Aflatoxin B1 (AFB1) is a mutagen that has been categorized as a group 1 human carcinogen by the International Agency for Research on Cancer. It is produced as a secondary metabolite by soil fungi Aspergillus flavus and Aspergillus parasiticus. Here, in this study, the effect of AFB1 on the structure and conformation of bovine serum albumin (BSA) using multispectroscopic tools like fluorescence spectroscopy, ultraviolet–visible absorption spectroscopy, and circular dichroism spectropolarimetry has been ascertained. Ultraviolet absorption spectroscopy revealed hyperchromicity in the absorption spectra of BSA in the presence of AFB1. The binding constant was calculated in the range of 104 M–1, by fluorescence spectroscopy suggesting moderate binding of the toxin to BSA. The study also confirms the static nature of fluorescence quenching. The stoichiometry of binding sites was found to be unity. The competing capability of warfarin for AFB1 was higher than ibuprofen as calculated from site marker displacement assay. Förster resonance energy transfer confirmed the high efficiency of energy transfer from BSA to AFB1. Circular dichroism spectropolarimetry showed a decrease in the α-helix in BSA in the presence of AFB1. The melting temperature of BSA underwent an increment in the presence of a mycotoxin from 62.5 to 70.3 °C. Molecular docking confirmed the binding of AFB1 to subdomain IIA in BSA.
Resveratrol is a polyphenol belonging to the class stilbenes. The active and stable form of resveratrol is trans-resveratrol. This polyphenol is bestowed with numerous biological properties. Aflatoxin B1 is a hepato-carcinogen and mutagen that is produced by Aspergillus species. In this study, the interaction of trans-resveratrol with HSA followed by competitive dislodging of AFB1 from HSA by trans-resveratrol has been investigated using spectroscopic studies. The UV-absorption studies revealed ground state complex formation between HSA and trans-resveratrol. Trans-resveratrol binds strongly to HSA with the binding constant of ~ 107 M-1 to a single binding site (n = 1.58), at 298.15 K. The Stern-Volmer quenching constant was calculated as 7.83 × 104 M-1 at 298.15 K, suggesting strong fluorescence quenching ability of trans-resveratrol. Site markers displacement assay projected subdomain IIA as the binding site of trans-resveratrol to HSA. The molecular docking approach envisages the amino acid residues involved in the formation of the binding pocket. As confirmed from the site marker displacement assays, both trans-resveratrol and AFB1 binds to HSA in the same binding site, subdomain IIA. The study explores the ability of trans-resveratrol to displace AFB1 from the HSA-AFB1 complex, thereby affecting the toxicokinetic behavior of AFB1 associated with AFB1 exposure.
4 beta, 15 Diacetoxyscirpenol (DAS) mycotoxin produced by Fusarium species was tested for detrimental effects on macrophage viability, phagocytosis, and Fc-receptor expression. Sephadex-elicited chicken abdominal cells were harvested to establish adherent macrophage monolayers on glass coverslips. Coverslips were then assigned randomly to treatment groups (0, 12.5 and 25 micrograms/mL DAS). Macrophage monolayers were exposed to treatments for 1 h, washed, and tested for various functional endpoints. Treatment with DAS resulted in decreased viability of macrophages (90.8% vs 81.5% vs 70.4% viable in the 0, 12.5 and 25 micrograms treatments, respectively) and decreased the percentage of macrophages phagocytizing sheep erythrocytes (81.6% vs 53.1% vs 46.0%. DAS also caused a decrease in the mean number of opsonized cells engulfed per phagocytic macrophage (5.7 vs 3.7 vs 2.9). A similar trend was observed using unopsonized sheep erythrocytes (15.4% vs 7.6% vs 5.5% phagocytic macrophages and 0.29 vs 0.11 vs 0.08 erythrocytes engulfed per macrophage). The incidence of Fc-receptor positive macrophages determined by sheep erythrocyte rosetting was also decreased in DAS-treated macrophages as compared to the control (49.2% vs 32.7% vs 24.2%). The findings of this study demonstrate that DAS exposure causes a suppression in macrophage phagocytic function and therefore may alter the first line of immunological defense in chickens.
genotypes for iNOS gene expression and activity. Two trials were conducted to compare these lines for immune parameters. For mononuclear phagocytic function quantification, 5 chickens per group were injected i.v. with Black India Ink at a rate of 1 mL/kg body weight (BW) at 4 wk. Blood samples were drawn and optical density measured at 640 nm on plate reader. K-strain was significantly better in clearing carbon (P = 0.04) than the GB1 and GB2 chickens at 10 min post i.v. injection. Lymphoproliferation was measured in vitro by incubating 0.2 x 10 peripheral blood leukocytes with 25 µg Concanavalin A. K-strain chicks had the highest 6 lymphoproliferation index than the GB1 and GB2. On the contrary, K-strain chicks had a weaker PHAPmediated toe-web swelling response as compared with the GB1 and GB2 chicks. No differences were observed between genotypes for the total or IgG anti-SRBC antibodies in primary or secondary immunization response (one trial). However, K-strain exhibited greater persistence (P < 0.05) of IgM levels than GB2 chicks during the decline phase of the booster response. While K chicks had heavier BW (P < 0.01), they had smallest bursal, thymic and splenic weights (relative to BW) vs. GB1 and GB2 in both trials. These findings show that the iNOS hyper responder K-strain chickens also perform better for monocyte/macrophage, Tlymphocyte and perhaps even B-lymphocyte functions even though their lymphoid organ growth lagged behind the iNOS hyporesponder (GB1 and GB2) genotypes.
A major goal of many poultry producers is to attain good flock liveability. Historically, most poultry producers have manipulated environmental conditions and management to maximize bird health. In the past two decades there has been much research into nutritional regimes that improve bird health through immunomodulation. Commercial poultry environments contain ubiquitous micro-organisms that continuously challenge the immune system. Nutritional supplements that enhance immune system function may improve flock performance and be economically advantageous. This paper reviews the literature on zinc-methionine and the avian cellular immune system. Current knowledge of the effects of zinc on many animal models is reviewed and a hypothetical mechanism for the action of zinc-methionine on this system is discussed.
Beta‐1,3/1,6‐glucan (β‐glucan) was tested as a possible immunomodulator. Chicken macrophages from a macrophage cell line MQ‐NCSU and from abdominal exudate of broiler chickens were exposed to various concentrations of β‐glucan in vitro. In addition, day‐old broiler chicks were fed a diet containing 0, 20, and 40 mg/kg β‐glucan in the starter and 0, 20, and 20 mg/kg in the grower diet. Several baseline immune parameters were examined following such exposures. The results showed that β‐glucan exposure increased nitrite and interleukin‐1 (IL‐1) production as well as induced macrophage to proliferate in culture. However, IL‐6 production was not affected. Dietary β‐glucan supplementation increased the macrophage phagocytic activity, anti‐sheep red blood cells antibody response post‐boost, as well as the PHA‐P‐mediated lymphoproliferative response measured as a toe‐web swelling. The percentage of CD4+, CD8+, and CD4+/CD8+ double positive lymphocytes in the intestinal intraepithelial leukocytes was increased in β‐glucan supplemented chicks. Furthermore, the primary and secondary lymphoid organs such as bursa of Fabricius, thymus and spleen were larger in β‐glucan‐supplemented chicks as compared to the chicks on basal diet. The findings of these studies which showed that β‐glucan improves several base‐line immune responses in the chicken imply that β‐glucan can be used as a possible immunomodulator in food animals such as the chicken.
Enteric disorders predispose poultry to malnutrition. The objectives of this paper were 1) to simulate the inanition of poult enteritis mortality syndrome by restricting feed intake and 2) to develop a drinking water supplement that supports the immune functions of poults with inanition. Poults were restricted to 14 g of feed/d for 7 d beginning at 14 d of age then fed ad libitum until 36 d (recovery). The control was fed ad libitum. During the feed-restriction period, duplicate groups of 6 poults received 1 of 5 drinking water treatments: 1) restricted feed, unsupplemented water; 2) restricted feed + electrolytes (RE); 3) RE + glucose + citric acid (REGC); 4) REGC + betaine (REGCB); or 5) REGCB + zinc-methionine (REGCBZ). Immunological functions were assessed by inoculating poults with SRBC and B. abortus (BA) antigen at 15, 22, and 29 d of age. Antibody (Ab) titers were determined 7 d later for primary, secondary, and recovery responses. The primary and secondary total Ab titers to SRBC for restricted feed were 4.71 and 6.16 log3, which where lower (P < 0.05) than for controls (8.00 and 9.66 log3) and the other treatments. The recovery Ab titer for controls was 10.7, significantly higher than restricted feed (8.71) and RE (8.10) groups but not different from other treatments. The primary total Ab responses to BA were significantly lower in the restricted feed and RE groups as compared with the control and other treatments. Although feed restriction of poults to maintenance reduces the humoral immune responses, these responses can be significantly improved by drinking water containing electrolytes and especially sources of energy such as glucose and citric acid.
The role of a novel "small round virus" (SRV) isolated from poult enteritis and mortality syndrome (PEMS) cases in inducing PEMS and associated immune alterations was examined in this study. Specific-pathogen-free and conventional poults were orally challenged with SRV and/or turkey coronavirus and monitored for clinical signs. Intestines, thymus, bursa, and spleens were examined for SRV antigen at various days postinoculation (DPI). Peripheral blood lymphocytes (PBLs), thymocytes, and splenic lymphocytes from inoculated poults or lymphocytes isolated from healthy poults after incubation with SRV in vitro were examined for lymphoproliferative potential against concanavalin A (Con A). The incidence of lymphocyte subpopulations in the peripheral blood and thymic lymphocytes of SRV-challenged poults was examined by flow cytometry. The results of these studies showed that the SRV challenge induced diarrhea, growth suppression, and atrophy of thymus and bursa resembling those of PEMS in field and/or experimental infections. The SRV antigen was detected in intestinal tissues soon after infection (i.e., at 2 and 4 DPI), whereas lymphoid tissues such as thymus, bursa, and spleen were positive for SRV antigen starting at 4 DPI until 8 DPI, suggesting virus translocation to lymphoid organs. The responsiveness of PBLs to Con A at 2 DPI was significantly reduced in all virus challenge groups (e.g., 28% and 22% in the SRV-alone group in studies 1 and 2, respectively) below the uninfected group. However, this suppressed response was no longer evident in the SRV group by 7 DPI. The SRV incubation with normal thymocytes and splenocytes in vitro resulted in significantly reduced lymphoproliferative response against Con A (41.2% and 10.49% reductions at 1:50 SRV dilution vs. controls in thymocytes and splenocytes, respectively). Flow cytometry analysis revealed a sudden decline at 2 DPI in the numbers of CD4- CD8+ lymphocyte subset in PBLs of SRV-infected poults. However, by 8 DPI, SRV-challenged poults had relatively higher CD4- CD8+ lymphocytes in PBLs. On the contrary, thymocytes had higher percentages of CD4- CD8+ lymphocytes at 2 and 4 DPI and reached comparable levels at 8 DPI in controls and SRV-infected poults. No differences were observed in CD4+ CD8- lymphocyte numbers in controls vs. SRV-infected poults. The findings of these studies imply that SRV may be a promising primary etiologic agent of PEMS. Furthermore, the SRV infection may compromise the lymphocyte-mediated immune defenses by reducing lymphoproliferation and the CD4- CD8+ (presumably T-cytotoxic cells) lymphocytes during the acute stage of SRV infection.
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