Refining and Improving BRSV Testing

Background

Bovine Respiratory Syncytial Virus (BRSV) is one of several viruses that play an important role in Bovine Respiratory Disease (BRD), and can cause big significant animal health and production losses. Current laboratory tests designed to diagnose BRSV are unable to distinguish between the disease-causing virus and the virus from a vaccine. This can lead to misdiagnoses. Genomic sequencing can distinguish between them.  

There are concerns whether the virus that the vaccine was originally based still provides protection against the strains of BRSV that are circulating in the environment. Viruses evolve over time, so if vaccines aren’t updated, they may provide less protection over time. Genomic sequencing can tell us how closely vaccine strains are related to circulating disease-causing field strains, and whether they can be expected to provide meaningful protection against disease.  

Objectives

•  Refine the current methods of diagnosing Bovine Respiratory Syncytial Virus (BRSV) with the ability to detect regional strains, differentiate strains causing disease from the vaccine strains and improve BRSV surveillance.

What they DID

This team will use existing known positive samples ranging from highly positive to borderline, from the Prairie This team used existing known positive samples ranging from highly positive to borderline, from the Prairie Diagnostic Services Inc. and other labs in western Canada and subjected them to two different methods of testing:  

• Metagenomic sequencing (nanopore technology) – sequencing all DNA and RNA from a sample, including BRSV sequences.  

• Targeted sequencing – only looking at BRSV 

The first method evaluated the limits of whole genome testing – how positive does a BRSV sample need to be to yield meaningful results. This method is less sensitive but can potentially detect all pathogens in a sample.  

The second method was less expensive and more sensitive. It doesn’t read the virus’ entire genome sequence, focusing instead on specific genes of interest. Samples that triggered positive results due to vaccination were also included to allow the test developers to differentiate between vaccinated vs. infected animals, and to identify the specific field strain responsible for the infection.  

What They Learned

When DNA sequences were compared for BRSV isolated throughout the world, Western Canadian field strains were most closely related to other North American field strains, while European field strains were more closely related to the vaccine strains. These findings provide the foundation for a database that can support future continued monitoring of BRSV in Canada. 

They successfully created sequencing methods for three key viral genes of BRSV: fusion (F), glycoprotein (G), and nucleoprotein (N). These methods were about 128 times more sensitive, 50% less expensive and more reliable than the metagenomic sequencing method. 

Comparisons of the F, G and N sequences from 43 clinical samples and two vaccine strains showed that Western Canadian field strains studied were highly similar to each other. The N genes of the field and vaccine strains were very similar, while there were slightly larger differences between the F (4.8%) and G (14%) genes of the field and vaccine strains. Some of these differences occurred in regions that may potentially be important for immune response, though evidence from other research in Saskatchewan suggest that current vaccines do provide effective protection against BRSV.  

What It Means

The good news from this study is that: 

• It seems BRSV has not (yet) mutated significantly in Canada 

• Now we have a tool to monitor the mutation of the field strains of BRSV 

• The monitoring is relatively simple and cost effective to conduct 

Because the Canadian strains are similar to each other, it means the vaccines should be similarly effective to all outbreaks without significant mutation events.  

Continuing monitoring field strains will help detect new strains entering the Canadian population, some of which may cause different and more severe clinical diseases and respond differently to vaccination.