Evaluation of virus isolation, one-step real-time reverse transcription polymerase chain reaction assay, and two rapid influenza diagnostic tests for detecting canine Influenza A virus H3N8 shedding in dogs

Abstract

Sustained transmission of canine Influenza A virus (CIV) H3N8 among U.S. dogs underscores the threat influenza continues to pose to canine health. Because rapid and accurate detection of infection is critical to the diagnosis and control of CIV, the 2 main objectives of the current study were to estimate and compare the sensitivities of CIV testing methods on canine swab samples and to evaluate the performance of Flu Detect™ (Synbiotics Corp., Kansas City, MO) for detecting CIV nasal shedding in high-risk shelter dogs. To address the first objective, nasal and pharyngeal swab samples were collected from 124 shelter and household dogs seen by Colorado State University Veterinary Teaching Hospital clinicians for canine infectious respiratory disease between April 2006 and March 2007 and tested for CIV shedding using virus isolation, the rapid influenza diagnostic test Directigen Flu A+B™ (BD Diagnostic Systems, Sparks, MD), and real-time reverse transcription polymerase chain reaction (RT-PCR). For the second objective, 1,372 dogs with unknown respiratory health status were sampled from 6 U.S. shelters from December 2009 to November 2010. Samples were tested for presence of CIV using real-time RT-PCR and Flu Detect. Using a stochastic latent class modeling approach, the median sensitivities of virus isolation, rapid influenza diagnostic test, and real-time RT-PCR were 72%, 65%, and 95%, respectively. The Flu Detect test performed poorly for detecting CIV nasal shedding compared to real-time RT-PCR. In conclusion, the real-time RT-PCR has the highest sensitivity for detecting virus nasal shedding and can be used as a rapid diagnostic test for CIV.

Epidemiology and Ecology of H3N8 Canine Influenza Viruses in US Shelter Dogs

“…Both CA and TX had no CIV‐seropositive samples and CA had only 1 dog shedding CIV upon admission into the shelter (April 2010) throughout the entire study. True prevalence estimates (Table 2) corroborate other reports suggesting CIV is endemic in CO and NY4, 5, 12, 13 with prevalence estimates of 2.7 and 2.5% for these shelters, respectively. The CIV‐positive nasal swabs collected from SC in the first 2 months of the study may explain the higher CIV‐shedding prevalence estimates for the SC shelter (1.9%) compared to the FL shelter (0.7%).…”

“…Water and viral transport medium were used as negative controls, whereas the positive control consisted of 10 TCID50 of CIV isolate A/canine/CO/224986/2006 (H3N8). For consistency, the epMotion Ep5070p4 automated system was used to load 96 well plates before amplification and detection by Mastercycler Realplex4 using previously described conditions 5…”

“…Assumptions for this method include constant sensitivity and specificity of the diagnostic test used and that only 1 population was sampled. For prior distributions, the rRT‐PCR mode was set at 0.96 for sensitivity and 0.74 for specificity at a 0.95 probability interval, based on previous studies 5, 20. Because each shelter is located in a distinct geographic region constituting a unique population, true prevalence was determined separately for each shelter.…”

“…In a NY and FL humane shelter study conducted shortly after CIV emergence, up to 97% of dogs were seropositive for CIV 1. Similarly, dogs from humane shelters in Colorado (CO) were infected with CIV in 5 of 5 humane shelters reporting CIRD outbreaks in 2006 and 2007 5. More recently, in Pennsylvania, 42% of dogs with and without clinical signs of CIRD were CIV seropositive 6…”

“…Studies conducted in nonshelter dog populations find very little CIV seropositivity in household, racing sled, and flyball tournament dogs,3, 10, 11 suggesting CIV infection to be relatively lower in dogs from the nonshelter community. In contrast, CIV is routinely detected in humane shelters throughout the United States, especially in CO and NY, where surveillance studies have been recently performed 4, 5, 12, 13…”