Specimen collection and laboratory procedure

A single, fresh, whole stool specimen (at least 3 ml or grams, ideally 10 ml or

grams) was collected from clinical diagnosed patient at general hospital makurdi.
A faecal swab was then placed in Cary-Blair medium in a plastic screw top test
tube. Stool samples were then transported to the central laboratory for screening
common enteric pathogens such as ETEC, Shigella spp. maintaining standard
protocols. Optimal cool temperature was strictly maintained from the point of
collection of stool sample to successful submission.
For ETEC, stool samples were plated onto MacConkey agar, and the plates
were incubated at 37C for 18 hours. Six lactose-fermenting individual colonies,
morphologically resembling E. coli, were tested.
ISOLATION AND PRESUMPTIVE IDENTIFICATION PROCEDURE
Stool Culture, Diarrheagenic E.coli
Test Stool Culture, Diarrheagenic E.coli
Indication Establish diarrheagenic E.coli as the cause of clinical illness.
Physiology
Diarrheagenic E.coli are Enterotoxigenic (ETEC)-Travelers diarrhea and infant
diarrhea in less developed countries.
Normal
Range
Normal colinc flora
Sample Rectal swab, fresh stool
Test Method Culture in macconkey agar.

Shigella species were isolated and identified in the enteric microbiology
laboratory by using standard biochemical and microbiological methods. Stool
specimens were inoculated in MacConkey and Shigella agar plates and incubated
overnight at 37C. Infectious dose above 10 to 200 cells can cause disease
Laboratory diagnosis:
Stool examination
a. Isolation of Shigella from feces or rectal swab specimen is diagnostic
but lacks specificity. Routine microscopy may reveal sheets of
leukocytes on methylene-blue stained stool smear, which is a
sensitive test for colitis but not specific for Shigella species.
b. In approximately 70% of patients with shigellosis, fecal blood or
leukocytes (confirming colitis) are detectable in the stool.


Stool culture
A sample for stool culture should be obtained in all suspected cases of
shigellosis.
The yield from stool cultures is greatest early in the course of disease.
Guidelines for obtaining specimens to improve the yield are as follows:
I. Process specimens immediately after collection.
II. If processing is delayed, use a transport medium (eg, buffered
glycerol saline).
III. Collect more than one stool or rectal (not anal) swab and
inoculate them promptly on at least 2 different culture media.
IV. Specimens should be plated lightly onto MacConkey, xylose-
lysine-deoxycholate, Hektoen enteric, or Salmonella-Shigella,
or eosin-methylene blue agars
If processing is delayed, a rectal-swab sample can be placed in Cary-Blair
transport medium or buffered glycerol saline.
After overnight incubation, colorless, nonlactose-fermenting colonies may
be tested by means of latex agglutination to establish a preliminary
identification of Shigella infection.
Antimicrobial susceptibility tests of all confirmed isolates should be
performed by using the agar diffusion technique. The agar and broth-
dilution methods are also widely used. The new Epsilometer strip method
(E test) is used to accurately determine the minimum inhibitory
concentration (MIC).
Despite meticulous care in obtaining and processing specimens from
patients infected with Shigella species, approximately 20% may fail to yield
Shigella organisms.

Enzyme immunoassay: An enzyme immunoassay for Stx is used to detect S
dysenteriae type 1 in the stool.
Rapid techniques: With rapid techniques, gene probes or polymerase chain
reaction (PCR) primers are directed toward virulence genes (invasion
plasmid locus).