Changes in heterophil to lymphocyte ratios of heat-stressed chickens in response to dietary supplementation of several related stress agents

The heterophil to lymphocyte ratio has become widely accepted as a reliable and accurate physiological indicator of the stress response in chickens (Gross and Siegel, 1983), because exposure to stressors causes it to increase progressively. Gross and Siegel (1993) suggested that reference values for the heterophil to lymphocyte ratio of about 0.2, 0.5 and 0.8 are characteristic of low, optimal and high degrees of stress, respectively. Several feed compounds are related to stress levels in animals and some may be used to alleviate stress; between them are: yeast, ascorbic acid, tocopherol, tryptophan, niacin, and lactic acid. Usually, poultry diets contain a-tocopherol, niacin and sometimes ascorbic acid and definitely tocopherol and niacin. The brewer’s yeast (Saccharomyces cerevisiae) is known to reduce stress in animals (Phillips and Von Tungeln, 1984). In broiler chickens, the use of yeast reduced significantly the effects of aflatoxicosis (Stanley et al., 1993) and the frequency of Salmonella and Campylobacter colonization in birds subjected to transport stress (Line et al., 1997). Furthermore, Chukwu and Stanley (1997) found that the performance of laying hens during severe high temperature could be maintained with dietary inclusion of yeast. On the other hand, avian species are not able to synthesise sufficient ascorbic acid (vitamin C) to replace the severe losses of this vitamin during stress (Pardue and Thaxton, 1986). Gross (1988) suggested that ascorbic acid may reduce the effect of Escherichia coli and Amakye-Anim et al. (2000) suggested that supplementation of ascorbic acid has beneficial effects on antibody response to infectious bursal disease. Furthermore, Kutlu and Forbes (1993) found that ascorbic acid ameliorated heat-induced deterioration in performance and metabolism of broilers. Similarly, dietary needs of a-tocopherol (vitamin E) increase during stress, and supplementation of vitamin E resulted in partially preventing a decrease in egg production (Schiedeler, 1996; Bollengier-Lee et al., 1998, 1999) and alleviating heat stress in hens (Williams, 1997; Lewis et al., 1999; Smith, 2000). Furthermore, vitamin E supplementation enhances macrophage activity (Konjufca et al., 1999) and alleviated the impact of E. coli infection (Yang et al., 2000). Increased dietary tryptophan markedly reduces aggression in broiler breeders (Shea et al., 1990) and the incidence of pecking damage in growing bantams (Savory et al., 1999). Furthermore, Mench (1991) showed that increasing the dietary tryptophan content to levels found to be effective in reducing aggressive behaviour reduced plasma corticosterone levels in feed-restricted broiler breeders, and Newberry and Blair (1993) suggested that the addition of trytophan to the diet might have a mild fearreducing effect. Although Shea-Moore et al. (1996) showed that decreases in aggression in tryptophan-supplemented broiler breeders were not due to increases in blood niacin levels (this vitamin is synthesised from the tryptophan), elevations in niacin levels have been reported to control nervousness and hysteria in chickens (Schutz, 1965; Hansen, 1976). Finally, high levels of lactic acid (metabolite from glycolysis) were reported in blood of broilers that suffered from sudden death syndrome (Imaeda, 2000a). Lactic acid levels in blood are elevated when chickens are reared under high levels of stress caused by high stocking density, warm ambient temperature (Imaeda, 2000b) and saturated dietary fat content (Rotter et al., 1985). Froning et al. (1978) and Ngoka and Froning (1982) observed deterioration of breast meat colour of birds that were allowed to struggle before slaughter compared to anaesthetised birds. The darker breast meat colour is caused by depletion of muscle glycogen and less lactic acid build-up during post mortem glycolysis. On the contrary, supplementation of lactic acid in drinking water during pretransport may reduce Salmonella and Campylobacter contamination of broiler carcass at processing (Byrd et al., 2001). Only a few studies have examined the effects of these compounds on the heterophil to lymphocyte ratio. The effect of yeast on the heterophil to lymphocyte ratio has been reported by Guo and Liu (1997), who indicated that the addition of yeast reduced significantly the heterophil to lymphocyte ratio in broilers under heat stress. A strong relationship between ascorbic acid and heterophil to lymphocyte ratio has been documented; ascorbic acid reduced significantly the heterophil to lymphocyte ratio in birds under heat, cold, sound, fasting, or management stress (Gross, 1992; Mckee and Harrison, 1995; Zulkifli et al., 2000). Vitamin E supplementation improved the het-