MT-PCR panel detection of canine parvovirus (CPV-2): Vaccine and wild-type CPV-2 can be difficult to differentiate in canine diagnostic fecal samples

Abstract

Canine parvovirus (CPV-2) remains an important cause of devastating enteritis in young dogs. It can be successfully prevented with live attenuated CPV-2 vaccines when given at the appropriate age and in the absence of maternal antibody interference. Rapid diagnosis of parvoviral enteritis in young dogs is essential to ensuring suitable barrier nursing protocols within veterinary hospitals. The current diagnostic trend is to use multiplexed PCR panels to detect an array of pathogens commonly responsible for diarrhea in dogs. The multiplexed PCR assays do not distinguish wild from vaccine CPV-2. They are highly sensitive and detect even a low level of virus shedding, such as those caused by the CPV-2 vaccine. The aim of this study was to identify the CPV-2 subtypes detected in diagnostic specimens and rule out occult shedding of CPV-2 vaccine strains. For a total of 21 samples that tested positive for CPV-2 in a small animal fecal pathogens diagnostic multiplexed tandem PCR (MT-PCR) panel during 2014-2016 we partially characterized the VP2 gene of CPV-2. Vaccine CPV-2 strain, wild type CPV-2a subtypes and vaccine-like CPV-2b subtypes were detected. High copy number was indicative of wild-type CPV-2a presence, but presence of vaccine-like CPV-2b had a variable copy number in fecal samples. A yardstick approach to a copy number or C-value to discriminate vaccine strain from a wild type virus of CPV-2 can be, in some cases, potentially misleading. Therefore, discriminating vaccine strain from a wild type subtype of CPV-2 remains ambitious.

Faecal PCR panel results and clinical findings in Western Australian dogs with diarrhoea

“…Since vaccine strain shedding has been associated with low copy numbers of viral DNA in faecal samples, fully quantitative fPCR could also play a role in ruling in or out vaccine interference. 1,33,34 Secondary vaccine failure, or waning of antibody-mediated protection over time, could also lead to CPV infection, and may account for some of the CPV-positive adult dogs in our study. Given the presence of diarrhoea at the time that these adult dogs tested CPV positive we do believe that these cases had true parvovirosis, however, other possible explanations include that CPV infection was present but they had another cause for their diarrhoea (i.e.…”

A history of canine parvovirus in Australia: what can we learn?

“…Also recently investigated was the ability of multiplexed PCR assays to differentiate between CPV shedding caused by wild‐type infection and vaccination 48 ; however, researchers reported difficulty confirming via PCR copy number or cycle‐threshold whether a detected CPV was of vaccine or wild‐type origin.…”

Multiplex amplification refractory mutation system PCR (ARMS-PCR) provides convenient method for differentiation of canine parvovirus vaccine and field strains

“…Vaccination with modified live attenuated vaccine is the most preferred way for controlling parvoviral infection in dogs. Fecal shedding of vaccine virus few days to couple of weeks post-vaccination is a common phenomenon [4,6,8]. There are instances of canine parvovirus outbreaks in vaccinated dogs probably due to interference of maternal antibodies in pups or lack of protective antibody titers against heterologous CPV antigenic types [4,10].…”

“…There are instances of canine parvovirus outbreaks in vaccinated dogs probably due to interference of maternal antibodies in pups or lack of protective antibody titers against heterologous CPV antigenic types [4,10]. Thus, exact determination of CPV-2 vaccine and field wild type strains is urgent requirement in CPV-2 diagnostics and for development of effective control strategies [8].…”

“…Vaccination with CPV-2 MLV causes transient viraemia and shedding of vaccine virus occurs a few days to a couple of weeks post-vaccination. The problem is complicated in situations where clinical signs appear (may be infection with virulent CPV-2 field strains before attaining protective antibody titer) after vaccination which posses diagnostic dilemma to differentiate the vaccine and the field strains [3][4][5][6]8]. Real-time PCR has also been used to estimate titer of viral load in fecal samples and correlate with vaccination or infection; as it is hypothesized that vaccine induced fecal shedding of virus is excreted in a lower titer [6].…”