MLAB 2434 –
MICROBIOLOGY
KERI BROPHY-MARTINEZ
General Concepts in Specimen
Collection and Processing
General Concepts in Specimen Collection and
Processing (cont’d)

Basic Principles of Specimen
Collection
Specimen should be taken in acute
phase of infection AND before
antibiotics are administered
 Written order must specify site of
culture (example: wound on left arm)
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General Concepts in Specimen Collection and
Processing (cont’d)
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Avoid normal flora and colonizing
organisms
Collect the appropriate quantity of
specimen
Package specimen in correct transport
media
Label with patient information & source
Transport the specimen to the lab asap to
avoid deterioration
General Concepts in Specimen Collection and
Processing (cont’d)

Appropriate Collection Techniques
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Aspirates and tissues
• Aspirates and tissues present few
problems, if collected using sterile
technique
• Lesions, wounds and abscesses; cultures
should be from as deep in the wound as
possible
General Concepts in Specimen Collection and
Processing (cont’d)

Swabs
• Used only as a last resort.
Dacron/polyester swabs preferred
• Steps
• Clean wound
• Explore wound
• Obtain fresh and quality culture material
• Should be placed in a holding medium to
protect pathogens without permitting
multiplication during transport
General Concepts in Specimen Collection and
Processing (cont’d)

Patient Education and Preparation
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If patient is responsible for collecting
specimen, good instructions are critical
Urine – midstream clean catch first
morning specimen
Sputum – collect sputum NOT spit,
morning specimens are preferred, deep
cough
Stools – usually 3 vials and at least 4 days
after barium X-rays, swabs are
discouraged
General Concepts in Specimen
Collection and Processing

Labeling & Requisition
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Specimen must have patient name,
time, date, source, location
Requisition
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must accompany specimen, include
diagnosis & antimicrobial history
General Concepts in Specimen
Collection and Processing

Safety
Standard Precautions
 Specimen processing should be
conducted in a Class II safety
cabinet (hood)
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General Concepts in Specimen Collection and
Processing (cont’d)

Preservation, Storage, and Transport of
Specimens
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Concerns
• Overgrowth
• Death of microorganisms: deliver to lab within
30 minutes of collection
• Inaccurate quantitation
• Loss of organisms from drying
• Protection from oxygen
• Protection from clotting
• Safety of transporter
General Concepts in Specimen Collection and
Processing (cont’d)

Anticoagulants
• Needed in any specimen that might clot
(blood, serum, joint fluids)
• Sodium polyanethol sulfonate (SPS)
• Most common anticoagulant in micro
General Concepts in Specimen Collection and
Processing (cont’d)

Preservatives
• Urine – boric acid
for up to 24 hours
• Stool – phosphatebuffered saline
(PBS) or O & P kit
General Concepts in Specimen Collection and
Processing (cont’d)

Use of Holding and Transport Media
 Media should provide viability without allowing
multiplication of bacteria
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Media should maintain pH, provide proper
atmospheric conditions and prevent drying
General Concepts in Specimen Collection and
Processing (cont’d)

Types of collection devices
General Concepts in Specimen Collection and
Processing (cont’d)

Blood culture bottles
General Concepts in Specimen
Collection and Processing (cont’d)

Sterile container for sputum, urine,
and other specimens
General Concepts in Specimen Collection and
Processing (cont’d)

Aspirates are usually collected and
transported in syringes
General Concepts in Specimen Collection and
Processing(cont’d)
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Storage of Specimens
• Urine, viral blood specimens, catheters and
swabs should be refrigerated (4oC)
• Blood and CSF should be processed ASAP
• Specimens for fungus cultures can be kept at
room temperature
• Respiratory and stool cultures should be
processed ASAP if at all possible, but
refrigerated if immediate processing is not
possible
• Anaerobes, genital, ear, eye cultures can be
held at room temperature
General Concepts in Specimen Collection
and Processing (cont’d)

Mailing specimens
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Regulated by U.S. Dept. of Health and
Human Services & U.S. postal Service
National guidelines issued by Department
of transportation
Retraining of employees must occur every
2 years
Packaging Infectious
Substances
General Concepts in Specimen Collection and
Processing (cont’d)

Unacceptable Specimens
 Labels on requisition and on specimen must match
(unlabeled, mislabeled)
• Noninvasive vs. invasive specimens
 Rejected specimens
• Leaking
• Syringes with needles attached
• Stools contaminated with urine or barium
• Anaerobic cultures on inappropriate sources
• Unpreserved specimens over 2 hours old
General Concepts in Specimen Collection
and Processing (cont’d)
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Unacceptable specimens
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Refrigerated blood cultures
Dried-up specimens
Specimens in formalin
Improper collection
Delay in transport
**Do not discard rejected specimen until a new one
is submitted, if original specimen is cultured,
notate in patients report
Do you feel like this?
General Concepts in Specimen Collection and
Processing
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Processing of Clinical Samples for
Optimal Organism Recovery
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Prioritization
• Level 1- Critical/invasive ( CSF)
• Level 2 – Unpreserved (sputum)
• Level 3 – Accuracy of quantitation
affected (urine, no preservative)
• Level 4 – Protected/Preserved (urine
with preservative)
General Concepts in Specimen Collection and
Processing (cont’d)

Gross Examination of Specimens
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Check specimen for:
• Volume submitted
• Presence of blood or mucous
• General appearance: cloudy, clear
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Check requisition against sample
General Concepts in Specimen Collection and
Processing (cont’d)

Direct Examination Techniques
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Direct Microscopic Examination
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Determine quality of specimen
Diagnose infectious disease
Guide routine culture interpretation
Dictate the need for nonroutine
processing
• Guide antibiotic therapy
General Concepts in Specimen Collection and
Processing (cont’d)
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Smear Preparation
Tissues
 Swabs
 Aspirates and body fluids
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Single drop smear
Centrifuged sediment smear
Layered smear
Cytocentrifuged smear
Additives
General Concepts in Specimen Collection and
Processing (cont’d)

Direct Smears NOT useful
Throats, nasopharyngeal swabs
 Urine
 Female genital tract
 Stools
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References
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Engelkirk, P., & Duben-Engelkirk, J. (2008). Laboratory Diagnosis of
Infectious Diseases: Essentials of Diagnostic Microbiology . Baltimore, MD:
Lippincott Williams and Wilkins.
http://catalog.bd.com/bdCat/search.doCustomer?searchText=urine+culture&
typeOfSearch=0&viewPageNum=0&sortByField=Category&x=0&y=0
http://coe.berkeley.edu/news-center/publications/engineeringnews/archive/engineering-news-vol-79-no-9f/need-a-study-break-here-area-few-ideas
http://www.firstqualitylaboratory.com/pages/specimenpreparation.html
http://www.lookfordiagnosis.com/images.php?term=Polyanetholesulfonate&lan
g=1
Mahon, C. R., Lehman, D. C., & Manuselis, G. (2011). Textbook of Diagnostic
Microbiology (4th ed.). Maryland Heights, MO: Saunders.