Further validation of 2 nonstructural protein-specific antibody tests for diagnosis and surveillance of foot-and-mouth disease in the United States.

Foot-and-mouth disease (FMD) is a highly contagious disease of cloven-hoofed animals. FMD poses an economic threat to the livestock industry in the United States. Due to the potential use of vaccines composed of partially purified structural proteins of the FMD virus (FMDV), it is important to test samples from infected and vaccinated animals with a competitive ELISA that detects antibodies against nonstructural proteins (NSPs) of FMDV. Our study extends the diagnostic validation of the Prionics ELISA (Thermo Fisher) and the VMRD ELISA. We used diverse serum sample sets from bovine, porcine, and other cloven-hoofed animals to evaluate the analytical specificity and sensitivity, diagnostic specificity and sensitivity, and differentiation of infected from vaccinated animals (DIVA) per validation guidelines outlined by the World Organisation for Animal Health (WOAH). The 2 tests were analytically 100% accurate. The VMRD test was diagnostically more sensitive than Prionics, but Prionics was diagnostically more specific than the VMRD test. Both tests could tell if animals were infected or vaccinated. Considering these data, both VMRD and Prionics ELISAs can be used for serodetection of FMDV antibodies at the Foreign Animal Disease Diagnostic Laboratory and within the National Animal Health Laboratory Network laboratories.

Evaluation of an in-house indirect immunoperoxidase test for detection of antibodies against African swine fever virus.

African swine fever (ASF) is a high-consequence transboundary animal disease caused by African swine fever virus (ASFV). Given that vaccines are not widely available, ASFV detection, including by molecular and serologic assays, is paramount to efficacious control and mitigation of ASF. ASFV-specific antibodies can be detected as early as 7-10 d postinfection in infected animals and may persist for several months or longer. Accurate detection of ASFV-specific antibody is critical for the identification of chronically infected, subclinically infected, or recovered animals. ELISAs are commonly used for the rapid screening of large numbers of animals for ASFV antibodies. The World Organisation for Animal Health recommends that ELISA-positive results should be confirmed with a second serologic method, such as an indirect immunofluorescent assay, indirect immunoperoxidase test (IPT), or immunoblot test. Commercial kits are not available for those tests. We developed and validated an in-house IPT by using a currently circulating genotype II ASFV strain as antigen. The sensitivity and specificity of the in-house IPT are comparable to the reference IPT developed by an international ASFV reference laboratory and superior to a commercial blocking ELISA.

Evaluation of oral fluid as an aggregate sample for early detection of African swine fever virus using four independent pen-based experimental studies.

The sustained spread of African swine fever (ASF) virus throughout much of the world has made ASF a global animal health priority, with an increased emphasis on enhancing preparedness to prevent, detect and respond to a potential outbreak of ASF virus (ASFV). In the event of ASFV entry to the North American swine population, enhanced surveillance and diagnostic testing strategies will be critical to facilitate progressive response and eradication of the disease. Compared to individual animal sampling, pen-based oral fluid collection for active surveillance is a non-invasive alternative that is less resource and time-intensive. To evaluate the feasibility of using rope-based oral fluid for early detection of ASFV, four independent animal experiments were conducted in weaned pigs housed in numbers that mimic the industry settings, utilising either highly virulent ASFV Georgia 2007/1 strain or moderately virulent ASFV Malta'78 strain. Pen-based oral fluid and individual oropharyngeal swabs were collected daily and blood samples from each animal were collected every other day. All samples were subsequently tested for ASFV by real-time PCR. ASFV genome was detected in individual blood samples as early as one day post-infection and detected in oral fluids at low-to-moderate levels as early as 3-5 days post-infection in all four independent experiments. These results suggest that pen-based oral fluid samples may be used to supplement the use of traditional samples for rapid detection of ASFV during ASF surveillance.