Influence of deoxynivalenol-contaminated feed on the immune response of pigs after PRRSV vaccination and infection.

The impact of the Fusarium mycotoxin deoxynivalenol (DON) on the immune response against porcine reproductive and respiratory syndrome virus (PRRSV) vaccination and infection was investigated. Forty-two weaned piglets were separated into seven groups and received three different diets: Low DON (1.09 ppm), High DON (2.81 ppm) or No DON. These three treatments were split further into either vaccinated (Ingelvac PRRSFLEX EU) and challenged with PRRSV 28 days post-vaccination, or only infected at day 28. A seventh group received no DON, no vaccination, and no infection. Two weeks after challenge infection, when pigs were euthanized, the number of IFN-γ producing lymphocytes in the blood of vaccinated animals was lower in pigs on High DON compared to animals on Low DON or No DON. Intracellular cytokine staining showed that vaccinated animals fed with the Low DON diet had higher frequencies of TNF-α/IFN-γ co-producing CD4+ T cells than the other two vaccinated groups, particularly in lung tissue. Vaccinated animals on High DON had similar viral loads in the lung as the non-vaccinated groups, but several animals of the Low DON or No DON group receiving vaccination had reduced titers. In these two groups, there was a negative correlation between lung virus titers and vaccine-specific TNF-α/IFN-γ co-producing CD4+ T cells located either in lung tissue or blood. These results indicate that after PRRSV vaccination and infection, high levels of DON negatively influence immune parameters and clearance of the virus, whereas low DON concentrations have immunomodulatory effects.

Proteomic and immunoproteomic insights into the exoproteome of Actinobacillus pleuropneumoniae, the causative agent of porcine pleuropneumonia.

Porcine pleuropneumonia caused by Actinobacillus pleuropneumoniae affects pig health status and the swine industry worldwide. Despite the extensive number of studies focused on A. pleuropneumoniae infection and vaccine development, a thorough analysis of the A. pleuropneumoniae exoproteome is still missing. Using a complementary approach of quantitative proteomics and immunoproteomics we gained an in-depth insight into the A. pleuropneumoniae serotype 2 exoproteome, which provides the basis for future functional studies. Label-free liquid chromatography-tandem mass spectrometry (LC-MS/MS) revealed 593 exoproteins, of which 104 were predicted to be virulence factors. The RTX toxins ApxIIA and ApxIIIA -were found to be the most abundant proteins in the A. pleuropneumoniae serotype 2 exoproteome. Furthermore, the ApxIVA toxin was one of the proteins showing the highest abundance, although ApxIVA is commonly assumed to be expressed exclusively in vivo. Our study revealed several antigens, including proteins with moonlight functions, such as the elongation factor (EF)-Tu, and proteins linked to specific metabolic traits, such as the maltodextrin-binding protein MalE, that warrant future functional characterization and might present potential targets for novel therapeutics and vaccines. Our Ig-classes specific serological proteome analysis (SERPA) approach allowed us to explore the development of the host humoral immune response over the course of the infection. These SERPAs pinpointed proteins that might play a key role in virulence and persistence and showed that the immune response to the different Apx toxins is distinct. For instance, our results indicate that the ApxIIIA toxin has properties of a thymus-independent antigen, which should be studied in more detail.

Establishment of an In Vitro Co-Culture Model of the Piglet Gut to Study Inflammatory Response and Barrier Integrity.

In intensive farming, piglets are exposed to various challenges that activate intestinal inflammatory processes, negatively affecting animal health and leading to economic losses. To study the role of the inflammatory response on epithelial barrier integrity, co-culture systems that mimic in vivo complexity are more and more preferred over cell monocultures. In this study, an in vitro gut co-culture model consisting of intestinal porcine epithelial cells and porcine peripheral blood mononuclear cells was established. The model provides an appropriate tool to study the role of the inflammatory response on epithelial barrier integrity and to screen for feed and food components, exerting beneficial effects on gut health. In the established model, inflammation-like reactions and damage of the epithelial barrier, indicated by a decrease of transepithelial electrical resistance, were elicited by activation of peripheral blood mononuclear cells via one of 3 stimuli: lipopolysaccharide, lipoteichoic acid, or concanavalin A. Two phytogenic substances that are commonly used as feed additives, licorice extract and oregano oil, have been shown to counteract the drop in transepithelial electrical resistance values in the gut co-culture model. The established co-culture model provides a powerful in vitro tool to study the role of intestinal inflammation on epithelial barrier integrity. As it consists of porcine epithelial and porcine blood cells it perfectly mimics in vivo conditions and imitates the inter-organ communication of the piglet gut. The developed model is useful to screen for nutritional components or drugs, having the potential to balance intestinal inflammation and strengthen the epithelial barrier integrity in piglets.

Reproductive failure in an Austrian piglet-producing farm due to porcine circovirus genotype 2d.

Infections of pigs with porcine circovirus type 2 (PCV2) can lead to various clinical conditions including reproductive disorders (PCV2-RD). In general, a transplacental infection of fetuses leads to mummification and stillbirth. So far, PCV2-RD has mainly been described in specific-pathogen-free (SPF) herds or farms with a high proportion of gilts. From December 2018 to February 2019, a high abundance of mummified fetuses (15.5%) was observed in two farrowing groups in an Austrian piglet-producing farm. PCV2 DNA was detected using qPCR in organs of all six investigated fetuses (2.07 × 108-1.09 × 1012 PCV2) genome equivalents/g tissue and via in situ hybridisation in organs from five fetuses, while histologic lesions were not observed in a single fetal heart. All isolates were sequenced and identified as PCV2d. After the implementation of a regular vaccination of all sows against PCV2, the abundance of mummified fetuses dropped to 3.5% in May 2019. In contrast to previous reports about PCV2-RD, this farm was neither an SPF herd nor a start-up herd with a high proportion of gilts. The implementation of regular PCV2 vaccination helped to reduce the abundance of mummified fetuses substantially.

Transglycosylated starch accelerated intestinal transit and enhanced bacterial fermentation in the large intestine using a pig model.

Resistant starch can alter the intestinal nutrient availability and bulk of digesta, thereby modulating the substrate available for microbial metabolic activity along the gastrointestinal tract. This study elucidated the effect of transglycosylated starch (TGS) on the retention of digesta in the upper digestive tract, ileal flow and hindgut disappearance of nutrients, and subsequent bacterial profiles in pigs. Fourteen ileal-cannulated growing pigs were fed either the TGS or control (CON) diet in a complete crossover design. Each period consisted of a 10-d adaptation to the diets, followed by 3-d collection of faeces and ileal digesta. Consumption of TGS decreased the retention of digesta in the stomach and small intestine, and increased ileal DM, starch, Ca and P flow, leading to enhanced starch fermentation in the hindgut compared with CON-fed pigs. TGS increased ileal and faecal total SCFA, especially ileal and faecal acetate and faecal butyrate. Gastric retention time positively correlated to Klebsiella, which benefitted together with Selenomonas, Lactobacillus, Mitsuokella and Coriobacteriaceae from TGS feeding and ileal starch flow. Similar relationships existed in faeces with Coriobacteriaceae, Veillonellaceae and Megasphaera benefitting most, either directly or indirectly via cross-feeding, from TGS residuals in faeces. TGS, in turn, depressed genera within Ruminococcaceae, Clostridiales and Christensenellaceae compared with the CON diet. The present results demonstrated distinct ileal and faecal bacterial community and metabolite profiles in CON- and TGS-fed pigs, which were modulated by the type of starch, intestinal substrate flow and retention of digesta in the upper digestive tract.

Listeriosis in fattening pigs caused by poor quality silage - a case report.

BACKGROUND: Listeria (L.) monocytogenes as the causative agent of listeriosis in humans and different animal species, has its reservoir in the environment. It can be found in the gut and faeces of healthy pigs, but under certain circumstances it may cause clinical disease. Fatteners are usually not known to get affected by Listeria-associated septicaemia and enteritis. This case report shows, that L. monocytogenes should be part of the list of differential diagnoses, when fattening pigs suffer from haemorrhagic diarrhoea and septicaemia. CASE PRESENTATION: Here, we report of an episode of fatal listeriosis in fattening pigs in a piglet producing farm in Lower Austria, which was combined with a fattening unit with space for 450 fatteners. The mortality rate resulted in 7.8% among fattening pigs after suffering from clinical symptoms such as anorexia, bloody diarrhoea and increased body temperature. Two fattening pigs with clinical symptoms and maize silage samples were used for further diagnostics. L. monocytogenes were isolated from serosa samples of the pigs and in the corresponding fed maize silage. One animal was positively tested for Brachyspira hyodysenteriae, which may have also been involved in the development of colitis. Immunohistochemically, L. monocytogenes could be detected in high amounts in lymphatic tissue of the gut. Molecular biological characterisation of the L. monocytogenes isolates from pigs and maize silage resulted in an identical DNA-fingerprint assigned to sequence type (ST) 21. Additionally, a high content of deoxynivalenol (3000 parts per billion) was found in maize silage. Therefore, the maize silage produced under inappropriate ensilaging conditions in a silo, was most likely the source of infection. Antimicrobial therapy with amoxicillin led to a fast cure of the remaining affected fatteners. CONCLUSION: To conclude, we were able to show, that L. monocytogenes can cause clinical disease in finishing pigs, which may have been a result of immunosuppression due to high deoxynivalenol exposure. When feeding silage it is important that all ensilaging procedures occur under appropriate anaerobic conditions to guarantee suppression of listerial growth.

Novel Pestivirus Species in Pigs, Austria, 2015.

A novel pestivirus species was discovered in a piglet-producing farm in Austria during virologic examinations of congenital tremor cases. The emergence of this novel pestivirus species, provisionally termed Linda virus, in domestic pigs may have implications for classical swine fever virus surveillance and porcine health management.

Influenza A Virus Infection in Pigs Attracts Multifunctional and Cross-Reactive T Cells to the Lung.

UNLABELLED: Pigs are natural hosts for influenza A viruses and play a critical role in influenza epidemiology. However, little is known about their influenza-evoked T-cell response. We performed a thorough analysis of both the local and systemic T-cell response in influenza virus-infected pigs, addressing kinetics and phenotype as well as multifunctionality (gamma interferon [IFN-γ], tumor necrosis factor alpha [TNF-α], and interleukin-2 [IL-2]) and cross-reactivity. A total of 31 pigs were intratracheally infected with an H1N2 swine influenza A virus (FLUAVsw) and consecutively euthanized. Lungs, tracheobronchial lymph nodes, and blood were sampled during the first 15 days postinfection (p.i.) and at 6 weeks p.i. Ex vivo flow cytometry of lung lymphocytes revealed an increase in proliferating (Ki-67(+)) CD8(+) T cells with an early effector phenotype (perforin(+) CD27(+)) at day 6 p.i. Low frequencies of influenza virus-specific IFN-γ-producing CD4(+) and CD8(+) T cells could be detected in the lung as early as 4 days p.i. On consecutive days, influenza virus-specific CD4(+) and CD8(+) T cells produced mainly IFN-γ and/or TNF-α, reaching peak frequencies around day 9 p.i., which were up to 30-fold higher in the lung than in tracheobronchial lymph nodes or blood. At 6 weeks p.i., CD4(+) and CD8(+) memory T cells had accumulated in lung tissue. These cells showed diverse cytokine profiles and in vitro reactivity against heterologous influenza virus strains, all of which supports their potential to combat heterologous influenza virus infections in pigs. IMPORTANCE: Pigs not only are a suitable large-animal model for human influenza virus infection and vaccine development but also play a central role in the emergence of new pandemic strains. Although promising candidate universal vaccines are tested in pigs and local T cells are the major correlate of heterologous control, detailed and targeted analyses of T-cell responses at the site of infection are scarce. With the present study, we provide the first detailed characterization of magnitude, kinetics, and phenotype of specific T cells recruited to the lungs of influenza virus-infected pigs, and we could demonstrate multifunctionality, cross-reactivity, and memory formation of these cells. This, and ensuing work in the pig, will strengthen the position of this species as a large-animal model for human influenza virus infection and will immediately benefit vaccine development for improved control of influenza virus infections in pigs.

Congenital infection with atypical porcine pestivirus (APPV) is associated with disease and viral persistence.

In 2013, several Austrian piglet-producing farms recorded outbreaks of action-related repetitive myoclonia in newborn piglets ("shaking piglets"). Malnutrition was seen in numerous piglets as a complication of this tremor syndrome. Overall piglet mortality was increased and the number of weaned piglets per sow decreased by more than 10% due to this outbreak. Histological examination of the CNS of affected piglets revealed moderate hypomyelination of the white substance in cerebellum and spinal cord. We detected a recently discovered pestivirus, termed atypical porcine pestivirus (APPV) in all these cases by RT-PCR. A genomic sequence and seven partial sequences were determined and revealed a 90% identity to the US APPV sequences and 92% identity to German sequences. In confirmation with previous reports, APPV genomes were identified in different body fluids and tissues including the CNS of diseased piglets. APPV could be isolated from a "shaking piglet", which was incapable of consuming colostrum, and passaged on different porcine cells at very low titers. To assess the antibody response a blocking ELISA was developed targeting NS3. APPV specific antibodies were identified in sows and in PCR positive piglets affected by congenital tremor (CT). APPV genomes were detected continuously in piglets that gradually recovered from CT, while the antibody titers decreased over a 12-week interval, pointing towards maternally transmitted antibodies. High viral loads were detectable by qRT-PCR in saliva and semen of infected young adults indicating a persistent infection.

Frequency of Th17 cells correlates with the presence of lung lesions in pigs chronically infected with Actinobacillus pleuropneumoniae.

Porcine contagious pleuropneumonia caused by Actinobacillus pleuropneumoniae (APP) remains one of the major causes of poor growth performance and respiratory disease in pig herds. While the role of antibodies against APP has been intensely studied, the porcine T cell response remains poorly characterized. To address this, pigs were intranasally infected with APP serotype 2 and euthanized during the acute phase [6-10 days post-infection (dpi)] or the chronic phase of APP infection (27-31 dpi). Lymphocytes isolated from blood, tonsils, lung tissue and tracheobronchial lymph nodes were analyzed by intracellular cytokine staining (ICS) for IL-17A, IL-10 and TNF-α production after in vitro stimulation with crude capsular extract (CCE) of the APP inoculation strain. This was combined with cell surface staining for the expression of CD4, CD8α and TCR-γδ. Clinical records, microbiological investigations and pathological findings confirmed the induction of a subclinical APP infection. ICS-assays revealed the presence of APP-CCE specific CD4+CD8αdim IL-17A-producing T cells in blood and lung tissue in most infected animals during the acute and chronic phase of infection and a minor fraction of these cells co-produced TNF-α. APP-CCE specific IL-17A-producing γδ T cells could not be found and APP-CCE specific IL-10-producing CD4+ T cells were present in various organs but only in a few infected animals. The frequency of identified putative Th17 cells (CD4+CD8αdimIL-17A+) in lung and blood correlated positively with lung lesion scores and APP-specific antibody titers during the chronic phase. These results suggest a potential role of Th17 cells in the immune pathogenesis of APP infection.

Genome-wide analysis of the transcriptional response to porcine reproductive and respiratory syndrome virus infection at the maternal/fetal interface and in the fetus.

BACKGROUND: Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) infection of pregnant pigs can result in congenital infection and ultimately fetal death. Little is known about immune responses to infection at the maternal-fetal interface and in the fetus itself, or the molecular events behind virus transmission and disease progression in the fetus. To investigate these processes, RNA-sequencing of two sites, uterine endothelium with adherent placental tissue and fetal thymus, was performed 21 days post-challenge on four groups of fetuses selected from a large PRRSV challenge experiment of pregnant gilts: control (CON), uninfected (UNINF), infected (INF), and meconium-stained (MEC) (n = 12/group). Transcriptional analyses consisted of multiple contrasts between groups using two approaches: differential gene expression analysis and weighted gene co-expression network analysis (WGCNA). Biological functions, pathways, and regulators enriched for differentially expressed genes or module members were identified through functional annotation analyses. Expression data were validated by reverse transcription quantitative polymerase chain reaction (RTqPCR) carried out for 16 genes of interest. RESULTS: The immune response to infection in endometrium was mainly adaptive in nature, with the most upregulated genes functioning in either humoral or cell-mediated immunity. In contrast, the expression profile of infected fetal thymus revealed a predominantly innate immune response to infection, featuring the upregulation of genes regulated by type I interferon and pro-inflammatory cytokines. Fetal infection was associated with an increase in viral load coupled with a reduction in T cell signaling in the endometrium that could be due to PRRSV-controlled apoptosis of uninfected bystander cells. There was also evidence for a reduction in TWIST1 activity, a transcription factor involved in placental implantation and maturation, which could facilitate virus transmission or fetal pathology through dysregulation of placental function. Finally, results suggested that events within the fetus could also drive fetal pathology. Thymus samples of meconium-stained fetuses exhibited an increase in the expression of pro-inflammatory cytokine and granulocyte genes previously implicated in swine infectious disease pathology. CONCLUSIONS: This study identified major differences in the response to PRRSV infection in the uterine endometrium and fetus at the gene expression level, and provides insight into the molecular basis of virus transmission and disease progression.

Relationships of CD163 and CD169 positive cell numbers in the endometrium and fetal placenta with type 2 PRRSV RNA concentration in fetal thymus.

Several routes of porcine reproductive and respiratory virus PRRSV transmission across the porcine diffuse epitheliochorial placentation have been proposed, but none have been proven. The objectives of this study were to investigate associations between numbers of CD163 and CD169 positive macrophages, cathepsin positive areolae, and type 2 PRRSV load at the maternal-fetal interface in order to examine important factors related to transplacental infection. On gestation day 85 ± 1, naïve pregnant gilts were inoculated with PRRSV (n = 114) or were sham inoculated (n = 19). At 21 days post-inoculation (dpi), dams and their litters were humanely euthanized and necropsied. Samples of the maternal-fetal interface (uterus with fully attached placenta) and fetal thymus were collected for analysis by RT-qPCR to quantify PRRSV RNA concentration. The corresponding paraffin-embedded uterine tissue sections were subjected to immunohistochemistry for PRRSV nucleocapsid N protein, CD163, CD169, and cathepsin. Our findings confirm significant increases in the numbers of PRRSV, CD163 and CD169 positive cells at the maternal-fetal interface during type 2 PRRSV infection in pregnant gilts. PRRSV load in fetal thymus was positively related to CD163(+) cell count in endometrium and negatively related to CD163(+) cell count in placenta, but unrelated to CD169 counts or cathepsin positive areolae. The endometrium:placenta ratio of CD163 cells, and to a lesser extent CD169 cells, was significantly associated with an increase fetal viral load in thymus. These findings suggest a more important role for CD163(+) cells following trans-placental PRRSV infection, but dichotomous responses in endometrium and placenta for both CD163 and CD169 cells.

Maternal and fetal predictors of fetal viral load and death in third trimester, type 2 porcine reproductive and respiratory syndrome virus infected pregnant gilts.

Minimal research has focused on understanding mechanisms underlying porcine reproductive and respiratory syndrome virus (PRRSV) induced reproductive failure. We have completed a large-scale project investigating phenotypic and genotypic predictors of reproductive PRRS severity in which numerous clinical, pathological, immunologic and viral responses were characterized in dams and fetuses. The goal was to determine which phenotypic responses were associated with fetal viral load and death after experimental infection of pregnant gilts with type 2 PRRSV, thereby elucidating mechanisms of reproductive PRRS in third trimester pregnant gilts. The presence of fetal infection and increasing RNA concentration at the maternal-fetal interface were strong predictors of the probability of fetal death, while PRRSV RNA concentration in dam sera and systemic tissues were not associated with the odds of fetal death. Fetal infection and death clustered, indicating that the status of adjacent fetuses is crucial for lateral transmission and fetal outcome. Several systemic immune responses of gilts were associated with fetal outcome and viral load: interferon-α contributed to the probability of fetal death, but absolute numbers of T helper cells in early infection, absolute numbers of myeloid cells over time and interleukin 12 levels appeared protective. These results suggest specific immune responses may either contribute to, or protect against, transplacental virus transmission. The WUR10000125 SNP on chromosome 4, associated with PRRS resilience in nursery pigs, was not associated with reproductive outcome. Whereas past research suggested that fetal death results from events occurring at the maternal-fetal interface, we conclude that viral replication within fetuses and spread of PRRSV to adjacent fetuses are pivotal events in the pathogenesis of reproductive PRRS.

PCV2 vaccination induces IFN-γ/TNF-α co-producing T cells with a potential role in protection.

Porcine circovirus type 2 (PCV2) is one of the economically most important pathogens for swine production worldwide. Vaccination is a powerful tool to control porcine circovirus diseases (PCVD). However, it is not fully understood how PCV2 vaccination interacts with the porcine immune system. Especially knowledge on the cellular immune response against PCV2 is sparse. In this study we analysed antigen-specific T cell responses against PCV2 in a controlled vaccination and infection experiment. We focused on the ability of CD4(+) T cells to produce cytokines using multicolour flow cytometry (FCM). Vaccination with a PCV2 subunit vaccine (Ingelvac CircoFLEX®) induced PCV2-specific antibodies only in five out of 12 animals. Conversely, vaccine-antigen specific CD4(+) T cells which simultaneously produced IFN-γ and TNF-α and had a phenotype of central and effector memory T cells were detected in all vaccinated piglets. After challenge, seroconversion occurred earlier in vaccinated and infected pigs compared to the non-vaccinated, infected group. Vaccinated pigs were fully protected against viremia after subsequent challenge. Therefore, our data suggests that the induction of IFN-γ/TNF-α co-producing T cells by PCV2 vaccination may serve as a potential correlate of protection for this type of vaccine.

Magnitude and kinetics of multifunctional CD4+ and CD8ß+ T cells in pigs infected with swine influenza A virus.

Although swine are natural hosts for influenza A viruses, the porcine T-cell response to swine influenza A virus (FLUAVsw) infection has been poorly characterized so far. We have studied Ki-67 expression and FLUAVsw-specific production of IFN-γ, TNF-α and IL-2 in CD4(+) and CD8ß(+) T cells isolated from piglets that had been intratracheally infected with a H1N2 FLUAVsw isolate. IFN-γ(+)TNF-α(+)IL-2(+) multifunctional CD4(+) T cells were present in the blood of all infected animals at one or two weeks after primary infection and their frequency increased in four out of six animals after homologous secondary infection. These cells produced higher amounts of IFN-γ, TNF-α and IL-2 than did CD4(+) T cells that only produced a single cytokine. The vast majority of cytokine-producing CD4(+) T cells expressed CD8α, a marker associated with activation and memory formation in porcine CD4(+) T cells. Analysis of CD27 expression suggested that FLUAVsw-specific CD4(+) T cells included both central memory and effector memory populations. Three out of six animals showed a strong increase of Ki-67(+)perforin(+) CD8ß(+) T cells in blood one week post infection. Blood-derived FLUAVsw-specific CD8ß(+) T cells could be identified after an in vitro expansion phase and were multifunctional in terms of CD107a expression and co-production of IFN-γ and TNF-α. These data show that multifunctional T cells are generated in response to FLUAVsw infection of pigs, supporting the idea that T cells contribute to the efficient control of infection.

Emergence of porcine epidemic diarrhea virus in southern Germany.

BACKGROUND: Over the last years, porcine epidemic diarrhea virus (PEDV) has caused devastating enteric diseases in the US and several countries in Asia, while outbreaks in Europe have only been reported sporadically since the 1980s. At present, only insufficient information is available on currently circulating PEDV strains in Europe and their impact on the European swine industry. In this case report, we present epidemic outbreaks of porcine epidemic diarrhea in three farms in South-Western Germany. CASE PRESENTATION: Epidemic outbreaks of diarrhea affecting pigs of all age groups were reported in three farms, one fattening farm and two piglet producing farms, in South-Western Germany between May and November 2014. In the fattening farm yellowish, watery diarrhea without evidence of mucus or blood was associated with a massive reduction of feed consumption. Severity of clinical signs and mortality in young suckling pigs varied significantly between the two affected sow farms. While mortality in suckling piglets reached almost 70 % in one sow herd, no increase in suckling piglet mortality was observed in the second sow farm. In all three cases, PEDV was confirmed in feces and small intestines by RT-qPCR. Phylogenetic analyses based on full-length PEDV genomes revealed high identity among strains from all three herds. Moreover, the German strains showed very high nucleotide identity (99.4 %) with a variant of PEDV (OH851) that was isolated in the United States in January 2014. This strain with insertions and deletions in the S-gene (so called INDEL strains) was reported to show lower virulence. Slightly lower identities were found with other strains from the US and Asia. CONCLUSION: Phylogenetic information on the distribution of PEDV strains in Europe is severely lacking. In this case report we demonstrate that acute outbreaks of PEDV occurred in southern Germany in 2014. Current strains were clearly different from isolates found in the 1980s and were closely related to a PEDV variant found in the US in 2014. Moreover, the present case report indicates that variant strains of PEDV, containing insertions and deletions in the S gene, which were reported to be of lower virulence, might be able to cause high mortality in suckling piglets.

Associations among Haemophilus parasuis, Mycoplasma hyorhinis, and porcine reproductive and respiratory syndrome virus infections in pigs with polyserositis.

The objective of this study was to determine the associations among Haemophilus parasuis, Mycoplasma hyorhinis, and porcine reproductive and respiratory syndrome (PRRS) virus (EU-field strain) infections in 95 pigs with polyserositis. A significant association between H. parasuis and M. hyorhinis was identified. H. parasuis and M. hyorhinis were significantly more often detected in PRRS virus positive pigs.

Association entre infections avecHaemophilus parasuis, Mycoplasma hyorhinis,et le virus du syndrome dysgénésique et respiratoire du porc chez les porcs atteints de polysérosite. L'objectif de l'étude était d'étudier l'association de Haemophilus parasuis, Mycoplasma hyorhinis, et du SDRP (souche européenne-sauvage) chez 95 porcs atteints de polysérosite. Une association significative a été mise en évidence entre H. parasuis et M. hyorhinis. H. parasuis et M. hyorhinis ont été significativement plus fréquemment détectés chez SDRP positif porcs.(Traduit par les auteurs).

Different sample types in pigs challenged with Haemophilus parasuis following two treatment schemes with tulathromycin.

This study aimed to test the efficacy of samplings for the detection of Haemophilus parasuis after metaphylactic treatment and subsequent challenge using an established model for Glässer's disease. In this model, 36 piglets were equally assigned to a negative control, a positive control, and two trial groups receiving tulathromycin 7 or 4 days prior to challenge. The piglets of three groups were challenged intratracheally with H. parasuis serovar 5. As a result, four pigs in each challenged group died or had to be euthanised within 10 days post challenge. The remaining 15 pigs of these challenged groups survived until termination of the experiment (days 14-15). All pigs were necropsied and collective swabs of serosal surfaces were tested by bacterial culture and PCR. Samples of tarsal synovial fluid and joint capsule, cerebrospinal fluid (CSF) and brain swabs were tested by PCR. A total of 22 out of the 27 challenged animals had macroscopically detectable polyserositis and all of them tested positive in the collective swab samples. Haemophilus parasuis was more frequently detected in pigs that died within the first 10 days compared to those surviving until days 14-15 (P < 0.001), and those that succumbed within 10 days showed higher positivity rates in the brain and CSF. All pigs which were positive in the CSF had detectable meningitis. At days 14-15, joint samples from 5 of the remaining 15 pigs tested positive for H. parasuis. Four of these five animals did not show any macroscopic or histological lesions in the joints. In conclusion, collective swabs were the best sample material in acute cases, whereas samples from the joints gave the best results in chronic cases. In this challenge model it was not possible to prove the metaphylactic effect of tulathromycin administered 4 and 7 days prior to infection with H. parasuis.

Changes in leukocyte subsets of pregnant gilts experimentally infected with porcine reproductive and respiratory syndrome virus and relationships with viral load and fetal outcome.

In spite of more than two decades of extensive research, the understanding of porcine reproductive and respiratory syndrome virus (PRRSv) immunity is still incomplete. A PRRSv infection of the late term pregnant female can result in abortions, early farrowings, fetal death, and the birth of weak, congenitally infected piglets. The objectives of the present study were to investigate changes in peripheral blood mononuclear cell populations in third trimester pregnant females infected with type 2 PRRSv (NVSL 97-7895) and to analyze potential relationships with viral load and fetal mortality rate. PRRSv infection caused a massive, acute drop in total leukocyte counts affecting all PBMC populations by two days post infection. Except for B cells, cell counts started to rebound by day six post infection. Our data also show a greater decrease of naïve B cells, T-helper cells and cytolytic T cells than their respective effector or memory counterparts. Absolute numbers of T cells and γδ T cells were negatively associated with PRRSv RNA concentration in gilt serum over time. Additionally, absolute numbers of T helper cells may be predictive of fetal mortality rate. The preceding three leukocyte populations may therefore be predictive of PRRSv-related pathological outcomes in pregnant gilts. Although many questions regarding the immune responses remain unanswered, these findings provide insight and clues that may help reduce the impact of PRRSv in pregnant gilts.

Cytokine profiles in pregnant gilts experimentally infected with porcine reproductive and respiratory syndrome virus and relationships with viral load and fetal outcome.

In spite of extensive research, immunologic control mechanisms against Porcine Reproductive and Respiratory Syndrome virus (PRRSv) remain poorly understood. Cytokine responses have been exhaustively studied in nursery pigs and show contradictory results. Since no detailed reports on cytokine responses to PRRSv in pregnant females exist, the objectives of this study were to compare host cytokine responses between PRRSv-infected and non-infected pregnant gilts, and to investigate relationships between cytokine levels in infected gilts and viral load or fetal mortality rate. Serum samples and supernatants of peripheral blood mononuclear cells (PBMC) either stimulated with PRRSv or phorbol myristate acetate/Ionomycin (PMA/Iono) were analyzed for cytokines/chemokines: interleukins (IL) 1-beta (IL1ß), IL4, IL8, IL10, IL12, chemokine ligand 2 (CCL2), interferon alpha (IFNα) and interferon gamma (IFNγ). Three cytokines (IFNα, CCL2, IFNγ) in gilt serum differed significantly in inoculated versus control gilts over time. In supernatants of PRRSv stimulated PBMC from PRRSv-infected gilts, levels of IFNα were significantly decreased, while IL8 secretion was significantly increased. PRRSv infection altered the secretion of all measured cytokines, with the exception of IFNα, from PBMC after mitogen stimulation, indicating a possible immunomodulatory effect of PRRSv. IFNα, CCL2, and IFNγ in serum, and IFNγ in supernatants of PMA/Iono stimulated PBMC were significantly associated with viral load in tissues, serum or both. However, only IFNα in supernatants of PRRSv stimulated PBMC was significantly associated with fetal mortality rate. We conclude that of the eight cytokines tested in this study IFNα was the best indicator of viral load and severity of reproductive PRRSv infection.