High ambient temperatures compromise performance and productivity by reducing feed intake and decreasing nutrient utilization, growth rate, feed efficiency and milk production which lead to economic losses in dairy farming. Environmental stress also leads to oxidative stress associated with a reduced antioxidant status in the animals in vivo, as reflected by increased oxidative damage and lowered plasma concentrations of antioxidant vitamins (e.g., vitamins E, A, and C) and minerals (e.g., Zn). Zinc has an important role in numerous biological processes in avian and mammalian species. For instance, Zn is an essential component of many enzymes, and it has both structural and catalytic functions in metalloenzymes. Furthermore, dietary Zn is required for normal immune function as well as proper skeletal development and maintenance. One of the most important functions of Zn is related to its antioxidant role and its participation in the antioxidant defense system. This work compiles past and present information about the role of Zn in heat-stressed dairy animal’s health.
The livestock sector is an integral component of agricultural production system and tool for socioeconomic development and it contributes 4.11% of total gross domestic product (GDP) and 27.25% of agricultural GDP (Jain et al., 2017). India ranks first in the milk production, with 163.7 million tones and per capita availability of 352 g/day (BAHS, 2017). Environmental factors, such as temperature and light, exert significant effects on the production, health and immunity of animals. Increase in earth atmospheric temperature by 0.8°C since 1880 (NASA, 2015) gives us a red signal about the maintenance of production level in animals. Heat stress during hot summer in tropical countries is a problem of great concern among farmers and livestock producers as it causes great economic loss in dairy farming. Spiers et al. (2004) reported that milk yield decreased by 0.41 kg/cow/day for each THI unit increase of above 69. Heat stress in dairy cows is caused by a combination of environmental factors (temperature, relative humidity, solar radiation and air movement). It affects the cellular physiology and systemic metabolism leading to reduction in dry matter intake (DMI) and milk production (Yousef, 1997; Collier et al., 2008). The lactating dairy cows have an increased sensitivity to heat stress compared with non lactating animals, due to milk production elevating metabolism (Purwanto et al., 1990). Moreover, because of the positive relationship between milk yield and heat production, higher yielding cows are more challenged by heat stress than lower yielding animals (Spiers et al., 2004). The upper critical temperature limit in cow may reach up to 25 to 26°C (West, 2003), whereas, comfort zone is between -0.5 to 20°C, however it varies between the breeds. The environmental temperature is not the only factor, relative humidity is also important to describe the cow’s comfort zone. The combined effect of temperature and humidity is quantified as Temperature Humidity Index (THI). The normal THI level of 72 is optimal to maintain the milk production of dairy animals (Prasad et al., 2012). Heat stress is one of the wide varieties of factors which causes oxidative stress in-vivo. Reactive oxygen species (ROS), the major culprits for causing oxidative stress, are constantly generated in vivo as an integral part of metabolism. ROS may cause oxidative stress when their level exceeds the threshold value. They trigger progressive destruction of polyunsaturated fatty acids (PUFA), ultimately leading to membrane destruction. Body employs anti-oxidants to quench these free radicals. The enzymatic antioxidants like superoxide dismutase (SOD), catalase and glutathione peroxidase (GPx) act by scavenging both intracellular and extracellular superoxide radical and preventing lipid peroxidation of plasma membrane. Non-enzymatic antioxidants include vitamins like vitamins C, A and E, minerals like zinc (Zn) and chromium (Cr), proteins like albumin, transferrin, glutathione (GSH) etc. The role of dietary supplements such as vitamins for alleviating the effect of heat stress has been reviewed extensively (Sahin and Kucuk, 2003b). However, effects of Zn on performance, oxidative stress markers, and immunity in heat-stressed animals remain rather limited. The objective of this article was to review studies dealing with the effect of Zn supplementation to heat-stressed animals with respect to its role in metabolic and antioxidant status, immune potency and performance.
The Effects of Heat Stress on Dairy Animals
Changes in physiological parameters give an immediate response to the climatic stress and consequently the level of discomfort to the animals. Rectal temperature (RT) is taken directly inside the body cavity which is typically slightly higher than skin temperature. Change in rectal temperature has been considered as an important measure of physiological status as well as ideal indicator for assessment of stress in animals (Johnson, 1980; West, 2003). Respiration rate is a sensitive indicator of heat stress and the respiration rate increases when environmental temperature increases. Silanikove (2000) reported that measuring respiration rate appears to be the most accessible and easiest approach for evaluating the degree of heat stress in farm animals (low: 40–60 breaths per min, medium high: 60 – 80, high: 80 – 120, and severe stress: above 150 breaths per minute in cattle). Upadhyay et al. (2009) reported that there is a significant (P<0.01) increase in pulse rate when the crossbred male calves were given treadmill exercise during hot dry season of the year. Ronchi et al. (2001) reported that dry matter intake decreased by 23% in heifers managed at 320C and 70% relative humidity. Holter et al. (1997) established a significant negative correlation between THI and DMI for cows and suggested that the effect of THI is probably mediated through the effects of increasing body temperature of cow. Heat stress affects the productive performance of dairy animals by reducing their dry matter intake (DMI), feed efficiency and milk yield (Gantner et al., 2011; Baumgard et al., 2012). Decreased DMI under heat stress indicates physiological adaptation of animal by decreasing nutrient metabolism and consequent heat increment. The optimum environmental temperature for lactation depends on species, breed and degree of tolerance to heat or cold. The milk yield of Holstein cattle declines at temperature above 21°C, in case of Brown Swiss and Jersey cattle it declines at about 24 to 27°C whereas milk yield of Zebu cattle declines only above 34°C (Hafez, 1968). Therefore high yielding crossbred animals maintained in the tropical countries seems to be under threat of heat stress. Amaral et al. (2009) suggested that milk production will be reduced whenever THI exceeds a value of 72. Kadzere et al. (2002) observed that at the temperature of 35°C and 40°C there is reduction of milk production by 33% and 50% respectively. The average milk production of Karan Fries and Sahiwal animals at around THI of 72 was 13.4 and 6.6 litre/day (Singh and Upadhyay, 2009). Other than milking period, even cows under dry period experiencing heat stressed has profound effects on milk production in the subsequent lactation in ruminants (Bernabucci et al., 2010). Plasma cortisol level considered as an indicator of heat stress in animals. Increased plasma cortisol indicates the higher heat stress (Patel et al., 2017). Marai and Habeeb, 2010 observed that cortisol values were 9.07 and 12.53 ng/ml during cold and hot season respectively. Similarly other studies also reported the physiological increase in serum cortisol under heat stress conditions (Abilay et al., 1975; Wise et al., 1988; Dhami et al., 2006). Thyroid hormones T3 and T4 were decreased from 4.55 ± 0.16 pmol/L and 21 ± 0.08 pmoil/L to 3.21 ± 0.08 and 16.70 ± 0.19 pmol/L respectively in goats under heat stress conditions (Sivakumar et al., 2010). Decreased thyroid hormone level seems to be logical as an adaptive mechanism to decrease the metabolic heat production under the stressed conditions (Mader et al., 2010). As a consequence of oxidative metabolism and decreased physiological antioxidants levels results in oxidative stress. Kumar et al. (2007) demonstrated the increase in indicator of oxidative stress thiobarbituric acid reactive substances (TBARS) concentration in cattle and buffaloes subjected to thermal stress. Reproductive status of the animals experiences the negative impact of heat stress by modified hormonal and reproductive characteristics. Acute heat stress cows experienced changes in circulating concentrations of progesterone, luteinizing hormone (LH), follicle-stimulating hormone (FSH) and estradiol (Bernabucci et al., 2010), which affect estrous cycle length, estrus expression, and conception rates.
Zn is a very important trace element, involved in wide range of metabolic activities and productive performance like growth (Underwood et al., 1999), reproduction and immune system (Abdel et al., 2011: Patel et al., 2017) were influenced. Zn involved in structure and function of many enzymes associated with the carbohydrate, protein metabolism, nucleic acid synthesis, carbon dioxide transport and many other reactions (Pond et al., 1995). Zn is widely distributed throughout the body as a component of metallo-enzymes and metallo-proteins. Zn finger proteins play an integral role in regulating gene expression, consequently impacting a wide variety of body functions including cell division, growth, hormone production, metabolism, appetite control and immune function (Vallee and Falchuk, 1993). Zn deficiency increases oxidative damage of cell membranes caused by free radicals (Oteiza et al., 1996; Salgueiro et al., 2000; Prasad and Kucuk, 2002). Zn plays a critical role in anti-oxidant defense as an integral part of the essential enzymes superoxide dismutase (SOD) (Underwood, 1999). SOD is a major scavenger of free radicals, present in the cytoplasm of many types of cells and in the extracellular space. It converts the superoxide free radical (O2-2) to hydrogen peroxide, which is further decomposed by catalase into water and oxygen (Leung, 1998). The role of Zn as an anti-oxidant has been predicted on its ability to be an intermolecular stabilizer, preventing the formation of disulfide bonds and either displacing or competing with cupric or ferric ions, which trigger the formation of free radicals. Zn increases the synthesis of metallothionein, a cystine-rich protein that acts as free radical scavenger (Oteiza et al., 1996). Zn deficiency has been reported to impair antioxidant functions and an increase in oxidative DNA damage (Jing et al., 2007). The major sources of Zn in the mineral supplements formulated for animal feeding are inorganic salts like Zinc sulphate (ZnSo4), Zn oxide (ZnO), Zn chloride (ZnCl2) etc. Because many natural feed ingredients are marginally Zn-deficient, this micronutrient is commonly supplemented to diets for livestock and poultry (Batal et al., 2001). However, recent research has shown that organic Zn sources may be more available to the chick than inorganic Zn sources (Kidd et al., 1996; Wedekind et al. (1992).
Zn Supplementation in Heat-Stressed Dairy Animals
Performance and Productivity
Zn deficiency in animals is characterized by decreased feed intake, growth, low circulating levels of growth hormone (GH) its receptor, insulin-like growth factor-I and GH binding protein (MacDonald, 2000). Jia et al. (2008) identified an increase in ADG and feed efficiency when 15-45 mg Zn/kg DM was supplemented to a basal diet containing 22 mg Zn/kg DM in Liaoning Cashmere goats. However, Mandal et al. (2007) and Salama et al. (2003) did not find any significant change in DMI of bulls and goats respectively. Anton (2013) reported that group of cows supplemented with ZnSo4 had higher milk production compared to that in the cows of the groups that have not received this mineral. This gain in milk yield was positively correlated with increase in the plasma zinc level. Cope et al. (2009) supplemented 60 ppm ZnO to lactating Holstein Friesen cows and observed increased milk yield and no changes in milk composition. However, Griffiths et al. (2007) reported an increase in milk production in cows fed a combination of organic sources of Zn, Mn, Cu and Co. Similarly, Habeeb et al. (2013) reported that supplemental Zn resulted in increased milk yield in goat. Nayeri et al. (2014) supplemented 75 ppm ZnSo4 to HF cow and reported an increase in milk yield. Chandra et al. (2013) and Patel et al. (2017) also reported significant increase in milk yield at 60 and 120ppm zinc respectively in dairy cows.
Somatic Cell Count (SCC)
Zn is also important for maintenance of proper immunity, udder health and reproduction in cows. Zn is involved in the catalytic, structural and regulatory processes of keratinization (Sordillo et al., 1997). Zn is important for synthesis of keratin which lines the inside of teat duct. Keratin lining helps to maintain a protective barrier on the surface epithelium of teat and keep out the microorganism that can cause mastitis (Boland, 2003).
Different breeds of cow’s viz. Friesian (40 ppm, Shakweer et al. 2010), Sahiwal (80 ppm, De et al., 2014) and in Karan Fries cow’s (80 and 120 ppm) supplemented with Zn significantly reduced the SCC in milk. Whereas, Cope et al. (2009) reported supplementation of 600 mg Zn/day as ZnO to lactating cows reduced milk SCC in treated groups. Indicating that fortification of diet with extra Zn in milking animals had added benefit on udder health and clean milk production. However, dose required for reduction in SCC is not specific. It varies between the breeds and it may also depend on the management conditions of the cows.
An increased free radical production lowers the concentrations of antioxidant vitamins and minerals such as E, C, A, and Zn in serum (Halliwell and Gutteridge, 1989; Sahin et al., 2005). Studies shown that concentrations of malondialdehyde (MDA), an indicator of lipid peroxidation, in serum decreases with dietary Zn picolinate supplementation in heat stressed animals. Similary, Nagalakshmi et al. (2012) supplemented zinc carbonate in female rats also found significantly (P<0.01) lower lipid peroxidation in supplemented groups as compared to control. Sahin and Kucuk (2003b) and Kucuk et al. (2003) reported that 30 or 60 mg/kg of Zn decreased serum and liver MDA levels in heat-stressed birds. Whereas, Patel et al. (2017) reported similar decreasing trend of MDA in postpartum heat stressed cows at 80 and 120 ppm of dietary Zn. Onderci et al. (2003) also reported that 30 mg/kg of ZnSO4 and 400 μg/kg of Cr supplementation decreased serum MDA concentrations and increased the concentrations of vitamins C, E, and A in cold-stressed laying hens. In addition to decreased MDA concentration Sahin et al. (2005) also reported linear increase in serum antioxidant vitamin C and E as dietary 30 or 60 mg/kg of ZnSO4 and Zn picolinate supplementation increased. Decreased MDA concentration in above said studies indicates the antioxidant role of Zn. Since Zn acts as a cofactor of the antioxidative enzyme Cu-Zn SOD speculated that increased scavenging of free radical and prevention of peroxidation. SOD also inhibits the NADPH-dependent lipid peroxidation (Prasad and Kucuk, 2002) and also via inhibiting glutathione depletion as well (Prasad, 1997). Due to the ability to replace Fe and Cu from binding sites, Zn can compete with these transition metals to bind to the cell membrane and decrease the production of free radicals and thus exert a direct antioxidant action (Kucuk, 2003). Zn also induces production of metallothionein, which is an effective scavenger for hydroxyl radical. It has been suggested that Zn-metallothionein complexes in the islet cells provide protection against immune-mediated free radical attack (Salgueiro et al., 2000; Prasad and Kucuk, 2002). Another mode of action proposed for Zn as an antioxidant is its interaction with vitamin E. During Zn deficiency, probably due to defective formation of chylomicrons in the enterocyte, absorption of lipid-soluble vitamins such as E and A is impaired. Thus, some of the oxidative damage in Zn-deficient animals may be linked to the impaired vitamin E status during Zn deficiency (Kim et al., 1998). Heat-stressed quail, shown that serum vitamin C, vitamin E, and Zn concentrations linearly increased, whereas MDA concentrations linearly decreased as dietary vitamin E (0, 250, and 500 mg/kg) and Zn picolinate (0, 30, and 60 mg/kg) supplementation increased (Sahin et al., 2006b). Heat shock proteins (HSP) are thought to play a role in cellular protection under high ambient temperature, with a proposed relationship between the development of thermo tolerance and HSP synthesis, especially HSP70 (Lindquist and Craig, 1988). A lower HSP70 expression was observed in animals receiving therapeutic levels of Zn picolinate (30 and 60 mg/kg) and in quails reared under TN conditions (Sahin et al., 2009).
Dang et al. (2013) supplemented 80 ppm ZnSO4 in Karan Fries cows and found reduced cortisol values under oxidative stress conditions in supplemented group compared to control (7.44 ± 0.62 vs. 9.95 ± 1.47). Patel et al. (2017) at 120ppm and Purwar et al. (2017) at 40 ppm along with the other feed additives in crossbred cows reported decreased serum cortisol level under heat stress conditions. Decreasing thyrotropin-releasing hormone (TRH) and target T3 andT4 levels under heat stress was considered to be due adaptive mechanism of animals for reducing heat production. However, Habeeb et al. (2013) and Patel et al. (2017) supplemented 30 ppm and 120 ppm respectively in goat and cows found increased T3 and T4 levels. It may be due to role of Zn in synthesis of T3 which in turn converted to T4 (Freake et al. 2001).
Numerous studies have evaluated the effects of heat stress on the immune responses in animals and chickens. The heterophil-to-lymphocyte ratio has been used as a sensitive indicator of heat stress, among chicken populations (Gross and Siegel, 1983; Mashaly et al., 2004). An increased heterophilto-lymphocyte ratio was observed under high ambient temperature, which indicates a relationship between heat stress and nonspecific immune reactive cells (McFarlane and Curtis, 1989; Mashaly et al., 2004). Zn is an important element for all aspects of immunity (Chandra and Dayton, 1982; Sherman, 1992) and is critical for the integrity of the cells involved in the immune response (Dardenne et al., 1985). Zn deficiency causes a decrease in cellular immunity (Prasad and Kucuk, 2002) and adversely affects thymus (Fraker et al., 1977), spleen (Luecke et al., 1978), and interleukin production (Dowd et al., 1986). Abnormal T-lymphocyte development is thought to be the primary consequence of Zn deficiency (Dardenne and Bach, 1993). Zinc deficiency causes an imbalance in functions of T helper-1 and T helper-2 cells (Shankar and Prasad, 1998). Heat stress condition mainly affects the epithelial barrier function, results in impairment in nutrient absorption and endotoxemia (Fernandez et al., 2014). Supplementing dietary Zn may aid in repair and maintenance of epithelial integrity in digestive tract.
The patterns of reproductive behavior of farm animals vary with the variation in climatic conditions. Pronounced variations observed in the signs of estrus, rates of conception and frequency of calving are generally attributed to climatic factors. Roth et al. (2001) reported that heat stress caused an immediate as well as a delayed effect on ovarian follicular growth in Holstein cows. However, Wolfenson et al. (1995) reported that there was no immediate effect of heat stress on the population of small, medium, and large follicles in Gir cows. Rabiee et al. (2010) reported that secretion of gonadotropin-releasing hormone from the hypothalamus and the gonadotropins, luteinizing hormone (LH) and follicle stimulating hormone (FSH) from the anterior pituitary gland affected by heat stress. Habeeb et al. (2013) supplemented zinc in diet of goat during transition period and found that days from weaning to estrous, duration of estrus and days of kidding interval were significantly lower in supplemented group compared to control. Kundu et al. (2014) evaluate the effect of dietary ZnO on post-partum reproductive performance of Teresa goat and found favorable result in terms of incidence of estrus, conception rate and kidding rate. Patel et al. (2017) also reported that supplementation of 80 and 120 ppm Zinc in feed during peripartum period to Karan fries cows is effective in improving post partum reproductive performance.
Heat stress in dairy animals results in compromised performance. This effect can be subsidized by supplementing dietary Zn. Based on few available research studies in animals under heat stress and Zn supplementation, it can be concluded that, expected increase in oxidative damage, decreased DMI, elevated cortisol, decreased thyroid hormones, reproductive performance and immunosuppression in heat stress can be revered by supplementing Zn. The role of dietary Zn supplementation for alleviating the effect of heat stress has been reviewed extensively in poultry, however, in animals it is scanty. Hence further studies are needed in animals to confirm the role of dietary Zn on heat stress alleviation.