Download shigella -study material-2012

SHIGELLA 1
Shigella
The organisms of genus Shigella differ from other. Enterobacteriaceae in that they are exclusively parasites of
human intestine and other primates and cause bacillary dysentery in man characterized by the passage of loose
motion mixed with blood and mucous. The dysentery bacilli belong to the genus Shigella, named after Shiga
who isolated the first member of the group in 1896 from epidemic dysentery in Japan which was then called
Shigelia shiga.
Morphology
The organisms are Gram-negative, aerobic, non- motile, non-flagellate and noncapsulated bacilli, measuring 1-3
x 0.5 μm.
Culture
The organisms grow in ordinary medium like nutrient agar, in which the colonies are about 2 mm in diameter,
circular, convex, smooth and translucent. For their isolation from faeces, MacConkey’s agar or Deoxycholate
citrate agar (DCA) media are used. In DCA and MacConkey’s agar the colonies are colorless except Shigella.
sonnei which is a late lactose fermenter. In Shigella-Salmonella agar (S. S. agar) the colonies are colorless.
Viability
The thermal death point for Shigelia is 55°C for one hour. They are killed by 1% phenol in 30 minutes. They
remain viable in water and ice for 1-6 months.
Antigenic structure
Shigellae are differentiated on the basis of their polysaccharide component of lipopolysaccharide complex of the
somatic 0-antigens into four groups A, B, C and D. Some strains produce K antigens which are usually of B variety
and cover the 0-antigen. The K-antigen may or may not be visible as capsule.
Classification
Antigenically, shigellae are divided into four groups (A, B, C & D) on the basis of serological specificity of
polysaccharide component of the somatic 0-antigen. These main groups or species of Shigella are better
differentiated by a combination of their biochemical reactions and antigenic structure. Mannitol fermentation
reactions are important which distinguish Group A strains (mannitol-negative) from Groups B, C and D
(mannitol-positive).
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Biochemical Reaction
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Antigenic structure
Based on 0 antigens shigellae are differentiated into serotypes, identified by agglutination absorbed antisera.
The antigenic structure of the shigellae is complex.
Subgroup A (Sh. dysenteriae)
Sh.. dysenteriae is subdivided into 12 serotypes, of which serotype 1 is known as Shiga’s bacillus and serotype 2
as Schmidtz’s bacillus.
Subgroup B (S/i. flexneri)
The members of the group have complex structures
There are six main serotypes (1-6) of S/i. flexneri, each characterized by different specific antigen, each of which
is further subdivided into sub serotypes (e.g. la and lb).
Subgroup C (S/i. boydii)
Eighteen serotypes of Sh. boydii are recognized. Except serotypes 10 and 11, all are antigenically distinct.
Subgroup D (Sh. sonnei)
Sh. sonnei strains are antigenically homogeneous but they may undergo an antigenic variation that affects the
somatic antigens and has been referred to as phase or form variation.
Toxin
1. Endotoxin: All shigellae release endotoxin after autolysis. It is the polysaccharide lipoprotein complex of cellwall. Endotoxin shows irritating action on the intestinal wall which causes diarrhoea and subsequent intestinal
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ulcers.
2. Exotoxin : Sh. shiga (Sh. dysenteriae type I) in addition to endotoxin, produces a powerful exotoxin in very
small amount in intestine. The exotoxin is a heat labile protein and acts as (a) enterotoxin as well as
(b) neurotoxin. As enterotoxin, it acts upon intestinal mucosa causing transudation of fluid in the lumen and as
neurotoxin it damages endothelial cells of small blood vessels of the central nervous system which results in
neurological complications like polyneuritis, coma and. meningism. The exotoxin is not primarily a neurotoxin.
Strains of Sh. dysenteriae 1 produce a cytotoxic which is active on Vero cells and is called verocytotoxin or VT. VT
also acts upon human colon and ileal cells in tissue culture.
Pathogenesis
The term dysentery is loosely used to mean diarrhoea with passage of pus, blood and or mucous in the stool
accompanied
by
abdominal
cramps
and tenesmus.
The
severity
of
the
illness
in bacillary dysentery
depends
on
the
infecting
species
of Shigella. Sh. shiga (Sh. dysenteriae type I) causes most severe and fulminating form of dysentery usually
associated with toxaemia. The infections due to Sh. flexneri and Sh. boydii are less severe and prevalent in
tropical and sub-tropical countries including India. The Sonnei infection is more frequently seen in children.
The incubation period of the disease is usually less than 48 hours but varies between 1 to 7 days and infection
occurs by ingestion of contaminated food. The food or drink is contaminated by faeces of infective cases or that
of carriers through contaminated objects in their environment and also through flies. Shigellae are pathogenic in
much smaller dose than the salmonellae and as few as 10 to 100 bacteria can cause dysentery. The organisms,
after reaching the terminal ileum and colon, first attach themselves to the surface of epithelial cells and
gradually they are taken by the epithelial cells where they multiply. The distal parts of the colon are more
severely affected. Then they spread laterally into adjacent cells as well as to the lamina propria where
replication and bacterial colonization take place. There is necrosis of surface epithelial cells. The
lamina propria and submucosa develop an acute inflammatory reaction with formation of micro-abscesses on
mucosal surface predominantly by polymorphonuclear leucocytes, along with capillary thrombosis. The necrotic
epithelia become soft and friable and are sloughed out in patches forming superficial snailtractor serpiginous ulcers which lead to bleeding from mucosa. Endotoxin irritates the bowel causing diarrhoea.
The ulcer surface is then covered by a pseudo membrane made up of fibrin, leucocytes, cell
debris, necrosed epithelia and bacteria. The intestinal ulceration is due to the activity of the bacterial endotoxin.
With remission, granulation tissue fills the defect and the ulcers heal by regeneration of the mucosal
epithelium.
The organisms tend to remain localized in the gut-wall and usually do not cause bacteraemia. In addition
to endotoxin, Sh. shiga produces a potent exotoxin which causes toxaemia. The toxin causes transudation of
fluid from intestinal mucosa leading to severe diarrhoea (enterotoxic action) and by toxaemia damages the
vascular endothelium causing neurological complication like polyneuritis and myocarditis or joint effusions.
The colonic ulcers are small, superficial with shallow depth and the intervening mucosa is oedematous. They are
multiple and are covered by pseudomembrane. In fulminating infection due to Sh. dysenteriae type I, the entire
mucous membrane of large gut may be affected and may slough out causing death. Fortunately milder forms of
disease due to Sh. flexneri or sonnei is common.
Dysentery carriers
Healthy carriers may be found in some community. Carrier state is acquired after recovery from acute attack of
dysentery.
Laboratory diagnosis
Specimens (a) For microscopy and culture, fresh stool, mucous flakes, rectal swabs are collected under aseptic
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precaution, (b) blood, serum may be examined for demonstration of rising titre of agglutinin antibodies.
A. Microscopical examination
of
stool
Coverslip preparation in saline and iodine shows large number of pus cells with degenerated nuclei. R.B.C. and
macrophages. The presence of protozoa, cysts or helminthic ova is excluded. The normal bacterial flora is
considerably diminished. Fluorescent antibody technique is of some help in rapid diagnosis of a case but is not
usually practiced
B. Bacteriological examination
I. Culture
Media :a) MacConkey’s agar ,b) DCA , c) S-S agar, d) Selenite F broth
A loopful of material is inoculated on selective media like MacConkey’s agar or S. S. agar media. Selenite-F broth
(0.4%) is used as enrichment and transport medium which permits rapid growth
of enteric pathogens. Organisms from selenite-F broth are sub cultured in MacConkey’s agar after 24 hours
incubation at 37°C.
Colorless colonies appear on MacConkey’s agar medium after 12 to 18 hours incubation which are further
tested by smear examination, motility preparation and biochemical reactions. Shigellae are Gram-negative nonmotile bacilli.
Biochemical reactions
Urease, Citrate, H2S and KCN negative; indole and M R positive organism is suggestive of a shigella strain.
Sh. sonnei is a late lactose fermenter.
Slide agglutination test
It is done by using polyvalent antisera of three groups (A, B and C) of shigella and antiserum of
group D (Sh. sonnei), one by one against the isolate. Monovalent antisera are used for the group for which
agglutination has occurred with polyvalent antisera. Then type specific antisera for strains belonging to group A,
B or C is used for agglutination test.
Occasionally, agglutination may not occur due to masking of 0 antigens by K antigen, which can be removed by
boiling the bacterial suspension at 100°C for 60 minutes.
Colicin typing
It is done for group D strains.
Prevention and control
As man is the major source of infection and the disease is transmitted by “food, fingers, faeces and flies” from
person to person, adequate sanitation together with detection and treatment of patients and carriers must be
undertaken to control the infection..
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