Transcript LAB DIAGNOSIS

Bacteriological diagnosis of enteric
fever consists of:
1. Isolation of bacilli
2. Demonstration of antibodies
3. Demonstration of circulating
antigen
4. Other laboratory test
ISOLATION OF BACILLI
Specimens:Blood
Urine
Faeces
Aspirated duodenal
fluid,etc..
BLOOD CULTURE
Positive in,
90%-1st week
75%-2nd week
60% -3rd week
25%-till the subsidence of pyrexia
contd…
• Blood collected is cultured on glucose
&taurocholate broth..
• We must take care of contamination
with external environment
• Dilute or add sodium polyanethol
sulphonate
• Incubated at 37degrees
• Castaneda’s method is also practiced
CLOT CULTURE
• Aseptically collected blood is allowed
to clot
• Serum-used in widal test
• Blood clot-added to bile broth
containing streptokinase(causes lysis
of blood clot)
FAECES CULTURE
• Salmonella are shed in the faeces
through out the disease even in
convalescence
• Antibiotic does not eliminate the
bacilli from the GUT as it does 4m
BLOOD
CULTURE
• On McConkey
agar & DCA media-Pale yellow colonies
• On wilson blair media
• S.typhi-black colinies(with metallic
sheen)
• S.paratyphi-green colonies(due to
absense of hydrogen sulphide
• Enrichment media are incubated and
then subcultured on selective media
On mac conkey agar
• S.typhi
• S.paratyhi
URINE CULTURE
• Not much more imp as blood & faeces
culture
• Positive only in 2nd& 3 rd weekes
• Urine sample is centrifuged
&inoculated into enrichment
&selective media
1.
2.
3.
4.
5.
6.
Other specimens
Bone marrow
Bile
Rose spots discharge
Pus
Suppurative lesions
Csf&sputum
At autopsy-gallbladder,liver,spleen
& mesenteric
lymphnodes
SLIDE AGGLUTINATION TEST
• A loopful of growth from a nutrient
agar is emulsified in two drops of
saline on a microscopic slide
CONTD..
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1st emulsion-control
To show the strain not
agglutable
2nd emulsion-test
Polyvalent “o”&polyvalent
“H”are added
Positive-presence of
salmonella sps
Then monovalent anti sera
belongs to S.typhi are added
If the isolate is fresh
agglutination is done with
“vi”antisera,alternatively the
bacterial suspension is boiled
for 20 mins which removes
the vi antigens
¤ Non typhoid Salmonella sproducing acid and
gas 4m carboydrates is tested for “O”
&”H”antisera for groups A,B,C…..
¤ NATIONAL SALMONELLA
REFERENCE CENTRE at the CRI
¤ INDIAN VETERINARY RESEARCH
INSTITUTE,Izatnagar
SLIDE
AGGLUTINATION
DEMONSTRATION OF
ANTIBODIES
WIDALTEST
PROCEDURE:
DREYER”S TUBE - H agglitination
FELIXTUBE
- O agglutination
28 test tubes are arranged in 4 rows each
row containing 7 tubes and named as
1st row-To
2nd row-Th
3rd row-Ah
4th row-Ao
Contd….
• To &Th for the detection of S.typhi
• Ao &Ah for the detection of
S.paratyphi
• Of the 7 tubes one acts as control
&remaining tubes as test
Contd….
• A low dilution of patient serum is
1:20 to 1:640
• All tubes in rack are incubated at 37
degrees for 24hrs&results are read
after over night refrigeration at 4
degrees
Contd….
• Positive H agglutination-formation of
loose,cotton wooly clumps
• Positive O agglutination-disc like pattern
at the bottom of tube
• In both supernatent fluid is clear
• Paratyphoid O antigens are not employed
because theycross react with typhoid O
antigen due to their haring of factor 12…
RESULT
INTERPRETATION
1. Titre depends on stage of disease
2. On immunistion with TAB vaccine
individuals may show high titres of
antibody
3. In non immunised individuals high
titre of H agglutinin indicates
entericfever or latent infection
4. ANAMNESTIC REATION:
5.
In carriers
DEMONSTRATION OF
ANTIGENS
• Ag present in blood and urine
• PREPARATION OF ANTIGENS:
S.aureus is stabilised with fprmaldehyde
&then coated with S.typhi antibody
• PROCEDURE:
Patients serum is mixed with staphylococcal
cells
Positive-1st week..
• CIEP,ELISA
OTHER….
1. TLC
2. DIAZO TEST IN URINE
• TLC-leucopenia with a relative
lymphocytosis is found
• DIAZO TEST:
• Positive -in b/w 5th &14th weeks
Duration of
diseaseS
Specimen
examination
%positivity
1st week
Blood culture
90%
2nd week
Blood culture, 75%
Faeces culture 50%
Widal test
Low titre
3rd week
Widal test
80-100%
Blood culture 60%
Faeces culture 80%
CARRIERS
Convalescent
carriers
Healthy carriers
Chronic
carriers
DIAGNOSIS…..
• SPECIMENS:
stool,urine,bile,duodenal drainage
Antibody ti vi antigen in serum is clamied indicate
thecarrier state
Convalescent &healthy carriers shed bacilli in
faeces from 2 months to 1 yr…
Where as chronic carriers shed bacilli in faeces
from 1yr to several years
• BACTERIOPHAGE TYPING:
• BIO TYPING
• PLASMID TYPING
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ANTOIBIOTICS:
chloramphenicol
ampicillin
furazolidone
cotrimoxazole
For multiresistent cases:
flouroquinolones
3rd generation cephalosporins
Resistence has been devoloped to antibiotics
In salmonella gastro enterits antibiotics should not
be used
1. Maintaining highgenic conditions
2. Carriers should not be engaged in
food preparation
3. Vaccination
TAB vaccine
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Heat killed,whole celled,
S.typhi-1,000 millions/ml
S.paratyphi A-750 millions/ml
S.paratyphi B-750 millions/ml
Dose schedule:
Protection:
Side effects:
TYPHOID VACCINE
• Live oral,
• Avirulent mutent strain
• S.typhi lacking UDP-galactose-4epimerase has been used`as live oral
vaccine
• Dose schedule:
• Protection:
TYPHIM-Vi
• Purified Vi antigen
• Dose schedule :im
•
singledose
•
25 micro grams