Microbiology Lab

JOCELYN YEO 0703359J



I was attached to microbiology lab for three days.



Upon receiving specimen, all request form is needed to clot in time of arrival in the lab. This is for tracibility and also to track the turn around time. Specimens received are sort according to high vaginal swab (HVS), ear swab, methicillin-resistant staphylococcus aureus (MRSA), urine specimen, blood specimen, Tip specimen and stool specimen. Before processing the specimens, we are suppose to check patient information labelled on the specimen against the request form. Different specimen will require different condition, procedure and agar plate for incubation. After incubation, medical technologists will read and type the result in a computer system.



Urine processing



Macconkey without salt and TSA blood agar plate is used to culture.

Shake the bottle of urine. Dip the 1µl disposable innoculation loop into the bottle. The loop should not be dipped all the way to the bottom. Then streak on the TSA blood agar first. Dip the same innoculation loop into the urine specimen again and streak on the Macconkey agar. Next, use a capillary tube to draw the urine and insert it into the disposable hemocytometer for cell counting.















Method to streak for urine specimen.

Stool specimen

Different form of stool specimen is needed to culture on different agar plate.
1. Formed or loosed stool : XLD, campylobacter plate and selenite F broth
2. Watery stool : XLD, campylobacter plate, selenite F broth, TCBS, TSA blood agar and alkaline peptone water
3. Bloody stool : (select mucus or blood flecked material to culture) XLD, campylobacter plate, selenite F broth, CIN, Macconkey sorbitol agar.
4. Stool from BMT or oncology patient : XLD, campylobacter plate selenite F broth, blood with gentamicin, blood agar, Macconkey plate and VRE plate.
5. Request for aeromonas or pesiomonas culture : XLD, campylobacter plate, selenite F broth, blood plate.
6. Request for VRE and ESBL screening : VRE plate and gentamicin blood agar plate.
(Macconkey sorbitol is to look for E.coli 0157 and campylobacter plate is to look for campylobacter.)
7. Rotavirus tested by using a commercial kit. Before test procedure, ensure test kit is at 20-25 degree celsius for optimum result. Add 1µl of diluent to 100µl of stool specimen and mix it. Allow sedimentation for 3 minute. Pipette 4 drops of mixture into the cassette. Read the result after 5 minutes. Inorder for the result to be valid, the control must be positive.



Ear culture

Culture on CDC and Macconkey without salt agar plate.



Wound specimen

Blood agar and Macconkey agar plate is used for superficial wound specimen. An additional CDC plate is needed for deep infection to check for anaerobic bacteria.



Tip specimen

Only blood agar plate is used as the aim is to check for any bacteria growth.



Upper respiratory system

Streak on blood plate and chocolate agar.
(chocolate agar is an enrichment media, hence Hemophilus influenza is able to grow.)



Endotrachel Aspirate (ETTA)

Culture on blood agar and Macconkey agar.



Subchronic specimen

This include specimen such as placenta, womb, etc.
Blood agar, Macconkey agar plate and CDC plate is used.



Sterile specimen

For all sterile specimen, gram staining must by carried out and analyze before plate reading.

1. CSF : Culture on blood agar, chocolate agar and CDC plate. (specimen is kept 3 days)
2. Pleural fluid : Culture on blood agar, chocolate agar, CDC plate and Macconkey agar.
3. Tissue (eg. lymph node) : Culture on blood agar, Macconkey agar and CDC plate.
4. Endocervical : Culture on Thayermartin media (MTM)















Method to streak for miscellanous specimen (all specimen except urine)



Results


Urine
1 colony is approximately 1000cfu/ml. If there is less than 10 colonies, report as no significant growth.
However, if bacteria is suspected to be yeast (candida) , perform wet mount and look under microscope. Germ test is then carry out if yeast is observed under microscope. Germ test is a test to confirm for Candida albicans.
Typical morphology of E.coli is flat, and lactose fermenter (pink on Macconkey plate). If bacteria is suspected to be E.coli, perform spot indole test by putting a drop of indole reagent on a strip of filter paper and scrap a colony. If colour changes to green, it is indole positve. Indole negative remain pink, hence send for identification of bacteria and susceptibility test.
Spot indole test should use colonies from the TSA plate as colonies on the Macconkey plate might be stained with colours thus affecting the result.



Stool
black colony may suggest Salmonella species. Other bacteria to look out for are E.coli and Acinetobacter baumanii.



Wound specimen
common bacteria are S. aureus, Group A Streptococcus, Pseudomonas aeruginosa and GMV.



Upper respiratory system specimen
look out for Streptococcus pneumonia, Hemophilus influenza and S. aureus.



Endotrachel aspirate (ETTA)
if Macconkey shows orange, it means the bacteria is a non-lactose fermenter, thus perform indole and oxidase test.
Colonies that look flat, spreading and metallic may suggest pseudomonas species. Thus perform oxidase test .
Catalyst test is to differentiate between Staphylococcus and Steptococcus species.



Ear specimen
look for Group B Streptococcus. There might be GMV and enterococcus.



Subchronic
look for gram negative bacteria.



MRSA
methicillin resistant organism will appear maurve colour (purplish pink) on MRSA select media.
If suspect methicillin resistant organism present, perform agglutination test with Bactistaph reagent. If Bactistaph reagent forms agglutinates with the colony, S. aureus is present.