Fractionation of swine barn dust and assessment of its impact on the respiratory tract following repeated airway exposure.

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.

Oligodeoxynucleotides containing CpG motifs (CpG-ODN) predominantly induce Th1-type immune response in neonatal chicks.

Earlier, we demonstrated that intramuscular administration of oligodeoxynucleotides containing CpG motifs (CpG-ODN) induces protection in neonatal chicks against a lethal challenge of Escherichia coli. However, the mechanism of induction of the protection was not clear. In an attempt to elucidate the mechanism of induced protection, we determined the kinetics of expression of cytokines/chemokines in the spleen and bursa of Fabricius of newly hatched chicks that had received intramuscular administration of CpG-ODN or non-CpG ODN compared to saline-treated controls. SyBr green, real-time quantitative reverse transcription polymerase chain reaction (RT-PCR) analysis of the RNA demonstrated increased expression of IL-1beta, IL-6, IL-8, IL-10, IL-18, IFN-gamma and MIP-3alpha mRNAs in the spleen and; IL-10 and IFN-alpha in bursa of Fabricious of chicks that had received CpG-ODN. However, non-CpG ODN failed to induce any of the cytokine. The increased level of IL-18 and IFN-gamma but not IL-4 mRNA suggests that the administration of CpG-ODN elicits a Th1 biased immune response, which may be important in inducing protection against infections in neonatal chicks. To our knowledge, this is the first report evaluating the induction of cytokines/chemokines in neonatal chicks following administration of CpG-ODN.

Acquisition of plasmin activity and induction of arachidonic acid release by Streptococcus suis in contact with human brain microvascular endothelial cells.

Infections caused by Streptococcus suis, a major swine pathogen, include meningitis, arthritis, pneumonia and septicaemia. In this study, we investigated interactions that may occur between human brain microvascular endothelial cells (HBMEC), the main constituent of the blood-brain barrier, and S. suis. We show that S. suis acquires plasmin activity in a time-dependent manner when in contact with cultured HBMEC. Cell-associated plasmin activity reached a plateau following a 48h co-incubation period. Zymography analysis revealed that HBMEC produce urokinase, which is probably involved in activation of plasminogen bound to S. suis. We also show that a S. suis culture supernatant which possesses both phospholipase C and haemolysin (suilysin) activities was able to induce the release of arachidonic acid from the membrane of HBMEC. Evidence suggests that the action of suilysin on HBMEC may be a prerequisite for the action of additional molecules such as phospholipase C. These new biological effects associated with S. suis may play an important role in the migration of S. suis through the blood-brain barrier and in the modulation of local inflammation.

Prevalence of chronic bronchitis in farm and nonfarm rural residents in Saskatchewan.

OBJECTIVE: To determine the prevalence of chronic bronchitis (CB) and associated risk factors in farm and nonfarm rural residents in Saskatchewan, Canada. METHODS: The questionnaire collected information about health, contextual, and individual factors from 8261 farm and nonfarm adult residents (18 years and older). RESULTS: 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. CONCLUSION: The results suggest that increasing household income and reducing smoking could be primary, modifiable determinants of CB prevalence.

Size distribution of particulate and associated endotoxin and bacteria in traditional swine barn rooms and rooms sprinkled with oil.

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., particles <4.7 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.

Gender-related tumor necrosis factor-alpha responses in naïve volunteers with Toll-like receptor 4 polymorphisms exposed in a swine confinement facility.

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.

Investigation of a novel DNase of Streptococcus suis serotype 2.

A secreted nuclease, SsnA, was identified in the virulent Streptococcus suis isolate SX332 and subsequently in each of the type strains of capsular serotypes 1 through 9. Screening of 258 porcine clinical isolates from surface (nasal mucosa or palatine tonsil) or internal (joint, brain or other internal organ) locations revealed a significant relationship (P < 0.001) between expression of nuclease and isolation from an internal site. A 3,126-bp gene, ssnA, was identified from a phenotypically nuclease-negative pGh9:ISS1 insertion mutant, and analysis of the predicted SsnA sequence revealed a 35-amino-acid (aa) secretion signal sequence, a 22-aa DNA-binding domain, and a typical gram-positive cell wall sorting motif. A requirement of Ca2+ and Mg2+ for SsnA activity was determined, and the substrate specificity was found to be for single- and double-stranded linear DNA. Reverse transcription-PCR experiments revealed that ssnA is expressed throughout all stages of S. suis growth, and Western blots with porcine anti-S. suis immune sera against a recombinant, truncated SsnA derivative (rSsnADelta) confirmed that SsnA is expressed in vivo. Furthermore, anti-rSsnADelta antibodies were sufficient to neutralize SsnA activity. Analyses of subcellular fractions of SX332 and derived mutants, on DNA-containing polyacrylamide gels and by Western blotting, suggest that SsnA is cell wall located.

Immunisation of dairy cattle with recombinant Streptococcus uberis GapC or a chimeric CAMP antigen confers protection against heterologous bacterial challenge.

The gapC genes, encoding the cell surface-associated GapC proteins of S. uberis and S. agalactiae, have been cloned and sequenced. To identify potential vaccine candidates against S. uberis-induced bovine mastitis, lactating dairy cows were vaccinated with either (6 x His)GapC of S. uberis or S. dysgalactiae, or with a chimeric CAMP-factor antigen, CAMP-3. For 7 days following heterologous challenge with S. uberis, milk somatic cell counts were determined to assess differences in the severity of mastitis between vaccinates and an unvaccinated control group. Vaccination with S. uberis (6 x His)GapC or CAMP-3 resulted in a significant reduction in inflammation on several days post-challenge, most significantly for the former antigen. Inflammation was not reduced in S. dysgalactiae (6 x His)GapC vaccinates, suggesting that it does not confer cross-species protection.