Manufacturer's Guidelines Research Articles

Clinical performance of polyethylenefiber reinforced resin composite restorations in endodontically treated teeth: (a randomized controlled clinical trial)

AimThe purpose of this study was to evaluate the performance of polyethylene fiber reinforced resin composite fillings compared to bulk fill resin composite fillings in endodontically treated teeth over a two-year monitoring period.MethodA total of 240 individuals with endodontically treated lower molars and a moderate amount of tooth structure were divided into two equal groups of 120 each. One group received polyethylene fiber reinforced bulk fill resin composite restorations while the other group received only bulk fill resin composite restorations, both applied as per manufacturer guidelines. Two proficient experienced blinded assessors assessed the restorations using modified USPHS criteria at baseline, 6, 12, and 24 months.Statistical analysis usedData analysis involved the utilization of Mann-Whitney U, Friedman’s test, and Nemenyi post hoc test, with age data being displayed as mean and standard deviation. The significance level was established as p < 0.05, and R software was utilized for statistical analysis.ResultsThere were no notable distinctions in any parameters or scores between the intervention and comparator groups at various time points. Alpha scores were present for retention, gross fracture, and secondary caries at all follow up intervals.ConclusionBoth direct resin composite restorations reinforced with polyethylene fibers and direct bulk fill resin composite restorations placed in endodontically treated molars with moderate remaining tooth structure demonstrated satisfactory clinical outcomes during a 24-month follow-up period.Clinical relevanceBulk fill resin composites directly placed in endodontically treated molars with moderate remaining tooth structure showed promise as a treatment option over a two-year period.Clinical trial registration(06-01-2022) on https://ClinicalTrials.gov with the ID (NCT05180903).

Impact of UV Light Exposure During Printing on Thermomechanical Properties of 3D-Printed Polyurethane-Based Orthodontic Aligners

Aim: Polyurethane-based aligners, created through photoinitiated free-radical polymerization, have been the subject of numerous studies focusing solely on their mechanical properties. In contrast, we investigate their thermomechanical properties, which are crucial for their efficacy. This paper aims to investigate the effects of different UV light exposure durations on the complex modulus of elasticity, tan delta, glass transition temperature, and the degree of conversion (DC). Methods: Aligners were printed using Tera Harz TC-85 and NextDent Ortho Flex resin with specific exposure times (2, 2.4, 3, 4, and 4.5 s for Tera Harz; 5, 6, 7, and 8 s for NextDent) and processed per manufacturer guidelines. The degree of conversion was analyzed using Attenuated Total Reflectance Fourier Transform Infrared (ATR-FTIR) spectroscopy, while Dynamic Mechanical Analysis (DMA) characterized the mechanical properties (complex modulus and tan delta) and the glass transition. Results: Tera Harz TC-85 showed a higher degree of conversion (90.29–94.54%), suggesting fewer residual monomers, which is potentially healthier for patients. However, its lower glass transition temperature (35.60–38.74 °C) might cause it to become rubbery in the mouth. NextDent Orto Flex, with a higher storage modulus (641.85–794.55 MPa) and Tg (49.36–50.98 °C), offers greater rigidity and stability at higher temperatures (greater than temperature in the oral cavity), ideal for orthodontic forces, though its lower degree of conversion raises health concerns. Conclusions: Tera Harz TC 85 generally achieves higher DC and more stable polymerization across different UV exposure times than NextDent Orto Flex. Optimal polymerization times significantly impact both the mechanical and thermal properties of these dental resins, with NextDent showing optimal properties at 7 s and Tera Harz benefiting from both very short and extended exposure times.

Enhancing Printability Through Design Feature Analysis for 3D Food Printing Process Optimization

We present a novel, systematic method for evaluating design printability in 3D food printing using a scoring system based on the Design for Additive Manufacturing (DfAM) guidelines. This study addresses a gap in the current literature by proposing a structured approach to assess and enhance the printability of 3D food designs. Our framework consists of a set of nine critical questions derived from the multi-level DfAM guidelines, focusing on key printability factors including unsupported features, geometric accuracy, and surface finish. The evaluation process converts qualitative assessments into numerical values, resulting in a comprehensive printability score that categorizes designs into high, moderate, or low printability levels. To validate the effectiveness of this method, we conducted a case study involving five different designs. The scoring system successfully explores the design space and maximizes the printability of 3D food products. This method alleviates the challenges in design evaluation compared with traditional trial-and-error approaches. The results demonstrate the practicality and efficiency of our framework’s output. The proposed methodology provides a structured approach to design evaluation, offering practical insights and a valuable tool for improving the success rate of 3D printed food products. This research contributes to the field by offering a systematic framework for assessing and enhancing the printability of 3D food designs, potentially accelerating the adoption and effectiveness of 3D food printing technology in various applications.

Remineralizing capacity of zinc oxide eugenol sealer following the addition of nanohydroxyapatite-tyrosine amino acid: An in vivo animal study

ObjectiveAlthough several sealers have been developed, the zinc oxide eugenol (ZOE) sealer is still used in many private dental clinics. This study aimed to compare the biocompatibility and remineralizing capacity οf ZOE sealer with the addition of nanohydroxyapatite-tyrosine amino acid. MethodsThis study was conducted on Twenty rabbits. The rabbits were divided into four groups based on the test observation period (3, 7, 21, and 28 days) following surgical implantation. General anesthesia was administered to each rabbit, and a subcutaneous incision of approximately 1±0.5 cm was made along the symphyseal area of the mandible of each rabbit. Four bone cavities were generated in the interdental space of the lower jaw between the central and molar teeth of each rabbit, with one longitudinal subcutaneous incision. The ZOE sealers were mixed according to the manufacturer's guidelines and directly inserted within the cavities generated at the end of each test period. The animals were sacrificed, and bone biopsy was carried out at the site of testing. The biopsy samples were obtained and subjected to histological analysis using a low-power light microscope (Olympus C ⅹ 21, Japan) and immunohistochemistry using Ki67 antibody. ResultsThe collected information was examined by parametric statistical tests using SPSS software version ‘‘22.” One-way ANOVA and post-hoc Duncan’s tests were used to measure the significance among various groups, with statistical significance set, when “P≤0.01”. ConclusionThe 20% mixed endodontic sealer displayed excellent outcomes compared to other experimental groups, as identified by higher new bone formation at every evaluation period.

Periodic phenotypic profile analysis of Acinetobacter baumannii drug resistance characteristics showing the emergence of PDR.

Acinetobacter baumannii, one of the ESKAPE pathogens, is on the World Health Organization (WHO) list of priorities needing urgent new effective antimicrobial agents due to exhibited high resistance by the bacterium to currently available antibiotics. This study examines the periodic changes of clinical A. baumannii isolates and their antimicrobial resistance patterns and types in Southeastern region of Saudi Arabia. One hundred and seventy-seven randomly selected A. baumannii isolates were used for the investigation with bacterial identities (IDs) and antimicrobial assay ascertained with Gram-negative (GN) ID cards and antimicrobial susceptibility test (AST) cards of Vitek 2 Compact Automated System according to manufacturer's guidelines. Descriptive phenotypic types of isolates were compared using comparison proportion and Fisher extraction test, while Morpheus versatile matrix visualization and analysis software was used for the dendrogram of hierarchical clustering. A significantly higher proportion of samples were from males compared to females (p = 0.025), with 33.33% of samples originating from patients aged 51-70 years. Resistance was high for imipenem (93%), meropenem (94%), levofloxacin, ciprofloxacin (99%), and aztreonam (98%). There was less percentage resistance to colistin (18%), tigecycline (23%), and minocycline (23%). Multidrug resistance (MDR)/carbapenem-resistant Acinetobacter baumannii (CRAB) was observed consistently across all years. There was no extensive drug resistance (XDR) among isolates from 2013 to 2014, but it was present in the 2016 to 2018 and 2019 to 2020 periods, while pandrug resistance was seen only in the 2019 to 2020 isolates. The study shows a clear trend of the isolates changing from MDR to XDR and then to pandrug resistance over the study period. Also, it indicates that carbapenems might no longer be a treatment choice in this study region. Although colistin exhibited less resistance, the toxicity of the drug reduces its usefulness. The development of pandrug resistance is a critical concern.

An improved high-resolution method for quantitative separation of empty and filled AAV8 capsids by strong anion exchange HPLC.

Cell and gene therapy (CGT) is a field of therapeutic medicine that aims to treat, prevent, and cure diseases using engineered cells (stem cells, immune cells, and differentiated adult or fetal cells), vectors [Adeno Associated Virus (AAV), Adeno Virus (AV), Herpes Simplex Virus (HSV), Baculo Virus (BV), Lenti Virus (LV), Retro Virus (RV), etc.], and other carriers [non-viral vectors, virus-like particles (VLP), Lipid Nano-Particles (LNP), etc.]. Among viral CGT vectors, adeno-associated viruses and lentiviruses (AAV and LV) are the most widely applied vector platforms. The presence of non-functional (empty or non-infectious) vectors that carry null or partial genes in the final drug product is classified as an impurity by the FDA. These impurities impair dosage accuracy and induce non-specific immunogenicity and variability in drug efficacy. These non-functional viral vectors in the drug product need to be elucidated following International Conference on Harmonization (ICH) guidelines for clinical manufacturing of the final drug product. This article showcases an ion-exchange chromatography (IEX) high-resolution method supporting ICH guidelines using commercially available AAV8 filled and empty capsids as reference standards. Our method successfully separated empty to full capsids with a resolution of 15 and sustained a linearity greater than 0.98 even under a wide range of empty or full viral particle concentrations (E+9 to E+13 vp/mL), which is an upgrade to other IEX capsid separation methods. The medium-throughput capacity and shorter sample processing time improve testing efficiency and save costs while delivering quality as value. The discussed method is a reliable and reproducible platform to precisely evaluate the presence of non-functional viral particles in AAV8 samples. Aligned with other orthogonal results, the method is a powerful tool to improve the quality of rAAV analytics.

A clinical investigation of force plate drift error on predicted joint kinetics during gait

Inverse dynamic analysis is a technique used during gait analysis to estimate intersegmental forces and net joint moments. Inverse dynamic calculations are susceptible to various forms of error. One such error is force plate drift, often produced by humidity condensing within the input connectors and electronics, causing an undesired change in output over time. This can be particularly concerning for movement laboratories where inverse dynamics are considered in clinical decision-making processes. Manufacturers will provide tolerance levels for drift. However, levels of acceptable drift are rarely considered from a clinical perspective. Therefore, this study aims to establish clinically acceptable limits of force plate drift error, induced by applying systematic errors to force plate channels, on predicted lower limb joint moments during gait. Gait data of 10 children with typical development were analysed and induced errors of 0.5 N, 1 N, 1.5 N, 3 N, 6 N and 12 N were incrementally applied to the horizontal and vertical force channels. Data were recalculated for each increment and mean profiles compared to an error free mean (±1SD) band. Error was deemed clinically significant when moments fell outside the mean (±1SD) band. Induced error at 6 N and above was sufficient to cause a clinically significant change. Sagittal and coronal plane moments at the hip were most affected, followed by the knee and then the ankle. While manufacturer guidelines for acceptable drift are usually well below 6 N, care is needed when using force plates over several minutes or more as drift may eventually exceed clinically acceptable limits.

Mesenchymal Stem Cell-Derived Exosomes as a Neuroregeneration Treatment for Alzheimer's Disease.

Alzheimer's disease (AD) is the most prevalent kind of dementia and is a long-term degenerative disease. Pathologically, it is defined by the development of extracellular amyloid-β plaques and intracellular neurofibrillary tangles made up of hyperphosphorylated tau protein. This causes neuronal death, particularly in the hippocampus and cortex. Mesenchymal stem cell (MSC)-derived exosomes have been identified as possibly therapeutic and have promise for Alzheimer's disease due to their regenerative characteristics. A systematic retrieval of information was performed on PubMed. A total of 60 articles were found in a search on mesenchymal stem cells, exosomes, and Alzheimer's disease. A total of 16 ongoing clinical trials were searched and added from clinicaltrials.gov. We added 23 supporting articles to help provide information for certain sections. In total, we included 99 articles in this manuscript: 50 are review articles, 13 are preclinical studies, 16 are clinical studies, 16 are ongoing clinical trials, and 4 are observational studies. Appropriate studies were isolated, and important information from each of them was understood and entered into a database from which the information was used in this article. The clinical trials on mesenchymal stem cell exosomes for Alzheimer's disease were searched on clinicaltrials.gov. Several experimental investigations have shown that MSC-Exo improves cognitive impairment in rats. In this review paper, we summarized existing understanding regarding the molecular and cellular pathways behind MSC-Exo-based cognitive function restoration, with a focus on MSC-Exo's therapeutic potential in the treatment of Alzheimer's disease. AD is a significant health issue in our culture and is linked to several important neuropathological characteristics. Exosomes generated from stem cells, such as mesenchymal stem cells (MSCs) or neural stem cells (NSCs), have been examined more and more in a variety of AD models, indicating that they may be viable therapeutic agents for the treatment of diverse disorders. Exosome yields may be increased, and their therapeutic efficacy can be improved using a range of tailored techniques and culture conditions. It is necessary to provide standardized guidelines for exosome manufacture to carry out excellent preclinical and clinical research.

Effect of Surface Treatments Applied to 3D Printed Permanent Resins on Shear Bond Strength.

3D-printed permanent resins have recently been introduced to produce permanent restorations. The aim of this study is to evaluate the effect of surface treatment methods on the shear-bond strength (SBS) between 3D-printed permanent resins and adhesive cement. In this study, samples were produced using digital light projection (DLP) and stereolithography (SLA) 3D printers with two permanent resins (Crowntec, Saremco and Permanent Crown, Formlabs) in accordance with manufacturer guidelines. The samples were separated into three groups: sandblasting, hydrofluoric acid and no surface treatment. The surface profile (Ra, Sa) of the samples was examined with a confocal microscope (Smartproof 5, Zeiss). Then, a self-adhesive resin cement was applied to the samples in a transparent mold (2.38 mm diameter) in accordance with ISO 29022:2013. A universal testing machine was used to perform SBS test. A stereomicroscope was used to analyze the different types of fractures. A two-way analysis of variance (ANOVA) test was used to assess the data (p<0.05). The samples with sandblasting applied to the surface showed higher surface roughness values than the samples with hydrofluoric acid (p<0.001). 3D printer technology and surface treatment methods affected SBS values (p<0.001). Sandblasting groups higher SBS values were than in the hydrofluoric acid group (p<0.001). Sandblasting revealed cohesion fractures, which indicated a stronger bond, while hydrofluoric acid displayed adhesive and mix fractures. When sandblasting was applied to the surface of the samples prepared using permanent resins, higher adhesion was achieved with adhesive cement. Key words:3D printing, Shear bond strength, Sandblasting, Stereolithography.

Enhancing the antibacterial and surface hardness of glass ionomer cement modified with Salvadora persica and Chlorhexidine: An in vitro study.

This laboratory study evaluated the effect of Salvadora persica (S. persica) root extracts and Chlorhexidine Digluconate (CHX) on the antibacterial and surface hardness properties of glass ionomer cement (GIC). The in vitro experimental study was conducted at the Baqai Institute of Pharmaceutical Sciences of Baqai Medical University, Karachi, Pakistan, from October 2022 to March 2023. There were a total four experimental groups. The first group consisted of ethanol extract (GIC-SPEE) and second group consisted of hexane extract as (GIC-SPHE) both prepared from Salvadora persica root respectively, and mixed with liquid of GIC separately. The third group comprised chlorhexidine (GIC-CHX) that was also mixing into liquid portion of GIC and the last group was Control i.e. (cGIC). The GIC samples were prepared by using stainless steel metallic moulds with dimension (5mm x 2mm), following the manufacturer guidelines. Antibacterial activity against Streptococcus mutans was done by disc diffusion test (DDT), and surface hardness test was done by Vickers hardness tester. Statistical analysis was performed using One-Way ANOVA and Tukey's post hoc tests (p<0.05). The antibacterial activity against S. mutans reported that the maximum zone of inhibition was obtained at 3 wt% by the GIC-SPEE, when compared with other experimental groups. For surface hardness, the highest mean and standard deviation and significant findings was reported by the group GIC-SPEE. Considering the outcome of this in vitro study, it can be concluded that the addition of 3 wt% GIC-SPEE increased the surface hardness and antibacterial activity against Streptococcus mutans.

USE OF SOLENSIA (FRUNEVETMAB) IN NONDOMESTIC FELIDS (PANTHERA TIGRIS, LYNX RUFUS, PANTHERA UNCIA) GUIDED BY PROTEIN SEQUENCE ANALYSIS.

Osteoarthritis and degenerative joint disease affect many species of nondomestic felid and negatively impact quality of life in managed care settings. Previously, pain management options were limited because of the frequency of comorbidities such as renal disease and common difficulty encountered in medication administration. SolensiaTM (frunevetmab) is a felinized monoclonal antibody that binds to nerve growth factor in arthritic joints, thereby inhibiting pain response cascades, and produces marked improvement in clinical signs and quality of life in domestic cats diagnosed with osteoarthritis. Protein sequence analysis was performed to inform the application of frunevetmab in four individuals from three nondomestic felid species (Panthera tigris, Lynx rufus, and Panthera uncia) diagnosed with degenerative joint disease to predict safety, contraindications, and likely response to treatment. Patients were then treated utilizing doses extrapolated from domestic cats and following manufacturer guidelines for administration, and clinical response was evaluated over a minimum 2-mon period. Significant improvement was noted in clinical signs in all four animals, resulting in marked improvement in mobility, lameness, activity level, demeanor, and overall quality of life. Frunevetmab presents an excellent adjunctive therapeutic alternative for nondomestic felids suffering from degenerative joint disease and may complement or decrease the need for other conventional therapeutic regimens.

Effects of Bacillus subtilis on Growth Performance, Metabolic Profile, and Health Status in Dairy Calves.

This study focused on assessing whether the inclusion of probiotics (B. subtilis) as feed additives during the preweaning stage can enhance the body weight and metabolic condition of neonatal calves. A total of 50 Holstein calves, all born on the same farm, were randomly divided into two homogeneous treatment groups after birth. The calves in the control group (CG) were fed a milk replacer (n = 25) (13 females and 12 males) and those in the B. subtilis-supplement-treated group (TG), (n = 25) (13 females and 12 males) were fed a milk replacer with 7.5 mL/calf/day of B. subtilis probiotic (complied with the manufacturer's guidelines). The probiotic was administered 24 h post-birth, signifying the start of the experimental period. It took one month to collect the animals. Body weight was measured at birth for all animals. A local veterinarian, working on the farm, conducted daily health checks of the calves, recording health parameters and any antibiotic treatments. Blood samples were collected from each calf at birth and 30, 60, and 90 days by puncturing the jugular vein using 10 mL evacuated serum tubes before morning feeding. Significant differences in body weight were observed between the CG and the TG at 30, 60, and 90 days of age. At 30 days, the TG had a 4.11% higher average body weight than the CG (54.38 kg vs. 52.71 kg). At 60 days, the TG's average weight was 3.75% higher (79.21 kg vs. 76.34 kg), and at 90 days, the TG had a 2.91% higher average weight (112.87 kg vs. 109.67 kg). At 30 days of age, the TG showed significantly lower AST activity, with a 41.12% decrease compared to the CG (51.02 IU/L vs. 72.00 IU/L). Conversely, GGT activity was significantly higher in the TG by 64.68% (40.64 IU/L vs. 14.35 IU/L). Phosphorus concentration at 30 days was also significantly higher in the TG by 9.36% (3.27 mmol/L vs. 2.99 mmol/L). Additionally, the TG had a significantly lower total protein concentration, with a 21.63% decrease at 30 days (46.32 g/L vs. 56.34 g/L) and a 20.28% decrease at 60 days (48.32 g/L vs. 58.12 g/L) compared to the CG. These findings indicate that dairy calves given conventional milk replacer along with a daily dose of 7.5 mL of B. subtilis probiotic experienced enhanced growth performance and a more favourable metabolic profile during the first 90 days of their lives.

Interdisciplinary Delphi study by PROSEC North America: Recommendations on single indirect restorations made from ceramic and nonmetallic biomaterials for posterior teeth.

This article puts forward consensus recommendations from PROSEC North America regarding single indirect restorations made from ceramic and nonmetallic biomaterials in posterior teeth. The consensus process involved a multidisciplinary panel and three consensus workshops. A systematic literature review was conducted across five databases to gather evidence. The recommendations, informed by findings from systematic reviews and formulated based on a two-phase e-Delphi survey, emphasize a comprehensive treatment strategy that includes noninvasive measures alongside restorative interventions for managing dental caries and tooth wear. The recommendations advocate for selecting between direct and indirect restorations on a case-by-case basis, favoring inlays and onlays over crowns to align with minimally invasive dentistry principles. The recommendations highlight the critical role of selecting restorative biomaterials based on clinical performance, esthetic properties, and adherence to manufacturer guidelines. They emphasize the importance of precision in restorative procedures, including tooth preparation, impression taking, contamination control, and luting. Regular follow-up and maintenance tailored to individual patient needs are crucial for the longevity of ceramic and nonmetallic restorations. These PROSEC recommendations provide a framework for dental practitioners to deliver high-quality restorative care, advocating for personalized treatment planning and minimally invasive approaches to optimize oral health outcomes. The PROSEC North America recommendations highlight the importance of minimally invasive techniques in posterior tooth restorations using ceramic and non-metallic biomaterials. These principles prioritize tooth structure conservation and personalized treatment planning, essential for enhancing clinical outcomes and long-term oral health.

Exploring the Impacts of Green Supply Chain Integration and Ambidextrous Green Innovation on the Financial Performance of China’s Pharmaceutical Manufacturing Enterprises

This study focuses on the factors affecting the financial performance of pharmaceutical manufacturing enterprises. Based on dynamic capability theory and ambidextrous innovation theory, this study adopts the Resource–Behavior–Performance framework to investigate how green supply chain integration (GSCI) and ambidextrous green innovation affect financial performance. The proposed hypotheses were tested through structural equation modeling using data from 400 China’s pharmaceutical manufacturing enterprises. The results indicate that the various dimensions of GSCI and ambidextrous green innovation yield distinct outcomes. Among the three dimensions of GSCI, only green supplier integration and green customer integration significantly impact financial performance directly, unlike green internal integration. Among the two dimensions of ambidextrous green innovation, only exploitative green innovation significantly influences financial performance, acting as a mediator between the GSCI dimensions and financial performance, while exploratory green innovation does not. The main advantages of this study include considering the connotation and value of GSCI from a green dynamic capability perspective, as well as the effects of exploratory and exploitative green innovation as intermediary behaviors while simultaneously considering the performance effects of GSCI and ambidextrous green innovation. This study offers novel academic insights and practical guidelines for pharmaceutical manufacturing enterprises to integrate GSCI and ambidextrous green innovation, with the aim of achieving better financial performance in their sustainable development efforts.

What About Water? Implications for Body Composition Assessment in Military Personnel.

Sergi, TE, Roberts, BM, and Heileson, JL. What About Water? Implications for Body Composition Assessment in Military Personnel. J Strength Cond Res 38(9): e534-e540, 2024-Body composition standards ensure service members maintain physical fitness, wellness, and support mission readiness. Anthropometric techniques (i.e., height/mass, circumference-based "tape test") have been the primary screening and percent body fat (%BF) assessment method in military personnel for about 4 decades. Recently, the Army and Marine Corps have implemented more advanced body composition assessment methods, such as air displacement plethysmography (ADP), multifrequency bioelectrical impedance analysis (MF-BIA), and dual-energy x-ray absorptiometry (DXA), to serve as supplemental %BF assessment after failing the tape test. Although supplemental assessments are intended to improve on the accuracy and precision of the tape test, preassessment standardization, specifically regarding acute water ingestion (AWI), is lacking. Thus, the purpose of this narrative review was to (a) summarize the available literature regarding the influence of AWI on body composition estimates derived from ADP, MF-BIA, and DXA and (b) provide evidence-based recommendations for researchers and practitioners. Studies indicate that AWI increases %BF estimates with ADP (4 of 6 [4/6] observations) and MF-BIA (6/7), whereas AWI increases muscle mass (6/6) and likely decreases %BF (2/3) when obtained by DXA. In conclusion, ADP, MF-BIA, and DXA are susceptible to confounding from AWI, leading to inaccurate body composition estimates that may negatively affect the careers of military personnel. Based on the findings from this narrative review, military practitioners and researchers should (a) follow manufacturer guidelines for calorie intake [food and fluid] and exercise avoidance, (b) conduct urine-specific gravity testing [if possible], and (c) limit AWI to <250 ml before assessment.

Tumour-induced alterations in single-nucleus transcriptome of atrophying muscles indicate enhanced protein degradation and reduced oxidative metabolism.

Tumour-induced skeletal muscle wasting in the context of cancer cachexia is a condition with profound implications for patient survival. The loss of muscle mass is a significant clinical obstacle and is linked to reduced tolerance to chemotherapy and increased frailty. Understanding the molecular mechanisms driving muscle atrophy is crucial for the design of new therapeutics. Lewis lung carcinoma tumours were utilized to induce cachexia and muscle atrophy in mice. Single-nucleus libraries of the tibialis anterior (TA) muscle from tumour-bearing mice and their non-tumour-bearing controls were constructed using 10X Genomics applications following the manufacturer's guidelines. RNA sequencing results were analysed with Cell Ranger software and the Seurat R package. Oxygen consumption of mitochondria isolated from TA muscle was measured using an Oroboros O2k-FluoRespirometer. Mouse primary myotubes were treated with a recombinant ectodysplasin A2 (EDA-A2) protein to activate EDA-A2 receptor (EDA2R) signalling and study changes in gene expression and oxygen consumption. Tumour-bearing mice were sacrificed while exhibiting moderate cachexia. Average TA muscle weight was reduced by 11% (P=0.0207) in these mice. A total of 12335 nuclei, comprising 6422 nuclei from the control group and 5892 nuclei from atrophying muscles, were studied. The analysis of single-nucleus transcriptomes identified distinct myonuclear gene signatures and a shift towards type IIb myonuclei. Muscle atrophy-related genes, including Atrogin1, MuRF1 and Eda2r, were upregulated in these myonuclei, emphasizing their crucial roles in muscle wasting. Gene set enrichment analysis demonstrated that EDA2R activation and tumour inoculation led to similar expression patterns in muscle cells, including the stimulation of nuclear factor-kappa B, Janus kinase-signal transducer and activator of transcription and transforming growth factor-beta pathways and the suppression of myogenesis and oxidative phosphorylation. Muscle oxidative metabolism was suppressed by both tumours and EDA2R activation. This study identified tumour-induced transcriptional changes in muscle tissue at single-nucleus resolution and highlighted the negative impact of tumours on oxidative metabolism. These findings contribute to a deeper understanding of the molecular mechanisms underlying muscle wasting.

P-535 Effect of advanced female age (AFA) on the ratio of patients receiving mosaic embryo transfers

Abstract Study question How does female age affect ratio of patients with at least one euploid embryo (≥1EE) and at least one mosaic embryo without any euploid ones(≥1ME)? Summary answer Percentage of patients receiving mosaic embryo transfers increases with female age. What is known already Introduction of Next Generation Sequencing (NGS) to the area of Preimplantation Genetic Testing for Aneuploidy (PGT-A) has enabled detection of mosaicism in human embryos. Studies reporting the birth of healthy babies after the transfer of embryos with a chromosomal mosaic result on PGT-A have been published (Greco et al., 2015; Kahraman et al., 2020; Viotti et al., 2021 and 2023). Data suggested lower implantation rates and higher miscarriage rates when mosaics were compared with euploid embryo transfers. The percentage of mosaic embryos in overall tested embryos decreases with increased maternal age, similar to euploid embryos’ decline (Sanders et al., 2023). Study design, size, duration This retrospective study includes 7287 cycles with PGT-A performed between January 2017 and December 2023. Number of biopsied blastocysts was 21,277 and all were tested with NGS. All PGT-A testing was conducted on Ion Torrent S5 (Thermo Fisher Scientific) according to the manufacturer guidelines. The mosaic embryo reporting thresholds were determined as 20 to 80%. Participants/materials, setting, methods Euploidy ratio (ER) was defined as (euploid embryos (n)/ all tested embryos(n)). Mosaicism ratio (MR) was defined as (mosaic embryos (n)/all tested embryos(n)). Alternative mosaicism ratio (AMR) was defined as the number of mosaic embryos divided by the sum of euploid embryos and mosaic embryos. In other words, AMR can be defined as the number of mosaic embryos divided by non-aneuploid embryos. Female ages were divided into five categories: FAG1:&amp;lt;35, FAG2:35-37, FAG3:38-40, FAG4:41-42, FAG5:&amp;gt;42. Main results and the role of chance The number of cycles, total number and average biopsied embryos with standard deviations were highest in FA1 and declined with AFA: FAG1:2537, 9298, 3.7±2.2; FAG2:1348, 4140, 3.1±1.7; FAG3:1701, 4298, 2.5±1.6; FAG4: 958,2116, 2.2±1.5; FAG5:738, 2407, 1.9±1.2. ER and MR were observed to decrease with AFA: FAG1:%48.4, %15.2; FAG2:%39.0, %12.7; FAG3:%26.5, %10.5; FAG4:%14.9, %6.0; FAG5:%5.0, %2.6. This decrease was statistically significant (p &amp;lt; 0.0001, Chi-Square Test). However, when AMR was calculated among age groups, AMR was increasing with AFA: FAG1:%23.9; FAG2:%24.6; FAG3:%28.5;FAG4:%28.7; FAG5:%34.5 and this was also statistically significant (p = 0.0002, Chi-Square Test). Correlation of MR and AMR with female age groups were tested with Pearson correlation coefficient and MR was found to be negatively correlated with AFA whereas AMR was positively correlated (r=-0.19, p &amp;lt; 0.0001; r = 0.07, p &amp;lt; 0.0001, respectively). The ratio of patients with ≥1EE and ≥1ME was found to be decreasing with AFA when compared to overall tested cycles: FAG1: %83%, %7; FAG2: %69, %9; FAG3: %47, %11; FAG4: %28, %9; FAG5: %9, %4. However, the ratio of ≥ 1ME in cycles with transferrable embryos significantly increased with AFA consisting of more than one fourth of embryo transfer cycles above 38 years of age: FAG1: %7; FAG2: %12; FAG3: %19; FAG4: %24; FAG5: 31%. Limitations, reasons for caution These are the results of a single ART and Reproductive Genetics Center with its strictly defined standard operating procedures for PGT-A testing and ovarian stimulation protocols. Although not expected, different PGT-A testing methodologies or stimulation protocols may influence the above-mentioned outcomes. Wider implications of the findings The alternative mosaicism ratio defined here is generally overlooked but impacts the treatment strategies of ART patients, specifically for the advanced maternal age groups. Trial registration number not applicable

P-222 Can embryo morphokinetics act as early warning key performance indicators in relation to consumable batching

Abstract Study question Can morphokinetics be used as an early warning indicator of a batch-related effect of oil currently in use in the laboratory on implantation rate? Summary answer Where morphokinetic timings were slower for a batch of oil, implantation rate was also reduced, suggesting they could be used as an early warning system. What is known already The commercialisation of oil means that there are strict manufacturing guidelines leading to consistency between batches. However, this does not mean that minor differences do not exist. Furthermore, oil can undergo peroxidation if stored incorrectly, causing it to become toxic to embryos. The introduction of timelapse incubation has allowed extensive research into several morphokinetic parameters. This research has established time-frames within which each developmental milestone should occur to indicate the likelihood of implantation. Implantation rate (IR) is often used as a key performance indicator in the laboratory however, potential systemic issues may be identified sooner if morphokinetic parameters were used. Study design, size, duration The following morphokinetic parameters were compared to IR to determine if a batch of oil was affecting embryo development; time of division to two cells (t2), five cells (t5), eight cells (t8), time to start of blastulation (tSB) and cell synchronicity between two and eight cells (CS2-8). Data from January 2019 to October 2019 from a single centre were used, and included data from 2008 embryos and 333 patients. Participants/materials, setting, methods Data was exported from the EmbryoScope + [Vitrolife, Sweden]. Data from RIWitness Laboratory Manager [CooperSurgical, Denmark] was utilised to identify when different batches of oil (Ovoil™, Vitrolife) were used within the specified time period. The data from the EmbryoScope+ was then separated by date to signify when a new batch of oil was open. A Kruskal-Wallis test was then used to identify whether the morphokinetic timings differed between batches of oil. Main results and the role of chance There was no significant difference in patient age (p = 0.178) or the number of oocytes collected (p = 0.097) between batches of oil used. There was no significant difference between the morphokinetic timings for each batch of oil used. However, the morphokinetic timings follow the same trend line as the IR for both t2 and t8; where t2 and t8 are higher, the IR for the corresponding batch of oil is reduced. The trend can also be seen in CS2-8. The batch of oil with the lowest IR (25.56%) also had the highest mean time to t2, t8 and tSB compared to the other batches of oil. This corresponds with previous research that identified a correlation between slower embryo development and reduced IR (Meseguer et al., 2011). A clear difference could be seen between the average time taken to reach each developmental milestone and each batch of oil used, perhaps suggesting oil can have an effect, however small, on embryo development. This study suggests that morphokinetic timings could be used as an early warning indicator that a new batch of oil may be affecting development allowing a rapid response by the laboratory team and possible cessation of use pending further investigation. Limitations, reasons for caution Different batches of culture medium were also used throughout this time period and were not introduced at the same time as oil batches. Therefore, the results of this investigation cannot be associated entirely to the introduction of a different batch of oil. Wider implications of the findings This information could be incorporated into an early warning algorithm that could be assessed after a batch of oil was introduced. The use of the batch could then be ceased immediately without waiting for a decrease in IR by which time it would have been used for many more embryos. Trial registration number Not applicable

First Report of Watermelon Crinkle Leaf-Associated Virus-2 (WCLaV-2) in Watermelon (Citrullus lanatus) in Georgia, USA.

Watermelon (Citrullus lanatus) is a key horticultural crop in Georgia with farmgate value of 142 million USD (2022 Farm Gate Value Report), yet faces challenges from whitefly-transmitted viruses, especially during periods of elevated whitefly populations in the fall. Foliar symptoms on watermelon plants including yellow mottling and chlorosis, wrinkling, bunching, and upward curling, were observed in experimental fields at UGA Tifton and commercial fields in Colquitt County in the fall of 2023. These were similar to those described for watermelon crinkle leaf-associated viruses (WCLaV-1 and WCLaV-2) from Florida (Hendrick et al, 2021) and Texas (Hernandez et al., 2021). The disease incidence reached 100% in both locations. WCLaV-1 was previously identified in Georgia (Adeleke et al., 2022a); however, WCLaV-2 remained undetected in further surveys (Adeleke et al., 2022b). Total nucleic acid was extracted from symptomatic leaf tissues with the MagMAXTM 96 Viral RNA isolation kit (ThermoFisher Scientific, USA), following the manufacturer's guidelines, with the omission of DNAse treatment. The presence of WCLaV-1 was identified through reverse transcription-polymerase chain reaction (RT-PCR) (Hernandez et al., 2021) in 17 out of 24 samples in Tift County, and 12 out of 15 samples from Colquitt County. Within the same set of samples, WCLaV-2 was identified in three samples from Tift County and four samples from Colquitt County by RT-PCR directed at the RdRp gene (Hernandez et al., 2021). WCLaV-1 was not detected in the three samples from Tift County that had WCLaV-2, while mixed infections of WCLaV-2 and WCLaV-1 were observed in the samples form Colquitt County. Two whitefly transmitted viruses, previously reported in Georgia were also identified as mixed infection in these samples (Table S1). The presence of WCLaV-2 was further confirmed by amplifying the movement protein (MP) gene of WCLaV-2 by RT-PCR assays (Hernandez et al., 2021). The amplicons, with expected sizes of 968bp for the RdRp gene and 562bp for the MP gene of WCLaV-2, located on RNA 1 and RNA 2 segments respectively, were directly sequenced from both directions (Genewiz, USA) from a sample collected in Tift County. The resulting data were analyzed via BLASTn search. The MP gene fragment (PP178543) shared 100% identity with isolates from Brazil (LC636074.1), Texas (MW559086.1), and Florida (MZ325858.1). RdRp gene (PP178542) shared >99.7% identity with isolates from Brazil (LC636073.1), Texas (MW559083.1) and Florida (MZ325855.1). WCLaV-1 and WCLaV-2, initially discovered in Asia (Xin et al., 2017), have been assigned to the genus Coguvirus, in the family Phenuiviridae (Walker et al., 2022). Subsequent reports from the USA (Hendrick et al., 2021; Hernandez et al., 2021), Australia (Mulholland et al., 2023), and Brazil (Maeda et al., 2022) indicate the global spread of these viruses. Watermelon is the primary host of WCLaV-2. Despite these findings, biological information, including vector relations, for both viruses and other members of the genus Coguvirus remains elusive. The impact of these viruses on watermelon production and yield in the identified regions remains largely unknown, underscoring the need for further investigations.

Poster 112: Toxicity of Chlorhexidine and Povidone-Iodine to Rotator Cuff Tendon

Objectives: Antimicrobial wound lavages containing chlorhexidine gluconate (CHG) or povidone-iodine (PI) are commonly used to prevent periprosthetic infection during shoulder arthroplasty. However, the potential toxicity of these solutions to the native surrounding rotator cuff tendon is not well understood. In total shoulder arthroplasty, the rotator cuff tendon is often concomitantly repaired, and any toxic effects from antimicrobial wound lavage during surgery may be detrimental to postoperative rotator cuff tendon healing. The purpose of this study was to determine the in vitro effects of a 1-minute CHG exposure and a 3-minute 0.35% PI exposure on the viability and biochemical content of rotator cuff tendon. Methods: Infraspinatus tendons (n=6 per group) were isolated from shoulders of Nubian goats (n=3; 2 female and 1 male). Tendons were submerged in Irrisept (0.05% CHG in sterile water) for 1 minute or 0.35% Betadine (PI) for 3 minutes per manufacturer guidelines, followed by phosphate-buffered saline wash and culture in tissue culture medium. Control tendon specimens were similarly exposed to tissue culture medium, followed by phosphate-buffered saline wash and culture in tissue culture medium. After seven days of culture, isolates from both the tendinous and enthesis portions of the rotator cuff were analyzed for viability, collagen content (hydroxyproline assay), and glycosaminoglycan (GAG) content (dimethylmethylene blue assay). Statistical analyses were performed via one-way ANOVA with post hoc Tukey’s test. Results: Within the tendinous portion of the rotator cuff, CHG-treated specimens demonstrated a decrease in mean tenocyte viability compared to controls (p=0.045) (Fig.1). While PI-treated specimens exhibited lower mean viability compared to controls, this difference was not statistically significant. Within the enthesis portion of the rotator cuff, tenocyte viability decreased in both CHG- and PI-treated groups; however, these decreases were not statistically significant (Fig. 2). There were no significant differences in mean collagen content/wet weight or GAG/wet weight among groups within the tendinous and enthesis portion of the rotator cuff (Fig. 3). Conclusions: Due to the potential catastrophic consequences associated with periprosthetic infection, CHG and PI antimicrobial lavages are commonly used to reduce the risk of periprosthetic infection during shoulder arthroplasty. However, in this study, brief 0.05% CHG and 0.35% PI exposures to rotator cuff tendon resulted in lower tenocyte viability 7 days after treatment. These results are consistent with the toxicity reported with exposure of CHG and PI to other musculoskeletal tissues, including articular cartilage. The toxicity of antimicrobial irrigation to the surrounding native rotator cuff tendon may be detrimental to the postoperative healing of concomitant rotator cuff repair in the setting of shoulder arthroplasty.