Summary A pilot study was conducted to evaluate the suitability of hepatic aflatoxin (AFB1) concentration as a biomarker to assess the in vivo efficacy of a mycotoxin binder in poultry when AFB1 dietary concentrations are low. Diets containing low doses of AFB1 without or with Mycosorb ® (MTB), a mycotoxin binder, were fed to broilers from 7 to 21 days of age. The accumulation of AFB1 in liver was measured by high performance liquid chromatography with a detection limit of 5 ng/kg liver. In response to 10 and 50 µg AFB1/kg feed, hepatic AFB1 accumulation was 27 and 145 ng AFB1/kg liver, respectively. At each dietary concentration of AFB1, the inclusion of 5 g MTB/kg of feed reduced ( P < 0.1 for 10 μg AFB1/kg feed and P < 0.05 for 50 μg AFB1/kg feed) hepatic AFB1 accumulation by at least 50%. These results suggest that hepatic AFB1 concentration is a suitable biomarker for evaluating mycotoxin binder efficacy in poultry fed the EU maximum dietary concentration of 10 µg of AFB1/kg feed.
This study explored the hypothesis that mannan oligosaccharide (MOS) acts to reduce systemic inflammation in pigs by evaluating cytokine production of alveolar macrophages (AM) and serum cytokine concentrations. A total of 160 pigs were fed diets containing 0.2 or 0.4% MOS for 2 or 4 wk postweaning compared with control diets without MOS. Dietary MOS did not affect the serum concentration of tumor necrosis factor (TNF)-α and tended (P = 0.081) to increase that of IL-10. These cytokine concentrations also changed over time (P < 0.001). After 2-wk feeding of the control or MOS diets, AM were collected and stimulated ex vivo with lipopolysaccharide (LPS) or polyinosinic:polycytidylic acid (PLIC) as infection models. The LPS-stimulated AM from MOS-fed pigs (n = 12) secreted less TNF-α (P < 0.001) and more IL-10 (P = 0.026) than those from control-fed pigs (n = 6). However, dietary MOS had less effect on ex vivo TNF-α and IL-10 production by PLIC-stimulated AM (P = 0.091 and P > 0.10, respectively. Further, effects of MOS were examined in 4 in vitro experiments. In Exp. 1 (n = 4 pigs), MOS and mannan-rich fraction (MRF), when added to AM cultures, were able to increase TNF-α production. This direct effect of MOS was not due to endotoxin contamination as verified in Exp. 2 (n = 6 pigs) using polymyxin B, an inhibitor of LPS activation of toll-like receptor 4. Polymyxin B inhibited production of TNF-α by AM after treatment with LPS (P < 0.001), but not after treatment with MOS in the absence of LPS (P > 0.70). In Exp. 3 (n = 6 pigs), when MOS was directly applied in vitro, the pattern of cytokine production by LPS-activated AM was similar to that observed ex vivo, as MOS suppressed LPS-induced TNF-α (P < 0.001) and enhanced LPS-induced IL-10 (P = 0.028). In Exp. 4 (n = 6 pigs), when MRF replaced MOS, AM-produced TNF-α induced by LPS or PLIC was suppressed by MRF (P = 0.015 or P < 0.001, respectively). These data establish that MOS and MRF suppress LPS-induced TNF-α secretions by AM. Generally, the study suggests that MOS may be a potent immunomodulator because it directly activates AM to secrete TNF-α and alters the cytokine responses of bacterial endotoxin-induced AM in both ex vivo and in vitro systems. In particular, feeding MOS to pigs for 2 wk reduces TNF-α and increases IL-10 concentrations after ex vivo treatment of AM with LPS. These immunomodulatory properties of MOS may have important implications for both host defense and avoidance of harmful overstimulation of the immune system.
The aim of this paper was to provide a comprehensive toxicological and safety evaluation of a yeast cell wall preparation (YCWP) for use as an animal feed ingredient. The following toxicological assessments were carried out: the mutagenic activity was tested using the Ames' Test in five Salmonella typhimurium strains; clastogenic activity was investigated using the mammalian micronucleus test in Swiss ICO OF1 (IOPS Caw) mice; genotoxic activity was assessed using the in vitro mammalian chromosomal aberration test in human lymphocytes; acute oral toxicity was tested by administration of a single dose of 2000 mg/kg BW. Eye and skin irritation were assessed in rabbits according to OECD guidelines; skin sensitivity was established in guinea pigs by means of the Buehler test, while acute dermal and inhalation studies in rats were further completed, also according to OECD guidelines. All conducted tests were considered valid under the experimental conditions. No significant mutagenic activity or genotoxic activity was observed, and it was concluded that the test article did not induce any clastogenic effect. YCWP was found to be mildly irritating to the eye, slightly irritating to the skin but was found to be non-sensitizing in the guinea pig. The acute oral, dermal and inhalation studies did not yield any evidence of gross toxicity or pharmacological effects.
In this paper we describe a study that evaluates the applicability of an in vitro fermentation model to assess the resistance of protein supplements to rumen degradation. The protein sources used were: soybean meal (SBM); whey protein (WHEY), which was expected to be rapidly degraded, and yeast-derived microbial protein (YMP), which was proposed to be resistant to rumen degradation. The basal diet was composed of grass silage and a commercial compound feed. The protein supplements were added at three isonitrogenous doses. Fermentation was monitored for 24 h and gas production, volatile fatty acids, lactic acid, and ammonia were analyzed at three timepoints. Protein degradation was estimated by determining the extent to which branched-chain amino acids (BCAA) introduced with the protein supplement were converted to corresponding branched-chain volatile fatty acids (BCVFA). At the highest dose of WHEY, 60% of introduced valine, leucine, and isoleucine was recovered as isobutyric, 2-methylbutyric, and isovaleric acid (products of BCAA decarboxylation and deamination), respectively. The BCVFA detected represented 50% of added BCAA with SBM, but <15% with YMP. Further indications that YMP protein is resistant to degradation were provided by analysis of ammonia. With YMP, the residual ammonia concentration only marginally exceeded that of the cultures with no protein supplementation, while it increased dose-dependently when the vessels were supplemented with WHEY or SBM. This suggests that with WHEY and SBM, the rate of deamination exceeded the rate of ammonia assimilation by bacteria. Residual ammonia and BCVFA, the two indicators of protein fermentation, were strongly correlated. Overall bacterial activity was monitored as yield of gas, volatile fatty acids, and bacteria. These three correlating parameters showed that WHEY only modestly stimulated fermentation, whereas SBM and YMP stimulated fermentation extensively, possibly owing to their higher carbohydrate content. The results presented suggest that the in vitro fermentation method was suitable for detecting differences in resistance of protein supplements to rumen degradation and following a full method validation could be a useful tool for diet formulation. The data obtained suggested that YMP was the most resistant and WHEY the most susceptible to degradation.
The consumption of long chain polyunsaturated fatty acids (LC-PUFA) is associated with several human health benefits. Most notable of these LC-PUFA is docosahexaenoic acid (DHA C22:6) whose inclusion is considered essential for optimum human health. Biofortification of common foods such as eggs with DHA has emerged as a specific approach to increase the intake of DHA in human populations. This can be achieved by supplementing poultry rations with feeds like microalgae or fish oil that are rich in DHA, which results in an increased uptake in the egg. Gas chromatography with flame ionization detection (GC-FID) is the method of choice when analyzing food such as eggs for DHA and other fatty acids. For regulatory studies it is desirable to demonstrate that the method is specifically suitable for the analysis of DHA and fatty acids in eggs. The purpose of this paper is to further extend the scope of the AOAC 996.06 methodology examined in the paper by Dillon et al., and to demonstrate the fitness for purpose of the method by examining specific validation parameters. It is a further objective to investigate the stability of DHA and other fatty acids of short and long timepoints. A validation of the method for the determination of DHA and three other fatty acids in eggs is thus presented.