Effect of ß-hydroxybutyric acid, parity, and body condition score on phenotype and proliferative capacity of colostral mononuclear leukocytes of high-yielding dairy cows.

In neonatal calves, the ingestion of colostrum is imperative for preventing infectious diseases. Investigations into the transfer of passive immunity of cattle have primarily focused on the importance of colostral immunoglobulins, with a recent increase in focus on understanding the role of colostral leukocytes. The main objective of the present study was to measure the influence of parity, body condition score, serum nonesterified fatty acids, and serum ß-hydroxybutyrate concentrations of periparturient cows on phenotype and mitogen- and antigen-induced proliferative capacity of bovine colostral leukocytes. Holstein-Friesian cows (n=141) were intramuscularly vaccinated at 60 and 30 d before the expected parturition date with a tetanus toxoid vaccine. Of these 141 animals, 28 primiparous and 72 multiparous cows were sampled immediately after parturition. Colostrum mononuclear cell populations were identified by flow cytometry using bovine cluster of differentiation markers, and the proliferative capacity of these cells was determined using a (3)H-thymidine proliferation assay. Under-conditioned cows had a significantly higher percentage of colostral macrophages than normal-conditioned animals, whereas over-conditioned cows had significantly more colostral B-lymphocytes. Serum ß-hydroxybutyrate was significantly associated with higher numbers of colostral T-lymphocytes and macrophages. Heifers had significantly higher mitogen- and antigen-induced proliferation of their colostral leukocytes than third parity or older cows. In conclusion, body condition score, parity, and serum ß-hydroxybutyrate concentration of periparturient high-yielding dairy cows were shown to influence the number of colostral macrophages or the mitogen- and antigen-induced proliferation of colostral leukocytes, possibly influencing the cellular immunity of the newborn calf.

Analysis of the mucosal immune responses induced by single and trickle infections with the bovine abomasal nematode Ostertagia ostertagi.

The purpose of this study was to provide more information on the kinetics of the immunological changes occurring in the abomasal mucosa after single and trickle infections with the bovine parasite Ostertagia ostertagi. The time course analysis of gene expression revealed that the major changes coincided with the emergence of adult worms from the gastric glands. These changes consisted of a simultaneous upregulation of Th1- and Th2-type cytokines. In addition, a single O. ostertagi infection elicited an upregulation of the epithelial-derived cytokine IL33, while TSLP expression levels were not impacted. Apart from the massive increase in inflammatory cytokines IL6, IL17 and IL21, O. ostertagi infection also elicited an upregulation of the immunosuppressors TGFB, IL10 and ARG1, as well as NK and γδ-T cell markers. Furthermore, the cytotoxic factors granulysin, perforin and granzyme B were upregulated following an O. ostertagi infection. Analysis of cytokine transcript levels in animals receiving trickle infections for 60 days showed a similar trend as observed following a single infection except for IL33, IL6, GATA-3, TBX21 and NCR1, which were no longer upregulated after trickle infections. Finally, the long trickle infections were associated with mucosal eosinophilia and mastocytosis.

Evaluation of different chromogenic media for the detection of methicillin-resistant Staphylococcus aureus CC398 in broilers.

Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) has emerged in a wide variety of animal species, including poultry. The objective of this study was to evaluate three different chromogenic media for MRSA clonal complex (CC) 398 detection in broilers. On three Belgian poultry farms, 50 broiler chickens were sampled per farm from both nose shell and cloaca. All swab specimens were enriched and inoculated the following day on three chromogenic media: chromID MRSA (bioMérieux), Brilliance MRSA 2 Agar (Oxoid) and MRSASelect (Bio-Rad). ChromID had the highest isolation rates, yet, Brilliance MRSA 2 Agar demonstrated the highest relative sensitivity, while MRSASelect and Brilliance MRSA 2 Agar showed the highest relative specificity. A subset of MRSA isolates was confirmed to be CC398 by the polymerase chain reaction (PCR) targeting sau1-hsdS1. In conclusion, Brilliance MRSA 2 Agar outperformed MRSASelect and chromID MRSA for the detection of MRSA in broilers.

Effect of a disinfection strategy on the methicillin-resistant Staphylococcus aureus CC398 prevalence of sows, their piglets and the barn environment.

AIMS: To assess, in a cleaned and disinfected barn environment, the efficacy of an animal disinfection strategy to reduce the livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) prevalence in sows, their offspring and the barn environment. METHODS AND RESULTS: On each farm, six sow rounds were sampled; sows were divided into either a test or control group. Per round, 20 sows and 40 of their piglets were sampled at different time points together with the barn environment. The disinfection strategy of the test groups consisted of washing the sows with a shampoo followed by disinfection of the skin with a solution containing chlorhexidine digluconate and isopropanol. On the first day of disinfection and 6 days after stopping the disinfection, a significant decrease (P < 0·01) of on average 68 and 66% in sow MRSA prevalence was observed on both farms, whereas no decrease was seen in the control groups. Just before weaning, 21-28 days after the end of the disinfection strategy, the difference in MRSA prevalence between both groups was reduced to 4% and no longer significant (P = 0·20). The MRSA prevalence of the piglets in the test groups was significantly lower (26%; P < 0·01) 6 days after the end of disinfection. Just before weaning, this difference was reduced to 5% but still significant (P < 0·01). In the swine nursery unit, no significant difference (P = 0·99) was seen between both groups. Based on semi-quantitative counts, a relationship (r(2)  > 0·6; P < 0·01) was seen between MRSA contamination in the barn environment and the MRSA prevalence in pigs. CONCLUSION: Results show that the tested disinfection strategy reduces temporarily the sow and piglet MRSA status, but does not result in a final reduction in MRSA at weaning or in the nursery unit. SIGNIFICANCE AND IMPACT OF THE STUDY: First report on the efficacy of an animal disinfection strategy to reduce LA-MRSA prevalence in sows, their offspring and the barn environment.

Seroepidemiological survey of trypanozoon infection in horses in the suspected dourine-infected Bale highlands of the Oromia region, Ethiopia.

This paper presents the results of a seroepidemiological survey of trypanozoon infection in horses carried out between September 2007 and June 2008. The survey was conducted to determine the seroprevalence of anti-trypanozoon antibodies in 880 serum samples collected randomly from selected horse-breeding districts of the Bale highlands of Ethiopia. The seroprevalence of trypanozoon infection was found to be 173 (19.66%) and 140 (15.91%) for the CATT/T. evansi and LATEX/T. evansi tests, respectively. The high seroprevalence of trypanozoon infection strongly indicates that the infection is endemic. Neither test can differentiate between anti-trypanozoon antibodies caused by infection with T. equiperdum (the causative agent of dourine) and those of T. evansi (the causative agent of surra). The findings of the present study suggest that field-applicable screening serological tests such as the CATT/T. evansi and LATEX/T. evansi could be useful for epidemiological studies and the control of trypanozoon infection.

Effect of beta-glucans on an ETEC infection in piglets.

The effect of orally administered beta-glucans in protecting pigs against an ETEC infection after weaning was analysed in this study. Three beta-glucans that differed in origin (Saccharomyces cerevisiae (MCG (Macrogard) and G2) or Sclerotium rolfsii (G3)) and/or extraction procedure were tested. Pigs fed for 2 weeks after weaning with these glucans were less susceptible to an F4+ ETEC infection in comparison with the control group. This was evidenced by a reduction in the faecal excretion of F4+ Escherichia coli as well as a reduced F4-specific serum antibody response. This decrease in faecal excretion was statistically significant for pigs fed with the MCG glucan in a first experiment and with the G3 glucan in a second experiment; diarrhoea was milder in the glucan-supplemented groups and was significantly reduced in the MCG-supplemented group. Furthermore, a lower amount of F4-specific IgM antibody-secreting cells (ASC) was found in the lymphoid tissues of pigs fed with G2 or G3 glucans in comparison with the control pig, as well as lower F4-specific IgA ASC in G3-fed pigs in comparison with the control pig. This study showed that beta-glucans can protect against an ETEC infection. Both MCG from S. cerevisiae and G3 from S. rolfsii, resulted in significant effects. To our knowledge, this is the first in vivo study, in which the use of beta-glucans as feed ingredient for just-weaned piglets was tested for their protective effects against ETEC infection.

Dietary ß-hydroxy-ß-methylbutyrate supplementation influences performance differently after immunization in broiler chickens.

Dietary addition of the leucine metabolite ß-hydroxy-ß-methylbutyrate (HMB) promotes growth in various species. In addition, HMB is described to enhance immune responses which might be associated with metabolic costs. We elaborated further on the role of HMB in growth, metabolism and immunity of meat-type chickens using the following parameters: zootechnical performance, blood chemistry and a specific immune responses after immunization with a human serum albumin (HSA)/Freund's (in) complete adjuvant combination. The chickens received commercial feeds either unsupplemented or supplemented with 300 mg HMB/kg feed. ß-hydroxy-ß-methylbutyrate-supplemented chickens were significantly heavier at 2 weeks of age but this difference was attenuated at later ages. Compared with their unsupplemented controls, cumulative feed conversion was significantly lower in HMB-supplemented chickens. There were no differences in blood chemistry between both dietary treatments. After immunization, HMB significantly attenuated the acute phase protein response at day 1 of post-immunization compared with that of their unsupplemented counterparts. After day 7 post-immunization, body weight gain of the immuno-challenged HMB-supplemented chickens was significantly depressed, but their specific anti-HSA IgG response was significantly enhanced compared with that of their immuno-challenged unsupplemented counterparts. The underlying mechanisms and signalling pathways for these phenomena need to be elucidated. Nevertheless, we are able to conclude that HMB is beneficial for performance under normal circumstances. On the other hand, HMB stimulates the immune response after an immunological challenge, though at the cost of reduced growth.

Effect of plasmid DNA encoding the porcine granulocyte-macrophage colony-stimulating factor on antigen-presenting cells in pigs.

We previously demonstrated that intradermal (ID) delivery of plasmid DNA encoding the porcine granulocyte-macrophage colony-stimulating factor (GM-CSF) 7 days before DNA vaccination enhances both cellular and humoral responses in pigs. In the present work, we studied the effect of the GM-CSF gene on antigen-presenting cells (APC) in pigs. We demonstrated that ID delivery of this gene significantly increased the number of epidermal CD1(+) cells (Langerhans' cells, skin dendritic cells) at the injection site at day 7. This was accompanied by an enhanced percentage of APC at the immune induction site following DNA vaccination, whereas a positive effect on APC maturation could not be demonstrated. Taken together, our data suggest that both DC recruitment to the immunization site and expansion of APC in the draining LN following DNA vaccination might contribute to the immune enhancing effect of plasmid encoded GM-CSF in pigs.

Screening of pigs resistant to F4 enterotoxigenic Escherichia coli (ETEC) infection.

The present study analysed quantitatively the mucin 4 polymorphism for determining the F4ac/ab receptor status of a total of 63 pigs by comparing it with the in vitro villous adhesion assay. The probability of a susceptible genotype for the mucin 4 increases significantly with increasing F4ab or F4ac ETEC adhesion per 250 microm villi (P=0.029 for F4ab, P=0.030 for F4ac), with the odds ratio for each unit increase of F4ab or F4ac equal to, respectively, 1.036 (95% CI [1.004-1.069]) and 1.018 (95% CI [1.002-1.034]). In the phenotypic in vitro villous adhesion test, a cut-off value of 5 bacteria was chosen as a criteria for the distinction between an F4R positive and F4R negative pig. The sensitivity and specificity for the in vitro villous adhesion test, with the genotyping test for mucin 4 as golden standard, is 100% and 24%, respectively, for F4ab as well as F4ac. Absence of adhesion of F4ac and F4ab ETEC to the villous brush borders was not associated with genotypic resistance suggesting that there is at least one other receptor for F4ab/ac Escherichia coli. As a consequence, not only mucin 4 gene polymorphism but also expression of these other receptor(s) has to be included in a screening assay for F4ac/ab receptor negative pigs.

The age-dependent expression of the F18+ E. coli receptor on porcine gut epithelial cells is positively correlated with the presence of histo-blood group antigens.

F18(+)Escherichia coli have the ability to colonize the gut and cause oedema disease or post-weaning diarrhoea by adhering to specific F18 receptors (F18R) on the porcine epithelium. Although it is well established that a DNA polymorphism on base pair 307 of the FUT1 gene, encoding an alpha(1,2)fucosyltransferase, accounts for the F18R phenotype, the F18R nature is not elucidated yet. The aim of the present study was to investigate the correlation between the presence of H-2 histo-blood group antigens (HBGAs) or its derivative A-2 HBGAs on the porcine gut epithelium and F18(+)E. coli adherence. A significant positive correlation was found between expression of both the H-2 (r=0.586, P<0.01) and A-2 (r=0.775, P<0.01) HBGAs and F18(+)E. coli adherence after examination of 74 pigs aged from 0 to 23 weeks. The majority of the genetically resistant pigs (FUT1M307(A/A)) showed no HBGA expression (91.7%) and no F18(+)E. coli adherence (83.3%). In addition, it was found that F18R expression levels rise with increasing age during the first 3 weeks after birth and that F18R expression is maintained in older pigs (3-23 weeks old). Taken together, these data suggest that, apart from H-2 HBGAs, A-2 HBGAs might be involved in F18(+)E. coli adherence.

Comparison of immune responses in parenteral FaeG DNA primed pigs boosted orally with F4 protein or reimmunized with the DNA vaccine.

We previously showed that an intradermal (i.d.) FaeG DNA prime (2x)-oral F4 protein boost immunization induces a systemic response and weakly primes a mucosal IgG response in pigs, especially when plasmid vectors encoding the A and B subunit of the E. coli thermo-labile enterotoxin (LT) are added to the DNA vaccine. In the present study, we evaluated whether addition of 1alpha,25-dihydroxyvitamin D(3) (vitD(3)) to the DNA vaccine could further enhance this mucosal priming and/or modulate the antibody response towards IgA. To further clarify priming of systemic and mucosal responses by the i.d. DNA vaccination, we firstly compared the localization of the F4-specific antibody response in pigs that were orally boosted with F4 to that in pigs that received a third i.d. DNA immunization and secondly evaluated cytokine mRNA expression profiles after i.d. DNA vaccination. The i.d. DNA prime (2x)-oral F4 boost immunization as well as the 3 i.d. DNA vaccinations induced mainly a systemic response, with a higher response observed following the heterologous protocol. Co-administration of vitD(3), and especially of the LT vectors, enhanced this response. Furthermore, only the heterologous immunization resulted in a weak mucosal priming, which appeared to require the presence of the LT vectors or vitD(3) as adjuvants. In addition, the LT vectors strongly enhanced the FaeG-specific lymphocyte proliferation and this was accompanied by the absence of a clear IL-10 response. However, despite two DNA immunizations in the presence of these adjuvants and an oral F4 boost, we failed to demonstrate the secretory IgA response needed to be protective against enterotoxigenic E. coli.

Mucosal immunization of piglets with purified F18 fimbriae does not protect against F18+ Escherichia coli infection.

Post-weaning diarrhoea and oedema disease in weaned piglets are caused by infection with F4+ or F18+ Escherichia coli strains. There is no commercial vaccine available, but it is shown that oral immunization of weaned piglets with purified F4 fimbriae induces a protective mucosal immune response. In the present study, piglets were orally and nasally immunized with purified F18 fimbriae in the presence of the mucosal adjuvant LT(R192G) or CTA1-DD, respectively. This immunization could not lead to protection against F18+ E. coli infection. The induced F18-specific immune response was directed towards the major subunit FedA and weakly towards the adhesive subunit FedF. The results of these experiments demonstrate that it is difficult to induce protective immunity against F18+ E. coli using the whole fimbriae due to the low response against the adhesin.

Perinatal priming of calves born to Schistosoma mattheei-infected dams.

The objective of this study was to elucidate whether calves born to infected dams had been primed against Schistosoma mattheei antigens. Infection-confirmed, pregnant cows were randomly selected for monitoring their offspring. Pre-colostral serum was collected from the neonates for the detection of specific antibodies at birth, as they indicate a transplacental transfer of schistosome-specific antibodies and antigen. At the age of approximately 2 months, peripheral blood mononuclear cells (PBMC) of calves were analysed for specific memory by antigen-specific stimulation in vitro. Twenty-six of the 30 calves demonstrated S. mattheei-specific proliferation. All 12 seropositive-born, as well as 14 of the 18 seronegative-born (before colostrum uptake) calves displayed mattheei-specific proliferation. The results indicate that the calves were primed against S. mattheei and might explain why seropositive-born calves from infected dams are better protected against S. mattheei, and query the impermeability of the damaged ruminant placenta with consequences for antigen transfer.

In vivo activation of chicken macrophages by infectious bursal disease virus.

Infectious bursal disease virus (IBDV) infects and replicates in the dividing B lymphocytes of chickens. In the present study, the in vivo effect of IBDV infection on chicken macrophage populations and macrophage activation were examined. Specific-pathogen-free chickens were exposed to virulent IBDV and splenic macrophages were recovered during the acute phase of the disease. At 3 and 5 days post-infection (dpi), spleens of virus-exposed chickens had fewer macrophages than those of virus-free controls (p < 0.05). Confocal microscopic examination revealed cells that were positive for both KUL01 (macrophage surface marker) and R63 (IBDVVP2), indicating presence of the virus in macrophages. MQ-NCSU cells, an avian macrophage cell line, were susceptible to replication of IBDV. In addition, splenic macrophages were activated and had temporarily increased levels of mRNA transcripts of pro-inflammatory mediators, including IL-1beta, IL-6, IL-18, and iNOS. The robust expression of proinflammatory cytokine transcripts, along with a decrease in macrophage numbers, suggest that IBDV activates and may lead to a reduction of resident macrophages in vivo.

Pathogenic interactions between Chlamydophila psittaci and avian pneumovirus infections in turkeys.

Both Chlamydophila psittaci and avian pneumovirus (APV) are highly prevalent in Belgian turkeys and might contribute to the respiratory disease complex observed in turkeys. Initial outbreaks of chlamydiosis occur mostly at the age of 4-8 weeks, often accompanied by an APV infection in APV non-vaccinated farms. Regardless APV vaccination, breakthroughs of APV infection from 8 weeks on do occur, a period when also a second C. psittaci infection appears. Therefore, this study examined the pathogenicity of an APV superinfection in C. psittaci predisposed turkeys. Turkeys were infected with C. psittaci, APV or with C. psittaci followed by APV. Simulating the impact of an APV infection during the acute phase or latent phase of a C. psittaci infection, turkeys have been infected with APV at 1 and 5 weeks post C. psittaci infection, respectively. APV infection during the acute phase of a C. psittaci infection aggravates the severity of clinical signs, macroscopic lesions, pharyngeal APV excretion and histological tracheae lesions. In contrast, no clear interaction could be established after APV infection in latently C. psittaci infected specific pathogen-free (SPF) turkeys. This study clearly demonstrates the exacerbating role of APV during acute C. psittaci infection, which can play an important role in the respiratory disease complex of turkeys.

Immunocompetence in organically fed finishing pigs: effect of corn cob mix.

Two consecutive experiments were performed to evaluate the effects on the immune response of corn cob mix (CCM) in an organic pig diet. The immunoglobulin (Ig) M, IgA and IgG responses against an intramuscularly injected model antigen, bovine thyroglobulin, were used as indicator. The experiments were performed in an organic barn with nine pens of four crossbred pigs (two barrows and two sows) from 45 kg to slaughter. In the first experiment, the organic concentrate was mixed with organic CCM-silage to obtain three concentrate: CCM ratios of 100:0, 80:20 and 60:40 (w:w). In the second experiment, three concentrates were produced to obtain diets with equal nutrient levels on a dry matter basis after 0%, 20% and 40% CCM inclusion. Higher inclusion rates of CCM in the ration were accompanied by lower thyroglobulin-specific IgG responses. These effects could not be attributed to one specific component of the CCM, such as fatty acid composition, although there was a degree of correlation with lower vitamin A concentrations. Mycotoxin concentrations were absent or minimal. The study indicated that dietary ingredient composition may affect immunocompetence.