Experimental veterinary SARS-CoV-2 vaccine cross neutralization of the Delta (B.1.617.2) variant virus in cats.

SARS-CoV-2 has exhibited varying pathogenesis in a variety of Mammalia family's including Canidae, Mustelidae, Hominidae, Cervidae, Hyaenidae, and Felidae. Novel SARS-CoV-2 variants characterized by spike protein mutations have recently resulted in clinical and epidemiological concerns, as they potentially have increased infectious rates, increased transmission, or reduced neutralization by antibodies produced via vaccination. Many variants have been identified at this time, but the variant of continuing concern has been the Delta variant (B.1.617.2), due to its increased transmissibility and infectious rate. Felines vaccinated using an experimental SARS-CoV-2 spike protein-based veterinary vaccine mounted a robust immune response to the SARS-CoV-2 spike protein. Using a reporter virus particle system and feline serum, we have verified that vaccinated felines produce antibodies that neutralize the SARS-CoV-2 Wuhan strain and variant B.1.617.2 at comparable levels.

Temporal patterns of colonization and infection with Mycoplasma hyorhinis in two swine production systems in the USA.

Control of Mycoplasma hyorhinis (M. hyorhinis) associated disease is currently hindered by limited knowledge of the epidemiology and ecology of this organism. A prospective longitudinal investigation was conducted to determine the dynamics of M. hyorhinis colonization in two swine production systems. In each system (A, B), 51 young sows (parities 1, 2) and 56 older sows (>parity 2) were selected at farrowing and tested by qPCR of nasal swabs and for antibodies by serum ELISA. From each sow, a piglet was randomly selected, and nasal and serum samples were collected at birth, weaning, and 10 days post-weaning. Two further samplings were performed in the nursery and finishing stages during the high-risk periods for M. hyorhinis-associated disease, and 12 pigs were euthanized and necropsied at these later sampling events. The prevalence of M. hyorhinis colonization in sows was low (<5%). No associations were found between sow parity or sow serum titer and piglet nasal colonization at either birth or weaning. In contrast to the low prevalence (0.95-2.70%) observed in piglets pre-weaning, most pigs became colonized during the first four weeks after weaning and remained positive throughout the nursery and finishing stages. The detection of M. hyorhinis in oral fluids followed similar patterns as those observed using nasal swabs. ELISA results showed decreased detection of maternal antibodies at around 3 weeks of age and a subsequent increase after natural exposure. The role of M. hyorhinis in polyserositis and arthritis was demonstrated in these two herds. Establishing the temporal dynamics of exposure and infection with M. hyorhinis in pigs will enable more strategic implementation of intervention strategies in affected herds.

Two clinical isolates of Mycoplasma hyosynoviae showed differing pattern of lameness and pathogen detection in experimentally challenged pigs.

Mycoplasma (M.) hyosynoviae is known to colonize and cause disease in growing-finishing pigs. In this study, two clinical isolates of M. hyosynoviae were compared by inoculating cesarean-derived colostrum-deprived and specific-pathogen-free growing pigs. After intranasal or intravenous inoculation, the proportion and distribution pattern of clinical cases was compared in addition to the severity of lameness. Tonsils were found to be the primary site of colonization, while bacteremia was rarely detected prior to the observation of clinical signs. Regardless of the clinical isolate, route of inoculation, or volume of inocula, histopathological alterations and tissue invasion were detected in multiple joints, indicating an apparent lack of specific joint tropism. Acute disease was primarily observed 7 to 10 days post-inoculation. The variability in the severity of synovial microscopic lesions and pathogen detection in joint cavities suggests that the duration of joint infection may influence the diagnostic accuracy. In summary, these findings demonstrate that diagnosis of M. hyosynoviae-associated arthritis can be influenced by the clinical isolate, and provides a study platform to investigate the colonization and virulence potential of field isolates. This approach can be particularly relevant to auxiliate in surveillance and testing of therapeutic and/or vaccine candidates.

Temperature-Sensitive Salmonella enterica Serovar Enteritidis PT13a Expressing Essential Proteins of Psychrophilic Bacteria.

Synthetic genes based on deduced amino acid sequences of the NAD-dependent DNA ligase (ligA) and CTP synthetase (pyrG) of psychrophilic bacteria were substituted for their native homologues in the genome of Salmonella enterica serovar Enteritidis phage type 13a (PT13a). The resulting strains were rendered temperature sensitive (TS) and did not revert to temperature resistance at a detectable level. At permissive temperatures, TS strains grew like the parental strain in broth medium and in macrophage-like cells, but their growth was slowed or stopped when they were shifted to a restrictive temperature. When injected into BALB/c mice at the base of the tail, representing a cool site of the body, the strains with restrictive temperatures of 37, 38.5, and 39°C persisted for less than 1 day, 4 to 7 days, and 20 to 28 days, respectively. The wild-type strain persisted at the site of inoculation for at least 28 days. The wild-type strain, but not the TS strains, was also found in spleen-plus-liver homogenates within 1 day of inoculation of the tail and was detectable in these organs for at least 28 days. Intramuscular vaccination of White Leghorn chickens with the PT13a strain carrying the psychrophilic pyrG gene provided some protection against colonization of the reproductive tract and induced an anti-S. enterica antibody response.

Protective immunity to Salmonella enterica is partially serogroup specific.

Pre-harvest reduction of Salmonella carriage by swine would benefit both animal health and food quality. While vaccination is an attractive pre-harvest intervention to reduce Salmonella levels in swine, the large number of potential Salmonella enterica serovars found in swine makes it critical that vaccines provide broad serotype efficacy. In order to directly compare the relative efficacy of Salmonella vaccines against serogroup-matched and serogroup-unmatched Salmonella, we vaccinated pigs with two commercially available Salmonella vaccines (either serogroup B or serogroup C1) and challenged with serovar-matched, serogroup-matched or serogroup-unmatched challenge strains. We found that while serogroup-matched vaccines provided relatively better efficacy than unmatched vaccines, serotype-unmatched vaccines also provided significant reduction of Salmonella carriage and shed. In addition, by measuring serogroup specific cell mediated (IFN-γ ELISPOT) and humoral (anti-LPS ELISA) immunity, we found that this serogroup specific efficacy correlates primarily with humoral immunity, while cell mediated immunity was mostly non-serogroup specific. While the practical relevance to pork quality of this serogroup-specific efficacy remains to be demonstrated, the large predominance of serogroup B Salmonella in swine suggests that a serogroup B Salmonella vaccine for swine would be of value to pre-harvest food safety interventions in swine.

Identification of novel pathogenicity loci in Clostridium perfringens strains that cause avian necrotic enteritis.

Type A Clostridium perfringens causes poultry necrotic enteritis (NE), an enteric disease of considerable economic importance, yet can also exist as a member of the normal intestinal microbiota. A recently discovered pore-forming toxin, NetB, is associated with pathogenesis in most, but not all, NE isolates. This finding suggested that NE-causing strains may possess other virulence gene(s) not present in commensal type A isolates. We used high-throughput sequencing (HTS) technologies to generate draft genome sequences of seven unrelated C. perfringens poultry NE isolates and one isolate from a healthy bird, and identified additional novel NE-associated genes by comparison with nine publicly available reference genomes. Thirty-one open reading frames (ORFs) were unique to all NE strains and formed the basis for three highly conserved NE-associated loci that we designated NELoc-1 (42 kb), NELoc-2 (11.2 kb) and NELoc-3 (5.6 kb). The largest locus, NELoc-1, consisted of netB and 36 additional genes, including those predicted to encode two leukocidins, an internalin-like protein and a ricin-domain protein. Pulsed-field gel electrophoresis (PFGE) and Southern blotting revealed that the NE strains each carried 2 to 5 large plasmids, and that NELoc-1 and -3 were localized on distinct plasmids of sizes approximately 85 and approximately 70 kb, respectively. Sequencing of the regions flanking these loci revealed similarity to previously characterized conjugative plasmids of C. perfringens. These results provide significant insight into the pathogenetic basis of poultry NE and are the first to demonstrate that netB resides in a large, plasmid-encoded locus. Our findings strongly suggest that poultry NE is caused by several novel virulence factors, whose genes are clustered on discrete pathogenicity loci, some of which are plasmid-borne.

Identification of multiple linear epitopes of the plasminogen activator A (PauA) of Streptococcus uberis with murine monoclonal antibodies.

Streptococcus (S.) uberis is a common cause of mastitis in cattle. A protein (PauA) secreted by this bacterium is capable of activating plasminogen from sheep and cattle. The PauA first binds to bovine plasminogen (b-plg) to form a PauA-plasminogen complex that subsequently binds to and activates b-plg to form plasmin. We have identified several linear epitopes of PauA that are recognized by murine monoclonal antibodies to PauA. Two of the monoclonal antibodies which neutralized the enzymatic activity of PauA, EC3 and 2.22, recognized common linear peptide sequences with similar charge and spacing patterns. These neutralization epitopes are located in the predicted alpha-domain of the PauA molecule. Further, these same epitopes are in critical structure/function domains identified in other studies. These characterizations may facilitate the design of an efficacious vaccine for streptococcal mastitis in the dairy cow.

Complex interactions between bovine plasminogen and streptococcal plasminogen activator PauA.

The interactions between bovine plasminogen and the streptococcal plasminogen activator PauA that culminate in the generation of plasmin are not fully understood. Formation of an equimolar activation complex comprising PauA and plasminogen by non-proteolytic means is a prerequisite to the recruitment of substrate plasminogen; however the determinants that facilitate these interactions have yet to be defined. A mutagenesis strategy comprising nested deletions and random point substitutions indicated roles for both amino and carboxyl-terminal regions of PauA and identified further essential residues within the alpha domain of the plasminogen activator. A critical region within the alpha domain was identified using non-overlapping PauA peptides to block the interaction between PauA and bovine plasminogen, preventing formation of the activation complex. Homology modelling of the activation complex based upon the known structures of streptokinase complexed with human plasmin supported these findings by placing critical residues in close proximity to the plasmin component of the activation complex.