Testing of umbilical cords by real time PCR is suitable for assessing vertical transmission of porcine reproductive and respiratory syndrome virus under field conditions

The objective of this study was to test the suitability of umbilical cord (UC) sampling and ear vein swabbing (EVS) as alternatives to jugular vein bleeding (JVB) for the assessment of vertical transmission of porcine reproductive and respiratory syndrome virus (PRRSV). Twelve farms suspected to be PRRSV-positive unstable were selected and the three types of samples were obtained from 21 batches of newborn piglets (n = 387). The proportions of positive results, viral loads and time spent to collect the samples were compared. UC yielded the highest detection rate, with 76 positives compared to 55 JVB- and 45 EV-positive results (P < 0.05). Average Ct values were 26.6 ± 8.5 for JVB, 30.8 ± 6.4 for EV and 32.1 ± 4.85 for UC (P < 0.01). UC was the fastest collection method (mean 24 s vs. 55 s for EV and 72 s for JVB; P < 0.05). In this study, UC testing was a faster and more sensitive alternative to JVB or EV for the detection of PRRSV in newborn piglets.info:eu-repo/semantics/acceptedVersio

Prevalence of enteric pathogens in diarrheic and non-diarrheic samples from pig farms with neonatal diarrhea in the North East of Spain

Diarrhea is one of the major causes of neonatal mortality in pigs. In the present study, 31 pig farms with outbreaks of neonatal diarrhea were investigated in Catalonia (NE Spain) from February 2017 until June 2018. Two hundred and fifteen diarrheic samples from 1 to 7 days old piglets were tested for a panel of enteric pathogens. In 19 of the studied farms additional fecal samples from apparently healthy pen-mates were collected and tested for the same panel of infectious agents. Samples were bacteriologically cultured and tested by PCR for E. coli virulence factors genes, C. perfringens types A and C toxins (Cpα, Cpβ, Cpβ2) and C. difficile toxins (TcdA, TcdB). Moreover, Rotavirus A (RVA), Rotavirus B (RVB), Rotavirus C (RVC), porcine epidemic diarrhea virus (PEDV) and transmissible gastroenteritis virus (TGEV) were also determined by RT-qPCR. More than one pathogen could be detected in all of the outbreaks. Nevertheless, RVA was the only agent that could be statistically correlated with the outcome of diarrhea. For the other viruses and bacteria analyzed significant differences between the diseased pigs and the controls were not found. In spite of this, the individual analysis of each of the studied farms indicated that other agents such as RVB, RVC, toxigenic C. difficile or pathogenic E. coli could play a relevant role in the outbreak of diarrhea. In conclusion, the large diversity of agent combinations and disease situations detected in neonatal diarrhea outbreaks of this study stand for a more personalized diagnosis and management advice at a farm level.info:eu-repo/semantics/acceptedVersio

Using commercial ELISAs to assess humoral response in sows repeatedly vaccinated with modified live porcine reproductive and respiratory syndrome virus

Background Sows in breeding herds are often mass vaccinated against porcine reproductive and respiratory syndrome (PRRS) every few months using modified live vaccines (MLV). Field veterinarians repeatedly report that multiple vaccinated sows test negative in ELISA. Obviously, this creates uncertainty when assessing the compliance of vaccination and the status of sows. Methods In the present study, four commercial ELISAs were used to assess the serological PRRS status in gilts and sows of three farms that were PRRS MLV vaccinated every four months. Animals were tested before vaccination (BV) and postvaccination (PV). Total and neutralising antibodies and cell-mediated responses were also measured in animals that yielded negative results in all ELISAs. Results The proportion of seronegative animals BV varied depending on the farm and the ELISA used. When samples were analysed using only one ELISA, a substantial number of negative results obtained BV remained as negative afterwards. Five animals were negative BV and PV with all the examined ELISAs. Those animals also yielded negative results in all the other immunological assays. Conclusion Our findings suggest that the use of ELISA for monitoring multiple PRRS MLV vaccinated sows is very limited due to the variability of the humoral responses and the moderate agreement between tests.info:eu-repo/semantics/acceptedVersio

Welfare Benefits of Intradermal Vaccination of Piglets

Vaccination is reported as a stressful and painful event for animals. This study investigated whether needle-free intradermal vaccination improves the welfare of weaned pigs through the reduction of stress and pain biomarkers and improvement of behavioural parameters compared to traditional intramuscular injection with a needle. A total of 339 weaned piglets were allocated to 3 treatment groups: Intradermal Application of Liquids (IDAL) pigs, vaccinated against Porcine Circovirus type 2 (PCV2) by means of intradermal vaccination using a needle-free device Porcilis® PCV ID; Intramuscular (IM) pigs vaccinated against PCV2 with Porcilis® PCV intramuscularly with a needle; CONTROL pigs were managed identically but did not receive any vaccine injection. At the time of the injection, the reaction of IDAL piglets was similar to control piglets, whereas a greater percentage of piglets vaccinated intramuscularly displayed high-pitch vocalizations (7% CONTROL, 7% IDAL, 32% IM) and retreat attempts (3% CONTROL, 7% IDAL, 39% IM). The day after vaccination, IDAL piglets did not differ from the control piglets for any of the behavioural variables studied through scan samplings. IM piglets showed a lower frequency of social negative interactions (p = 0.001) and rope manipulation (p = 0.04) compared to the CONTROL group. Resting postures did not differ between treatments. At 28 h post-vaccination, IDAL piglets presented lower blood C-reactive protein levels (CONTROL = 20 μg/mL; IDAL = 39 μg/mL; IM = 83 μg/mL, p < 0.0001) and blood Haptoglobin (CONTROL = 1.8 mg/mL; IDAL = 1.9 mg/mL vs. IM = 3.1 mg/mL, p < 0.0001) compared to IM piglets. Salivary chromogranin A and alpha-amylase did not differ between treatment groups when measured 25 min post-vaccination. The method of vaccination did not affect the growth of the piglets or their rectal temperature. These results support that needle-free intradermal vaccination reduces vaccination-related pain in growing pigs.info:eu-repo/semantics/publishedVersio

Diversity of respiratory viruses present in nasal swabs under influenza suspicion in respiratory disease cases of weaned pigs

Respiratory diseases in weaned pigs are a common problem, with a complex etiology involving both viruses and bacteria. In the present study, we investigated the presence of eleven viruses in nasal swabs, collected from nurseries (55 cases) under the suspicion of swine influenza A virus (swIAV) and submitted by swine veterinarians for diagnosis. The other ten viruses included in the study were influenza B (IBV) and D (IDV), Porcine reproductive and respiratory syndrome virus (PRRSV), Porcine respiratory coronavirus (PRCV), Porcine cytomegalovirus (PCMV), Porcine circovirus 2 (PCV2), 3 (PCV3) and 4 (PCV), Porcine parainfluenza 1 (PPIV1) and Swine orthopneumovirus (SOV). Twenty-six swIAV-positive cases and twenty-nine cases of swIAV-negative respiratory disease were primarily established. While IBV, IDV, PCV4 and PPIV1 were not found in any of the cases, PRCV, SOV, and PCMV were more likely to be found in swIAV-positive nurseries with respiratory disease (p < 0.05). Overall, PCV3, PRRSV, and PCMV were the most frequently detected agents at herd level. Taken individually, virus prevalence was: swIAV, 48.6%; PRCV, 48.0%; PRRSV, 31.6%; SOV, 33.8%; PCMV, 48.3%, PCV2, 36.0%; and PCV3, 33.0%. Moreover, low Ct values (<30) were common for all agents, except PCV2 and PCV3. When the correlation between pathogens was individually examined, the presence of PRRSV was negatively correlated with swIAV and PRCV, while was positively associated to PCMV (p < 0.05). Also, PRCV and SOV were positively correlated between them and negatively with PCMV. Besides, the analysis of suckling pig samples, collected in subclinically infected farrowing units under an influenza monitoring program, showed that circulation of PRCV, PCMV, SOV, and PCV3 started during the early weeks of life. Interestingly, in those subclinically infected units, none of the pathogens was found to be correlated to any other. Overall, our data may contribute to a better understanding of the complex etiology and epidemiology of respiratory diseases in weaners. This is the first report of SOV in Spain and shows, for the first time, the dynamics of this pathogen in swine farms.info:eu-repo/semantics/publishedVersio

Welfare benefits of intradermal vaccination of piglets

Vaccination is reported as a stressful and painful event for animals. This study investigated whether needle-free intradermal vaccination improves the welfare of weaned pigs through the reduction of stress and pain biomarkers and improvement of behavioural parameters compared to traditional intramuscular injection with a needle. A total of 339 weaned piglets were allocated to 3 treatment groups: Intradermal Application of Liquids (IDAL) pigs, vaccinated against Porcine Circovirus type 2 (PCV2) by means of intradermal vaccination using a needle-free device Porcilis® PCV ID; Intramuscular (IM) pigs vaccinated against PCV2 with Porcilis® PCV intramuscularly with a needle; CONTROL pigs were managed identically but did not receive any vaccine injection. At the time of the injection, the reaction of IDAL piglets was similar to control piglets, whereas a greater percentage of piglets vaccinated intramuscularly displayed high-pitch vocalizations (7% CONTROL, 7% IDAL, 32% IM) and retreat attempts (3% CONTROL, 7% IDAL, 39% IM). The day after vaccination, IDAL piglets did not differ from the control piglets for any of the behavioural variables studied through scan samplings. IM piglets showed a lower frequency of social negative interactions (p = 0.001) and rope manipulation (p = 0.04) compared to the CONTROL group. Resting postures did not differ between treatments. At 28 h post-vaccination, IDAL piglets presented lower blood C-reactive protein levels (CONTROL = 20 µg/mL; IDAL = 39 µg/mL; IM = 83 µg/mL, p < 0.0001) and blood Haptoglobin (CONTROL = 1.8 mg/mL; IDAL = 1.9 mg/mL vs. IM = 3.1 mg/mL, p < 0.0001) compared to IM piglets. Salivary chromogranin A and alpha-amylase did not differ between treatment groups when measured 25 min post-vaccination. The method of vaccination did not affect the growth of the piglets or their rectal temperature. These results support that needle-free intradermal vaccination reduces vaccination-related pain in growing pigs

In the presence of non-neutralising maternally derived antibodies, intradermal and intramuscular vaccination with a modified live vaccine against porcine reproductive and respiratory syndrome virus 1 (PRRSV-1) induce similar levels of neutralising antibodies or interferon-gamma secreting cells

The purpose of this study was to compare the immune response generated by the intramuscular and the intradermal vaccination route against the porcine reproductive and respiratory syndrome virus (PRRSV). Piglets from a seronegative and a seropositive farm were selected (n = 28 piglets per farm), and each group was divided into two groups and vaccinated after weaning with modified live vaccine Unistrain® PRRS (Laboratorios Hipra Amer, Spain) by the intramuscular (IM) or the intradermic (ID) route. For the following 6 weeks, animals were weekly bled to assess the humoral response by PRRSV-specific antibody ELISA and viral neutralisation test. At 0-, 3-, 4- and 6 weeks post-vaccination, peripheral mononuclear blood cells (PBMC) from eight animals per group were recovered to analyse cellular response by IFN-γ ELISPOT and lymphoproliferation. Serum IL-12 was also quantified by ELISA. Seroconversion was first detected 14 days post-vaccination (dpv) for both IM and ID routes, and peaked at 35 dpv (both IM groups and ID seropositive) or 42 dpv (ID seronegative). At 3 weeks after vaccination, 6/27 (22.22%) animals from negative origin had not seroconverted, and neutralising titres were significantly lower at 35 dpv compared to the seropositive origin (mean log2 titres of 1.36 and 4.25 respectively) Also, it was 10 times more probable for them to have high levels of IL-12 a week after vaccination than for animals of seropositive origin. Cellular immune response analysed by lymphoproliferation and IFN-γ ELISPOT was already present at 21 dpv and until 42 dpv, with no significant differences between groups except for a higher lymphoproliferation at 35 dpv in the IM seropositive group (Kruskal-Wallis, p < 0.05). These results indicate that the intradermal route induces an immune response equivalent to the classical intramuscular route even in presence of non-neutralising maternal immunity, which in this study has proven to facilitate seroconversion after vaccination with an heterologous strain

Bottlenecks in the transmission of porcine reproductive and respiratory syndrome virus (PRRSV1) to naïve pigs and the quasi-species variation of the virus during infection in vaccinated pigs

This paper describes the results of two experiments regarding porcine reproductive and respiratory syndrome virus (PRRSV1): the first one studied the existence of bottlenecks in an experimental one-to-one model of transmission in pigs; while the second analysed the differences between viral quasi-species in vaccinated pigs that developed shorter or longer viraemias after natural challenge. Serum samples, as well as the initial inoculum, were deep-sequenced and a viral quasi-species was constructed per sample. For the first experiment, the results consistently reported a reduction in the quasi-species diversity after a transmission event, pointing to the existence of bottlenecks during PRRSV1 transmission. However, despite the identified preferred and un-preferred transmitted variants not being randomly distributed along the virus genome, it was not possible to identify any variant producing a structural change in any viral protein. In contrast, the mutations identified in GP2, nsp9 and M of the second experiment pointed to changes in the amino acid charges and the viral RNA-dependent RNA polymerase structure. The fact that the affected proteins are known targets of the immunity against PRRSV, plus the differential level of neutralizing antibodies present in pigs developing short or long viraemias, suggests that the immune response selected those changes.info:eu-repo/semantics/publishedVersio

Introduction of a PRRSV-1 strain of increased virulence in a pig production structure in Spain : virus evolution and impact on production

A strain of Porcine reproductive and respiratory syndrome virus (PRRSV), showing characteristics of enhanced virulence, affected a pyramidal production system from Spain with 7600 sows in 4 genetic nuclei and 13,000 sows in multipliers. Different PRRSV strains circulating in this production system from December 2020 to October 2021 were detected and sequenced. The spread of each isolate was examined and their impact on health and production in three of the affected farms was evaluated. The newly emerged PRRSV isolate with enhanced virulence entered the system before the onset of the study (January 2020) and afterwards four significantly different clades were detected during the study period in different farms, probably because of independent introduction events. The diversification of the enhanced virulence strain was higher for those clades (substitution rates up to 1.1% nucleotides/year) compared to other PRRSV strains present in the production system (up to 0.17%), suggesting a faster spread and adaptation. The impact of the infection in the first affected farm was dramatic, with an average abortion rate above 27% during 17 weeks before returning to the baseline production. Fertile sow mortality reached 6.5% for 39 weeks. In two farms infected later by other clades of this enhanced virulence strain, the impact was less acute; despite the fact that for parameters such as the proportion of stillbirths or mummies, more than 10 months were needed to recover pre-outbreak values. In the examined nurseries, mortalities reached peaks between 28 and 50% and several months were needed to return to normality. Introduction of a PRRSV strain of enhanced virulence in a production system where several farms were previously positive for other PRRSV strains, resulted in a fast spread such as would be observed in naïve farms. The productive and health impact was very high taking several months to return to normality. The online version contains supplementary material available at 10.1186/s40813-022-00298-3

Diversity of respiratory viruses present in nasal swabs under influenza suspicion in respiratory disease cases of weaned pigs

Respiratory diseases in weaned pigs are a common problem, with a complex etiology involving both viruses and bacteria. In the present study, we investigated the presence of eleven viruses in nasal swabs, collected from nurseries (55 cases) under the suspicion of swine influenza A virus (swIAV) and submitted by swine veterinarians for diagnosis. The other ten viruses included in the study were influenza B (IBV) and D (IDV), Porcine reproductive and respiratory syndrome virus (PRRSV), Porcine respiratory coronavirus (PRCV), Porcine cytomegalovirus (PCMV), Porcine circovirus 2 (PCV2), 3 (PCV3) and 4 (PCV), Porcine parainfluenza 1 (PPIV1) and Swine orthopneumovirus (SOV). Twenty-six swIAV-positive cases and twenty-nine cases of swIAV-negative respiratory disease were primarily established. While IBV, IDV, PCV4 and PPIV1 were not found in any of the cases, PRCV, SOV, and PCMV were more likely to be found in swIAV-positive nurseries with respiratory disease (p < 0.05). Overall, PCV3, PRRSV, and PCMV were the most frequently detected agents at herd level. Taken individually, virus prevalence was: swIAV, 48.6%; PRCV, 48.0%; PRRSV, 31.6%; SOV, 33.8%; PCMV, 48.3%, PCV2, 36.0%; and PCV3, 33.0%. Moreover, low Ct values (<30) were common for all agents, except PCV2 and PCV3. When the correlation between pathogens was individually examined, the presence of PRRSV was negatively correlated with swIAV and PRCV, while was positively associated to PCMV (p < 0.05). Also, PRCV and SOV were positively correlated between them and negatively with PCMV. Besides, the analysis of suckling pig samples, collected in subclinically infected farrowing units under an influenza monitoring program, showed that circulation of PRCV, PCMV, SOV, and PCV3 started during the early weeks of life. Interestingly, in those subclinically infected units, none of the pathogens was found to be correlated to any other. Overall, our data may contribute to a better understanding of the complex etiology and epidemiology of respiratory diseases in weaners. This is the first report of SOV in Spain and shows, for the first time, the dynamics of this pathogen in swine farms