Microbiology

1. SPECIMEN COLLECTION &
TRANSPORT,
CRITERIA FOR ACCEPTANCE AND
REJECTION OF SPECIMEN
Presented by :
Dr. APURBADAS
JR1 , Dept. of Microbiology
SVNGMC, Yavatmal
Date : 24th January, 2023

2. SPECIMEN COLLECTION AND
TRANSPORT

3.  Cultivation (growth), identification, and antimicrobial susceptibility testing of
microorganisms
Direct detection of infecting organisms by microscopy
 Direct detection of specific products of infecting organisms using chemical,
immunologic, or molecular techniques
 Detection of antibodies produced by the patient in response to an infecting
organism (serology)
Objectives

4. GENERAL PRINCIPLES
Standard precautions should be followed.
Culture specimen should be collected during the acute phase of
infect and prior to administration of antimicrobial agents.
Contamination with indigenous flora should be avoided (eg. Urine &
blood )
Specimens should be collected in *Sterile * tightly sealed * leak
proof
* wide mouth * Screw capped containers.

5. GENERAL PRINCIPLES
All specimens must be appropriately labelled.
Specimens grossly contaminated or compromised or improperly labelled
may be rejected.
If anaerobic culture is requested, proper anaerobic collection containers
with media should be used.

6. PROCEDURES FOR
SAMPLE COLLECTION &
TRANSPORT

7. ABSCESS
(also Lesion, Wound, Pustule, Ulcer)

8. SPECIMEN
Superficial Deep
CONTAINER
Aerobic swab moistened with
Stuart’s or Amies medium.
Anaerobic trasnporter
PATIENT
PREPARATION Wipe area with sterile saline or 70% alcohol
SPECIAL
INSTRUCTIONS
Swab along
leading edge of wound
Aspirate material from wall or
excise tissue
TRANSPORT TO
LABORATORY Within 24 hr in Room temperature
STORAGE BEFORE
PROCESSING 24 hr in Room temperature

9. HOLDING MEDIA
holding media is a nonnutritional medium
which maintains the viability of organisms
without significant multiplication.
The small agar content provides a semisolid
consistency which prevents oxidation and
drying during transport.
Stuart’s medium and Amie’s medium are two
common holding media.

10. BLOOD

11. SPECIMEN BLOOD
CONTAINER
Blood culture media set (aerobic and anaerobic bottle) or
Vacutainer tube with SPS
PATIENT
PREPARATION
Disinfect venipuncture site with 70% alcohol
SPECIAL
INSTRUCTIONS
•Draw blood at time of febrile episode
•draw two sets from right and left arms
•do not draw more than 3 sets in a 24-hr period
•draw >_20 mL/set (adults) or 1-20 mL/set (pediatric)
depending on patient’s weight
TRANSPORT TO
LABORATORY
Within 2 hr in Room temperature
STORAGE BEFORE
PROCESSING
<2 hr in Room temperature
Must be incubated at 37°C on receipt in laboratory.

12. Venepuncture technique for blood collection
Blood culture transport media
Vacutainer tubes

13. BODY FLUIDS

14. SPECIMEN
Body Fluids Amniotic,
abdominal, ascites
(peritoneal), bile, joint (synovial), pericardial, pleural
CONTAINER
Sterile, screwcap tube or anaerobic transporter or
direct inoculation into blood culture bottles
PATIENT
PREPARATION
Disinfect skin with iodine preparation before aspirating
specimen
SPECIAL
INSTRUCTIONS Needle aspiration
TRANSPORT TO
LABORATORY
<15 min
STORAGE BEFORE
PROCESSING
<24 hrs in room temperature, Plate as soon as received.

15. ASCITIC FLUID ASPIRATION

16. CEREBROSPINAL FLUID (CSF)

17. SPECIMEN CEREBROSPINAL FLUID (CSF)
CONTAINER
Sterile, screwcap tube
PATIENT
PREPARATION
Disinfect skin with iodine or
chlorhexidine before aspirating specimen
TRANSPORT TO
LABORATORY
<15 min
STORAGE BEFORE
PROCESSING
<24 hrs, Routine incubate at 37°C,
except for viruses,
which can be held at 4°C for up to 3 days.

18. LUMBAR PUNCTURE FOR CSF
COLLECTION

19. RESPIRATORY TRACT
UPPER

20. SPECIMEN
NASOPHARYNX
CONTAINER
Swab moistened with Stuart’s or Amie’s medium
SPECIAL
INSTRUCTIONS Insert flexible swab through nose into posterior
nasopharynx and
rotate for 5 seconds;
TRANSPORT TO
LABORATORY <15 min in room temperature without transport media;
<2 hrs in room temperature using transport media
STORAGE BEFORE
PROCESSING 24 hrs in room temperature

21. SPECIMEN
PHARYNX (THROAT)
CONTAINER
Swab moistened with Stuart’s or Amie’s medium
SPECIAL
INSTRUCTIONS Swab posterior pharynx and tonsils;
TRANSPORT TO
LABORATORY <2 hrs in room temperature
STORAGE BEFORE
PROCESSING 24 hrs in room temperature

22. THROAT SWAB COLLECTION

23. RESPIRATORY TRACT
LOWER

24. SPECIMEN BRONCHOALVEOLAR LAVAGE,
BRONCHIAL BRUSH, BRONCHIAL WASH
CONTAINER
Sterile, screw-top container
SPECIAL
INSTRUCTIONS Anaerobic culture appropriate only if sheathed (protected)
catheter used
TRANSPORT TO
LABORATORY <2 hrs in room temperature
STORAGE BEFORE
PROCESSING 24 hrs in 4°C temperature

25. SPECIMEN SPUTUM
CONTAINER Sterile, screw-top container
PATIENT
PREPARATION Patient should brush teeth and then
rinse or gargle with water before collection.
SPECIAL
INSTRUCTIONS
Sputum collection from deep cough;
At least 3 consecutive sputum specimens are needed,
each collected in 8- to 24- hour intervals,
with at least one being an early morning specimen.
TRANSPORT TO
LABORATORY
<2 hrs in room temperature
STORAGE BEFORE
PROCESSING
24 hrs in 4°C temperature

27. Gastrointestinal Tract

28. SPECIMEN
GASTRIC ASPIRATE
CONTAINER
Sterile, screwcap tube
PATIENT
PREPARATION
Collect in early morning before patient eats or gets out of
bed
SPECIAL
INSTRUCTIONS Most gastric aspirates are on infants or for AFB
TRANSPORT TO
LABORATORY
<15 min in room temperature
STORAGE BEFORE
PROCESSING
<24 hrs/4°C Must be neutralized with sodium
bicarbonate within 1 hr of collection

29. SPECIMEN
RECTAL SWAB
CONTAINER
Swab placed in enteric transport medium
SPECIAL
INSTRUCTIONS
Insert swab ,
1-1.5 cm past anal sphincter;
feces should be visible on swab.
TRANSPORT TO
LABORATORY <2 hrs in room temperature
STORAGE BEFORE
PROCESSING <24 hrs in room temperature

30. Enteric transport medium

31. SPECIMEN
STOOL
CONTAINER
Clean, leakproof container;
transfer feces to enteric transport medium (Cary-Blair)
if transport will exceed 1 hr
TRANSPORT TO
LABORATORY Within 24 hrs in room temperature
STORAGE BEFORE
PROCESSING
24 hrs in 4°C temperature,
<48 hrs in room temperature or 4°C.

32. GENITAL TRACT
FEMALE

33. SPECIMEN
CERVIX
CONTAINER
Swab moistened with Stuart’s or Amie’s medium
PATIENT
PREPARATION
Remove mucus before
collection of specimen.
SPECIAL
INSTRUCTIONS
Do not use lubricant on speculum;
use viral/chlamydial transport medium, if necessary;
swab deeply into endocervical canal.
TRANSPORT TO
LABORATORY
<2 hrs in room temperature
STORAGE BEFORE
PROCESSING
24 hrs in room temperature

34. Cervical swab collection

36. SPECIMEN
URETHRA
CONTAINER
Swab moistened with Stuart’s or Amie’s medium
PATIENT
PREPARATION
Collect 1 hour after patient’s last urination.
Remove exudate from urethral opening.
SPECIAL
INSTRUCTIONS
Collect discharge by massaging urethra against pubic symphysis or
insert flexible swab 2-4 cm into urethra and rotate swab for 2 sec;
collect at least 1 hr after patient has urinated.
TRANSPORT TO
LABORATORY
<2 hrs in room temperature
STORAGE BEFORE
PROCESSING
24 hrs in room temperature

37. SPECIMEN
VAGINA
CONTAINER
Swab moistened with Stuart’s or Amie’s medium
or JEMBEC transport system
PATIENT
PREPARATION
Remove exudate
SPECIAL
INSTRUCTIONS
Swab secretions and mucous membrane of vagina.
If a smear is also required, use a second swab.
TRANSPORT TO
LABORATORY
<2 hrs in room temperature
STORAGE BEFORE
PROCESSING
24 hrs in room temperature

38. Improved transport system for Neisseria gonorrhoeae
The JEMBEC system is made up of a rectangular plastic tray
which contains modified Thayer-Martin medium.
The tray has a well into which a CO2-generating tablet is placed.

39. GENITAL TRACT
MALE

40. SPECIMEN
PROSTATE
CONTAINER
Swab moistened with Stuart’s or Amie’s medium or sterile,
screw-cap tube
PATIENT
PREPARATION
Clean urethral meatus with soap and water and
massage the prostrate through the rectum.
SPECIAL
INSTRUCTIONS Collect secretions on swab or in tube
TRANSPORT TO
LABORATORY
<2 hrs in room temperature for swab; immediately if in tube
in room temperature
STORAGE BEFORE
PROCESSING
24 hrs in room temperature for swab;
plate secretions immediately if in tube

41. SPECIMEN
URETHRA
CONTAINER
Swab moistened with Stuart’s or Amie’s
medium or JEMBEC transport system
SPECIAL
INSTRUCTIONS Insert flexible swab 2-4 cm into urethra and rotate for 2
seconds or
collect discharge on JEMBEC transport system.
TRANSPORT TO
LABORATORY <2 hrs in room temperature for swab; within 2 hrs for
JEMBEC system
STORAGE BEFORE
PROCESSING 24 hrs in room temperature for swab;
put JEMBEC at 37°C immediately on receipt in laboratory

42. Urine

43. SPECIMEN Clean-voided midstream (CVS)
CONTAINER Sterile, screw-cap container Containers that
include a variety of chemical urinalysis preservatives may also be used
PATIENT
PREPARATION
Females:
clean area with soap and water,
then rinse with water;
hold labia apart and begin voiding in
commode;
after several mL have passed, collect
midstream
Males:
clean glans with soap and water,
then rinse with water;
retract foreskin and begin voiding
in commode;
after several mL have passed,
collect midstream.
TRANSPORT TO
LABORATORY
Preserved : within 24 hrs in room temperature
Unpreserved : <2 hrs in room temperature
STORAGE BEFORE
PROCESSING 24 hrs in 4°C temperature
Boric acid is used for Urine sample presevation

44. SPECIMEN STRAIGHT CATHETER (IN AND OUT)
CONTAINER Sterile, screw-cap container
PATIENT
PREPARATION Clean urethral area (soap and water) and rinse (water).
SPECIAL
INSTRUCTIONS
Insert catheter into bladder;
allow first 15 mL to pass;
then collect remainder.
TRANSPORT TO
LABORATORY <2 hrs in room temperature
STORAGE BEFORE
PROCESSING 24 hrs in 4°C temperature

45. SPECIMEN
SUPRAPUBIC ASPIRATE
CONTAINER
Sterile, screwcap container or anaerobic transporter
PATIENT
PREPARATION
Disinfect skin.
SPECIAL
INSTRUCTIONS Needle aspiration above the symphysis pubis
through the abdominal wall into the full bladder.
TRANSPORT TO
LABORATORY
Immediately in room temperature
STORAGE BEFORE
PROCESSING
Plate as soon as received

47. OPHTHALMOLOGICAL SPECIMENS

48. SPECIMEN
CONJUNCTIVA
CONTAINER
Aerobic swab moistened with Stuart’s or Amie’s medium
SPECIAL
INSTRUCTIONS Sample both eyes;
use separate swabs premoistened with sterile saline.
TRANSPORT TO
LABORATORY <2 hrs in room temperature
STORAGE BEFORE
PROCESSING 24 hrs in room temperature

49. SPECIMEN AQUEOUS/ VITREOUS FLUID
CONTAINER Sterile, screw-cap tube
PATIENT
PREPARATION Prepare eye for needle aspiration
TRANSPORT TO
LABORATORY <15 min in room temperature
STORAGE BEFORE
PROCESSING <24 hrs in room temperature
Set up immediately on receipt.

50. SPECIMEN
CORNEAL SCRAPINGS
CONTAINER
Bedside inoculation of BHI 10%
PATIENT
PREPARATION
Clinician should instill local anesthetic before collection
TRANSPORT TO
LABORATORY
<15 min in room temperature
STORAGE BEFORE
PROCESSING
<24 hrs in room temperature
Must be incubated at 28°C (SDA) or
37°C (everything else) on receipt in laboratory.

51. EAR

52. SPECIMEN INNER EAR OUTER EAR
CONTAINER
Sterile, screwcap tube or anaerobic
transporter
Aerobic swab moistened
with Stuart’s or Amie’s
medium
PATIENT
PREPARATION
Clean ear canal with mild soap
solution.
Wipe away crust with sterile
saline.
SPECIAL
INSTRUCTIONS
Aspirate material behind drum with
syringe if ear drum intact;
flexible shaft swab to collect
material from ruptured ear drum
Firmly rotate swab in outer
canal.
TRANSPORT TO
LABORATORY
<2 hrs in room temperature
STORAGE BEFORE
PROCESSING
24 hrs in room temperature 24 hrs in 4° C temperature

53. FOREIGN BODIES

54. SPECIMEN
IUD, IV catheters, ET tube etc.
CONTAINER
Sterile, screw-cap container
PATIENT
PREPARATION
Disinfect skin with alcohol before removal.
TRANSPORT TO
LABORATORY <15 min in room temperature
STORAGE BEFORE
PROCESSING Plate as soon as received.

55. HAIR, NAILS, OR SKIN SCRAPINGS
(for fungal culture)

56. SPECIMEN
HAIR, NAILS, OR SKIN SCRAPINGS
CONTAINER
Clean, screw-top tube
PATIENT
PREPARATION
Nails or skin: wipe with 70% alcohol
SPECIAL
INSTRUCTIONS
Hair: collect hairs with intact shaft
Nails: send clippings of affected area
Skin: scrape skin at leading edge of lesion
TRANSPORT TO
LABORATORY
Within 72 hrs in room temperature
STORAGE BEFORE
PROCESSING
Indefinitely in room temperature

57. Specimen Labeling
Specimens should be labeled with
 patient’s name,
 identifying number (hospital or sample number),
 date and time of collection,
 source,
 the initials of the individual that collected the sample.
 Bio hazard
Enough information must be provided on the specimen label so that the
specimen can be matched up with the test requisition when it is received
in the laboratory.

58. Specimen Requisition
 The specimen (or test) requisition is an order form that is sent to the laboratory
along with a specimen.
 A complete requisition should include the following:
• The patient’s name
• Hospital identification number
• Age and date of birth
• Sex
• Collection date and time
• Ordering physician
• Exact nature and source of the specimen
• Diagnosis (may be ICD-10-CM code)
• Current antimicrobial therapy

60. CRITERIAS FOR REJECTION OF
SPECIMEN
Specimens are unacceptable if any of the following conditions apply:
 The information on the label does not match the information on
the requisition or the specimen is not labeled at all.
 The specimen has been transported at the improper temperature.
 The specimen has not been transported in the proper medium

61. CRITERIAS FOR REJECTION OF
SPECIMEN
 The quantity of specimen is insufficient for testing (the
specimen is considered quantity-not-sufficient [QNS]).
The specimen is leaking.
The specimen transport time exceeds prescribed time or the
specimen is not preserved.
 The specimen was received in a fixative (formalin), which, in
essence, kills any microorganism present.

62. CRITERIAS FOR REJECTION OF
SPECIMEN
 The specimen has been received for anaerobic culture from a
site known to have anaerobes as part of the normal microbiota
(vagina, mouth).
The specimen is dried.
Processing the specimen would produce information of
questionable medical value (e.g., Foley catheter tip).

63. Reference
s
Koneman’s Color Atlas and Textbook of Diagnostic Microbiology.
BAILEY & SCOTT'S Diagnostic Microbiology.

64. THANK YOU…..