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Classical swine fever (CSF) is one of the most important viral diseases in swine, causing severe economic losses in the swine industry. In China, CSF is one of the key diseases that needs to be controlled; the government has implemented control measures, and vaccination with C-strain vaccines (C-vacs) has been compulsory since the 1950s. C-vacs do not allow the differentiation of field virus-infected and vaccinated animals (DIVA). In 2012, China proposed a goal of eradicating CSF. Additionally, a baculovirus-expressed E2 subunit vaccine (E2-vac) was licensed in 2018. However, the C-vac and E2-vac characteristics have not been compared. Here, we demonstrate that both the C-vac and E2-vac provide complete protection against CSF in pigs. The E2-vac allows DIVA, and the E2 antibody responses of stimulated pigs are developed earlier and are stronger than the C-vac antibody responses. Therefore, the E2-vac is a new candidate licensed vaccine to completely eradicate CSF on pig farms. [ABSTRACT FROM AUTHOR]

African Swine Fever (ASF) is an infectious lethal disease affecting domestic pigs and wild boar. In the EU the infection perpetuates predominantly in wild boar populations. ASF control comprises wild boar population reduction measures,e.g. pre‐emptive culling in delineated zones, called white zones (WZ).TheseWZ areplaced geographically adjacent to an area with ASF circulating in wild boar (ASF positive area).The ideal WZ would be depopulated of wild boar without possibility of recolonization. However, WZ may still harbour live wild boar after its establishment and the functionality of the WZ inherently foresees ASF enteringit. But the spread of the infection is expected tostop within an effective WZ. The concept mustnotmatch legislative zones, likewise infected area, Part I, Part II, etc. In order to compare different approaches to implement a WZ(e.g. targets and speed of population reduction in the WZ, width of the WZ, and distance of the WZ from the ASF‐positive area), an individual‐based spatially explicit model was adjusted to four historic WZ‐like situations in the EU, i.e. Estonia 2014, Latvia 2016, Czech Republic 2017, and France 2018. The model was used to simulate the reported spatio‐temporal layout and targeted measures. The stochasticity of the model provided understanding of the general efficiency of these WZ. Alternatives of the local measures were simulated as scenarios to identify caveats of the settings and derive improvements in future applications. The simulation outcome suggests issues to be addressed in implementing future WZ: i) distance between ASF‐positive area and the WZ was adequate if adapted to the speed of propagation according to the local wild boar density, and the time horizon of depopulation measures envisaged for the WZ; ii) the width of the WZ was adequately set if everywhere it was preventedthat short infection chains already led out of the zone, iii) the WZ around focal introductions was most efficient if depopulated by culling the maximum of a defined (or fenced) population in shortest time with minimal disturbance, for instance, by trapping, sharp shooting or using silencers.Aspects of density, timing and spatial distribution in relation to the efficiency of WZ layout are explored. [ABSTRACT FROM AUTHOR]

Porcine reproductive and respiratory syndrome virus (PRRSV) is a notable threat to the pig industry. Long-term epidemiological investigations and genetic variation analyses of PRRSV isolates benefit PRRSV prevention and control. In our study, 43 PRRSV strains were successfully isolated from the lungs of sick pigs, and the genetic variations of these isolates were analyzed. Phylogenetic analysis showed that the isolates belonged to PRRSV2 and that lineage 8 (8.7) subgroup III strains remained the dominant type circulating in South China. In addition, sequence alignment analysis identified many novel deletions and mutations in the Nsp2 and GP5 genes. Furthermore, phylogenetic analysis showed that highly frequent recombination events of PRRSV between different lineages might occur in Guangdong Province. These results may help to elucidate the epidemiology and genetic variation of PRRSV isolates in Guangdong Province. [ABSTRACT FROM AUTHOR]

Antecedentes: la colibacilosis clasifica entre las enfermedades catastróficas en las producciones porcinas. Pese a las numerosas investigaciones encaminadas a su diagnóstico, prevención y control durante años no ha logrado erradicarse. Objetivo. Analizar cuatro elementos que favorecen la persistencia de la enfermedad en Camagüey. Desarrollo: se revisaron 115 fuentes especializadas. En base a las normas editoriales de la Revista de Producción Animal se seleccionaron 50 como referencias, para lo cual el elemento de discriminación fue el objetivo propuesto. Las referencias, por su actualidad, correspondieron a tres grupos: 19 (último quinquenio), 11 (2000-2010) y 20 (anteriores al 2000). En el período 1985-2005 se prestó mayor atención a la colibacilosis en la provincia, como confirman la vacuna VACOLI, el desarrollo de diagnosticadores, entre los que destaca AuBIODOT-ECET, y antidiarreicos de origen vegetal sustentables como el Eucabev. A finales de la etapa se acrecienta: la subvaloración de la enfermedad, el desconocimiento del agente etiológico, el detrimento en los métodos diagnósticos; las medidas preventivas y de control se enfocan al efecto, no a la causa de la enfermedad. Conclusiones: cuatro elementos coadyuvan a que las colibacilosis porcinas en crías persistan en Camagüey con las consecuentes pérdidas económicas: subvaloración de la enfermedad; desconocimiento de los patotipos de E. coli involucrados; ausencia de sistemas de diagnóstico precisos y rápidos y la aplicación de medidas preventivas y de control inadecuadas. El empleo de prebióticos, probióticos, o simplemente microorganismos autóctonos multipropósito, ayudaría a reducir el impacto negativo de esta enfermedad. [ABSTRACT FROM AUTHOR]

Recent studies have identified a new genus of the Orthomyxoviridae family, Influenza D virus (IDV). This virus was shown to infect farm animals including swine and cattle, and to efficiently replicate and transmit in ferrets, the animal model of choice for transmission of influenza A virus to humans. This partnering grant on IDV addressed the need for capacity building at EU level to improve the EU's scientific assessment capacity and international competitiveness. We have promoted cross‐disciplinary cooperation between the partner institutes representing six Member States (BE, FR, IT, LU, NL and SE). We have shown that the available antigen and genome test systems allow reliable influenza D diagnostics in partners laboratories, while for a few of the applied antibody testing methods adjustments are recommended. Tools were developed to study virus‐host range, with a gain of knowledge on host and tissue tropism of IDV in farm animals but also in wild life and very preliminary data was generated on human tissues. Serological results in European cattle suggest that influenza D virus is enzootic. Virus diversity is still unfolding: new virus introductions were identified, as well as new reassortants whose differential clinical impact or cross‐protection levels are still poorly understood. Considering drivers of emergence, IDV was in the top five in comparison with 29 other diseases. The main risk factors of IDV in cattle are related to the animal density, presence of respiratory clinical signs in cattle and contact rates between animals. Simplified quantitative IDV risk assessment exposure model indicated a possible infection of human by IDV through aerosols in cattle farms. Further studies are warranted to fully assess the risk of IDV for both animal and Human health in Europe. [ABSTRACT FROM AUTHOR]

To study the influence of vaccinations against porcine cycrovirus 2 (PCV2) by vaccines of two different manufacturers on leukogram and level of specific antibodies in blood of pigs at the Pavlovsky Pork Complex of the Bryansk Oblast 2 groups of 15 heads of weaning pigs of 21 day old were formed. Inactivated vaccines were injected intramuscularly into the neck region: animals of group 1 - Suvaxyn PCV2, pigs of group 2 - Ingelvac CircoFLEX. Blood was taken from 15 animals from each group from the femoral vein in the morning before feeding: for morphological study - before the beginning of the experiment and further every 14-15 days up to 125 days after vaccination; for determination of antibodies titer - 21, 60, 90 and 120 days after vaccination. It has been established that the mechanism of cellular antiviral protection is expressed to the greatest extent when using the Ingelvac CircoFLEX vaccine, and the highly specific humoral protection and higher reactivity of pigs' organism in relation to technological stress in a remote period after vaccination - after application of Suvaxyn PCV2. [ABSTRACT FROM AUTHOR]

Porcine epidemic diarrhoea (PED) is a highly contagious and devastating enteric disease of pigs caused by porcine epidemic diarrhoea virus (PEDV), an enveloped, single-stranded RNA virus belonging to the Alphacoronavirus genus of the Coronaviridae family. The disease is clinically similar to other forms of porcine gastroenteritis. Pigs are the only known host of the disease, and the occurrence of PED in wild boars is unknown. The virus causes acute diarrhoea, vomiting, dehydration, and high mortality in suckling piglets reaching 100%. Heavy economic losses in the pig-farming industry were sustained in the USA between 2013 and 2015 when PEDV spread very quickly and resulted in epidemics. The loss in the US pig industry has been estimated at almost seven million pigs. The purpose of this review is a description of the current status of porcine epidemic diarrhoea in European pigs and the risk presented by the introduction of PEDV to Poland in comparison to the epidemics in the USA. [ABSTRACT FROM AUTHOR]

Defatting of seabass skins using pulsed electric field (PEF)‐assisted process at different electric field strengths (16 and 24 kV/cm) and times (36, 72, and 108 ms) in combination with porcine pancreas lipase (PPL) at 25 U/g dry matter was investigated. PEF‐treated skin at 24 kV/cm for 72 ms followed by PPL treatment removed 86.93% lipids. PEF‐treated skin was further optimized for lipid reduction by response surface methodology. Central composite design was adopted to establish treatments based on two independent variables, involving PPL concentration (30 to 55 U/g dry matter) and hydrolysis time (60 to 180 min). Second‐order polynomial model was used for predicting the response. The highest lipid removal (91.96 ± 1.70%) was attained when the optimal condition (42.36 PPL units/g dry skin matter for 139.78 min) was used. The experiment value was in accordance with the predicted value. PEF‐PPL‐treated skin had lower monounsaturated and polyunsaturated fatty acids than the solvent‐extracted skin (P < 0.05). When PEF‐PPL‐treated skin was hydrolyzed using papain at 0.30 U/g dry matter, lower fishy odor/flavor of resulting hydrolyzed collagen (PEF‐PPL‐HC) was found than other samples (P < 0.05). Lower total volatile compounds were also obtained in PEF‐PPL‐HC sample. Thus, the use of PEF pretreatment along with PPL before papain hydrolysis effectively prevented the formation of fishy odor/flavor in hydrolyzed collagen from seabass skin. Practical Application: Fishy odor/flavor caused by lipid oxidation of fish skin hydrolysates limits their applications in foods. Defatting process is the significant step for skin pretreatment. Although several methods could remove lipids from fish skins, either by lipase or solvent extraction, fishy odor/flavor is still detected in hydrolysate. Pulsed electric field‐assisted process in combination with porcine lipase is another approach that can be used to enhance efficiency via electroporation, causing the loosened skin matrix and facilitating the migration of lipase into the skin. Consequently, the resulting hydrolysate might have the lowered fishy odor/flavor and could be used in foods, especially for fortification. [ABSTRACT FROM AUTHOR]

Highlights • PAMs from young pigs are more susceptible to PRRSV infection than adult PAMs. • PRRSV receptor expression was similar between PAMs from young and adult pigs. • Age-dependent resistance to PRRSV infection is not due to receptor-mediated entry. Abstract Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) is the most economically important disease affecting swine production worldwide. The severity and susceptibility of PRRSV infection varies with age. Nursery pigs have been shown to be more susceptible to PRRSV infection and a more severe and prolonged infection is observed as compared to growing or adult pigs. However, antibody responses to PRRSV are observed independent of age. Swine are the only known hosts of PRRSV, infection is restricted to cells of monocytic lineage, and fully differentiated porcine alveolar macrophages are the primary target of natural infection. Pulmonary intravascular macrophages from young pigs have been shown to be more susceptible to infection than those from adult pigs. A better understanding of why young pigs and macrophages from young pigs are more susceptible to PRRSV infection is critical to identify mechanisms of infection that can be explored for enhanced treatment or prevention of disease. This study examined PRRSV susceptibility of porcine alveolar macrophages isolated from the lungs of pigs of different age groups, and the presence of cell surface receptors to determine if differences correlated with infection level. The younger the pigs were, the more susceptible the macrophage were to PRRSV infection, but no differences in cellular receptor expression were observed between pigs of different ages. Resistance to infection is likely related to intracellular innate immune mechanisms rather than receptor-mediated entry. [ABSTRACT FROM AUTHOR]

Highlights • Ivermectin inhibits NLS-mediated nuclear location of protein Cap. • Ivermectin inhibits PCV2 replication in PK-15 cells. • Ivermectin treatments mitigate impact of PCV2 infection in piglets. Abstract Porcine circovirus 2 (PCV2) capsid protein (Cap) has a nuclear localization signal (NLS) and can enter the nucleus. In this study, ivermectin, a small-molecule nuclear import inhibitor of proteins was used to determine the role of nuclear localization of Cap on PCV2 replication. Observation by fluorescence microscopy of the intracellular localization of Cap and Cap NLS in cells cultured with ivermectin (50 μg/mL) determined that Cap and Cap NLS were located in the cytoplasm; in contrast, for cells cultured without ivermectin, they accumulated in the cell nucleus. Ivermectin treatment also reduced nuclear transport of Cap derived from PCV2 infection as well as PCV2 replication in PK-15 cells. In addition, lower levels of PCV2 in tissues and sera of piglets treated with ivermectin were detected by qPCR. These results established for the first time that ivermectin has potent antiviral activity towards PCV2 both in vitro and vivo. [ABSTRACT FROM AUTHOR]

Abstract Classical swine fever (CSF) continues to be a devastating infectious disease for the swine industry in China and commonly exists as wild or atypical types. From June 3rd to October 3rd, 2018, outbreaks of typical CSF cases with mortality rates of 42–86% occurred in 11 swine herds in five cities of Guangdong province, and were confirmed by RT-PCR. Phylogenetic analyses based on the nucleotide sequences of full-length E2 genes showed that the CSFV isolates collected in Guangdong, 2018 grouped into sub-subgenotype 2.1c and formed a separate clade from previously identified 2.1c isolates. Sequence comparison further confirmed the distance between the novel emergent and previously identified 2.1c isolates, with shared 94.5–98.2% and 97.8–99.7% identities at the nucleotide and amino acid levels respectively. Furthermore, 2.1c isolates collected in 2018 from Guangdong province contained a unique amino acid substitution (K174R) in the E2 protein in comparison with other 2.1c representative strains and CSFV 2.1, 2.2, 2.3 strains. Of note, the novel emergent 2.1c isolates are neutralized by sera from C-strain vaccinated sows, indicating that C-strain is still efficacious for protection against field isolates of CSFV. Highlights • CSFV sub-subgenotype 2.1c is dominant in Guangdong province. • The novel identified 2.1c isolates contained a unique substitution K174R in the E2 proteins. • The emergent 2.1c isolates can still be neutralized by sera from vaccinated pigs. [ABSTRACT FROM AUTHOR]

Abstract Hepatitis E virus (HEV) is an important pathogen causing public health burden. Swine has been recognized as a main reservoir. Interestingly, genotype 1 HEV only infects human; whereas genotype 3 and 4 are zoonotic. However, there is a lack of in-depth understanding in respect to the transmission from swine to human. Codon usage patterns generally participate in viral survival and fitness towards its hosts. We have analyzed codon usage patterns of the three open reading frames (ORFs) for 243 full-length genomes of HEV genotypes 1, 3 and 4. The divergence of synonymous codon usage patterns is different in each ORF for genotypes 1, 3 and 4, but the genotype-specific codon usage bias in genotype 1 is stronger than those of genotypes 3 and 4. In respect to genotypes 3 and 4, compared with strains isolated from human, HEV isolated from swine shows appreciable variation in adaptation of codon usages to human or swine. These results may help to understand the transmission and host adaptation of HEV genotypes 3 and 4 from swine to human. Highlights • Nucleotide usage bias at the third codon position is different from genotype 1 to genotypes 3 and 4. • Codon usage bias of HEV genotype 1 is significant different from those of genotypes 3 and 4. • Diversity of synonymous codon usages may help the transmission from swine to human. [ABSTRACT FROM AUTHOR]

Recently, emerging porcine parvovirus species (PPVs) have been identified. These species have designated as PPV2 to PPV8. At present, PPV5 and PPV8 remained unclassified by ICTV. The criteria for PPV taxonomy utilize the amino acid (aa) sequence of NS1 which is a multifunctional protein essential for viral replication. In this scenario, a PPV member is ascribed to the same genre and species if it shares 35–45 % and ≥ 85 % identity of NS1, respectively. To perform a NS1 molecular analysis, three PPV5+ DNA samples were used to achieve the complete ORF1 lineal nucleotide sequence via amplification of three overlapping PCR products. At editing, a NS1 lineal sequence of 1750 nucleotides comprising 96.9 % of ORF1 was obtained from each DNA sample. The aa matrices showed that the Mexican sequences have a 96.7–99.8 % identity with previously reported PPV5 sequences, and high similarity with PPV4 (76.0−87.3 %) and PPV6 (54.9−55.4 %) sequences. The analysis of aa sequences pointed at five conserved motifs and arginine fingers which are portrayed for superfamily 3 (SF3) helicases. The current work findings suggest that PPV5 might belong to the Copiparvovirus genre due to its proximity to PPV4 and PPV6 and that NS1. Most likely, NS1 is a SF3 helicase. [ABSTRACT FROM AUTHOR]

Porcine Reproductive and Respiratory Syndrome (PRRS) is a highly significant disease due to the economic losses it causes globally. PRRS affects pigs of all ages, primarily causing reproductive failure in sows and respiratory disease in piglets. The etiological agent, PRRS virus (PRRSV), belongs to the order Nidovirales, family Arteriviridae, genus Porartevirus, with a positive-sense single-stranded RNA genome of 15 kb. The glycoprotein GP5 is encoded in the ORF5 gene, whose sequence is commonly used to establish phylogenetic relationships and classify PRRSV isolates. During routine monitoring of various swine production units in Mexico, a total of 5,840 samples, mainly serum/whole blood, oral fluids, and lung tissue, were analyzed for PRRSV diagnosis using real-time RT-PCR, resulting in an average positivity of 18.51 %. Positive samples were selected for Sanger sequencing of the ORF5 gene. The obtained sequences were aligned, and a maximum likelihood phylogenetic tree was constructed. The circulating PRRSV strains in Mexico were found to belong to lineages 1 (57 %) and 5 (43 %). The predominant RFLP pattern within lineage 1 was 1-7-4 type, while in lineage 5, 2-5-2 type. These findings shed light on the genetic diversity of PRRSV circulating in Mexico and provide valuable insights for the surveillance and control of the disease. [ABSTRACT FROM AUTHOR]

Porcine hemagglutinating encephalomyelitis virus (PHEV) is a highly neurovirulent coronavirus that invades the central nervous system in piglets. The incidence of PHEV among pigs in many countries is rising, and the economic losses to the pig industry may be significant. Serological studies suggest that PHEV is spread worldwide. However, no surveillance has been carried out in the Czech Republic. In this study, eight pig farms were screened for the presence of members of the Coronaviridae family with the use of reverse transcription PCR. A collection of 123 faecal samples and 151 nasal swabs from domestic pigs were analysed. In PHEV-positive samples, almost the complete coding sequence of the nucleocapsid gene was amplified and the acquired sequences were compared to those of geographically dispersed PHEV strains; phylogenetic analyses were also performed. PHEV was present in 7.9% of nasal swabs taken from different age categories of pigs. No other swine coronaviruses were detected. The amino acid sequence of the Czech PHEV strains showed 95.8-98.1% similarity to other PHEV reference strains in GenBank. PHEV strains collected from animals on the same farm were identical; however, strains from different farms have only exhibited only 96.7-98.7% amino acid sequence identity. Our study demonstrates the presence of PHEV in pigs in the Czech Republic. The Czech PHEV strains were evolutionarily closest to the Belgium strain VW572. [ABSTRACT FROM AUTHOR]

The alphaherpesvirus pseudorabies virus (PRV) is the causative agent of pseudorabies, a disease of great economic and welfare importance in swine. Other alphaherpesviruses, including herpes simplex virus (HSV), utilize low pH-mediated endocytosis to enter a subset of cell types. We investigated whether PRV used this entry pathway in multiple laboratory model cell lines. Inhibition of receptor-mediated endocytosis by treatment with hypertonic medium prevented PRV entry. PRV entry into several cell lines including porcine kidney (PK15) cells, and African green monkey kidney (Vero) cells was inhibited by noncytotoxic concentrations of the lysosomotropic agents ammonium chloride or monensin, which block the acidification of endosomes. Inactivation of virions by acid pretreatment is a hallmark of viruses that utilize a low pH-mediated entry pathway. Exposure of PRV virions to pH 5.0 in the absence of host cell membrane reduced entry into PK15 and Vero cells by greater than 80%. Together, these findings suggest that endocytosis and subsequent fusion with host membranes triggered by low endosomal pH is an important route of entry for PRV. [ABSTRACT FROM AUTHOR]

Porcine reproductive and respiratory syndrome virus (PRRSV) is an economically important pathogen that affects the global swine industry. The continuous evolution of this virus has made control and prevention difficult, which emphasizes the importance of monitoring currently circulating PRRSV strains. In this study, we investigated the genetic characteristics of whole structural genes of 35 PRRSV-2 isolates that circulated between 2012 and 2017 in Korea. Genetic and phylogenetic analysis demonstrated that a recently identified PRRSV-2 shared a relatively low level of nucleotide sequence identity that ranged from 86.2% to 92.8%; however, they were clustered into four distinct Korean field clades, except KU-N1702, in ORF2-7-based phylogeny. KU-N1702 was closely related to the NADC30-like strains that were identified in the USA and China. Amino acid sequence analysis showed that the GP5 neutralizing epitope was conserved among the KU viruses. In contrast, the viruses had genetic mutations in key residues for viral neutralization within GP5 and M. For minor structural proteins, neutralizing epitopes, aa 41-55 of GP2, 61-75 of GP3, and 51-65 of GP4, were variable among the KU viruses. Bioinformatics demonstrated diversifying evolution within the GP2 and GP4 neutralizing epitopes and the emergence of a novel glycosylation site within the GP3 and GP4 neutralizing epitopes. Taken together, these data provide evidence that Korean PRRSV-2 evolved independently in Korea, with genetic heterogeneity in antigenic regions of structural proteins. [ABSTRACT FROM AUTHOR]

This case study report describes a transmissible gastroenteritis coronavirus (TGEV) infection presented in a commercial pig herd. The clinical signs of infection appeared in newborn piglets, including medium morbidity and low mortality rates. Rectal swabs were collected from five different affected litters for laboratory examinations. Samples from two dead piglets and two euthanized affected piglets were collected for gross and histopathological examinations. All fecal samples were tested TGEV positive by real-time polymerase chain reaction (RT-PCR). Necropsy revealed nonspecific gross lesions. The histopathological examinations revealed villi fused with denuded tips and severe villus atrophy, leading to extensive epithelial flattening in middle and lower small intestine. The architecture pattern of villi presented columnar and cuboidal poorly differentiated enterocytes with mild subepithelial edema. In some enterocytes, pycnotic nuclei were detected. Microscopic examination of brain tissue revealed diffuse gliosis in the area of pia matter with mild congestion of the meningeal and parenchymal vessels and neuronal degeneration. In conclusion, this case study reported an epidemic TGEV infection in piglets, characterized by low mortality and medium morbidity rates accompanied by typical histopathological lesions in small intestine, as well as by coexisting brain lesions, that are described for the first time. [ABSTRACT FROM AUTHOR]

The phospholipase C (PLC) is a family of kinases that hydrolyze phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] to generate two second messengers, inositol 1,4,5-trisphosphate (IP3) and diacylglycerol (DAG), which stimulate distinct downstream signaling. Recently, it has been reported that PLC signaling is activated by multiple viruses for efficient replication and the virus-induced inflammatory response. In this study, we demonstrated that PLC-specific inhibitor U73122 strongly suppressed porcine reproductive and respiratory syndrome virus (PRRSV) productive infection in cell cultures. The inhibitor affected both viral post-binding cell entry and post-entry processes. The virus infection led to an early transient activation of PLCγ-1 at 0.5 h post-infection (hpi), and sustained event at a stage from 4 to 16 hpi in MARC-145 cells. In addition, U73122 inhibited the activation of p38 MAPK signaling stimulated by PRRSV infection, suggesting that PLC signaling may be associated with the virus infection-induced inflammatory response. Taken together, these studies suggested that PLC signaling played an important role in PRRSV infection or pathogenesis. [ABSTRACT FROM AUTHOR]

The presence of porcine astroviruses in diarrheic and healthy pigs has been reported, however, the consequences of the astrovirus infection during the weaning process have not been described. In this study, eight healthy conventional newly-weaned piglets were used to evaluate effects of astrovirus infection during the first five days. Four piglets were infected with the porcine astrovirus PoAstV/PUJP5 strain and the other four represented the control group. Body weight, rectal temperature, diarrhea and other clinical signs were monitored every 24 hours. The results showed that all animals gained body weight, the occurrence of mild diarrhea on the 3rd day post-infection, and the astroviral presence in diarrheic samples. On the 5th day post-infection all the piglets were euthanized and then intestinal and extra-intestinal tissues were analyzed for the presence of PoAstV/PUJP5. The cytoplasmic antigen of PoAstV/PUJP5 was observed in the enterocytes of infected piglets from jejunum, ileum, colon and in inflammatory cells from mesenteric lymph nodes. In addition, villi atrophy, fusion, epithelial hyperplasia and incipient virus detection in mesenteric lymph were observed. RNAemia could not be proved. This study shows for the first time the effects of porcine astrovirus infection on conventional newly-weaning piglets. [ABSTRACT FROM AUTHOR]

The main immunogenic protein of the porcine epidemic diarrhea virus (PEDV) is the spike protein (S protein), which plays an important role in receptor binding, membrane fusion, and viral invasion of the host. In this paper, the linear epitope of the 3F10 non-neutralizing monoclonal antibody that was previously prepared in our laboratory was identified. The expression of truncated forms of the S protein for 3F10 reaction studies proved that the epitope was located between amino acids (aa) 674 and 791. To further locate the core aa of the 3F10 epitope, 12 random peptide libraries were used, and the result showed that the key aa located at aa 685-688 of the S protein (LLAF) were recognized by 3F10. Homology analysis of the regions corresponding to 20 typical strains of different PEDV subtypes showed that the epitope is highly conserved. Identifying the epitope recognized by an antibody helps to improve our understanding of the structure and function of the antigen. [ABSTRACT FROM AUTHOR]

Classical swine fever (CSF) is a highly contagious viral infection that affects domestic and wild pig population. The classical swine fever virus (CSFV) targets immune cells which perturb the immune functions causing immunopathological disorders such as immunosuppression, leukopenia and haemorrhage. In the present study, ELISA and quantitative real-time reverse transcription PCR (qRT-PCR) analysis was employed to determine cytokine profiles in pigs naturally infected with CSFV using whole blood assay (WBA) under field conditions. Significantly higher TNF-α, IL-10, and IL-6 expression levels were found in unvaccinated pigs infected with CSFV (group B) compared to vaccinated pigs that recovered after CSF (group A), the difference being statistically significant (p = 0.001). However, the expression of IFN-γ was significantly higher in group A compared to group B (p = 0.001). The findings of this field-supported study will help us to understand the immune biology of CSFV infection in infected pigs. The WBA technique can be used as a reliable, fast and feasible in vitro method to assess porcine cellular immune responses as it imitates the porcine blood conditions. Such studies could be of some value in determining the immune status of the ailing animals infected with CSFV. [ABSTRACT FROM AUTHOR]

Abstract Pseudorabies virus (PRV) is one of the most vital pathogens of swine, leading to huge economic losses to the pig industry. Functioning in innate immunity, type-I interferon (IFN) plays a vital role in initial stage of viral infection. ISG15, an IFN-stimulated ubiquitin-like protein, is highly increased during virus infection and participates in the IFN-mediated antiviral immune response. However, limited attention has been paid to the functional role of porcine ISG15 (pISG15) in PRV infection. In this study, we generated a PK15 inducible cell line stably expressing the pISG15 gene and investigated the potential anti-PRV response of pISG15. We demonstrated that pISG15 was upregulated in an early stage of PRV infection, and pISG15 overexpression efficiently inhibited PRV replication by reducing the viral titers and mRNA levels of PRV, and also increased expression of IFN- β and activation of the ISRE promoter. However, knockdown of pISG15 by siRNA did not affect PRV replication, and potentiated IFN-I-mediated signaling, resulting in an increase in antiviral response in the process of PRV infection. The results showed that pISG15 has a potential immunodulatory role in cellular antiviral response against PRV. Highlights • The PK15 inducible cell line stably expressing the pISG15 gene was constructed to investigate the potential anti-PRV response of pISG15. • pISG15 was upregulated in an early stage of PRV infection. • pISG15 overexpression efficiently inhibited PRV replication and increased expression of IFN- β and activation of the ISRE promoter. • Knockdown of pISG15 did not affect PRV replication and enhanced type I IFN-mediated signaling during PRV infection. [ABSTRACT FROM AUTHOR]

Highlights • APPV-induced infection in newborn piglets produces neuronal necrosis of the brain. • APPV infection induces gliosis, neuronophagia, and satellitosis. • APPV-associated histopathologic findings are predominantly necrotic and degenerative. • The proliferation of GFAP + cells is an important feature of APPV infection. Abstract Atypical porcine pestivirus (APPV) has been associated with congenital tremor (CT) type A-II in newborn piglets. Although the number of APPV-based studies is increasing, the associated pathologic findings in infected piglets are underreported. This study describes the histopathologic features of spontaneous APPV infection in CT-affected piglets and complements a previous report by our group. Four two-day-old piglets with CT were evaluated by histopathology, immunohistochemistry (IHC), and molecular assay. The main histopathologic findings at the brain and spinal cord included neuronal necrosis, gliosis, neuronophagia, satellitosis, demyelination, Wallerian degeneration, and Purkinje cell necrosis. An IHC assay designed to detect the proliferation of glial fibrillary acidic protein (GFAP) in affected areas of the brain and spinal cord revealed that the proliferation of GFAP + cells and fibers was predominant in APPV-infected piglets relative to asymptomatic piglets of the same age group. The RT-nested-PCR assays identified APPV RNA in the cerebrum, cerebellum, and brainstem of all piglets; other viruses known to produce similar manifestations were not detected. These results suggest that the APPV-induced histopathologic findings are predominantly degenerative and necrotic and correlate with our previous findings. Consequently, it is proposed that neuronal necrosis, gliosis, neuronophagia, and satellitosis should be considered as important histologic features of APPV-induced infection in symptomatic CT piglets. [ABSTRACT FROM AUTHOR]

Highlights • PRRSV infection induces IL-1β mRNA and protein expression in microglia. • Myeloid differentiation primary response gene 88 (MyD88), extracellular signal-regulated kinase1/2 (ERK) and AP-1 signaling pathways are involved in PRRSV induced IL-1β production in microglia. • In microglia, PRRSV infection actually increased the phosphorylation of ERK1/2 and c-Jun. • NLRP3 inflammasome was proved to be activated by PRRSV and required for IL-1β secretion in microglia. Abstract Porcine reproductive and respiratory syndrome virus (PRRSV) infection which caused severe reproductive failure and respiratory disorders in swine is accompanied with severe nervous symptoms. Our previous studies demonstrated that microglia, the resident innate immune cells in central nervous system (CNS), could support PRRSV infection and replication in vitro. And PRRSV infection led to the increased expressions of large amounts of proinflammatory cytokines and chemokines which contributed to neuropathogenesis of PRRSV. Interleukin-1β (IL-1β) is one of the increased proinflammatory cytokines, which possesses diverse functions in immune response upon virus infection, including activation of innate immune and modulation of adaptive immune responses. Importantly, considerable evidences indicated that 1L-1β is involved in neuronal injury. Here, we demonstrated that PRRSV infection up-regulated IL-1β expression at both the mRNA and protein levels in microglia in a dose-dependent manner. Myeloid differentiation primary response gene 88 (MyD88), extracellular signal-regulated kinase1/2 (ERK) and activator protein 1 (AP-1) were involved in PRRSV induced IL-1β production in microglia. Moreover, NOD-like receptor protein 3 (NLRP3) inflammasome is activated by PRRSV in microglia, which is required for IL-1β secretion. Taken together, our data indicated that PRRSV infection could induce IL-1β up-regulation, which was likely mediated by MyD88/ERK/AP-1 and NLRP3 inflammasome. These findings will provide new insights into the molecular mechanisms of IL-1β production and some implications for neuropathogenesis of PRRSV. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • Sixteen outbreaks of diarrhoea in suckling piglets and sows were investigated. • Deep-sequencing was performed on 24 rotavirus A positive faecal samples. • A new strain was introduced in Spain shortly before the onset of the outbreaks. • A high diversification of the VP7 and VP4 genes occurred in a short time. • A high number of changes in the epitopes compared to vaccine strains was reported. Abstract Since early 2017, in Spain there was an apparent increase in reports on rotavirus involvement in neonatal diarrhoea outbreaks, affecting also adult sows. In this study, 16 unrelated outbreaks of diarrhoea in suckling pigs and sows, where rotavirus A was the only pathogen detected, were investigated. Deep-sequencing was performed on total RNA from twenty-four positive faecal samples. Genotyping, phylogenetic and bayesian analyses showed that all isolates had a common ancestor of porcine, or human porcine-like, origin. The new strain was introduced in the population shortly before the onset of the outbreaks. Besides, a high diversification of the VP7 and VP4 genes occurred in a short time. Isolates presented a high number of amino acid changes in the neutralizing epitopes compared to vaccine sequences. The present report illustrates how a new rotavirus A strain may disseminate rapidly and the extremely high diversification that this pathogen may undergo in a short period. [ABSTRACT FROM AUTHOR]

Abstract Porcine reproductive and respiratory syndrome virus (PRRSV) is the causative agent of porcine reproductive and respiratory syndrome (PRRS), which results in immense economic losses in the swine industry. Outbreaks of disease caused by NADC30-like PRRSV are of great concern in China. Here, a novel variant, NADC30-like PRRSV strain HB17A, was analyzed and its pathogenicity in pigs was examined. The full-length genome sequence of HB17A shared 83.6–95.1% nucleotide similarity with NADC30-like and NADC30 PRRSV without any gene insertions, but with a unique 2-amino acid deletion in Nsp2. A phylogenetic analysis showed that HB17A clustered with NADC30 strains. Different degrees of variation in the signal peptide, transmembrane region (TM), primary neutralizing epitope (PNE), non-neutral epitopes, and N-glycosylation sites were observed in GP5. Challenge experiments showed that HB17A infection resulted in persistent fever, moderate respiratory clinical signs, low levels of viremia and viral loads in serum, and mild gross and microscopic lung lesions. Moreover, IFN-γ, IL-6, and IL-10 cytokine levels were significantly elevated in serum, but the levels of IFN-α and IL-2 were similar to those of the negative controls. HB17A was less pathogenic but was secreted longer in nasal discharge than HP-PRRSV FZ06A. Our findings indicate that HB17A is a novel NADC30-like strain with certain deletions and mutations but with no evidence of genomic recombination. This strain exhibits intermediate virulence in pigs. This research will be help define the evolutionary characteristics of Chinese NADC30-like PRRSV. Highlights • A novel variant, NADC30-like PRRSV strain HB17A, has certain deletions and mutations but with no evidence of genomic recombination and also showed intermediate virulence in pigs. [ABSTRACT FROM AUTHOR]

The present study was conducted to examine whether cellular and/or viral cholesterol levels play a role in porcine deltacoronavirus (PDCoV) replication. Our results showed that depletion of cholesterol from cells or virions by treating them with methyl-β-cyclodextrin (MβCD) diminished PDCoV infection in a dose-dependent manner. The addition of exogenous cholesterol to MβCD-treated cells or virions moderately restored PDCoV infectivity. Furthermore, the pharmacological sequestration of cellular or viral cholesterol efficiently blocked both virus attachment and internalization. Taken together, the current data indicate that the cholesterol present in the cell membrane and viral envelope contributes to PDCoV replication by acting as a key component in viral entry. [ABSTRACT FROM AUTHOR]

Patients with congenital heart disease with a pressure-overloaded right ventricle can develop liver disease and would benefit from non-invasive diagnostic modalities such as ultrasound shear wave elastography (US SWE). We sought to investigate the ability of US SWE to measure dynamic changes in liver stiffness with an acute fluid bolus in an animal model. Three piglets underwent surgical intervention to create a pressure-overloaded right ventricle and, 12 wk later, underwent US SWE, both pre- and post-intravenous infusion of a saline bolus. Ultrasound measures of shear modulus, velocity and attenuation were taken to characterize hepatic mechanical properties. Liver stiffness exhibited a dynamic component that increased after fluid bolus, although not reaching statistical significance with our small sample size, and these changes were greater in more diseased livers. US SWE may provide a promising non-invasive method for assessing dynamic changes in hydration status and degree of liver disease. [ABSTRACT FROM AUTHOR]

Feed has been identified as a vector of transmission for porcine epidemic diarrhea virus (PEDV). The objective of this study was to determine if feed batch sequencing methods could minimize PEDV cross-contamination. Porcine epidemic diarrhea virus-free swine feed was manufactured to represent the negative control. A 50 kg feed batch was mixed in a pilot scale feed mill for 5 min, sampled, then discharged for 10 min into a bucket elevator and sampled again upon exit. Next, a pathogenic PEDV isolate was used to inoculate 49.5 kg of PEDV-free feed to form the positive control. The positive control was mixed, conveyed and sampled similar to the negative control. Subsequently, 4 sequence batches (sequence 1 to 4) were formed by adding a 50 kg batch of PEDV-negative feed to the mixer after the prior batch was mixed and conveyed; all sequences were mixed, conveyed, and sampled similar to the negative and positive control batches. None of the equipment was cleaned between batches within a replicate. This entire process was replicated 3 times with cleaning the feed mill between replicates. Feed was then analyzed for PEDV RNA by real-time reverse transcriptase semiquantitative polymerase chain reaction (rRTPCR) as measured by cycle threshold (Ct) and for infectivity by bioassay. Sequence 1 feed had higher (P . 0.05) rRT-PCR Ct values than the positive batch and sequence 2 feed had higher (P . 0.05) Ct values than sequence 1, regardless of sampled location. Feed sampled from the mixer from sequence 2, 3, and 4 was rRT-PCR negative whereas feed sampled from the bucket elevator was rRT-PCR negative from sequence 3 and 4. Bioassay was conducted using 66 mixed sex 10-d-old pigs confirmed negative for PEDV allocated to 22 different rooms. Pigs were initially 10-d old. Control pigs remained PEDV negative for the study. All pigs from the mixer positive batch (9/9) and bucket elevator positive batch (3/3) were rRT-PCR positive on fecal swabs by the end of the study. One replicate of pigs from mixer sequence 1 was rRT-PCR positive (3/3) by 7 dpi. One replicate of mixer pigs from sequence 2 was rRT-PCR positive (3/3) by 7 dpi although no detectable PEDV RNA was found in the feed. The results demonstrate sequenced batches had reduced quantities of PEDV RNA although sequenced feed without detectible PEDV RNA by rRT-PCR can be infectious. Therefore, a sequencing protocol can reduce but not eliminate the risk of producing infectious PEDV carryover from the first sequenced batch of feed. [ABSTRACT FROM AUTHOR]

Japanese encephalitis virus (JEV) is a zoonotic mosquito-borne virus, persisting in pigs, Ardeid birds and Culex mosquitoes. It is endemic to China and Southeastern Asia. The case-fatality ratio (CFR) or the rate of permanent psychiatric sequelae is 30% among symptomatic patients. There were no reported local JEV human cases between 2006 and 2010 in Hong Kong, but it was followed by a resurgence of cases from 2011 to 2017. The mechanism behind this “skip-and-resurgence” patterns is unclear. This work aims to reveal the mechanism behind the “skip-and-resurgence” patterns using mathematical modelling and likelihood-based inference techniques. We found that pig-to-pig transmission increases the size of JEV epidemics but is unlikely to maintain the same level of transmission among pigs. The disappearance of JEV human cases in 2006–2010 could be explained by a sudden reduction of the population of farm pigs as a result of the implementation of the voluntary “pig-rearing licence surrendering” policy. The resurgence could be explained by of a new strain in 2011, which increased the transmissibility of the virus or the spill-over ratio from reservoir to host or both. [ABSTRACT FROM AUTHOR]

Porcine epidemic diarrhoea is caused by the porcine epidemic diarrhoea virus, and is a highly contagious disease which affects the intestines of new-born piglets resulting in intense diarrhoea. Historically, the virus has caused enormous economic losses in the pig industry. In particular, the emergence of new epidemic strains means there is a pressing need for prevention and control of the disease. Owing to the specificity of the monoclonal antibodies now available, study of the pathogenesis, immune mechanisms and new diagnostic methods can be performed. In this study, 13 strains of positive hybridoma cells were prepared by immunising mice with purified whole porcine epidemic diarrhoea virus, and analysis was performed using ELISA and Western blotting. Three cell strains specifically recognised the porcine epidemic diarrhoea virus nucleocapsid protein (N protein). In this study, we report the characterisation of effective tools for the establishment of porcine epidemic diarrhoea virus diagnostic methods and we have specifically generated primary antibodies for ELISA, IFA, test strip and Western blotting. [ABSTRACT FROM AUTHOR]

Natural antisense transcripts (NATs) are widely present in mammalian genomes and act as pivotal regulator molecules to control gene expression. However, studies on the NATs of pigs are relatively rare. Here, we identified a novel antisense transcript, designated PLA2G16 -AS, transcribed from the phospholipase A2 group XVI locus ( PLA2G16 ) in the porcine genome, which is a well-known regulatory molecule of fat deposition. PLA2G16 -AS and PLA2G16 were dominantly expressed in porcine adipose tissue, and were differentially expressed between Tibetan pigs and Rongchang pigs. In addition, PLA2G16 -AS has a weak sequence conservation among different vertebrates. PLA2G16 -AS was also shown to form an RNA–RNA duplex with PLA2G16 , and to regulate PLA2G16 expression at the mRNA level. Moreover, the overexpression of PLA2G16 -AS increased the stability of PLA2G16 mRNA in porcine cells. We envision that our findings of a NAT for a regulatory gene associated with lipolysis might further our understanding of the molecular regulation of fat deposition. [ABSTRACT FROM AUTHOR]

Porcine reproductive and respiratory syndrome (PRRS), caused by the PRRS virus (PRRSV), is a widespread disease that affects domestic pigs of all ages. Accurate and rapid detection of PRRSV specific neutralizing antibodies levels in a pig herd is beneficial for the evaluation of the herd’s immunity to combat the specific viral infection. However, the current methods for viral detection, including fluorescent focus neutralization (FFN) and cytopathic effect (CPE) reduction neutralizing assays, are subjective and time-consuming. Therefore, a Clover-tagged PRRSV virus neutralization assay were developed that instrumentally measures the fluorescence signal of Clover stably expressing by a PRRSV infectious clone for at least 10 passages. Herein, the results showed that the proposed Clover-tagged PRRSV neutralization assay is reliable using instrumental measurements of the fluorescence signal of Clover and allows for rapid detection of neutralizing antibodies against PRRSV. The assay was evaluated by testing swine sera from experimental and field samples, and comparisons were made with the traditional FFN and CPE reduction assays. These results suggest that the Clover-tagged PRRSV infectious clone offers a fast and reliable testing method for neutralizing antibodies and could permit high-throughput screening of new antiviral agents. [ABSTRACT FROM AUTHOR]

Since late 2011, a pseudorabies virus (PRV) variant with increased virulence in old pigs had caused major disease outbreaks and great economic losses to the pig industry in China. The gene mutations that contributed to the increased virulence were unclear. To study the basis of the enhanced pathogenicity, an infectious bacterial artificial chromosome (BAC) clone consisting of the complete genome of the PRV variant was developed. Using homologous recombination and Cre/LoxP recombination, the recombinant virus rJS-2012-BAC carrying a BAC insertion downstream of the open reading frame (ORF) of gG was constructed. The circular genome of rJS-2012-BAC was extracted from infected Vero cells and transformed into Escherichia coli DH10B, generating the BAC clone pBAC-JS2012. The loxP sites flanking the BAC vector were used to excise the BAC sequences using Cre recombinase. The reconstituted BAC-excision virus, vJS2012 L, from pBAC-JS2012 exhibited similar biological properties to the wild-type virulent strain JS-2012. To manipulate the BAC clone pBAC-JS2012, the galK selection system was adopted to delete the gI/gE gene from pBAC-JS2012 in E. coli and to generate the gI/gE-deleted virus vJS2012-ΔgE/gI. The BAC clone, pBAC-JS2012, retained the same level of virulence as its parent strain and was easily manipulated using a galK system which would facilitate the study of the enhanced pathogenicity of the PRV variant as well as other studies on PRV. [ABSTRACT FROM AUTHOR]

The goals of this study were to compare the pathogenicity and infection dynamics of a historical and a contemporary SVA strains (SVV 001 and SD15-26) and to assess cross-neutralizing and cross-reactive T cell responses following experimental infection in pigs. Both SVA strains successfully infected all inoculated animals, resulting in viremia and robust antibody and cellular immune responses. SVA SD15-26 infection resulted in characteristic clinical signs and vesicular lesions, however, SVA SVV 001 did not cause overt clinical disease with inoculated animals remaining clinically normal during the experiment. Notably, neutralization- and -recall IFN-γ expression-assays revealed marked cross-neutralizing antibody and cross-reactive T cell responses between the two viral strains. Together these results demonstrate that the historical SVA SVV 001 strain presents low virulence in pigs when compared to the contemporary SVA SD15-26 strain. Additionally, immunological assays indicate that SVA SVV 001 and SD15-26 are antigenically related and share conserved antigenic determinants. [ABSTRACT FROM AUTHOR]

Highly contagious classical swine fever (CSF) remains a major trade and health problem in the pig industry, resulting in large economic losses worldwide. In CSF-endemic countries, attenuated CSF virus (CSFV) vaccines have been routinely used to control the disease. However, eradication of CSFV in a geographical area would require permanent reduction to zero presence of the virus. It is therefore of paramount importance to develop a safe, potent, and non-infectious CSF vaccine. We have previously reported on a cost-effective CSF E2 subunit vaccine, KNB-E2, which can protect against CSF symptoms in a single dose containing 75 μg of recombinant CSFV glycoprotein E2. In this study, we report on a series of animal studies undertaken to elucidate further the efficacy of KNB-E2. We found that pigs vaccinated with a single KNB-E2 dose containing 25 μg of recombinant CSFV glycoprotein E2 were protected from clinical symptoms of CSF. In addition, KNB-E2-mediated reduction of CSF symptoms was observed at two weeks post-vaccination and the vaccinated pigs continued to exhibit reduced CSF clinical signs when virus challenged at two months and four months post-vaccination. These results suggest that KNB-E2 effectively reduces CSF clinical signs, indicating the potential of this vaccine for safely minimizing CSF-related losses. [ABSTRACT FROM AUTHOR]

Two strains of porcine reproductive and respiratory syndrome virus (PRRSV) were isolated in 2006 and 2016 and designated as FZ06A and FZ16A, respectively. Inoculation experiments showed that FZ06A caused 100% morbidity and 60% mortality, while FZ16A caused 100% morbidity without death. By using genomic sequence and phylogenetic analyses, close relationships between a Chinese highly pathogenic PRRSV strain and the FZ06A and FZ16A strains were observed. Based on the achieved results, multiple genomic variations in Nsp2, a unique N-glycosylation site (N33→K33), and a K151 amino acid (AA) substitution for virulence in the GP5 of FZ16A were detected; except the 30 AA deletion in the Nsp2-coding region. Inoculation experiments were conducted and weaker virulence of FZ16A than FZ06A was observed. Based on our results, a 30 AA deletion in the Nsp2-coding region is an unreliable genomic indicator of a high virulence PRRSV strain. The Nsp2 and GP5 differences, in addition to the virulence difference between these two highly pathogenic PRRSV strains, have the potential to be used to establish a basis for further study of PRRSV virulence determinants and to provide data useful in the development of vaccines against this economically devastating disease. [ABSTRACT FROM AUTHOR]

Summary: Porcine epidemic diarrhoea virus (PEDV) is a globally emerging and re‐emerging enteric coronavirus in pigs causing serious economic threats to the world swine industry. Since the re‐emergence of massive PEDV outbreaks in South Korea in 2013−2014, domestic pig farms have continued to experience PED epidemics or endemics. This study represents the molecular characterization of PEDV isolates identified in diarrhoeic animals collected across the country in 2017. Initial sequencing analysis of the full‐length S genes revealed that 70% of the 2017 isolates (7/10) belong to the G2b subgroup, while the remaining isolates were classified as G1b. The data indicated that both variant G1b and global epidemic G2b strains were responsible for current PED outbreaks in South Korea. The 2017 G1b and G2b isolates shared 98.7%–99.4% and 98.1%–99.2% amino acid sequence identity at the S gene level and 99.3% and 99.0%–99.6% nucleotide sequence homology at the genome level compared to the corresponding Korean prototype G1b and G2b strains, respectively. In an interesting manner, one G2b‐like KNU‐1705 strain was found to possess a large 39‐nucleotide deletion in the ORF1a region theoretically encoding nonstructural protein 3. Phylogenetic analysis based on the entire genome and spike protein sequences indicated that the 2017 isolates were most closely related to other global G1b or G2b strains but formed different branches within the same genogroup. These results indicate that PEDVs undergo continuous evolution in the field. In addition, one 2017 PEDV strain, KOR/KNU‐1705/2017, was successfully isolated and propagated in Vero cells. The antisera raised against the Korean prototype 2014 G2b strain efficiently neutralized KNU‐1705 virus infection, suggesting antigenic homology between the 2014 and 2017 PEDV strains. Our data advance the understanding of the molecular epidemiology and antigenicity of PEDV circulating in South Korea. [ABSTRACT FROM AUTHOR]

Summary: Since the introduction of African swine fever virus (ASFV) into the Baltic states and Poland in 2014, the disease has continued to spread within these regions. In 2017, the virus spread further west and the first cases of disease were reported in the Czech Republic and Romania, in wild boar and domestic pigs, respectively. To control further spread, knowledge of different modes of transmission, including indirect transmission via a contaminated environment, is crucial. Up until now, such an indirect mode of transmission has not been demonstrated. In this study, transmission via an environment contaminated with excretions from ASFV‐infected pigs was investigated. Following euthanasia of pigs that were infected with an isolate of ASFV from Poland (POL/2015/Podlaskie/Lindholm), healthy pigs were introduced into the pens, in which the ASFV‐infected pigs had been housed. Introduction was performed at 1, 3, 5 or 7 days, following euthanasia of the infected pig groups. Pigs, that were introduced into the contaminated environment after 1 day, developed clinical disease within 1 week, and both ASFV DNA and infectious virus were isolated from their blood. However, pigs introduced into the contaminated pens after 3, 5 or 7 days did not develop any signs of ASFV infection and no viral DNA was detected in blood samples obtained from these pigs within the following 3 weeks. Thus, it was shown that exposure of pigs to an environment contaminated with ASFV can result in infection. However, the time window for transmissibility of ASFV seems very limited, and, within our experimental system, there appears to be a rapid decrease in the infectivity of ASFV in the environment. [ABSTRACT FROM AUTHOR]

Background: The large livestock operations and dense human population of Southeast Asia are considered a hot-spot for emerging viruses. Objectives: To determine if the pathogens adenovirus (ADV), coronavirus (CoV), encephalomyocarditis virus (EMCV), enterovirus (EV), influenza A-D (IAV, IBV, ICV, and IDV), porcine circovirus 2 (PCV2), and porcine rotaviruses A and C (RVA and RVC), are aerosolized at the animal-interface, and if humans working in these environments are carrying these viruses in their nasal airways. Study: This cross-sectional study took place in Sarawak, Malaysia among 11 pig farms, 2 abattoirs, and 3 animal markets in June and July of 2017. Pig feces, pig oral secretions, bioaerosols, and worker nasal wash samples were collected and analyzed via rPCR and rRT-PCR for respiratory and diarrheal viruses. Results: In all, 55 pig fecal, 49 pig oral or water, 45 bioaerosol, and 78 worker nasal wash samples were collected across 16 sites. PCV2 was detected in 21 pig fecal, 43 pig oral or water, 3 bioaerosol, and 4 worker nasal wash samples. In addition, one or more bioaerosol or pig samples were positive for EV, IAV, and RVC, and one or more worker samples were positive for ADV, CoV, IBV, and IDV. Conclusions: This study demonstrates that nucleic acids from a number of targeted viruses were present in pig oral secretions and pig fecal samples, and that several viruses were detected in bioaerosol samples or in the nasal passages of humans with occupational exposure to pigs. These results demonstrate the need for future research in strengthening viral surveillance at the human-animal interface, specifically through expanded bioaerosol sampling efforts and a seroepidemiological study of individuals with exposure to pigs in this region for PCV2 infection. [ABSTRACT FROM AUTHOR]

Colistin resistance in Enterobacteriaceae is a public health problem. The present study was designed to evaluate the dissemination of a colistin-resistant Escherichia coli strain and its resistance gene, mcr-1 , between orally inoculated pigs and their contacts. A non-inoculated control group, one low-dose and one high-dose group—both including two pens of two inoculated and three contact pigs—were raised in separate rooms. After inoculation of a colistin- and rifampicin-resistant E. coli suspension (2.5 × 10 5 CFU/pig for the low-dose group and 2.5 × 10 8 CFU/pig for the high-dose group), feces from inoculated and non-inoculated contact pigs were collected and inoculated on colistin- and rifampicin-supplemented media directly or after enrichment in rifampicin-supplemented media, then the isolates were characterized. PCR was used to detect the mcr-1 gene in lysates from feces cultivated in colistin-supplemented broth and DNA prepared from feces. Results showed that the low-dose inoculum was probably insufficient to obtain durable colonization, but could lead to the temporary presence of mcr-1 -positive E. coli strains. The high-dose inoculum resulted in durable colonization of both inoculated and contact animals. In all groups, the mcr-1 gene was also detected in rifampicin-susceptible strains, suggesting its transfer to several commensal strains. A comparison of detection methods showed that more positive samples were obtained with cultures in rifampicin-supplemented media and suggests that current methods to evaluate the prevalence of colistin resistance in fecal samples suffer from poor sensitivity. [ABSTRACT FROM AUTHOR]

The porcine small intestinal epithelial cell line, IPEC-J2, is useful to characterize the interactions of enterocytes with enteric viruses in vitro . We investigated whether IPEC-J2 cells are susceptible to porcine deltacoronavirus (PDCoV) infection. We conducted quantification of infectious virus or viral RNA, immunofluorescent (IF) staining for the detection of PDCoV antigens, and TUNEL assay in IPEC-J2 cells inoculated with the strain OH-FD22-P8 grown in LLC-PK cells, and supplemented with 10 μg/ml of trypsin in the cell culture medium. Cytopathic effects (CPE) that consisted of enlarged and rounded cells followed by cell shrinkage and detachment, were identified by the 3rd viral passage in the IPEC-J2 cells. PDCoV antigen was detected in the cells showing CPE. By double IF and TUNEL staining, most PDCoV antigen-positive IPEC-J2 cells failed to show TUNEL-positive signals, indicating that PDCoV-infected IPEC-J2 cells may not undergo apoptosis, but rather necrosis, similar to necrotic cell death of infected enterocytes in vivo. There was increased interleukin-6 in PDCoV-infected IPEC-J2 cell culture supernatants at post-inoculation hour (PIH) 48–96, as evaluated by ELISA, concurrent with increased titers of PDCoV at PIH 24–72. The susceptibility of IPEC-J2 cells to PDCoV infection supports their usefulness to characterize the interactions of enterocytes with PDCoV. We also demonstrated that IPEC-J2 cell culture-passaged PDCoV (OH-FD22-P8-I-P4) was enteropathogenic in 10-day-old gnotobiotic pigs, and induced systemic innate and pro-inflammatory cytokine responses during the acute PDCoV infection. [ABSTRACT FROM AUTHOR]

The efficacy of the CA-2-MP120 vaccine, a cell culture-attenuated strain of virulent porcine reproductive and respiratory syndrome virus (PRRSV), was assessed in pigs. Despite the persistence of viremia in all vaccinated animals during the immunization period, the virus was not detected in vaccinated pigs following challenge. Furthermore, no pigs in the vaccinated group shed PRRSV nasally, orally or rectally throughout the experiment. Moreover, histopathological lung and lymph node lesions in the immunized group were much milder than those in the unimmunized and challenged group. These results indicated that CA-2-MP120 can provide effective protection against virulent wildtype PRRSV-2. [ABSTRACT FROM AUTHOR]

Porcine Reproductive and Respiratory Syndrome (PRRS) is an elusive model of host/virus relationship in which disease is determined by virus pathogenicity, pig breed susceptibility and phenotype, microbial infectious pressure and environmental conditions. Successful disease control in PRRS-endemic Countries corresponds to “stability”, i.e. a condition with no clinical signs of PRRS in the breeding-herd population and no viremia in weaning-age pigs. The aim of this work was to compare the profile and time-course of humoral and cell-mediated immunity in stable and unstable herds, respectively. In particular, we investigated PRRS virus (PRRSV) in serum and group oral fluid samples by Real-time RT-PCR, PRRSV-specific IgA and IgG in oral fluids, serum IgG antibody and the cell-mediated response (PRRSV-specific release of interferon-gamma) in whole blood samples. These parameters were measured in order to identify possible discrepancies in the development and kinetics of the immune response against PRRSV. PRRS-free gilts got regularly infected after entering PRRS-stable and unstable farms. In an open cycle, unstable pig farm PRRSV infection could be demonstrated in all groups of pigs, including suckling piglets. Four main results should be highlighted: A) the precocity of the Ab response in group oral fluids was generally similar to that recorded in sera; B) circulation of PRRSV was consistently detected in all age groups in the unstable herds, as opposed to the stable ones; C) an early, balanced, IgA and IgG response in oral fluids was only observed in the stable herds; D) an early IFN-gamma response after PRRSV infection was often observed in stable herds, as opposed to the unstable ones. These were characterized by IFN-gamma responses in piglets, likely due to transfer of maternal immunity. Most important, the mucosal IgA response was associated with cessation of virus excretion in oral fluid samples of PRRS-unstable herds. The above findings indicate that a peculiar profile of immune response to PRRSV can be found in PRRS-stable herds. Therefore, the outlined immune parameters can represent a useful readout system to evaluate successful adaptation to PRRSV based on acclimatization of breeding animals and management of pig flow. [ABSTRACT FROM AUTHOR]

The purpose of this case report is to describe for the first time concurrent porcine reproductive and respiratory syndrome virus (PRRSV) and porcine circovirus 2 (PCV-2) infections in a commercial farrow-to-finish pig farm in Albania, as well as the phylogenetical analysis of isolated PRRSV strain. The present study reports on a farrow-to-finish commercial pig farm, located in South Albania. In a percentage of about 40% of weaners in each batch (60–70 piglets per batch), clinical signs, including fever, severe respiratory signs, wasting, jaundice, rough hairy coat, palpable inguinal lymphadenopathy, and high mortality rate, were performed. The clinical signs of sows included sporadic premature farrowings (22%), with increased number of stillbirth (3.3%) and weak piglets (4.1%) based on the record system of the farm. Blood samples were obtained from 8 sows (4 lactating and 4 dry-period sows), 25 piglets of 5 different batches (5 at 15–20 days, 5 at 40 days, 5 at 50 days, 5 of 60 days, and 5 of 70 days), and 5 finishers of 130–150 days of age. Moreover, tissue samples were collected from five weaners at 20–70 days of age. Histopathological examination of lung and lymph node sections revealed findings compatible with PRRSV and PCV-2 infection. Pigs between 15 and 130–140 days of age were positive for type 1 (European) PRRSV and pigs between 50 and 130–140 days of age were positive for PCV-2. Blood serum samples were tested by real-time polymerase chain reaction (PCR) for PCV-2 and one real-time reverse transcription–PCR-positive sample was selected for subsequent complete ORF5 (Gp5) gene sequencing. The results of this case report confirm the detection of PRRSV and PCV-2 concurrent infection in an Albanian farrow-to-finish pig farm. The full-length ORF5 sequence of the detected PRRSV strain (named “Mursi/AL/15”) was successfully determined, revealing high nucleotide identity with other type 1 European isolates. [ABSTRACT FROM AUTHOR]

Influenza A virus commonly circulating in swine (IAV-S) is characterized by large genetic and antigenic diversity and, thus, improvements in different aspects of IAV-S surveillance are needed to achieve desirable goals of surveillance such as to establish the capacity to forecast with the greatest accuracy the number of influenza cases likely to arise. Advancements in modeling approaches provide the opportunity to use different models for surveillance. However, in order to make improvements in surveillance, it is necessary to assess the predictive ability of such models. This study compares the sensitivity and predictive accuracy of the autoregressive integrated moving average (ARIMA) model, the generalized linear autoregressive moving average (GLARMA) model, and the random forest (RF) model with respect to the frequency of influenza A virus (IAV) in Ontario swine. Diagnostic data on IAV submissions in Ontario swine between 2007 and 2015 were obtained from the Animal Health Laboratory (University of Guelph, Guelph, ON, Canada). Each modeling approach was examined for predictive accuracy, evaluated by the root mean square error, the normalized root mean square error, and the model’s ability to anticipate increases and decreases in disease frequency. Likewise, we verified the magnitude of improvement offered by the ARIMA, GLARMA and RF models over a seasonal-naïve method. Using the diagnostic submissions, the occurrence of seasonality and the long-term trend in IAV infections were also investigated. The RF model had the smallest root mean square error in the prospective analysis and tended to predict increases in the number of diagnostic submissions and positive virological submissions at weekly and monthly intervals with a higher degree of sensitivity than the ARIMA and GLARMA models. The number of weekly positive virological submissions is significantly higher in the fall calendar season compared to the summer calendar season. Positive counts at weekly and monthly intervals demonstrated a significant increasing trend. Overall, this study shows that the RF model offers enhanced prediction ability over the ARIMA and GLARMA time series models for predicting the frequency of IAV infections in diagnostic submissions. [ABSTRACT FROM AUTHOR]

The nucleocapsid (N) protein is the most abundant protein of porcine reproductive and respiratory syndrome virus (PRRSV). It has been shown to be multiphosphorylated. However, the phosphorylation sites are still unknown. In this study, we used liquid chromatography tandem mass spectrometry (LC–MS/MS) to analyze the phosphorylation sites of N protein expressed in Sf9 cells. The results showed that N protein contains two phosphorylation sites. Since N protein can regulate IL-10, which may facilitate PRRSV replication, we constructed four plasmids (pCA-XH-GD, pCA-A105, pCA-A120 and pCA-A105-120) and transfected them into Pig alveolar macrophages (PAMs,3D4/2). The qPCR results showed that mutations at residues 105 and 120 were associated with down-regulation of the IL-10 mRNA level, potentially decreasing the viral growth ability. Then, we mutated the phosphorylation sites (S105A and S120A) and rescued three mutated viruses, namely, A105, A120 and A105-120. Compared with wild-type virus titers, the titers of the mutated viruses at 48 hpi were significantly decreased. The Nsp(non-structural protein) 9 qPCR results were consistent with the multistep growth kinetics results. The infected PAMs (primary PAMs) results were similar with Marc-145.The findings indicated that the mutations impaired the viral replication ability. The confocal microscopy results suggested that mutations to residues 105 and 120 did not affect N protein distribution. Whether the mutations affected other functions of N protein and what the underlying mechanisms are need further investigation. In conclusion, our results show that residues 105 and 120 are phosphorylation sites and are important for N protein function and for viral replication ability. [ABSTRACT FROM AUTHOR]

Porcine reproductive and respiratory syndrome (PRRS) is one of the most important swine diseases affecting pig industry worldwide. Sialoadehesin (Sn) and CD163 are the two specific receptors for PRRSV infection of porcine alveolar macrophages. Our previous study showed that the soluble Sn receptor Sn4D-Fc and soluble CD163 receptor SRCR59-Fc expressed by the two recombinant adenoviral (rAd) vectors have an additive anti-PRRSV effect in vitro . In the present study, rAd-Sn4D-Fc and rAd-SRCR59-Fc were inoculated into pigs, and the efficient expression of Sn4D-Fc and SRCR59-Fc proteins was detected by ELISA. Then, PRRSV-naïve pigs were inoculated with rAd-Sn4D-Fc and/or rAd-SRCR59-Fc before contagious infection with different PRRSV strains. Among the three rAd inoculation groups, simultaneous inoculation with the two rAd vectors provided the best protection against highly pathogenic JXA1 strain PRRSV, followed by rAd-SRCR59-Fc inoculation and rAd-Sn4D-Fc inoculation. Clinical observation and quantitative RT-PCR analyses showed that all of the double rAd-inoculated pigs (n = 9) survived from the contagious infection with highly pathogenic JXA1, JS07 or SH1705 strain PRRSV with significantly alleviated clinical scores, viremia, fecal viral emission and tissue virus loads. These data suggest that rAd-Sn4D-Fc and rAd-SRCR59-Fc can be developed further as the universal therapeutic vaccine to facilitate PRRSV eradication. [ABSTRACT FROM AUTHOR]