The document is a detailed overview of Shigella and Salmonella, highlighting their morphology, classification, pathogenesis, clinical manifestations, and laboratory diagnosis. It discusses Shigella as an agent of bacillary dysentery and Salmonella's role in enteric fevers, including typhoid and non-typhoidal infections. Treatment options and diagnosis methods, including culture and serological testing, are also covered.

1. SHIGELLA AND SALMONELLA
Ms. Clemencia Tjazuko
MSc. Medical Microbiology

2.  Morphology
 Short gram-negative rods
 0.5 by 1-3 µm in size
 Non-motile
 Non-sporing
 Non-capsulated
 Fimbriae may be present
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3. Shigella
 Agent of bacillary dysentery
 Dysentery- infection of the intestines which causes
the passage of bloody stools mixed with mucus.
 Named after Japanese microbiologist Kiyoshi
Shiga, who isolated the first member 
S.dysenteriae in 1886 from epidemic dysentery
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4.  Antigenic Structure
 Simple
 ‘Major’ antigens and ‘Minor’ somatic antigens
 Some have K antigens
 Fimbrial antigens
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5. Classification
 Classified into four species based on combination
of biochemical and serological characteristics
 Serotypes distinguished within the species
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6. 1. S.dysenteriae (Group A)
 Does not ferment mannitol
 15 serotypes
 Serotype 1- S.shigae
 Only shigella producing shiga toxin (exotoxin)
 Only member of enterobacteriaceae that is catalase
negative
 Indole negative
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7.  Serotype 2- S.schmitzi
 Forms indole
 Ferments sorbitol and rhamnose
 Serotype 3-7
 Formerly Large-Sachs group
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8. 2. S.flexneri (Group B)
 Based on specific type antigen
 6 serotypes
 Complex antigenically
 Serotype 6
 Indole negative and has three biotypes
 Manchester, Newcastle and Boyd 88
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9. 3. S. Boydii (Group C)
 19 serotypes
 Isolated least frequently
 Resemble S.flexneri biochemically but not
antigenically
 Described by Boyd (1931) from India
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10. 4. S. Sonnei (Group C)
 Sonne (1915), Denmark
 Ferments lactose and sucrose late
 Indole negative
 Antigenically homogenous
 2 forms: Phase I and Phase II
 Phase II colonies are larger, flatter and irregular
 Mildest form of bacillary dysentry
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11.  Toxins
1. Endotoxin
 May be due to the LPS of Gram-negative cell wall
 Has irritating effect on intestinal wall which causes
diarrhoea and subsequently intestinal ulcers
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12. 2. Exotoxin
 Produced by S.dysenteriae type 1
 Also called Shiga toxin or verotoxin
 Acts as enterotoxin and neurotoxin
 Inhibit protein synthesis
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13. 3. Verocytotoxin (VT)
 Cytotoxin produced by S.dysenteriae type 1
 Acts on vero cells
 VT1 and VT2
 VT1 two subunits A and B.
A binds cytotoxin to cells
B inhibit protein synthesis
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14. Pathogenesis of Shigella
 Mode of transmission:
infection through contaminated fingers
contaminated food/water
flies (rare)
transmitted sexually (homosexuals), gay bowel
syndrome
 Minimum Infective Dose:
10-100 bacilli
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15.  Entry via M cell:
Bacilli enter mucosa via M cell
Cross the basolateral side of M cells to reach submucosa where
they are engulfed by macrophages
Bacilli released by macrophages
Cytokines released which attract inflammatory cells to infected
site inflammation acute collitis
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16.  Invasion:
Inside submucosa, shigellae induce own uptake
into adjacent epithelial
Determined by large virulence plasmid, codes
for ipa proteins and Type III secretion system
 Direct cell-to-cell spread:
from one host cell to the other by inducing actin
polymerisation cells of host
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17. Bacillary dysentery
 Short incubation period (1-7 days)
 Clinical features:
 Frequent passage of loose, scanty feces containing
blood and mucus, with abdominal cramps and
tenesmus
 Complications:
 Arthritis, toxic neuritis, conjunctivitis, parotitis,
intussusception in children
 Hemolytic uremic syndrome in severe cases
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19. Laboratory Diagnosis
 Specimen
 Isolating bacillus from feces
 Fresh feces inoculated or transported in Sach’s
buffered glycerol saline or Gram-negative broth @
pH7.0-7.4
 Rectal swabs not satisfactory
 Gram-negative broth sodium deoxycholate to
inhibit gram-positive bacteria, subcultured 6-8hrs
after incubation
 Selenite F Broth/Salmonella Shigella (SS) broth
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20.  Microscopy- plenty of pus cells, erythrocytes and
macrophages
 Culture- best to use mucus flakes
 MacConkey, DCA/ XLD
 Incubation @ 37 ºC, plates inspected for pale or pink-
coloured colonies, identified by biochemical reactions
 Non-motile bacillus that is : urease, citrate, H2S and
KCN negative further investigated by biochemical tests
 Confirmed by slide agglutination with sera
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21.  Salmonella Shigella Agar - colonies colourless
 Hektoen enteric (HE) Agar – contains bile salts as
inhibitory agents and some dyes, colonies appear
green with fading colour to the periphery
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22.  Treatment
 Acute cases infants and young children,
dehydration corrected propmptly
 Antibiotics reserved for severe toxic cases
 Many strains still sensitive to Nalidixic acid and
Norfloxacin
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23. Salmonella
 Morphology
 Gram-negative rods,1-3 by 0.5 µm in size
 Fimbriae – present
 Motile with petrichate flagella
 But S.Gallinarum and S.Pullorum are non-motile
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24. Clinical Classification
1. Typhoidal Salmonella
 Serotypes S.Typhi and S.Paratyphi restricted to
human hosts causing enteric fever
(typhoid/paratyphoid fever)
2. Non-typhoidal Salmonella
 Remaining serotypes are the food poisoning
group, infect human beings causing
gastroenteritis, septicemia or localised infections
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25. Antigenic Classification
 Kauffmann-White scheme
 Based on the presence of different somatic (O) and
flagellar (H) which is detected by agglutination with
respective antisera
 Based on O antigen:
 Serogroup 2
 Serogroup 4
 Serogroup 9
 Within each group further diffrentiation by Phase 1
and Phase 2 flagellar antigens
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27. Antigenic Structure
Flagellar antigen H
Somatic antigen O
Surface antigen Vi
H antigen
Present on flagella, heat labile
Destroyed by boiling or alcohol
H suspensions agglutinate rapidly to form large, loose,
fluffy clumps
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28.  Strongly immunogenic, induce antibody formation
 Occurs in two phases
O antigen
 Phosholipid-protein-polysaccharide complex
 Identical to endotoxin
 Heat and alcohol stable
 67 O antigens
 Less immunogenic
 O suspensions form compact, chalky, granular clumps
with antisera
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29. Vi antigen
 Surface polysachharide envelope or capsular antigen
covering the O antigen
 Inhibits phagocytosis, resists complement
activation, bacterial lysis
 Heat labile, Poorly immunogenic
 Present bacilli is inagglutinable with O antisera.
Becomes agglutinable after boiling or heating at 100
ºC for 1hr
 S.Paratyphi C
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31. Antigenic Variation
 H-O Variation
 Associated with the loss of flagella. Induced by:
Phenol agar: inhibits flagella
Mutation: Non-motile mutant of S.Typhi 901-O
strain subcultured in Craigie’s tube
 Phase variation:
Phase 1: serotype specific (a, b,c etc)
Phase 2: nospecific/group antigens (1,2, etc)
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32.  V-W Variation
 V forms of S.Typhi isolates with Vi antigens are
agglutinated by Vi antisera but not O antisera
 After serial subcultures, Vi antigen is lost W
forms agglutinable by O antisera but not Vi
antisera
 Also VW forms agglutinable with both
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33.  S-R Variations
 Smooth to rough variation, due to loss of of O
antigen side chain from LPS leading to exposure of
core polysaccharide portion
 Smooth colonies produced by virulent strains
 Rough strains form large, round and irregular
colonies, avirulent due to loss of O antigen
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34. Pathogenesis of Salmonella
 S.Typhi and S.Paratyphi A, B, C  Typhoidal
salmonellae
 Cause Enteric fever also known as Typhoid fever
 Infective Dose:103 – 106 bacilli
 Risk factors: stomach acidity, intestinal integrity
 Entry through epithelial cells (M cells): lines the
mucosa. Salmonellae cause formation of
membrane ruffles. Ruffles enclose adherent
bacteria bacteria mediated endocytosis (BME)
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35.  Mechanism of BME: Type III secretion system
delivery of bacterial proteins into cytoplasm of
epithelial cells alters actin cytoskeleton required for
bacterial uptake
 Entry into macrophages: Salmonellae containing
vacuoles cross epithelial layersubmucosa
phagocytosed
 Survival inside macrophages:
 S.Typhi alters its surface so that it is no longer
suspectible to lysosomal enzymes of macrrophages
 Encode Type III secretion system that delivers bacterial
proteins into macrophage cytoplasm
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36.  Primary bacteremia: Salmonellae inside
macrophages spread via lymphatics to enter blood
stream
 Spread: disseminate throughout liver, spleen,
lymph nodes and bone marrow; and gallbladder,
kidneys and lungs further multiplication
 Secondary bacteremia: from clinical organs
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38. Clinical Manifestations
 Gradual onset with headac he,
malaise,anorexia, coated tongue and abdominal
discomfort with constipation or diarrhea
 Soft, palpable spleen. Hepatomegally
 Rashes/Rose spots that fade on pressure
 Complications: gastrointestinal bleeding and
intestinal perforation in third or fourth week of
illness
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39.  Carriage
 10% of untreated patients become carriers and excrete
S.Typhi in urine or feces
 Two types of carriers:
 Fecal carriers: bacilli multiply in gall bladder and
excreted in feces. Common
 Urinary carriers: multiplication in kidneys and bacilli
excreted in urine
 Tyhoid Mary-chronic carrier, a New York cook who
caused 1300 cases outbreaks
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40. Laboratory Diagnosis
 Specimen and Culture Isolation
 Blood and Bone marrow culture:
 In first week of illness in blood culture bottle
(BHI broth, automated BACTEC)
 Stool Culture (3-4 weeks of illness)
 Enrichment broth such as Selenite F Broth,
tetrathionate broth and gram-negative broth
 MacConkey Agar NLF
 DCA, XLD agar, SS agar and Wilson Blair’s
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43.  Urine Culture
 In 3-4 weeks of illness
 On MacConkey agar
 Culture Smear and Motility
 Motile, gram-neg bacilli
 Biochemical Identification
 Catalase positive and oxidase negative
 Nitrate reduced to nitrite
 Indole and Urease negative
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44.  Slide Agglutination
 Confirm the serotype
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45.  Serum Ab detection (Widal Test)
 2-3 weeks of illness, Abs detected against TO, TH,
AH, BH antigens
 S.Typhi infection: high TO and TH Abs
 S.Paratyphi A infection: high TO and AH Abs
 S.Paratyphi B infection: high To and BH Abs
 Antigen detection (serum and urine)
 ELISA, CIEP
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46.  Molecular Methods
 PCR detecting flagellin gene, iroB, and fliC
gene
 Antimicrobial testing
 Done on Mueller-Hinton Agar
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47. Assignment due 22 Feb 2017
1. To which family does Shigella and Salmonella
belong to? What are the general properties of
that family?
2. Name the Non-Typhoidal Salmonellae
species? Write short notes about what
diseases they cause
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