Sampling Time Points Research Articles

Simulation of spatially varying non-stationary seismic waves using random frequencies and non-uniform fast fourier transform

The simulation of spatially varying non-stationary seismic waves is a crucial consideration for the seismic analysis of engineering structures. The traditional approach often incurs substantial computational expenses and suffers from low efficiency due to the extensive implementation of large-scale matrix factorizations at numerous sampling points in both time and frequency domains. In this study, an enhanced approach is developed for the efficient simulation of non-stationary seismic waves. The core of this approach lies in the utilization of random frequency sampling coupled with time-domain interpolation to reduce matrix factorizations, and non-uniform fast Fourier transform for expediting the summation of trigonometric functions. A parametric analysis is conducted to investigate the effect of vital parameters, including the decoupling strategy of time-frequency spectrum, the number of random frequencies, and the number of interpolation knots over time, on both the efficiency and accuracy of the enhanced approach. Consequently, the preferred values for these parameters are determined. By considering the simulation of seismic waves acting on a group of bridges, the enhanced approach is validated in the view of correlation functions. The results demonstrate a high level of fidelity of the simulated seismic waves and proves the effectiveness of the enhanced approach in the efficient simulation of spatially varying non-stationary earthquake ground motions. The developed approach can be used for the simulation of non-stationary seismic waves for a wide range of engineering structures.

Defining the optimal setting for transcriptomic analyses on blood samples for response prediction in immunotherapy-treated NSCLC patients

Transcriptomic profiling of blood immune cells offers a promising alternative to invasive, sampling bias-prone tissue-based biomarker assays for predicting immune checkpoint inhibitor (ICI) therapy response in non-small cell lung cancer (NSCLC) patients. However, the optimal analytical approach to identify systemic correlates of response still needs to be explored. We collected peripheral blood mononuclear cells and whole blood (WB) samples from 33 ICI-treated NSCLC patients before ICI treatment and at the first response evaluation. After bulk polyadenylated RNA-sequencing, we assessed differences in gene expression profiles between non-responders and responders using differential expression analysis, single sample gene set enrichment analysis (ssGSEA), and cell type deconvolution. We evaluated gene expression values, ssGSEA scores, and deconvolved cell type proportions to distinguish non-responders from responders via ROC curve (AUC) analysis, training a logistic regression classification model. Gene expression values and deconvolved proportions yielded the best results with WB samples after treatment (AUC = 0.87 and 0.85, respectively). Overall, ssGSEA scores showed superior classification performance across all sample types and timepoints (AUC > 0.7). In conclusion, transcriptomic analysis through ssGSEA demonstrated the best performance as a non-invasive biomarker for predicting clinical benefit in ICI-treated NSCLC patients, with gene expression and deconvolution on post-treatment WB samples also showing promising results.

Calcium propionate is an alkalizing agent in exercising Standardbreds

The objective of this study was to determine if calcium propionate is an alkalizing agent in exercising Standardbreds and if it alters plasma glucose and serum insulin concentrations. This study used a randomized crossover design to test the hypotheses that calcium propionate alters total CO2 (tCO2), Ca++, pH, strong ion difference (SID), glucose, and insulin in Standardbreds completing a simulated race test (SRT) on a high-speed equine treadmill. Blood was collected from eight horses (mean age ± SD = 16 ± 2.7 years; range = 13 - 21 years) at 10 and 5 min prior to treatment or control administration, just prior to the subsequent SRT, directly after the SRT, and at 60- and 90-min post SRT. Plasma pH and plasma concentrations of tCO2, glucose, Ca++, and Na+, K+, Cl-, Lac- (for SID calculation) were measured in duplicate by blood gas analyzer and serum insulin by radioimmunoassay. Data were analyzed by two-way repeated-measures ANOVA. For plasma pH, Na+, K+, Cl-, Lac-, SID, insulin and glucose, no effects of treatment or treatment by sampling-timepoint interaction were found (p>0.05). Plasma tCO2, Ca++, pH, Na+, SID, and glucose concentrations were significantly lower, and Lac- was significantly higher, directly after the SRT compared to all other timepoints (p<0.05). Plasma tCO2, HCO3- and Ca++ were significantly higher in treated than in control horses at multiple sampling timepoints after treatment/control administration (p<0.05). Serum insulin concentration, measured only pre and 30 min post treatment/control administration, was unaffected by treatment. Calcium propionate is an alkalizing agent in horses.

Comparative Preclinical Pharmacokinetics and Disposition of Favipiravir Following Pulmonary and Oral Administration as Potential Adjunct Therapy Against Airborne RNA Viruses.

Favipiravir is administered orally, even against airborne RNA viruses, in a loading-dose/maintenance dose regimen. We investigated whether-(a) pulmonary delivery of favipiravir would generate high concentrations in the luminal side of the respiratory tract; and (b) avoiding first-pass metabolism by the liver by inhaled drug would generate comparable pharmacokinetics (PK) with doses significantly smaller than the oral maintenance dose. A dry powder inhalation (DPI) of favipiravir formulated by mixing with Inhalac 400® was prepared and characterized. Inhalations of ~ 120µg dose strength, with or without a prior oral loading dose were administered to mice. Comparator mice received human-equivalent oral doses (3mg). Three mice per sampling time point were sacrificed and favipiravir concentrations in the blood plasma, bronchio-alveolar lavage fluid (BALF) and lung tissue homogenate determined by HPLC. One-compartment PK modeling of concentration-time data indicated that the area under the curve (AUC0-24h) generated in the BALF recovered from mice receiving inhalations of ~ 1/25th of the oral dose subsequent to an oral loading dose was 86.72 ± 4.48µg⋅mL-1⋅h. This was consistently higher than the AUC observed in the BALF of orally-dosed mice (56.71 ± 53.89µgmL-1⋅h). In blood serum, the respective values of AUC were 321.55 ± 124.91 and 354.71 ± 99.60µg⋅mL-1⋅h. Pulmonary delivery of significantly smaller doses of favipiravir generates meaningful drug disposition and pharmacokinetics at the site of respiratory viral infections. We provide the rationale for designing a self-administered, non-invasive, low-cost, targeted drug delivery system against airborne RNA virus infection.

Protocol of clinical population pharmacokinetics/pharmacodynamics analysis of alkaloids from the leaves of alstonia scholaris in acute bronchitis patients

Objective: To validate whether orally administered Alkaloid Capsules from the leaves of Alstonia scholaris (CALAS) can improve the clinical indices of acute bronchitis and to investigate the alterations in the relationship between its composition and pharmacodynamic (PD) markers, thereby providing a clinical reference for the administration of this medication. Methods: This is a prospectively planned, blinded, placebo-controlled, parallel-grouped clinical trial with aggregated population pharmacokinetics/PD (PPK/PPD) data. A total of 55 subjects will be randomly allocated into four arms; specifically, 10 of the 55 subjects will be selected randomly for the placebo arm, and will be orally administered placebo (Tid), and 45 subjects will be randomly assigned to CALAS treatment groups (20 mg, 40 mg, and 80 mg Tid, at 15 subjects per group). The medication, presence of cough and phlegm, as well as body temperature of every subject will be recorded daily during treatment. About 3–4 blood samples will be collected from each subject at the following points for PPK/PPD parameters analysis: at pre-dose (0 h) and post-dose at 15 min, 40 min, 1 h, 1.5 h, 2 h, 3 h, 4 h, 6 h, 8 h, 12 h, 24 h, 30 h, and 48 h after last dosing. All the subjects will be subjected to a laboratory examination and efficacy evaluation on day 8. Discussion: A new integrating strategy to explore the relationship among drug components, action pathways, and clinical efficacy will be established through this study. We aim to explore the mechanism of action of CALAS in the treatment of acute bronchitis on the premise of definite active ingredients and reliable clinical efficacy. It is difficult to enrol patients in classical PK research because it adopts an intensive sampling method, and it cannot quantify the variability of PK parameters (Intra-individual variation, inter-individual variation and weekly variation). Moreover, the extrapolation and prediction of dosage regimens in different species and populations cannot be achieved. Therefore, the PPK/PPD method, which takes advantage of sparse data (3–5 time points sampling per patient), is adopted to determine the measurable pathological and physiological factors that can influence dose concentration to guide reasonable dose adjustment towards achieving optimal clinical effects.

Land use drives prokaryotic community composition of directly adjacent grasslands

AbstractUnderstanding the impact of agricultural land use on the soil prokaryotic communities in connected downslope sites is crucial for developing sustainable strategies to preserve ecosystem properties and mitigate agriculture’s environmental impacts. In this study, we investigated topsoil samples collected at three time points in 2022 (March, June, and November) from two adjacent catenas, reaching from hillslope to floodplain. The catenas differed in land use (extensive grassland vs. extensive cropland) at the top and middle parts, while the floodplain remained an extensive grassland due to legal restrictions. Using quantitative real-time PCRs and metabarcoding, we assessed prokaryotic abundance and prokaryotic community composition. Results show higher bacterial abundance in the cropland-influenced floodplain part across all time points compared to the grassland-influenced floodplain part. Temporal dynamics revealed a progressive decrease in the shared prokaryotic communities of the floodplain parts, peaking at the summer sampling time point, indicating a significant influence of the respective management type of the agricultural sites over the bacterial and archaeal communities of the floodplain parts. Differential abundance analyses identified several nitrifying taxa as more abundant in the cropland-influenced floodplain. Upstream land use also influenced the prokaryotic network of the cropland-floodplain, with some cropland taxa becoming keystone taxa and altering network morphology, an effect not observed in the grassland-influenced floodplain. These findings suggest that upstream agricultural land use practices have exerted a long-term influence on the floodplain prokaryotic communities over the past three decades. Moreover, there is evidence suggesting that these prokaryotic communities may undergo a potential reset during winter, which requires further investigation.

A systematic evidence map protocol for mapping global exposure to bisphenols and their alternatives and social and environmental justice implications

BackgroundBisphenol A (BPA) is one of the highest-volume chemicals produced worldwide, and human exposure to BPA is thought to be ubiquitous. Stricter regulations around the use of BPA have led many manufacturers to switch to other bisphenol chemicals with similar functions such as bisphenol S and F. Even though exposure to BPA, other bisphenol chemicals and bisphenol alternatives poses a health risk for humans, very little is known about the granular exposure levels of different populations around the world. AimThis systematic evidence map (SEM) will identify human studies reporting concentrations of bisphenols and their alternatives measured in human bio-samples with the aim to chart the global human exposure levels by country and population characteristics to identify research gaps and discuss any social and environmental injustice implications. Search strategy and eligibility criteriaMEDLINE®, Embase and Web of Science (WoS) databases as well as grey literature sources will be searched using predefined search strings. The database search results will be supplemented by backward and forward citation tracking on included studies. A scoping exercise conducted during planning identified 90 bisphenol chemicals and alternatives used in plastics. These include BPA, other bisphenol chemicals with/without similar functions to BPA as well as alternatives with similar functions to bisphenol chemicals. Eligible studies must report concentrations of at least one relevant bisphenol chemical/alternative measured in human bio-samples. Study selectionOnly primary studies published in English since 2010 will be considered. The title, abstract and keywords will be screened by the DistillerAI tool and two independent reviewers. Grey literature will be screened by two reviewers for inclusion and exclusion. The full text of the included studies will then be screened by two independent reviewers. Study appraisalStudy quality will not be evaluated in this SEM. Data extraction and codingData extraction and coding will be performed by two independent reviewers. Parameters of interest will include the following: study characteristics (e.g., year of publication, sampling timepoints and study design), population information (e.g., country, age, sex, ethnicity, number of participants) and exposure information (sources of exposure, bio-sample analyzed, chemical name, concentration, and detection frequencies). Synthesis and visualizationThe results will be presented using a narrative summary, tables, bar plots and color-coded maps. The interactive database will be available on a dedicated freely accessible website. Systematic map protocol registry and registration numberThis protocol has been registered on Open Science Framework (OSF) and is available at https://doi.org/10.17605/OSF.IO/MNWTD.

The impact of an oral glucose load on IFN-γ-release in persons infected with Mycobacterium tuberculosis

Background and objectiveTo diagnose tuberculosis infection (TBI), whole blood is incubated with M.tuberculosis (Mtb)-specific peptides and the release of interferon-γ (IFN-γ) is measured in IFN-γ-release assays (IGRAs). Hyperglycaemia and fluctuations in blood glucose may modulate IFN-γ-release. Here, we investigated if glucose intake affects IFN-γ-release or IGRA results in IGRAs taken during an oral glucose tolerance test (OGTT).MethodsPersons with TB disease (TB) or TBI underwent a standard 75-g OGTT at the start and end of treatment for TB or TBI. Blood for the IGRA QuantiFERON-TB Gold Plus (QFT) containing Mtb-specific tubes (TB1 and TB2), a non-specific mitogen tube (MIT) and an empty control tube (NIL) was drawn at sample-timepoints -15 (baseline), 60, 90, 120 and 240 min during the OGTT. Blood glucose was measured in parallel at all timepoints. IFN-γ-release (after subtraction of NIL) at each timepoint was compared with baseline using linear-mixed-model analysis.ResultsTwenty-four OGTTs from 14 participants were included in the final analysis. Compared to baseline, IFN-γ-release was increased at sample-timepoint 240 min for TB1; geometric mean (95% confidence interval) 3.0 (1.5–6.2) vs 2.5 (1.4–4.4) IU/mL (p = 0.047), and MIT; 182.6 (103.3–322.9) vs 146.0 (84.0–254.1) IU/mL (p = 0.002). Plasma glucose levels were not associated with IFN-γ-release and the QFT test results were unaffected by the OGTT.ConclusionIngestion of glucose after a 10-h fast was associated with increased IFN-γ-release after 240 min in the MIT tube. However, there was no association between plasma glucose levels at the QFT sampling timepoint and IFN-γ-release. Furthermore, the QFT test results were not affected by glucose intake. The overall effect of an OGTT and prevailing plasma glucose levels on IFN-γ-release in IGRAs seem limited.Trial registrationTrial registration ID: NCT04830462 (https://clinicaltrials.gov/study/NCT04830462). Registration date: 05-Apr-2021.

Application of shadowgraph imaging (SGI) particle characterisation data to interpret the impact of varying test conditions on powder dissolution and to develop an automated agglomeration identification method (AIM) in the USP flow-through apparatus

The aims of this work were 1) to explore the application of shadowgraph imaging (SGI) as a real time monitoring tool to characterize ibuprofen particle behaviour during dissolution testing under various conditions in the USP 4 flow-through apparatus and 2) to investigate the potential to develop an SGI-based automated agglomeration identification method (AIM) for real time agglomerate detection during dissolution testing. The effect of surfactant addition, changes in the drug mass and flow rate, the use of sieved and un-sieved powder fractions, and the use of different drug crystal habits were investigated. Videos at every sampling time point during dissolution were taken and analysed by SGI. The AIM was developed to characterize agglomerates based on two criteria – size and solidity. All detections were confirmed by manual video observation and a reference agglomerate data set. The method was validated under new dissolution conditions with un-sieved particles. Characterisation of particle dispersion behaviour by SGI enabled interpretation of the impact of dissolution test conditions. Higher numbers of early detections reflected greater dissolution rates with increased surfactant concentration, using sieved fraction or plate-shaped crystals, but was impacted by drug mass tested. An AIM was successfully developed and applied to detect agglomerates during dissolution, suggesting potential, with appropriate method development, for application in quality control.

Surveillance of tick-borne pathogens in domestic dogs from Chad, Africa

BackgroundTick-borne pathogens are understudied among domestic animals in sub-Saharan Africa but represent significant threats to the health of domestic animals and humans. Specifically, additional data are needed on tick-borne pathogens in Chad, Africa. Surveillance was conducted among domestic dogs in Chad for selected tick-borne pathogens to measure (1) the prevalence of antibodies against Anaplasma spp., Borrelia burgdorferi, and Ehrlichia spp.; (2) the prevalence of infections caused by Hepatozoon spp., Ehrlichia canis, Anaplasma platys, and Babesia spp.; and (3) associations of pathogens with demographic, spatial, and temporal factors. Blood samples were collected from domestic dogs at three time points (May 2019, November 2019, June 2020) across 23 villages in southern Chad.ResultsOf the 428 dogs tested with the IDEXX SNAP 4Dx test in May 2019, 86% (n = 370, 95% CI = 83–90%) were positive for antibodies to Ehrlichia spp., 21% (n = 88, 95% CI = 17–25%) were positive for antibodies to Anaplasma spp., and 0.7% (n = 3, 95% CI = 0.1–2%) were positive for antibodies to Borrelia burgdorferi. Four different pathogens were detected via PCR. Hepatozoon spp. were most commonly detected (67.2–93.4%, depending on the time point of sampling), followed by E. canis (7.0-27.8%), A. platys (10.1–22.0%), and Babesia vogeli (0.4–1.9%). Dogs were coinfected with up to three pathogens at a single time point, and coinfections were most common in May 2019 compared to November 2019 and May 2020.ConclusionsOverall, this study provides new data about the epidemiology of tick-borne pathogens in domestic dogs in Chad, with potential implications for dog and human health.

Evaluation of stress in farmed Atlantic salmon (Salmo salar) using different biological matrices

Atlantic salmon were subjected to an acute crowding scenario, and their subsequent stress responses were observed under three distinct swimming speed/water flow (WF) conditions: 0.5, 1, and 1.5 body lengths per second (BL/s). Feces, dermal mucus, and plasma were collected for analysis at 1, 6, and 24 h (h) post-stress. Additionally, the head kidney and two regions of the brain (pituitary and POA) were collected for transcript expression analysis. Fish swimming at 0.5 BL/s exhibited higher pre-stress (baseline) cortisol levels. Across all groups and matrices, the highest cortisol/cortisol metabolites (CM) levels were observed at the 1 h post-stress sampling point. At 6 h (second sampling time point), a clear decline toward baseline levels were observe in all groups. Significant increases in mean plasma glucose levels were observed at 1 h post-stress for all groups. The mean plasma lactate levels varied based on WF treatments, with a significant increase observed at 1 h only for the 1.5 BL/s group. Additionally, significant decreases in mean plasma lactate were noted at 6 and 24 h post-stress for some groups. The mRNA abundances of the tested genes (star, cyp17a1, hsd11β2, srd5a1) increased following the stress events. These changes were not uniform across all groups and were tissue dependent. In summary, the results indicate that mucus and feces can be used as potentially less invasive matrices than blood for evaluating stress and, consequently, the welfare of Atlantic salmon in captivity.

PSIV-15 Effect of transient fetal bovine viral diarrhea virus infection on postnatal growth, estimated dry matter digestibility, glucose concentrations, and carcass characteristics

Abstract Fetal infection of bovine viral diarrhea virus (BVDV) before d 125 to 150 of gestation results in the birth of immunotolerant, persistently infected (PI) calves. Infection of BVDV during late gestation results in transient fetal infections (TI). Unvaccinated, yearling Hereford heifers (n = 25), seronegative for antibodies to BVDV1 and BVDV2, were bred by artificial insemination with X chromosome-bearing sperm from an Angus sire to examine the impact of TI on postnatal growth, estimated dry matter digestibility, blood glucose concentrations, and carcass characteristics. On d 175 of pregnancy, heifers were intranasally inoculated with either DMEM + 2% horse serum (sham control) to generate control calves or 4.0 log TCID50 noncytopathic type2 BVDV to generate TI calves. All sham-inoculated control dams remained seronegative, and all BVDV-inoculated dams seroconverted by d 14 post-inoculation. Sham-inoculated control dams (n = 12) and BVDV-inoculated dams (n = 12) gave birth to live calves. All control offspring were seronegative, and all TI offspring were seropositive for antibodies to type 2 BVDV at birth. All offspring were raised on pasture until weaning. At weaning, all calves were transported to our feedlot research facility, housed in one group feedlot pen, and transitioned to a high-energy concentrate-based diet until reaching an approximate BW of 600 kg. Upon arrival at the feedlot, all animals received a standard heifer growth implant, a modified live viral vaccine containing IBR-BRSV-PI3 and were dewormed. Heifer BW and jugular blood samples were collected every 28 d. On d 84 of the feeding period, titanium dioxide was added to the diet of 12, age-paired, individually fed, heifers (3 control and 3 TI heifers; approximately 1 yr of age) for 28 d and used to estimate dry matter digestibility. After approximately 280 d on feed heifers were transported to a USDA-inspected abattoir and harvested. The TI heifers had lighter birth weights (P &amp;lt; 0.03) and final BW (P &amp;lt; 0.04) when compared with control heifers. Average daily gain was greater (P &amp;lt; 0.01) in control compared with TI heifers. Blood glucose concentrations were similar between control and TI heifers at all sampling time points. Dry matter intake of individually fed heifers was similar across treatments. TI heifers had a 2.2% lesser (P &amp;lt; 0.05) dry matter digestibility and lighter (P &amp;lt; 0.01) hot carcass weights compared with controls. These data suggest that TI fetal BVDV infection negatively impacts growth throughout the feeding period, possibly by impacting gastrointestinal tract function. This research was supported by USDA-NIFA Grant # 2019-67015-29866.

PSV-16 Evaluation of upper respiratory and fecal microbiomes in neonatal beef calves

Abstract There is a critical lack of information on the timing of colonization of bovine respiratory disease-related bacteria in the upper respiratory tract, potentially affecting calf health and disease onset. The objective of this study was to evaluate the upper respiratory tract and fecal microbiomes of neonatal beef calves during the first 24 h of life compared with their dams at the time of birth. Late-gestation commercial beef cows (n = 28) were assigned to individual, soil-surfaced pens (6.25 x 2.29 m) fitted with canvas tarps on shared fences to limit contact between unrelated pairs. Left nasal (LN), right nasal (RN), fecal, and vaginal swab samples were collected from cows at a single time point promptly after parturition. Swabs of the LN, RN, and rectum were collected from calves at birth (0 h), 6, 12, and 24 h post-parturition. Samples were flash-frozen and stored at -80°C until processing. DNA was extracted from swabs using Power Soil Pro kit. Extracted DNA was used to prepare DNA libraries for full-length 16S rRNA gene sequencing on a MinION Mk1C device. Sequenced reads were classified using Centrifuge and used in microbiome analyses conducted in R to estimate the changes in the microbiome according to animal type (cow versus calf), time of collection, and sample type (nasal, fecal, or vaginal). Upper respiratory microbiomes significantly differed (P &amp;lt; 0.005) between cows and their offspring at birth. In nasal microbiomes, animal type explained 12.6% (P = 0.002) and 18.5% (P = 0.001) of variance in beta diversity in left and right cavities, respectively. Animal type explained 42.1% of the beta diversity variance in fecal microbiomes (P = 0.001). Nasal microbiomes at birth were significantly more dispersed in dams compared with their calves in both the LN (P = 0.03) and RN (P = 0.03). Similarly, fecal microbiomes of dams were significantly more dispersed (P &amp;lt; 0.0001) than those of their offspring. There was a clear transition of the microbial communities associated with calf age in the upper respiratory and fecal niches. Calf age accounted for 2% (P = 0.02), 3.5% (P = 0.001), and 28% (P = 0.001) of the variance in the beta diversity of the LN, RN, and fecal microbiomes, respectively. After adjusting for calf age, the laterality of nasal microbiomes was not significantly (P = 0.682) associated with the composition of microbial communities. In conclusion, calf microbial communities were dissimilar to that of their dam at birth. Additionally, microbial community transition was evident with each subsequent sampling time point, potentially influenced by the presence of the dam.

PSLBI-13 Characterizing the seminal microbiota in mature rams managed on divergent planes of nutrition, and in their male offspring

Abstract Increasing evidence suggests that the bovine semen harbors commensal microbiota, and it may be important in male fertility. However, much is unknown regarding the seminal microbiota, its evolution and factors shaping this community in sheep. The objective of this study was to evaluate whether managing the rams on divergent planes of nutrition can influence the seminal microbiota in mature rams and their male offspring. For this, Rambouillet rams (n = 24; 1.5 to 4 yr old) were randomly assigned to one of three groups: 1) a positive plane of nutrition [target 12% gain in their initial body weight (BW; POS, n = 8); 2) a maintenance [maintain the initial BW (MNT, n = 8)]; and 3) a negative [target a 12% reduction in BW (NEG; n = 8)] plane of nutrition over 84 d. The rams were individually housed and fed a common diet, and feed allocations were adjusted weekly. Following the 84-d feeding period, these rams were used to breed 240 mature Rambouillet ewes (1:10 ram:ewe ratio) over 28 d of breeding. After lambing, the male lambs were maintained on a similar diet for 124 d. Semen samples from the mature rams (F0 generation) were collected on d 28, 56, and 84, and semen samples from offspring rams (F1 generation) were collected at 330 d of age via electroejaculation. Genomic DNA was extracted from semen samples and the seminal microbiota characterized using 16S rRNA gene (V3-V4) sequencing. Several environmental and negative extraction controls were also sequenced to assess potential contaminants in these samples. Overall community structure of the seminal microbiota in F0 rams was not affected by the divergent planes of nutrition (PERMANOVA: P ≥ 0.05) or by sampling time (P &amp;gt; 0.05). Microbial richness (observed ASVs) was also not different (P &amp;gt; 0.05) between the three groups of F0 rams at any of the sampling time points; however, microbial diversity (P &amp;lt; 0.05) in the F0 rams was influenced by sire dietary treatment. Microbial community structure, richness and diversity did not differ among the POS, MNT and NEG F1 generation rams (P &amp;gt; 0.05). Actinobacteriota, Firmicutes, Bacteroidota, Proteobacteria and Fusobacteriota were the most abundant phyla. Only Bacteroidota abundance was significantly (P &amp;lt; 0.05) different between three F0 groups, with POS rams had greater (18.6%) of this phylum on d 28 as compared with MNT (2.1%) and NEG (5.1%). The predominant genera identified included Fastidiosipila, Corynebacterium, Trueperella, Arthrobacter, Dietzia, Bifidobacterium, Streptobacillus, Ornithinimicrobium, and Porphyromonas, and their relative abundance was not affected (P &amp;gt; 0.05) by the dietary treatment in F0 rams. Overall, rams managed on positive and negative planes of nutrition harbored similar seminal microbiota to control rams, and divergent nutrition also had no effect on the offspring ram seminal microbiota.

208 Assessment of a new oral neonatal supplement for lambs

Abstract The objective of this study was to evaluate an easy to administer and economical oral nutritional supplement for neonatal lambs. Neonates are at risk of nutrient deficiencies because of maternal deficiencies, minimal body stores at birth and decreased nutritional intake. The pre-ruminant lamb relies solely on milk for adequate immunity and nutrition until they begin ingesting forages. Insufficiencies can lead to increased mortality and morbidity. However, deficiencies in the ewe are common due to variation in management, feeding and health protocols. A total of 10 lambs (5 sets of twins) were enrolled into the study. The lambs were divided into two groups (one twin from each set) received a treatment of an oral vitamin-mineral supplement (VitaFerst-Care) 3 mL and the control lambs a saline solution of 3 mL, all at 3 d of age (d0). Blood samples were collected, prior to, and again following supplementation (d 0, d 21) by jugular venipuncture. Vitamin A, vitamin E, iron (Fe) and selenium (Se) concentrations were determined in blood serum/plasma of the lambs by HPLC. Kidney and liver function markers were analyzed using HESKA DC5X veterinary analyzer to confirm the safety of the supplement. Variables tested included alkaline phosphatase (ALP), alanine aminotransferase (ALT), blood urea nitrogen (BUN), creatinine (CREA), glucose (GLU), total protein (TP), total bilirubin (T-BILI), albumin (ALB), phosphorus (PHOS), calcium (CA), cholesterol (CHOL), and gamma-glutamyl transferase (GGT). All lambs remained with their mothers. The average difference was calculated between d 21 and d 0 (baseline) for treated versus control animals for vitamin A, vitamin E, Se, and Fe. Significance was calculated using a two-tailed student’s t-test where P &amp;lt; 0.10 between d 21 control and treated animals. Control animals had an average vitamin E plasma concentration of 1.168 + 0.355 ppm on d 21, whereas treated animals had an average of 1.547 + 0.559 ppm. Significance was calculated at P = 0.09. Vitamin A, Fe and Se concentrations did not reach significance. Kidney and liver panel checks for animals in each group were not concerning. Future studies should look at modifying first and second sampling timepoints to be closer together. Based on results in calves with a similar study design, shortening the time between treatment and second sampling would give a clearer picture of how the neonatal supplement influenced the nutritional status of the treatment versus control animals.

Urine-derived stem cells serve as a robust platform for generating native or engineered extracellular vesicles

BackgroundMesenchymal stromal cell (MSC) therapy holds great potential yet efficacy and safety concerns with cell therapy persist. The beneficial effects of MSCs are often attributed to their secretome that includes extracellular vesicles (EVs). EVs carry biologically active molecules, protected by a lipid bilayer. However, several barriers hinder large-scale MSC EV production. A serum-free culturing approach is preferred for producing clinical-grade MSC-derived EVs but this can affect both yield and purity. Consequently, new strategies have been explored, including genetically engineering MSCs to alter EV compositions to enhance potency, increase circulation time or mediate targeting. However, efficient transfection of MSCs is challenging. Typical sources of MSC include adipose tissue and bone marrow, which both require invasive extraction procedures. Here, we investigate the use of urine-derived stem cells (USCs) as a non-invasive and inexhaustible source of MSCs for EV production.MethodsWe isolated, expanded, and characterized urine-derived stem cells (USCs) harvested from eight healthy donors at three different time points during the day. We evaluated the number of clones per urination, proliferation capacity and conducted flow cytometry to establish expression of surface markers. EVs were produced in chemically defined media and characterized. PEI/DNA transfection was used to genetically engineer USCs using transposon technology.ResultsThere were no differences between time points for clone number, doubling time or viability. USCs showed immunophenotypic characteristics of MSCs, such as expression of CD73, CD90 and CD105, with no difference at the assessed time points, however, male donors had reduced CD73 + cells. Expanded USCs were incubated without growth factors or serum for 72 h without a loss in viability and EVs were isolated. USCs were transfected with high efficiency and after 10 days of selection, pure engineered cell cultures were established.ConclusionsIsolation and expansion of MSCs from urine is non-invasive, robust, and without apparent sex-related differences. The sampling time point did not affect any measured markers or USC isolation potential. USCs offer an attractive production platform for EVs, both native and engineered.

Temporal stability of the rumen microbiome and its longitudinal associations with performance traits in beef cattle

The rumen microbiome is the focus of a growing body of research, mostly based on investigation of rumen fluid samples collected once from each animal. Exploring the temporal stability of rumen microbiome profiles is imperative, as it enables evaluating the reliability of findings obtained through single-timepoint sampling. We explored the temporal stability of rumen microbiomes considering taxonomic and functional aspects across the 7-month growing-finishing phase spanning 6 timepoints. We identified a temporally stable core microbiome, encompassing 515 microbial genera (e.g., Methanobacterium) and 417 microbial KEGG genes (e.g., K00856—adenosine kinase). The temporally stable core microbiome profiles collected from all timepoints were strongly associated with production traits with substantial economic and environmental impact (e.g., average daily gain, daily feed intake, and methane emissions); 515 microbial genera explained 45–83%, and 417 microbial genes explained 44–83% of their phenotypic variation. Microbiome profiles influenced by the bovine genome explained 54–87% of the genetic variation of bovine traits. Overall, our results provide evidence that the temporally stable core microbiome identified can accurately predict host performance traits at phenotypic and genetic level based on a single timepoint sample taken as early as 7 months prior to slaughter.

Nanopore sequencing provides snapshots of the genetic variation within salmonid alphavirus-3 (SAV3) during an ongoing infection in Atlantic salmon (Salmo salar) and brown trout (Salmo trutta)

Frequent RNA virus mutations raise concerns about evolving virulent variants. The purpose of this study was to investigate genetic variation in salmonid alphavirus-3 (SAV3) over the course of an experimental infection in Atlantic salmon and brown trout. Atlantic salmon and brown trout parr were infected using a cohabitation challenge, and heart samples were collected for analysis of the SAV3 genome at 2-, 4- and 8-weeks post-challenge. PCR was used to amplify eight overlapping amplicons covering 98.8% of the SAV3 genome. The amplicons were subsequently sequenced using the Nanopore platform. Nanopore sequencing identified a multitude of single nucleotide variants (SNVs) and deletions. The variation was widespread across the SAV3 genome in samples from both species. Mostly, specific SNVs were observed in single fish at some sampling time points, but two relatively frequent (i.e., major) SNVs were observed in two out of four fish within the same experimental group. Two other, less frequent (i.e., minor) SNVs only showed an increase in frequency in brown trout. Nanopore reads were de novo clustered using a 99% sequence identity threshold. For each amplicon, a number of variant clusters were observed that were defined by relatively large deletions. Nonmetric multidimensional scaling analysis integrating the cluster data for eight amplicons indicated that late in infection, SAV3 genomes isolated from brown trout had greater variation than those from Atlantic salmon. The sequencing methods and bioinformatics pipeline presented in this study provide an approach to investigate the composition of genetic diversity during viral infections.

Metals and other trace elements in plasma and feathers of seabirds breeding in Svalbard

We assessed the concentrations of metals and other trace elements in two of the most common seabird species breeding on Svalbard, the black-legged kittiwake (Rissa tridactyla) and the Brünnich's guillemot (Uria lomvia). Both of these species feed mostly on fish and crustaceans but have different foraging strategies, kittiwakes being surface feeders while guillemots are divers. We investigated the concentrations of arsenic (As), cadmium (Cd), chromium (Cr), copper (Cu), mercury (Hg), lead (Pb), selenium (Se) and zinc (Zn) in the plasma and body feathers of black-legged kittiwakes (n = 17), as well as in the body feathers of Brünnich's guillemots (n = 13). Samples were collected from adult birds at two time points, one week apart during July 2017 in Kongsfjorden, Svalbard. Of the non-essential trace elements, As was found at the highest median concentration at both the first (56.23 ng/g ww) and second (39.99 ng/g ww) sampling timepoints in the kittiwake plasma. When separating for the sexes, as well as sampling time, males sampled at the first sampling time point had significantly higher concentrations of As (median at 0.087 ng/g versus 0.039 ng/g) and Se (median 0.26 ng/g versus 0.16 ng/g) compared to males sampled at the second time point. There was no significant difference in plasma concentrations between females at first and second sampling time points. Kittiwake feathers contained significantly higher concentrations of As, Cd and Hg than guillemot feathers, while guillemot feathers had significantly higher concentrations of Cu, Pb and Zn. However, of the non-essential elements in both kittiwake and guillemot feathers Hg was found with the highest median concentrations at 5160 and 1080 ng/g, respectively, thus in kittiwakes exceeding the level of 5000 ng/g associated with adverse effect (e.g., impaired reproduction). Levels of Hg and Se found in the kittiwake feathers were higher than previous studies on seabirds in the Arctic.

The impact of sampling time point on the lipidome composition

Lipidomic profiling has been reported as an effective approach for characterizing and differentiating brain tumors. However, since lipids can undergo non-specific enzymatic and nonenzymatic reactions due to tissue disruption, it is critical to consider the preanalytical phase of the diagnostic process (e.g., optimizing the sampling time and sampling conditions). Thus, this study assesses the ways in which the time point of sampling impacts the lipidome composition of brain tumors. Two histologically distinct brain tumors—namely, meningiomas and gliomas—were sampled using solid-phase microextraction (SPME) fibers at two time points: on-site directly after removal, and after 12 months of storage at −30 °C. The samples were analyzed via HILIC chromatography coupled with HRMS, which enabled the detection of a wide range of features, including phospholipids and sphingolipids, as well as changes in the profiles of these compounds. The samples obtained from the stored tissues tended to have elevated levels of analytes with lower m/z values. In addition, the samples obtained from the fresh and stored tissues were easily distinguished based on their lipidome compositions, regardless of the histological tumor type. Notably, while storage did not affect the possibility of differentiating meningiomas and gliomas, the biological interpretation of the obtained results were prone to bias.