Efficient influence of ssDNA virus PCV2 replication by CRISPR/Cas9 targeting of the viral genome.

Porcine circovirus type 2 (PCV2), a ubiquitous pathogen that primary cause of postweaning multisystemic wasting syndrome (PMWS), had caused significant morbidity and mortality in swine populations with huge economic losses in the worldwide swine industry. Currently, looking for effective antiviral drugs for PCV2 infection remains an important works. In our study, CRISPR/Cas9 system was used to further detected the key sites of PCV2 replication. We designed 8 single guide RNAs (sgRNA) by targeting essential genes across the genome of PCV2. Western-blot(WB), Cell counting kit-8 for high-throughput sgRNA screening were applied to detect PCV2 replication levels. The results showed that Oc8, O13, O134, NQT and NPS sgRNAs can edit the PCV2 genome efficiently and inhibit PCV2 replication in PK-15 cell; H3 sgRNA cannot edit the PCV2 genome successfully; NAT sgRNA can edit the PCV2 genome efficiently to improve the PCV2 replication in PK-15 cell; O26 sgRNA can edit the PCV2 genome successfully but it is not known yet of its effect on PCV2 replication, besides the Cas9 expression had no effect on cell viability. These data suggest that CRISPR/Cas9 system targeting PCV2 essential genes may serve as a novel therapeutic agent against PCV2 infection in the future.

Field evaluation of a sing-dose bivalent vaccine of porcine circovirus type 2b and Mycoplasma hyopneumoniae.

BACKGROUND: The field efficacy of a bivalent vaccine containing porcine circovirus type 2b (PCV2b) and Mycoplasma hyopneumoniae was evaluated on three pig farms. METHODS: Three pig farms were used, two of which had a history of subclinical PCV2 and clinical M. hyopneumoniae infections between 84 and 126 days of age while concurrent porcine circovirus-associated disease and clinical M. hyopneumoniae infection between 70 and 105 days of age. Each farm vaccinated pigs with a single dose of a bivalent vaccine at 10 days of age while unvaccinated pigs were administered a single dose of phosphate buffered-saline at the same age. RESULTS: Vaccination improved growth performance and reduced clinical scores significantly (p < .05) when compared with unvaccinated animals. The amount of PCV2d loads in blood and M. hyopneumoniae loads in nasal swabs of vaccinated animals were also significantly lower (p < .05) when compared with unvaccinated animals. Immunologically, vaccinated groups elicited a significantly higher (p < .05) level of protective immunity against PCV2d such as neutralizing antibodies and interferon-γ secreting cells (IFN-γ-SC), as well as protective immunity against M. hyopneumoniae such as IFN-γ-SC when compared with unvaccinated animals. Pathologically, vaccination significantly lowered (p < .05) the scores of M. hyopneumoniae-induced pneumonia and PCV2-associated lymphoid lesions when compared with unvaccinated animals. CONCLUSIONS: The evaluated bivalent vaccine provided good protection against PCV2d and M. hyopneumoniae infection under field conditions.

The epidemiology, molecular characterization and clinical pathology of circovirus infections in pigeons - current knowledge.

The first cases of circovirus infections in pigeons were documented less than 25 years ago. Since then, circovirus infections have been reported on nearly all continents. The specificity of pigeon breeding defies biosecurity principles, which could be the reason for the high prevalence of PiCV infections. PiCV infections in pigeons lead to atrophy of immune system organs and lymphocyte apoptosis. Infected birds could be more susceptible to infections of the respiratory and digestive tract. PiCV has been associated with the young pigeon disease syndrome (YPDS). PiCVs are characterized by high levels of genetic diversity due to frequent point mutations, recombination processes in the PiCV genome and positive selection. Genetic recombinations and positive selection play the key role in the evolution of PiCV. A protocol for culturing PiCV under laboratory conditions has not yet been developed, and traditional vaccines against the infection are not available. Recombinant capsid proteins for detecting anti-PiCV antibodies have been obtained, and these antigens can be used in the production of diagnostic tests and subunit vaccines against PiCV infections. However, YPDS has complex etiology, and it remains unknown whether immunization against PiCV alone will contribute to effective control of YPDS.