1. MICROBIOLOGY (NRS 212)
Sample collection and shipping
Collection, Labelling and Transportation of
Microbiological samples

2. Collection and transport of microbiological
samples

3. • Collection of adequate and appropriate specimens
• Sufficient documentation
• Biosafety and decontamination
• Correct packaging
• Rapid transport
• Choice of a laboratory that can accurately perform the tests
• Timely communication of results
• Advance planning
Successful laboratory investigations:-

4. Consider differential diagnoses:-
• Decide on test(s) to be conducted
• Decide on clinical samples to be collected to conduct these tests
• Consultation between microbiologist, clinicians and epidemiologist
Specimen collection: key issues

5. • Allows organisms (pathogens and contaminants) to survive.
• Non-nutritive - does not allow organisms to proliferate.
• For bacteria – i.e., Cary Blair
• For viruses - virus transport media (VTM)
Transport medium:-

6. Collection:-
Capillary blood from finger prick
• make smear
• fix with methanol or other fixative
Handling and transport:-
Transport slides within 24 hours
Do not refrigerate (can alter cell morphology)
Blood for smears:-

7. Collection
• Venous blood
• infants: 0.5 – 2 ml
• children: 2 – 5 ml
• adults: 5 – 10 ml
• Requires aseptic technique.
• Collect within 10 minutes of fever.
• if suspect bacterial endocarditis: 3 sets of blood culture
Blood for cultures:-

8. Handling and Transport:-
Blood for cultures Collect into bottles with
infusion broth
• change needle to inoculate the broth
Transport upright .
• prevents hemolysis
Blood for cultures:-

9. Collection
Venous blood in sterile test tube
• Let clot for 30 minutes at surrounding temperature
• glass better than plastic
Handling
Place at 4-8oC for clot retraction for at least 1-2 hours
Centrifuge at 1500 RPM for 5-10 min
• separates serum from the clot
Transport
4-8oC if transport lasts less than 10 days
Freeze at -20oC if storage for weeks or months before processing
and shipment to reference laboratory.
Avoid repeated freeze-thaw cycles
• destroys IgM
To avoid hemolysis: do not freeze unseparated blood
Serum:-

10. Collection procedure:-
• Lumbar puncture
• Sterile tubes
• Aseptic conditions
• Trained person
Cerebrospinal fluid (CSF):-

11. CSF
Handling and transportation
Bacteria
• preferably in Transporter -isolate medium,
pre-warmed to 25-37°C before inoculation
OR
• transport at surrounding temperature (relevant pathogens
do not survive at low temperatures)
Viruses
• transport at 4-8oC (if up to 48hrs or -70oC for longer
duration)

12. Collection:
Freshly passed stool samples
• avoid specimens from a bed pan
Use sterile or clean container
• do not clean with disinfectant
During an outbreak - collect from 10-20 patients
Stool samples:-

13. Advantage:-
• suitable
• Adapted to small children, impaired patients and other situations
where voided stool sample not feasible (possible)
Disadvantage:-
• No macroscopic assessment possible
• not recommended for viruses
Rectal swabs:-

14. Timing
 within 48 hours of onset
Sample amount
 5-10 ml fresh stool from patients (and controls)
Methods
 fresh stool unmixed with urine in clean, dry and sterile container
Storage
 refrigerate at 4oC; do not freeze
 store at -15oC - for Ag detection, polymerase chain reaction (PCR)
Transport
 4oC (do not freeze); dry ice for (Ag detection and PCR)
Stool samples for viruses:-

15. Timing
 during active phase
Sample amount and size
 fresh sample and two swabs from patients, controls and carriers (if
indicated)
Method
 Cary-Blair medium
 For Ag detection/PCR – no transport medium
Storage
 refrigerate at 4oC if testing within 48 hours, -70oC if longer; store at -
15oC for Ag detection and PCR
Transport
 4oC (do not freeze); dry ice for (Ag detection and PCR)
Stool samples for bacteria:-

16. Timing
 as soon as possible after onset
Sample amount and size
 at least 3 x 5-10 ml fresh stool from patients and controls
Method
 Mix with 10% formalin or polyvinyl chloride, preservative
 Unpreserved samples for Ag detection and PCR
Storage
 refrigerate at 4oC; store at -15oC for Ag detection and
PCR
Transport
 4oC (do not freeze); dry ice for (Ag detection and PCR)
Stool samples for parasites:-

17. • Hold tongue away with tongue
depressor
• Locate areas of inflammation and
exudate in (posterior pharynx,
tonsillar region of throat behind
uvula)
• Avoid swabbing soft palate (uvula);
do not touch tongue
• Rub area back and forth with cotton
or Dacron swab.
WHO/CDS/EPR/ARO/2006.1
Throat swab (posterior pharyngeal swab):-
Dacron swab

18. • Tilt (slope) head backwards
• Insert flexible fine-shafted polyester
swab into nostril and back to
nasopharynx
• Leave in place a few seconds
• Withdraw slowly; rotating motion
Nasopharyngeal swab:-

19. • Tilt (slope) head slightly backward.
• Instill (inject) 1-1.5 ml of VTM (Viral
Transport Medium) /sterile normal saline
into one nostril.
• Use aspiration device.
• Insert silicon catheter in nostril and
aspirate the secretion gently by suction
in each nostril
Naso-pharyngeal aspirate:-

20. Collection
• Instruct patient to take a deep breath and cough up sputum directly
into a wide-mouth sterile container.
• avoid saliva or postnasal discharge.
• 1 ml minimum volume
Sputum:-

21. Handling and Transport:-
- All respiratory specimens except sputum are transported
in appropriate media.
• bacteria: Amie’s or Stuart’s transport medium
• viruses: viral transport medium (VTM)
- Transport as quickly as possible to the laboratory to
reduce overgrowth by oral flora
- For transit periods up to 24 hours
• surrounding temperature for bacteria
• 4-8°C for viruses
Respiratory samples:-

22. Collection:-
Biopsy relevant tissues
• place in formalin for histopathology
• place in transport medium for microbiological testing
• place in sterile saline for isolation of viral pathogens
Post-mortem samples:-
Handling and transportation:-
• Fixed specimens can be transported at surrounding
temperatures.
• Transport specimens in transport media within 24h at
surrounding temperature.
• Transport specimens in sterile saline at 4-8oC within 48h.

23. • Patient’s name
• Clinical specimen
• Unique ID number (Research/Outbreak)
• Specimen type
• Date, time and place of collection
• Name/ initials of collector
Labeling specimens:-

24. Label slides individually:
• use glass marking pencil
• ensure markings don’t interfere with staining process
Each slide should bear:
• Patient name
• Unique identification number
• Date of collection
Glass slides for microscopy:-

25. Patient information:-
 age (or date of birth), sex, complete address
Clinical information:-
 date of onset of symptoms, clinical and immunization history, risk
factors or contact history where relevant, anti-microbial drugs taken
prior to specimen collection
Laboratory information:-
 acute or recovering specimen
 other specimens from the same patient
Receiving laboratory records:
 Date and time when specimen was received
 Name and initials of the person receiving specimen
 Record of specimen quality
(Line listing – if large number of patients)
Case investigation form:-

26. • Use single use equipment
• Disinfect
• Work in a clean, specific area
Biosafety: protect the patient:-

27. Use personal protective equipment
• disposable gloves
• laboratory coats / gown
• mask
• protective eyewear .
• Face shields if procedure is likely to generate aerosols.
Note:-
• If no sharps container: collect sharps immediately to prevent needle-
stick injury
• First aid kit readily accessible
• Do not reuse contaminated equipment
Biosafety: protect yourself:-

28. • Package samples appropriately for transport.
• Decontaminate spills - 10% bleach wash in soapy water before re-
use, sterilize if necessary
• Place waste in leak-proof biohazard bags - ensure safe final
management of waste
• Protect cleaning/decontamination personnel with protective
coat, thick rubber gloves
Biosafety: protect others, the environment:-

29. • Mismatch of information on the label and the request.
• Inappropriate transport temperature
• Excessive delay in transportation
• Inappropriate transport medium
1. specimen received in a fixative
2. dry specimen
3. sample with questionable relevance
• Insufficient quantity.
Criteria for rejecting samples:-

30. THANKS