main principles of specimen collection for microbiology tests with examples of collection rules and techniques for various types of samples

1. Collection and transport of
biological specimens

2. Biological specimens
Depending on source:
• sterile collection sites:
– Blood, CSF, urine
• contaminated collection sites:
– Faeces, naso-pharyngeal secretion, pus, vaginal
secretion
+ specimens from sterile collection sites which may be
contaminated during elimination e.g. sputum, urine

3. Parameters to be considered when
collecting biological specimens
• WHAT: choice of biological specimen depending on
clinical stage
• WHEN: optimal moment of collection; maximum time to
reach the laboratory
• HOW: collection procedure (avoid contamination of
specimen); instruments (sterile !!); manipulation and
packaging (no contamination of staff and environment)
• HOW MUCH: adequate quantity for required tests

4. Minimal Information in Request form
• About patient: ID, age, gender
• About disease: date of onset, suspected
diagnosis/causative pathogen
• About specimen: type of specimen, collection
site, time of collection, time of shipment, storage
conditions (if applicable)
• About referring physician – contact data (name,
phone no)

5. Microbiological tests: Most often referred
specimens
• Biological samples: secretions, excretions,
tissues, organ fragments
• Food samples
• Pharmaceutic products, cosmetics
• Environmental samples: water, air, soil

6. General requirements for collecting
pharyngeal, naso-pharingeal exudate
• For the patient:
– in the morning, before feeding, before brushing teeth;
alternatively: at least 4 hours since last meal & teeth
brushing
– No mouth rinse, no chewing gum!
– No antibiotics during the last 7-10 days
• For medical staff:
– Wear gloves, face protection (mask, eye
protection/face shield), protective lab coat

7. Collection of pharyngeal exudate
• Dacron or Rayon swab
• Tongue blade & good light
• Insert swab behind uvula
without touching it
• Swab tonsils, posterior
pharynx + lesions (if any)
• Avoid touching tongue,
cheeks, teeth
• Place swab in sterile tube
• Transport to lab (RT/2-8°C)

8. Collection of pharyngeal exudate

9. Collection of nasal/nasopharyngeal exudate
• Head of patient imobilised in neck extension
• Sterile swab gently inserted through nostril along nasal
floor until reaching posterior pharingeal wall
• Rotate swab, withdraw slowly + reinsert and repeat steps
• Take out and insert into sterile tube
• Send to lab

10. Collection of sputum
• Challenging! – must avoid contamination of sputum with
saliva and secretions from upper air ways
Optimal moment: in the morning (higher amount of sputum
secreted during the night and stagnant in lower
respiratory ways)
Indirect method:
• Patient energically rinses mouth with saline solution
• Coughs and expectorates in sterile container (Petri dish)
Direct method:
• Bronchoscopy / tracheal punctioning

11. Collection of Blood
• Normally sterile biological product
• Collection technique depends on the required tests:
– A. bacteriology = hemoculture
– B. Serology

12. A. Blood collection for hemoculture
Blood injected in 2
sets of sealed bottles
containing liquid culture
medium for aerobic and
anaerobic bacterial
growth

13. Collection of blood for hemoculture
• Wear gloves + PPE
• Thoroughly wipe skin with antiseptic (chlorhexidine,
iodine, alcohol)
• During 3 hours, draw blood by venipuncture from up to 3
different sites at 1 hour interval (3 sets of 2 bottles each)
– around 5 ml blood per bottle
• After drawing the blood, dispose of the syring needle and
attach new, sterile needle
• Disinfect cap of each culture medium bottle and inject 5
ml blood/bottle

14. Collection of blood for hemoculture

15. Automated systems for detection of bacteria in blood and
other normally sterile body fluids

16. B. Blood collection for serology
• similar PPE used by person collecting sample (gloves,
lab coat, eye protection, etc)
• Thorough skin decontamination with antiseptic
• Blood drawn by venipuncture with:
• sterile syringe and needle and injected/inserted in sterile
container
• Vacuum collecting system (tube + needle): blood drawn
by vacuum into tube containing clot activating substance
(facilitates clot formation & separation of serum)

17. B. Blood collection for serology

18. Collection of Pus
• Composition: white blood cells, microbes, cell debris,
fibrin
Closed purulent lesions (abscesses):
• surgical collection:
– rigurous cleaning and disinfection of skin (iodine)
– Incision and aspiration of pus
Open lesions:
• Cleaning and disinfection of skin around lesion (iodine)
• Collection of pus with sterile swab / loop

19. Collection of stool (faeces)
• Disposable stool collection containers (simple / with
transportation medium Carry Blair: non-nutritive medium
which prevents overgrowth of Enterobacteriaceae but
preserves viable enteric pathogens (Salmonella,
Shigella, Vibrio, etc)

20. Collection of urine
When?:
- in the morning (first miction)
How?:
- clean uro-genital area
- eliminate first flow
- collect middle flow in
sterile container
Send to lab immediately or store
at 2-8°C

21. Collection of cerebrospinal fluid (CSF)
Lumbar punction (spinal tap)
• patient lies on the side, knees pulled up toward
chest, chin tucked downward
• back cleaned and disinfected (iodine) + health
care provider injects local anesthetic into lower
spine
• spinal needle inserted into lower back area
• needle properly positioned, CSF pressure
measured and sample collected in sterile tube
• needle removed, area cleaned, bandage placed
over puncture site