Global warming is a significant threat to the sustainability and profitability of the dairy sector, not only in tropical or subtropical regions but also in temperate zones where extreme summer temperatures have become a new and challenging reality. Prolonged exposure of dairy cows to high temperatures compromises animal welfare, increases morbidity, and suppresses fertility, resulting in devastating economic losses for farmers. To counteract the deleterious effects of heat stress, cattl e employ various adaptive thermoregulatory mechanisms including molecular, endocrine, physiological, and behavioral responses. These adaptations involve the immediate secretion of heat shock proteins and cortisol, followed by a complex network of disrupted secretion of metabolic and reproductive hormones such as prolactin, ghrelin, ovarian steroid, and pituitary gonadotrophins. While the strategic heat stress mitigation measures can restore milk production through modifications of the microclimate and nutritional interventions, the summer fertility records remain at low levels compared to those of the thermoneutral periods of the year. This is because sustainment of high fertility is a multifaceted process that requires appropriate energy balance, undisrupted mode of various hormones secretion to sustain the maturation and fertilizing competence of the oocyte, the normal development of the early embryo and unhampered maternal—embryo crosstalk. In this review, we summarize the major molecular and endocrine responses to elevated temperatures in dairy cows, as well as the impacts on maturing oocytes and early embryos, and discuss the consequences that heat stress brings about in dairy cattle fertility.

Heat shock protein 70 (HSP70) is a chaperon that stabilizes unfolded or partially folded proteins, preventing inappropriate inter- and intramolecular interactions. Here, we examined the developmental competence of in vitro matured oocytes exposed to heat stress with or without HSP70. Bovine oocytes were matured for 24 h at 39 °C without (group C39) or with HSP70 (group H39) and at 41 °C for the first 6 h, followed by 16 h at 39 °C with (group H41) or without HSP70 (group C41). After insemination, zygotes were cultured for 9 days at 39 °C. Cleavage and embryo yield were assessed 48 h post insemination and on days 7, 8, 9, respectively. Gene expression was assessed by RT-PCR in oocytes, cumulus cells and blastocysts. In C41, blastocysts formation rate was lower than in C39 and on day 9 it was lower than in H41. In oocytes, HSP70 enhanced the expression of three HSP genes regardless of incubation temperature. HSP70 at 39 °C led to tight coordination of gene expression in oocytes and blastocysts, but not in cumulus cells. Our results imply that HSP70, by preventing apoptosis, supporting signal transduction, and increasing antioxidant protection of the embryo, protects heat stressed maturing bovine oocyte and restores its developmental competence.

The aims of the present study were to examine the effects of HSP70 addition in the in vitro culture medium of day 3 embryos on their developmental competence and quality. Bovine oocytes (n = 1442) were in vitro matured, inseminated and cultured for the first two days according to standardized methods. The presumptive zygotes were randomly allocated in three experimental groups: Control, C (embryos cultured at 39 °C throughout the culture period), group C41 (temperature was raised to 41 °C from the 48th to 72nd h post insemination (p.i.) and then it returned at 39 °C for the remaining culture period), and group H41 (the temperature modification was the same as in C41 and during heat exposure, HSP70 was added in the culture medium). Cleavage and embryo yield were assessed 48 h p.i. and on days 7, 8, 9, respectively and gene expression in day 7 blastocysts was assessed by RT-PCR. Blastocyst yield was the highest in group C39; and higher in group H41 compared to group C41. From the gene expression analyses, altered expression of 11 genes was detected among groups. The analysis of the orchestrated patterns of gene expression differed between groups. The results of this study confirm the devastating effects of heat stress on embryo development and provide evidence that HSP70 addition at the critical stages can partly counterbalance, without neutralizing, the negative effects of the heat insult on embryos, acting mainly through mechanisms related to energy deployment.

Flaxseed and lupin seed were offered as an alternative dietary approach in dairy cows, through the partial substitution of soybean meal. Milk production and fertility traits were investigated. A total of 330 animals were allocated into two groups, treated (n = 176) and control (n = 154). From each group, 30 animals were selected for hematological and cytological studies. The experimental feeding period lasted for 81 days (25 days prepartum and 56 days postpartum). The control ration (group C) contained corn, barley, soybean meal, rapeseed cake, corn silage and lucerne hay; whereas, in the treatment group (group T), 50% of the soybean meal was replaced by an equal mixture of flaxseed and lupins. The two rations were formulated to be isonitrogenous and isoenergetic. Milk samples were analyzed for chemical composition, somatic cell count (SCC) content and total colony forming units (CFU). Blood samples were collected, and serum was analyzed for non-esterified fatty acids (NEFA), acute phase proteins (haptoglobin and serum amyloid) and lipid oxidation indices, namely thiobarbituric-acid-reactive substances (TBARS) and catalase activity. To assess polymorphonuclear neutrophils (PMN) numbers, endometrial samples from each cow were collected on days 21 and 42. No difference was recorded between groups in milk yield (p > 0.05). In multiparous cows, NEFA (mMol/L) concentrations were significantly lower in group T than in group C on day 14 (p > 0.009) and on day 42 (p = 0.05), while no difference was detected in the group of primiparous cows. At all time points, serum TBARS and catalase values were similar in both groups (p > 0.05). Multiparous cows in group T expressed the first postpartum estrus and conceived earlier than cows in group C (p ≤ 0.05). Between days 21 to 42 postpartum, the PMN reduction rate was higher in group T animals (p ≤ 0.05). Acute phase protein levels were in general lower in group T animals, and at specific time points differed significantly from group C (p ≤ 0.05). It was concluded that the partial replacement of soybean meal by flaxseed and lupins had no negative effect on milk yield or milk composition, and improved cow fertility; which, along with the lower cost of flaxseed and lupins mixture, may increase milk production profitability.

The effect of partial substitution of soybean meal by equal quantities of flaxseed and lupins in diets of Holstein dairy cows and heifers was investigated. A total of 6 animals (30 multiparous and 30 primiparous) were allocated into two equal groups in a randomised block design and fed control (group CO) or modified (group FL) TMR diets from three weeks prior to calving until day 40 postpartum. The TMR of group CO contained corn, barley, soybean meal, rapeseed cake, corn silage, and Lucerne hay, whereas in group FL equal quantities of whole flaxseed and lupins were used to replace 50% of the soybean meal in the TMR. All animals were fed twice daily with a daily allowance of 24 kg dry matter intake per animal. Milking was carried out three times daily and milk yield was recorded during every milking. Milk samples were analysed for chemical composition and SCC content. White cheeses were manufactured from bulk milk of each group at industrial level. Bulk milk and white cheese were analysed for chemical composition and fatty acid profile; cheese was also assessed for its organoleptic properties. Results indicate that milk yield did not differ among groups. Lipid oxidation values were similar among the groups, for both milk and cheese. However, FL inclusion resulted in lower (p < 0.05) protein carbonyls and higher (p < 0.05) phenolic compounds in both milk and cheese samples. Milk from the FL group had decreased palmitic (p < 0.05) and myristic (p < 0.05) and increased oleic (p < 0.05) and linolenic acid (p < 0.05) when compared to group CO. White cheese from group FL showed a decrease in saturated fatty acids (SFA) (p < 0.05), an increase in monounsaturated fatty acids (MUFA) (p < 0.05), and a higher increase in polyunsaturated fatty acids (PUFA) (p < 0.05) when compared with that of group CO. The white cheese of cows fed diets with flaxseed and lupins showed compositional and organoleptic properties quite similar to control group cheese; aroma, texture, and color were acceptable and desirable in both cheeses. However, increased levels of n-3 polyunsaturated fatty acids were found in the cheese of FL fed animals. The substitution of soybean meal by flaxseed and lupins in diets of Holstein cows warrants further investigation, especially towards the production of cheese that meet the consumers’ demand for novel and healthier dairy products.

Olive cake was incorporated at a low inclusion rate (3.7%) into the rations of dairy cows through partial substitution of maize, and its effects on milk production, general health, and fertility traits were investigated. Multiparous purebred Holstein dairy cows (n = 148) were divided into two groups: a treated group (n = 86) and a control group (n = 62). The control ration (group C) was modified by replacing 1 kg of maize with an equal quantity of olive cake (group T). The experiment lasted from 60 days prepartum to 150 days postpartum. Electronic aids were utilized to quantify daily milk yield and detect estrus expression, while clinical and uterine examinations were performed weekly. Blood samples, uterine epithelial cells, and endometrial tissue samples were collected from a subgroup of healthy animals in both groups at specific time points. Blood samples were examined to determine the β-hydroxybutyric acid (BHBA), acute phase proteins (haptoglobin [Hpt] and serum amyloid A [SAA]), progesterone, and pregnancy-associated glycoproteins concentrations. Uterine epithelial cells were assessed for polymorphonuclear neutrophil (PMN) counts, and the expression of nine genes encoding inflammatory cytokines and immune system activation was analyzed in uterine biopsy tissue. No significant differences (p > 0.05) were observed between groups in milk yield, general morbidity, clinical endometritis, or conception rates. However, animals in group T came to estrus approximately 6 days earlier (p = 0.013) than those in group C; progesterone concentrations on day 7 of the subsequent cycle tended (p = 0.07) to be higher in group T. On day 21, BHBA concentrations were higher in group C than in group T (p < 0.05). Throughout the experiment, Hpt levels in group T were consistently lower (p < 0.001), while SAA was lower on day 7 compared to group C. From days 21 to 42 postpartum, there was a significant reduction in PMN numbers (group C p = 0.02; group T p < 0.0001), with a tendency for a greater reduction rate in group T (p = 0.08). Among the genes studied, a significant difference was revealed in the expression of the ILA1 gene, with strong correlations of gene expression in group C and weak to moderate correlations in group T. In conclusion, under the conditions of this experiment, the inclusion of olive cake into dairy cows’ rations did not affect milk production but improved certain health and fertility parameters, making olive cake a suitable alternative feedstuff for high-producing animals.