Greater than 3-Log Reduction in Viable Coronavirus Aerosol Concentration in Ducted Ultraviolet-C (UV-C) Systems.

Control technologies to inactivate airborne viruses effectively are needed during the ongoing SARS-CoV-2 pandemic, and to guard against airborne transmitted diseases. We demonstrate that sealed UV-C flow reactors operating with fluences near 253 ± 1 nm of 13.9-49.6 mJ cm-2 efficiently inactivate coronaviruses in an aerosol. For measurements, porcine respiratory coronavirus (PRCV) was nebulized in a custom-built, 3.86 m wind tunnel housed in a biosafety level class II facility. The single pass log10 reduction of active coronavirus was in excess of 2.2 at a flow rate of 2439 L min-1 (13.9 mJ cm-2) and in excess of 3.7 (99.98% removal efficiency) at 684 L min-1 (49.6 mJ cm-2). Because virus titers resulting from sampling downstream of the UV-C reactor were below the limit of detection, the true log reduction is likely even higher than measured. Comparison of virus titration results to reverse transcriptase quantitative PCR and measurement of fluorescein concentrations (doped into the nebulized aerosol) reveals that the reduction in viable PRCV is primarily due to UV-C based inactivation, as opposed to physical collection of virus. The results confirm that UV-C flow reactors can efficiently inactivate coronaviruses through incorporation into HVAC ducts or recirculating air purifiers.

Wind tunnel-based testing of a photoelectrochemical oxidative filter-based air purification unit in coronavirus and influenza aerosol removal and inactivation.

Recirculating air purification technologies are employed as potential means of reducing exposure to aerosol particles and airborne viruses. Toward improved testing of recirculating air purification units, we developed and applied a medium-scale single-pass wind tunnel test to examine the size-dependent collection of particles and the collection and inactivation of viable bovine coronavirus (BCoV, a betacoronavirus), porcine respiratory coronavirus (PRCV, an alphacoronavirus), and influenza A virus (IAV), by a commercial air purification unit. The tested unit, the Molekule Air Mini, incorporates a MERV 16 filter as well as a photoelectrochemical oxidating layer. It was found to have a collection efficiency above 95.8% for all tested particle diameters and flow rates, with collection efficiencies above 99% for supermicrometer particles with the minimum collection efficiency for particles smaller than 100 nm. For all three tested viruses, the physical tracer-based log reduction was near 2.0 (99% removal). Conversely, the viable virus log reductions were found to be near 4.0 for IAV, 3.0 for BCoV, and 2.5 for PRCV, suggesting additional inactivation in a virus family- and genus-specific manner. In total, this work describes a suite of test methods which can be used to rigorously evaluate the efficacy of recirculating air purification technologies.

Comparison of dermoscopic features between congenital and acquired acral melanocytic nevi in Korean patients.

BACKGROUND: The dermoscopic patterns of acral melanocytic nevi (AMNs) are crucial in differentiating them from acral melanoma. Several studies have reported the dermoscopic patterns of acquired acral melanocytic nevi (AAMNs). However, few have investigated the dermoscopic patterns of congenital acral melanocytic nevi (CAMNs). OBJECTIVE: To compare the clinical and dermoscopic features of CAMNs and AAMNs. METHODS: The present study included 43 patients with CAMNs and 40 with AAMNs. We reviewed their medical records as well as their clinical and dermoscopic findings. RESULTS: Congenital acral melanocytic nevis were more asymmetrical than AAMNs (P = 0.002) and presented more frequently as comma-shaped (P = 0.005). Regarding dermoscopic findings, globular pattern (55.8%) was the most common feature of CAMNs, while parallel furrow pattern (37.5%) was the most common feature of AAMNs. The presence of fibrillar, globular, and parallel ridge patterns, and diffuse multi-component pigmentation differed significantly between the groups (P < 0.05). Furthermore, CAMNs showed melanoma-specific dermoscopic patterns, such as parallel ridge (18.6%) and diffuse multi-component pigmentation (25.6%). CONCLUSION: The dermoscopic patterns of CAMNs and AAMNs differed markedly. In terms of dermoscopic patterns, CAMNs resembled acral melanoma more often than AAMNs did.

Longitudinal study of Staphylococcus aureus colonization and infection in a cohort of swine veterinarians in the United States.

BACKGROUND: People working with pigs are at elevated risk of harboring methicillin resistant S. aureus (MRSA) in their nose, which is attributable to occupational exposure to animals harboring livestock adapted S. aureus. To obtain insight into the biological nature of occupationally related nasal culture positivity, we conducted a longitudinal study of 66 swine veterinarians in the USA. METHODS: The study cohort resided in 15 US states and worked predominantly with swine. Monthly for 18 months, participants self-collected nasal swabs and completed a survey to report recent exposure to pigs and other animals; the occurrence of work related injuries; and any relevant health events such as skin and soft tissue infections or confirmed staphylococcal infections. Nasal swabs were cultured using selective methods to determine the presence of MRSA and methicillin susceptible S. aureus (MSSA), and isolates were characterized by spa typing and MLST. RESULTS: Prevalences of S. aureus (64%, monthly range from 58 to 82%) and MRSA (9.5%; monthly range from 6 to15%) were higher than reported for the US population (30% and 1.5% respectively). Predominant spa types were t034 (ST398, 37%), t002 (ST5, 17%) and t337 (ST9/ST398 13%), a distribution similar to that found in a concurrent study in pigs in the USA. Veterinarians were classified into three groups: Persistent carriers (PC, 52%), Intermittent carriers (IC, 47%) and Non-carriers (NC, 1%). Persistent carriage of a single spa type was observed in 14 (21%) of participants, and paired (first and last) isolates from PC subjects had minor genetic differences. Swabs from PC veterinarians carried higher numbers of S. aureus. Among IC veterinarians, culture positivity was significantly associated with recent contact with pigs. CONCLUSIONS: Exposure to pigs did not lead to prolonged colonization in most subjects, and the higher numbers of S. aureus in PC subjects suggests that unknown host factors may determine the likelihood of prolonged colonization by S. aureus of livestock origin. Exposure to S. aureus and persistent colonization of swine veterinarians was common but rarely associated with S. aureus disease.

Evaluation of biosecurity measures to prevent indirect transmission of porcine epidemic diarrhea virus.

BACKGROUND: The effectiveness of biosecurity methods to mitigate the transmission of porcine epidemic diarrhea virus (PEDV) via farm personnel or contaminated fomites is poorly understood. This study was undertaken to evaluate the effectiveness of biosecurity procedures directed at minimizing transmission via personnel following different biosecurity protocols using a controlled experimental setting. RESULTS: PEDV RNA was detected from rectal swabs of experimentally infected (INF) and sentinel pigs by real-time reverse transcription polymerase chain reaction (rRT-PCR). Virus shedding in INF pigs peaked at 1 day post infection (dpi) and viral RNA levels remained elevated through 19 dpi. Sentinel pigs in the low biosecurity group (LB) became PEDV positive after the first movement of study personnel from the INF group. However, rectal swabs from pigs in the medium biosecurity (MB) and high biosecurity (HB) groups were negative during the 10 consecutive days of movements and remained negative through 24 days post movement (dpm) when the first trial was terminated. Viral RNA was detected at 1 dpm through 3 dpm from the personal protective equipment (PPE) of LB personnel. In addition, at 1 dpm, 2 hair/face swabs from MB personnel were positive; however, transmission of virus was not detected. All swabs of fomite from the HB study personnel were negative. CONCLUSIONS: These results indicate that indirect PEDV transmission through contaminated PPE occurs rapidly (within 24 h) under modeled conditions. Biosecurity procedures such as changing PPE, washing exposed skin areas, or taking a shower are recommended for pig production systems and appear to be an effective option for lowering the risk of PEDV transmission between groups of pigs.

Live Animal Markets in Minnesota: A Potential Source for Emergence of Novel Influenza A Viruses and Interspecies Transmission.

BACKGROUND: Live animal markets have been implicated in transmission of influenza A viruses (IAVs) from animals to people. We sought to characterize IAVs at 2 live animal markets in Minnesota to assess potential routes of occupational exposure and risk for interspecies transmission. METHODS: We implemented surveillance for IAVs among employees, swine, and environment (air and surfaces) during a 12-week period (October 2012-January 2013) at 2 markets epidemiologically associated with persons with swine-origin IAV (variant) infections. Real-time reverse transcription polymerase chain reaction (rRT-PCR), viral culture, and whole-genome sequencing were performed on respiratory and environmental specimens, and serology on sera from employees at beginning and end of surveillance. RESULTS: Nasal swabs from 11 of 17 (65%) employees tested positive for IAVs by rRT-PCR; 7 employees tested positive on multiple occasions and 1 employee reported influenza-like illness. Eleven of 15 (73%) employees had baseline hemagglutination inhibition antibody titers ≥40 to swine-origin IAVs, but only 1 demonstrated a 4-fold titer increase to both swine-origin and pandemic A/Mexico/4108/2009 IAVs. IAVs were isolated from swine (72/84), air (30/45), and pen railings (5/21). Whole-genome sequencing of 122 IAVs isolated from swine and environmental specimens revealed multiple strains and subtype codetections. Multiple gene segment exchanges among and within subtypes were observed, resulting in new genetic constellations and reassortant viruses. Genetic sequence similarities of 99%-100% among IAVs of 1 market customer and swine indicated interspecies transmission. CONCLUSIONS: At markets where swine and persons are in close contact, swine-origin IAVs are prevalent and potentially provide conditions for novel IAV emergence.

Circulating Mycobacterium bovis peptides and host response proteins as biomarkers for unambiguous detection of subclinical infection.

Bovine tuberculosis remains one of the most damaging diseases to agriculture, and there is also a concern for human spillover. A critical need exists for rapid, thorough, and inexpensive diagnostic methods capable of detecting and differentiating Mycobacterium bovis infection from other pathogenic and environmental mycobacteria at multiple surveillance levels. In a previous study, Seth et al. (PLoS One 4:e5478, 2009, doi:10.1371/journal.pone.0005478) identified 32 host peptides that specifically increased in the blood serum of M. bovis-infected animals). In the current study, 16 M. bovis proteins were discovered in the blood serum proteomics data sets. A large-scale validation analysis was undertaken for selected host and M. bovis proteins using a cattle serum repository containing M. bovis (n = 128), Mycobacterium kansasii (n = 10), and Mycobacterium avium subsp. paratuberculosis (n = 10), cases exposed to M. bovis (n = 424), and negative controls (n = 38). Of the host biomarkers, vitamin D binding protein (VDBP) showed the greatest sensitivity and specificity for M. bovis detection. Circulating M. bovis proteins, specifically polyketide synthetase 5, detected M. bovis-infected cattle with little to no seroreactivity against M. kansasii- and M. avium subsp. paratuberculosis-infected animals. These data indicate that host and pathogen serum proteins can serve as reliable biomarkers for tracking M. bovis infection in animal populations.

Decreased expression of PIAS1 and PIAS3 in essential thrombocythemia patients.

Gain of function mutation of Janus kinase 2 (JAK2V617F) has been identified in Philadelphia-negative myeloproliferative diseases; about half of essential thrombocythemia (ET) patients harbor this mutation. The activated JAK-STAT pathway promotes cell proliferation, differentiation and anti-apoptosis. We studied the role of negative regulators of the JAK-STAT pathway, PIAS, and SOCS in ET patients. Twenty ET patients and 20 healthy individuals were enrolled in the study. Thirteen of the ET patients harbored the JAK2V617F mutation based on mutation analysis. Quantitative-PCR was applied to assay the expression of SOCS1, SOCS3, PIAS1, PIAS3. The expression levels of PIAS1 and PIAS3 were significantly lower in ET groups than that in normal individuals. There was no significant difference between JAK2V617F (+) and JAK2V617F (-) patients. SOCS1 and SOCS3 expression did not differ between ET patients and normal individuals, or between JAK2V617F (+) and JAK2V617F (-) patients. We suggest that failed negative regulators of the JAK-STAT pathway take part in the pathomechanism of ET.

Infection dynamics and incidence of wild-type porcine reproductive and respiratory syndrome virus in growing pig herds in the U.S. Midwest.

Porcine reproductive and respiratory syndrome virus (PRRSV) infections greatly impact the health and productivity of growing pigs. The introduction and persistence of wild-type PRRSV (WT-PRRSV) strains in growing pig populations is poorly understood. In an observational prospective cohort study, we monitored and surveyed 63 wean-to-finish (WTF) herds across 10 companies located in medium to high pig dense areas in the U.S. Midwest. All herds received weaned pigs from PRRSV-negative or positive-stable breeding herds. Herds were monitored monthly using oral fluids collected following a fixed spatial sampling regime and samples were tested by PRRSV ELISA, RT-PCR and ORF5 sequencing. In most (90%) of the herds, pigs were vaccinated with PRRSV modified-live vaccines either at processing, weaning or shortly after weaning. Wild type PRRSV (WT-PRRSV) infections were defined by the criterion of having more than 2% nucleotide differences in the ORF-5 region compared with reference vaccine strain sequences. Wild type PRRSV was detected in 42% of the herds with infections being more prevalent in the mid to late growing period, with a mean of 20 weeks post placement. Nineteen distinct WT-PRRSV were identified in seven out of 10 production companies with an average of 3 distinct WT-PRRSV strains per company. Vaccinated WTF herds with and without WT-PRRSV detection were compared to each other showing different PCR and ELISA infection patterns. Close-out mortality in vaccinated herds with WT-PRRSV was numerically higher (6.5%) than mortality in those sites where WT-PRRSV was not detected (5.0%) (p = 0.07). Mortality was also higher (10.5%) when WT-PRRSV was detected earlier at eight weeks post-placement compared to late finishing at 20 and 25 weeks post-placement, 2.9% and 4.5% respectively (p = 0.017). Overall, this study sheds light on WT-PRRSV infection dynamics in vaccinated populations of growing pigs, reinforces the importance of biosecurity practices in this phase of production and calls for better understanding of risk factors associated with PRRSV introductions in growing pig sites.

Control technologies to prevent aerosol-based disease transmission in animal agriculture production settings: a review of established and emerging approaches.

Transmission of infectious agents via aerosols is an ever-present concern in animal agriculture production settings, as the aerosol route to disease transmission can lead to difficult-to-control and costly diseases, such as porcine respiratory and reproductive syndrome virus and influenza A virus. It is increasingly necessary to implement control technologies to mitigate aerosol-based disease transmission. Here, we review currently utilized and prospective future aerosol control technologies to collect and potentially inactivate pathogens in aerosols, with an emphasis on technologies that can be incorporated into mechanically driven (forced air) ventilation systems to prevent aerosol-based disease spread from facility to facility. Broadly, we find that control technologies can be grouped into three categories: (1) currently implemented technologies; (2) scaled technologies used in industrial and medical settings; and (3) emerging technologies. Category (1) solely consists of fibrous filter media, which have been demonstrated to reduce the spread of PRRSV between swine production facilities. We review the mechanisms by which filters function and are rated (minimum efficiency reporting values). Category (2) consists of electrostatic precipitators (ESPs), used industrially to collect aerosol particles in higher flow rate systems, and ultraviolet C (UV-C) systems, used in medical settings to inactivate pathogens. Finally, category (3) consists of a variety of technologies, including ionization-based systems, microwaves, and those generating reactive oxygen species, often with the goal of pathogen inactivation in aerosols. As such technologies are typically first tested through varied means at the laboratory scale, we additionally review control technology testing techniques at various stages of development, from laboratory studies to field demonstration, and in doing so, suggest uniform testing and report standards are needed. Testing standards should consider the cost-benefit of implementing the technologies applicable to the livestock species of interest. Finally, we examine economic models for implementing aerosol control technologies, defining the collected infectious particles per unit energy demand.

Assessing the role of sow parity on PRRSv detection by RT-qPCR through weekly processing fluids monitoring in breeding herds.

The use of processing fluids to monitor the breeding herd's porcine reproductive and respiratory syndrome (PRRS) status has gained industry acceptance. However, little is known about PRRS virus RT-qPCR detection dynamics in processing fluids and factors that may contribute to maintain PRRS virus in the herd after an outbreak. This study aimed to describe weekly RT-qPCR processing fluid results in breeding herds after an outbreak and to evaluate the proportion of RT-qPCR positive results among parity groups. Processing tissues of 15 first parity (P1), 15 second parity (P2), and 15 third parity or higher (P3+) litters (parity groups) were collected weekly for between 19 and 46 weeks in nine breeding herds. Processing fluids were aggregated, and RT-qPCR tested by parity group weekly. Additionally, a subset of 743 processing fluid samples of litters that formed 50 parity groups, as previously described, were RT-qPCR tested individually at the litter level. The agreement between RT-qPCR results of processing fluid samples of parity groups (15 litters) and results based on individual litter testing was assessed using overall percent of agreement, Kappa statistic, and McNemar test. The association between RT-qPCR results and the parity group was evaluated using a generalized estimating equations model, after accounting for the effects of sampling week, breeding herd PRRS control strategy (i.e., open to replacements v/s closed) and herd. An autoregressive correlation structure was used to account for the repeated samplings within a herd in time. The overall agreement was 98 %, and Kappa statistic 0.955 (McNemar p = 1.0). Sensitivity of parity group processing fluid samples was estimated at 100 % (95 % CI 89-100 %), while specificity was estimated at 94 % (95 % CI 71-100 %). Although P1 aggregated litters had on average a higher proportion of RT-qPCR positive results from outbreak week 25 onwards, the proportion was not significantly different to the one observed for P2 and P3+ aggregated litters (p > 0.13). Additionally, herds that interrupted gilt entry had lower odds of PRRS RT-qPCR positivity than herds that continued entering gilts (OR = 0.35, 95 % CI 0.16-0.78). PRRS virus persistence in processing fluids was not affected by the sow parity effect in most of the breeding herds studied. No evidence of disagreement between RT-qPCR results of an aggregated sample of 15 litters and those of individual litters was observed. This level of litter aggregation testing strategy may be of particular use at the last stages of an elimination program under low PRRS virus prevalence.

Porcine Reproductive and Respiratory Syndrome (PRRSV2) Viral Diversity within a Farrow-to-Wean Farm Cohort Study.

Describing PRRSV whole-genome viral diversity data over time within the host and within-farm is crucial for a better understanding of viral evolution and its implications. A cohort study was conducted at one naïve farrow-to-wean farm reporting a PRRSV outbreak. All piglets 3-5 days of age (DOA) born to mass-exposed sows through live virus inoculation with the recently introduced wild-type virus two weeks prior were sampled and followed up at 17-19 DOA. Samples from 127 piglets were individually tested for PRRSV by RT-PCR and 100 sequences were generated using Oxford Nanopore Technologies chemistry. Female piglets had significantly higher median Ct values than males (15.5 vs. 13.7, Kruskal-Wallis p < 0.001) at 3-5 DOA. A 52.8% mortality between sampling points was found, and the odds of dying by 17-19 DOA decreased with every one unit increase in Ct values at 3-5 DOA (OR = 0.76, 95% CI 0.61-0.94, p = 0.01). Although the within-pig percent nucleotide identity was overall high (99.7%) between 3-5 DOA and 17-19 DOA samples, ORFs 4 and 5a showed much lower identities (97.26% and 98.53%, respectively). When looking solely at ORF5, 62% of the sequences were identical to the 3-5 DOA consensus. Ten and eight regions showed increased nucleotide and amino acid genetic diversity, respectively, all found throughout ORFs 2a/2b, 4, 5a/5, 6, and 7.

Reassortant influenza A viruses in wild duck populations: effects on viral shedding and persistence in water.

Wild ducks of the genus Anas represent the natural hosts for a large genetic diversity of influenza A viruses. In these hosts, co-infections with different virus genotypes are frequent and result in high rates of genetic reassortment. Recent genomic data have provided information regarding the pattern and frequency of these reassortant viruses in duck populations; however, potential consequences on viral shedding and maintenance in the environment have not been investigated. On the basis of full-genome sequencing, we identified five virus genotypes, in a wild duck population in northwestern Minnesota (USA), that naturally arose from genetic reassortments. We investigated the effects of influenza A virus genotype on the viral shedding pattern in Mallards (Anas platyrhynchos) and the duration of infectivity in water, under different temperature regimens. Overall, we found that variation in the viral genome composition of these isolates had limited effects on duration, extent and pattern of viral shedding, as well as on the reduction of infectivity in water over time. These results support that, in wild ducks, functionally equivalent gene segments could be maintained in virus populations with no fitness costs when genetic reassortments occur.

Evidence of influenza A infection and risk of transmission between pigs and farmworkers.

Interspecies transmission of influenza A virus (IAV) between pigs and people represents a threat to both animal and public health. To better understand the risks of influenza transmission at the human-animal interface, we evaluated 1) the rate of IAV detection in swine farmworkers before and after work during two human influenza seasons, 2) assessed risk factors associated with IAV detection in farmworkers and 3) characterized the genetic sequences of IAV detected in both workers and pigs. Of 58 workers providing nasal passage samples during 8-week periods during the 2017/18 and 2018/19 influenza seasons, 33 (57%) tested positive by rRT-PCR at least once. Sixteen (27%) workers tested positive before work and 24 (41%) after work. At the sample level, 58 of 1,785 nasal swabs (3.2%) tested rRT-PCR positive, of which 20 of 898 (2.2%) were collected prior to work and 38 of 887 (4.3%) after work. Although farmworkers were more likely to test positive at the end of the working day (OR = 1.98, 95% CI 1.14-3.41), there were no influenza-like illness (ILI) symptoms, or other risk indicators, associated with IAV detection before or after reporting to work. Direct whole-genome sequencing from samples obtained from worker nasal passages indicated evidence of infection of a worker with pandemic 2009 H1N1 of human-origin IAV (H1-pdm 1A 3.3.2) when reporting to work, and exposure of several workers to a swine-origin IAV (H1-alpha 1A 1.1) circulating in the pigs on the farm where they were employed. Our study provides evidence of 1) risk of IAV transmission between pigs and people, 2) pandemic H1N1 IAV infected workers reporting to work and 3) workers exposed to swine harbouring swine-origin IAV in their nasal passages temporarily. Overall, our results emphasize the need to implement surveillance and transmission preventive protocols at the pig/human interface.

Detection of Multiple Lineages of PRRSV in Breeding and Growing Swine Farms.

The detection and co-circulation of multiple variants of porcine reproductive and respiratory syndrome virus (PRRSV) have been observed and reported in swine. However, the potential long-term impact of multiple prevailing PRRSV variants on pig-performance is not yet fully understood. The primary objective of this study was to describe the genetic variation of PRRSV in processing fluid (PF), oral fluid (OF), and tonsil scraping (TS) specimens from five swine farms with different production types and PRRS status over a period of time (~1 year). Furthermore, the association between PRRSV prevalence and production parameters was investigated. Results showed that PRRSV was detected by RT-qPCR in 21-25% of all types of specimens. In breeding farms, PRRSV detection in PF and/or TS samples was correlated with stillborn and mummified fetuses, and pre-weaning mortality throughout the study period. Although ORF5 sequences were obtained in <16% of all sample types, simultaneous detection of PRRSV variants including field and vaccine strains within a single sampling event was identified in both breeding and growing pig farms. Phylogenetic analyses based on the ORF5 sequence classified the detected field PRRSV into L1A and L1H, two sub-lineages of lineage 1 (L1). Our study demonstrated the presence of multiple PRRSV lineages, sub-lineages, and variants in swine herds and its potential association with swine reproductive performance under field conditions.

Effect of pooling udder skin wipes on the detection of influenza A virus in preweaning pigs.

Influenza A virus (IAV) active surveillance in pigs prior to weaning is commonly conducted by collecting individual samples, mostly nasal swabs. Recently, the use of udder skin wipes collected from lactating sows was identified as an effective sampling method to indicate IAV status of suckling piglets prior to weaning. However, there is limited information on the effect of pooling multiple udder wipes on the ability to detect IAV. We evaluated the effect of pooling 3, 5, or 10 udder wipes on the sensitivity of detecting IAV and compared the results with testing the wipes individually. The likelihood of detecting positive udder wipes decreased with pooling when the initial positive cycle threshold value was ≥31.5; pooling of up to 3 samples could be performed without affecting sensitivity significantly. Our results support pooling of udder skin wipes to conduct surveillance of IAV in pigs prior to weaning.

Comparison of samplers collecting airborne influenza viruses: 1. Primarily impingers and cyclones.

Researchers must be able to measure concentrations, sizes, and infectivity of virus-containing particles in animal agriculture facilities to know how far infectious virus-containing particles may travel through air, where they may deposit in the human or animal respiratory tract, and the most effective ways to limit exposures to them. The objective of this study was to evaluate a variety of impinger and cyclone aerosol or bioaerosol samplers to determine approaches most suitable for detecting and measuring concentrations of virus-containing particles in air. Six impinger/cyclone air samplers, a filter-based sampler, and a cascade impactor were used in separate tests to collect artificially generated aerosols of MS2 bacteriophage and swine and avian influenza viruses. Quantification of infectious MS2 coliphage was carried out using a double agar layer procedure. The influenza viruses were titrated in cell cultures to determine quantities of infectious virus. Viral RNA was extracted and used for quantitative real time RT-PCR, to provide total virus concentrations for all three viruses. The amounts of virus recovered and the measured airborne virus concentrations were calculated and compared among the samplers. Not surprisingly, high flow rate samplers generally collected greater quantities of virus than low flow samplers. However, low flow rate samplers generally measured higher, and likely more accurate, airborne concentrations of Infectious virus and viral RNA than high flow samplers. To assess airborne viruses in the field, a two-sampler approach may work well. A suitable high flow sampler may provide low limits of detection to determine if any virus is present in the air. If virus is detected, a suitable lower flow sampler may measure airborne virus concentrations accurately.

Phylogenetically Distinct Near-Complete Genome Sequences of Porcine Reproductive and Respiratory Syndrome Virus Type 2 Variants from Four Distinct Disease Outbreaks at U.S. Swine Farms over the Past 6 Years.

Porcine reproductive and respiratory syndrome virus (PRRSV) continues to mutate, causing disruptive PRRS outbreaks in farms that lead to reproductive failure and respiratory disease-associated mortality. We present four new PRRSV type 2 variants in the United States belonging to four distinct orf5 sublineages within lineage 1.

Immune responses to porcine epidemic diarrhea virus (PEDV) in swine and protection against subsequent infection.

Understanding the immune responses against Porcine epidemic diarrhea virus (PEDV) is important to prevent infection and to design control strategies. We evaluated both systemic and mucosal immune responses to PEDV in pigs and assessed if prior exposure to virus protects against re-infection. Three-week-old pigs were infected with PEDV and immune response in blood, intestine, and mesenteric lymph node (MLN) was evaluated. At 30 dpi, virus exposed pigs were challenged with a field isolate of PEDV and immune response at 5 d post challenge was evaluated. We found that PEDV RNA persists in the intestine even after fecal shedding of the virus was stopped at 28 dpi and pigs previously exposed to PEDV are protected from virus shedding after re-infection. PEDV infection induced both humoral and cell mediated immune response with an increase in PEDV specific IgA and IgG antibodies in intestine and serum. Flow cytometry analysis showed a significantly higher frequency of B cells and lower frequency of T cells at 4 dpi. The frequency of CD4/CD8 double positive (DP) memory T cells was significantly increased in the MLN of challenged animals. These studies may provide further insights into understanding the mucosal immune response to PEDV and its role in protection against disease.