100K Pathogen Genome Project: 306 Listeria Draft Genome Sequences for Food Safety and Public Health.

Listeria monocytogenes is a food-associated bacterium that is responsible for food-related illnesses worldwide. This is the initial public release of 306 L. monocytogenes genome sequences as part of the 100K Pathogen Genome Project. These isolates represent global genomic diversity in L. monocytogenes

Draft Genome Sequences of 1,183 Salmonella Strains from the 100K Pathogen Genome Project.

Salmonella is a common food-associated bacterium that has substantial impact on worldwide human health and the global economy. This is the public release of 1,183 Salmonella draft genome sequences as part of the 100K Pathogen Genome Project. These isolates represent global genomic diversity in the Salmonella genus

Effects of Age and Mitochondrial Functionality on Canine Sperm Quality: Implications for Canine Fertility

Not all sires have sperm suitable for chilled or frozen storage, and success in artificial insemination (AI) varies highly among individual dogs and breeds. Fertilizing potential is further complicated as sperm quality declines with the aging process. The overall objective of this work was to study the effects of age and mitochondrial function on sperm quality and bioenergetics using traditional methods assessed in the clinical setting, as well as emerging methods of investigating metabolic flexibility and the heterogenous nature of sperm. This dissertation explores the physiology and pathophysiology of canine sperm through the study of a breeding colony of highly fertile Labrador retrievers, and a rapidly aging purebred population of North American Great Danes. This was accomplished in three chapters. Chapter 1, Comparative Oxidative Metabolism in Mammalian Sperm, provides a review of the literature and current understanding of mitochondrial physiology and metabolic control of essential sperm functions in mammalian species of veterinary importance. We discuss the differences in fundamental oxygen and ATP substrate balance in stallions, bulls, and dogs, with an emphasis on the maintenance of sperm motility, electron transport chain (ETC) function, reactive oxygen species (ROS) production, and the balance of glycolytic and oxidative phosphorylation production of ATP in sperm. In Chapter 2, Sperm Parameters in the Great Dane: Influence of Age on Semen Quality, I characterized the distribution of sperm quality parameters within a purebred population of actively showing North American Great Danes, a rapidly aging breed currently suffering declining fertility. I identified progressive declines in sperm quality that accompany aging, detailed the relationship between ROS production and sperm motility and morphology, and distinguished sources of variation in sperm quality within the breed. Age was negatively associated with several sperm motility parameters associated with sperm fertility such as total and progressive motility, and amplitude of lateral head displacement (p<.05), with a predicted -9.9%, -9.0%, and +8.3% change per year of age. Sperm of younger GD dogs aged 12 ≤ x <24 months had significantly higher total and progressive motility (TM, PM), amplitude of lateral head displacement (ALH), and nonlinear motility (p<.05) than older dogs (x ≥ 48 months). Results support anecdotal reports of decline of the fertility with the advance of age in this breed, and indicate that age and ROS have significant influences on sperm parameters in the GD. The influence of selection for breed specific phenotypes could help explain the functional significance of the diversity among GD males. In Chapter 3, Effects of Substrate Availability and Mitochondrial Disruption on Oxidative Metabolism and Sperm Motility in Fertile Dogs, I investigated the adaptability of canine sperm bioenergetics under differing nutrient conditions and the effects of mitochondrial dysregulation on sperm motility. With simultaneous measurement of mitochondrial oxygen consumption (MITOX) and sperm kinematics, I observed a high adaptability and metabolic flexibility of high-quality canine sperm. While all energetic substrates tested successfully maintained sperm motility, significant differences were found in MITOX and mitochondrial spare respiratory capacity (SRC), the strength with which mitochondrial energy production may rise to meet energetic challenge. In addition, the ability to maintain sperm kinematics when supplemented with various energetic substrates differed significantly with mitochondrial disruption by mitochondrial effectors. A population of Labrador retrievers from a highly fertile breeding colony managed by Guide Dogs for the Blind was chosen for this study due to their high semen quality, good health, and known fertility

Assessment of an iPad-based sperm motility analyzer for determination of canine sperm motility.

The objective of this study was to evaluate the repeatability and accuracy of canine sperm motility (total and progressive) assessment with a tablet-based Canine iSperm instrument compared to computer-assisted sperm analysis (CASA). The experiment used fresh and frozen/thawed canine semen samples for comparisons of semen analysis parameters (concentration, total motility, and progressive motility) between a CASA system, iSperm, and NucleoCounter SP-100 (concentration) instruments. Spearman's Rho correlational analysis was used to identify significant associations between motility assessment methods. Significant positive correlations were found between CASA assessment and iSperm for both progressive and total motility measurements. We also determined the coefficient of variation (CV) for repeatability of sample analysis for iSperm and CASA for fresh sperm, wherein each sample was assessed 10 times on both devices. For fresh and frozen-thawed samples, concentration assessment by iSperm showed high variability (CV= 19.9 ± 1.5%). For iSperm assessment of total and progressive motility, the CVs were 6.3 ± 0.5% and 10.7 ± 0.8%, respectively. The results indicate that the iSperm application offers an accurate and alternative measurement of motility to traditional CASA analysis, though caution should be taken when assessing concentration due to the high CV observed in this study

Assessment of an iPad-based sperm motility analyzer for determination of canine sperm motility.

The objective of this study was to evaluate the repeatability and accuracy of canine sperm motility (total and progressive) assessment with a tablet-based Canine iSperm instrument compared to computer-assisted sperm analysis (CASA). The experiment used fresh and frozen/thawed canine semen samples for comparisons of semen analysis parameters (concentration, total motility, and progressive motility) between a CASA system, iSperm, and NucleoCounter SP-100 (concentration) instruments. Spearman's Rho correlational analysis was used to identify significant associations between motility assessment methods. Significant positive correlations were found between CASA assessment and iSperm for both progressive and total motility measurements. We also determined the coefficient of variation (CV) for repeatability of sample analysis for iSperm and CASA for fresh sperm, wherein each sample was assessed 10 times on both devices. For fresh and frozen-thawed samples, concentration assessment by iSperm showed high variability (CV= 19.9 ± 1.5%). For iSperm assessment of total and progressive motility, the CVs were 6.3 ± 0.5% and 10.7 ± 0.8%, respectively. The results indicate that the iSperm application offers an accurate and alternative measurement of motility to traditional CASA analysis, though caution should be taken when assessing concentration due to the high CV observed in this study

100K Pathogen Genome Project: 306 Listeria Draft Genome Sequences for Food Safety and Public Health

Listeria monocytogenes is a food-associated bacterium that is responsible for food-related illnesses worldwide. This is the initial public release of 306 L. monocytogenes genome sequences as part of the 100K Pathogen Genome Project. These isolates represent global genomic diversity in L. monocytogenes

Quality Control of High-Throughput Library Construction Pipeline for KAPA HTP Library Using an Agilent 2200 TapeStation

<p>Next Generation Sequencing requires the input of high molecular weight genomic DNA to construct quality libraries for whole genome bacterial sequencing. Large scale sequencing projects, such as the 100K Pathogen Genome Project, require<br>100K Pathogen Genome Project methods to rapidly assess the quantity and quality of the input DNA using high-throughput methods that are fast and cost effective. In this study, the Agilent 2200 TapeStation and Agilent 2100 Bioanalyzer Systems were used to assess a few critical quality control steps for library construction. With minimal manual interven-tion, the Agilent 2200 TapeStation System determined the quality of genomic DNA, fragmented DNA, and final libraries constructed from multiple types of foodborne pathogens. The Agilent 2200 TapeStation System provided a single platform that effectively evaluated the necessary quality control steps, which provided a distinct advantage to decrease the time needed for library construction and a common<br>instrument methodology for quality control.</p> <p> </p

Draft Genome Sequences of 1,183 Salmonella Strains from the 100K Pathogen Genome Project.

Salmonella is a common food-associated bacterium that has substantial impact on worldwide human health and the global economy. This is the public release of 1,183 Salmonella draft genome sequences as part of the 100K Pathogen Genome Project. These isolates represent global genomic diversity in the Salmonella genus