Implementation of PRRSV status classification system in swine breeding herds from a large integrated group in Spain

Background: Porcine Reproductive and Respiratory Syndrome (PRRS) is an endemic swine disease causing significant productive and economic losses. Knowledge of PRRS epidemiology is crucial to develop control strategies against this disease. In that regard, classifying farms according to PRRS virus (PRRSV) shedding and exposure, and understanding key drivers of change in status over time, provides great applied knowledge for developing disease control programs. In most European countries, PRRSV monitoring is performed most frequently at the individual farm level although criteria selected for monitoring varies among different regions and farms. The aim of this study was to implement a systematic monitoring program for PRRSV in Spanish sow farms. Breeding herds were classified according to a standardized PRRSV infection status using sampling programs and terminology currently adopted in the United States (US), which allowed an evaluation of PRRSV epidemiology in a large integrated Spanish group during a one-year study period (February 2017–March 2018). Results: Fifteen farms achieved a stable PRRSV status after the first 4 consecutive samplings and 20 farms were classified as unstable. One of the farms maintained a stable status throughout the duration of the whole monitoring period. Among the 20 farms classified as unstable at the beginning of the monitoring protocol, 9 farms (45%) never reached the stable status and 11 farms (55%) reached stable status afterwards during the monitoring study period. From PRRSV PCR positive pools, there were 47 different PRRSV nucleotide sequences from 24 different farms. More than one PRRSV sequence was obtained from 15 farms. In the farms with more than one sequence detected, we observed recirculation of the same PRRSV field strain in 7 farms and introduction of a different PRRSV strain in 5 farms and both events in 3 farms. Conclusions: Systematic monitoring for PRRSV in breeding herds established a basis of knowledge of PRRSV epidemiology at the farm level and provided key data to classify farms according to PRRSV exposure and shedding status. These data allow further evaluation of the impact of the PRRSV farm status on production and economic performance in breeding herds and additional investigation of factors related to PRRSV epidemiology

Porcine Epidemic Diarrhea Virus Surface Decontamination Strategies Using Chemical Sanitizing to Reduce the Quantity of PEDV RNA on Feed Manufacturing Surfaces with Environmental Swabbing

Porcine Epidemic Diarrhea virus (PEDV) is a possible hazard in feed mills that could impact pig health. If the virus enters a feed mill, it quickly becomes widely distributed and is difficult to decontaminate from surfaces.6,7 The objective of this study was to evaluate a variety of liquid and dry chemical treatments that could be used as sanitizers to reduce the amount of PEDV found on feed manufacturing surfaces in mills. This experiment was replicated 3 times and was designed in a 5 × 10 factorial with main effects of 5 different feed manufacturing surfaces and 10 sanitizing treatments. Surfaces included stainless steel, plastic, rubber, woven polypropylene tote bag, and sealed concrete coupons (4 × 4 in). One mL (1×105 TCID50/mL) of stock PEDV was applied to each surface and allowed to dry completely for 60 min. Next, a mitigation treatment was applied for 15 min: 1) no sanitation treatment (control); 2) untreated rice hulls; 3) rice hulls treated with formaldehyde-based commercial product (Sal CURB; Kemin Inc., Des Moines, IA); 4) liquid formaldehyde-based commercial product (Sal CURB; Kemin Inc., Des Moines, IA); 5) dry commercial benzoic acid and probiotic blend (VevoVitall and CRINA; DSM Nutritional Products Inc., Parsippany, NJ); 6) liquid ammonium chloride, isopropanol, and hydrogen peroxide-based commercial food-grade sanitizer (DrySan Duo; Ecolab, St. Paul, MN); 7) liquid hydrogen peroxide commercial product (INTERvention; Virox Technologies Inc. Ontario, Canada); 8) liquid quaternary ammonium glutaraldehyde commercial product (Synergize; Preserve International, Reno NV); 9) liquid sodium hypochlorite commercial sanitizer (Bleach; Clorox, Oakland, CA); and 10) liquid medium chain fatty acid blend of caprylic, caproic, and capric acids. There were 3 replicates per treatment. The quantity of PEDV RNA was determined using qRT-PCR. All main effects, interaction, and comparisons were highly significant (P ≤ 0.001). Liquid Sal CURB and liquid bleach were the most effective chemical treatments to reduce the quantity of detectable PEDV RNA, but their application is limited due to their liquid state and potential corrosiveness. Additional research is necessary to identify the role of sanitizer on PEDV infectivity, even if RNA residue remains, and to develop dry sanitizers capable of removing PEDV RNA on swine feed manufacturing surfaces that are not corrosive

Utilizing Feed Sequencing to Decrease the Risk of Porcine Epidemic Diarrhea Virus (PEDV) Cross-contamination During Feed Manufacturing

Understanding key points of potential cross-contamination during the feed manufacturing process is important to developing efficacious methods to control or prevent transmission of pathogens into swine diets. In this study, an experiment was conducted involving 30 crossbred 10-d-old pigs that were used as a bioassay model for Porcine Epidemic Diarrhea Virus (PEDV) to determine the effects of feed batch sequencing on PEDV cross-contamination and subsequent infectivity. PEDV with a PCR cycle threshold value (Ct) of 11 was uniformly mixed into 4.5 kg of swine diet using a stainless steel bench top mixer validated for mixing efficiency. The inoculated feed was then added to 45 kg of swine diet and mixed using a 4 ft3 electric paddle mixer validated for mixing efficiency to form the positive experimental treatment. Feed was discharged, carried into a bucket elevator, and exited through a downspout. Subsequent treatment batches were formed when 50 kg of PEDV-free swine diet was sequenced immediately after the PEDV-inoculated batch without cleaning the equipment to replicate the batching process used in commercial feed mills. The subsequent sequence batches (1-4) mixed, discharged, and sampled similar to the PEDV-positive batch. Feed samples were analyzed for the presence of PEDV using PCR and bioassay. Pigs were then orally challenged with harvested supernatant. Fecal swabs were collected for PEDV PCR testing. At seven days after challenge, all pigs were necropsied. Cecum contents, ileum, and jejunum were collected for PCR, histologic, and immunohistochemistry (IHC) evaluation. Overall, the results indicate that sequencing reduced but did not eliminate the risk of PEDV transmission. All pigs (9/9) challenged with the positive treatment were infected with PEDV with feed that had a Ct mean of 31.7. The discharge for the first sequence had a Ct value of 38.1 and infected pigs were noted in pigs from one of three rooms used to bioassay the feed. The second sequence did not have detectable PEDV RNA by using PCR. Interestingly, feed from the second sequence was infectious as verified by infection in pigs from one of three rooms used for bioassay. This study is the first to demonstrate feed without detectable PEDV RNA can be infective but is similar to other research using tissue homogenates and cell culture as bioassay material. In summary, feed batch sequencing should be considered a risk mitigation strategy that can be incorporated into feed mill biosecurity programs but should not be considered a risk elimination strategy

Determining the Minimum Infectious Dose of Porcine Epidemic Diarrhea Virus (PEDV) in a Feed Matrix

Understanding the magnitude of transmissible risk Porcine Epidemic Diarrhea Virus (PEDV)-infected feed imposes and establishing the minimum infectious dose of PEDV in a feed matrix are important components in strengthening virus prevention and control methods. In this study, an experiment was performed involving 30 crossbred, 10-d-old pigs that were used as a bioassay model for the minimum infectious dose of PEDV in feed. The PEDV was first diluted using tissue culture media to form 8 serial 10-fold dilutions. An aliquot of the original stock virus at 5.6 x 105 tissue culture infectious dose/ml (TCID50/ml), each serial PEDV dilution, and one virus-negative culture medium were mixed into separate 4.5 kg batches of swine diet to form 10 experimental treatments. The feed was then subsequently evaluated for infectivity using bioassay. Fecal swabs were collected at 0, 2, 4, 6, and 7 d after challenge for PCR testing. At 7 d after challenge, all pigs were necropsied. Cecum contents, ileum and jejunum were collected for PCR, histologic and immunohistochemistry (IHC) evaluation. Overall, the results indicate 5.6 × 101 TCID50/g was the minimum PEDV dose in which infection was detected. This feed had a corresponding PCR cycle threshold (Ct) of 37. This is a relatively low dose. To illustrate, using this dose, approximately 1 g of PEDV-infected baby piglet feces could contaminate up to 500 tons of feed. The data confirm that detectable Ct values in feed can result in pig infection. Our results also illustrate that the Ct in feed that was detected as infectious can be above the detection threshold used by some diagnostic laboratories

Evaluating the Effect of Manufacturing Porcine Epidemic Diarrhea Virus (PEDV)-Contaminated Feed on Subsequent Feed Mill Environmental Surface Contamination

This study aimed to utilize the only known pilot feed mill facility approved for pathogenic feed agent use in the United States to evaluate the effect of manufacturing Porcine Epidemic Diarrhea Virus (PEDV)-contaminated feed on subsequent feed mill environmental surface contamination. In this study, PEDV inoculated feed was manufactured and conveyed on equipment along with four subsequent batches of PEDV-free feed. Equipment and environmental surfaces were sampled using swabs and analyzed for the presence of PEDV RNA by PCR. The experiment was replicated three times with decontamination of the feed mill and all equipment between replications. Overall, environmental swabs indicated widespread surface contamination of the equipment and work area after a PEDV contaminated batch of feed was processed. There was little difference in environmental sample cycle threshold (Ct) values after manufacturing each of the subsequent PEDV-negative feed batches. In summary, introduction of PEDV-infected feed into a feed mill will likely result in widespread contamination of equipment and surfaces, even after several batches of PEDV-free feed are produced. Eliminating the PEDV RNA from the feed mill environment was challenging and required procedures that are not practical to apply on a regular basis in a feed mill. This data suggests that it is extremely important to prevent the introduction of PEDV-contaminated feed, ingredients, or other vectors of transmission to minimize PEDV-risk. More research should be conducted to determine if contaminated surfaces can lead to PEDV infectivity and to determine the best feed mill PEDV-decontamination strategies

Efficacy in Pigs of Inactivated and Live Attenuated Influenza Virus Vaccines against Infection and Transmission of an Emerging H3N2 Similar to the 2011-2012 H3N2v

Vaccines provide a primary means to limit disease but may not be effective at blocking infection and pathogen transmission. The objective of the present study was to evaluate the efficacy of commercial inactivated swine influenza A virus (IAV) vaccines and experimental live attenuated influenza virus (LAIV) vaccines against infection with H3N2 virus and subsequent indirect transmission to naive pigs. The H3N2 virus evaluated was similar to the H3N2v detected in humans during 2011-2012, which was associated with swine contact at agricultural fairs. One commercial vaccine provided partial protection measured by reduced nasal shedding; however, indirect contacts became infected, indicating that the reduction in nasal shedding did not prevent aerosol transmission. One LAIV vaccine provided complete protection, and none of the indirect-contact pigs became infected. Clinical disease was not observed in any group, including nonvaccinated animals, a consistent observation in pigs infected with contemporary reassortant H3N2 swine viruses. Serum hemagglutination inhibition antibody titers against the challenge virus were not predictive of efficacy; titers following vaccination with a LAIV that provided sterilizing immunity were below the level considered protective, yet titers in a commercial vaccine group that was not protected were above that level. While vaccination with currently approved commercial inactivated products did not fully prevent transmission, certain vaccines may provide a benefit by limitating shedding, transmission, and zoonotic spillover of antigenically similar H3N2 viruses at agriculture fairs when administered appropriately and used in conjunction with additional control measures

Evaluating the Impact of VevoVitall and/or CRINA as Potential Porcine Epidemic Diarrhea Virus Mitigation Strategies as Determined by Polymerase Chain Reaction Analysis and Bioassay

Feed and feed ingredients have been shown to be potential vectors of porcine epidemic diarrhea virus (PEDV). Potential strategies to mitigate the risk of disease transmission via feed and feed ingredients would be valuable to the swine and feed milling industries. Therefore, the objective of this experiment was to determine the impact of VevoVitall (5,000 ppm; DSM Nutritional Products Inc., Parsipanny, NJ), CRINA (200 ppm; DSM Nutritional Products Inc., Parsipanny, NJ), and a combination of both products (COMBINATION; 5,000 ppm VevoVitall and 200 ppm CRINA) as feed additives with potential to mitigate the risk of PEDV, in swine gestation diet (FEED) and spraydried porcine plasma (SDPP) as determined by real time quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) analyzed at seven sampling days post laboratory inoculation (d 0, 1, 3, 7, 14, 21, and 42) and bioassay. There was a marginally significant treatment × feed matrix × day interaction (P = 0.082), in which the cycle threshold (Ct) value increased over time in the diet when treated with the COMBINATION, whereas, there was no increase over time observed in SDPP. There was a highly significant (P \u3c 0.001) feed matrix × day interaction in which the Ct increased over time in FEED, whereas, there was very little increase over time observed in SDPP. Additionally, there was a marginally significant treatment × feed matrix interaction (P = 0.079). Overall, the COMBINATION was most effective at reducing the quantity of genetic material as detected by qRT-PCR (P \u3c 0.001). Virus shedding was observed in the d 7 post-inoculation SDPP COMBINATION treatment, as well as d 0 FEED COMBINATION treatment. No other treatment bioassay room had detectable RNA shed and detected in fecal swabs or cecal contents (d 1, 3, 7, 14, and 21 post-laboratory inoculation FEED, COMBINATION). In summary, the combination of CRINA and VevoVitall enhanced degradation of PEDV RNA in swine feed, but had no impact on RNA degradation in SDPP. Furthermore, both untreated feed and feed treated with the combination of CRINA and VevoVitall caused infection at d 0 post-laboratory inoculation; however, neither set of samples was infective at d 1 post-laboratory inoculation