ABSTRACT
Background and Aim: Various methods can detect foot-and-mouth disease (FMD) in cows, but they necessitate resources, time, costs, laboratory facilities, and specific clinical specimen submission, often leading to FMD virus (FMDV) diagnosis delays. The 2022 FMD outbreak in East Java, Indonesia, highlighted the need for an easy, inexpensive, rapid, and accurate detection approach. This study aims to devise a one-step reverse transcriptase loop-mediated isothermal amplification (RT-LAMP) technique and phylogenetic analysis to detect the serotype O FMDV outbreak in East Java. Materials and Methods: Swab samples were collected from the foot vesicles, nasal secretions, and saliva of five suspected FMDV-infected cows in East Java between June and July 2022. The RT-LAMP design used hydroxy naphthol blue dye or SYBR Green I dye, with confirmatory analysis through reverse transcriptase polymerase chain reaction (RT-PCR) targeting 249 base pairs. PCR products underwent purification, sequencing, and nucleotide alignment, followed by phylogenetic analysis. Results: The RT-LAMP method using hydroxy naphthol blue dye displayed a positive reaction through a color shift from purple to blue in the tube. Naked-eye observation in standard light or ultraviolet (UV) light at 365 nm, with SYBR Green I stain, also revealed color change. Specifically, using SYBR Green I dye, UV light at 365 nm revealed a color shift from yellow to green, signifying a positive reaction. Nucleotide alignment revealed mutations and deletion at the 15th sequence in the JT-INDO-K3 isolate from the East Java FMDV outbreak. Despite differing branches, the phylogenetic tree placed it in the same cluster as serotype O FMDV from Malaysia and Mongolia. Conclusion: JT-INDO-K3 exhibited distinctions from Indonesian serotype O FMDV isolates and those documented in GenBank. Then, the RT-LAMP method used in this study has a detection limit 10 times higher latter than the conventional RT-PCR limit, without any cross-reactivity among strains.
ABSTRACT
Madura cattle are the germplasm of native cattle on the verge of extinction because of crossbreeding. Therefore, the present study was aimed to determine the serum nerve growth factor (NGF) concentration as a predictor of fresh ejaculate fertility parameters in Madura bull candidates. Eleven Madura bull candidates used for frozen semen production were selected for the study. Blood samples were collected using a vacutainer from the jugular vein for analyzing serum NGF and testosterone levels. Meanwhile, semen collection was conducted using an artificial vagina for sperm motility, viability, and concentration assessment. Data were analyzed to determine the correlation among variables and the linear regression of NGF concentration to other significant variables. The result showed that NGF had a significant correlation (p < 0.05) with testosterone levels, sperm motility, viability, and concentration. A significant correlation was observed between testosterone levels, sperm concentration, and sperm viability. The regression equation among significantly correlated variables was determined. For artificial insemination, suitable bull ejaculates for obtaining frozen semen should reach at least 2.12 ng/mL of NGF levels, with sperm viability, sperm concentration, and testosterone levels of more than 78.63%, 1,462.177 million/mL ejaculate, and 25.67 ng/mL, respectively. This is the first study to identify NGF as a predictor of male fertility in bull candidates of Madura cattle. Therefore, NGF levels could be used as a marker of male fertility in Madura bull cattle candidates. Thus, based on the minimum NGF levels, the ejaculate of Madura bull candidate that meets the requirements for frozen semen production could predict fertility.
ABSTRACT
Background and Aim: The production of male calf beef cattle is an agricultural innovation needed to increase the farm's productivity as a provider of meat sources. This study aimed to determine the sex ratio of the offspring of cows inseminated with Y-bearing sperm enriched by Percoll density gradient centrifugation and swim-up, combined with delayed fixed-time artificial insemination (FTAI). Materials and Methods: Ejaculates of Simmental bulls were divided into four equal portions and grouped as T0 (control, non-sexed semen), T1 and T2 were sexed semen using Percoll density gradient centrifugation three and five levels, respectively, and T3 was sexed semen using swim-up. After the sex was sorted, the semen was diluted in a tris-egg yolk extender, packaged in French mini-straws containing 50 million live sperm cells, and frozen. Pre-sexed, post-sexed, and post-thawed spermatozoa were evaluated based on progressive motility, viability, intact plasma membrane, and abnormality. The post-thawed semen of T0 was artificially inseminated to recipient cows at 12 h after onset of estrus (not delayed FTAI). Meanwhile, the delayed FTAI was conducted 18-20 h after onset of estrus using the T0, the best of T1 and T2, and the T3 post-thawed semen. Results: The Percoll density gradient centrifugation reduced motility, viability, and intact plasma membrane but increased sperm abnormalities. Meanwhile, the swim-up process increased motility, viability, and intact plasma membrane of sperm cells but decreased sperm abnormalities. Post-thawed semen decreased motility, viability, and intact plasma membrane of sperm cells but increased sperm abnormalities. The sex ratio of the Simmental crossbred offspring was 96.08% and 100% in T1 and T3, respectively, compared to 48.25% and 67.39% in T0 not delayed and delayed FTAI, respectively. Conclusion: The Percoll density gradient centrifugation and swim-up methods are prospective for obtaining male offspring.
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BACKGROUND AND AIM: Persistent corpus luteum (PCL) causes anestrus in mares. This study aimed to determine the effect of intrauterine prostaglandin F2α (PGF2α) treatment on PCL of racing mares to restore fertility. MATERIALS AND METHODS: Twelve racing mares suspected with PCL were diagnosed using transrectal palpation and confirmed by serum progesterone (P4) concentration measurement. PGF2α was infused intrauterine, followed by serum collection at 24, 48, and 72 h after. Estrous symptoms were monitored, and mating was conducted on day 3 of estrus with an earlier injection of 8.4 µg gonadotropin-releasing hormone twice a day. Transrectal palpation was performed on days 21-30 to observe the corpus luteum. Pregnancy diagnosis was performed rectally on 40-45 days post-mating and confirmed using Doppler ultrasound scanning. RESULTS: Eleven of the 12 mares had PCL. There was a dramatic reduction in the P4 concentration following PGF2α treatment of mares with PCL. All mares exhibited estrus 2.6±0.55 days post-treatment with a P4 concentration of 0.12±0.12 ng/mL. Rectal palpation and P4 concentration on 21-30 days after estrous onset showed that all mares were ovulating. The evaluation of P4 concentration on days 40-45 post-mating showed that all mares were still in the luteal phase. However, the pregnancy rate was only 54.5% based on rectal palpation and Doppler ultrasound scanning. CONCLUSION: Treatment of PCL in racing mares with an intrauterine infusion of PGF2α restored the estrous cycle and induced ovulation and pregnancy.
ABSTRACT
BACKGROUND AND AIM: Kacang buck sperm is cryosensitive due to the seminal plasma of semen itself. Meanwhile, bull seminal plasma contains the insulin-like growth factor-1 (IGF-1) complex, which is cryoprotective. The addition of the crude protein of Simmental bull seminal plasma increased the quality of post-thawed semen of Kacang buck. The study was conducted to determine the effects of Simmental bull seminal plasma with IGF-1 on the fertility of post-thawed Kacang buck semen. MATERIALS AND METHODS: Buck semen was diluted in the following skim milk-egg yolk extender preparations: Without the addition of Simmental bull seminal plasma IGF-1 complex protein (T0); with the addition of 12-µg Simmental bull seminal plasma IGF-1 complex protein (T1); and with the addition of 24-µg Simmental bull seminal plasma IGF-1 complex protein (T2). The extended semen was packed in 0.25-mL straws and frozen. Post-thawed semen fertility was evaluated based on the following variables: Sperm motility, viability, intact plasma membrane (IPM), malondialdehyde (MDA) levels, capacitation status, and acrosome reaction. The difference in each variable among the groups was evaluated using analysis of variance, followed by Tukey's honestly significant difference test, at a 95% level of significance. Meanwhile, principal component analysis (PCA) was used to identify the principal component of semen fertility among the seven parameters. RESULTS: The T1 group showed the highest sperm motility, viability, IPM, and percentage of incapacitated sperm and the lowest MDA levels, percentage of capacitated sperm, and acrosome reaction. PCA revealed that sperm motility had a moderate to very robust correlation with other variables and is the most crucial parameter, accounting for 80.79% of all variables. CONCLUSION: The IGF-1 complex in Simmental bull seminal plasma was useful for increasing the fertility of post-thawed Kacang buck semen, and sperm motility was the principal component of semen fertility.
ABSTRACT
BACKGROUND AND AIM: The genetic improvement of cattle through livestock section is based on quantitative, qualitative, and molecular characteristics. This study examined polymorphisms of the melanocortin-4 receptor (MC4R) and leptin genes as a reference for the selection of superior breeds in Madrasin cattle. MATERIALS AND METHODS: The leptin and MC4R genes of Madrasin cattle were amplified using polymerase chain reaction (PCR); then, restriction fragment length polymorphism of the leptin gene was performed using the restriction enzyme BsaA1, at site 2793 with ACGT point position. RESULTS: The leptin gene was divided into three bands, namely, AA with one fragment (522 bp), CG with two fragments (441 bp and 81 bp), and AG with three fragments (522 bp, 441 bp, and 81 bp). The MCR-4 gene was divided into three bands, namely, 493 bp, 318 bp, and 175 bp. CONCLUSION: The MC4R and leptin genes can act as molecular markers for growth traits in Madrasin cattle and can be used to genetically optimize and improve growth. The GG allele of the MC4R gene and the AA allele of the leptin gene can be used in Madrasin cattle.
