In this article we will discuss about Yersinia Pestis in Humans:- 1. Morphology and Staining of Yersinia Pestis 2. Cultural Characteristics of Yersinia Pestis 3. Biochemical Reaction 4. Resistance 5. Antigenic Structure and Toxins 6. Bacteriocines and Bacteriophages 7. Epidemiology 8. Yersinia Pseudo-Tuberculosis and Yersinia Entero-Colitica 9. Y. Pseudo-Tuberculosis 10. Y. Enterocolitica and Other Details.
Contents:
- Morphology and Staining of Yersinia Pestis
- Cultural Characteristics of Yersinia Pestis
- Biochemical Reaction of of Yersinia Pestis
- Resistance of Yersinia Pestis
- Antigenic Structure and Toxins of Yersinia Pestis
- Bacteriocines and Bacteriophages of Yersinia Pestis
- Epidemiology of Yersinia Pestis
- Yersinia Pseudo-Tuberculosis and Yersinia Entero-Colitica
- Y. Pseudo-Tuberculosis
- Y. Enterocolitica
- Pathogenicity
- Pasteurella Multocida (P. Septica)
- Laboratory Diagnosis of Yersinia Pestis
1. Morphology and Staining of Yersinia Pestis:
Y. pestis is short oval bacillus with rounded ends—i.e. coccobacillary—1.5 µ * 0.7 µ, occurring singly and in pairs. In the tissue, a capsule may be formed; in cultures grown at 37°C capsule may be demonstrated by India Ink method.
It is Gram-negative and, when stained with Wayson’s stain or methylene blue, it shows typical bipolar staining which is an important feature of metachromatic granules. It is highly pleomorphic, non-motile and non-sporing.
2. Cultural Characteristics of Yersinia Pestis:
Y. pestis grows aerobically and anaerobically on ordinary media (blood agar, MacConkey’s agar) at an optimum temperature of 27°C. Colonies on agar are at first very small, transparent, white, circular disks (1 mm or less) later becoming larger (3-4 mm) and opaque.
If cultured in a flask of broth with drops of sterile oil or ghee on the surface without shaking the flask, a characteristic growth develops like stalactites hanging down in the fluid from the oil-drop. On Cary Blair transport medium (Fig. 5.2(b)), Y. pestis can survive for at least 75 days. Sunlight drying kills Y. pestis quickly.
3. Biochemical Reaction of Yersinia Pestis:
Y. pestis is a non-lactose fermenter and ferments glucose, maltose, mannitol, with production of acid and no gas (Table 36.2). MR is positive, and catalase activity is more prominent in virulent than the avirulent strains. Indole is negative. On the basis of glycerol fermentation and nitrate reduction, Y. pestis is grouped in three epidemiological types, each one has a specific geographical distribution (Table 36.3).
4. Resistance of Yersinia Pestis:
The thermal death point for Y. pestis is 55°C for 15 minutes. It is destroyed by heat, sunlight, drying and chemical disinfectants (0.5% phenol in 15 minutes). It can survive in the soil of rodent (rat) burrow and is sensitive to gentamycin, streptomycin, tetracycline, chloramphenicol.
5. Antigenic Structure and Toxins of Yersinia Pestis:
Y. pestis recovered from man and rodents from different parts of the world are biologically identical and serotypes do not exist.
They have a complex antigenic structure and some 30 different antigens have been identified which include:
1. A heat labile envelope antigen (protein) produced at 37°C, inhibits the phagocytosis and is usually found in virulent strains. It is F1 (fraction 1) glycoprotein antigen complex virulent strains carry plasmid. The virulent or wild type Y. pestis produce VW antigen (somatic antigens) which are encoded by genes of plasmid.
2. Virulent strains of Y. pestis produce a bacteriocin (pesticin), coagulase and fibrinolysin. Coagulase is produced by Y. pestis at 28°C (body temperature of rat flea).
3. Exotoxins are of many types, one is lethal for mice in a dose of 1 microgram. This lethal toxin (protein in nature, M.W. 74,000) produce beta-adrenergic block and is cardio-toxic in animal.
4. Plague toxins found in culture filtrates are both exotoxins and endotoxins.
6. Bacteriocines and Bacteriophages:
Y. pestis liberate two types of bacteriocins (pesticin I and pesticin II). The production of pesticin I is plasmid mediated and inhibits the growth of Y. pseudo-tuberculosis, Y. enterocolitica and E. coli. Pesticin II is also produced by Y. pseudo tuberculosis and is active against certain strains of Y. pestis. Bacteriophage typing of Y. pestis is useful for identification.
Pathogenicity and Clinical Features:
Plague may occur mainly in two forms:
1. Pestis major (bubonic, septicaemic, pneumonic plague);
2. Pestis minor (ambulatory form with mild pyrexia and lymphadenitis).
Sylvatic Plague:
Occurs in wild rats.
When a host specific rat flea (Xenopsylla cheopis) attracted towards rat bites an infected rat, it ingests Y. pestis which multiply in the gut of the flea and produce a coagulase enzyme at 28°C (normal temperature of the flea) which helps the blocking of the proventriculus (stomach) so that the food cannot pass through.
The coagulase is not produced at 35°C. Hence the transmission via flea is low or absent in very hot weather. The low temperature of rat burrows is congenial for the survival of Y. pestis in the absence of fleas and rats.
The blocked, hungry flea becomes angry and bites the exposed parts (leg or hand) of the human body. The inoculated organisms are phagocytosed, carried to inguinal or axial lymph nodes. An intense haemorrhagic inflammation develops into enlarged lymph nodes which may necrose and become fluctuant.
The resultant mass is the primary “bubo” (1) Bubonic plague. Secondary buboes may develop in other lymph nodes. Y. pestis reach the blood stream and become widely disseminated. Haemorrhagic and necrotic lesions may develop in all organs, resulting into 2) Septicaemic, and 3) Pneumonic plague.
The last report of Plague in India (Karnataka) was in 1966. In 1994, there was an epidemic of pneumonic plague in Gujarat (Surat) and bubonic plague in Maharashtra (Mamala, Beed, Majalgaon), India.
An incubation period of 2-7 days is followed by high fever, painful lymphadenopathy (greatly enlarged tender lymph nodes—”buboes” in the groin and axilla), vomiting, diarrhoea, hypotension, altered mental status, renal and cardiac failure.
Signs of pneumonia and meningitis can appear terminally and Y. pestis can be seen in blood smear. Massive involvement of blood vessels may result into purpuric lesions in the skin—known as “black death.”
Laboratory Diagnosis:
Blood is taken for culture and lymph node aspirate for smear and culture. Sputum culture for pneumonic plague, Cerebrospinal fluid (CSF) for culture and smear in possible meningitis, stained smear will show Y. pestis with bipolar deep staining. Y. pestis can be grown on blood agar and MacConkey’s agar.
All cultures are highly infectious and must be handled with care. Experimental animals should be deeply buried with lime. The disease can be contracted if the infected material comes in contact with broken skin (bite or scratch).
A presumptive diagnosis can be confirmed in about 80% of cases by fluorescent antibody staining. Microscopical examination of buffy coat smear of peripheral blood may show Y. pestis in septicaemic cases.
Antigen Detection:
F1 glycoprotein antigen complex may be detected in sputum and aspirated fluid from bubo by immunofluorescence and ELISA test.
Serological Tests:
Antibodies to F1 antigen appear towards the end of first week of the illness and may be detected by haemagglutination or complement fixation tests which are useful to identify plague foci as the test remains positive for several years after recovery from plague. It has a limited use in the laboratory diagnosis as the antibody levels (1:16 or more) reach only after 2-3 weeks following onset of disease.
Treatment and Prophylaxis:
Tetracycline is the drug of choice, which is followed by streptomycin, sulphonamides. Haffkine killed plague vaccine is recommended and is available at Haffkine Institute, Parel, Mumbai, its immunity lasts for 6 months; live attenuated vaccine may provoke unacceptable reactions.
Rat fleas can be killed by insecticides, though there is the possibility to develop the resistance to the insecticides. The rat hole/burrows may be fumigated by formaldehyde gas to destroy Y. pestis which survive at low temperature of rat holes, as well as rats and fleas by suffocation due to fumigation.
7. Epidemiology of Yersinia Pestis:
Eradication of plague is never possible as it is disease of the earth, of creatures that run and burrow and of the fleas that live on them.
The disease persists in natural foci because of latent infection in resistant hosts during hibernation, by prolonged survival of Y. pestis in soil of rodent burrows and by survival of infected fleas.
When sylvatic rodents spread the infection into urban, rats via rat flea, the domestic plague may occur. This is the basic cycle of epidemic bubonic plague. Because of his occupation, man may get accidently infected when he comes in close contact with wild rodents.
Plague persisted in urban areas of different parts of the world during 1980-87.
8. Yersinia Pseudo-Tuberculosis and Yersinia Entero-Colitica:
These two organisms differ from Y. pestis morphologically; rodents, birds, wild and domestic animals are reservoirs of infection. They sometimes cause gastroenteritis with abdominal pain and fever in man. They differ from Y. pestis in the following aspects—they are non-capsulated, motile at 22°C (not at 37°C), urease positive, oxidase negative and are not lysed by Y. pestis bacteriophage (Table 15.2).
9. Y. Pseudo-Tuberculosis:
On the basis of somatic and flagella antigens they are distinguished into serological groups and nine serotypes have been distinguished. Antigenically they cross-react with Y. pestis and Salmonella. Infection occurs in animals through alimentary tract in enzootic and epizootic forms.
Multiple tubercle like lesions are seen in liver, spleen and lungs of infected animals, hence it is named as Y. pseudo tuberculosis. Most probably, man gets infected via ingestion of water contaminated with animal faeces.
10. Y. Enterocolitica:
On the basis of cultural and biochemical characters, they are distinguished into five serotypes and they are antigenically distinct: 50 O serotypes and 19 H factors have been identified. Serotypes 03, 08 and 09 are mostly responsible for human infection. There is serological cross-reaction between 09 and Brucella contaminated food may probably cause human infection.
11. Pathogenicity:
The roots of infection is by alimentary tract. The bacteria multiply in the mucosa of the intestine during the incubation period of 5-10 days. The symptoms are fever, abdominal pain and diarrhoea. The severe watery to bloody stool is probably due to an enterotoxin or invasion of mucosa by bacteria which may progress to septicaemia in individuals with lowered body resistance.
Y. enterocolitica are susceptible to antibiotics. Systemic infection has high mortality. They can be isolated from faeces, blood, mesenteric lymph nodes. There is a rise of antibody titre.
Epidemiology occurs through contaminated food or water or via faeco-oral route.
12. Pasteurella Multocida (P. Septica):
They are non-motile, Gram-negative (0.7 x 0.4 µ m) with bipolar staining resembling Y. pestis. They fail to grow on MacConkey’s agar medium. They are Oxidase and Indole positive. They cause chicken cholera and haemorrhagic septicaemia (very common in India) in various animals.
They are usually normal inhabitants of the respiratory tract of animals. Similarly, they occur in the throat of humans. Human infection is rare causing septic wound after cat or dog bite, meningitis after head injury. Penicillin is effective.
Other pasteurella species (P. haemolytica, P. pneumotropica, P. ureae) differ from P. multocida in several aspects. Their pathogenic role in man is yet to be known.
13. Francisella Tularensis:
It causes tularaemia, a zoonotic disease, in rabbits (mainly) in USA. Human infection is by handling or eating infected rabbits or drinking contaminated water. They enter the host via skin or mucous membrane through the arthropod bite or via respiratory or gastrointestinal tract.
After an incubation period of 3-5 days, an ulcerating papule develops at the primary site of penetration on finger or hand. The regional lymph node enlarges and suppurates like bubos of plague. There is bacteriaemia with headache, chills and high fever. The organisms are seeded in various organs with granulomatous lesions which ultimately undergo necrosis.
14. Laboratory Diagnosis of Yersinia Pestis:
Examination of smear from blood, pus, sputum will reveal Gram-negative cocco-bacillus (0.7 x 0.2 µ m) with capsule which can be demonstrated by India Ink method.
The isolated cultures in blood-glucose-cysteine can be tested by
(a) Biochemical reactions
(b) Specific agglutination test
(c) Fluorescent antibody staining.
Intra-peritoneal injection of bacilli into guinea pig may cause death of animal.
Serology:
A titre of 1: 80 or above can be considered positive, provided there is no brucella agglutinins since this organisms shares somatic antigen with Br. abortus and Br meltensis. The skin test is specific.
Epidemiology:
Tularaemia is worldwide in distribution.
Prophylaxis and Chemotherapy:
A live vaccine prepared from avirulent strain can be used. Streptomycin is the drug of choice.
