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Salmonella enterica serotype Heidelberg is primarily a poultry adapted serotype of Salmonella that can also colonize other hosts and cause human disease. In this study, we compared the genomes of outbreak associated non-outbreak causing Salmonella ser. Heidelberg strains from diverse hosts and geographical regions. Human outbreak associated strains in this study were from a 2015 multistate outbreak of Salmonella ser. Heidelberg involving 15 states in the United States which originated from bull calves. Our clinicopathologic examination revealed that cases involving Salmonella ser. Heidelberg strains were predominantly young, less than weeks-old, dairy calves. Pre-existing or concurrent disease was found in the majority of the calves. Detection of Salmonella ser. Heidelberg correlated with markedly increased death losses clinically comparable to those seen in herds infected with S. Dublin, a known serious pathogen of cattle. Whole genome based single nucleotide polymorphism based analysis revealed that these calf isolates formed a distinct cluster along with outbreak associated human isolates. The defining feature of the outbreak associated strains, when compared to older isolates of S . Heidelberg , is that all isolates in this cluster contained Saf fimbrial genes which are generally absent in S . Heidelberg . The acquisition of several single nucleotide polymorphisms and the gain of Saf fimbrial genes may have contributed to the increased disease severity of these Salmonella ser. Heidelberg strains.

Background: Porcine reproductive and respiratory syndrome virus (PRRSV) causes reproductive failure in pregnant sows and acute respiratory disease in young pigs. It is a leading infectious agent of swine respiratory complex, which has significant negative economic impact on the swine industry. Commercial markets currently offer both live attenuated and killed vaccines; however, increasing controversy exists about their efficacy providing complete protection. Virus-like particles (VLPs) possess many desirable features of a potent vaccine candidate and have been proven to be highly immunogenic and protective against virus infections. Here we explored the efficacy of PRRSV VLPs together with the use of a novel 2', 3'-cGAMP VacciGrade™ adjuvant., Methods: Animals were immunized twice intranasally with phosphate buffered saline (PBS), PRRSV VLPs, or PRRSV VLPs plus 2', 3'-cGAMP VacciGrade™ at 2 weeks apart. Animals were challenged with PRRSV-23983 at 2 weeks post the second immunization. PRRSV specific antibody response and cytokines were measured. Viremia, clinical signs, and histological lesions were evaluated., Results: PRRSV N protein specific antibody was detected in all animals at day 10 after challenge, but no significant difference was observed among the vaccinated and control groups. Surprisingly, a significantly higher viremia was observed in the VLPs and VLPs plus the adjuvant groups compared to the control group. The increased viremia is correlated with a higher interferon-α induction in the serum of the VLPs and the VLPs plus the adjuvant groups., Conclusions: Intranasal immunizations of pigs with PRRSV VLPs and VLPs plus the 2', 3'-cGAMP VacciGrade™ adjuvant exacerbates viremia. A higher level of interferon-α production, but not interferon-γ and IL-10, is correlated with enhanced virus replication. Overall, PRRSV VLPs and PRRSV VLPs plus the adjuvant fail to provide protection against PRRSV challenge. Different dose of VLPs and alternative route of vaccination such as intramuscular injection should be explored in the future studies to fully assess the feasibility of such a vaccine platform for PRRSV control and prevention.

Background: This study describes a model developed to evaluate the transboundary risk of PEDV-contaminated swine feed ingredients and the effect of two mitigation strategies during a simulated transport event from China to the US., Results: Ingredients imported to the USA from China, including organic & conventional soybeans and meal, lysine hydrochloride, D-L methionine, tryptophan, Vitamins A, D & E, choline, carriers (rice hulls, corn cobs) and feed grade tetracycline, were inoculated with PEDV. Control ingredients, and treatments (ingredients plus a liquid antimicrobial (SalCURB, Kemin Industries (LA) or a 2% custom medium chain fatty acid blend (MCFA)) were tested. The model ran for 37 days, simulating transport of cargo from Beijing, China to Des Moines, IA, US from December 23, 2012 to January 28, 2013. To mimic conditions on land and sea, historical temperature and percent relative humidity (% RH) data were programmed into an environmental chamber which stored all containers. To evaluate PEDV viability over time, ingredients were organized into 1 of 4 batches of samples, each batch representing a specific segment of transport. Batch 1 (segment 1) simulated transport of contaminated ingredients from manufacturing plants in Beijing (day 1 post-contamination (PC)). Batch 2 (segments 1 and 2) simulated manufacturing and delivery to Shanghai, including time in Anquing terminal awaiting shipment (days 1-8 PC). Batch 3 (segments 1, 2 and 3) represented time in China, the crossing of the Pacific and entry to the US at the San Francisco, CA terminal (day 1-27 PC). Batch 4 (segments 1-4) represented the previous events, including transport to Des Moines, IA (days 1-37 PC). Across control (non-treated) ingredients, viable PEDV was detected in soybean meal (organic and conventional), Vitamin D, lysine hydrochloride and choline chloride. In contrast, viable PEDV was not detected in any samples treated with LA or MCFA., Conclusions: These results demonstrate the ability of PEDV to survive in a subset of feed ingredients using a model simulating shipment from China to the US. This is proof of concept suggesting that contaminated feed ingredients could serve as transboundary risk factors for PEDV, along with the identification of effective mitigation options.

Background: Contaminated complete feed and porcine plasma are risk factors for PEDV introduction to farms and a liquid antimicrobial has been proven useful for reducing risk. This study provides information on the survivability of PEDV across common swine feed ingredients in the presence or absence of the liquid antimicrobial., Results: Eighteen ingredients commonly included in commercial swine diets were selected, including 3 grain sources (corn, soybean meal (SBM), dried distillers grains with solubles (DDGS)), 5 porcine by-products (spray-dried plasma, purified plasma, intestinal mucosa, meat and bone meal and red blood cells (RBCs)), 3 vitamin/trace mineral (VTM) mixes (sow, nursery, finishing), 2 fat sources (choice white grease and soy oil), 3 synthetic amino acids (lysine HCL, D/L methionine, threonine), as well as limestone and dry choline chloride. Complete feed and stock PEDV served as controls. Thirty grams of each ingredient were inoculated with 2 mL PEDV. A matched set of samples were treated with the formaldehyde-based liquid antimicrobial SalCURB® (LA). All samples ( n  = 320) were stored outdoors under winter time ambient conditions for 30 days. Samples were submitted on 1, 7, 14 and 30 days post-inoculation (DPI) and tested by PCR and virus isolation (VI). All VI-negative samples were tested by swine bioassay. Viable PEDV was detected by VI or swine bioassay at 1, 7, 14 and 30 DPI from SBM, DDGS, meat & bone meal, RBCs, lysine HCL, D/L methionine, choice white grease, choline chloride, complete feed and stock virus control and at 7 DPI in limestone and at 14 DPI in threonine. Supplementary testing of complete feed and SBM indicated viable virus out to 45 and 180 DPI, respectively. All other samples were negative by VI and bioassay. In contrast, treatment with LA inactivated PEDV across all ingredients on 1 DPI and induced RNA reduction over time., Conclusions: Under the conditions of this study, PEDV viability in feed was influenced by ingredient with extended survival in SBM. Furthermore, LA treatment rendered virus inactive, independent of ingredient type.

Background: Porcine epidemic diarrhea virus (PEDV), a highly pathogenic and transmissible virus in swine, was first detected in the U.S. in May, 2013, and has caused tremendous losses to the swine industry. Due to the difficulty in isolating and growing this virus in cell culture, few vaccine studies using cell culture propagated PEDV have been performed on U.S. strains in pigs. Therefore, the objective of this study was to evaluate the humoral immune response to the selected inactivated PEDV vaccine candidate in a dose-titration manner., Results: PEDV was isolated from a pig with diarrhea and complete genome sequencing found >99% nucleotide identity to other U.S. PEDV. Inactivated adjuvanted monovalent vaccines were administered intramuscularly to five week old pigs in a dose titration experimental design, ranging from 6.0-8.0 log10 tissue culture infective dose (TCID50/mL), to evaluate immunogenicity using a fluorescent foci neutralization assay (FFN), fluorescent microsphere immunoassay (FMIA), and enzyme-linked immunosorbent assay (ELISA) on sera. Pigs vaccinated with 8.0 log10 TCID50/mL inactivated virus showed significantly higher FFN titers as well as FMIA and ELISA values than 6.0 log10 TCID50/mL vaccinates and the negative controls., Conclusions: These results demonstrate the immunogenicity of a PEDV inactivated viral vaccine with a U.S. strain via dose-titration. A future vaccination-challenge study would illustrate the efficacy of an inactivated vaccine and help evaluate protective FFN titers and ELISA and FMIA responses.

Background: Since its initial detection in May 2013, porcine epidemic diarrhea virus (PEDV) has spread rapidly throughout the US swine industry. Recently, contaminated feed was confirmed as a vehicle for PEDV infection of naïve piglets. This research provides in vivo data supporting the ability of a liquid antimicrobial product to reduce this risk., Results: Sal CURB® (Kemin Industries, Des Moines, IA, USA) is a FDA-approved liquid antimicrobial used to control Salmonella contamination in poultry and swine diets. To test its effect against PEDV, Sal CURB®-treated feed was spiked with a stock isolate of PEDV (Ct = 25.22), which PEDV-naïve piglets were allowed to ingest via natural feeding behavior (ad libitum) for a 14-day period. For the purpose of a positive control, a separate group of piglets was allowed to ingest non-treated (Sal CURB®-free) feed also spiked with stock PEDV (Ct = 25.22). A negative control group received PEDV-free feed. Clinical signs of PEDV infection (vomiting and diarrhea) and viral shedding in feces were observed in the positive control group 2-3 days post-consumption of non-treated feed. In contrast, no evidence of infection was observed in pigs fed Sal CURB®-treated feed or in the negative controls throughout the 14-day study period. In addition, the Sal CURB®-treated feed samples had higher (p < 0.0001) mean PEDV Ct values than samples from the positive control group., Conclusions: These data provide proof of concept that feed treated with Sal CURB® can serve as a means to reduce the risk of PEDV infection through contaminated feed. Furthermore, the results from the positive control group provide additional proof of concept regarding the ability of contaminated feed to serve as a risk factor for PEDV infection of naïve piglets.

Background: Since its initial detection in May 2013, porcine epidemic diarrhea virus (PEDV) has spread rapidly throughout the US swine industry. Initially, contaminated feed was proposed as a risk factor for PEDV; however, data were not available to support this theory. Here we provide proof of concept of this risk by describing a novel means for recovering PEDV-contaminated complete feed material from commercial swine sites and conducting an in vivo experiment to prove its infectivity., Results: For on-farm detection of PEDV RNA in feed, paint rollers were used to collect material from at-risk feed bins from 3 clinically affected breeding herds. This material was tested by PCR and determined to be positive for PEDV-RNA (Ct = 19.50-22.20 range). To test infectivity, this material was pooled (Ct = 20.65) and a Treatment group of 3-week old PEDV-naïve piglets were allowed to consume it via natural feeding behavior. For the purpose of a Positive control, piglets were allowed to ingest feed spiked with stock PEDV (Ct = 18.23) while the negative control group received PEDV-free feed. Clinical signs of PEDV infection (vomiting and diarrhea) and viral shedding were observed in both the Positive control and Treatment group' post-consumption with virus and microscopic lesions detected in intestinal samples No evidence of infection was observed in the Negative controls., Conclusions: These data provide proof of concept that contaminated complete feed can serve as a vehicle for PEDV infection of naïve pigs using natural feeding behavior.

Background: Parkinson's disease (PD) has been linked with exposure to a variety of environmental and immunological insults (for example, infectious pathogens) in which inflammatory and oxidative processes seem to be involved. In particular, epidemiological studies have found that pesticide exposure and infections may be linked with the incidence of PD. The present study sought to determine whether exposure to a viral mimic prior to exposure to pesticides would exacerbate PD-like pathology., Methods: Mice received a supra-nigral infusion of 5 μg of the double-stranded RNA viral analog, polyinosinic: polycytidylic acid (poly(I:C)), followed 2, 7 or 14 days later by administration of the pesticide, paraquat (nine 10 mg/kg injections over three weeks)., Results: As hypothesized, poly(I:C) pre-treatment enhanced dopamine (DA) neuron loss in the substantia nigra pars compacta elicited by subsequent paraquat treatment. The augmented neuronal loss was accompanied by robust signs of microglial activation, and by increased expression of the catalytic subunit (gp91) of the NADPH oxidase oxidative stress enzyme. However, the paraquat and poly(I:C) treatments did not appreciably affect home-cage activity, striatal DA terminals, or subventricular neurogenesis., Conclusions: These findings suggest that viral agents can sensitize microglial-dependent inflammatory responses, thereby rendering nigral DA neurons vulnerable to further environmental toxin exposure.