Sequencing PRRS diagnostics

Genomic variation is used to understand virus transmission and to identify similarities in virus over time and space. Both virus sequencing and restriction fragment length polymorphism (RFLP) are used.
 

Virus sequencing compares amplified nucleic acids and their corresponding amino acids in a target region based on the percent homology and clustering. The OFR5 is most commonly used. For RFLP, enzymes are used to cut the nucleic acids and the patterns are compared. Virus sequencing is more precise than RFLP. Similar strains can have different RFLP patters and strains with a similar RFLP pattern can have sufficiently dissimilar nucleic acids to be placed in different phylogenic groupings. For these reasons, virus sequencing is becoming the recommended approach. Neither tool is used to monitor for the presence of PRRSV however they have been considered a gold-standard test to confirm RT-PCR specificity.

Sequence analysis of viral genomes is now the method of choice for characterisation of PRRSv isolates. Sequencing and phylogenetic analysis is performed on PCR products from test samples, with serum and infected tissues providing the most reliable data.  Typically, data on the highly variable ORF5 sequence are the most reliable for distinction of strains, and a large library of ORF5 sequences is available for comparison. Phylogenetic relationships between strains are visualised using dendograms, and these can be generated at the individual farm level to monitor local changes within the herd, such as the appearance of a new strain or the reappearance of an infection. However, non-random mutations, such as through recombination or genetic selection, can impact on dendogram patterns. Therefore caution must be exercised when interpreting data on potential new viruses within the herd.