**FACT SHEET**

**SARS Laboratory Diagnostics**

**When to Test for SARS**

In the absence of documented SARS transmission, diagnostic testing for
SARS-associated coronavirus (SARS-CoV) should NOT be considered unless
the clinician and health department have a high index of suspicion for
SARS (e.g., a hospitalized pneumonia patient has a possible SARS
exposure during travel and no other explanation for their pneumonia).

**Successful SARS Diagnosis**

A successful laboratory diagnosis of SARS infection requires obtaining
the correct type of specimen(s), based on the stage of the illness,
using validated assay methods, and understanding the strengths and
limitations of the diagnostic assays used. All laboratory results should
be interpreted in the context of the clinical and epidemiologic
findings.

**Current Diagnostic Methods for SARS-CoV**

Laboratory assays for SARS-CoV are based on either the detection of the
virus or virus products, or detection of an antibody response to viral
infection. Isolation in Vero E6 cells and electron microscopy played a
critical role in the early identification of SARS-CoV, however, these
methods are not suitable for routine diagnoses because they lack
sensitivity and viral culture requires biosafety level III containment.
Current detection methods for SARS-CoV include real-time reverse
transcription polymerase chain reaction (RT-PCR) assay for detection of
viral RNA and enzyme immunoassay (EIA) for detection of antibodies to
SARS-CoV. The real-time RT-PCR assay is highly sensitive, detecting
between 1 and 10 RNA transcript copies per reaction, and utilizes primer
& probe sets to three independent sites along the SARS-CoV genome to
assure specific detection of SARS CoV. Serology is the "gold-standard"
for diagnosis of SARS-CoV infection. The SARS EIA uses a lysate of
SARS-CoV infected Vero E6 cells as antigen. Serosurveys with the SARS
EIA have demonstrated low or undetectable levels of antibody to the
SARS-CoV in the general population. No cross-reactivity has been
observed in validation studies with serum specimens containing
antibodies to other human coronaviruses.

**Specimen Selection for RT-PCR**

Detection of SARS-CoV infection by RT-PCR within the first few days
following onset of symptoms has been problematic because of extremely
low virus titers in respiratory specimens during the early stages of
infection. To increase the rate of detection of SARS positive cases, it
is recommended that more specimens and multiple specimen types be
collected. Respiratory tract specimens are valuable, with lower
respiratory tract specimens (e.g., sputum, bronchial alveolar lavage)
preferred over upper respiratory tract specimens (e.g., washes,
aspirates, swabs) when available. New evidence suggests that blood
collected within the first few days of onset of illness may be useful
for detection of SARS-CoV RNA. Stool may be the best specimen for
detection of SARS-CoV beginning about one week after onset of illness
and may continue to be positive for several weeks thereafter.

For the patient to be considered positive for SARS-CoV infection by
RT-PCR, current recommendations specify that a positive result on one
specimen should be confirmed in a reference laboratory and a second
specimen, either from another site or collected at a different time,
should also be confirmed positive.

**Special Considerations for Interpreting SARS RT-PCR Results**

The CDC real-time RT-PCR assay has proven both sensitive and specific
for detection of SARS-CoV. However, as with all PCR assays, there is
potential for both false positive and false negative results. False
positive results can occur from contamination with previously amplified
DNA during specimen processing or preparation of the amplification
reaction. Cross-contamination between patient specimens can also occur
during the course of collection, transport, storage, and processing.
Careful monitoring of all steps between specimen collection and RT-PCR
should be made to minimize the possibility of false positive results.
All positive results should be confirmed by repeat testing from a new
extraction of the original sample AND having the sample retested in a
second qualified laboratory (e.g., CDC) before reporting. False negative
results can occur from insufficient SARS-CoV RNA in the sample or
degradation of RNA during storage and transport of the specimen. Optimal
specimen collection as described above and careful handling of the
specimen can help minimize false negative results.

**Serology Testing**

Detecting SARS-CoV antibodies by EIA is a less ambiguous approach to
diagnosing SARS-CoV infection than is RT-PCR, but antibodies are often
not be detectable early in the course of illness or if the patient is
immune suppressed and unable to mount a good antibody response.
Seroconversion from negative to positive or a four-fold rise in antibody
titer from acute to convalescent serum specimens confirms recent
infection. At the present time, when prior infection is exceedingly
rare, a positive serology result is also considered indicative of acute
infection with SARS-CoV in a patient with a SARS-like illness. Although
many SARS patients develop antibodies to SARS-CoV within as few as 8 to
10 days, some patients do not test positive until more than 28 days
after the onset of illness. For patients with a negative antibody test
result with specimens collected 28 days after illness onset, an
additional serum specimen collected \>28 days after onset should be
tested. A negative antibody test after 28 days from onset of illness can
be used to rule out SARS-CoV infection.

**How CDC is Improving SARS Diagnostics**

CDC is actively working to improve existing assays and to develop new
tools for diagnosis of SARS-CoV infection. Serologic EIAs using
recombinant viral antigens to replace cell culture derived virus and to
detect SARS-CoV IgM antibodies are currently under evaluation. CDC is
also preparing new protocols to guide the type and timing of specimen
collection and processing to try to improve the ability to detect low
levels of SARS-CoV early in infection and is developing a quality
assessment program to monitor the accuracy and reliability of the tests.
By working with partners, CDC will continue to disseminate the most up
to date assay protocols and reagents to the laboratory response network
(LRN) and state and local public health laboratories.

**CDC WEB Resources**

**Interim Guidelines for Laboratory Diagnosis of SARS-CoV Infection**

<http://www.cdc.gov/ncidod/sars/labdiagnosis.htm>

**Handling & Processing Specimens Associated with SARS**

<http://www.cdc.gov/ncidod/sars/sarslabguide.htm>

**Guidelines for Collection of Specimens from Potential Cases of SARS**

<http://www.cdc.gov/ncidod/sars/specimen_collection_sars2.htm>

**Specimen Submission Form for Potential Cases of SARS**

<http://www.cdc.gov/ncidod/sars/pdf/specimensubmissionform-sars.pdf>

**Guidelines for International Specimens Associated with SARS**

<http://www.cdc.gov/ncidod/sars/intspecimens.htm>

**Packing Diagnostic Specimens for Transport**

<http://www.cdc.gov/ncidod/sars/packingspecimens-sars.htm>

**Emerging Infectious Diseases (Journal)**

<http://www.cdc.gov/ncidod/EID/sars_links.htm>