Improved reproductive performance achieved in tropical dairy cows by dietary beta-carotene supplementation.

Dairy farming in tropical climates is challenging as heat stress can impair reproduction in cows. Previous studies have demonstrated the beneficial effects of beta-carotene supplementation on bovine reproductive performance. This study was performed in Thailand, where the temperature-humidity index (THI) during the experimental periods was measured to range from 78.4 to 86.1. Lactating Holstein cows classified as repeat breeders (previous artificial insemination [AI] failures) were randomly assigned into two treatments, control treatment (T1; received placebo, n = 200) and test treatment (T2; received 400 mg/h/day of beta-carotene, n = 200). All cows were subjected to a protocol for synchronization of ovulation and timed artificial insemination (TAI). The day of the 1st ovulation synchronized protocol was defined as day 0, and the total experimental period was 160 days. Daily placebo or beta-carotene supplements were given orally on day 0 and each subsequent day of the experiment. Diagnosis of pregnancy was performed using ultrasound on day 30 after insemination. Non-pregnant cows were subjected to further ovulation synchronizations (maximum of four) and TAI over a period of 160 days. Milk samples were collected every ten days throughout the experiment. The samples were analyzed for beta-carotene concentration, superoxide dismutase (SOD) and glutathione peroxidase (GPx) activities. The pregnancies per AI of the cows in T2 were significantly greater than that of T1 from the 2nd to 4th TAI. During the entire experimental period, the pregnancies in T2 were significantly greater than that of T1. Cox's proportional hazards regression model data indicated a 44% greater probability of pregnancy for cows receiving beta-carotene. The concentrations of milk beta-carotene in T2 were significantly greater than T1 from the 2nd to 4th TAI. Significantly greater SOD and GPx activities were observed in T2 than T1, suggesting a reduction of oxidative stress in cows treated with beta-carotene. Dietary supplementation with beta-carotene thus improves the reproductive performance of repeat breeders exposed to heat stress, possibly by reducing oxidative stress.

Comparison of the Effects of Microbial Inoculants on Fermentation Quality and Microbiota in Napier Grass (Pennisetum purpureum) and Corn (Zea mays L.) Silage.

Forage preservation for livestock feeding is usually done by drying the plant material and storing it as hay or ensiling it into silage. During the ensiling process, the pH in the system is lowered by the activities of lactic acid-producing bacteria (LAB), inhibiting the growth of spoilage microorganisms and maintaining the quality of the ensiled product. To improve this process, inoculation of LAB could be used as starter cultures to shorten the ensiling time and control the fermentation process. Here, we compared fermentation quality and bacterial dynamics in two plant materials, whole-plant corn (Zea mays L.) and Napier grass (Pennisetum purpureum), with and without starter inoculation. The efficacy of Lactobacillus plantarum, L. brevis, and Pediococcus pentosaceus as starter cultures were also compared in the ensiling system. In whole-plant corn, pH decreased significantly, while lactic acid content increased significantly on Day 3 in both the non-inoculated and LAB-inoculated groups. Prior to ensiling, the predominant LAB bacteria were Weissella, Enterococcus, and Lactococcus, which shifted to Lactobacillus during ensiling of whole-plant corn in both the non-inoculated and LAB inoculated groups. Interestingly, the epiphytic LAB associated with Napier grass were much lower than those of whole-plant corn before ensiling. Consequently, the fermentation quality of Napier grass was improved by the addition of LAB inoculants, especially L. plantarum and a combination of all three selected LAB strains showed better fermentation quality than the non-inoculated control. Therefore, the different abundance and diversity of epiphytic LAB in plant raw materials could be one of the most important factors determining whether LAB starter cultures would be necessary for silage fermentation.

Comparison of Cryotop and micro volume air cooling methods for cryopreservation of bovine matured oocytes and blastocysts.

This study was designed to compare the efficiency of the Cryotop method and that of two methods that employ a micro volume air cooling (MVAC) device by analyzing the survival and development of bovine oocytes and blastocysts vitrified using each method. In experiment I, in vitro-matured (IVM) oocytes were vitrified using an MVAC device without direct contact with liquid nitrogen (LN2; MVAC group) or directly plunged into LN2 (MVAC in LN2 group). A third group of IVM oocytes was vitrified using a Cryotop device (Cryotop group). After warming, vitrified oocytes were fertilized in vitro. There were no significant differences in cleavage and blastocyst formation rates among the three vitrified groups, with the rates ranging from 53.1% to 56.6% and 20.0% to 25.5%, respectively; however, the rates were significantly lower (P < 0.05) than those of the fresh control group (89.3% and 43.3%, respectively) and the solution control group (87.3% and 42.0%, respectively). In experiment II, in vitro-produced (IVP) expanded blastocysts were vitrified using the MVAC, MVAC in LN2 and Cryotop methods, warmed and cultured for survival analysis and then compared with the solution control group. The rate of development of vitrified-warmed expanded blastocysts to the hatched blastocyst stage after 24 h of culture was lower in the MVAC in LN2 group than in the solution control group; however, after 48-72 h of culture, the rates did not significantly differ between the groups. These results indicate that the MVAC method without direct LN2 contact is as effective as the standard Cryotop method for vitrification of bovine IVM oocytes and IVP expanded blastocysts.

Bovine embryo sex determination by multiplex loop-mediated isothermal amplification.

In cattle, the ability to determine the sex of embryos before embryo transfer is beneficial for increasing the number of animals with the desired sex. This study therefore developed a new modification of loop-mediated isothermal amplification in a multiplex format (multiplex LAMP) for highly efficient bovine embryo sexing. Two chromosomal regions, one specific for males (Y chromosome, S4 region) and the other common to both males and females (1.715 satellite DNA), were amplified in the same reaction tube. Each target was amplified by specifically designed inner primers, outer primers, and loop primers, where one of the S4 loop primers was labeled with the fluorescent dye 6-carboxyl-X-rhodamine (emitting a red color), whereas both satellite loop primers were labeled with the fluorescent dye fluorescein isothiocyanate (emitting a green color). After amplification at 63 °C for 1 hour, the amplified products were precipitated by a small volume of cationic polymer predispensed inside the reaction tube cap. Green precipitate indicated the presence of only control DNA without the Y chromosome, whereas orange precipitate indicated the presence of both target DNAs, enabling interpretation as female and male, respectively. Accuracy of the multiplex LAMP assay was evaluated using 46 bovine embryos with known sex (25 male and 21 female) generated by somatic cell nuclear transfer and confirmed by multiplex polymerase chain reaction. The multiplex LAMP showed 100% accuracy in identifying the actual sex of the embryos and provides a fast, simple, and cost-effective tool for bovine embryo sexing.

Determination of Sperm Sex Ratio in Bovine Semen Using Multiplex Real-time Polymerase Chain Reaction.

Gender selection is important in livestock industries; for example, female calves are required in the dairy industry. Sex-sorted semen is commonly used for the production of calves of the desired gender. However, assessment of the sex ratio of the sorted semen is tedious and expensive. In this study, a rapid, cost effective and reliable method for determining the sex ratio was developed using a multiplex real-time polymerase chain reaction (PCR) assay. In this assay, the X and Y chromosome-specific markers, i.e., bovine proteolipid protein (PLP) gene and sex-determining region Y (SRY) were simultaneously quantified in a single tube. The multiplex real-time PCR assay was shown to have high amplification efficiencies (97% to 99%) comparable to the separated-tube simplex real-time PCR assay. The results obtained from both assays were not significantly different (p>0.05). The multiplex assay was validated using reference DNA of known X ratio (10%, 50%, and 90%) as templates. The measured %X in semen samples were the same within 95% confidence intervals as the expected values, i.e., >90% in X-sorted semen, <10% in Y-sorted semen and close to 50% in the unsorted semen. The multiplex real-time PCR assay as shown in this study can thus be used to assess purity of sex-sorted semen.

Changes in reproductive physiology of lactating dairy cows due to elevated steroid metabolism.

This manuscript focuses on potential changes in reproductive physiology that occur due to high milk production in lactating dairy cows. Four reproductive measures are discussed: interval to first ovulation, conception rate, duration of estrus, and multiple ovulation rate. The last two responses have now been closely linked to level of milk production. In contrast, time to first ovulation does not appear to be associated with level of milk production, and the association of conception rate with level of milk production is still controversial. In an attempt to explain some of the changes in reproductive physiology caused by high milk production a model of elevated steroid metabolism in lactating dairy cows is presented. Although many aspects of this model remain to be tested, a central role for elevated steroid metabolism in lactation-induced reproductive changes seems likely.