Thesis Abstract

Abstract
The aim of this study was to investigate the responses of broiler chickens fed with betaine hydrochloride supplementation under dexamethasone-induced stress conditions. Stress is a significant factor affecting poultry production and welfare. Betaine, a natural and widely used feed additive, has been reported to have potential stress-relieving effects in various animal species. In this experiment, a total of 200 broiler chickens were randomly divided into four treatment groups control group (CON), stress group (ST), stress group supplemented with 500 mg/kg betaine hydrochloride (ST+B500), and stress group supplemented with 1000 mg/kg betaine hydrochloride (ST+B1000). The stress was induced by administering dexamethasone at 2 mg/kg body weight for five consecutive days. The results indicated that dexamethasone administration significantly increased the serum corticosterone levels and decreased the serum total protein levels in the ST group compared to the CON group. However, betaine hydrochloride supplementation at both levels (500 mg/kg and 1000 mg/kg) significantly attenuated the dexamethasone-induced elevation of corticosterone levels and restored the total protein levels in the serum. Furthermore, the ST group exhibited reduced growth performance parameters such as body weight gain and feed intake compared to the CON group, whereas betaine supplementation improved these parameters in a dose-dependent manner. Histopathological examination of the liver tissues revealed that dexamethasone induced hepatic damage characterized by vacuolization and inflammatory cell infiltration in the ST group, while betaine supplementation mitigated these pathological changes. Additionally, the oxidative stress markers such as malondialdehyde (MDA) levels were elevated in the liver tissues of the ST group, which were ameliorated by betaine supplementation. Moreover, betaine supplementation enhanced the activities of antioxidant enzymes including superoxide dismutase (SOD) and catalase (CAT) in the liver tissues under stress conditions. In conclusion, betaine hydrochloride supplementation demonstrated protective effects against dexamethasone-induced stress in broiler chickens by regulating the stress-related hormones, improving growth performance, preserving liver histology, and enhancing antioxidant defense mechanisms. These findings suggest that betaine supplementation could be a potential strategy to alleviate stress and improve the welfare and productivity of broiler chickens in stressful production environments.

Thesis Overview

It is estimated that world food consumption will double by 2050 as more developing nations improve their economic status and per capital meat consumption increase (Godfray et al., 2010). This places an increasing pressure on animal producers especially poultry farmers to maximize output levels of their animals to meet this growing demand. Physiological stress is one of many concerns facing the modern broiler producer. The term stress is very familiar to most researchers, but there is no universal definition for stress. When a stressor is actually causing a negative impact on the well-being of an animal, this can be defined as distress (Moberg, 2000). Another broader definition states that stress is any biological response elicited when an animal perceives a threat to its homeostasis (Moberg, 2000). Extremes of ambient temperature is an important stressor that confronts poultry in many regions of the world and large economic losses occur because of mortality and decreased production (Altan et al., 2000).

The thermoneutral zone for poultry is 18°C–24°C in the tropics and 12°C–26°C in the temperate zones, but this often gets exceeded in the tropics, resulting in heat stress (Holik, 2009; Dei and Bumbie, 2011). When the hypothalamic-pituitary-adrenocortical axis is activated, the hypothalamus produces corticotrophin-releasing factor, which in turn stimulates the pituitary to release adrenocorticotropic hormone (ACTH) (Mormède et al., 2007). Secretion of ACTH causes the cells of the adrenal cortical tissue to proliferate and to secrete corticosteroids. The main active hormone of the axis is cortisol in cattle, sheep, pig, mink, fox and fish, and corticosterone (CS) in birds and rodents (Mormède et al., 2007). These are cholesterol-derived steroids synthesized in the fascicular zone of the adrenal cortex under the control of the pituitary hormone. ACTH is synthesized by specialized cells (corticotrophs) of the anterior pituitary gland (Mormède et al., 2007).The production of glucocorticoids is increased by stress; therefore, corticosterone can be used as a biomarker of stress in poultry.

In chickens, adrenal corticosteroids are secreted shortly after exposure to stress and elevated levels of plasma glucocorticoids have been used as an index of the response to stress in poultry (Siegel, 1995). By elevating circulatory corticosteroids and decreasing thyroid activity, heat stress impairs broiler performance, especially adult birds, because the ability to dissipate heat decreases with age (Mahmoud et al., 2014). Drastic decline in feed intake occurring in heat-stressed birds is a physiological response to minimize intrinsic heat production. It is aimed at maintaining thermal homeostasis, thus decreasing feed efficiency, live weight gain, and survival rates (Faria-Filho et al., 2007). Lower breast-meat yield and higher carcass-fat deposition are the other deleterious effects of heat stress that lower the economic value of broiler carcasses (Geraert et al., 1996; Ain-Baziz et al., 1996). Corticosterone has an indirect role in lipid metabolism bycausing the rate of fat deposition to increase in poultry (Jiang et al., 2008; Yuan et al., 2008). Glucocorticoid hormone secretion also has implications for mineral metabolism, thus corticosteroids have been directly implicated in the development of osteoporosis in stressed animals (Siegel and Latimer, 1970). It has also been shown that glucocorticoids support the action of catecholamines, which have been shown to increase urinary calcium and sodium excretion (Fink and Brody, 1978).

Corticosteroids have been shown to inhibit several immune system functions in various species, as demonstrated in depressed number of circulating lymphocytes, which results in an increase in the ratio of circulating heterophils to lymphocytes, the most recognizable symptom of stress in poultry (Siegel, 1995). Regression of the thymus, bursa, and spleen has also been demonstrated in chickens after corticosterone or ACTH administration (Puvadolpirod and Thaxton, 2000a).

Similarly, synthetic glucocorticoid dexamethasone administration mimics the adverse effects of increased corticosterone. Dexamethasone (doses ranging from 0.2 to 4.0 mg/kg) has been used as an immune suppressive agent (Fowles et al., 1993), mediator of prenatal stress (Maccari et al., 2003) and to induce oxidative stress in laying hens (El-Habbak et al. 2005) and cockerels (Eid et al., 2006). Aengwanich (2007) demonstrated that synthetic glucocorticoid; dexamethasone indoses of up to 6 mg/kg in their diets had many effects on broilers like internal glucocorticoid.

Certain feed additives (selenium, vitamins C and E, α-lipoic acid, α-tocopherol, prebiotics and probiotics) enhance performance in heat-stressed broilers (Ghazi Harsini et al., 2012; Hamano, 2012; Imik et al., 2012; Khan et al., 2012; Sandhu et al., 2012; Sohail et al., 2012). Betaine, a methyl group donor, functions in lipid metabolism by stimulating oxidative catabolism of fatty acids through carnitine synthesis. Betaine is also an organic osmolyte and does not interfere with enzyme function or upset metabolism (Simon, 1999). As an osmolyte, betaine may have a stabilizing function on cells subjected to osmotic stressors, such as in the case of coccidiosis infection (Klasing et al., 2002) by regulating the water balance, resulting in the stability of tissue metabolism especially in the gastrointestinal tract (Lipinski et al., 2012). Dietary supplementation of betaine presumably reduces the requirement for other methyl-group donors, such as methionine and choline (Siljander-Rasi et al., 2003). Florou-Paneri et al. (1997) showed that between 30 and 80 percent of supplemental methionine can be substituted by betaine without negative effects on performance. As a feed additive, betaine is most commonly added to animal diets as anhydrous betaine; betaine monohydrate, and betaine hydrochloride (Kidd et al., 1997; Eklund et al., 2005). Within the body, betaine is synthesized from choline (Sakomura et al., 2013).