test

As H5N1 avian influenza continues to spread in wild birds and spills over into domestic birds and mammals of many kinds, we’re becoming more aware of the risks to domestic mammals and there are more questions about test selection and interpretation. Fortunately, testing for H5N1 influenza is relatively straightforward.

  • For cats and dogs, the recommended samples for testing are typically oropharyngeal swabs, plus or minus nasal swabs (plus or minus other tissue samples of the animal is deceased. 

What tests are currently being run by veterinary diagnostic labs?

PCR is the most accessible and useful test; diagnostic labs basically offer two types of influenza PCR: matrix / pan-influenza A PCR, and strain-specific PCRs (see below). Different commercial labs offer different tests or combinations of these tests, and the tests offered may also vary by species (i.e. what’s routinely done for dogs can be different from cats). Test offerings may also change over time as labs adapt to the ever-changing situation with flu in different populations. If influenza is a consideration in your patient, check with your lab regarding which tests they will run (particularly if you are submitting samples for a respiratory PCR panel) and how to interpret the results, including whether or not the test will detect H5N1 flu if that’s a concern.

Influenza A matrix PCR (aka pan-influenza A PCR)

This test will detect RNA from any / all influenza A viruses. A positive test confirms that flu virus is present, but not the strain (and not that the virus present is necessarily viable). Knowing the strain is important to understand how the animal might have been exposed and transmission risk. It’s a good first screening step, but if it’s positive we need more testing. If a cat was positive, it could mean it has a human seasonal flu strain (people sometimes infect their cats, and ’tis the season), or it could have H5N1 influenza, or another flu strain (e.g. a low pathogenicity flu strain which can cats sometimes get from wild birds, or potentially a swine flu virus if they have contact with pigs). Really rarely, but importantly, an animal could be infected with a combination of different flu viruses. Our concerns about and responses to these different scenarios are really different, so it’s important information to get. 

Strain-specific influenza PCR

These tests target specific influenza strains in different species, like canine H3N2, canine H3N8, avian H7N2 (found sometimes in cats) and human H1N1 (which can spill over sometimes into dogs and cats). It’s important to be aware that these are strain-specific tests, so an animal that just has H5N1 influenza will test negative on the H3N2 test, for example.

H5N1-specific influenza PCR 

As the name suggests, this test is specifically for H5N1 influenza, and we can be more confident in the result if that’s the strain for which we’re looking. The downside is that it won’t detect co-infections with multiple flu strains. The odds of a dog having H3N2 canine flu or H1N1 human flu and H5N1 avian flu at the same time are REALLY low, but that would be a REALLY concerning situation, so it would be nice to know. It’s more relevant when there’s higher non-H5N1 flu activity in the area as well, since that would mean there’s a greater chance of a hidden co-infection.

In order to provide the best possible information without over-testing every sample, labs will sometimes perform different tests in sequence: 

Run matrix PCR; if positive, then test for H5N1

This adds a step, but it’s usually a quick one, and we find out whether the animal has a flu virus and if so, whether its H5N1. The same issue with not identifying co-infections applies here, because there’s no testing for other flu strains.

Run matrix PCR, then test for non-H5N1 strains; if negative, then test for H5N1 

This works too. It adds a bit of extra time/work, and the more steps that are required, the greater the chance of a test error, but it gets to the same result pretty quickly and gives us a specific H5N1 result. The main theoretical issue is that we could still miss a coinfection (in this case because the H5N1 test isn’t run if any of the other strains are detected). 

Run matrix PCR, then test for non-H5N1 strains; if negative, then refer to another lab for H5N1 testing

This slows things down more and adds in some uncertainty as it requires sending the sample out for follow-up testing, which is another step where human error could affect things. It also misses co-infections.

Run matrix PCR, then test for non-H5N1 strains, then test for H5N1; if negative, stop and call it a generic influenza A positive

This isn’t ideal but still tells us a lot. In most cases nowadays, if the sample is matrix positive and negative for all the other main flu strains, it’s probably H5N1 flu, but pets can get spillover infection of other strains that aren’t included in strain-specific tests. If they stop here, it’s functionally okay but not ideal. I’d want to try to get followup testing of any matrix positive, type-specific negative samples (and would treat them as H5N1 positive until proven otherwise).

Any of these combinations would be okay for testing an animal in which there’s suspicion of H5N1 flu exposure / infection, but I’d rather have a quick H5N1-specific result in these cases, and I’d want to make sure that the lab will forward any positive sample for further genomic testing, so we can better understand the situation with H5N1 flu. 

Take home message

  • Talk to your lab to know what they can (and can’t) do in terms of testing for H5N1 flu, and follow up testing.
  • We can’t just stop at “flu positive.” Any such result needs to be scrutinized to make sure we know the strain (or strains) involved.