UNASSIGNED: The intracytoplasmic sperm injection (ICSI) technique has low efficiency in cattle. This has mainly been attributed to the oocyte activation failure due to oocyte and/or sperm factors.
UNASSIGNED: Our aim was to evaluate the effect of conventional ICSI and Piezo-ICSI with bull or human sperm on bovine oocyte activation and embryo development and to assess its relationship with the phospholipase C zeta (PLCɀ) activity of both species.
UNASSIGNED: In vitro matured bovine oocytes were randomly divided into five groups and were fertilized as follows: conventional ICSI using bovine sperm with chemical activation (control), conventional ICSI using bovine sperm, Piezo-ICSI using bovine sperm, conventional ICSI using human sperm, and Piezo-ICSI using human sperm. PLCɀ activity was determined in bull and human sperm samples.
UNASSIGNED: Within the groups using bull sperm, the oocytes fertilized by conventional ICSI had the lowest values of 2 pronuclei (PN) formation and cleavage, Piezo-ICSI increased both percentages and ICSI + chemical activation presented the highest 2 PN, cleavage, and blastocyst rates (p < 0.05). Within the groups using human sperm, the oocytes fertilized by Piezo-ICSI presented higher 2 PN and cleavage rates than those activated by conventional ICSI (p < 0.05). Piezo-ICSI with human sperm increased bovine oocyte activation as much as conventional ICSI + chemical activation with bovine sperm (p < 0.05). Higher values of PLCɀ activity were found in human sperm compared with bovine sperm (p < 0.05).
UNASSIGNED: Our results suggest that the higher stability of the bovine sperm in combination with its relatively low content of PLCɀ impairs bovine oocyte activation after ICSI.

The primary objective was to investigate the association between delayed milk ejection (DME) and the average milk flow rate, milking unit-on time, and duration in a low milk flow rate in Holstein dairy cows in a large dairy herd with suboptimal premilking teat stimulation. Our second objective was to study the association between peak lactation milk yield and the occurrence of DME. This longitudinal field study was conducted at a 4300-cow dairy farm with a thrice-daily milking schedule over a 1-week period. We analyzed data from 61,677 cow milking observations from 2937 cows. Delayed milk ejection was defined as present if the 30-60 s milk flow rate was ≤3.1 kg/min. The mean average milk flow rate (MAMF, kg/min), mean milking unit-on time (MMUT, s), and mean duration of a low milk flow rate (MLMF, s) were calculated as the mean values from the 21 milking observations. General linear multivariable models revealed associations of DME with MAMF, MMUT, and MLMF. A multivariable ordinal logistic regression model revealed an association between peak lactation milk yield and DME. Cows with lower peak lactation milk yield had greater odds of exhibiting a higher frequency level of DME. The observed associations between DME and milking performance indices suggest that DME can negatively affect milking and parlor efficiency. Peak lactation milk yield may serve as a proxy to estimate cows\' risk of recurrent DME. Future research is warranted to test if alleviating DME through, for example, a modified milking routine influences the milking performance indices described herein.

Ischemic teat necrosis (ITN) is a growing problem in the dairy industry characterized by teat lesions, necrosis, pruritus and automutilation. Despite the economic and welfare consequences, there is no treatment, and the etiology of the disease remains poorly understood. The aim of this study was to investigate ITN by analyzing its clinical presentation, potential risk factors and microbial involvement. Methods included collection of milk and swab samples from affected cows over a period of one-and-a-half years and completion of questionnaires by veterinarians and farmers. Microbial testing included PCR testing for Treponema spp. and cultural testing by anaerobic and aerobic incubation on blood agar. The results showed a high and significant prevalence of Treponema spp. and Staphylococcus aureus in affected teats compared to non-ITN-affected control teats, indicating their potential role in the development of ITN. Other factors such as edema and milking practices also appear to contribute to the tissue damage. First-lactation and early-lactation heifers are particularly at risk. In conclusion, ITN appears to have a multifactorial etiology with both infectious and non-infectious factors playing a role. Further research is needed to better understand the complex interplay of these factors and to develop effective prevention and management strategies.

This study aimed to enhance our understanding of the agreement between two sampling methods for the detection of bovine respiratory disease (BRD) pathogens in calves using high-throughput real-time qPCR (ht-RT-qPCR). In total, 233 paired nasal swab (NS) and non-endoscopic bronchoalveolar lavage (nBAL) samples were collected from 152 calves from 12 Danish cattle herds. In 202 of the observations, the calves were examined using a standardized clinical protocol. Samples were tested for three viruses (bovine respiratory syncytial virus, bovine corona virus, and influenza D virus) and six bacteria (Histophilus somni, Mannheimia haemolytica, Mycoplasma bovis, Mycoplasma species, Pasteurella multocida, and Truepurella pyogenes). The results showed age-related differences in disease and pathogen occurrence, with the highest detection rates in calves aged 35 days or older. Poor to moderate agreement was found between the NS and nBAL results. The presence of Mannheimia haemolytica in both NS and nBAL in younger calves and in nBAL in older calves was associated with clinical BRD. There was a potential link between BRD and influenza D virus in older calves, although it was only found in one herd in a small sample size. Overall, NS was a relatively poor predictor of pathogens in the lower respiratory tract. The present study confirms the complexity of pathogen detection in BRD, with marked influences of age and the sampling method on pathogen detection and disease associations.

BACKGROUND: Acinetobacter lwoffii (A. lwoffii) is a Gram-negative bacteria common in the environment, and it is the normal flora in human respiratory and digestive tracts. The bacteria is a zoonotic and opportunistic pathogen that causes various infections, including nosocomial infections. The aim of this study was to identify A. lwoffii strains isolated from bovine milk with subclinical mastitis in China and get a better understanding of its antimicrobial susceptibility and resistance profile. This is the first study to analyze the drug resistance spectrum and corresponding mechanisms of A. lwoffii isolated in raw milk.
RESULTS: Four A. lwoffii strains were isolated by PCR method. Genetic evolution analysis using the neighbor-joining method showed that the four strains had a high homology with Acinetobacter lwoffii. The strains were resistant to several antibiotics and carried 17 drug-resistance genes across them. Specifically, among 23 antibiotics, the strains were completely susceptible to 6 antibiotics, including doxycycline, erythromycin, polymyxin, clindamycin, imipenem, and meropenem. In addition, the strains showed variable resistance patterns. A total of 17 resistance genes, including plasmid-mediated resistance genes, were detected across the four strains. These genes mediated resistance to 5 classes of antimicrobials, including beta-lactam, aminoglycosides, fluoroquinolones, tetracycline, sulfonamides, and chloramphenicol.
CONCLUSIONS: These findings indicated that multi-drug resistant Acinetobacter lwoffii strains exist in raw milk of bovine with subclinical mastitis. Acinetobacter lwoffii are widespread in natural environmental samples, including water, soil, bathtub, soap box, skin, pharynx, conjunctiva, saliva, gastrointestinal tract, and vaginal secretions. The strains carry resistance genes in mobile genetic elements to enhance the spread of these genes. Therefore, more attention should be paid to epidemiological surveillance and drug resistant A. lwoffii.

Global methylation levels differ in in vitro- and in vivo-developed embryos. Follicular fluid (FF) contains extracellular vesicles (EVs) containing miRNAs that affect embryonic development. Here, we examined our hypothesis that components in FF affect global DNA methylation and embryonic development. Oocytes and FF were collected from bovine ovaries. Treatment of zygotes with a low concentration of FF induced global DNA demethylation, improved embryonic development, and reduced DNMT1/3A levels. We show that embryos take up EVs containing labeled miRNA secreted from granulosa cells and the treatment of zygotes with EVs derived from FF reduces global DNA methylation in embryos. Furthermore, the methylation levels of in vitro-developed blastocysts were higher than those of in their vivo counterparts. Based on small RNA-sequencing and in silico analysis, we predicted miR-29b, -199a-3p, and -148a to target DNMTs and to induce DNA demethylation, thereby improving embryonic development. Moreover, among FF from 30 cows, FF with a high content of these miRNAs demethylated more DNA in the embryos than FF with a lower miRNA content. Thus, miRNAs in FF play a role in early embryonic development.

The majority of species previously categorized as Bacteroides have been reassigned into new genera. Bacteroides levii (Holdeman, Cato, and Mooretaxonomic)\'s status has remained uncertain. This species shares a high degree of similarity with members of the genus Porphyromonas based on biochemical, chemical, and comparative 16s rRNA sequence analysis. As a result, Bacteroides levii (Holdeman, Cato, and Moore) was reclassified as Porphyromonas levii comb. now under the genus Porphyromonas.

Recent progress in bovine intestinal organoid research has expanded opportunities for creating improved in vitro models to study intestinal physiology and pathology. However, the establishment of a culture condition capable of generating organoids from all segments of the cattle intestine has remained elusive. Although previous research has described the development of bovine jejunal, ileal, and colonic organoids, this study marks the first report of successful bovine duodenal and rectal organoid development. Maintenance of these organoids through serial passages and cryopreservation was achieved, with higher success rates observed in large intestinal organoids compared to their small intestinal counterparts. A novel approach involving the use of biopsy forceps during initial tissue sampling streamlined the subsequent tissue processing, simplifying the procedure compared to previously established protocols in cattle. Additionally, our study introduced a more cost-effective culture medium based on Advanced DMEM/F12, diverging from frequently used commercially available organoid culture media. This enhancement improves accessibility to organoid technology by reducing culture costs. Crucially, the derived organoids from jejunum, ileum, colon and rectum faithfully preserved the structural, cellular, and genetic characteristics of in vivo intestinal tissue. This research underscores the significant potential of adult bovine intestinal organoids as a physiologically and morphologically relevant in vitro model. Such organoids provide a renewable and sustainable resource for a broad spectrum of studies, encompassing investigations into normal intestinal physiology in cattle and the intricate host-pathogen interactions of clinically and economically significant enteric pathogens.

Infections caused by tick-borne haemoparasites pose a significant global threat to both human and animal health. Within this category, various haemoparasites species belonging to genera like Anaplasma sp., Babesia sp., Ehrlichia sp., Hepatozoon sp., and Theileria sp., are particularly concerning due to their ability to cause diseases in a wide range of hosts, including mammals, birds, and reptiles. The present cross-sectional study involving 580 animals provides annual insights into the prevalence of major haemoparasites infections in the Bathinda region of Punjab. The observed trends indicate that haemoparasites infections were most common in cattle, followed by buffalo and canines. Risk factor analysis revealed that crossbreed cattle were more susceptible to infection, with a prevalence of 35.73% (95% CI 4.28-45.17). Amongst the cattle, adults exhibited a higher vulnerability to haemoparasites infections, with a prevalence of 35.89% (95% CI 5.50-33.64). Conversely, companion animals showed the opposite pattern, with a prevalence of 18.18% (95% CI 9.11-169.27). Furthermore, female dogs had a higher risk of haemoparasites infection, with a prevalence of 16.28% (95% CI 8.36-218.7). In light of these findings, it is imperative to emphasize early diagnosis, prompt antiprotozoals drug treatment, and effective control of tick vectors for the successful recovery of animals afflicted by haemoparasites infections.

In markets for beef and sheep meat, an appropriate level of intramuscular fat (IMF) is highly desirable for meat-eating quality, but strategies to improve it usually lead to an undesirable excess in carcase fat, presenting a major challenge to livestock producers. To solve this problem, we need to understand the partitioning of fat among the major fat depots: IMF, subcutaneous fat (SCF) and visceral fat (VF). In most genotypes of cattle and sheep, the rate of accretion is lower for IMF than for SCF and VF, so genetic selection for a high level of IMF, or the use of an increased dietary energy supply to promote IMF deposition, will increase overall fatness and feed costs. On the other hand, feeding postnatal calves with excessive concentrates promotes IMF deposition, so a nutritional strategy is feasible. With genetic strategies, several problems arise: 1) positive genetic correlations between IMF, SCF and VF differ among genotypes in both cattle and sheep; 2) genotypes appear to have specific, characteristic rates of accretion of IMF during periods of growth and fattening; 3) most breeds of cattle and sheep naturally produce meat with relatively low levels of IMF, but IMF does vary substantially among individuals and breeds so progress is possible through accurate measurement of IMF. Therefore, an essential prerequisite for selection will be knowledge of the genetic correlations and fat accretion rates for each genotype. Currently, selection for IMF is based on existing technology that directly measures IMF in the progeny or siblings, or estimates IMF in live animals. New technology is needed to permit the simultaneous measurement of SCF and IMF in the field, thus opening up the possibility of accurate selection, particularly for fat partitioning in live animals. Specifically, there would be great value in detecting individuals with an IMF advantage at an early age so the generation interval could be shortened and genetic gain accelerated. Genetic gain would also be greatly aided if we could select for genes that control adipogenesis and lipogenesis and are also differentially expressed in the various depots.