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SALMONELLA

SALMONELLA. SALMONELLA. Tribe – Salmonellaea Enteric fever, gastroenteritis, septicemia Salmonella typhi – typhoid fever DE Salmon – American microbiologist Eberth (1880) , Gaffky (1884) isolated – Eberth–Gaffky bacillus 2000 serotypes – two groups:

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SALMONELLA

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  1. SALMONELLA

  2. SALMONELLA Tribe – Salmonellaea Enteric fever, gastroenteritis, septicemia Salmonella typhi – typhoid fever DE Salmon – American microbiologist Eberth (1880) , Gaffky (1884) isolated – Eberth–Gaffky bacillus 2000 serotypes – two groups: Enteric fever, S. Typhi, S. paratyphi A and B Food poisoning, animal parasites – gastroenteritis, septicemia

  3. MORPHOLOGY Gram-negative, non-sporing, non-capsulated 1–3 x 0.5 micron Fimbriae – present Motile, peritrichate flagella S. Gallinarum and S. Pullorum – non-motile

  4. CULTURAL CHARACTERISTICS Nutrient agar: convex, translucent, moist MacConkey agar: Non-Lactose fermenting colonies. Selective media: XLD agar: Red coloured colonies Deoxycholate Citrate agar (DCA): Colourless colonies Wilson and Blair medium: Colourless colonies with black centres: Metallic sheen in S.typhi

  5. CULTURAL CHARACTERISTICS Wilson, Blair bismuth sulphide S. Typhi – jet black colonies, sulphite to sulphide S. paratyphi A – green colonies S. paratyphi B – black colonies Salmonella Shigella agar Enrichment media – selenite F broth, tetrathionate broth

  6. BIOCHEMICAL REACTIONS Oxidase negative Catalase positive Ferment glucose Mannitol, maltose – acid and gas Lactose, sucrose – negative S. Typhi anaerogenic Indole –ve, MR +ve,VP −ve, citrate +ve (S. Typhi – citrate −ve) H2S +ve, except S. paratyphi A

  7. BIOCHEMICAL CHARACTERISTICS

  8. ANTIGENIC STRUCTURE Flagellar antigen H Somatic antigen O Surface antigen Vi H antigen – heat labile Destroyed by boiling, alcohol Strongly immunogenic Antisera–large loose fluffy clumps

  9. ANTIGENS O antigen – somatic Phospholipid, protein polysaccharide Heat and alcohol stable 67 O antigens Less immunogenic Antisera – compact, chalky, granular clumps

  10. ANTIGENIC STRUCTURE Vi Ag – surface Ag Heat labile acidic polysaccharide Virulence – inhibits phagocytosis, resists complement activation, bacterial lysis Similar Ag: S. paratyphi C, Escherichia, Citrobacter Epidemiological typing/carrier

  11. ANTIGENIC VARIATIONS H–O variation: Associated with loss of flagella phenol agar (1:800) – phenotypic change 901 O strain – stable, non-motile, mutant – Craigie’s tube, U tube Phase variation: Flagellar Ags two phases Phase 1 – specific/few species – a, b, c, d, etc. Phase 2 – non-specific/fewer – 1, 2, etc. Diphasic, monophasic – S. Typhi

  12. ANTIGENIC VARIATIONS V–W variation: V form agglutinate – Vi antiserum and not O. After subcultures ,Vi lost; W form – O antiserum; V–W both O and Vi S–R variation: Smooth to rough, loss of O Ag virulence, autoagglutinable Prevention – Use Dorsett’s egg, lyophilisation

  13. ANTIGENIC VARIATIONS Variation O Ag: Lysogenisation phages S. anatum-------serotype – 3,10 e, h;1, 6 | lysogenisation – phage 15 | S. newington------serotype – 3,15; e h:1,6 | lysogenisation – phage 34 S. minneopolis------serotype –3,15, 34; e, h: 1, 6

  14. CLASSIFICATION AND TAXONOMY OF SALMONELLA Old: Serotyping & biochemical assays used to name individual species within genus (e.g., Salmonella enteritidis, S. choleraesuis, S. typhi) Over 2400 O-serotypes : Kauffman-White antigenic scheme New: DNA homology shows only two species Salmonella enterica (six subspecies) and S. bongori Most pathogens in S. enterica ssp. enterica

  15. CLASSIFICATION Kauffman White scheme O antigen – 1, 2, 3 etc factors Factor 2 – group A Factor 4 – group B Factor 9 – group C

  16. SALMONELLA CLASSIFICATION Antigenic characterisation – Kauffmann White scheme SEROGRPS SEROTYPE ANTIGEN O H Phase I Phase II 2-A S. para A 1,2,12 a - 4-B S. para B 1,4,5,12 b 1,2 S. typhimurium 1,4,5,12 I 1,2 S. chester 4,5,12 e,h e,n,x 7-C S. para C 6,7(Vi) c 1,5 S. cholera –suis 6,7 c 1,5 8-C2 S. muenchen 6,8 d 1,2 9-D S. Typhi 9,12(Vi) d - S. enteritidis 1,9,12 g,m - S. Gallinarum 1,9,12 - - 10-E S. anatum 3,10 e,h 1,6

  17. CLINICAL CLASSIFICATION Enteric fevers: S. typhi S. paratyphi A S. paratyphi B S. paratyphi C Gastroenteritis: S. typhimurium S. choleraesuis S. enteritidis Septicemia: S. choleraesuis S. typhimurium S. enteritidis

  18. PATHOGENICITY Enteric fever – S. Typhi – typhoid S. paratyphi A S. paratyphi B Paratyphoid fever S. paratyphi C Clinical syndromes in humans: Enteric fever Gastroenteritis or food poisoning Septicemia

  19. ENTERIC FEVER Includes typhoid fever caused by S. Typhi, and paratyphoid fever caused by S. paratyphi A, B and C.

  20. ENTERIC FEVER ID: 50-103- 106 bacilli Ingestion – attachment microvilli – penetration lamina propria – resist intracellular killing – mesentric lymph nodes – multiply – thoracic duct – bloodstream (transient bacteremia – different organs – massive bacteremia – gall bladder – Payers patches

  21. EPIDEMIOLOGY ENTERIC FEVER • Person-to-person spread • No animal reservoir • Contamination with human faeces • Usual vehicle is contaminated water. • Occasionally, contaminated food (usually handled by an individual who harbours s. Typhi)

  22. INFECTIOUS DOSE • Typically about 1,000,000 bacteria • Much lower if the stomach ph is raised • Much lower if the vehicle for infection is milk product • Protects the bacteria in their passage through the stomach • An infectious dose of about 100 bacteria

  23. PATHOGENESIS • Virulence factors • Invasiveness • Endotoxin • Enterotoxin

  24. PATHOGENESIS ENTERIC FEVER • Bacteria invade mucosa or peyer's patches of small intestine (?M cells), • Pass into mesenteric lymph nodes where they multiply and • Then enter the blood stream via the thoracic duct • Primary bacteraemia cleared by RES,

  25. PATHOGENESIS ENTERIC FEVER • Bacteria multiply in RES cells and destroy them • Facultative intracellular parasites Secondary bacteraemia occurs and results in spread to other organs. • Infection of the biliary tract. • Multiplication in biliary tract leads to seeding the intestine with large numbers of bacteria.

  26. PATHOGENESIS ENTERIC FEVER • Involvement of intestinal lymphoid tissue may lead to necrosis and ulceration. • In untreated nonfatal cases, temperature drops in 3 to 4 weeks (onset on immunity)

  27. Ingestion of S. typhi Incubation period Hemorrhage; perforation Inflammation and ulceration of Peyer’s patches Small intestine Lymph nodes Bile Mesenteric Lymph nodes Cholecystitis; Carrier state gallbladder Thoracic duct Multiplication in macrophages in liver, spleen, kidney,and bone marrow Bile Fever; Relatively slow pulse; Enlarged liver And spleen; Rose spots; Normal or low WBC count Transient (primary) bacteremia Septicemia (second) Fever, malaise Signs and symptoms

  28. CLINICAL FEATURES Mild undifferentiated pyrexia – fatal fulminating disease Onset – headache, malaise, anorexia, coated tongue, abdominal discomfort – constipation/diarrhea Temperature – step ladder rise – first week, high – 7–10 days, falls by third/fourth week Physical signs – relative bradycardia, hepatomegaly, splenomegaly, rose spots – second/third week

  29. COMPLICATIONS Intestinal perforation Hemorrhage Circulatory collapse Cholecystitis, arthritis, abscess Periosteitis, nephritis Venous thromboses, peripheral neuritis

  30. EPIDEMIOLOGY Endemic in all parts of India Typhoid : paratyphoid : 10 :1 Paratyphoid B – rare Carriers – convalescent, temporary, chronic

  31. TYPHOID MARY Mary Mallon – New York cook – 15 years – 7 outbreaks – 200 persons affected Carriers – convalescent – 3 weeks–3 months – temporary >3 months – <1 year – chronic >1 year, 2–4% – fecal and urinary carriers

  32. LABORATORY DIAGNOSIS Blood culture Clot culture Feces culture Urine culture Other material – bone marrow, bile

  33. LABORATORY DIAGNOSIS Serology PCR-based tests Demonstration of circulating antigen Other tests

  34. BLOOD CULTURE Positive blood culture diagnostic 90% positive – first week 75% positive – second week 60% positive – third week 5–10 ml blood collected – venipuncture aseptically, inoculated to 50–100 ml of 0.5% bile broth

  35. BLOOD CULTURE After 24 hours’ incubation – bile broth subcultured on MacConkey agar Subculture repeated – 10 days – culture negative Castaneda’s method – biphasic medium Serotyping – slide agglutination National reference centre – CRI Kasauli

  36. CULTURE Clot culture – serum – Widal test, clot – streptokinase Feces culture – carriers DCA, XLD, Wilson Blair Urine culture

  37. WIDAL Tube agglutination test O agglutination – chalky granular deposit Felix tube H agglutination – fluffy, cottony deposit Dreyers tube

  38. DIAGNOSIS OF CARRIERS Fecal carriers – fecal culture, bile Urinary carriers – urine culture Vi antibody detection – 1:40 SEWER SWAB TECHNIQUE – gauze pads, millipore membranes National Salmonella Reference Centre – Central Research Institute, Kasauli

  39. TYPING METHODS BACTERIOPHAGE TYPING Craigie and Yen (1937) – bacteriophage Vi II, 97 Vi phage – S. Typhi Tracing epidemics Trends and patterns epidemiology National phage typing centre – Lady Hardinge Medical College, New Delhi A-E-A-India

  40. TYPING METHODS Other typing methods Nicolle’s complementary phage typing – Kristensen’s biotyping – xylose and arabinose Bacteriocin Antibiogram

  41. PROPHYLAXIS General measures – sanitation, protected drinking water Vaccine – TAB vaccine, S.Typhi 1000 million and S. Paratyphi A and B 750 million Dose – 0.5 ml SC 4–6 week interval

  42. PROPHYLAXIS Live oral vaccine – (typhoral) – stable mutant – S.Typhi strain Ty2a Dose – one capsule orally Vi vaccine – injectable – purified Vi polysaccharide antigen

  43. TREATMENT Rx – chloramphenicol, ampicillin, amoxycillin, streptomycin. Carriers – vaccine + combination therapy, cholecystectomy, pyelolithotomy, nephrectomy DRUG RESISTANCE – 1972 –chloramphenicol, Mexico and India

  44. Calicut, Kerala – drug-resistant plasmids 1980 – multi drug resistance (MDR) – ‘R’ factor – England – 1960 – S. typhimurium phage 29 Rx – fluoroquinolones (ciprofloxacin, pefloxacin, ofloxacin),third generation cephalosporins, ceftazidime, ceftriaxone Hospital cross-infections – neonates – septicemia TREATMENT

  45. SALMONELLA GASTROENTERITIS Zoonotic first food – Gartner, Germany S. typhimurium – meat – 1898 – Durham, England S. enteritidis– meat from cow Source – poultry, meat, milk, cream, eggs, rat droppings Clinically – incubation period – 24 hrs/less, diarrhea, vomiting/pain, half loose stools,subsides – 2–4 days LABORATORY DIAGNOSIS Feces culture, article food Rx – symptomatic – never antibiotics – increase period of fecal shedding of bacilli

  46. SALMONELLA SEPTICEMIA S. cholera suis – osteomyelitis, deep abscesses, endocarditis Diagnosis – blood, pus, fecal cultures Rx – chloramphenicol

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