New insights into AI semen use and objective semen evaluation in UK Veterinary Practice

The reproductive performance of breeding cattle is a key driver of sustainable beef and dairy production. Good fertility performance in breeding cattle is a multifactorial phenomenon and the male factors of fertility should not be overlooked. The aim of this study was to investigate the semen handling and storage practices on 47 farms utilising Artificial Insemination (AI) through a questionnaire, and then to use objective multi parametric semen analysis equipment in a veterinary practice to assess the quality of the AI semen in storage on the same beef and dairy farms in North Yorkshire. From these data, the proportion still above breeding company pre-release standards, and differences between conventional and sexed semen, beef and dairy, breed of bull, beef and dairy type of farm were investigated for all assays performed using Students t-test. Fresh semen from bull breeding soundness evaluations was also assessed and in the same laboratory and subjective and objective assessments of motility and morphology assessments compared. Finally semen evaluation results were compared with field fertility data on one farm and correlations reported. There were inconsistencies in how farmers stored and handled semen in storage on their farms. 12.1% of flasks in use were over 14 years old, over half of the herds surveyed (51.7%) did not respond with the age of flask, but 13.8% were new in the last 4 years. Of the herds that responded, the most common response (45.1%) was that liquid nitrogen levels were not checked at all and 6.1% herds checked their liquid nitrogen levels weekly. The majority (19.5%) of farms used a thaw temperature between 370C and 37.90C, next most common was 35 – 35.90C, 36 -36.90C accounted for 2.4% of farms, and only 1.2% of farms used >380C. The majority (28%) of farms that responded thawed straws for between 21 and 30 seconds, but some farms (2.4%) were using short thaw times of less than 10 seconds. Only 9.8% of farms thawed straws for longer than 30 seconds. 70.7% of herds in the study were using DIY AI; less than 25% of herds were using a technician service. 40.2% of herds were carrying out less than 50 serves/month. 30.5% of herds were serving more than 99 times per month. Semen analysis of straws of frozen semen taken from farm storage in North Yorkshire, UK was compared to pre-release standards used in UK and N America. Analysis of conventional and sex sorted semen showed respectively 65% and 70% were above the viability standard; 46% and 0% were above mitochondrial activity standard; 44% and 10% above CASA motility standard; 32% and 10% above CASA progressive motility standard in the UK; 86% and 40% above CASA progressive motility standard in North America (see table 3). Categories of semen were then compared using students t test, following visual assessment of normality. Comparisons between sex sorted and conventional straws showed significant differences in CASA motility (p=0.0182), progressive motility (p=0.0024), mitochondrial activity (p<0.0001) and morphology (p=0.0257). 11 | Page - Straws from dairy sires had significantly greater viability (p=0.0432) and morphology (p<0.0001) than beef sires, whereas straws from dairy farms had significantly greater acrosome integrity (p=0.0043), CASA motility (p=0.0129), progressive motility (p=0.0243) and morphology (p=0.0271) than straws from beef farms. There was no significant difference in motility (p=0.1001) or progressive motility between dairy and beef sires (p=0.0804). There was no significant difference in acrosome integrity between beef and dairy sire (p=0.1959). Investigation of field fertility outcomes on one dairy farm showed significant positive correlations between the flow cytometry viability assay and conception rate (r2 = 85.3% and P = 0.025), and also the CASA % motile at 2 hours post thaw and conception rate (r2 = 90% and P = 0.05). 70 semen samples from bull breeding soundness evaluations were evaluated in the same manner, as well as having manual assessment of morphology performed. Significant differences were present when on farm subjective motility assessments were compared with laboratory-based assessments (CASA motility and subjective), (p<0.0001) but not between CASA motility assessment in the lab and subjective assessment in the lab. There was no difference between morphological evaluations assessed by CASA or manually, despite CASA limitations in picking up sperm head abnormalities. Multiparametric objective semen analysis in a Veterinary practice-based laboratory can offer additional information on the semen in use on farm (AI and natural service), to complement traditional subjective methods. This may allow investigation as to what factors impact on semen quality and which parameters may therefore be most important when selecting semen to use on farm

High-throughput characterisation of bull semen motility using differential dynamic microscopy

<div><p>We report a high-throughput technique for characterising the motility of spermatozoa using differential dynamic microscopy. A movie with large field of view (∼10mm²) records thousands of cells (e.g. ≈ 5000 cells even at a low cell density of 20 × 10⁶ cells/ml) at once and yields averaged measurements of the mean () and standard deviation (<i>σ</i>) of the swimming speed, head oscillation amplitude (<i>A</i>₀) and frequency (<i>f</i>₀), and the fraction of motile spermatozoa (<i>α</i>). Interestingly, we found that the measurement of <i>α</i> is facilitated because the swimming spermatozoa enhance the motion of the non-swimming population. We demonstrate the ease and rapidity of our method by performing on-farm characterisation of bull spermatozoa motility, and validate the technique by comparing laboratory measurements with tracking. Our results confirm the long-standing theoretical prediction that for swimming spermatozoa.</p></div

Programmable RNA Shredding by the Type III-A CRISPR-Cas System of Streptococcus thermophilus

Immunity against viruses and plasmids provided by CRISPR-Cas systems relies on a ribonucleoprotein effector complex that triggers the degradation of invasive nucleic acids (NA). Effector complexes of type I (Cascade) and II (Cas9-dual RNA) target foreign DNA. Intriguingly, the genetic evidence suggests that the type III-A Csm complex targets DNA, whereas biochemical data show that the type III-B Cmr complex cleaves RNA. Here we aimed to investigate NA specificity and mechanism of CRISPR interference for the Streptococcus thermophilus Csm (III-A) complex (StCsm). When expressed in Escherichia coli, two complexes of different stoichiometry copurified with 40 and 72 nt crRNA species, respectively. Both complexes targeted RNA and generated multiple cuts at 6 nt intervals. The Csm3 protein, present in multiple copies in both Csm complexes, acts as endoribonuclease. In the heterologous E. coli host, StCsm restricts MS2 RNA phage in a Csm3 nuclease-dependent manner. Thus, our results demonstrate that the type III-A StCsm complex guided by crRNA targets RNA and not DNA. Highlights • Streptococcus thermophilus type III-A Csm (StCsm) complex targets RNA •Multiple cuts are introduced in the target RNA at 6 nt intervals •Csm3 protein subunits are responsible for endoribonuclease activity of the complex •StCsm complex offers a programmable tool for RNA degradatio

An Instrumental Case Study: Examining the Impact of Coaching on Building Middle School Teacher Agency

Background: According to Noonan (2014) and Calvert (2016), agency gives teachers the capacity to direct their professional growth in ways that positively influence student achievement. Agency is the combination of one’s belief in their effectiveness, or self-efficacy, and their actual knowledge and skills. Teachers who have acquired agency can identify ways to enhance their professional growth and positively impact student learning. As teachers develop their agency by finding effective ways to advance their professional learning, it is also important for school leaders to contribute to the improvement of teacher agency through coaching and development. The need for this support by school leaders is critical to ensure that teacher agency is acquired for the purpose of assisting with effective instruction for all learners in school settings. This research study is significant due to its focus on matters of literacy at the middle-school level. Literacy is the essential foundation for learning across the educational continuum. Purpose: The purpose of this study was to investigate how sustained coaching develops teacher agency. Research Question: The proposed study addressed the following question: What is the impact of instructional coaching on building agency among middle school literacy teachers? Methods: The qualitative instrumental case study was used to determine the impact of instructional coaching in developing teacher agency. This research design was appropriate because the research focused on the impact of instructional coaching provided for teachers who focus on literacy skills (Creswell, 2018). The convenience sampling strategy was used to select participants from schools that have received instructional coaching from the researcher. Teachers who accepted invitations to participate in the study partook in one (1) one-on-one semi-structured interview and one (1) focus group using the interview guidelines outlined by Creswell (2018). Qualitative data consisted of interview data and researcher journal entries and instructional artifacts. This data was analyzed utilizing constant comparative method for interview data, focus group, and content analysis for document data (journal entries and instructional artifacts) to identify emerging themes and key concepts. To ensure objectivity for this research study, a former NASA research methodologist was utilized for data review and analysis. This individual is adept at deriving common themes from standardized methods of inquiry and had no direct contact with any research participants. Findings: The study found that coaching significantly impacts teacher agency, teacher retention, and student achievement. Research data revealed five emerging themes: 1) coaching positively impacts teacher knowledge acquisition; 2) coaching positively impacts instruction delivery; 3) coaching positively impacts the way teacher’s viewed themselves (self-confidence); 4) coaching positively impacts teachers’ beliefs in their ability to deliver quality instruction (self-efficacy); and 5) coaching positively impacts student achievement. Conclusion: Instructional coaching has a significantly positive impact on building teacher agency