Comparative analysis of routes of immunization of a live porcine reproductive and respiratory syndrome virus (PRRSV) vaccine in a heterologous virus challenge study.

Porcine reproductive and respiratory syndrome (PRRS) is caused by PRRS virus (PRRSV), which infects primarily the respiratory tract of pigs. Thus intranasal (IN) delivery of a potent vaccine-adjuvant formulation is promising. In this study, PRRS-MLV (VR2332) was coadministered ± an adjuvant Mycobacterium vaccae whole cell lysate or CpG ODN through intramuscular (IM) or IN route as a mist, and challenged with a heterologous PRRSV 1-4-4 IN at 42 days post-vaccination (dpv). At 14 and 26 dpv, vaccine viral RNA copies were one log greater in the plasma of PRRS-MLV IM compared to IN vaccinated pigs, and the infectious replicating vaccine virus was detected only in the IM group. In PRRS-MLV ± adjuvant IM vaccinated pigs, reduced viral RNA load and absence of the replicating challenged virus was observed at 7, 10 and 14 days post-challenge (dpc). At 14 dpc, in BAL fluid ≥ 5 log viral RNA copies were detected in all the pig groups, but the replicating challenged virus was undetectable only in IM groups. Immunologically, virus neutralizing antibody titers in the plasma of IM (but not IN) vaccine groups was ≥ 8 against the vaccine and challenged viruses. At 26 dpv, PRRS-MLV IM (without adjuvant) received pigs had significantly increased population of CD4 and CD8 T cells in PBMC. At 14 dpc, relatively increased population of IFN-γ(+) total lymphocytes, NK, CD4, CD8 and γδ T cells were observed in the MLV-IM group. In conclusion, PRRS-MLV IM vaccination induced the virus specific T cell response in pigs, but still it is required to improve its cross-protective efficacy.

Pretreatment of epithelial cells with live Streptococcus pneumoniae has no detectable effect on influenza A virus replication in vitro.

Influenza A virus (IAV) and Streptococcus pneumoniae (pneumococcus) are two major upper respiratory tract pathogens responsible for exacerbated disease in coinfected individuals. Despite several studies showing increased susceptibility to secondary bacterial infections following IAV infection, information on the direct effect of S. pneumoniae on IAV in vitro is unknown. This is an important area of investigation as S. pneumoniae is a common commensal of the human upper respiratory tract, present as an important coinfecting pathogen with IAV infection. A recent study showed that S. pneumoniae enhances human metapneumovirus infection in polarized bronchial epithelial cells in vitro. The aim of the current study was to determine whether treatment of epithelial cells with S. pneumoniae affects IAV replication using a standard immunofluorescence assay (IFA). For this study we used four IAV permissive epithelial cell lines including two human-derived cell lines, 12 pneumococcal strains including recent human clinical isolates which represent different genetic backgrounds and serotypes, and six IAV strains of varying genetic nature and pathogenic potential including the pandemic 2009 H1N1 virus. Our results suggested that pretreatment of MDCK cells with 7.5×10(6) colony-forming units (CFUs) of live S. pneumoniae resulted in gradual cell-death in a time-dependent manner (0.5 to 4 hr). But, pretreatment of cell lines with 7.5×10(5) and lower CFUs of S. pneumoniae had no detectable effect on either the morphology of cells or on the IAV replication. However, unlike in epithelial cell lines, due to influence of secreted host factors the effect of pneumococci on IAV replication may be different during coinfections in vivo in the human upper respiratory tract, and in vitro with primary human polarized bronchial epithelial cells.

Development and validation of an assay to detect porcine reproductive and respiratory syndrome virus-specific neutralizing antibody titers in pig oral fluid samples.

Porcine reproductive and respiratory syndrome virus (PRRSV)-specific neutralizing antibodies (NA) are important for clearing the virus. Pen-based pig oral fluid samples for disease surveillance are gaining in importance due to the ease of collection and low cost. The aim of this study was to develop a PRRSV-specific NA assay to determine NA titers in pig oral fluid samples. At first, we standardized the PRRSV NA assay using pen-based pig oral fluid samples collected over a period of 3 months from a herd of swine that received a PRRSV modified live vaccine (PRRS-MLV), and we also used oral fluid and serum samples collected from individual boars that were vaccinated with PRRS-MLV or infected with a virulent PRRSV strain. Our results suggest that a PRRSV NA titer of >8 in oral fluid samples is virus specific and can be detected beginning at 28 days after vaccination or infection. To validate the assay, we used 104 pen-based pig oral fluid and five representative serum samples from each pen of unknown history, as well as 100 serum samples from repeatedly vaccinated sows and oral fluid samples of their respective litters belonging to four different swine-breeding farms. Our results demonstrated that PRRSV NA titers in oral fluid samples are correlated with serum sample titers, and maternally derived PRRSV-specific NA titers could be detected in litters at the time of weaning. In conclusion, we have standardized and validated the pig oral fluid-based PRRSV NA assay, which has 94.3% specificity and 90.5% repeatability. The assay can be used to monitor herd immunity against PRRSV in vaccinated and infected herds of swine.