Polioencephalomyelitis in Domestic Swine Associated With Porcine Astrovirus Type 3.

In the past decade, different members of the genus Mamastrovirus have been associated with outbreaks of neurologic disease in humans, cattle, sheep, mink, and, most recently, porcine astrovirus 3 (PoAstV3) in swine. We performed a retrospective analysis of 50 cases of porcine neurologic disease of undetermined cause but with microscopic lesions compatible with a viral encephalomyelitis to better understand the role and pathogenesis of PoAstV3 infection. Nucleic acid was extracted from formalin-fixed paraffin-embedded (FFPE) tissue for reverse transcription quantitative polymerase chain reaction (RT-qPCR) testing for PoAstV3. In addition, 3 cases with confirmed PoAstV3-associated disease were assayed by RT-qPCR to investigate PoAstV3 tissue distribution. PoAstV3 was detected in central nervous system (CNS) tissue via RT-qPCR and in situ hybridization in 13 of 50 (26%) FFPE cases assayed. PoAstV3 was rarely detected in any tissues outside the CNS. Positive cases from the retrospective study included pigs in various production categories beginning in 2010, the earliest year samples were available. Based on these results, PoAstV3 appears to be a recurring putative cause of viral encephalomyelitis in swine that is rarely detected outside of the CNS at the time of clinical neurologic disease, unlike other common viral causes of neurologic disease in swine.

Recombination between Vaccine and Field Strains of Porcine Reproductive and Respiratory Syndrome Virus.

We isolated and plaque purified IA76950-WT and IA70388-R, 2 porcine reproductive and respiratory syndrome viruses from pigs in the same herd in Iowa, USA, that exhibited coughing and had interstitial pneumonia. Phylogenetic and molecular evolutionary analysis indicated that IA70388-R is a natural recombinant from Fostera PRRSV vaccine and field strain IA76950-WT.

First retrospective studies with etiological confirmation of porcine transmissible gastroenteritis virus infection in Argentina.

BACKGROUND: In 2014, a notification of porcine transmissible gastroenteritis virus (TGEV) was made by the National Services of Animal Health of Argentina (SENASA) to the World Organization of Animal Health (OIE). The notification was based on a serological diagnosis in a small farm with a morbidity rate of 2.3% without enteric clinical signs. In order to determine if TGEV was circulating before the official report, a retrospective study on cases of neonatal diarrhea was performed. The selection criteria was a sudden increase in mortality in 1- to 21-day-old piglets with watery diarrhea that did not respond to antibiotics. Based on these criteria, three clinical cases were identified during 2010-2015. RESULTS: All animals that were evaluated presented histological lesions consistent with enteric viral infection. The feces and ultrathin sections of intestine that were evaluated by electron microscopy confirmed the presence of round particles of approximately 80 nm in size and characterized by finely granular electrodense nucleoids consistent with complete particles of coronavirus. The presence of the TGEV antigen was confirmed by monoclonal specific immunohistochemistry, and final confirmation of a metabolically-active virus was performed by in situ hybridization to detect a TGE mRNA encoding spike protein. All sections evaluated in this case were negative for PEDV and rotavirus A. CONCLUSIONS: This is the first case series describing neonatal mortality with etiological confirmation of TGEV in Argentina. The clinical diagnosis of TGEV infections in endemic regions is challenging due to the epidemiological distribution and coinfection with other enteric pathogens that mask the clinical presentation.

Pathogenesis comparison between the United States porcine epidemic diarrhoea virus prototype and S-INDEL-variant strains in conventional neonatal piglets.

At least two genetically different porcine epidemic diarrhoea virus (PEDV) strains have been identified in the USA: US PEDV prototype and S-INDEL-variant strains. The objective of this study was to compare the pathogenicity differences of the US PEDV prototype and S-INDEL-variant strains in conventional neonatal piglets under experimental infections. Fifty PEDV-negative 5-day-old pigs were divided into five groups of ten pigs each and were inoculated orogastrically with three US PEDV prototype isolates (IN19338/2013, NC35140/2013 and NC49469/2013), an S-INDEL-variant isolate (IL20697/2014), and virus-negative culture medium, respectively, with virus titres of 104 TCID50 ml- 1, 10 ml per pig. All three PEDV prototype isolates tested in this study, regardless of their phylogenetic clades, had similar pathogenicity and caused severe enteric disease in 5-day-old pigs as evidenced by clinical signs, faecal virus shedding, and gross and histopathological lesions. Compared with pigs inoculated with the three US PEDV prototype isolates, pigs inoculated with the S-INDEL-variant isolate had significantly diminished clinical signs, virus shedding in faeces, gross lesions in small intestines, caeca and colons, histopathological lesions in small intestines, and immunohistochemistry staining in ileum. However, the US PEDV prototype and the S-INDEL-variant strains induced similar viraemia levels in inoculated pigs. Whole genome sequences of the PEDV prototype and S-INDEL-variant strains were determined, but the molecular basis of virulence differences between these PEDV strains remains to be elucidated using a reverse genetics approach.

Evaluation of serological cross-reactivity and cross-neutralization between the United States porcine epidemic diarrhea virus prototype and S-INDEL-variant strains.

BACKGROUND: At least two genetically different porcine epidemic diarrhea virus (PEDV) strains have been identified in the United States (U.S. PEDV prototype and S-INDEL-variant strains). The current serological assays offered at veterinary diagnostic laboratories for detection of PEDV-specific antibody are based on the U.S. PEDV prototype strain. The objectives of this study were: 1) isolate the U.S. PEDV S-INDEL-variant strain in cell culture; 2) generate antisera against the U.S. PEDV prototype and S-INDEL-variant strains by experimentally infecting weaned pigs; 3) determine if the various PEDV serological assays could detect antibodies against the U.S. PEDV S-INDEL-variant strain and vice versa. RESULTS: A U.S. PEDV S-INDEL-variant strain was isolated in cell culture in this study. Three groups of PEDV-negative, 3-week-old pigs (five pigs per group) were inoculated orally with a U.S. PEDV prototype isolate (previously isolated in our lab), an S-INDEL-variant isolate or virus-negative culture medium. Serum samples collected at 0, 7, 14, 21 and 28 days post inoculation were evaluated by the following PEDV serological assays: 1) indirect fluorescent antibody (IFA) assays using the prototype and S-INDEL-variant strains as indicator viruses; 2) virus neutralization (VN) tests against the prototype and S-INDEL-variant viruses; 3) PEDV prototype strain whole virus based ELISA; 4) PEDV prototype strain S1-based ELISA; and 5) PEDV S-INDEL-variant strain S1-based ELISA. The positive antisera against the prototype strain reacted to and neutralized both prototype and S-INDEL-variant viruses, and the positive antisera against the S-INDEL-variant strain also reacted to and neutralized both prototype and S-INDEL-variant viruses, as examined by IFA antibody assays and VN tests. Antibodies against the two PEDV strains could be detected by all three ELISAs although detection rates varied to some degree. CONCLUSIONS: These data indicate that the antibodies against U.S. PEDV prototype and S-INDEL-variant strains cross-reacted and cross-neutralized both strains in vitro. The current serological assays based on U.S. PEDV prototype strain can detect antibodies against both U.S. PEDV strains.

Bacterial probiotics as an aid in the control of Clostridium difficile disease in neonatal pigs.

Although Clostridium difficile infection (CDI) is a common disease in swine, there is a lack of prevention strategies. The objectives of this study were to evaluate: i) the effectiveness of Lactobacillus spp. and ii) non-toxigenic C. difficile (NTCD) as prevention for the development of CDI in piglets. Cesarean-derived piglets (N = 150) were randomly assigned to 6 groups: GROUP 1 - negative control (n = 10); GROUP 2 - NTCD only (n = 13); GROUP 3 - Lactobacillus spp. only (n = 14); GROUP 4 - positive control (challenged with toxigenic C. difficile strain) (n = 35); GROUP 5 - NTCD and challenged with the toxigenic C. difficile strain (n = 34); and GROUP 6 - Lactobacillus spp. and challenged with the toxigenic C. difficile strain (n = 44). Piglets which received NTCD showed lower prevalence of toxin-positive feces, mesocolonic edema, and microscopic lesions compared with positive control piglets. Administration of Lactobacillus spp. did not reveal clear benefits.

Probiotiques bactériens pour faciliter le contrôle de la maladie àClostridium difficilechez les porcelets néonataux. Même si l'infection par Clostridium difficile (ICD) est une maladie commune chez les porcs, il existe une absence de stratégies de prévention. Les objectifs de cette étude consistaient à évaluer: i) l'efficacité de Lactobacillus sp. et de ii) C. difficile non toxinogène (CDNT) comme méthode de prévention contre le développement de l'ICD chez les porcelets. Les porcelets délivrés par césarienne (N = 150) ont été assignés au hasard à 6 groupes: GROUPE 1 ­ groupe témoin négatif (n = 10); GROUPE 2 ­ CDNT seulement (n = 13); GROUPE 3 ­ Lactobacillus sp. seulement (n = 14); GROUPE 4 ­ groupe témoin positif (avec épreuve pour la souche toxinogène de C. difficile) (n = 35); GROUPE 5 ­ CDNT et avec épreuve pour la souche toxinogène de C. difficile (n = 34); et GROUPE 6 ­ Lactobacillus sp. et avec épreuve pour la souche toxinogène de C. difficile (n = 44). Les porcelets ayant reçu CDNT ont affiché une prévalence inférieure de fèces positives pour les toxines, de l'œdème du mésocôlon et de lésions microscopiques comparativement aux porcelets du groupe témoin positif. L'administration de Lactobacillus sp. n'a pas révélé de bienfaits évidents.(Traduit par Isabelle Vallières).

RETROSPECTIVE EVALUATION OF CASES OF NEOPLASIA IN A CAPTIVE POPULATION OF EGYPTIAN FRUIT BATS (ROUSETTUS AEGYPTIACUS).

Reports of neoplasia in Chiroptera species are rare. (6, 10) This retrospective study describes five types of neoplasia identified within a captive population of male Egyptian fruit bats (Rousettus aegyptiacus) housed in a zoo from 2004 through November of 2014. Tumor types identified include fibrosarcoma, cutaneous lymphoma, benign focal bronchioloalveolar neoplasm, anaplastic sarcoma, and sebaceous epithelioma. To the author's knowledge, aside from a recent report of focal brochioloalveolar adenoma, (8) these tumor types have not previously been described in the Rousettus species, nor in chiropterans in general. Based upon these findings and other recent publications regarding R. aegyptiacus, neoplasia does appear to be a significant cause of morbidity and mortality in captive members of this megachiropterid species.

Isolation and characterization of porcine epidemic diarrhea viruses associated with the 2013 disease outbreak among swine in the United States.

Porcine epidemic diarrhea virus (PEDV) was detected in May 2013 for the first time in U.S. swine and has since caused significant economic loss. Obtaining a U.S. PEDV isolate that can grow efficiently in cell culture is critical for investigating pathogenesis and developing diagnostic assays and for vaccine development. An additional objective was to determine which gene(s) of PEDV is most suitable for studying the genetic relatedness of the virus. Here we describe two PEDV isolates (ISU13-19338E and ISU13-22038) successfully obtained from the small intestines of piglets from sow farms in Indiana and Iowa, respectively. The two isolates have been serially propagated in cell culture for over 30 passages and were characterized for the first 10 passages. Virus production in cell culture was confirmed by PEDV-specific real-time reverse-transcription PCR (RT-PCR), immunofluorescence assays, and electron microscopy. The infectious titers of the viruses during the first 10 passages ranged from 6 × 10(2) to 2 × 10(5) 50% tissue culture infective doses (TCID50)/ml. In addition, the full-length genome sequences of six viruses (ISU13-19338E homogenate, P3, and P9; ISU13-22038 homogenate, P3, and P9) were determined. Genetically, the two PEDV isolates were relatively stable during the first 10 passages in cell culture. Sequences were also compared to those of 4 additional U.S. PEDV strains and 23 non-U.S. strains. All U.S. PEDV strains were genetically closely related to each other (≥99.7% nucleotide identity) and were most genetically similar to Chinese strains reported in 2011 to 2012. Phylogenetic analyses using different genes of PEDV suggested that the full-length spike gene or the S1 portion is appropriate for sequencing to study the genetic relatedness of these viruses.

Effect of age, dose and antibiotic therapy on the development of Clostridium difficile infection in neonatal piglets.

Piglet diarrhea is associated with increased pre-weaning mortality, poor growth rates, and variation in weight at weaning. Clostridium difficile is a known cause of enteric disease in neonatal piglets, yet risk factors associated with C. difficile infection in piglets are unknown. The objectives of this study were (1) to evaluate the consistency and severity of lesions in piglets challenged with C. difficile at different bacterial doses (DOSAGE experiment), (2) evaluate the use of antibiotics as a contributing risk factor in 1-day-old piglets (ANTIMICROBIAL experiment), and (3) to provide a clinical and histological evaluation of C. difficile infection in 10-day-old piglets (AGE experiment). One hundred and eleven conventional neonatal pigs were snatch farrowed and divided into experimental groups addressing the objectives. In the DOSAGE experiment, 40 1-day-old piglets were sham inoculated or challenged with varying amounts of C. difficile heat shocked spores and euthanized 72 h post infection. Results indicate a clear trend for disease development as bacterial numbers increase. In the ANTIMICROBIAL experiment, 39 1-day-old piglets were challenged and then treated with one of four different antibiotics after 16 h. No significant difference in disease development was found. Thirty-three 10-day-old piglets were given varying doses of C. difficile in the AGE experiment. Disease and lesions were reproduced in 10-day-old piglets. Combined results indicate that C. difficile dosage appears to be an important factor that influences the appearance and severity of lesions, 10-day-old pigs can develop disease associated with C. difficile, and antibiotic administration following inoculation may not be a major contributor for disease in neonatal piglets.

Systemic Conidiobolus incongruus infection and hypertrophic osteopathy in a white-tailed deer (Odocoileus virginianus).

Postmortem examination of a free-range white-tailed deer (Odocoileus virginianus) revealed severe emaciation, bilateral firm proliferation of the metatarsal diaphyses, and a large intrathoracic mass associated with the accessory lung lobe. Smaller masses were evident in the abomasum, duodenum, omentum, and the capsular surface of the liver. Microscopically, the masses were similar and were diagnosed as eosinophilic granulomas with intralesional fungal hyphae characteristic of Zygomycetes spp. Fungal hyphae were identified as Conidiobolus incongruus by 18S ribosomal RNA sequencing on fresh lung tissue. Furthermore, the proliferative lesions of the metatarsal bones along with the intrathoracic mass were compatible with hypertrophic osteopathy.

Characterization of shedding patterns of Porcine circovirus types 2a and 2b in experimentally inoculated mature boars.

Porcine circovirus-2 (PCV-2) is an economically important swine pathogen and causes PCV-associated disease (PCVAD) in pigs worldwide. Currently, 2 genotypes of PCV-2, PCV-2a and -2b, are circulating in U.S. swine herds. The objectives of the current study were to evaluate the amount of PCV-2 DNA present in semen over time, compare and correlate incidence and amount of PCV-2 present in semen samples to that present in serum samples and blood swabs, and determine if there are differences in shedding patterns between PCV-2a and -2b. Fifteen 7-month-old PCV-2-naïve Landrace boars (Sus scrofa) were randomly allocated to 3 treatment groups. The boars in group 1 (n = 3) served as negative controls, and those in groups 2 (n = 6) and 3 (n = 6) were intranasally and intramuscularly inoculated with PCV-2a and -2b, respectively. Semen, serum, and blood swab samples were collected up to 90 days postinoculation (DPI), and necropsies were performed on DPI 23, 48, and 90. Larger quantities of both PCV-2a and -2b DNA were detected earlier in serum and blood swab samples than in raw semen of experimentally inoculated boars. The incidence and duration of presence of PCV-2 DNA in semen varied among boars; however, intermittent shedding was not observed. In all sex glands, PCV-2 DNA was detected by polymerase chain reaction; however, PCV-2 antigen was not detected by immunohistochemistry, and PCV-2 had no effect on sperm morphology. Differences in shedding patterns between PCV-2a and -2b were not observed under the study conditions.

Determination of reference intervals for fluid analysis and cytologic evaluation variables in synovial fluid samples obtained from carpal and tarsal joints in commercial nonlame growing swine.

OBJECTIVE To determine reference intervals for total nucleated cell count, total protein concentration, pH, RBC count, and percentages of neutrophils, lymphocytes, and large mononuclear cells in synovial fluid samples (SFSs) obtained from the carpal and tarsal joints of healthy swine. ANIMALS 54 healthy commercial finisher pigs that had no evidence of lameness or gross joint swelling. PROCEDURES Each pig was anesthetized, and SFSs were collected from 1 carpal and 1 tarsal joint for fluid analysis, cytologic evaluation, bacterial culture, and PCR analyses for common swine joint pathogens. Each pig was euthanized after SFS collection, and synovial tissue samples were collected for histologic assessment. If necessary, postmortem SFSs were collected. RESULTS Overall, 37 of 50 tarsal and 46 of 53 carpal SFSs met inclusion criteria of sufficient volume, no gross blood contamination, and negative results of bacterial culture and PCR analyses, and were from joints with histologically normal synovial tissues. For the carpal and tarsal joints, upper reference limits were as follows: total nucleated cell count, 3,281 cells/µL and 2,368 cells/µL, respectively; total protein concentration, 3.6 g/dL and 3.6 g/dL, respectively; pH, 7.2 and 7.0, respectively; RBC count, 0.8 × 106 cells/µL and 0.1 × 106 cells/µL, respectively; and percentage of neutrophils, 46.5% and 33.7%, respectively; percentage of lymphocytes, 40.6% and 56.3%, respectively; and percentage of large mononuclear cells, 92.0% and 95.3%, respectively. CONCLUSIONS AND CLINICAL RELEVANCE Results have provided reference intervals for selected variables in SFSs obtained from the carpal and the tarsal joints of healthy swine, which should be useful in diagnostic investigations of swine lameness and arthritis.

Dendritic cell-targeted porcine reproductive and respiratory syndrome virus (PRRSV) antigens adjuvanted with polyinosinic-polycytidylic acid (poly (I:C)) induced non-protective immune responses against heterologous type 2 PRRSV challenge in pigs.

Porcine Reproductive and Respiratory Syndrome (PRRS) is an economically important swine viral disease worldwide. Current modified live-attenuated vaccines are ineffective against heterologous strains of PRRS virus (PRRSV) circulating in the field. In this study, we evaluated three dendritic cell (DC)-targeted vaccine candidates for their protective efficacy against heterologous PRRSV challenge. Ectodomain regions of DNA-shuffled structural proteins GP3, GP4, GP5 and M of PRRSV were fused together to form the vaccine antigen which was in turn fused with one of three recombinant antibodies each specific to a DC receptor: DC-SIGN, Langerin, and DEC205. The recombinant antibody-fused vaccine antigens were co-administered with polyinosinic-polycytidylic acid (poly (I:C)) adjuvant and subsequently challenged with a heterologous type 2 PRRSV strain (NADC20) in pigs. Our results demonstrate that pigs in DC-SIGN- and DEC205-targeted, but not Langerin- and non-targeted, vaccine groups showed significant IFN-γ- and IL-4-specific CD4T cell immune responses against the vaccine antigen in 7days post-challenge. Pigs in DC-SIGN- and Langerin-targeted vaccine groups showed greatly reduced IgG responses as compared to the DEC205- and non-targeted vaccine groups. The immune responses induced by DC-targeted vaccines did not reduce viremia and lung pathological lesions in type 2 PRRSV-challenged pigs. In contrast, pigs in Langerin-targeted vaccine group showed significantly increased serum viral titers and viral antigen in lung tissues at 7 and 14days post-challenge respectively. In conclusion, specific targeting of PRRSV antigen through DC-SIGN or DEC205 or Langerin-specific antibodies in the presence of poly (I:C) adjuvant induced immune responses that failed to protect pigs against heterologous type 2 PRRSV challenge.

Development of Polioencephalomyelitis in Cesarean-Derived Colostrum-Deprived Pigs Following Experimental Inoculation with Either Teschovirus A Serotype 2 or Serotype 11.

Teschovirus encephalomyelitis is a sporadic disease associated with Teschovirus A (PTV) serotype 1 and, less frequently, other serotypes. In recent years, the number of cases submitted to the Iowa State University Veterinary Diagnostic Laboratory with a history of posterior paresis has increased. Submission histories from various regions of the United States suggest a trend for clinical disease to persist in herds and affect a wider age-range of pigs than historically reported. Polioencephalitis and/or myelitis was consistently present and PTV was detected in affected neural tissue by PCR in a portion of cases. Sequencing from two clinical cases identified PTV-2 and PTV-11. To assess neuropathogenicity of these isolates, 5-week-old cesarean derived and colostrum-deprived pigs were assigned to three groups: negative control (n = 4), PTV-2-inoculated (n = 7), and PTV-11-inoculated (n = 7). Three PTV-2-inoculated pigs developed mild incoordination of the hind limbs, one of which progressed to posterior ataxia. While all PTV-11-inoculated pigs showed severe neurological signs consistent with Teschovirus encephalomyelitis, no evidences of neurological signs were observed in sham-inoculated animals. All PTV-2- and PTV-11-inoculated pigs had microscopic lesions consistent with Teschovirus encephalomyelitis. To our knowledge, this is the first description of PTV-11 and experimental study demonstrating the neuropathogenicity of PTV-11 in the United States.

Effects of label-dose permethrin administration in yearling beef cattle: I. Bull reproductive function and testicular histopathology.

Pyrethroid administration to a wide variety of laboratory animals has been shown to cause detrimental effects on male fertility, including sperm quality, by means of endocrine disruption. The objective of this experiment was to study the effects of a commercial, permethrin-containing pour-on product on reproductive variables and testicular histopathology of yearling beef bulls. Black Angus bulls (n = 60; aged 369 ± 17 days; 511 ± 33 kg; 6.2 ± 0.5 body condition scores) were assigned to either (1) saline control (CON) or (2) permethrin pour-on administered at label dose (PYR). Blood samples were collected, and industry standard breeding soundness examinations (BSE), via electroejaculation, were performed on all bulls at 5 days before and 14 days after treatment. Progressive sperm motility and eosin-nigrosin-stained sperm were analyzed using high-power phase-contrast microscopy. Plasma testosterone concentrations were analyzed via radioimmunoassay. Bulls were slaughtered at 34 days, and one testicle per bull was randomly collected for histologic examination. Change in sperm motility between BSEs was not different because of treatment; sperm morphology however improved across treatments, but PYR bulls had less improvement in percent of head (P < 0.001) sperm abnormalities compared to CON, resulting in less improvement of primary abnormalities (P = 0.04). Nonetheless, morphological differences did not change the overall outcome for satisfactory breeder status. Change in testosterone concentration did not differ because of treatment. Histopathologic examination identified that testicular degeneration and tubule diameter did not differ as a result of treatment. It should be noted, however, that degeneration score (higher score having more degeneration) was positively correlated with primary abnormalities (P < 0.01; r = 0.35) and negatively correlated with normal sperm cells (P < 0.001; r = -0.43). In summary, these data indicate that a single use of permethrin at label dose in yearling Angus bulls results in minimal detrimental effects on sperm morphology but not to a degree that impacts the ability of bulls to pass a standard BSE.

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.

Prophylactic Administration of Vector-Encoded Porcine Granulocyte-Colony Stimulating Factor Reduces Salmonella Shedding, Tonsil Colonization, and Microbiota Alterations of the Gastrointestinal Tract in Salmonella-Challenged Swine.

Salmonella colonization of food animals is a concern for animal health and public health as a food safety risk. Various obstacles impede the effort to reduce asymptomatic Salmonella carriage in food animals, including the existence of numerous serovars and the ubiquitous nature of Salmonella. To develop an intervention strategy that is non-specific yet effective against diverse Salmonella serovars, we explored the prophylactic use of a cytokine to decrease Salmonella in swine by boosting the host's innate immune system. Granulocyte-colony stimulating factor (G-CSF) is the major cytokine regulating the production, differentiation, function, and survival of neutrophils. Neutrophils play a critical role in the response to Salmonella; therefore, we evaluated the vectored-delivery of porcine G-CSF as a prophylactic to reduce Salmonella in pigs. Crossbred pigs, 5 weeks of age, were intramuscularly injected with a replication-defective human adenovirus (Ad5) engineered to express porcine G-CSF (Ad5-G-CSF, n = 9). Control pigs received the same Ad5 vector lacking the gene encoding G-CSF (Ad5-empty, n = 7). Four days later, all pigs (n = 16) were intranasally inoculated with 1 × 10(7) colony forming unit (CFU) of Salmonella enterica serovar Typhimurium UK1. At 2 and 3 days post-challenge with Salmonella, Ad5-G-CSF-treated pigs shed significantly less Salmonella (~10(3) CFU/g) in their feces than Ad5-empty-treated pigs (~10(4)-10(5) CFU/g; P < 0.05). A significant 4-log reduction in tonsil colonization was also observed in the Ad5-G-CSF-treated pigs at 7 days post-challenge (P < 0.05). In the gastrointestinal tract, the Peyer's patch region of the ileum exhibited a significant 0.5-log reduction in colonization in the Ad5-G-CSF-treated pigs (P < 0.05). The microbiota of all challenged pigs was assessed by sequencing and analyzing the V1-V3 region of the 16S rRNA gene from fecal DNA samples. The microbial community structure of Salmonella-challenged pigs was less disturbed post-challenge in the Ad5-G-CSF-treated pigs than the Ad5-empty-treated pigs. This suggests that Ad5-G-CSF administration mitigated changes in the microbial community structure caused by Salmonella challenge. Collectively, these data suggest that delivery of a targeted immunostimulant to enhance neutropoiesis may be a strategy to reduce Salmonella colonization, potentially during periods of immunological stress.

Does Circulating Antibody Play a Role in the Protection of Piglets against Porcine Epidemic Diarrhea Virus?

The contribution of circulating antibody to the protection of naïve piglets against porcine epidemic diarrhea virus (PEDV) was evaluated using a passive antibody transfer model. Piglets (n = 62) derived from 6 sows were assigned to one of 6 different treatments using a randomized block design which provided for allocation of all treatments to all sows' litters. Each treatment was designed to achieve a different level of circulating anti-PEDV antibody via intraperitoneally administration of concentrated serum antibody. Piglets were orally inoculated with PEDV (USA/IN/2013/19338E, 1 x 103 TCID50 per piglet) 24 hours later and then monitored for 14 days. Piglets remained with their dam throughout the experiment. Sow milk samples, piglet fecal samples, and data on piglet clinical signs, body weight, and body temperature were collected daily. Fecal samples were tested by PEDV real-time reverse transcriptase PCR. Serum, colostrum, and milk were tested for PEDV IgG, IgA, and virus-neutralizing antibody. The data were evaluated for the effects of systemic PEDV antibody levels on growth, body temperature, fecal shedding, survival, and antibody response. The analysis showed that circulating antibody partially ameliorated the effect of PEDV infection. Specifically, antibody-positive groups returned to normal body temperature faster and demonstrated a higher rate of survivability than piglets without PEDV antibody. When combined with previous literature on PEDV, it can be concluded that both systemic antibodies and maternal secretory IgA in milk contribute to the protection of the neonatal pig against PEDV infections. Overall, the results of this experiment suggested that passively administered circulating antibodies contributed to the protection of neonatal piglets against PEDV infection.

Tissue localization, shedding, virus carriage, antibody response, and aerosol transmission of Porcine epidemic diarrhea virus following inoculation of 4-week-old feeder pigs.

We determined tissue localization, shedding patterns, virus carriage, antibody response, and aerosol transmission of Porcine epidemic diarrhea virus (PEDV) following inoculation of 4-week-old feeder pigs. Thirty-three pigs were randomly assigned to 1 of 3 groups for the 42-day study: inoculated (group A; n = 23), contact transmission (group B; n = 5), and aerosol transmission (group C; n = 5). Contact transmission occurred rapidly to group B pigs whereas productive aerosol transmission failed to occur to group C pigs. Emesis was the first clinical sign noted at 3 days postinoculation (dpi) followed by mild to moderate diarrhea lasting 5 more days. Real-time PCR detected PEDV in fecal and nasal swabs, oral fluids, serum, and gastrointestinal and lymphoid tissues. Shedding occurred primarily during the first 2 weeks postinoculation, peaking at 5-6 dpi; however, some pigs had PEDV nucleic acid detected in swabs collected at 21 and 28 dpi. Antibody titers were measurable between 14 and 42 dpi. Although feces and intestines collected at 42 dpi were PEDV negative by PCR and immunohistochemistry, respectively, small intestines from 70% of group A pigs were PCR positive. Although disease was relatively mild and transient in this age group, the results demonstrate that 4-week-old pigs are productively infected and can sustain virus replication for several weeks. Long-term shedding of PEDV in subclinically affected pigs should be considered an important source for PEDV transmission.