Avoid turmoil when testing for heartworm disease


Heartworm disease is accompanied by few, if any, clinical signs, so proper diagnostic testing is a must. Here's the 411 for veterinary professionals on which tests to use (and why), including the best tests for cats.

Cut down on your and veterinary clients' confusion with the right tests at the right time to diagnose heartworm disease. (PixieMe/stock.adobe.com)

Most veterinary professionals know that heartworm disease is serious and progressive, and that early diagnosis improves prognosis significantly. At Fetch dvm360 conference in Baltimore, parasitologist Brian Herrin, DVM, PhD, outlined current recommendations for heartworm diagnosis, including how to interpret false-positive and false-negative results.

Dr. Herrin, an assistant professor at Kansas State University College of Veterinary Medicine, reminded the audience of the American Heartworm Society's recommendations: yearly heartworm disease screening for dogs over 7 months of age, using both the antigen and microfilaria tests.

The antigen test

Antigen testing screens for a uterine antigen produced by adult female Dirofilaria immitis. Based on the D. immitis life cycle, female worms begin producing this antigen about six or seven months after infection occurs. Antigen tests can also detect occult infections, or those in which adult worms are present but microfilariae are not. All available in-clinic antigen tests have similar sensitivities and can detect infections reliably with three or more adult female D. immitis.

Reference laboratory well-type tests have higher sensitivity and specificity values compared with in-clinic tests, and they can detect infections with a single adult female worm. A weakly positive in-clinic test result is likely positive, but the result can be confirmed by repeating the test or ordering a reference lab test. If results are discordant between in-clinic and laboratory tests, Dr. Herrin advises trusting the reference lab result.

Heartworm diagnosis should always be confirmed before treatment is administered, using a different manufacturer's antigen test, a microfilaria test or a reference lab test.

False-positive antigen test results, although rare, may occur due to cross-reactivity from other parasites, such as Spirocerca lupi. Heartworm diagnosis should always be confirmed before treatment is administered, using a different manufacturer's antigen test, a microfilaria test or a reference lab test.

False-negative results are also possible, especially in cases of early (i.e. prepatent) infections or small worm burdens. Also, doxycycline and/or preventive treatment of heartworm disease can cause chemical interference with antigen tests.

Administering preventive to a heartworm-positive dog may mask a positive antigen test in certain patients, particularly those with a highly active immune response after treatment. Once the circulating antigen binds to host antibody, it cannot attach to the antibody substrate of the diagnostic test. However, heat or acid treatment (offered by most diagnostic labs) can be performed to disrupt the immune complexes and free up antigen for detection.1 One study using Oklahoma shelter dogs found that all negative antigen test results from microfilaria-positive dogs reverted to positive after heat treatment.2 The rate of reversion was generally much lower for samples from client-owned dogs, because pets are more likely than are shelter dogs to receive regular heartworm prevention. At this time, there are insufficient data to know how long chemical interference lasts after administration of doxycycline or preventive.

To avoid confusion, heartworm diagnosis should always be confirmed immediately with a follow-up test, before doxycycline and/or preventive treatment is administered. Dr. Herrin recommended heat treatment for any samples from patients that test antigen-negative but have clinical signs consistent with heartworm disease. This step can also help confirm weakly positive tests or resolve discordant test results.

The microfilaria test

The American Heartworm Society recommends using either filtration or the modified Knott method for visualizing microfilaria. According to Dr. Herrin, the microfilaria test can be both an excellent indicator of heartworm infection and a useful visual tool to educate clients. The test is particularly valuable for dogs with an unknown preventive history.

Antigen-positive dogs may test negative for microfilaria, particularly if they receive heartworm preventive sporadically or have a small worm burden. Therefore, Dr. Herrin asserted that a negative microfilaria test result “doesn't mean anything” on its own and should always be interpreted alongside an antigen test. It's also important to remember that microfilariae of other filiarial nematodes, particularly Acanthocheilonema reconditum, can resemble D. immitis and result in a false-positive microfilaria test result.

Radiography and echocardiography

Neither thoracic radiography or echocardiography can be used alone to diagnose heartworm disease, but both tools can help with clinical staging and determining whether a patient is stable enough for treatment. In some patients, particularly small dogs, adult worms may also be visible on echocardiography.

Diagnosing feline heartworm disease

For cats, a positive antibody test indicates only that heartworm-specific antibody (not current infection) is present and that further diagnostics should be pursued. A negative test lowers suspicion of disease, because antibody tests can generally detect heartworm infection as early as two months post infection.

Unlike in dogs, the antigen test is not an effective screening tool for cats, Dr. Herrin warned. Although a positive antigen test result confirms heartworm infection, it's important to remember that the cat is a suboptimal host for D. immitis; therefore, infections are often nonpatent (i.e. nonmicrofilaremic) and involve small worm burdens. However, because 50% to 70% of feline infections include at least one female worm, a negative antigen test has some utility for lowering suspicion of infection.3,4 Just like with canine samples, heat treatment may be necessary to disrupt immune complexes.


1. Little SE, Raymond MR, Thomas JE, et al. Heat treatment prior to testing allows detection of antigen of Dirofilaria immitis in feline serum. Parasit Vectors 2014;7:1.

2. Little S, Saleh M, Wohltjen M, Nagamori Y. Prime detection of Dirofilaria immitis: understanding the influence of blocked antigen on heartworm test performance. Parasit Vectors 2018;11:186.

3. Snyder PS, Levy JK, Salute ME, et al. Performance of serologic tests used to detect heartworm infection in cats. J Am Vet Med Assoc 2000;216:693-700.

4. Berdoulay P, Levy JK, Snyder PS, et al. Comparison of serological tests for the detection of natural heartworm infection in cats. J Am Anim Hosp Assoc 2004;40:376-384.

Dr. Stilwell provides freelance medical writing and aquatic veterinary consulting services through her business, Seastar Communications and Consulting. In addition to her DVM obtained from Auburn University, she holds a MS in fisheries and aquatic sciences and a PhD in veterinary medical sciences from the University of Florida.

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