Today's Veterinary Practice

JUL-AUG 2017

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28 AHS HEARTWORM HOTLINE PEER REVIEWED excess, disrupting the normal clearance mechanisms and promoting continued immune complex creation. Presence of Microfilariae The presence of detectable circulating microfilariae in canine blood samples also appears to influence the interference of immune complexes with antigen detection. Circulating microfilariae contribute to the chronic inflammatory response of heartworm disease and are therefore theorized to contribute to continued antibody production, allowing continued immune complex formation. One report of 26 shelter dogs with circulating microfilariae demonstrated 32 times greater odds of converting to a positive antigen test result after HPT compared with dogs that tested antigen negative both before and after HPT. 2 Other reports have also identified circulating microfilariae in dogs whose test results converted from negative to positive after HPT. 1,3,4 Interestingly, one of these reports included microfilariae of D immitis, Dirofilaria repens, and Acanthocheilonema reconditum, 1 which suggests the potential for decreased specificity of antigen tests after HPT. CAN I PERFORM HEAT PRETREATMENT IN MY CLINIC? A variety of veterinary diagnostic laboratories offer HPT panels upon request. In general, costs are minimal (<$30) and results are available within 1 to 3 days. Perhaps the biggest benefit of using diagnostic laboratories for HPT is the consistency and reliability of sample handling and testing techniques. Testing methods are typically validated and concomitantly run with positive controls to ensure accuracy of results. Diagnostic laboratory services are preferred, but when these are not available or feasible, a simplified HPT protocol has been successfully used ( Box 1 , Figure 2 ). In a study of shelter dogs, 616 samples underwent the simplified HPT protocol. 2 Of these, 13 samples could not undergo repeat testing after heating. Five "untestable" samples were presumed to be directly related to inconsistencies in the application of heat, resulting in solidification of the serum sample. The remaining 8 samples were untestable for a variety of reasons presumably unrelated to the heating protocol (eg, insufficient serum, invalid controls). WHEN SHOULD I CONSIDER HEAT PRETREATMENT? Heat pretreatment is likely not indicated in most heartworm screening test scenarios. It should be considered when screening test results conflict (eg, antigen negative and microfilariae positive), when patients are receiving an alternative treatment protocol (eg, macrocyclic lactone and/or doxycycline), when the patient has known chronic inflammatory diseases (eg, pyoderma, otitis, endoparasitism), or when dogs test negative but originate from a known heartworm- endemic region and have a history of lapsed or no preventive administration. An NAD result on a pretreated sample by a diagnostic laboratory can also rule out suspicion of heartworm infection. BOX 1. Simplified Heat Pretreatment Protocol 1. Dilute serum sample with approximately equal volume of 0.9% NaCl. 2. Place ~250 mL tap water into 500-mL glass beaker. 3. Microwave beaker to the point of boiling (approximately 2 min in 1000 W). 4. Remove heated water from microwave and place tube with diluted sample in the heated water for 10 minutes. (Note: Place serum in a glass collection tube and remove the rubber stopper before heating. Some warping of tube is expected.) 5. Repeat antigen test. FIGURE 2. Diluted serum samples undergo a simplified heat pretreatment protocol. Note the use of glass tubes and the absence of rubber stoppers.

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