SPECIMEN HANDLING, COLLECTION,
TRANSPORT, PRESERVATION, PROCESSING,
AND DISPOSAL - CRITERIA FOR
ACCEPTABILITY OF SAMPLES
FACTORS THAT MAY ALTER TEST RESULT
GROUP 10 - PARASITOLOGY
MARTINEZ, KIM CLAIRE O.
CUERDO, MARRY FLOR T.
FONTANILLA, NATHALIE T.
MARTISANO, ANTONETTE C.
OBJECTIVES
1. To discuss the basic techniques in proper specimen handling.
2. To properly identify the specimens to be accepted and
rejected.
3. To maintain the integrity of the specimen to be send in the
laboratory.
4. To know safety precautions in handling specimen.
5. To learn the important factors to avoid specimen alteration.
TOPIC OUTLINE
SPECIMEN HANDLING
SPECIMEN COLLECTION
SPECIMEN TRANSPORT
PRESERVATION OF THE SPECIMEN
SPECIMEN PROCESSING
DISPOSAL OF SPECIMEN
ICE BREAKER
TIME
GUESS WHAT
PARASITE?
___ ___ ___ ___ ___ ___ ___ ___ ___ ___ ___ ___ ___ ___
Answer:
Taenia saginata
___ ___ ___ ___ ___ ___ ___ ___ ___ ___ ___ ___ ___ ___ ___
Answer:
Hymenolopis nana
ova
___ ___ ___ ___ ___ ___ ___ ___ ___ ___ ___ ___ ___ ___ ___ ___
___ ___ ___ ___ ___
Answer:
Diphyllobothrium
latum
SPECIMEN HANDLING
Laboratory examination may influence whether organisms will be
identified. The stool should be delivered to the laboratory as soon as
possible after collection or a should be portion placed immediately in a
preservative. Trophozoites, the motile and reproductive form of some
amebae, or eggs of some helminths may disintegrate if not preserved or
examined within a short time. Because many intestinal organisms are shed
into the stool irregularly, a single stool specimen may be insufficient to
detect an intestinal parasite.
SPECIMEN COLLECTION
The appropriate collection container for feces should be clean, dry, sealed tightly,
and waterproof (e.g., a plastic container with lid). Commercial systems that
incorporate collection container and preservatives are also available. Stool
specimens should never be collected from bedpans or toilet bowls; such practice
might contaminate the specimen with urine or water, resulting in the destruction of
trophozoites or introduction of free-living protozoa. As an alternative, the specimen
could be collected on a clean piece of waxed paper or newspaper and transferred to
the container. Another alternative is to use a disposable collection container that can
be fitted under the toilet bowl rim. The specimen should be submitted as soon as
possible after passage. Information on the container should include the patient's
name and the date and time the stool was collected.
SPECIMEN COLLECTION
Stool specimens for parasites should be collected before a barium enema, certain
procedures using dyes, or the start of antimicrobial therapy. Antimicrobials can
reduce the number of organisms present. If the patient has undergone a barium
enema, stool examination should be delayed for 7 to 10 days because barium
obscures organisms when specimens are examined microscopically, even after
concentration procedures. If a purged specimen is to be collected, it is recommended
that a saline or phosphosoda purgative be used because mineral oil droplets interfere
with identification of parasites, especially protozoan cysts, an infective dormant form
resistant to environmental stress. The second or third specimen after the purge is
more likely to contain trophozoites that inhabit the cecum.
Stool Specimens – Specimen Collection According to CDC
1. Collect the stool in a dry, clean, leakproof container. Make sure no urine, water, soil or other material gets in the
container.
2. The image on the right demonstrates the distribution of protozoa in relation to stool consistency and should be
taken into consideration when specimens are received.
3. Fresh stool should be examined, processed, or preserved immediately. An exception is specimens kept under
refrigeration when preservatives are not available; these specimens are suitable for antigen testing only.
4. Preserve the specimen as soon as possible. If using a commercial collection kit, follow the kit’s instructions. If kits
are not available, the specimen should be divided and stored in two different preservatives, 10% formalin and PVA
(polyvinyl-alcohol), using suitable containers. Add one volume of the stool specimen to three volumes of the
preservative.
5. Insure that the specimen is mixed well with the preservative. Formed stool needs to be well broken up.
6. Insure that the specimen containers are sealed well. Reinforce with parafilm or other suitable material. Insert the
container in a plastic bag.
7. Certain drugs and compounds will render the stool specimens unsatisfactory for examination. The specimens
should be collected before these substances are administered, or collection must be delayed until after the effects
have passed. Such substances include: antacids, kaolin, mineral oil and other oily materials, non-absorbable
antidiarrheal preparations, barium or bismuth (7-10 days needed for clearance of effects), antimicrobial agents (2-3
weeks), and gallbladder dyes (3 weeks).
8. Specimen collection may need to be repeated if the first examination is negative. If possible, three specimens
passed at intervals of 2-3 days should be examined.
SPECIMEN TRANSPORT
Clinical specimens should be transported to the laboratory immediately to
increase the likelihood of finding intact organisms. Because a lag time often
occurs between collection of the specimen and its arrival in the laboratory,
most facilities routinely use preservatives for collection and transport. This
approach ensures that any parasites present maintain their morphology and
can be identified after processing.
SPECIMEN PRESERVATION
Preservation several methods are available for stool preservation if the specimen will not be
delivered immediately to the laboratory. The preservative used is determined by the
procedure to be performed on the fecal sample. Regardless of the preservative used, the
ratio of three parts preservative to one part feces should be maintained for optimal fixation.
Table 28.1 presents some of the more common preservatives and their appropriate use. The
time that the stool was passed and the time it was placed in the fixative should be noted on
the laboratory requisition and container. A commercially available two-vial system using
polyvinyl alcohol (PVA), a resin polymer, in one vial and 10% formalin in the other vial is
commonly used. The system comes with patient instructions and a self-sealing plastic bag
for transport. The classic PVA fixative, which consists of mercuric chloride (for fixation) and
PVA (to increase adhesion of the stool to the slide), was traditionally used when a
permanently stained smear was to be made.
SPECIMEN PROCESSING
Diagnostic parasitology procedures designed to detect organisms in clinical
specimens typically depend on morphologic criteria and visual identification
(Evolve Procedures 46.1 to 46.10). Many clinical specimens, such as those from
the intestinal tract, contain numerous artifacts that complicate the differentiation
of parasites from surrounding debris. Specimen preparation may require
concentration methods designed to increase the chance of finding the
organism(s) by removing some fecal debris. Microscopic examination requires
review of the prepared clinical specimen using multiple magnifications;
organism identification also depends on the skill of the microbiologist. Final
identification is based on microscopic examination of stained preparations to
identify key characteristics of the parasite form. (Table 46.3 includes specific
details on specimen processing.
Specimen Disposal
Wastes should be separated according to the following criteria and
necessary actions should be taken.
1. Infectious, potentially infectious, recombinant or synthetic nucleic
acid biological wastes
2. Non-infectious biological wastes
1-Infectious, Potential Infectious, Recombinant or Synthetic Nucleic Acid
biological wastes: Such wastes are substances that contain or are
contaminated with:
• Human, animal and plant pathogens
• Recombinant or synthetic nucleic acids and recombinant organisms
• Laboratory and clinical wastes containing human and animal blood, blood
products, tissues, cell cultures and other potentially infectious materials
• Cultures
Infectious, potentially infectious, recombinant or synthetic nucleic acid biological
laboratory wastes must be inactivated before removal from the facility. Although
combustion and chemical inactivation are appropriate in some cases, the most
preferred method is steam sterilization (autoclave). The storage of all wastes in
this category which are not inactivated should be in the laboratory. Infectious or
pathogenic waste should be kept in closed/enclosed waste cans. Such wastes
should not be stored for more than 24 hours until inactivation. Biological waste
bins or bags should be marked with a biohazard mark. Full or partially filled waste
bins should not be kept for more than 30 days
2-Non-infectious biological wastes Such wastes are used in clinical and
research laboratories and are not defined as infectious biological wastes:
• Laboratory materials (cell culture tubes and flasks, vials, centrifuges and test
tubes)
• Unused medical devices • Gloves and other protective equipment the
personnel
• Blood, blood products or tissues that are not contaminated with or are not
known to be contaminated with pathogens.
Inactivation of such wastes is not necessary before being removed from the
plant. Such products should be placed in red biological waste containers or
sachets
SPECIMENS AVAILABLE FOR PARASITIC EXAMINATIONS
STOOL
Most common utilize specimen in the parasitology laboratory.
BLOOD
Plasmodium spp., Babesia spp., Trypanosoma spp., Leishmania spp., Wuchereria bancrofti,
Brugia spp., Loa, and Mansonella spp.
SPUTUM
First morning specimen
Paragonimus westermani, Strongyloides stercoralis
Other parasitic infections that may be found in sputum samples include microsporidia, E.
histolytica, Entamoeba gingivalis, Ascaris lumbricoides, and hookworm
SPECIMENS AVAILABLE FOR PARASITIC EXAMINATIONS
URINE
For the detection of Schistosoma haematobium eggs and may also yield Trichomonas
vaginalis trophozoites.
CSF
Trypomastigotes of Trypanosoma cruzi, Trypanosoma brucei rhodesiense, and
Trypanosoma brucei gambiense
Trophozoites of Naegleria
TISSUE BIOPSY
Muscle biopsy specimens for Trichinella spp.
SPECIMENS AVAILABLE FOR PARASITIC EXAMINATIONS
PERIANAL SWAB
Used to recover eggs of Enterobius vermicularis.
TISSUE ASPIRATES
1. Duodenal Aspirate
There are occasions when duodenal aspirates are better specimens to use in the diagnosis
of Giardia lamblia and Strongyloides stercoralis.
2. Cutaneous or Skin Aspirates
cutaneous leishmaniasis
CRITERIA FOR ACCEPTABILITY OF
SAMPLES
The specimen container should be properly labeled.
Stool specimens should be collected in a clean, watertight container with a tight-fitting
lid.
The acceptable amount of stool required for parasite study is 2 to 5 g.
Urine should not be allowed to contaminate the stool specimen.
Stool should not be retrieved from toilet bowl water.
Deliver samples to the laboratory within a specified timeframe to minimize parasite
deterioration.
FACTORS THAT MAY ALTER TEST RESULT
Sample rejection criteria would include the following (although testing of specific samples may be done upon consultation with the
Clinical Lead):
1. Wrong sample type
Plasma instead of serum or EDTA blood for most serology tests (unless discussed beforehand with the Parasitology Clinical Lead).
Blood samples other than Citrate blood for Microfilarial microscopy
Blood samples other than EDTA blood for malaria microscopy and PCR
Peripheral blood for Leishmania microscopy and PCR (except where specifically agreed with the Clinical Lead)
2. Sample incorrectly stored/treated
Refrigerated stool sample for stool culture
Fixed sample for Leishmania culture (PCR will be performed)
Fixed sample for stool PCR (Microscopy will be performed)
3. Contaminated or broken container/slide
Container contaminated due to leaking sample.
Container/slide poses safety risk due to breakage.
4. Insufficient sample volume (especially for Stronygloides Culture which requires around 20ml of stool)
5. Heavily lipemic or hemolyzed serum sample
6. Sample delivery delayed beyond viable processing time
15 minutes for Hot Stool sample
24 hours for Trypanosomal blood microscopy
7. Sample unaccompanied by request form or accompanied by an incorrect or incorrectly filled out request form. Such errors
would include lack of specific tests requested. Please ensure that suitable travel history is completed as specified by individual
tests (such as African and South American Trypanosomiasis).
REFERENCES:
Bailey & Scott's Diagnostic Microbiology 15th edition
Mahon 6th edition
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