Specimen Selection
Select the appropriate specimens for viral isolation based on the clinical symptoms as
outlined in the Viral Specimen Selection Table.
Specimen Collection
Collect specimens as outlined in the Viral
Specimen Collection Table, as soon after onset as possible. The best specimens for
viral isolation are those collected during the first 3 days after onset of symptoms.
Collect all specimens as aseptically as possible. Swabs used to obtain specimens for
viral isolation should be made of sterile cotton or Dacron with plastic or aluminum
shafts. Avoid swabs with wooden shafts. DO NOT USE CALCIUM ALGINATE SWABS FOR VIRAL
ISOLATION SPECIMENS.
All swabs and small tissue specimens for viral isolation should be placed in viral
transport media (VTM). Typically, viral transport media consists of protein such as bovine
serum albumin or gelatin to stabilize the virus and antimicrobial agent in a buffered salt
solution. Transport medium can be purchased commercially. Swabs should be placed in 2-3 ml
of medium, larger volumes of medium should not be used because of the dilution effect.
Place each specimen into a separate, leakproof container labeled with the
patients name, the collection site, and the date of collection. For additional
information on the packaging of specimens, see The
TDSHS Manual of Reference Services.
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Specimen Submission Form
Complete a G-1a specimen submission form for each specimen submitted to the Viral
Isolation Section. Specimens will not be tested without a completed G-1a form. Follow the
Guidelines for Specimen Shipping and Mailing for
shipping specimens to the laboratory. Forms may
be obtained from the Laboratory Office.
Specimen Transport and Handling
Specimens for viral isolation should be transported to the laboratory as quickly as
possible after collection since the chance of isolating a virus declines with increased
time in transit. Samples containing labile viruses (i.e. respiratory syncytial virus,
cytomegalovirus and varicella zoster virus) at low titers are those most likely to show
loss of infectivity with delayed transport. For short-term transit or storage, specimens
should be kept at refrigerator temperatures on wet ice or cold packs. If a longer transit
time (greater than 2-3 days) is anticipated, the specimen should be frozen at -70° C and transported to the laboratory on dry ice.
Specimens for chlamydia isolation should be put in chlamydia transport media and kept
at refrigerator temperatures on wet ice or cold packs if they can be processed within 48
hr after collection. If not, they should be frozen at -70° C.
Specimens for direct detection (i.e. EM, PCR) should be kept at refrigerator
temperatures on wet ice or cold packs. Freezing specimens for direct detection is not
optimal.
Specimens must be transported in accordance with all safety and labeling regulations. The
TDSHS Manual of Reference Services includes detailed instructions for
packaging and shipping specimens and isolates.
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Testing Methods
Viral Isolation
Viruses cause a wide range of clinical symptoms and are shed from various anatomical
sites during the infection. The site selected for specimen collection should be based on
the clinical symptoms (see Viral
Specimen Collection Table) and the seasonal and/or local epidemiological trends. Once a specimen
is received in the laboratory, it is processed and inoculated into living host systems
(i.e. cell cultures). Inoculated host systems are incubated to allow any virus present in
the specimen to grow and replicate. During the incubation period, if observation indicates
the presence of a virus, additional laboratory tests are performed to confirm the
viruss presence and identity.
Reference Culture: Viral Isolation
Laboratories who would like final identification and/or confirmation of their results
may send isolates to the TDSHS Laboratory.
Isolates can be sent either frozen on dry ice or at ambient temperatures in cell
culture monolayers that are shipped when CPE is first evident in the monolayers. Fill the
tube or vial with media to prevent drying of the monolayer during transit if the tube or
vial gets turned during transit.
Although the Viral Isolation laboratory will confirm an adenovirus identification,
prior approval is required for typing of adenovirus isolates since they are ubiquitous.
Typing of adenoviruses is limited to situations when the virus is the suspect cause of
death, is involved in a significant outbreak, or under other unusual circumstances.
Contact the Medical Virology Branch of the Laboratory at (512) 458-7515 to discuss the
possibility of typing an adenovirus isolate.
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Electron Microscopy
The electron microscope has been an invaluable tool for the basic understanding of
viruses. Clinical application of the electron microscope has yielded another use: the
rapid and unequivocal recognition of viruses. The results of studies with the electron
microscope during the past have ranged from describing a new virus for the first time to
the rapid recognition of viruses in clinical specimens.
The use of the electron microscope in the detection and identification of viruses
offers many advantages. Using a negative stain technique, results are generally available
in hours, not days. The infectivity of the virus does not need to be maintained, and
viruses which are difficult or impossible to culture can be identified. The approach is
open-ended which allows a wide variety of agents to be visualized, and since the actual
agent is visualized, there are no false positive results.
Specimens for electron microscopy should not be placed into transport medium or
fixative. Package specimens on wet ice or cold packs and transport as quickly as possible.
Do not freeze specimens for electron microscopy.
See
sample electron micrographs
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Dengue Isolation
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of the Texas Department of State Health Services. These sites may also not be accessible to
people with disabilities.
Dengue is an
acute febrile illness resulting from infection with 1 of 4 closely related arboviruses
(dengue serotypes 1,2,3, and 4).
A laboratory diagnosis of dengue is obtained by a rise in antibody titer between acute
and convalescent sera (drawn at least 14 days apart), the presence of IgM, the isolation
of dengue virus from an acute phase serum specimen, or the detection of RNA in the serum
by PCR.
Submission of specimens for dengue isolation should be coordinated with Infectious Disease
Epidemiology and the Laboratory.
Serum is the optimal specimen for isolation of dengue virus. Isolation of dengue virus
from serum is most successful when the blood is collected within 3 days of onset of
symptoms, although virus has been recovered from sera collected up to 5 days after onset.
Ship the serum cold, on ice packs, by overnight carrier to the TDSHS Laboratory if the
specimens will arrive at the laboratory within 2-3 days of collection. If the specimen
will be delayed more than 2-3 days, ship the specimens frozen, on dry ice. However, if the
specimen is being shipped as whole blood rather than serum, send it only on cold packs. DO
NOT FREEZE WHOLE BLOOD OR SEND WHOLE BLOOD ON DRY ICE. This will lyse the red blood cells
and be toxic to the cell cultures, compromising the ability to isolate the virus.
Specimens will be rejected for isolation if the specimen is not received in a sterile
container, there is excessive delay in transport, the specimen is not refrigerated or
frozen during transport, or if the date of collection is not within 5 days of onset of
symptoms.
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