Sequivity(TM) RNA Particle Technology from Merck Animal Health

Whole Genome Sequencing Provides Answer in PRRSV Investigation

Porcine reproductive and respiratory syndrome continues to challenge the swine industry in many ways. As an RNA virus, it is well-documented that this virus has the ability to change constantly. As these changes accumulate through time, viral diversity increases which in turn becomes a challenge for the pig as the immune system does not recognize the virus and thus, previously generated immunity will only protect partially. [Source: National Hog Farmer, 11 Feb 2020, by Sunil Mor and Albert Rovira, University of Minnesota]

PRRS virus changes through either mutation (e.g. minor changes in its genome) or through recombination which occurs when two different viral strains are coinfecting the same pig, they exchange fragments of their genome and a whole new virus is then generated.

Currently, producers and practitioners work intensely to protect herds from new introduction of viruses as the cost of an outbreak continues to be significant due to throughput disruption, morbidity and interventions aimed at controlling the outbreak. Within the prevention measures adopted, biosecurity programs play an important role as they are a first line of defense in avoiding transmission.

However, in certain regions of the country the establishment of herd immunity as a mitigating factor also plays a role. Immunity can be established after an outbreak of PRRS virus, or through exposure to the virus by the use of live and killed vaccines. Presently, PRRSV vaccines are being used in different ways and on different animals (e.g. sows, gilts and growing pigs). In certain occasions, both the wildtype virus and the vaccine strain virus can be present in one pig which may increase the chances of new strain generation.

Recently, we developed the capability of sequencing the whole virus as opposed to small portions as it has been done for the last couple of decades. This breakthrough has allowed us to open a whole new chapter in describing the diversity of the virus and at the same time it has enabled practitioners to better understand field scenarios, such as outbreak investigations as it will be summarized in this article.

The case described here summarizes the recombination of two PRRSV vaccine strains. This case occurred in a PRRSV seropositive sow farm that had been weaning negative piglets. The herd had been meeting production targets until an increase in abortions was observed. The veterinarian rapidly collected serum from 10 sows that had aborted and sent it to the University of Minnesota Veterinary Diagnostic Laboratory for PRRSV diagnostics.

While all samples yielded a polymerase chain reaction-positive result, seven out of the 10 had a cycle threshold value below 30. A portion of the virus was sequenced from seven samples using the popular approach (e.g. Open reading frame -- ORF5 sequence) together with virus isolation in three samples. All of the ORF5 sequences were 98% similar to one of the modified-live PRRSV commercial vaccines. However, the farm had not used this modified-live vaccine (Vaccine A) in this herd for two years which generated many questions as to why were they finding such a strain. It was unclear whether this PRRSV strain was a close relative to Vaccine A, and if so, why was it associated with clinical signs.

The diagnostic laboratory team proceeded to try to solve these questions by sequencing the whole genome through next generation sequencing. The results from this investigation clearly demonstrated a recombination event between two vaccines. As with ORF5, other genes such as ORF4, ORF5a, ORF6 and ORF7 were found to be similar to Vaccine A. However, ORF1b, ORF2, ORF2b and ORF3 genes were similar to another vaccine (Vaccine B), the one that the farm was currently using. A recombination analysis comparing this PRRSV genome with that of Vaccine A and Vaccine B showed that there was not just one but multiple recombination events that had accumulated in this PRRSV strain.

Additionally, a deletion in ORF1b gene was detected. Hence, whole genome sequence analysis clearly showed that this was not Vaccine A, but a new PRRSV strain derived from the mutation (deletion) and recombination of two commercially available vaccines. This new strain may be more virulent than the original vaccine strains, as it was isolated from sows that had abortions.

Recombination of RNA viruses, especially PRRSV is not a new event as it has been previously reported on different occasions. The possibility of viral recombination should be considered with the use of a live vaccine. In this case, whole genome sequencing had a deeper discriminatory power and provided more information when compared to classical ORF5 Sanger sequencing. The use of this technology and the higher level of discernment it provides can aid in the understanding of PRRSV dynamics in pig populations.