Folic acid enhances proinflammatory and antiviral molecular pathways in chicken B-lymphocytes infected with a mild infectious bursal disease virus.

1. This study evaluated the effect of folic acid (FA) supplementation on the proinflammatory and antiviral molecular pathways of B-lymphocytes infected with a modified live IBDV (ST-12) mild vaccine strain during a timed post-infection analysis.2. A chicken B-lymphocytes (DT-40) cell line was cultured in triplicate at a concentration of 5 × 105 cells per well in 24-well plates; and was divided into three groups: 1: No virus, FA; 2: Virus, no FA; 3: Virus + FA at a concentration of 3.96 mM. The experiment was repeated three times.3. Cells in groups 2 and 3 were infected with a modified live IBDV (ST-12) mild vaccine strain at one multiplicity of infection (MOI: 1). After 1 hour of virus adsorption, samples were collected at 0, 3, 6, 12, 24 and 36 hours post-infection (hpi).4. The modified live IBDV (ST-12) mild vaccine strain triggered a B-lymphocyte specific immune response associated with the upregulation of genes involved in virus recognition (Igß), virus sensing (TLR-2, TLR-3, TLR-4 and MDA5), signal transduction and regulation (TRIF, MyD88 and IRF7), and the antiviral effector molecules (IFN-α, OAS, PKR, and viperin).5. FA supplementation modulated IBDV replication and regulated the proinflammatory and antiviral downstream molecular pathways.6. In conclusion, the low virulent pathotype serotype I modified live IBDV (ST-12) mild vaccine strain was able to trigger and mount an immune response in chicken B-lymphocytes without affecting B-cell viability. FA supplementation modulated B lymphocytes response and improved their innate immune proinflammatory and antiviral response molecular pathways.

Differential immunomodulatory effect of vitamin D (1,25 (OH)2 D3) on the innate immune response in different types of cells infected in vitro with infectious bursal disease virus.

It has been demonstrated that vitamin D (Vit D) included in diets offers a beneficial effect by improving innate immune responses in chickens. However, its mechanisms of action and the effect on immunosuppressive pathogens, such as infectious bursal disease virus, are not yet known. In the present study, we have studied the immunomodulatory effect of Vit D on the innate immune response in 3 cell lines: fibroblast cells (DF-1), macrophages (HD11), and B cells (DT-40) infected with IBDV (intermediate vaccine) at 2 multiplicity of infections (MOI) (1 and 0.1). Genes associated with innate immune responses (TLR-3, TLR-21, MDA-5, MyD88, TRIF, IRF-7, INF-α, INF-ß, PKR, OAS, viperin, IL-1ß, IL-6, and IL-12) were evaluated at different time points (3, 6, 12, 24, and 36 h after infection, h.p.i). Virus production reached a maximum at 24 h.p.i., which was significantly (P < 0.05) higher in DF-1 cells, followed by HD-11 and DT-40 cells. Mainly in HD-11 cells, there was a significant (P < 0.05) effect of Vit D supplementation on receptors TLR-3, TLR-21, and MDA-5 after 12 h.p.i, independent of MOI. DT-40 cells showed the highest antiviral activity, with a significant (P < 0.05) effect on IRF-7, IFN-ß, OAS, and PKR gene expression, where expression of IRF-7 and IFN-ß correlated positively with Vit D supplementation, while OAS and PKR were independent of Vit D. Proinflammatory cytokines were significantly (P < 0.05) upregulated and found to be Vit D and MOI dependent. In conclusion, this study demonstrated the capacity of IBDV to trigger a strong innate immune response in chicken cells and contributes to the understanding of the activation pathways of innate immunity induced by IBDV and further shows the benefitial effect of Vit D supplementation as an immunomodulator.

Bioassay of estrogenicity and chemical analyses of estrogens in streams across the United States associated with livestock operations.

Animal manures, used as a nitrogen source for crop production, are often associated with negative impacts on nutrient levels in surface water. The concentrations of estrogens in streams from these manures also are of concern due to potential endocrine disruption in aquatic species. Streams associated with livestock operations were sampled by discrete samples (n = 38) or by time-integrated polar organic chemical integrative samplers (POCIS, n = 19). Samples were analyzed for estrogens by gas chromatography-tandem mass spectrometry (GC-MS(2)) and estrogenic activity was assessed by three bioassays: Yeast Estrogen Screen (YES), T47D-KBluc Assay, MCF-7 Estrogenicity Screen (E-Screen). Samples were collected from 19 streams within small (≈ 1-30 km(2)) watersheds in 12 U.S. states representing a range of hydrogeologic conditions, dominated by: dairy (3), grazing beef (3), feedlot cattle (1); swine (5); poultry (3); and 4 areas where no livestock were raised or manure was applied. Water samples were consistently below the United Kingdom proposed Lowest Observable Effect Concentration for 17ß-estradiol in fish (10 ng/L) in all watersheds, regardless of land use. Estrogenic activity was often higher in samples during runoff conditions following a period of manure application. Estrone was the most commonly detected estrogen (13 of 38 water samples, mean 1.9, maximum 8.3 ng/L). Because of the T47D-KBluc assay's sensitivity towards estrone (1.4 times 17ß-estradiol) it was the most sensitive method for detecting estrogens, followed by the E-Screen, GC-MS(2), and YES. POCIS resulted in more frequent detections of estrogens than discrete water samples across all sites, even when applying the less-sensitive YES bioassay to the POCIS extracts.