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The majority of infectious bursal disease virus (IBDV) strains circulating in the broiler chicken industry in Canada are variant strains (varIBDV). Despite high levels of maternally derived antibodies (MtAb), the circulating varIBDVs can establish infection and cause severe immunosuppression in broiler chicks. The objective of this study was to evaluate circulating varIBDVs as broiler breeder vaccine candidates and investigate their protective efficacy against varIBDV challenge in their progeny chicks. Six groups of breeders (20 females/group) were vaccinated with varIBDV strains, SK09, SK10, SK11, SK12, and SK13 or saline at the age of 13 weeks and antibody response was determined by ELISA at 3-7-, and 20- weeks post-vaccination. We also included commercial chicks for the comparison. Results showed that SK-09 is the most antigenic strain, followed by SK-10, SK-12, and SK-13. In contrast, SK-11 showed the lowest antibody response, and over time, antibody titers steadily decreased. Eggs from breeders were collected at 21-week post-vaccination and incubated to produce their respective progenies. The serum antibody titer in day-old chicks showed a successful MtAb transfer. Progeny chicks (n = 40/group) were orally challenged with varIBDV-SK-09 strain at 6 days of age and serum antibody titer (19 d and 35 d of age), bursa to body weight ratio (19 d and 35 d of age), bursal viral load (9 d and 19 d of age) was examined to assess the protection against IBDV. Following the challenge, we found a significant increase in the antibody titers in MtAb-free and commercial vaccine groups than in the varIBDV groups, both at 19 d and 35 d of age. The BBW ratio and viral load data indicated a significant homologous and heterologous protection against varIBDV-SK-09 challenge by SK-09 and SK-10 MtAbs, respectively. Overall, this study demonstrated the feasibility of developing breeder vaccines using circulating varIBDV as candidate vaccine antigens.

Antimicrobial resistance (AMR) is a global issue, posing a grave threat to the public, animal, and environmental health. The AMR surveillance at the level of the hatchery is crucial to develop an AMR control strategy in the poultry industry. The objective of this study was to investigate the AMR profiles of bacteria isolated from yolk material of non-viable broiler chicken embryos at hatch from commercial hatcheries in western Canada. Antimicrobial susceptibility testing was done using the Kirby–Bauer disk diffusion method focusing on Escherichia coli (n = 170) and Enterococcus (n = 256) species, which are commonly used as indicators of AMR evolution. E. coli isolates were resistant to tetracycline, ampicillin, amoxycillin-clavulanic acid, triple sulpha, ceftiofur, gentamycin, and spectinomycin at the rate of 52.9%, 50.6%, 40.0% 31.8%, 29.4%, 29.4%, 21.8% respectively. Among those, 37.1% of E. coli were multidrug resistant. The descending order of antimicrobial resistance of E. faecalis was; tetracycline (61.9%), ceftiofur (46.2%), bacitracin (43.9%), erythromycin (31.4%) and tylosin (27.4%). Multidrug resistance was detected in 40.4% of E. faecalis isolates, and 85.7% of E. faecium isolates. To the best of our knowledge, this is the first report on AMR surveillance of non-viable chicken embryos. Overall, the present study revealed that non-viable chicken embryos, an overlooked niche for AMR surveillance, harbour multidrug-resistant E. coli, and enterococci that can be a substantial source of superbugs in the environment. Our data also highlight the urgency of including non-viable chicken embryos in AMR surveillance programme to understand AMR dissemination and its control.

Synthetic CpG-ODNs can promote antimicrobial immunity in neonatal chicks by enriching immune compartments and activating immune cells. Activated immune cells undergo profound metabolic changes to meet cellular biosynthesis and energy demands and facilitate the signaling processes. We hypothesize that CpG-ODNs induced immune activation can change the host’s metabolic demands in neonatal chicks. Here, we used NMR-based metabolomics to explore the potential of immuno-metabolic interactions in the orchestration of CpG-ODN-induced antimicrobial immunity. We administered CpG-ODNs to day-old broiler chicks via intrapulmonary (IPL) and intramuscular (IM) routes. A negative control group was administered IPL distilled water (DW). In each group (n = 60), chicks (n = 40) were challenged with a lethal dose of Escherichia coli, two days post-CpG-ODN administration. CpG-ODN administered chicks had significantly higher survival (P P P P

Synthetic DNA containing CpG motifs (CpG-ODN) are potent innate immune stimulators in neonatal and adult broiler chickens against bacterial septicemia. We have recently demonstrated that intrapulmonary (IPL) delivery of CpG-ODN as microdroplets under laboratory conditions can protect neonatal chickens against lethal Escherichia coli septicemia. The objectives of this study were to develop a commercial-scale poultry nebulizer (CSPN) that can deliver CpG-ODN as microdroplets in neonatal broiler chicks in the hatcheries and study the efficacy of CSPN in inducing immune-protective effects under different environmental conditions in 2 geographical locations in Canada. Three field experiments were conducted in commercial poultry hatcheries during different seasons of the year in Saskatchewan and British Columbia, Canada. Neonatal broiler chicks (n = 8,000/experiment) received CpG-ODN by the IPL route in the CSPN chamber for 30 min, and control chicks received distilled water (DW) for 30 min. Broiler chicks (CpG-ODN—240 chicks/experiment and DW—40 chicks/experiment) were randomly sampled from all locations of the CSPN after nebulization and challenged with a lethal dose of E. coli to examine the CpG-ODN nebulization induced protection. We found a significant level (P < 0.05) of protection in broiler chicks against E. coli challenge, suggesting that the newly built CSPN successfully delivered CpG-ODN via the IPL route. We found that when the CSPN was maintained at humidex 28°C or below and relative humidity (RH) between 40 and 60%, neonatal birds were significantly (P < 0.05) protected against E. coli septicemia after IPL delivery of CpG-ODN. By contrast, protection in chicks was adversely affected when the CSPN was maintained at the humidex of 29°C or higher and RH of 70%. Overall, the present study successfully built a CSPN for CpG-ODN delivery in chicks at the hatchery and revealed that the temperature, humidity, and humidex were critical parameters in CSPN for efficient delivery of CpG-ODN.

Losses due to infectious diseases are one of the main factors affecting productivity in the swine industry, motivating the investigation of disease resilience-related traits for genetic selection. However, these traits are not expected to be expressed in the nucleus herds, where selection is performed. One alternative is to use information from the commercial level to identify and select nucleus animals genetically superior for coping with pathogen challenges. In this study, we analyzed the genetic basis of antibody (Ab) response to common infectious pathogens in health-challenged commercial swine herds as potential indicator traits for disease resilience, including Ab response to influenza A virus of swine (IAV), Mycoplasma hyopneumoniae (MH), porcine circovirus (PCV2), and Actinobacillus pleuropneumoniae (APP; different serotypes). Ab response was measured in blood at entry into gilt rearing, post-acclimation (∼40 days after entering the commercial herd), and parities 1 and 2. Heritability estimates for Ab response to IAV, MH, and PCV2 ranged from 0 to 0.76. Ab response to APP ranged from 0 to 0.40. The genetic correlation (rG) of Ab response to IAV with MH, PCV2, PRRSV, and APPmean (average Ab responses for all serotypes of APP) were positive (>0.29) at entry. APPmean was negatively correlated with PCV2 and MH at entry and parity 2 but positively correlated with MH at post-acclimation and parity 1. Genomic regions associated with Ab response to different APP serotypes were identified on 13 chromosomes. The region on chromosome 14 (2 Mb) was associated with several serotypes of APP, explaining up to 4.3% of the genetic variance of Ab to APP7 at entry. In general, genomic prediction accuracies for Ab response were low to moderate, except average Ab response to all infectious pathogens evaluated. These results suggest that genetic selection of Ab response in commercial sows is possible, but with variable success depending on the trait and the time-point of collection. Future work is needed to determine genetic correlations of Ab response with disease resilience, reproductive performance, and other production traits.

The transition to antibiotic-free poultry production in the face of pathogenic threats is a very challenging task. We recently demonstrated that mucosal delivery of CpG-ODN alone by the intrapulmonary route (IPL) has potential as an effective alternative to antibiotics in neonatal chicks against Escherichia coli septicemia. How exactly mucosal delivery of CpG-ODN elicits, protective antibacterial immunity remained poorly understood. In this study, CpG-ODN or saline was delivered via the intrapulmonary route to day-old chicks (n = 80/group) using a compressor nebulizer in an acrylic chamber (1 mg/mL CpG-ODN for 15 minutes). In the first part of the study, two days after mucosal CpG-ODN delivery, 40 chicks from each group were challenged subcutaneously with 1 × 105 cfu (n = 20) or 1 × 106 cfu (n = 20) of E. coli and the mortality pattern was monitored for seven days. We found significantly higher survival, better clinical conditions and lower bacterial loads in chicks that received mucosal CpG-ODN. To explore the mechanisms behind this protective immunity, we first looked at the kinetics of the cytokine gene expression (three birds/ group/ time for 10 time-points) in the lungs and spleens. Multiplex gene analysis demonstrated a significant elevation of pro-inflammatory cytokine genes mRNA in the CpG-ODN group. Interleukin (IL)-1β robustly upregulated many folds in the lung after CpG-ODN delivery. Lipopolysaccharide-induced tumor necrosis factor (LITAF) and IL-18 showed expression for an extended period in the lungs. Anti-inflammatory cytokine IL-10 was upregulated in both lungs and spleen, whereas IL-4 showed upregulation in the lungs. To investigate the kinetics of immune enrichment in the lungs and spleens, we performed flow cytometry, histology, and immunohistochemistry at 24, 48 and 72 hrs after CpG-ODN delivery. CpG-ODN treated lungs showed a significant enrichment with monocytes/macrophages and CD4+ and CD8+ T-cell subsets. Macrophages in CpG-ODN treated group demonstrated mature phenotypes (higher CD40 and MHCII expression). Importantly, mucosal delivery of CpG-ODN via the intrapulmonary route significantly enriched immune compartment in the spleen as well, suggesting a systemic effect in neonatal chicks. Altogether, intrapulmonary delivery of aerosolized CpG-ODN orchestrates protective immunity against E. coli septicemia by not only enhancing mucosal immunity but also the systemic immune responses.

Immunoprotective function of oligodeoxynucleotides containing CpG motifs (CpG-ODN) has been demonstrated in neonatal chickens against common bacterial pathogens such as E.coli and Salmonella sp. Our recent study reported that CpG-ODN administration enriches immune compartments in neonatal chicks. However, a causal relationship between CpG-ODN-induced immune enrichment and protective mechanisms remains unestablished. In this study, we investigated in ovo administered CpG-ODN-mediated immune cell recruitment in the immunological niches in lymphoid (spleen) and nonlymphoid (lungs) organs using various doses of CpG-ODN and examined whether the immunological profiles have any correlation with immunoprotection against E.coli infection. Eighteen-day-old embryonated eggs were injected with either 5, 10, 25, and 50 μg of CpG-ODN or saline (n=~40 per group). On the day of hatch (72 hr after CpG-ODN treatment), we collected the spleen and lungs (n=3‐4 per group) and examined the recruitment of macrophages/monocytes, their expression of MHCII and CD40, and the number of CD4+ and CD8+ T-cell subsets in the immunological niches in the spleen and lungs using flow cytometry. We observed the dose-dependent recruitment of immune cells, wherein 25 μg and 50 μg of CpG-ODN induced significant enrichment of immunological niches in both the spleen and the lungs. Four days after the CpG-ODN treatment (1-day after hatch), chicks were challenged with a virulent strain of E. coli (1×104 or 1×105 cfu, subcutaneously). Clinical outcome and mortality were monitored for 8 days postchallenge. We found that both 25 μg and 50 μg of CpG-ODN provided significant protection and reduced clinical scores compared to saline controls against E. coli infection. Overall, the present study revealed that CpG-ODNs orchestrate immunological niches in neonatal chickens in a dose-dependent manner that resulted in differential protection against E. coli infection, thus supporting a cause and effect relationship between CpG-ODN-induced immune enrichment and the antibacterial immunity.

Historically, fowl adenovirus (FAdV) associated inclusion body hepatitis (IBH) was considered a secondary disease in broiler chickens associated with immunosuppression. However, we previously reported the occurrence of IBH as a primary disease in the broiler chicken industry in Canada as a result of infections with various FAdV serotypes. Therefore, the objectives of this study were to develop an immunization strategy in broiler breeders using live FAdV 11-1047 and FAdV8a-TR59 to confer homologous and heterologous protection in broiler progeny against IBH and to study the efficacy of natural exposure of naive broiler breeders to a vaccine virus from live FAdV vaccinated birds as an immunization technique. Broiler breeders vaccinated orally with FAdV8a-TR59 (1 × 104 TCID50/bird) and FAdV11–1047 (1 × 104 TCID50/bird), FAdV8a-TR59 (1 × 106 TCID50/bird) and FAdV11-1047 (1 × 106 TCID50/bird) or FAdV8b (1 × 106 TCID50/bird) transferred substantial levels of neutralizing antibodies to their progeny. The efficacy of maternal antibodies was studied by challenging 14-day old broiler chickens with 1 × 107 TCID50 of FAdV2–685, FAdV7-x11a like, FAdV8a-TR59, FAdV8b-SK or FAdV11-1047 which are the dominant serotypes causing IBH outbreaks in Canada. Broiler chickens from the low and high dose vaccinated breeders were significantly protected against all serotypes of FAdV (P

Fowl adenovirus (FAdV) is comprised of five species (A to E) and 12 serotypes (1–7, 8a, 8b, 9–11). Inclusion body hepatitis (IBH) is caused by FAdV-7, 8a, 8b (species E) and FAdV-2 and 11 (species D). Commercial vaccines against IBH are not available in Canada. Autogenous FAdV broiler breeder vaccines are now used in some areas where outbreaks of IBH are occurring. The objective of this study was to evaluate the efficacy of a bivalent (species D and E) live and an inactivated FAdV broiler breeder vaccine in protecting broiler chicks against IBH through maternal antibody (MtAb) transfer. FAdV seronegative broiler breeders (n = 300/group) received either a live or inactivated bivalent (FAdV-8b-SK + FAdV-11-1047) vaccine. The live vaccine (1 × 104 TCID50 of each virus/bird) was given orally once at 16 weeks of age and the inactivated vaccine (1 × 106TCID50 of each virus + 20% Emulsigen D) was given intramuscularly at 16 and 19 weeks of age. Controls (n = 150) were given saline orally. The inactivated vaccine group was boosted 3 weeks later with the same vaccine. Neutralizing antibodies (NAb) in sera (n = 10) were detected at 19, 22, 30 and 48 weeks of age. NAb were able to neutralize various FAdV serotypes within species D and E. Mean NAb were similar in the both live and killed vaccine groups at 19, 30 and 48 weeks and ranged from 2.4 to 3.7 log10. Approximately 26 ± 7% of MtAbs were passively transferred through eggs to day-old chicks. Progeny challenged with a lethal dose (1 × 107 TCID50/bird intramuscularly) of FAdV-8b-SK, FAdV-11-1047, or FAdV-2-685 (n = 90/group) at 14 days post-hatch (dph) showed 98–100% protection in broiler chicks to homologous or heterologous FAdV challenges. Our data suggests that a bivalent live and an inactivated FAdV vaccine are equally effective and have the potential for the control of IBH.

The emergence of enterococcal infections in neonatal broiler chickens in the poultry industry has become common in many countries, including Canada. The objective of this study was to examine the bacterial infections in nonviable broiler chicken embryos in three western Canadian poultry hatcheries using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The pattern of embryo mortality that occurred during incubation and the breakout analysis results were similar in all three hatcheries. The majority of embryo mortality occurred during the late stage of incubation (35.08%), followed by the early stage of incubation (15.35%). The breakout analysis showed that 65.82% of swabs had at least one type of bacterial growth while 34.17% of swabs were negative for bacterial isolation. Of those 65.82% swabs with bacterial growth, 34.3% of swabs yielded a mixed bacterial population while 31.52% yielded one type of bacterial growth. The frequency of bacterial isolation from hatch debris (60%-75%) increased with the age of broiler breeders. MALDI-TOF MS was able to provide genus-level identification of 83.13% of isolates among all bacterial types isolated. MALDI-TOF MS identified Enterococcus and Escherichia coli isolates with 97.18% and 100% accuracy at species level, respectively, whereas Staphylococcus species were identified with 62.59% accuracy. The congruence between MALDI-TOF MS identification and 16S rRNA or cpn60 universal gene target sequencing was 100% or 90%, respectively. Of all bacteria isolated, Enterococcus species (29.71%) were the most prevalent, followed by E. coli (19.46%). About 56% of E. coli-infected samples were coinfected with Enterococcus species. Among all Enterococcus species isolated, Enterococcus faecalis (79.58%) was the most prevalent, followed by Enterococcus faecium (8.1%). Overall, our study showed that Enterococcus-associated embryo mortality was predominant in all three hatcheries investigated and suggests that MALDI-TOF MS technology can be applied to identify bacteria such as Enterococcus species isolated from poultry.

Inclusion body hepatitis (IBH) is an economically important diseases in broiler chicken industry. Several serotypes of fowl adenovirus (FAdV) can cause IBH, among them, serotype FAdV-8b is associated with the majority of the IBH cases in Canada. Here, we evaluated FAdV-8b virus-like particles (VLPs) and recombinant FAdV-8b fiber proteins (expressed in E. coli) as potential broiler-breeder vaccines against IBH. For assessing the immunogenicity of vaccines, we investigated both humoral and cellular immunity. The humoral immune response was evaluated by determining total IgY and virus-neutralizing antibody in serum at 14, 28, 35 and 60days post-immunization (dpi). We examined cellular immunity using flow cytometry by determining CD4:CD8 ratio change in peripheral blood after the booster vaccination. The protective effect of vaccines was tested by challenging 14day-old progeny (n=30/group) carrying maternal antibodies (MtAb) by challenging with virulent FAdV-8b virus (1×107 TCID50, FAdV-8b-SK). Although total IgY levels were comparable in all groups, the neutralizing antibody response in broiler-breeders at 35 and 60 dpi was significantly (p

Previous results indicated that antibody response to PRRSV has moderate genomic prediction accuracy; however, little is known about this for other common infectious diseases. Therefore, the objective of this study was to estimate the accuracy of genomic prediction for antibody response to infectious diseases in commercial sows. A total of 2,848 Large White x Landrace replacement gilts were sourced from 17 high-health multipliers (7 breeding companies; BC) and introduced to 23 commercial farms with a history of common diseases, following standard acclimation procedures. Serum was used to quantify antibody response to swine influenza virus (SIV), Mycoplasma hyopneumoniae (MH), porcine circovirus type 2 (PCV2), and 8 serotypes of Actinobacilluspleuropneumoniae(APP1-3, 5, 7, 10, 12, and 13) at entry (S/PEntry), following acclimation (S/PAcclimation), and during parities 1 (S/PParity1) and 2 (S/PParity2). All animals were genotyped for 38,191 SNPs. Genomic prediction was performed using BayesB (pi=0.99), with the fixed effect of CG and random effects of SNPs included in the model. Training and validation were performed using 7-fold cross-validation, with data from each BC used as the validation dataset in one-fold. In general, prediction accuracies were low: SIV, from 0.13 (S/PAcclimation) to 0.26 (S/PParity1); MH, -0.07 (S/PAcclimation) to 0.13 (S/PParity2); PCV2, 0.04 (S/PParity1) to 0.32 (S/PAcclimation); APP, -0.08 (S/PEntry, APP10) to 0.26 (S/PAcclimation, APP7). At each point, average accuracies were 0.06 for S/PEntry, 0.09 for S/PAcclimationand S/PParity1, and 0.08 for S/PParity2, showing small increases in accuracy after the acclimation period. Among diseases, average accuracies ranged from 0.01 (APP1) to 0.22 (PCV2). Results show that, overall, the accuracy of genomic prediction of antibody response to common infectious diseases in commercial gilts is limited. The authors thank PigGen Canada, Genome Canada, and the Canadian Swine Health Board for financial support, and the late Dr. Stephen Bishop for his scientific contributions.

The objective of this study was to perform genomic predictions for reproductive performance of sows under natural health challenge. Reproductive performance (1 to 4 parities) and genotype (~40K SNPs) were available for 2,604 crossbred sows, for a total of 7,635 farrowing records. Animals from 17 high-health multipliers from 7 breeding companies (PigGen Canada) were shipped to 23 commercial farms with recent history of common infectious diseases. Gilts entered farms with an average of 53 animals per contemporary group (CG). Traits included total number of piglets: born (TB), born alive (NBA), stillborn (SB), mummified (MUM), born dead (NBD), and weaned (NW). Genomic predictions were performed using Bayes-B (pi=0.995) with a seven-fold cross-validation using each company in turn for validation and the others for training. The model included the effects of CG (fixed) and SNP (random), and net number of fosters (covariate) for NW. Genomic predictions were done for animal lifetime performance (sum performance of parities) for each trait and using first parity performance as the training set to predict subsequent parity performance. Accuracy was calculated as the weighted average correlation between GEBV and adjusted phenotype across validation sets divided by the square root of heritability. Lifetime performance accuracies were low to moderate, ranging from 0.11 (TB) to 0.45 (NBD). Accuracies using parity 1 to predict subsequent performance were low, ranging from -0.07 (SB in parity 3) to 0.19 (NBD in parity 2), with average accuracies per trait ranging from 0.04 (SB) to 0.16 (NBD).Although most accuracies were low, the moderately high accuracies for some lifetime performance shows that genomic prediction can be used to improve performance under natural health challenge in sows. We appreciate the financial support of PigGen Canada, Canadian Swine Health Board, Genome Alberta and Swine Innovation Porc, and the late Dr. Stephen Bishop for his scientific contributions.

Oligodeoxynucleotides containing CpG motifs (CpG-ODN) induce innate immunity against bacterial infections. Despite recent advances, how CpG-ODN alone protects against bacterial infections remained elusive. Here, we report for the first time, to our knowledge, that CpG-ODN orchestrates anti-microbial protective immunity by inducing a rapid enrichment of various immune compartments in chickens. In this study, eighteen-day-old embryonated eggs were injected with either 50 µg of CpG-ODN or saline (~n = 90 per group). In the first experiment, four days after CpG-ODN treatment, chicks were challenged subcutaneously with a virulent strain of Escherichia coli (E. coli) and mortality was monitored for 8 days. We found significant protection, and reduced clinical scores in CpG-ODN treated chicks. To gain insights into mechanisms of protection induced by CpG-ODN, first we investigated cytokine expression kinetics elicited by CpG-ODN. The spleen and lung were collected from embryos or chicks (n = 3–4 per group) at 10 time points post-CpG-ODN inoculation. Multiplex gene analysis (interleukin (IL)-1, IL-4, IL-6, IL-10, IL-18, interferon (IFN)-γ, IFN-α, and lipopolysaccharide induced tumor necrosis factor (LITAF), revealed a significantly higher expression of pro-inflammatory cytokines following CpG-ODN treatment compared to the saline controls. In our study, LITAF stands out in the cytokine profiles of spleen and lungs, underscoring its role in CpG-ODN-induced protection. The third experiment was designed to examine the effects of CpG-ODN on immune cell populations in spleen, lungs, and thymus. Flow cytometry analysis was conducted at 24, 48 and 72 hrs (thymus only collected at 72 hr) after CpG-ODN administration to examine the changes in CD4+ and CD8+ T-cell subsets, monocyte/macrophage cell populations and their expression of maturation markers (CD40 and CD86). Flow cytometry data indicated a significant enrichment of macrophages, CD4+ and CD8+ T-cell subsets in both spleen and lungs of CpG-ODN treated embryos and chicks. Macrophages in spleen and lungs showed an upregulation of CD40 but not CD86, whereas thymocytes revealed significantly high CD4 and CD8 expression. Overall, the present study has demonstrated that CpG-ODN provides protection in neonatal chicks against E. coli infection not only by eliciting cytokine responses and stimulating immune cells but also through enriching immunological niches in spleen and lungs.

Current knowledge regarding the association between indoor mold exposures and asthma is still limited. The objective of this case-control study was to investigate the relationship between objectively measured indoor mold levels and current asthma among school-aged children. Parents completed a questionnaire survey of health history and home environmental conditions. Asthma cases had a history of doctor-diagnosed asthma or current wheeze without a cold in the past 12 months. Controls were age- and sex-matched to cases. Vacuumed dust samples were collected from the child's indoor play area and mattress. Samples were assessed for mold levels and quantified in colony-forming units (CFU). Sensitization to mold allergens was also determined by skin testing. Being a case was associated with family history of asthma, pet ownership, and mold allergy. Mold levels (CFU/m2 ) in the dust samples of children's mattress and play area floors were moderately correlated (r = 0.56; P

Synthetic oligodeoxynucleotides (ODN) containing unmethylated cytosine phosphodiester guanine (CpG) motifs (CpG-ODN) are effective immunostimulatory agents against a variety of viral, bacterial, and protozoan diseases in different animals including poultry. We have recently demonstrated that in ovo injection of CpG-ODN confers protection in neonatal chickens against bacterial septicemias. The objective of this study was to investigate the effectiveness of needle-free intrapulmonary (IPL) delivery of CpG-ODN microdroplets against Escherichia coli infection in neonatal chicks. In the present study, we used 880 chicks in total keeping 40 chicks per group. Chicks were delivered CpG-ODN or saline by IPL at the day 1 of hatch. Three days later, chicks were challenged with two doses (1 × 104 CFU, n = 20 or 1 × 105 CFU, n = 20) of E. coli. Chicks treated with CpG-ODN by the IPL route had significantly lower clinical signs and bacterial load compared to the group treated with saline ( P < 0.05). CpG-ODN-treated groups were significantly protected against E. coli septicemia. We observed dose- and exposure time-dependent immunoprotective effects of IPL CpG-ODN in chicks. We found that IPL delivery of CpG-ODN can induce protective immunity as early as 6 hr that remains effective at least until day 5 post-treatment. Moreover, there were no adverse effects of IPL delivery of CpG-ODN on growth or mortality up to 42 days of age. Based on these findings, it can be suggested that CpG-ODN delivery by IPL route can be a promising alternative to antibiotics for inducing protective immunity in chicks during the critical first week of neonatal life.

The objective of this study was to perform genome-wide association studies (GWAS) to identify Quantitative Trait Loci (QTL) associated with antibody response to infectious diseases in commercial sows. A total of 2,848 Large White x Landrace replacement gilts were sourced from 17 high-health multipliers (7 breeding companies; BC) and introduced to 23 commercial farms with a history of common pig diseases, following the standard acclimation procedures with an average of 53 animals per entry group (CG). Serum was used to quantify antibody response to swine influenza virus (SIV), Mycoplasma hyopneumoniae (MH), porcine circovirus type-2 (PCV2), and 8 serotypes of Actinobacillus pleuropneumoniae (APP1-3, 5, 7, 10, 12, and 13) at entry (S/PEntry), following acclimation (S/PAcclimation), and during parities 1 (S/PParity1) and 2 (S/PParity2). All animals were genotyped for 38,191 SNPs. GWAS was performed using BayesB (pi=0.99), with the fixed effect of CG and the random effects of SNPs in the model. For APP, QTL were only identified at S/PAcclimation; on SSC14 (2Mb) for APP3, APP7, APP10, and APP13 that explained 5.6, 4.7, 2.8, and 3.6% of the genetic variance, respectively. A gene within this QTL region is SYK, involved in the control of immune-receptors. For APP5, a QTL that explained 4.2% of the genetic variance was identified on SSC4 (105Mb), which co-localizes with two genes associated with immune-response: SIKE1and NRAS. For SIV, no QTL was identified. A QTL on SSC7 (130-131Mb) was identified for MH (S/PParity1, 5.1%) and PCV2 (S/PEntry, 34%; S/PAcclimation, 43.4%). These results provide new information on the genetic basis of response to infectious diseases in sows. The identified QTL have the potential to be used to select for improved immune response. The authors thanks PigGen Canada, Genome Canada, and the Canadian Swine Health Board for financial support, and the late Dr. Stephen Bishop for his scientific contributions.

Background Our recent research showed that antibody response to porcine reproductive and respiratory syndrome (PRRS), measured as sample-to-positive (S/P) ratio, is highly heritable and has a high genetic correlation with reproductive performance during a PRRS outbreak. Two major quantitative trait loci (QTL) on Sus scrofa chromosome 7 (SSC7; QTLMHC and QTL130) accounted for ~40 % of the genetic variance for S/P. Objectives of this study were to estimate genetic parameters for PRRS S/P in gilts during acclimation, identify regions associated with S/P, and evaluate the accuracy of genomic prediction of S/P across populations with different prevalences of PRRS and using different single nucleotide polymorphism (SNP) sets. Methods Phenotypes and high-density SNP genotypes of female pigs from two datasets were used. The outbreak dataset included 607 animals from one multiplier herd, whereas the gilt acclimation (GA) dataset included data on 2364 replacement gilts from seven breeding companies placed on health-challenged farms. Genomic prediction was evaluated using GA for training and validation, and using GA for training and outbreak for validation. Predictions were based on SNPs across the genome (SNPAll), SNPs in one (SNPMHC and SNP130) or both (SNPSSC7) QTL, or SNPs outside the QTL (SNPRest). Results Heritability of S/P in the GA dataset increased with the proportion of PRRS-positive animals in the herd (from 0.28 to 0.47). Genomic prediction accuracies ranged from low to moderate. Average accuracies were highest when using only the 269 SNPs in both QTL regions (SNPSSC7, with accuracies of 0.39 and 0.31 for outbreak and GA validation datasets, respectively. Average accuracies for SNPALL, SNPMHC, SNP130, and SNPRest were, respectively, 0.26, 0.39, 0.21, and 0.05 for the outbreak, and 0.28, 0.25, 0.22, and 0.12, for the GA validation datasets. Conclusions Moderate genomic prediction accuracies can be obtained for PRRS antibody response using SNPs located within two major QTL on SSC7, while the rest of the genome showed limited predictive ability. Results were obtained using data from multiple genetic sources and farms, which further strengthens these findings. Further research is needed to validate the use of S/P ratio as an indicator trait for reproductive performance during PRRS outbreaks. Electronic supplementary material The online version of this article (doi:10.1186/s12711-016-0230-0) contains supplementary material, which is available to authorized users.

While the prevalence of infectious bursal disease virus (IBDV) on chicken farms in some provinces of Canada has been documented, the economic impact of variant IBDV infection on the broiler chicken industry in Saskatchewan has not. The objectives of this study were to identify the variant strains of IBDV circulating on Saskatchewan chicken farms and evaluate their economic impact on broiler production. Infection due to IBDV was detected in 43% of Saskatchewan chicken farms, with variant strains detected in infected birds closely related predominantly to NC171, 586, and Delaware-E. Infected flocks showed an IBDV antibody titer of 4236 geometric mean (GM), whereas an antibody titer of 157 GM was measured in uninfected flocks. Infected flocks had very low (0.06) bursa-to-body-weight (BBW) ratio (an indicator of immunity) compared to high BBW ratio (0.17) in uninfected flocks, which suggests a significant immunosuppression in the former. Flocks positive for IBDV had mean mortality of 8.6% and mean condemnation of 1.5%. In contrast, mean mortality in uninfected flocks was 6.1% and mean condemnation was 1.1%. The live market weight per grow area at 37 d of age was 29.3 kg/mBien que la prévalence du virus de la maladie infectieuse de la bourse (VMIB) sur les fermes de poulets dans quelques provinces canadiennes ait été documentée, l’impact économique d’infections par des variants du VMIB sur l’industrie du poulet à griller en Saskatchewan ne l’est pas. Les objectifs de la présente étude étaient d’identifier les souches variantes du VMIB circulant au sein des fermes de poulet de la Saskatchewan et d’évaluer leur impact économique sur la production de poulets à griller. L’infection due au VMIB a été détectée sur 43 % des fermes de poulet de la Saskatchewan, avec des souches variantes détectées chez des oiseaux infectés étant fortement apparentées en prédominance aux souches NC171, 586, et Delaware-E. Les troupeaux infectés présentaient une moyenne géométrique (MG) des titres d’anticorps contre le VMIB de 4236, alors que la MG des titres d’anticorps des oiseaux provenant de troupeaux non-infectés était de 157. Les troupeaux infectés avaient un très faible ratio (0,06) du poids bourse-poids corporel (BPC) (un indicateur de l’immunité) comparativement au ratio BPC élevé (0,17) dans les troupeaux non-infectés, ce qui suggère une immunosuppression significative dans les troupeaux infectés. Les troupeaux positifs au VMIB avaient un taux de mortalité moyen de 8,6 % et un taux de condamnation moyen de 1,5 %. À l’opposé, dans les troupeaux non-infectés le taux de mortalité moyen était de 6,1 % et le taux de condamnation moyen de 1,1 %. Le poids vif de marché à 37 j d’âge par surface de croissance était de 29,3 kg/m

Chickens are commonly processed around 35-45days of age in broiler chicken industry hence; diseases that occur at a young age are of paramount economic importance. Early age infection with infectious bursal disease virus (IBDV) results in long-lasting immunosuppression and profound economic losses. To our knowledge, this is the first study comparing the protection efficacy of modified live (MdLV) IBDV and herpesvirus turkey (HVT)-IBDV vaccines against early age variant IBDV (varIBDV) infection in chicks. Experiments were carried out in IBDV maternal antibody (MtAb) positive chicks (n=330), divided into 6 groups (n=50-60/group), namely Group 1 (saline), Group 2 (saline+varIBDV), Group 3 (HVT-IBDV), Group 4 (HVT-IBDV+varIBDV), Group 5 (MdLV) and Group 6 (MdLV+varIBDV). HVT-IBDV vaccination was given via the in ovo route to 18-day-old embryonated eggs. MdLV was administered via the subcutaneous route in day-old broilers. Group 2, Group 4 and Group 6 were orally challenged with varIBDV (SK-09, 3×103 EID50) at day 6 post-hatch. IBDV seroconversion, bursal weight to body weight ratio (BBW) and bursal histopathology were assessed at 19 and 35days of age. Histopathological examination at day 19 revealed that varIBDV-SK09 challenge caused severe bursal atrophy and lower BBW in HVT-IBDV but not in MdLV vaccinated chicks. However by day 35, all challenged groups showed bursal atrophy and seroconversion. Interestingly, RT-qPCR analysis after varIBDV-SK09 challenge demonstrated an early (9days of age) and significantly high viral load (∼5744 folds) in HVT-IBDV vaccinated group vs unvaccinated challenged group (∼2.25 folds). Furthermore, flow cytometry analysis revealed inhibition of cytotoxic CD8+ T-cell response (CD44-downregulation) and decreased splenic lymphocytes counts in chicks after HVT-IBDV vaccination. Overall, our data suggest that MdLV delays varIBDV pathogenesis, whereas, HVT-IBDV vaccine is potentially immunosuppressive, which may increase the risk of early age varIBDV infection in broilers.

Antibiotic-free and safe animal products are most desirable among consumers. However, ensuring safe poultry products is a challenging task when the chicken immune system is compromised. Infectious bursal disease virus (IBDV) causes immunosuppression and predisposes chickens to secondary infections. Breeder vaccination against IBDV is routinely practiced for producing chicks with maternally-derived antibody (MAb) to prevent infection in newly hatched chicks. The majority of IBDV circulating in Canadian farms are variant strains (vIBDV). Whether circulating vIBDV strains are immunosuppressive in chicks or are amenable to current vaccine regimens has not previously been tested through challenge studies. In this study, one-day-old broiler chicks (n=240) carrying MAb were obtained from broiler breeders vaccinated with commercial IBDV vaccines. In the first set of experiments (n=40/group), at six days post-hatch, one group was challenged with a Canadian field isolate, vIBDV (strain-SK09) (3×10(3) EID50). The second and the third groups (controls) were inoculated with non-immunosuppressive IBDV D-78 (10×10(3) TCID50) and saline, respectively. Histopathological examination on days 14 and 30 post-challenge revealed that despite the high level of MAb, vIBDV (SK09) caused severe bursal damage in chicks. Another set of experiments with treatment groups as above, demonstrated that pre-exposure of chicks with vIBDV (SK09) caused immunosuppression resulting in significantly higher mortality and disease severity in chicks challenged with a virulent strain of Escherichia coli (E. coli). Our data provide evidence that IBDV strains circulating in Canada are immunosuppressive, not amenable to current anti-IBDV vaccination strategy, and a potential threat to antibiotic-free chicken farming.

To determine the prevalence of chronic bronchitis (CB) and associated risk factors in farm and nonfarm rural residents in Saskatchewan, Canada.The questionnaire collected information about health, contextual, and individual factors from 8261 farm and nonfarm adult residents (18 years and older).The prevalence of CB was 5.3% among farm residents and 6.4% among nonfarm residents. We found a greater prevalence of CB associated with household income adequacy, increasing age, allergies, history of lung disease in a parent, exposure to stubble smoke, obesity, prenatal exposure to smoking, and female sex. Smoking interacted with occupational exposure to wood dust and solvents, and allergic reaction to molds.The results suggest that increasing household income and reducing smoking could be primary, modifiable determinants of CB prevalence.

The objective of this pilot study was to investigate if a once-a-day sprinkling of canola oil in a swine confinement facility alters the airborne concentration and distribution of particulate matter and associated compounds (endotoxin and culturable microbes). Particulate was collected using an eight-stage cascade impactor in four identical swine grower/finisher rooms of a swine barn. Particulate (mg/m(3)) and endotoxin (EU/m(3) and EU/mg) distribution was determined. A six-stage viable cascade impactor was used to quantify total bacteria, enteric bacteria, and fungi. Microbes were characterized from subcultures prepared from the 10 most predominant colony types on each stage 3 (aerodynamic size 3.3-4.7μm) of the collection plates. Results indicated that oil sprinkling reduced total dust by 86% and total endotoxin concentration by 82.5%. However, the distribution patterns indicate that reduction is observed predominantly on large dust particles. In addition, the proportion of endotoxin associated with smaller particulate sizes (i.e., particles4.7 μm) was higher in the oil-sprinkled rooms. Oil sprinkling does not markedly alter distribution of total bacteria, enteric bacteria, or fungi. The most frequently identified species were gram-positive genera. Oil sprinkling in swine confinement grower/finisher rooms can significantly reduce airborne total dust and endotoxin; however, smaller particles and associated endotoxin appear to remain in suspension, suggesting the overall improvement in air quality is uncertain. Further distribution studies and exposure outcome studies would need to be undertaken to determine the impacts of oil sprinkling.

The aim of this work is to better understand the responses of people that are exposed to agricultural organic dust and other factors in modern swine production. We investigated the effects of toll-like receptor 4 (TLR4) genotype and gender on respiratory responses of naïve volunteers (18-28 years) to swine barn exposure. Non-smoking healthy subjects (16 men and 13 women) with TLR4 299 (Asp299Gly) and/or 399 (Thr399Ile) polymorphisms (TLR4 299/399) and age-sex matched subjects with TLR4 wild-type alleles spent 5 h in a nonexposed environment (baseline day) and 5 h in a swine facility (exposure day). The results showed significant decreases between baseline and exposure days in across-shift forced vital capacity (FVC), forced expiratory volume in 1 second (FEV(1)), forced midexpiratory flow rate (FEF(25-75)), and FEV(1)/FVC ratio and in methacholine concentration that reduced FEV1 by 20% (PC(20)) in all groups; however, there were no differences by sex or genotype. Similarly, nasal cytokines, serum cytokines, and blood neutrophil count increased after exposure; in contrast, however, these were influenced by gender. The increase in serum tumor necrosis factor-alpha (TNF-alpha) between baseline and exposure was gender-dependent with male sex associated with a significant increase in the wild-type group and female sex associated with a significant increase in the polymorphic group. These results suggest that for persons exposed to a swine facility, one's immunological response varies with gender as well as TLR4 genotype.

Recently there has been interest in the air quality in and around intensive livestock production facilities, such as modern swine production barns, where agricultural workers and surrounding residents may be exposed to elevated levels of organic dusts. The health effects of these exposures are not completely understood. The study that is reported here is a component of a larger investigation of the relationships among the acute effects of high-concentration endotoxin exposure (swine barn dust), polymorphisms in the TLR4 gene, and respiratory outcomes following exposure to swine confinement buildings. The relationships among a mediator of acute lung inflammation, tumor necrosis factor alpha (TNF-alpha), and clinical responses to acute swine barn exposure were characterized. Analysis of the results showed that in vitro stimulation of human monocytes with as little as 1 ng/ml of lipopolysaccharide (LPS) produced a significant increase in the monocytes that produced TNF-alpha. Although the proportion of TNF-alpha-positive monocytes after in vitro stimulation with 1 ng/ml of LPS was not associated with gender or TLR4 genotype, it was positively associated with the concentration of monocytes in blood after barn exposure. Thus, these two responses to different forms of LPS exposure are significantly correlated, and more responsive monocytes in vitro indicate a forthcoming relative monocytosis, post barn exposure, which may initiate a cascade of chronic inflammation.

Campylobacter jejuni , a commensal Gram-negative motile bacterium commonly found in chickens is a frequent cause of human gastrointestinal infections. The polar flagellum of C. jejuni is an important virulence and colonization factor, providing motility to the cell as well as a type III secretion function. The flagellar biosynthesis genes fliA ( σ 28 ) and rpoN ( σ 54 ) of C. jejuni regulate a large number of genes involved in motility, protein secretion and invasion, which have been shown to be important factors for the virulence of this organism. To understand the role of the flagellar sigma factors, σ 28 and σ 54 , in regulating colonization of the chicken intestinal tract, we assessed fliA and rpoN mutants of C. jejuni NCTC11168 for their ability to secrete Cia proteins and to adhere to and invade Hela cells. The mutants were also tested for their in vivo colonization potential in a chicken model with two different challenge doses. The fliA mutant showed reduced motility (25% that of the wild type) but secreted Cia proteins, yet it did not colonize the chicken cecum. The rpoN mutant cells lacked the spiral shape of C. jejuni and motility was reduced to 10% of the wild-type. The rpoN mutant did not secrete any Cia proteins but RT-PCR analysis showed the presence of c iaB mRNA, indicating that c iaB gene expression was independent of σ 54 . Not surprisingly, the colonization defects of both fliA and rpoN mutants were more severe than the flgK mutant. We also demonstrated that FlgK, the hook filament junction protein of C. jejuni , is required for assembly of the flagellar secretory apparatus and an flgK mutant of C. jejuni expressing only the hook showed diminished motility and was completely attenuated for cecal colonization in chickens.

The flagellum of Campylobacter jejuni is not only responsible for initiating colonization of the gastrointestinal tract of host animals but is also a major antigen that induces protective immune responses. However, protection is limited to the homologous strain and the ability to protect against multiple serotypes has yet to be determined. In this study, we have shown that FlaA is an immmunodominant protein on NCTC11168 CJ1 flagella and we mapped the immunoreactive epitopes on the protein by probing a series of overlapping synthetic peptides spanning the entire sequence with sera against multiple C. jejuni serotypes. Amino acid residues 176–205 (P8), 376–405 (P16) and 501–530 (P21) were immunodominant and cross-reactive. The mucosal IgA in the intestinal secretions of CJ1-infected birds reacted significantly with peptides P16 and P21 indicating that the specificity of the mucosal response is different from the systemic response. Antisera raised against formalin-killed CJ1 cells and purified flagellin showed positive reactivity with a subset of peptides identified by antisera against live C. jejuni. This study provides insight into the specificity of the host immune responses to the FlaA protein of C. jejuni and suggests that these sequences merit further testing for their immunogenicity and potential as subunit vaccine candidates for multiple serotypes.

Sao is a Streptococcus suis surface protein recently identified as a potential vaccine candidate. In this study, recombinant Sao in combination with Quil A provided cross-protection against S. suis serotype 2 disease in mouse and pig vaccination protocols. Subcutaneous immunization of mice elicited strong immunoglobulin G (IgG) antibody responses. All four IgG subclasses were induced, with the IgG2a titer being the highest, followed by those of IgG1, IgG2b, and IgG3. Challenge of the mice with S. suis strain 31533 resulted in a mortality rate of 80% for the control group, which received Quil A only. In contrast, all of the mice immunized with Sao survived. In a pig vaccination protocol, intramuscular immunization with Sao also elicited significant humoral antibody responses, and both the IgG1 and IgG2 subclasses were induced, with a predominance of IgG2 production. In vitro assay showed that Sao-induced antibodies significantly promoted the ability of porcine neutrophils in opsonophagocytic killing of S. suis . An aerosol challenge of the pigs with S. suis strain 166 resulted in clinical signs characteristic of S. suis infection in diseased pigs. The vaccine group showed significantly better survival, lower clinical scores, and less S. suis recovery from postmortem tissue samples than did the control group. Furthermore, this study also revealed that although challenge S. suis strains express Sao size variants, recombinant Sao conferred cross-protection. These data demonstrate that recombinant Sao formulated with Quil A triggers strong opsonizing antibody responses which confer efficient immunity against challenge infection with heterologous S. suis type 2.

Synthetic oligodeoxynucleotides (ODN) containing cytosine-phosphodiester-guanine (CpG) motifs (CpG-ODN) have been shown to be effective immunoprotective agents and vaccine adjuvants in a variety of bacterial, viral, and protozoan diseases in different animal species. The objective of this study was to compare the immune response of chickens to a killed Escherichia coli vaccine combined with oil in water emulsion or with CpG-ODN. Birds were vaccinated with killed E. coli antigens with either 10 or 50 microg of CpG-ODN on days 10 and 20 of age. At day 30, a virulent isolate of homologous E. coli was applied on a scratch site on the caudal abdominal region. Birds were examined for 10 days post-E. coli challenge, and pathologic and bacteriologic assessments were conducted on all birds that were either found dead or euthanized. The E. coli vaccine group that received no CpG-ODN had a survival rate of 65%. In contrast, groups that received the vaccine with CpG-ODN adjuvant had significantly higher survival rate of 92% (P0.01) with isolation of low numbers of E. coli from internal organs. Total IgG against E. coli antigens was highest in groups that received CpG-ODN as an adjuvant. Birds that received vaccine containing CpG-ODN had minimal inflammatory reaction without tissue necrosis at the injection site. Severe tissue necrosis was present in birds that received vaccine containing oil in water emulsion adjuvant. This study demonstrated that CpG-ODN is an effective vaccine adjuvant in chickens and results in minimal tissue destruction. This study is the first study in which CpG-ODN has been demonstrated to produce an adaptive immune response, at a significant level, against an extracellular bacterial infection in chickens.

The mechanisms used by Campylobacter jejuni to colonize the (chicken) intestinal tract have not been defined. In this study, we obtained evidence that in the presence of chicken serum and mucus, C. jejuni secreted proteins that may play a role in the colonization of chicken gut (Campylobacter invasion antigen = Cia). C. jejuni strains NCTC11168V1 and 81-176, as well as an NCTC11168V1 flaA mutant, were found to colonize intestinal tract and secrete proteins in the presence of chicken mucus, chicken serum, or fetal bovine serum in cell culture-conditioned medium. C. jejuni strain NCTC11168V26, which was observed to be a poor colonizer compared with the other C. jejuni isolates, did not secrete Cia proteins. Secreted proteins were also recognized by Western immunoblot using sera from birds that had been colonized by C. jejuni. These data suggest that C. jejuni secretes Cia proteins during colonization of chicken gut and that these Cia proteins play an important role in colonization.

The objective of this study was to identify the causative agents of hepatitis observed in broiler chickens at processing. Livers of chickens from 16 broiler farms in Saskatchewan with gross lesions of hepatitis were collected at processing. In addition to routine bacterial isolation and histopathological examination, serologic studies for infectious bursal disease virus (IBDV) and Chicken anaemia virus (CAV), calculation of the ratio of the weight of the bursa of Fabricius (BF) to body weight (BBW), and histopathological examination of the BF were done. Of the 264 livers with gross lesions, 83% had multifocal to coalescing necrotizing hepatitis, 16% had perihepatitis, and 1% had hemorrhages. No definitive causative microorganisms were isolated from the hepatic lesions; however, no significant bacterial isolations were made. Bursal atrophy, low BBW ratio, and high titer of antibody against IBDV each correlated with the rate of total condemnations (P = 0.0188, P = 0.0001, and P = 0.0073, respectively). Nucleotide sequencing of IBDV isolated from the BF identified the variant strains Delaware-E and 586. Condemnation because of hepatic lesions was correlated with titer of antibody against IBDV and BBW (P = 0.016 and P = 0.027). The results of this study demonstrate that hepatic lesions in Saskatchewan chickens are not currently caused by a primary bacterial pathogen but are associated with indicators of immunosuppression that is likely due to variant IBDV.L’objectif de la présente étude était d’identifier les agents causals de l’hépatite observée chez des poulets à griller au moment de la transformation. Les foies de poulets provenant de 16 fermes de poulets à griller en Saskatchewan avec des lésions macroscopiques d’hépatite furent prélevés. En plus de l’isolement bactérien de routine et de l’examen histopathologique, on effectua des analyses sérologiques pour le virus de la bursite infectieuse aviaire (VBIA) et le virus de l’anémie du poulet (VAP), le calcul du ratio du poids de la bourse de Fabricius (BF) sur le poids corporel (BPC), et l’examen histopathologique de la BF. Sur les 264 foies ayant des lésions macroscopiques, 83 % avaient des lésions multifocales à coalescentes d’hépatite nécrosante, 16 % de la péri-hépatite et 1 % des hémorragies. Aucun agent causal définitif ne fut isolé des lésions hépatiques; toutefois, aucun agent bactérien significatif ne fut isolé. Une atrophie de la bourse, un faible ratio BPC, et un titre élevé d’anticorps dirigé contre VBIA corrélaient tous avec le taux de condamnation totale (

Unformulated oligodeoxynucleotides (ODN) containing CpG motifs (CpG-ODN) have been shown to stimulate the innate immune system against a variety of bacterial, viral, and protozoan infections in a variety of vertebrate species. We have previously shown that in ovo delivery of unformulated CpG-ODN was able to significantly protect neonatal broiler chickens against Escherichia coli or Salmonella Typhimurium infections. The objectives of this study were to examine the safety and immunoprotective effects of CpG-ODN formulated with 2 types of carbon nanotubes (CNTs) or 2 types of lipid-surfactant (LSC) delivery systems in neonatal broilers against E. coli septicemia. Embryonated eggs, which had been incubated for 18 days, received either 50 μg of CNT-CpG-ODN, 50 μg of LSC-CpG-ODN, 50 μg of unformulated CpG-ODN, or saline. Four days after exposure to CpG-ODN (day 1 posthatch), 1 x 10(4) or 1 x 10(5) colony-forming units of a virulent strain of E. coli isolated from a turkey with septicemia were inoculated subcutaneously in the neck. Clinical signs, pathology, bacterial isolations from the air sacs, and mortality were observed for 8 days following challenge with E. coli. Bacterial isolations and pathologic observations were conducted immediately after birds were dead or euthanatized. The survival rate of birds in groups receiving saline following E. coli infection was 20% to 30%. In contrast, birds receiving CpG-ODN formulations had a significantly higher survival rate of 60% to 80% (P0.01). Bacterial loads and clinical scores were significantly lower (P0.05) in groups treated with CNT- or LSC-CpG-ODN compared to the groups receiving CpG-ODN or saline. Moreover, there is no evidence of any adverse effects of these formulations in any organs or in growth rates of birds until 42 days of age. This is the first time that CpG-ODN formulated with CNT and LSC have been demonstrated to have an immunomodulatory effect against an E. coli infection in neonatal broiler chickens following in ovo delivery.

Objective: The objective of this study was to assess respiratory outcomes and environmental exposure levels of workers in cage-housed and floorhoused poultry operations. Methods: Poultry operations were evaluated for total dust, endotoxin, and ammonia, and respiratory symptoms and lung function tests of workers were conducted. Results: Workers in floor-housed poultry operations had significantly greater exposures to total dust and ammonia, whereas workers from cage-housed poultry operations reported greater frequency of current and chronic symptoms overall and significantly greater current and chronic phlegm (39% vs 18% and 40% vs 11%, respectively). Endotoxin concentration (EU/mg) was a significant predictor (P 0.05) of chronic phlegm for all poultry workers. Conclusions: Greater endotoxin concentration in the presence of significantly lower total dust, in conjunction with greater respiratory symptoms in workers from cage-housed poultry operations, as compared with workers from floor-housed poultry operations, appears to indicate that differences in environmental exposures may impact respiratory outcomes of workers. (J Occup Environ Med. 2006;48:741–748)

The ineffectiveness of simple delivery of soluble antigens to mucosal membranes for immunization has stimulated extensive studies of strategies for appropriate delivery systems and adjuvants. Biphasic lipid vesicles are formulations suitable for the delivery of proteins, peptides, and oligo/polynucleotides. The purpose of these studies was to investigate the ability of biphasic lipid vesicles (as vaccine-targeting adjuvants) containing a bacterial antigen and unmethylated oligonucleotides containing CGdinucleotides - CpG motifs (CpG ODNs) to induce systemic and mucosal immune responses in pigs. Results showed that while the protein, either alone or with CpG ODNs, did not induce mucosal immune responses, administration of antigen and CpG ODNs in biphasic lipid vesicles resulted in induction of both systemic and local antibody responses after immunization using a combined mucosal/systemic approach.

The nasal mucosa is an important arm of the mucosal system since it is often the first point of contact for inhaled antigens. The ineffectiveness of the simple delivery of soluble antigens to mucosal membranes for immunization has stimulated extensive studies in appropriate delivery systems and adjuvants. We have evaluated biphasic lipid vesicles as a novel intranasal (i.n.) delivery system (designated as vaccine targeting adjuvant, VTA) containing bacterial antigens and CpG oligode-oxynucleotides (ODNs). Results show that administration of antigen and CpG ODNs in biphasic lipid vesicles resulted in greater induction of IgA levels in serum (P

Streptococcus dysgalactiae is a significant pathogen associated with bovine mastitis in lactating and nonlactating dairy cows, causing a severe inflammatory response of the mammary gland, which results in major economic losses to the dairy industry. Two proteins from S. dysgalactiae strain SDG8 were tested for their protective capacity against a homologous bacterial challenge in a dry cow model. The first was a bovine plasmin receptor protein (GapC), which shares 99.4% sequence identity to the plasmin-binding Plr protein of group A streptococci. The second protein product was Mig, a α2-M-, IgG-, and IgA-binding protein present on the cell surface of SDG8. We investigated the efficacy of immunization with purified recombinant forms of GapC and Mig by measuring the number of somatic cells and assessing the presence of the challenge strain in mammary secretions following challenge. In this model, we found that, although the number of quarters containing SDG8 was significantly reduced in the GapC- but not in the Mig-immunized animals, the somatic cell counts from teat secretions were significantly decreased in both the GapC and Mig vaccinates.Key words: Streptococcus dysgalactiae, bovine mastitis, Mig, GapC, GAPDH.

We examined two variants of the genome-sequenced strain, Campylobacter jejuni NCTC11168, which show marked differences in their virulence properties including colonization of poultry, invasion of Caco-2 cells, and motility. Transcript profiles obtained from whole genome DNA microarrays and proteome analyses demonstrated that these differences are reflected in late flagellar structural components and in virulence factors including those involved in flagellar glycosylation and cytolethal distending toxin production. We identified putative sigma(28) and sigma(54) promoters for many of the affected genes and found that greater differences in expression were observed for sigma(28)-controlled genes. Inactivation of the gene encoding sigma(28), fliA, resulted in an unexpected increase in transcripts with sigma(54) promoters, as well as decreased transcription of sigma(28)-regulated genes. This was unlike the transcription profile observed for the attenuated C. jejuni variant, suggesting that the reduced virulence of this organism was not entirely due to impaired function of sigma(28). However, inactivation of flhA, an important component of the flagellar export apparatus, resulted in expression patterns similar to that of the attenuated variant. These findings indicate that the flagellar regulatory system plays an important role in campylobacter pathogenesis and that flhA is a key element involved in the coordinate regulation of late flagellar genes and of virulence factors in C. jejuni.

Synthetic oligodeoxynucleotides (ODN) containing CpG motifs (CpG-ODN) have been shown to be effective immunoprotective agents in murine models for a variety of viral, intracellular bacterial, and protozoan infections. Until now, the use of CpG-ODN to protect against extracellular bacterial infections has not been reported. The objective of this study was to investigate the effect of CpG-ODN against cellulitis and colibacillosis in broiler chickens, using a well-established model. At 22 days of age, birds received CpG-ODN by either the subcutaneous or intramuscular route. Three days later, a virulent isolate of Escherichia coli was applied to a scratch site on the caudal abdominal skin. Birds were examined for 10 days after the E. coli challenge, and pathological and bacteriological assessments were conducted on all birds. The control group of birds receiving no CpG-ODN (2007) had a survival rate of 15%. In contrast, groups that received CpG-ODN (2007) , by either subcutaneous or intramuscular injection, had significantly higher survival rates ( P < 0.0001). Furthermore, the size of the cellulitis lesion was significantly smaller in groups that received CpG-ODN (2007) by the subcutaneous route ( P < 0.01). A dose of as little as 3.16 μg of CpG-ODN (2007) , delivered 3 days prior to challenge by either the subcutaneous or intramuscular route, significantly protected birds against E. coli infection ( P < 0.01). This study demonstrates that CpG-ODN (2007) has both local and systemic protective effects in broiler chickens. This is the first time that CpG-ODN (2007) has been demonstrated to have an immunoprotective effect against an extracellular bacterial infection in any food animal species.

The adjuvanticity of a synthetic oligodeoxynucleotide containing unmethylated CpG motifs (CpG ODN) was determined in cattle. Calves were immunized with a truncated secreted version of glycoprotein D (tgD) of bovine herpes virus-1 (BHV-1) formulated with alum, CpG ODN, or a combination of both. BHV-1 tgD formulated with CpG ODN or with alum and CpG ODN induced a stronger and more balanced immune response than tgD in alum. This level of immunity was of sufficient magnitude to minimize weight loss and significantly reduce the duration of virus shedding after intranasal viral challenge. Local tissue reactions generated by CpG ODN were very mild and transient, whereas reactions induced by alum or a combination of CpG ODN and alum were moderate in severity and duration. These data demonstrate that CpG ODN causes minimal injection site reactions and yet acts as an effective adjuvant in cattle.

We tested the capacity of biochar (made at 450 °C from a common reed species) to neutralise pH and remove metals in two acid drainage waters (pH 2.6 and 4.6) using column leaching and batch mixing experiments. In the column experiments, the acid drainage water was neutralised upon passage through the biochar with substantial increases (4–5 pH units) in the leachate pH. In the batch experiments, the leachate pH remained above 6.5 when the drainage:biochar ratio was less than approximately 700:1 (L acid drainage:kg biochar) and 20:1 for the pH 4.6 and pH 2.6 drainage waters, respectively. Dissolved metal concentrations were reduced by 89–98 % (Fe ≈ Al > Ni ≈ Zn > Mn) in the leachate from the biochar. A key mechanism of pH neutralisation appears to be solid carbonate dissolution as calcite (CaCO3) was identified (via X-ray diffraction) in the biochar prior to contact with acid drainage, and dissolved alkalinity and Ca was observed in the leachate. Proton and metal removal by cation exchange, direct binding to oxygen-containing functional groups, and metal oxide precipitation also appears important. Further evaluation of the treatment capacity of other biochars and field trials are warranted.

A study was conducted to evaluate the acute health effects of wearing an N-95 disposable respirator in a swine confinement facility.A crossover trial design was used in the study.The study was carried out at the research facilities of the Centre for Agricultural Medicine, the Royal University Hospital, and the Prairie Swine Centre Inc, Saskatoon, Saskatchewan, Canada.Twenty-one nonsmoking healthy male subjects with no previous swine barn exposure participated in the study.The subjects participated in a laboratory session (baseline day), a 4-h exposure in a traditional swine room wearing the respirator (intervention day), and a 4-hour exposure in a traditional swine room without a respirator (nonintervention day).Lung function, methacholine challenge tests, blood counts, nasal lavage, and cytokines in serum and nasal lavage fluid.Mean (+/- SE) shift change in FEV(1), from preexposure to postexposure, was highest on nonintervention day (-8.1+/-1.01%) and was significantly different from intervention day (0.32+/-0.62%; p0.0001) and baseline day (1.57+/-0.51%; p0.0001). Similar patterns were observed in the mean values of the provocative concentration of a substance (methacholine) causing a 20% fall in FEV(1) (nonintervention day, 130.4+/-36.9 mg/mL; intervention day, 242.0+/-38.0 mg/mL; and baseline day, 328.0 mg/mL +/-34.1 mg/mL). Significant increases in serum neutrophil levels and nasal cell counts were observed on the nonintervention day in comparison to the baseline and intervention days. Significant increases also were found in the levels of cytokines interleukin (IL)-6 and IL-8 in nasal lavage fluid and in the levels of IL-6 in serum for the nonintervention day in comparison to the other 2 days.The results demonstrate that an N-95 disposable respirator can help to significantly reduce acute negative health effects in subjects not previously exposed to a swine barn environment.

An expression library was constructed from an Actinobacillus pleuropneumoniae serotype 1 clinical isolate using a plasmid vector. The library was screened with serum raised against the culture supernatant of this strain. One Escherichia coli transformant which also reacted with convalescent serum was isolated and found to express a protein with an electrophoretic mobility of approximately 50,000. The A. pleuropneumoniae-derived DNA encoding the protein was localized and characterized by nucleotide sequence analysis and primer extension mapping. One open reading frame of 1,095 bases was detected and confirmed by TnphoA insertion mutagenesis. It encoded a protein with a calculated molecular mass of 40 kDa which was lipid modified and present in the outer membrane and in membrane blebs of A. pleuropneumoniae. This protein was designated as outer membrane lipoprotein A (OmlA), and the encoding gene as omlA. Southern blotting under low-stringency conditions revealed the presence of hybridizing sequences in all A. pleuropneumoniae type strains, and a specific serum detected a homologous protein in serotypes 2, 8, 9, 11, and 12 type strains. Pigs immunized with this recombinant protein preparation were protected from death in an aerosol challenge experiment with an A. pleuropneumoniae serotype 1 isolate.

The effects of repeated exposure to a range of doses of swine barn dust (SBD) on airway hyperresponsiveness (AHR) and inflammation were evaluated using a mouse model system. A number of components, including endotoxin and a number of feed proteins, were identified in SBD, and mice were exposed 20 min/d for 14 d to a log dilution series of nebulized SBD suspensions. AHR to methacholine was measured using head-out whole-body plethysmography, and the methacholine concentration inducing a 20% decrease in pulmonary airflow (PC(20) MCh) was calculated. At the end of the 14-d exposure period, bronchoalveolar lavage (BAL) fluids were recovered, cytokines (interleukin [IL]-1beta, IL-6, keratinocyte-derived chemokine [KC], and tumor necrosis factor [TNF]) in BAL were measured by enzyme-linked immunosorbent assay (ELISA), and leukocytes in BAL were counted. The PC(20) MCh was significantly lower in the group of mice that were exposed to the highest concentration of SBD than in controls or the group exposed to the lowest level of dust. Likewise, the group that was exposed to the highest level of SBD had significantly higher levels of IL-1beta, KC, and TNF than controls and some other groups. There were substantially more lymphocytes and monocytes in the BAL from mice that were exposed to the higher levels of SBD for the 14-d period, but neutrophils were not a part of this response. The SBD exposures used in these experiments induced chronic inflammatory phenotype responses, as indicated by the predominance of lymphocytes and monocytes, but not neutrophils, in BAL and by inflammatory cytokines detected. The association between the PC(20)MCh and dose of SBD suggests that a threshold of susceptibility occurs after a relatively low, chronic exposure to SBD.

Background: Individuals engaged in work in intensive animal houses experience some of the highest rates of occupationally related respiratory symptoms. Organic dust and in particular endotoxin has been mostclosely associatedwith respiratorysymptomsand lung function changes in workers. It has previously been shown that for intensive poultryoperations, type of poultry housing [cage-housed (CH) versus floor-housed (FH)] can influence the levels of environmental contaminants. The goal of the study was to determine the differences in endotoxin and dust levels at different size fractions between CH and FH poultry operations. Methods:FifteenCH and15FHpoultryoperations weresampledforstationarymeasurements (area) of dust and associated endotoxin. Fractioned samples were collected utilizing Marple cascade impactors. Gravimetric and endotoxin analysis were conducted on each of the filters. Results: When assessed by individual Marple stage, there was significantly greater airborne endotoxin concentration (endotoxin units per cubic meter) in the size fraction >9.8 mm for the FH operations whereas at the size fraction 1.6‐3.5 mm, the CH operations had significantly greater airborne endotoxin concentration than the FH operations. Endotoxin concentration in the dust mass (endotoxin units per milligram) was significantly greater in the CH operations as compared to the FH operations for all size fractions >1.6 mm. As such, endotoxin in the respirable fraction accounted for 24% of the total endotoxin in the CH operations whereas it accounted for only 11% in the FH operations. There was significantly more dust in all size fractions in the FH operations as compared to the CH poultry operations. Conclusions: There is more endotoxin in the presence of significantly lower dust levels in the respirable particle size fractions in CH poultry operations as compared to the FH poultry operations. This difference in respirable endotoxin may be important in relation to the differential respiratory response experienced by CH and FH poultry operation workers.