During heat stress, broilers are subjected to osmotic stress, which can cause water imbalances and osmotic changes in cells through dehydration. It was reported that betaine may help as a protector against osmolarity changes under heat stress and protect the cells and improve the body weight of broilers under high ambient temperature (Zhou et al., 2012; Liu et al., 2019). Previous studies also showed that heat acclimation during the incubation period can improve the adaptation of chickens to heat stress through epigenetic mechanisms without changing the DNA sequence (Xu et al., 2022; Yalçın et al., 2022). It was hypothesized that the combined effect of in ovo betaine and heat acclimation would result in better growth performance in broilers under cyclic heat stress.
Gholami et al. (2015) and El-Shater et al. (2024) obtained on effect of in ovo injection of betaine at different concentrations on hatching performance of chicks. However, in ovo betaine injection at high dose (100 mg of betaine/egg) led to reduce hatchability compared, with control (uninjected) (Kadam et al., 2013). On the other hand, betaine spraying of quail eggs and incubation temperature increased hatchability (Abuoghaba et al., 2016). In the present study, the hatchability of fertile eggs and embryo mortality were found to be similar among the groups in agreed with Gholami et al. (2015) and El‐Shater et al. (2024).
Tanimowo and Longe (2020) reported that 5 or 10 mg of betaine-hydrochloride injection on 18 d of incubation did not affect the weights of the residual yolk sac, heart, and liver of chicks. Hu et al. (2015) found no effect of in ovo betaine at 2.5 mg/egg on chick and liver weight. Similarly, in ovo betaine did not influence chick and organ weights in the present study. Conflicted results were found related to the effect of HA on the weights of chick, relative yolk sac, and organ in the literature. No effect (Yahav et al., 2004; Akşit et al., 2010) or heavier chick weight but lighter heart weight (Yalçın et al., 2008a; Vitorino Carvalho et al., 2020) was found depending on the genotype, duration of the acclimation period, and incubation temperature. In the present study, neither HA nor betaine injection affected chick weight, and gastrointestinal tract weight of day-old chicks at hatch. Besides, the embryos from the experimental groups utilized the egg yolk to the same extent.
The weight of peripheral immune organs such as the spleen and central immune organs such as the thymus and bursa could be indicators of the immune system. Heavier thymus and spleen weights of day-old chicks obtained by in ovo betaine and HA would indicate a better immune system. Moreover, the day-old chicks from HA + in ovo betaine had significantly higher IgG levels compared to the CI. This indicates that HA + in ovo betaine activated specific immune responses in chicks, therefore, these chicks may have a higher chance of survival. Several studies showed that heat stress had a negative effect on the immune system of birds (Mashaly, 2004; Lara and Rostagno, 2013). It was reported that the relative weights of the thymus, a primary lymphoid organ, were decreased in birds exposed to heat stress (Jahanian and Rasouli, 2015). Al-Zghoul et al. (2019) found that HA increased heat tolerance by enhancing the immunological response of chickens to heat stress. The results of the present study showed that the decrease in thymus weight due to cyclic heat stress would be enhanced by HA. This result suggested that HA would contribute to the adaptation of broiler chickens to stressful conditions.
Maternal dietary betaine supplementation and in ovo injection of betaine increased cholesterol accumulation in the liver of neonatal piglets and day-old chicks, respectively (Cai et al., 2014; Hu et al., 2015; Idriss et al., 2017). Cai et al. (2014) reported that this increase in the hepatic cholesterol accumulation by maternal betaine did not affect the serum cholesterol level of piglets. Abobaker et al. (2019) found that maternal betaine did not affect blood cholesterol but suppressed cholesterol trafficking genes in offspring. In our study, there was no effect of in ovo betaine injection on the blood cholesterol of either day-old chicks or broilers at 42 d. However, liver cholesterol accumulation or cholesterol traffic genes were not investigated in the present study.
The decrease in the body weight and feed intake in birds after CHS agreed with the previous studies (Yalçın et al., 2005, 2008b). This reduction was accompanied by a decrease in feed consumption. Similar to previous findings, the results of the present study showed that HA has a beneficial effect on body weight gain under CHS conditions, but the effect was not significant at optimum rearing temperatures (Yalçın et al., 2008b). In the present study, the body weights of HA birds at 28 and 35 d were similar for broilers reared under OPT and CHS conditions, while the body weight of CI chickens exposed to CHS decreased by 84 and 173 g at 28 and 35d, respectively. Moreover, HA improved immunity modulating thymus weight of chickens at CHS. The similar body weight of chickens from OPT and CHS at 42d was probably related to the adaptation of birds to cyclic heat stress.
The increased growth performance under heat stress conditions through dietary betaine supplementation could be related to better intestinal structure and gut villi integrity thus digestive function of birds (Eklund et al., 2005; Liu et al., 2019). Ghasemi and Nari (2020) reported that daily feed intake and weight gain were reduced in response to heat stress while dietary betaine ameliorated these negative effects. In the present study, under CHS conditions, in ovo chickens were heavier at days 28 and 35 than those non injected indicating positive effect of in ovo betain injection as a response to heat stress. Increased feed intake of in ovo or non-injected HA chickens resulted similar FCR to CI chicks. Moreover, total feed consumption and feed conversion from 21 to 42 and 0 to 42d were similar among the treatment groups. This result may also link to the genetic selection program of the strains, as Attia et al. (2018) showed that the growth performance of Ross-308 broilers was better than Cobb 500 under heat stress.
In conclusion, the results of the present study showed that HA and in ovo betaine positively affected thymus weight and IgG level of day-old chicks and enhanced broiler performance of birds under cyclic heat stress. The results revealed that was higher feed consumption of those birds compared to those from CI. Further studies would be necessary to gain a better understanding of the combined effect of in ovo and HA on broiler performance under chronic heat stress.