Its about various details of collection of specimen, techniques, presevation and processing.

1. WINTER
GE RAL CONCE T F
NE P S OR SP CIM N
E E
Template SPECIMEN
COL E ION , H
L CT ANDLING OF
AND SP CIM N P
E E ROCE SSING
By
Serena Lobo
DOS in Microbiology
Mysore University

2. INTRODUCTION
Laboratory investigations use scientific methods to help us
arrive rapidly at a diagnosis of an infectious disease.
Once the causative agent for a particular infection has been
identified, appropriate and rational therapy can be instituted.
Before requesting for laboratory investigations, the clinician
should have arrived at a provisional diagnosis of the probable
cause of an infection, based on accurate and detailed history
taking and careful physical examination.
Anyone connected with medical care, whether a doctor,
nurse or a laboratory technician may collect a specimen.
Hence all medical personnel should have a clear
understanding of the principles behind the collection of
appropriate specimen for microbiological examination.

3. GE RAL CONSIDE AT
NE R IONS R GAR
E DING COL E ION
L CT
OF SP CIM N
E E
ῼCollect culture specimens before antibiotic therapy.
ῼThe specimen must be representative of the infection
with as little contamination from normal flora as
possible.
ῼUtilize appropriate collection devices, use sterile
equipment and aseptic technique to collect specimens.
ῼCollect an adequate amount of specimen.
ῼCollect specimens in sturdy, sterile, leak proof
containers.
ῼClear and specific instructions on proper collection
techniques must be given to patients when they must
collect their own specimens.

4. ῼThe specimen must be transported promptly and
maintained in a manner that allows survival of fastidious
organisms and prevents overgrowth by more hardy
organisms.
ῼSpecimens that are collected in syringes are best
maintained by dead-capping the syringe, expelling
excess air, and transporting to the laboratory in the
syringe.
ῼA fluid or tissue specimen is generally a more desirable
specimen than a swab.

5. DOCUMENTATION
WINTER
Each tube or container with obtained specimen must be
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provided with a stick - on label with written name of the
patient, date of taking the specimen and exact description of
the source of the specimen.
Other details to be noted are:
♣ Name and address of the patient, his/her birth number,
his/her health insurance number.
♣Number of days from the onset of symptoms of disease.
♣ Clinical diagnosis.
♣Exact description of the obtained specimen, date (hour) of
taking it and sending to the lab.
♣Requested examination.
♣The name and stamp of the ordering doctor, his/her identity
number and address of the health facility .

6. TRANSPORTATION OF SPECIMEN
•Specimen should be packed well and labeled
‘HAZARDOUS’ on the packing.
•The specimen must be delivered to the laboratory as
soon as possible
•To overcome any delay and drawbacks due to delay in
reaching the microbiology department, the following
methods may be used.
☺Transport media
☺Refrigeration
☺Freezing

7. TRANSPORT MEDIA
oTransport media is a medium for transporting clinical
specimen to the laboratory for examination for examination.
oTransport media should fulfill the following criteria:
♠Temporary storage of specimen being transported to the
laboratory for cultivation.
♠Maintain the viability of the organisms in the specimen
without altering their concentration.
♠Contains only buffers and salts.
♠Lack of carbon, nitrogen, organic growth factors so as to
prevent microbial multiplication.
♠Transport media used in the isolation of anaerobes must
be free of molecular oxygen.
Examples :Thioglycolate broth for strict anaerobes.
Venkat-Ramakrishnan medium for Vibrio cholarae.

8. CONTAINERS FOR COLLECTING SPECIMEN

9. TECHNIQUESOF SPECIMEN COLLECTION
FROM SKIN: WINTER
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☻The area of the skin from which specimen is collected is
first cleaned using soap and water.
☻Swabs are firmly rubbed over the affected part of the
skin and sent at once to the laboratory for processing.
☻Only swabs which have been moisturized in sterile
broth or saline should be used.
☻If a viral infection is suspected, crusts, scabs or vesicle
fluids are collected in capillary tubes.
☻If a fungal lesion is suspected, infected hairs and nails
as well as scrapings from the affected part of the skin may
be collected.

10. F OM T E UP E RE IR ORY T ACT
R H P R SP AT R
Oral cavity :
Swabs are rubbed firmly over the
ulcerated or patch like lesions.
Anterior nares:
If pus is present is present, collect it
on the swabs.
If no pus is present, moisten the
swabs and then swab the anterior
nares.
Troat:
The mouth is held wide open and
the tongue is depressed. Swabs are
firmly rubbed over the tonsils and
pharyngeal mucosa.

11. Nasopharynx :
A pernasal swab is used. This is made from fairly flexible wire
which is bent at one end and is covered with sterile cotton at this
end. The swab is carefully passed through the cavity till it
impinges on the nasopharynx and then firmly rubbed over the
area.
Epiglottis :
Swabs are firmly rubbed over the inflamed and ulcerated areas.
Before collecting the swabs precautions should be taken to
maintain the airways in case a laryngeal spasm suddenly
occurs. Ideally the swabs should be taken by an ENT surgeon.
Paranasal sinuses:
If pus is present in these sinuses , it is collected on swabs or
aspirated with a syringe and needle.

12. FROM LOWER RESPIRATORY TRACT
ᵜCommonly only sputum is collected. This should be
coughed up from far down the bronchial tree and
expectorated immediately. It should not be mixed with
saliva or other oropharyngeal secretions.
ᵜAn even better specimens material that is aspirated
directly from the bronchi or trachea. This is collected by
using a flexible fibreoptic bronchoscope.
ᵜAn alternative is to collect bronchial washings.
ᵜThese procedures should be conducted by an
experienced pulmonologist or cardio thoracic surgeon.

13. FROM THE GASTROINTESTINAL TRACT
WINTER
Feces is collected on a sterile wide mouthed , leak
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proof container preferably provided with a small spoon
to transfer material to any other container.
If feces are semi solid , a small quantity is sufficient:
if liquid, it should fill a third of the specimen jar.
Fresh feces may be collected by inserting a short
catheter into the rectum.
A rectal swab can be collected as an alternative : this
is gently inserted into the rectum and turned clockwise
and anti clockwise ensuring the rectal mucosa is firmly
rubbed.

14. FROM URINARY TRACT
Contamination of urine by bacteria colonizing the
distal parts of the urethra and the perenium is
prevented by clean catch technique.
Here the periurethral area is cleaned twice with soap
and water.
The urethra is flushed by voiding the first portion of
the urine , which is discarded.
The subsequent mid stream urine is collected
directly in a sterile container and is used for culture
and colony counting.
It is essential for cultural purposes that urine be
processed within one hour of collection or stored in a
refrigerator at 40 C till it can be cultured.

15. FROM THE GENITAL TRACT
In women:
The best specimen is a cervical swab. A sterile bivalve
speculum is moistened with warm water and inserted into the
vagina. The cervical mucus plug is removed with a cotton ball
and forceps: the external surface of the cervix is then cleaned
with a large cotton swab.
In men:
If the patient has purulent urethral exudates, culture is not
necessary. A Gram stained direct smear is enough for clinical
diagnosis of Gonorrhea if intracellular gram negative diplococcic
are seen. In men, uncentifuged first voided urine may be
cultured for N. gonorrhea .

16. FROM CENTRAL NERVOUS SYSTEM
ᴪThe Cerebro Spinal Fluid is usually collected from
lumbar puncture.
ᴪThe dural sheath is pierced by a needle and CSF is
allowed to drip from it into a sterile container.
ᴪIt is essential to avoid introducing contaminating
organisms into the subdural space or the specimen.
ᴪTherefore the procedure should be viewed as a minor
surgical operation.
ᴪThe technique should be vigourously aseptic.

17. WINTER
FROM THE BLOODSTREAM
Blood is collected by strict aseptic technique and care
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should be taken to avoid introducing organisms into the
blood stream as well as prevent contamination of the
specimen.
The vein from which the blood is to be taken should be
clearly seen and the skin overlying the vein is vigorously
wiped with 70% alcohol.
Once the area is dry, the specimen is collected using a
perfectly dry, sterile syringe and needle.
After this, the area is cleaned with 70% alcohol and
finally wiped with povidine-iodine in alcohol.

18. FROM PLEURAL AND PERITONEAL CAVITIES
۞. A sample of pleural fluid is collected by a doctor with a
syringe and needle using a procedure called thoracentesis.
۞The health care provider will clean the skin around the
insertion site and drape the area.
۞A local anesthetic is injected into the skin.
۞The thoracentesis needle is inserted above the rib into the
pocket of fluid. As fluid drains into a collection bottle, many
people cough a bit as the lung reexpands to fill the space
where fluid had been.
۞This sensation normally lasts for a few hours after the test
is completed.

19. FROM THE CONJUNCTIVA, LID MARGINS, CORNEA
AND INTRAOCULAR STRUCTURES
Material from the lid margin is collected by firmly rubbing a pre
moisturized swab from the Inferior lid margin to superior lid
margin. The swab is inoculated into appropriate culture media.
Any visible purulent conjunctival discharge is collected on a
swab and inoculated at once.
If pus discharge is not present, smears can be made and
culture media is inoculated with material taken directly from the
conjunctival surface by a sterile bacteriological loop (made of
platinum and not nichrome).
FROM THE EAR
A swab can be used to collect material from the external ear.
The swab on the wire is inserted into the external auditory canal
with a twisting motion to collect a secretion from it.

20. CRITERIA FOR REJECTION OF SPECIMEN
WINTER
Criteria should be developed by a laboratory on the
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basis of which the processing of a specimen may not
be done by the laboratory. The following are some
examples:
§ Missing or inadequate identification.
§ Insufficient quantity.
§ Specimen collected in an inappropriate
container.
§ Contamination suspected.
§ Inappropriate transport or storage.
§ Unknown time delay.
§ Haemolysed blood sample.

21. PROCESSING OF SPECIMEN
¶ Once the appropriate specimen has been
collected, they are processed as speedily as
possible to ensure that organisms that the
organisms do not die before being transferred
to the culture media, and that the reports are
available at the earliest.
¶ Generally material is placed on slides and
stained appropriately, and then examined under
the microscope.
¶ If bacteria and fungi are detected, specific
therapy can be started at once. Material is also
inoculated onto appropriate culture media.

22. CONCLUSION
Specimen collection is an essential prerequisite that
the laboratory must be given sufficient clinical
information to guide the microbiologist in selection of
the suitable media and appropriate techniques.
Also, it is important for the clinician to appreciate the
limitations and potentials of the microbiological
laboratory and to realize that a negative report does
not necessarily invalidate the diagnosis.
It is essential that a close cooperation and frequent
consultation among the clinician, nurse and
microbiologist are the rule rather than exception.