Module 11: Rapid Point of Care (POC) Testing

Module 11.5: Heartworm Testing

Heartworm testing

Heartworm disease (HWD) is a mosquito-borne filarial disease that infects dogs and cats worldwide caused by the nematode Dirofilaria immitus. (See Agents of Disease 2 notes for more specifics on the disease and disease presentation) As a result of climate change and the increased movement of both dogs and cats from regions of the United States (and the world) that have a high prevalence of HWD to regions of the country that have traditionally had a low prevalence, HWD has become more widespread. In fact, CAPC has reported an overall 20% increase in HWD prevalence between 2013-2017 for the United States overall, but especially in parts of the country that were considered to have low prevalence.

As HWD becomes more prevalent throughout the United States and worldwide, the need for routine HWD (and other vector-borne diseases) testing will become more common in clinical practice.

Heartworm Disease
Heartworm Disease (HWD)

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Heartworm ELISA (LFA) testing

In today’s laboratory, you will be using the IDEXX SNAP 4Dx Plus results to answer the questions in your case study. However, there are several other tests that are available on the market for heartworm screening.

The following chart summarizes the 3 most common LFA tests available on the market for in-clinic testing.

Table 11.1: The 3 Most Common LFA Tests for In-Clinic Testing

HW test

Heska SOLO®


Zoetis Witness®

What sample (s) can you use to run this test (blood, serum, saliva, etc.)
Plasma, serum, anticoagulated whole blood Plasma, serum, anticoagulated whole blood Plasma, serum, anticoagulated whole blood
What does this test specifically detect?
Antigen secreted by the adult worm Antigen secreted by the adult worm Antigen secreted by the adult worm
Does this test detect any other diseases?
No Anaplasma phagocytophilium (Ab), Anaplasma platys (Ab), Ehrlichia canis (Ab), Ehrlichia ewingii (Antibody), Borellia burgdorferi (Ab), Dirofilaria immitis No

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4Dx Plus Procedure

  1. If stored in a refrigerator, allow all components to equilibrate at room temperature (18–25°C) for 30 minutes before use. Do not heat.
  2. Using the pipette provided, dispense 3 drops of the patient sample into a new sample tube.
  3. Holding the bottle vertical, add 4 drops of the conjugate to the sample tube.
  4. Cap the sample tube and mix it thoroughly by inverting it 3–5 times.
  5. Place the device on a horizontal surface. Add the entire contents of the sample tube to the sample well, being careful not to splash the contents outside of the sample well. The sample will flow across the result window, reaching the activation circle in 30–60 seconds. Some samples may remain in the sample well.
  6. When color FIRST appears in the activation circle, push the activator firmly until it is flush with the device’s body.

Note: Some samples may not flow to the activation circle within 60 seconds, and therefore, the circle may not turn color. In this case, press the activator after the sample has flowed across the result window.

  1. Read the test result at 8 minutes.

Note: The positive control may develop sooner, but results are not complete until 8 minutes.

Fun fact: The SNAP test conjugate is BLUE, the word blue has 4 letters and you also use 4 drops of the conjugate to run this test. Blood is RED and the word red has 3 letters in it therefore you only need 3 drops of blood product (serum, plasma, or whole blood) to run the test.

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Interpretation of 4Dx Plus

The SNAP 4Dx Plus detects 4 pathogens including D. immitusBorrelia burgdorferiAnaplasma sp., and Ehrlichia sp

SNAP 4Dx Plus Test with labels
SNAP 4Dx Plus Interpretation

For dogs that test positive for HWD using an LFA, prior to administering adulticidal therapy a secondary confirmatory test needs to be performed to rule out a false positive.

A few secondary tests available includes:

  • Modified Knott’s test
    • This test looks for living larvae in blood
    • Concentrates and allow speciation of microfilariae in regions of the country where other microfilariae species aside from D. immitus are present. (i.e. Acanthocheilonema reconditum
  • ELISA testing performed in a reference laboratory

Other diagnostic aids

  • Thoracic radiography
  • Echocardiogram to detect adult worms

To complete the question in your laboratory case, please refer to the American Heartworm Society’s guidelines and website.

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