An ELISA to Detect Antibodies to Bovine Alphaherpesviruses 1 and 5 and Bubaline Alphaherpesvirus 1 in Cattle Sera

Simple Summary Bovine alphaherpesviruses type 1 (bovine herpesvirus type 1 subtype 1.1, 1.2a, and 1.2b), type 5 (subtypes 5a, 5b, and 5c), and bubaline herpesvirus type 1 (BuHV-1) induce highly, though not fully, cross-reactive antibody responses, but no antibody assay has, to date, been evaluated against all recognized types/subtypes of these viruses. Six hundred bovine field serum samples were screened in serum neutralization tests (SN) performed against the seven virus types/subtypes and tested in seven ELISAs prepared with each of the virus types/subtypes. A combined, multiple antigens ELISA (mAgELISA) was prepared by mixing five viral antigens. In comparison to SN, the mAgELISA sensitivity was 96.5% with 96.1% specificity (kappa = 0.93; PPV = 95.0%; NPV = 97.3%). The findings reveal that the mAgELISA developed here is suitable for the detection of antibodies to BoAHV-1, BoAHV-5, and BuHV-1, with sensitivity and specificity comparable to SN, performed with all recognized types of BoAHV-1, BoAHV-5, and BuAHV-1 as challenge viruses. Bovine alphaherpesvirus 1 (subtypes 1.1, 1.2a, and 1.2b), type 5 (subtypes 5a, 5b, and 5c), and bubaline herpesvirus 1 (BuHV-1) induce highly, though not fully cross-reactive serological responses. Most types and subtypes of these viruses circulate particularly in countries of the southern hemisphere, notably Brazil and Argentina. Therefore, the detection of infected animals is important in defining prevention and control strategies, particularly when flocks are destined for international trade. Identification of infected herds is most often achieved by assays that detect antibodies, such as enzyme immunoassays (ELISAs). However, to date, no ELISA has been evaluated in its capacity to detect antibodies to these alphaherpesviruses. Here, an ELISA was developed to detect antibodies to all currently recognized BoAHV-1, BoAHV-5, and BuAHV-1 types/subtypes, and its sensitivity and specificity were determined. Six hundred bovine sera were screened in serum neutralization tests (SN) against the seven viruses. ELISAs prepared with each of the viruses were compared to SN. Subsequently, a combined assay with multiple antigens LISA was prepared by mixing five viral antigens, chosen for their highest sensitivity in the preparative assays. In comparison to SN, the mAgELISA sensitivity was 96.5% with 96.1% specificity (kappa = 0.93; PPV = 95.0%; NPV = 97.3%). The findings reveal that the mAgELISA developed here is highly suitable for the detection of antibodies, comparable in sensitivity and specificity to that of SN when performed with all known types and subtypes of bovine and bubaline alphaherpesviruses.

Automated summary

A highly sensitive and specific multi-antigen ELISA method has been developed for detecting antibodies to Bovine alphaherpesviruses type 1, type 5, and bubaline herpesvirus type 1. The sensitivity and specificity of this method are comparable to serum neutralization tests.