Spore Forming Bacilli: Features and Importance

In this article we will discuss about Spore Forming Bacilli which causes Anthrax, a Zonotic Disease in Animals and Malignant Pustules or Wool Sorter Disease in Man:- 1. Morphology and Staining of Bacilli 2. Cultural Characteristics of Bacilli 3. Biochemical Reaction 4. Resistance 5. Antigenic Structure 6. Toxin 7. Pathogenicity and Clinical Features 8. Laboratory Diagnosis 9. Immunity 10. Epidemiology and Other Details.

Contents:

1. Morphology and Staining of Bacilli
2. Cultural Characteristics of Bacilli
3. Biochemical Reaction of Bacilli
4. Resistance
5. Antigenic Structure of Bacilli
6. Toxin
7. Pathogenicity and Clinical Features of Bacilli
8. Laboratory Diagnosis of Bacilli
9. Immunity of Bacilli
10. Epidemiology of Bacilli
11. Prophylaxis of Bacilli

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1. Morphology and Staining of Bacilli:

Gram-positive, straight, rectangular (4-8 x 1-1.4µ ) bacilli arranged in long chains with square ends and central spores (Fig. 42.1), they are non-motile. In blood and tissue, they exhibit a distinct capsule composed of D-glutamic acid which, when stained with polychrome methylene blue, is purplish and the bacillus is blue — this is known of Methylene blue reaction of Mc Fadyean.

2. Cultural Characteristics of Bacilli:

Bacillus anthracis is aerobe and facultative anaerobe, optimum temperature 37 C, grows well on all ordinary media; on nutrient agar, colonies are white, granular, circular disks which—under the low power of the microscope—show a wavy margin like locks of hair — “medusa head” appearance; On agar stroke, the growth is of ground glass appearance; in gelatin stab, “inverted fir-tree” growth appears haemolysis on blood agar medium is uncommon with Bacillus anthracis, but common with the saprophyte bacilli.

3. Biochemical Reaction of Bacilli:

It has little value. Glucose, sucrose and maltose and fermented with acid production only. B. anthracis are catalase positive and reduce nitrates to nitrites.

4. Resistance of Bacilli:

(a) Vegetative forms are killed at 56 C in 30 minutes.

(b) Spore forms (Figs. 42.1, 2) resist dry heat at 140 C for 1-3 hour and boiling and steaming at 100 C for 5-10 min. They remain viable in environment in dry state for years.

Spores are destroyed by:

1. Chemicals (mercuric chloride, 1 in 1,000); potassium permanganate solution, 4%; formaldehyde, 10%) in half an hour.

2. Autoclaving at 121 C (15 lbs./in²) for 15 minutes.

B. Anithracis are sensitive to several antibiotics penicillin, streptomycin, erythromycin, tetracycline and chloramphenicol. Since these antibiotics have no effect on “Anthrax toxin complex“, it is necessary to administrate immuno serum along with antibiotics.

5. Antigenic Structure of Bacilli:

B. anthracis have three main antigens:

I. Capsular antigen is present in virulent strains and consists of a polypeptide D-glutamic acid which acts as a hapten.

II. Cell wall antigen is a polysaccharide.

III. Somatic antigen is present in tissue or oedema fluid of anthrax lesion; it is heat labile protein. It is protective antigen, it stimulates immunity, the antibody thus produced is protective.

6. Toxin:

From culture filtrate (anthrax toxin), three substrates have been separated by glass filtration and chromatography:

1. Protective antigen (a protein),

2. oedema factor;

3. toxic factor;

Mixture of 1, 2 and 3 is more toxic in animals and more immunogenic than single substances. It is known as “Anthrax toxin complex”.

The toxic complex is lethal and produces intensive vascular permeability and oedema which results into death of infected animal. Besides, this toxin has, probably, an affinity for cells of mononuclear phagocytic system. The capsular antigen inhibits opsonisation and phagocytosis. The production of toxin by B. anthracis is controlled by a plasmid, loss of which renders the strain avirulent.

7. Pathogenicity and Clinical Features of Bacilli:

The infection is usually acquired by the entry of spores through injured skin and mucous membrane, rarely by inhalation of spores in the lung. The spores germinate in the tissue at the site of entry and the growth of the vegetative organism results in the formation of a gelatinous oedema and congestion.

Men (farmers, veterinarians) acquire the infection from contact of the infected animals. The resulting lesion is malignant pustule in which the papule changes into vesicle, then pustule and, finally, into a necrotic ulcer which is responsible for the dissemination of infection resulting into septicaemia.

Another type of anthrax is inhalation anthrax (Wool sorter disease). The inhalation of anthrax spores from the dust of wool, hair or hides results in the germination of spores in the lungs and the production of pneumonia, meningitis and sepsis which are rapidly fatal. In anthrax sepsis, the number of organisms in the blood is more than 10⁷/ml just before the death.

8. Laboratory Diagnosis of Bacilli:

A. Specimens:

Fluid or pus from local lesion, blood sputum.

B. Smears:

Anthrax bacilli occur in long chains of large Gram-positive bacilli and can be identified by immunofluorescence staining technique.

C. Culture:

On blood agar, they are non-haemolytic, grey colonies. On nutrient agar, medusa head appearance colonies.

In semi-solid medium, they are non-motile but saprophytic bacilli are motile. Virulent anthrax culture kills mice or guinea pig upon intra-peritoneal injection.

D. Ascoli test:

Extract of infected tissues show a ring of precipitate when layered over immune serum.

E. Serological tests:

Precipitating or haemagglutinating antibodies can be demonstrated in the patient serum.

Treatment:

Many antibiotics are effective.

9. Immunity:

Immunity is produced and the “toxigenic complex” acts as immunogen when susceptible animals are vaccinated with the attenuated B. anthracis. Alum precipitated toxoid prepared from the protective antigen has been successfully used in occupational ‘risk groups’. It is administrated intramuscularly in three doses at intervals of 6 weeks and 6 months. After one year a booster may be given.

A “Sterne vaccine” has been developed containing spores of a non-capsulated a virulent mutant strain of B. anthracis.

A “Mazucchi vaccine” contains spores of stable attenuated Carbazoo strain of B. anthracis in a preservative of 2% saponin. The animal is protected for a year with a single injection of spore vaccine which has been widely used with good result. But it is unsafe for human use.

Treatment:

Penicillin is the drug of choice. The case fatality of malignant pustule has been brought down from 20% to 5% by the treatment with penicillin and streptomycin. Antibodies are of no use when the toxin is formed. Scalvo immune serum—prepared in ass by active immunization—can neutralize toxin septicaemia; can be treated with antibiotics and together with immune serum.

10. Epidemiology of Bacilli:

Anthrax is primarily a disease of animals transmitted secondarily to human beings (Zoonosis). WHO reported in Asia and African many agricultural anthrax which were contacted by the animals after grazing the pasture contaminated with the anthrax spores.

These spores may enter the oral mucosa damaged by dry sharp grass and germinate in the wound into vegetative forms liberating toxin thus causing Anthrax (enzootic anthrax). In 1980, in Zimbabwe, during epizootie of anthrax, about 9,000 human cases were reported.

Industrial anthrax is due to contaminated imported hides, hairs, bone meals which can be diagnosed by Ascoli’s test.

11. Prophylaxis of Bacilli:

1. Animal died of Anthrax should be disposed off by burning or deep burial in a lime pit at a depth of 6 feet (2 meters).

2. During epizootie, all animals should be protected by live attenuated vaccine.

3. Animal products (wool, horse, hair etc.) should be disinfected.

4. During outbreak, antibiotics along with immune serum should be administrated to person at risk.