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Maternal recognition of pregnancy (MRP) in the mare is not well defined. In a non-pregnant mare, prostaglandin F2α (PGF) is released on day 14 post-ovulation (PO) to cause luteal regression, resulting in loss of progesterone production. Equine MRP occurs prior to day 14 to halt PGF production. Studies have failed to identify a gene candidate for MRP, so attention has turned to small, non-coding RNAs. The objective of this study was to evaluate small RNA (

Equine maternal recognition of pregnancy (MRP) is a process whose signal remains unknown. During MRP the conceptus and endometrium communicate to attenuate prostaglandin F2α (PGF) secretion, sparing the corpus luteum and maintaining progesterone production. Recognition of a mobile conceptus by the endometrium is critical by days 14-16 post-ovulation (PO), when endometrium produces PGF, initiating luteolysis. The objective of this study was to evaluate endometrial gene expression changes based upon pregnancy status via RNA sequencing. This experiment utilized a cross-over design with each mare serving as both a pregnant and non-mated control on days nine, 11, and 13 PO (n = 3/status/day). Mares were randomly assigned to collection day and pregnancy confirmed by terminal uterine lavage at the time of endometrial biopsy. Total RNA was isolated and libraries prepared using Illumina TruSeq RNA sample preparation kit. Reads were mapped and annotated using HISAT2 and Stringtie. Expression values were evaluated with DESEQ2 (P ≤ 0.05 indicated significance). On day nine, 11, and 13 there were 1435, 1435 and 916 significant transcripts, respectively. Multiple genes with splice variants had different expression patterns within the same day. These are the first data to evaluate the endometrial transcriptome during MRP on days nine, 11, and 13.

Abstract Background Small RNA (sRNA) regulatory pathways (SRRPs) are important to anti-viral defence in mosquitoes. To identify critical features of the virus infection process in Dengue serotype 2 (DENV2)-infected Ae. aegypti, we deep-sequenced small non-coding RNAs. Triplicate biological replicates were used so that rigorous statistical metrics could be applied. Results In addition to virus-derived siRNAs (20-23 nts) previously reported for other arbovirus-infected mosquitoes, we show that PIWI pathway sRNAs (piRNAs) (24-30 nts) and unusually small RNAs (usRNAs) (13-19 nts) are produced in DENV-infected mosquitoes. We demonstrate that a major catalytic enzyme of the siRNA pathway, Argonaute 2 (Ago2), co-migrates with a ~1 megadalton complex in adults prior to bloodfeeding. sRNAs were cloned and sequenced from Ago2 immunoprecipitations. Viral sRNA patterns change over the course of infection. Host sRNAs were mapped to the published aedine transcriptome and subjected to analysis using edgeR (Bioconductor). We found that sRNA profiles are altered early in DENV2 infection, and mRNA targets from mitochondrial, transcription/translation, and transport functional categories are affected. Moreover, small non-coding RNAs (ncRNAs), such as tRNAs, spliceosomal U RNAs, and snoRNAs are highly enriched in DENV-infected samples at 2 and 4 dpi. Conclusions These data implicate the PIWI pathway in anti-viral defense. Changes to host sRNA profiles indicate that specific cellular processes are affected during DENV infection, such as mitochondrial function and ncRNA levels. Together, these data provide important progress in understanding the DENV2 infection process in Ae. aegypti.

Abstract Background Due to the large number of hypothesis tests performed during the process of routine analysis of microarray data, a multiple testing adjustment is certainly warranted. However, when the number of tests is very large and the proportion of differentially expressed genes is relatively low, the use of a multiple testing adjustment can result in very low power to detect those genes which are truly differentially expressed. Filtering allows for a reduction in the number of tests and a corresponding increase in power. Common filtering methods include filtering by variance, average signal or MAS detection call (for Affymetrix arrays). We study the effects of filtering in combination with the Benjamini-Hochberg method for false discovery rate control and q-value for false discovery rate estimation. Results Three case studies are used to compare three different filtering methods in combination with the two false discovery rate methods and three different preprocessing methods. For the case studies considered, filtering by detection call and variance (on the original scale) consistently led to an increase in the number of differentially expressed genes identified. On the other hand, filtering by variance on the log2 scale had a detrimental effect when paired with MAS5 or PLIER preprocessing methods, even when the testing was done on the log2 scale. A simulation study was done to further examine the effect of filtering by variance. We find that filtering by variance leads to higher power, often with a decrease in false discovery rate, when paired with either of the false discovery rate methods considered. This holds regardless of the proportion of genes which are differentially expressed or whether we assume dependence or independence among genes. Conclusion The case studies show that both detection call and variance filtering are viable methods of filtering which can increase the number of differentially expressed genes identified. The simulation study demonstrates that when paired with a false discovery rate method, filtering by variance can increase power while still controlling the false discovery rate. Filtering out 50% of probe sets seems reasonable as long as the majority of genes are not expected to be differentially expressed.

Background: Rehabilitation of tendon injuries in horses often involves cryotherapy to reduce inflammation and occasionally tissue heating to increase collagen extensibility. The application of alternating cold and hot (ie contrast therapy) is widely used in human physical therapy; however, its utility in equine rehabilitation is largely unknown. Objectives: The objectives of this study were to (a) assess if the equipment could achieve therapeutic tissue temperatures (<15 and >40°C) at different tissue depths relative to the digital flexor tendons and (b) evaluate the time‐temperature profiles during serial heating and cooling cycles using a contrast therapy device. Study design: In vivo experiment. Methods: In 4 adult horses with normal forelimb digital flexor tendons, fine‐wire temperature probes were placed superficially on the skin and implanted subcutaneously, deep to the superficial digital flexor tendon (SDFT) and deep to the deep digital flexor tendon (DDFT). Temperatures were recorded over three complete thermal (hot‐cold) cycles. Minimum and maximum temperatures were recorded and the rate of temperature changes and the areas underneath the time‐temperature curves (ie thermal load) were calculated. Results: Minimum and maximum tissue temperatures (°C) included: superficial skin [12.6 ± 1.0; 42.4 ± 2.4], subcutaneous tissues [14.1 ± 0.8; 42.3 ± 2.2], deep to the SDFT [15.6 ± 0.8; 41.7 ± 2.6] and deep to DDFT [25.1 ± 2.0; 38.0 ± 3.5]. An initial rapid rate of tissue temperature change between 3.2 and 4.3°C/min occurred within tissues to the depth of the DDFT. Tissue thermal loads during heating ranged from 255 to 607°C*second and from 309 to 780°C*second during tissue cooling, with the lower values noted deep to the DDFT. Main limitations: Unknown clinical efficacy in diseased tissues. Conclusions: The applied contrast therapy was consistently able to induce cooling and heating of tissues to the depth of the DDFT. [ABSTRACT FROM AUTHOR]

Abstract Background Small RNA regulatory pathways (SRRPs) control key aspects of development and anti-viral defense in metazoans. Members of the Argonaute family of catalytic enzymes degrade target RNAs in each of these pathways. SRRPs include the microRNA, small interfering RNA (siRNA) and PIWI-type gene silencing pathways. Mosquitoes generate viral siRNAs when infected with RNA arboviruses. However, in some mosquitoes, arboviruses survive antiviral RNA interference (RNAi) and are transmitted via mosquito bite to a subsequent host. Increased knowledge of these pathways and functional components should increase understanding of the limitations of anti-viral defense in vector mosquitoes. To do this, we compared the genomic structure of SRRP components across three mosquito species and three major small RNA pathways. Results The Ae. aegypti, An. gambiae and Cx. pipiens genomes encode putative orthologs for all major components of the miRNA, siRNA, and piRNA pathways. Ae. aegypti and Cx. pipiens have undergone expansion of Argonaute and PIWI subfamily genes. Phylogenetic analyses were performed for these protein families. In addition, sequence pattern recognition algorithms MEME, MDScan and Weeder were used to identify upstream regulatory motifs for all SRRP components. Statistical analyses confirmed enrichment of species-specific and pathway-specific cis-elements over the rest of the genome. Conclusion Analysis of Argonaute and PIWI subfamily genes suggests that the small regulatory RNA pathways of the major arbovirus vectors, Ae. aegypti and Cx. pipiens, are evolving faster than those of the malaria vector An. gambiae and D. melanogaster. Further, protein and genomic features suggest functional differences between subclasses of PIWI proteins and provide a basis for future analyses. Common UCR elements among SRRP components indicate that 1) key components from the miRNA, siRNA, and piRNA pathways contain NF-kappaB-related and Broad complex transcription factor binding sites, 2) purifying selection has occurred to maintain common pathway-specific elements across mosquito species and 3) species-specific differences in upstream elements suggest that there may be differences in regulatory control among mosquito species. Implications for arbovirus vector competence in mosquitoes are discussed.

Objective: To describe and compare normative anterior segment parameters between canine age groups using the Pentacam® HR Scheimpflug camera (Pentacam). Animals Studied: Thirty‐six sedated dogs (60 eyes) of varying ages and breeds were imaged with the Pentacam; only nondiseased anterior segments were included. Procedures: Dogs were divided into three age groups: Group 1 (1‐5 years), Group 2 (6‐10 years), and Group 3 (11‐15 years). Values assessed included central corneal thickness (CCT), anterior and posterior corneal elevation (ACE/PCE), anterior and posterior corneal curvature metrics, corneal volume (CV), anterior and posterior corneal astigmatism (AA/PA), anterior chamber depth (ACD), anterior chamber volume (ACV), and anterior chamber angle (ACA). Tukey‐adjusted pairwise comparisons were performed. Results: Overall CCT (mean ± SD) was 631.07 ± 59.91 µm. Central corneal thickness was 608.60 ± 48.63 µm for Group 1, 648.57 ± 51.06 µm for Group 2, and 635.37 ± 73.71 µm for Group 3. Anterior corneal elevation (ACE) measured 9.08 ± 0.58 mm, PCE measured 8.04 ± 0.50 mm, and CV was 58.13 ± 5.39 mm3. Astigmatism values were 1.34 ± 0.94 D for AA and 0.46 ± 0.44 D for PA. Anterior chamber values were 3.76 ± 0.56 mm for ACD, 383.68 ± 66.24 mm3 for ACV, and 23.62 ± 29.33˚ for ACA. Significant differences were found between Groups 1 and 2 for CV (55.08 ± 4.08 mm3 and 60.32 ± 4.19 mm3, respectively), (P = 0.02). Conclusions: Corneal volume significantly increased between Group 1 and Group 2. Central corneal thickness increased from Group 1 to Group 3, but was not significant with the current sample size. There were no other differences between age groups. [ABSTRACT FROM AUTHOR]

Purpose: Identify functional factors that are important correlates to physical activity levels among people with multiple sclerosis. Methods: A total of eight functional tests were conducted and physical activity was objectively measured (Actigraph GT3X accelerometer) for one week in 34 people with multiple sclerosis. A corrected Akaike Information Criterion analysis was performed to identify the strongest correlates with moderate-to-vigorous physical activity, total activity and sedentary time. Results: The multiple regression analysis converged on a model for moderate-to-vigorous physical activity (R2 = 0.31, F = 6.97, p= 0.003) that included total strength of the less-affected leg (partial r = 0.46, p = 0.007) and average peg test performance (partial r = −0.30, p = 0.087). The model for total activity (R2 = 0.40, F = 10.51, p < 0.001) included five times sit-to-stand performance (partial r= −0.44, p = 0.010) and total strength of the less-affected leg (partial r = 0.31, p = 0.077). The model for sedentary time (R2=0.22, F = 9.23, p = 0.005) only included total strength of the more affected leg (r= −0.47, p = 0.005). Conclusion: These results suggest that leg strength, manual dexterity and the ability to perform functional tasks may be important correlates with physical activity levels in people with multiple sclerosis. The findings of this pilot study can inform future investigations aiming to increase physical activity levels or develop improved rehabilitation protocols for people with multiple sclerosis. Implications for Rehabilitation: Physical activity is an effective means of improving the symptoms associated with multiple sclerosis. Participation in physical activity by people with multiple sclerosis may be affected by functional factors such as leg strength, manual dexterity and the ability to rise from a seated position. Bilateral leg strength differences should be assessed and addressed in people with multiple sclerosis. [ABSTRACT FROM AUTHOR]

Abstract: Objective: To compare central corneal thickness (CCT) values in canine eyes using Pentacam‐HR® Scheimpflug imaging (Pentacam), Optovue® iVue spectral‐domain optical coherence tomography (SD‐OCT), and high‐resolution ultrasound biomicroscopy (UBM) and generate normative canine Pentacam CCT values. Animals studied: Twenty‐four client‐owned dogs (37 eyes) with nondiseased cornea(s) presenting to the Colorado State University Veterinary Teaching Hospital. Procedures: Corneal images were acquired via Pentacam, SD‐OCT, and UBM in the listed order. Machine‐calculated values of CCT from Pentacam and SD‐OCT were compared to operator‐measured values from UBM. Bland–Altman analysis was performed to evaluate agreement between instruments. Results: Mean CCT ± SD measured by Pentacam was 629.73 ± 64.57 μm, by SD‐OCT was 610.56 ± 57.48 μm, and by UBM was 689.77 ± 55.93 μm. On average, Pentacam CCT was 19.17 ± 32.90 μm (3%) thicker than SD‐OCT and 65.12 ± 44.52 μm (10.3%) thinner than UBM. SD‐OCT was on average 82.47 μm (13.5% ) thinner than UBM. The 95% limits of agreement were (−45.31, 83.65), (−152.38, 22.13), and (−126.674, −38.270) for Pentacam vs. SD‐OCT, Pentacam vs. UBM, and SD‐OCT vs. UBM respectively. All differences were statistically significant (P < 0.01). Conclusions: Considering there is an average of 7.5% normal diurnal variation in canine CCT, a 3.0% difference between Pentacam and SD‐OCT values is likely not clinically relevant. However, Pentacam measurements were both statistically and clinically significantly different from UBM and SD‐OCT measurements. [ABSTRACT FROM AUTHOR]

Background. Early Lyme disease patients often present to the clinic prior to developing a detectable antibody response to Borrelia burgdorferi, the etiologic agent. Thus, existing 2-tier serology-based assays yield low sensitivities (29%-40%) for early infection. The lack of an accurate laboratory test for early Lyme disease contributes to misconceptions about diagnosis and treatment, and underscores the need for new diagnostic approaches. Methods. Retrospective serum samples from patients with early Lyme disease, other diseases, and healthy controls were analyzed for small molecule metabolites by liquid chromatography-mass spectrometry (LC-MS). A metabolomics data workflow was applied to select a biosignature for classifying early Lyme disease and non-Lyme disease patients. A statistical model of the biosignature was trained using the patients' LC-MS data, and subsequently applied as an experimental diagnostic tool with LC-MS data from additional patient sera. The accuracy of this method was compared with standard 2-tier serology. Results. Metabolic biosignature development selected 95 molecular features that distinguished early Lyme disease patients from healthy controls. Statistical modeling reduced the biosignature to 44 molecular features, and correctly classified early Lyme disease patients and healthy controls with a sensitivity of 88% (84%-95%), and a specificity of 95% (90%-100%). Importantly, the metabolic biosignature correctly classified 77%-95% of the of serology negative Lyme disease patients. Conclusions. The data provide proof-of-concept that metabolic profiling for early Lyme disease can achieve significantly greater (P < .0001) diagnostic sensitivity than current 2-tier serology, while retaining high specificity. [ABSTRACT FROM AUTHOR]