Group A rotaviruses (GARs) are one of the most common causes of diarrhea in suckling pigs. Although a number of G and P genotypes have been identified in porcine GARs, few attempts have been made to study the molecular epidemiology of these viruses associated with diarrhea outbreaks within a farm over an extended period of time. Here, we investigated the molecular characteristics of GARs that caused four outbreaks of diarrhea among suckling pigs in a farrow-to-finish farm over the course of a year. G and P genotyping of GARs detected at each outbreak demonstrated genetic diversity in this farm as follows: G9P[23] was detected at the first outbreak, G9P[13]/[22] and G9P[23] at the second, G3P[7] at the third, and G9P[23], G5P[13]/[22], and P[7] combined with an untypeable G genotype at the fourth. Sequence analysis of the detected GARs revealed that such genetic diversity could have resulted not only from the introduction of new GAR strains, but also from gene reassortment between GAR strains within the farm. Further, the GAR strain carrying the untypeable G genotype was shown to be a novel porcine GAR bearing a new G26 genotype, as confirmed by the Rotavirus Classification Working Group.

A total of 2,703 pig sera from 171 farms in six regions in Japan were screened for virus-neutralizing (VN) antibody against transmissible gastroenteritis virus (TGEV). Although none of the farms had clinical signs of transmissible gastroenteritis (TGE) or vaccination against TGEV, VN antibody was detected in 14.4% of sera at 30 farms (17.5%) across all six regions. On testing of 263 VN antibody-positive sera from 27 farms with a commercial blocking ELISA to distinguish TGEV and porcine respiratory coronavirus (PRCV) antibodies, 78.3% were positive for PRCV antibody only, while 12.5% were positive for TGEV antibody only or both TGEV and PRCV antibodies. Seven of the eight TGEV antibody-positive farms were also positive for PRCV antibody. Five months after the antibody examination, a TGE outbreak occurred at one of these seven farms. These results suggest that most of the detected VN antibodies were to PRCV, and that TGEV infection might be present at some PRCV-positive farms in Japan.

Macrophages are a heterogeneous population of cells that are present in all vertebrate tissues. They play a key role in the innate immune system, and thus, in vitro cultures of macrophages provide a valuable model for exploring their tissue-specific functions and interactions with pathogens. Porcine macrophage cultures are often used for the identification and characterization of porcine viral pathogens. Recently, we have developed a simple and efficient method for isolating primary macrophages from the kidneys and livers of swine. Here, we applied this protocol to fetal porcine intestinal tissues and demonstrated that porcine intestinal macrophages (PIM) can be isolated from mixed primary cultures of porcine small intestine-derived cells. Since the proliferative capacity of primary PIM is limited, we attempted to immortalize them by transferring the SV40 large T antigen and porcine telomerase reverse transcriptase genes using lentiviral vectors. Consequently, immortalized PIM (IPIM) were successfully generated and confirmed to retain various features of primary PIM. We further revealed that IPIM are susceptible to infection by the African swine fever virus and the porcine reproductive and respiratory syndrome virus and support their replication. These findings suggest that the IPIM cell line is a useful tool for developing in vitro models that mimic the intestinal mucosal microenvironments of swine, and for studying the interactions between porcine pathogens and host immune cells.

Peptidoglycan recognition proteins (PGLYRPs) are a family of pattern recognition receptors (PRRs) that are able to induce innate immune responses through their binding to peptidoglycan (PGN), lipopolysaccharide, or lipoteichoic acid, or by interacting with other PRR-ligands. Recently, progress has been made in understanding the immunobiology of PGLYRPs in human and mice, however, their functions in livestock animals have been less explored. In this study, we characterized the expression patterns of PGLYRPs in porcine intestinal epithelial (PIE) cells and antigen-presenting cells (APCs) and their modulation by the interactions of host cells with PRR-ligands and non-viable immunomodulatory probiotics referred to as paraimmunobiotics. We demonstrated that PGLYRP-1, -2, -3, and -4 are expressed in PIE cells and APCs from Peyer’s patches, being PGLYPR-3 and -4 levels higher than PGLYRP-1 and -2. We also showed that PGLYRPs expression in APCs and PIE cells can be modulated by different PRR agonists. By using knockdown PIE cells for TLR2, TLR4, NOD1, and NOD2, or the four PGLYRPs, we demonstrated that PGLYRPs expressions would be required for activation and functioning of TLR2, TLR4, NOD1, and NOD2 in porcine epitheliocytes, but PGLYRPs activation would be independent of those PRR expressions. Importantly, we reported for the first time that PGLYRPs expression can be differentially modulated by paraimmunobiotic bifidobacteria in a strain-dependent manner. These results provide evidence for the use of paraimmunobiotic bifidobacteria as an alternative for the improvement of resistance to intestinal infections or as therapeutic tools for the reduction of the severity of inflammatory damage in diseases in which a role of PGLYRPs-microbe interaction has been demonstrated.

Human rotavirus (HRV) infection is a major cause of viral gastroenteritis in young children worldwide. Current oral vaccines perform poorly in developing countries where efficacious vaccines are needed the most. Therefore, an alternative affordable strategy to enhance efficacy of the current RV vaccines is necessary. This study evaluated the effects of colonization of neonatal gnotobiotic (Gn) pigs with Escherichia coli Nissle (EcN) 1917 and Lacticaseibacillus rhamnosus GG (LGG) probiotics on immunogenicity and protective efficacy of oral attenuated (Att) HRV vaccine. EcN-colonized pigs had reduced virulent HRV (VirHRV) shedding and decreased diarrhea severity compared with the LGG-colonized group. They also had enhanced HRV-specific IgA antibody titers in serum and antibody secreting cell numbers in tissues pre/post VirHRV challenge, HRV-specific IgA antibody titers in intestinal contents, and B-cell subpopulations in tissues post VirHRV challenge. EcN colonization also enhanced T-cell immune response, promoted dendritic cells and NK cell function, reduced production of proinflammatory cytokines/Toll like receptor (TLR), and increased production of immunoregulatory cytokines/TLR expression in various tissues pre/post VirHRV challenge. Thus, EcN probiotic adjuvant with AttHRV vaccine enhances the immunogenicity and protective efficacy of AttHRV to a greater extent than LGG and it can be used as a safe and economical oral vaccine adjuvant.