Duration of maternally derived antibodies of porcine parvovirus in growing pigs and presence of antibodies in gilts and sows vaccinated with three different parvovirus vaccines.

While gilts and sows are regularly vaccinated against the porcine parvovirus (PPV), little is known on the presence of antibodies in vaccinated sows nor the decline of maternally derived antibodies (MDA) in their offspring. On twelve farms serum samples were taken from 180 gilts and sows vaccinated at least twice with one of three different commercial PPV vaccines. On nine farms, additional 270 serum samples were collected from growing pigs of three different age categories. All 450 samples were examined for PPV antibodies (Abs) by ELISA and haemagglutination inhibition (HI) assay. In total, 65% of all gilts vaccinated twice with either vaccine 1 or vaccine 3 were seronegative by HI assay. In each farm, there were at least three animals with high Ab titres (≥ 1:1280) indicating the presence of PPV in all twelve study farms. However, PPV DNA could not be detected in collected faecal samples. While low to moderately high Ab titres (1:10-1:640) were measured in 98% of twelve-weeks-old pigs, ELISA was only positive in 30% of the same pigs. Though, the statement on the duration of MDA may depend on the applied test, we could confirm an exponential decay of MDA. In addition, we could demonstrate that applied serological tools are insufficient for the confirmation of successful vaccination.

Chlamydia suis survival in dust: First insights.

Chlamydia (C.) suis, a zoonotic intracellular bacterium, is described as a causative agent for conjunctivitis, particularly in nursery and fattening pigs. Chlamydiaceae are claimed to survive drying and to persist in dust. The objective of this study was to evaluate the viability of Chlamydia in dust sampled in a fattening pig farm with a high appearance of chlamydial-induced conjunctivitis. Dust was collected and stored at room temperature. To evaluate bacterial load and survival over time, quantitative PCR (Chlamydiaceae, C. suis) and isolation in cell culture were performed every week for up to 16 weeks. While qPCR results remained highly positive with consistent bacterial loads between 103 and 104 copy numbers/100 µL eluate over a period of 16 weeks and even after 40 weeks, it was not possible to isolate Chlamydia except for the initial sample. These results show only short-term viability of C. suis in dust. This is an important information regarding reduction of chlamydial loads in pig farms and risk for pigs and people to get infected via dust.

Influence of PRRSV-1 vaccination and infection on mononuclear immune cells at the maternal-fetal interface.

Porcine reproductive and respiratory syndrome virus (PRRSV) is one of the most devastating viruses for the global swine industry. Infection during late gestation causes reproductive failure but the local immune response in utero remains poorly understood. In this study, an experimental PRRSV-infection model with two different PRRSV-1 field isolates was used to investigate the immune cell phenotypes at the maternal-fetal interface during late gestation. In addition, phenotypic changes induced by a modified live virus (MLV, ReproCyc® PRRS EU) vaccine were studied. Vaccinated (n = 12) and non-vaccinated pregnant gilts (n = 12) were challenged with either one of the PRRSV-1 field isolates (low vs. high virulent, LV or HV) or sham-inoculated at day 84 of gestation. Twenty-one days post infection all gilts were euthanized and the fetal preservation status for all fetuses per litter was assessed. Leukocytes from the maternal-fetal interface were isolated and PRRSV-induced changes were investigated using ex vivo phenotyping by flow cytometry. PRRSV load in tissue from the maternal endometrium (ME) and fetal placenta (FP) was determined by RT-qPCR. In the ME, a vast increase in CD8ß T cells with CD8αposCD27dim early effector phenotype was found for fetuses from the non-vaccinated LV and HV-challenged gilts, compared to non-treated and vaccinated-only controls. HV-challenged fetuses also showed significant increases of lymphocytes with effector phenotypes in the FP, including NKp46pos NK cells, CD8αhigh γδ T cells, as well as CD8αposCD27pos/dim CD4 and CD8 T cells. In vaccinated animals, this common activation of effector phenotypes was more confined and the fetal preservation status significantly improved. Furthermore, a negative correlation between the viral load and CD163highCD169pos mononuclear phagocytic cells was observed in the FP of HV-infected animals. These results suggest that the strong expansion of effector lymphocytes in gilts that were only infected causes immune-pathogenesis rather than protection. In contrast, the attenuated MLV seems to dampen this effect, yet presumably induces memory cells that limit reproductive failure. This work provides valuable insights into changes of local immune cell phenotypes following PRRSV vaccination and infection.

Efficacy of a Modified Live Virus Vaccine against Porcine Reproductive and Respiratory Syndrome Virus 1 (PRRSV-1) Administered to 1-Day-Old Piglets in Front of Heterologous PRRSV-1 Challenge.

PRRSV is one of the most important viruses in the global swine industry and is often controlled by the use of modified live virus (MLV) vaccines. This study assessed the impact of a PRRSV-1 MLV vaccine applied to 1-day-old piglets challenged on day 28 of life with a PRRSV-1 field isolate (AUT15-33). Twenty-one piglets were vaccinated within 24 h of birth (T02), whereas 20 piglets were left unvaccinated (T01). Necropsy was performed two weeks post-challenge. Comparing the two groups, T02 piglets showed significantly higher (p = 0.017) average daily weight gain. In addition, significantly lower (p < 0.0001) PRRSV RNA loads were measured in serum of T02 piglets at all investigated time points. All T01 piglets were viremic and shed virus in nasal swabs, whereas only 71.4% and 38.1% of the T02 group were viremic or shed virus, respectively. Piglets from T02 had significantly higher numbers (p < 0.0001) of IFN-γ producing lymphocytes compared to T01. At necropsy, differences in gross and histologic lung lesions were statistically significant (p = 0.012 and p < 0.0001, respectively) between the two groups. Hence, this MLV vaccine administered to 1-day-old piglets was able to protect piglets against PRRSV infection at weaning.

Comparison of different detection methods for Ascaris suum infection on Austrian swine farms.

BACKGROUND: Ascaris suum, the large roundworm of pigs, is one of the economically most important pig parasites worldwide. In Austria it is commonly diagnosed by monitoring livers for milk spots at the slaughterhouse and intravital diagnosis (flotation for detection of fecal egg shedding). Recently, serological diagnosis based on the detection of specific antibodies with an ELISA (SERASCA®) with high sensitivity has been developed. To introduce and evaluate serology for A. suum screening in Austrian pigs, blood (for serology) (n = 177) and feces (for copromicroscopy) (n = 177) were taken from randomly selected slaughter pig batches from 18 farms at a slaughterhouse in Lower Austria. In addition, livers presented at slaughter (n = 844; max. 70/farm) were evaluated for milk spots. RESULTS: Overall, 19% of the livers were milk spot-positive (22% of those with complete diagnostic evaluations). Thirteen percent of the fecal samples contained A. suum eggs, while 69% of the blood samples were serologically positive. Despite we did not determine the sensitivity of the ELISA specifically, results ouf our study confirmed the high sensitivity of the ELISA, which was claimed by the manufacturer prior to our work (sensitivity: liver assessment: 23.5-27.0%; copromicroscopy: 8.5-9.0%; ELISA: 99.5%), and a high percentage of A. suum infections that remained undetected by standard liver assessment. CONCLUSIONS: This suggests that the current method of roundworm diagnostics is insufficient and antibody detection at the end of the fattening period should be established as the standard procedure.

T-Cell Cytokine Response in Salmonella Typhimurium-Vaccinated versus Infected Pigs.

Vaccination with the live attenuated vaccine Salmoporc is an effective measure to control Salmonella Typhimurium (STM) in affected swine populations. However, the cellular immune response evoked by the Salmoporc vaccine including differences in vaccinated pigs versus non-vaccinated pigs upon STM infection have not been characterized yet. To investigate this, tissue-derived porcine lymphocytes from different treatment groups (vaccination-only, vaccination and infection, infection-only, untreated controls) were stimulated in vitro with heat-inactivated STM and abundances of IFN-γ, TNF-α and/or IL-17A-producing T-cell subsets were compared across organs and treatment groups. Overall, our results show the induction of a strong CD4+ T-cell response after STM infection, both locally and systemically. Low-level induction of STM-specific cytokine-producing CD4+ T cells, notably for the IFN-γ/TNF-α co-producing phenotype, was detected after vaccination-only. Numerous significant contrasts in cytokine-producing T-cell phenotypes were observed after infection in vaccinated and infected versus infected-only animals. These results suggest that vaccine-induced STM-specific cytokine-producing CD4+ T cells contribute to local immunity in the gut and may limit the spread of STM to lymph nodes and systemic organs. Hence, our study provides insights into the underlying immune mechanisms that account for the efficacy of the Salmoporc vaccine.

NK and T Cell Differentiation at the Maternal-Fetal Interface in Sows During Late Gestation.

The phenotype and function of immune cells that reside at the maternal-fetal interface in humans and mice have been, and still are, extensively studied with the aim to fully comprehend the complex immunology of pregnancy. In pigs, information regarding immune cell phenotypes is limited and mainly focused on early gestation whereas late gestation has not yet been investigated. We designed a unique methodology tailored to the porcine epitheliochorial placenta, which allowed us to address immune phenotypes separately in the maternal endometrium (ME) and fetal placenta (FP) by flow cytometry. In-depth phenotyping of NK cells, non-conventional and conventional T cells within maternal blood (mBld), ME, FP, and fetal spleen (fSpln) revealed major differences between these anatomic sites. In both maternal compartments, all NK cells were perforin+ and had NKp46-defined phenotypes indicative of late-stage differentiation. Likewise, T cells with a highly differentiated phenotype including CD2+CD8α+CD27dim/-perforin+ γδ T cells, CD27-perforin+ cytolytic T cells (CTLs), and T-bet+ CD4+CD8α+CD27- effector memory T (Tem) cells prevailed within these compartments. The presence of highly differentiated T cells was also reflected in the number of cells that had the capacity to produce IFN-γ. In the FP, we found NK cells and T cell populations with a naive phenotype including CD2+CD8α-CD27+perforin- γδ T cells, T-bet-CD4+CD8α-CD27+ T cells, and CD27+perforin- CTLs. However, also non-naive T cell phenotypes including CD2+CD8α+CD27+perforin- γδ T cells, T-bet+CD4+CD8α+CD27- Tem cells, and a substantial proportion of CD27-perforin+ CTLs resided within this anatomic site. Currently, the origin or the cues that steer the differentiation of these putative effector cells are unclear. In the fSpln, NKp46high NK cells and T cells with a naive phenotype prevailed. This study demonstrated that antigen-experienced immune cell phenotypes reside at the maternal-fetal interface, including the FP. Our methodology and our findings open avenues to study NK and T cell function over the course of gestation. In addition, this study lays a foundation to explore the interplay between immune cells and pathogens affecting swine reproduction.

Comparison of clinical and immunological findings in gnotobiotic piglets infected with Escherichia coli O104:H4 outbreak strain and EHEC O157:H7.

BACKGROUND: Shiga toxin (Stx) producing Escherichia coli (E. coli) (STEC) is the most frequent cause of diarrhoea-positive haemolytic uraemic syndrome (D + HUS) in humans. In 2011, a huge outbreak with an STEC O104:H4 strain in Germany highlighted the limited possibilities for causative treatment of this syndrome. The responsible STEC strain was found to combine Stx production with adherence mechanisms normally found in enteroaggregative E. coli (EAEC). Pathotypes of E. coli evolve and can exhibit different adhesion mechanisms. It has been shown previously that neonatal gnotobiotic piglets are susceptible for infection with STEC, such as STEC O157:H7 as well as for EAEC, which are considered to be the phylogenetic origin of E. coli O104:H4. This study was designed to characterise the host response to infection with the STEC O104:H4 outbreak strain in comparison to an STEC O157:H7 isolate by evaluating clinical parameters (scoring) and markers of organ dysfunction (biochemistry), as well as immunological (flow cytometry, assessment of cytokines/chemokines and acute phase proteins) and histological alterations (light- and electron microscopy) in a gnotobiotic piglet model of haemolytic uraemic syndrome. RESULTS: We observed severe clinical symptoms, such as diarrhoea, dehydration and neurological disorders as well as attaching-and-effacing lesions (A/E) in the colon in STEC O157:H7 infected piglets. In contrast, STEC O104:H4 challenged animals exhibited only mild clinical symptoms including diarrhoea and dehydration and HUS-specific/severe histopathological, haematological and biochemical alterations were only inconsistently presented by individual piglets. A specific adherence phenotype of STEC O104:H4 could not be observed. Flow cytometric analyses of lymphocytes derived from infected animals revealed an increase of natural killer cells (NK cells) during the course of infection revealing a potential role of this subset in the anti-bacterial activity in STEC disease. CONCLUSIONS: Unexpectedly, E. coli O104:H4 infection caused only mild symptoms and minor changes in histology and blood parameters in piglets. Outcome of the infection trial does not reflect E. coli O104:H4 associated human disease as observed during the outbreak in 2011. The potential role of cells of the innate immune system for STEC related disease pathogenesis should be further elucidated.