MT KENYA UNIVERSITY

SCHOOL CLINICAL MEDICINE

DEPARTMENT CLINICAL MEDICINE AND SURGERY

UNIT NAME LABARATORY TECHNIQUE 1

UNIT CODE DML 2104

NAME SUZAN MURIMI CHACHA

REG NO DCM/2017/72179

TASK: CAT
1. Write brief notes on what needs consideration when determining if a blood
culture isolate represents a pathogen or a skin contaminant (10 marks)
2. Describe the procedures and tests that are used in the laboratory culture and
identification of suspected non technical salmonella enterica infections (10
marks)
3. Discuss the diagnosis of bacterial vaginosis and the expected results of any
laboratory test which may be performed (10 marks)
1. Write brief notes on what needs consideration when determining if a blood
culture isolate represents a pathogen or a skin contaminant (10 marks)

 Presence of disposing factors and a consistent clinical presentation can help clinicians
interpret test results in deciding whether a blood isolate is a pathogen or contaminant.
 The identity of microorganisms provides important information.
 Micro-organisms that represent true infection when isolated from blood cultures
include Staphylococcus aureus,Streptococcus pyogens,Streptococcus
agalactiae,Streptococcus pneumonia,escherichia coli and other members of the family
enterobacteriae,pseudomonas aureginosa and candida species
 In contrast, coagulase-negative staphylococci, cornebacterium species, bacillus species
represent contamination.
 Corynebacterium are normal skin flora but can cause clinically significant infections in
presence of medical devices such as catheters, heart valves, joint prostheses.
 In true endovascular infections, either all or most of blood cultures obtained at the time
of diagnosis will be positive, whereas when a blood culture is contaminated, usually only
one of several blood sets will be positive.
 It is important for the health worker to use strict aseptic technique when obtaining
blood specimen to avoid contaminants.
 Use of efficacious antiseptic preparations for skin sterilization e.g. iodine tincture should
be considered.
 Means by which blood is obtained for culture should be considered; obtaining blood
cultures from existing in dwelling I.V catheter or other access devices are more
contaminated, they should therefore be obtained by peripheral venepuncture.

2. Describe the procedures and tests that are used in the laboratory culture and
identification of suspected non technical salmonella enterica infections (10
marks)

Test: Stool culture

Procedure

 Weigh 25 gram of faeces with a sterile wood spatula.

 If there will be unavoidable delay in culturing of the specimen, then place it in a 5ml
buffered glycerol transport media.
 Place the faeces directly on McConkey agar and deoxylate citrate agar and Wilson and Blairs
bismuth sulphite agar medium.
 For enrichment, one tube each of Selenite F and tethrionate broth are also inoculated.
 Incubate at 37 degrees Celsius for 8-12 hrs. or overnight with subsequent subculture on
MacConkey medium.
 Tetrathionate broth and selenite F broth are responsible for the inhibition of replication of
normal intestinal bacteria and permits multiplication of salmonella species.
 The specimen is plated on salmonella shigella agar or hektoen enteric agar.

Results and Identification

 On deoxycholate citrate agar, colonies are pale(straw colour) and non lactose fermenting.
 On xylose lysine deoxycholate agar-colonies are red and black centre.
 Bismuth sulphite medium permits rapids rapid detection of salmonella which forms black
colonies because of hydrogen sulphide gas production.
 Suspected colonies are identified by biochemical reaction pattern and slide agglutination
test with specific sera.

Blood specimen
Procedure

 Blood should be collected when the patient’s body temperature is rising.

 It should also be collected before any microbial treatment.
 Two samples should be collected at different times and should be collected aseptically as
possible.
 Swab once and insert the needle on the cubital fossa of the forearm and draw ml/s of blood
 Dispose 5 millilitres in each of the culture bottles.
 Gently mix the blood with the broth, the broth does not allow the blood to clot and also
serves to dilute the bactericidal nature present in the blood.
 Inoculate the sample on thioglycolate broth medium and incubate
 Incubate the inoculated media at 35-37 degrees for up to duration of two weeks and
observe for overnight growth.
 If there is turbidity and colonies present, subculture in the blood and mcConkey Agar and
incubate at 37 degrees for 24 hours.
 If there is no growth then report the subculture every day for up to a period of 7 days.
Identification and results

 If there is no growth after the 7th day then the culture is negative.
 If there is growth subculture in blood and macConkey agar.
 In blood agar colonies are large about 2-3cm if salmonella is present.
 In MacConkey agar the colonies are 1-3mm in diameter and pale.
 Salmonella enteridis does not ferment inositol

Agglutination test
Procedure

 For slide agglutination test a loopful of growth from a nutrient agar plate or slope is
emulsified in two separate drops of saline on a clean slide.
 One emulsion acts as a control to show that the strain is not auto agglutinable.
 If the isolate is anaerogenic, a loopful of factor 9 anti-sera is added to one drop of a bacteria
emulsion on the slide.
 Immediate agglutination indicates that the isolate belongs to salmonella serogroup D.

Identification

 Its identity as Salmonella typhi is established by agglutination with the flagellar antiserum
(anti-d) serum.
 If the isolate is non typhoidal salmonella,it is tested for agglutination with D and H antisera
for serogroups A,B,C.

3. Discuss the diagnosis of bacterial vaginosis and the expected results of any
laboratory test which may be performed (10 marks)
Diagnosis of bacterial vaginosis is made by the following signs or symptoms:

 A vaginal pH of greater than 4.5.

 A positive whiff test (fishy amine odour) when the discharge is mixed with 10% Potassium
hydroxide.
 A homogenous, thin white discharge that smoothly coats the vaginal walls.
 At least 20% clue cells on microscope exam(Vaginal epithelial cells with borders obscured by
adherent cocobacilli on went mount preparation on gram stain).
 Burning sensation during urination.
Lab test and results

 Wet preparation(wet mount/smear):A drop of vaginal discharge is placed on a glass slide

and examined under a microscope. Results indicate the presence of ‘clue cells’ which are
cells from lining of the vagina.
 Gram Stain:Vaginal fluid is placed on a glass slide and stained with a gram stain. Positive
findings show 20% or more of cells in the vaginal lining covered by bacteria
 pH test: the vaginal discharge is checked for ph. Results include a pH Of greater than 4.5
 Potassium hydroxide prep: A sample of vaginal discharge is placed on a glass slide with a
drop of 10% Potassium Hydroxide. Positive findings include chemical compounds called
amines which are released causing a fishy odour.
 Vaginal discharge culture: involves successful culture and identification of Gardnerella
vaginalis.
 Molecular methods e.g. nucleic acid amplification for identification of bacteria vaginosis