OPPCONCERNED SHEEP BREEDERS SOCIETY
OPP TESTING 101
Since the OPP Concerned Sheep Breeders Society is often asked where samples should be sent for testing — and which test should be used — we have compiled the following information to assist you and your veterinarian in deciding what’s best for your flock. It is important to remember that no diagnostic test is 100% accurate in all cases. Over time, however, you should see consistency in test reports. If you experience wide, unexplainable swings in results from test to test, you may want to consider a different lab.
SPEND WISELY! Eradication of OPP is a significant investment that requires careful planning and vigorous
follow-through. When animals are tested for the first time, a partial-flock screening (see more below) can save
considerable $$$. If infection is found, explore all options and plan before you proceed. Be aware that there
are various paths to success — no one-size-fits-all — and multiple tests will be needed to confirm OPP status.
Always use an accredited lab!
We recommend that samples be sent only to a lab that is accredited by the American Association of Veterinary Laboratory Diagnosticians (AAVLD). Our members report excellent results from the University of Minnesota, Cornell University in New York, Colorado State University-Fort Collins, and Washington State University.
ELISA (enzyme-linked immunosorbent assay):
The ELISA detects viral infection by identifying antibody response. The primary advantage of this test is that it is read by a machine rather than by a human, thus eliminating the subjectivity factor. One drawback is that ELISAs can be overly sensitive, sometimes giving results that are falsely positive.
The first ELISA for OPP, using antigen produced by tissue culture, was developed at Cornell University in the mid-1980s but it was discontinued due to high cost. Subsequently, various more sophisticated ELISAs have been used in research facilities and two are now commercially available in the U.S. Most recently (fall 2013) the University of Minnesota imported Elitest® which is the only OPP/CAE ELISA to have been validated according to World Organization for Animal Health (OIE) standards. Elitest® is available to any producer at a cost of $6.25 per sample. Also available in the U.S. is a competitive ELISA (cELISA), developed for CAE/goats but also licensed for use in sheep.3
For accurate ELISAs, antigen purity and adequate lab controls are extremely important. Sample quality may also be a factor. When collecting blood for ELISA tests, every effort should be made to avoid hemolysis (handle tubes with care, never freeze whole blood, and remove clot unless serum-separator tubes are used). Submit 1 ml of serum for this test.
AGID (agar gel immunodiffusion):
Like the ELISA, the AGID detects infection by identifying antibody response. AGID has proven effective in eradicating OPP for more than 30 years and is the prescribed test for international trade. It is simple, reliable, inexpensive, and “standard” in that most labs use a USDA-licensed test kit — including OPP antigen — from the same manufacturer.
Since it can take 12 weeks or longer following infection with the OPP virus before antibodies are detectable by AGID, two or three subsequent negative AGID tests done at 6-12 month intervals will increase confidence in the negative status of a given animal. The AGID does not give false positives. And seroconversion is permanent, meaning that an AGID test-positive animal will remain positive on future tests.1,2 The cost of the AGID typically runs from $5.50 and up depending on the lab.
For the highest degree of accuracy, the AGID must be done by a properly trained, experienced technician with good eyesight. So again, always use an AAVLD-accredited lab! Submit 1 ml of serum for the AGID.
PCR (polymerase chain reaction):
The PCR has long been used for OPP testing in research facilities and first became available to producers in 1999. While no silver bullet — like other tests, the PCR is not infallible — this test detects the actual presence of the virus rather than relying on antibody response to infection. Due to its relatively high cost, the PCR is usually reserved for especially valuable animals, or those for whom other tests have been inconclusive.
While no formal validation studies have been done on the PCR for OPP testing, Colorado State University has published recommendations for its use as well as results of a test run comparing AGID and PCR.4,5 They suggest using the AGID for initial whole-flock screening, then verifying with PCR those animals already tested negative by AGID.
To our knowledge, the only lab offering this test for OPP is Colorado State University, at a charge of $37.50/sample. Draw 5-10 ml of blood into lavender-top EDTA tubes (the more blood, the better). Immediately after collecting each sample, thoroughly mix blood by gently rocking the tube back and forth several times. Ship whole blood with ice pack, but do not freeze.
1. Juste RA, Kwang J, de la Concha-Bermejillo A: Comparative evaluation of the agar gel immunodiffusion test and recombinant ELISA for the diagnosis of ovine progressive pneumonia. In Proceedings of the 99th Annual Meeting of the U.S. Animal Health Association, 1995; 536-545.
2. Wolf C: Testing and control of ovine progressive pneumonia. In Proceedings of the Minnesota Veterinary Medical Association Convention, 2000; 389-391.
3. Herrmann LM, Cheevers WP, Marshall KL, McGuire TC, Hutton MM, Lewis GS, Knowles DP: Detection of serum antibodies to ovine progressive pneumonia virus in sheep by using a caprine arthritis-encephalitis cirus competitive-inhibition enzyme-linked immunosorbent assay. Clin Diag Lab Immunol, Sept. 2003; 862-865.
4. Collins J: Eradication of OPP or CAE: Testing with AGID vs PCR. In Lab Lines, Newsletter of the Colorado State University Veterinary Diagnostic Laboratories, Volume 4, Number 1, Fall 1999.
5. Van Campen H: Diagnostic tests for caprine arthritis-encephalitis and ovine progressive pneumonia. In Lab Lines, Newsletter of the Colorado State University Veterinary Diagnostic Laboratories, Volume 5, Number 2, Spring 2001.
Recommendations for Partial-Flock Testing:
When a flock of unknown status is tested for the first time, only a portion of the animals may be sampled for the initial screening. The table below shows the number of animals 12 months of age and older that needs to be randomly sampled and tested in order to be 95% confident of detecting at least one positive animal if 5% or more of the flock is infected
NOTE: Since sampling a truly random subset can present difficulties at the management level, it is recommended to select for testing only those animals that have been in the flock for at least two years, with ample representation of all ages. This may increase the odds of detecting infection at the flock level.
<30 (test all)
Collecting Blood and Submitting Samples for OPP Testing:
Regardless of which AAVLD-accredited lab you and your veterinarian select, high quality samples and carefully completed paperwork will help to ensure fast turnaround.
For collecting blood samples, use silicone treated glass tubes with red or mottled red/grey rubber stopper caps. (Be aware that the latter, often called “tiger-tops,” are serum-separator tubes which must be centrifuged prior to submitting to the lab.) The 13 x 100 mm size (1/2” diameter, 4” long) works whether collecting with a syringe or directly into the tube using a Vacutainer needle and holder. For submitting serum, tubes can be either glass or plastic. Please do not submit slim (pencil diameter) tubes or those less than 3” in length.
A 3 ml draw is adequate and can be done using a disposable syringe or a double-ended Vacutainer needle with holder. In either case, use a new, sterile needle/syringe and specimen tube for each animal. If collecting with a syringe, blood must then be transferred immediately and carefully into a tube by inserting the needle through the rubber cap. While transferring, be aware that the tube has a vacuum and you want to avoid “splashing” which can cause red blood cells to burst. So try to direct the needle toward the inside wall of the tube, allowing blood to flow gently down the side. If red blood cells do burst, and this may be unavoidable, your serum will be pink or red. That’s OK, but straw colored serum is the goal.
No blood clots! Submit only serum separated from clotted blood (1 ml is adequate). Your veterinarian can spin samples in a centrifuge to separate the clots, which must then be removed unless samples have been collected in serum-separator tubes. Alternatively, blood can be allowed to clot at room temperature in the red-top tubes (tubes upright) and then serum poured into a fresh tube. Serum may be refrigerated or frozen. If samples have been frozen, be sure to note this on your submission form.
Using a permanent marker so the ink won’t smear when wet, number tubes 1 through XXX. This number should be dark, written with care so it’s easy to read, and placed at the very top of the label (nearest the rubber stopper/cap) while the tube is held upright. Then, turning the tube on its side (horizontal), write the date drawn, the individual animal ID, and your last name. Organize tubes in numerical order, matching the sequence noted on your submission form.
Screen-fillable submission forms can be found on the lab’s website (see below), or you can print a copy and complete it by hand. All information on the form must match the labels on the tubes, and the completed form should be placed in a Ziploc bag separate from the sample tubes. Submit under your veterinarian’s name, noting whether blood was drawn by you or by the veterinarian. If no vet, leave that section blank. Do not include payment; the lab will bill you.
Pack tubes carefully in a strong carton to avoid breakage (mailing boxes may be available at no charge; see the lab’s website), enclosing absorbent material inside a leak-proof inner wrap. Tubes may be shipped in a horizontal position but be sure to keep them in numerical order. For highest quality samples, ship with a frozen gel pack and mail early in the week to avoid weekend layover.
U.S. Priority Mail is fast and economical. Mark box with the words “Exempt Animal Specimens.” Since many states do not have an AAVLD accredited laboratory, suggested regional labs are noted below. For a complete list, see www.aavld.org
University of Minnesota
**Elitest ELISA available
— Click here to view our new 30-minute video on YouTube —
Video also available as a DVD. See ‘Membership’ page for more information.