Dietary supplementation of di- and trimethylglycine to attenuate the effects of pulmonary hypertension syndrome (ascites) in broilers

Abstract

Both N, N-dimethylglycine (DMG) and trimethylglycine (Betaine; TMG) are intermediary metabolites in the pathway of choline-to-glycine metabolism. DMG possess non-enzymatic anti-oxidant properties and have been implicated in enhanced oxygen utilisation while TMG posses osmolytic, methyl-donating, lipolytic and also potentially anti-oxidant-like properties. Recently, DMG has been supplemented to broiler chicken diets as a means to alleviate pulmonary hypertension syndrome (PHS or ascites) in rapidly growing broiler chickens. The present trial aimed to evaluate the effects of these different methylamines on production and slaughter performance and ascites-relates traits in broilers, and to assess oxidative stress in broilers with ascites. A challenge study was conducted where broilers were subjected to ascites-inducing conditions (AIC) from embryonic stage until 40 days of age. These AIC included high (>38.8°C) eggshell temperatures during mid-late incubation (Embryonic Day 11 (ED11) to Embryonic Day 18 (ED18)) and cold temperatures during brooding which increases the bird?s relative oxygen requirements and basal metabolic rate, as a means to incite pulmonary hypertension syndrome (PHS). A total of 1,632-day-old separate sex (males vs. females) broiler chicks were distributed equally amongst 96 pens. Birds were allocated to treatments based on a factorial arrangement comprising of two sexes and four dietary treatment groups of which each group had 12 replicate pens with 17 birds each. The treatment diets comprised of the following: a negative control group (NC; basal diet which was low in supplemental methionine (0.16 % methionine) and no additional choline chloride; a betaine group (NC + Betaine, where 1.042 g/kg of pure betaine (100 %) was added to the basal diet); a DMG group (NC + DMG, where equal molar equivalents of DMG (1.380 g/kg) was added to the basal diet); a positive control (PC) group (basal diet which contained additional levels of DL-methionine (0.21 % methionine) and additional choline chloride (250mg/kg)). Both DMG and TMG were fed at 100 % methyl group donors, i.e. concentrations higher than recommended levels. The experiment was divided into three phases: a starter phase (1-14 d), a grower phase (15-28 d) and a finisher phase (29-40 d). All birds were fed basal diet consistinf of maize-soybean meal. Animal performance was determined at weekly intervals. At termination of the study (d40 & 41), slaughter performance traits were determined on two birds per replicate. All birds were monitored twice daily for overt signs of PHS and mortality were recorded. During both processing stage 1 (d 20 & 21) and 2 (d 40 & 41) of the experiment, the risk of heart failure associated with the abovementioned dietary treatments was tested followed by gross and biochemical investigation from randomly selected individuals to determine the progression of PHS. Overall, body weight (BW), body weight gain (BWG) and cumulative BWG (CumBWG) were consistently higher in birds fed dietary betaine in comparison with birds fed the negative control (basal) or DMG-supplemented diets, whereas dietary treatment appeared to have little to no effect on feed intake. Weekly and cumulative feed conversion ratios (cumFCR) were consistently lower (P <0.05) in the betaine group compared with both the NC and DMG-supplemented groups, but only during the starter period (up to 14 d of age). It was also demonstrated that the use of betaine significantly (P <0.05) improved carcass parameters such as dressing percentage, breast meat weight and breast meat yield in comparison with both the NC (basal) and DMG-supplemented diet. Dietary treatment had no effect on any of the abdominal fat parameters. Furthermore, there was a highly significant (P <0.05) response following dietary betaine supplementation in pre-slaughter live BW, carcass weight, leg portion- and wing weight but not yield, in contrast to birds fed DMG. CumBWG, cumulative feed intake (CumFI) and CumFCR were significantly better (P <0.0001) for the commercial Ross male broiler chickens in contrast to the females. The heavier male broilers had significantly higher pre-slaughter live body- and carcass weights (P <0.0001), lower abdominal fat content (P <0.0001), and higher carcass cut (leg portion and wing) yields (P <0.05) than the females at slaughter. The incidence of PHS was not significantly different in broilers fed either of the control diets as compared to the betaine-supplemented group although a 19 % lower mortality rate was observed (P >0.05) in the betaine-supplemented group compared to the NC treatment group. Broilers fed dietary betaine was less likely to succumb to heart failure due to PHS compared to birds fed dietary DMG (i.e. 33% lower mortality rate) (P <0.05). Male broilers showed a ~4 times higher (P <0.001) ascites mortality rate compared to their female counterparts, confirming male broiler chickens to be more prone to develop ascites due to their inherently faster growth rate. The beneficial effects of betaine supplementation were also evident by a lower concentration of total serum homocysteine concentration (an independent risk factor for cardiovascular disease) compared to the control and DMG-supplemented groups during both processing stages. The level of 4-hydroxy-2-nonenal (4-HNE), an indicator of lipid peroxidation, was significantly lower for both the betaine-supplemented and PC treatment groups compared to either the NC or DMG treatment groups during processing stage 1 (d 20 & 21) (P <0.05). No significance was observed between the different treatment diets fed at 40 and 41 days of age (P >0.05). TBARS, an additional indicator of lipid peroxidative damage, was significantly lower for the DMG-supplemented group at 40 and 41 days of age compared to birds that received either the betaine-supplemented or Control diets. A slightly elevated AMPK concentration following oxidative-induced stress was obtained in birds fed dietary betaine at 40 and 41 days of age compared to birds fed either the DMG-supplemented or either of the control diets; however this difference was not significant (P>0.05). In conclusion, current data from the present study demonstrates the beneficial effects of supplementing diets with betaine in contrast to its methyl derivative, DMG, and methyl-inadequate diets on production and slaughter performance and attenuating PHS in broilers. Oxidative stress contributed to the pathogenesis of PHS in broilers in this study and supplementation of betaine can, to some extent, prevent oxidative damage and alleviate cumulative mortalities in broilers that develop PHS.