil, vegetables and a wide variety of foods including milk, cereals, spices, poultry & meat. • Causes foodborne gastroenteritis – 2 patterns of disease (diarrhoeal & emetic); both types are mild & self limited, requiring no specific therapy.
Bacillus cereus clinical presentation Gastroenteritis DIARRHOEALFORM EMETIC FORM Incubation period > 6 hours Diarrhoea Lasts 20-36 hours Incubation period < 6 hours Severe vomiting Lasts 8-10 hours
Type I Wide range of foods including cooked meat & vegetables Diarrhoea & abdominal pain develops 8 –16 hrs after consumption Few bacilli seen in fecal specimens Caused by serotypes 2,6,8,9,10 or 12. Enterotoxin resembles LT of E.coli Type II Chinese fried rice exclusively. Acute nausea & vomiting 1-5 hrs after meals, diarrhoea rare Large no of bacilli in cooked rice & fecal samples. Caused by serotypes 1,3 or 5 Toxin resembles staphylococcal enterotoxin Types of Gastroenteritis
Diagnosis • Primarily depends on clinical diagnosis & food sources • Laboratory Diagnosis • Specimen – stool, vomitus, food, blood • Microscopy – not of much help • Culture • Test for toxin – to differentiate from staphylococcal food poisoning.
Culture • Blood agar • Special MYPA medium: Mannitol - egg yolk - phenol red – polymyxin agar : to isolate B.cereus from feces & other sources.
