Polymerase Chain Research Articles

Defense strategies against sweet potato chlorotic stunt and pakakuy virus coinfection unraveled

AbstractThis study into the response of two Ipomoea batatas (L.) Lam cultivars, Melinda and Tio Joe, to coinfection with sweet potato chlorotic stunt virus (SPCSV) and sweet potato pakakuy virus (SPPV), employed a comprehensive approach encompassing symptomatology, real‐time quantitative polymerase chain reaction, metagenomics, and transcriptomics. SPCSV is a quarantine virus with synergistic effects, which decimate yields. SPPV is the most prevalent DNA virus in sweet potato germplasm, with a tendency to persist in meristems, posing a significant risk for germplasm transfer between territories. Graft inoculation experiments revealed that while Tio Joe remained asymptomatic for 12 weeks and suppressed virus replication, Melinda displayed symptoms early on and exhibited high virus titers. Metagenomic analyses corroborated these observations and confirmed that SPCSV and SPPV were responsible. Transcriptomic analysis unveiled disparities in gene expression between Melinda and Tio Joe. Differential gene expression was heightened and altered in Melinda as the viruses disrupted its gene expression. Its defense strategies, such as inducing abscisic acid signaling, were insufficient to overcome disruptive viral effects like oxidative stress, rendering it susceptible. In contrast, Tio Joe had relatively stable differential gene expression, indicating resistance to SPPV–SPCSV coinfection. Overexpressed genes such as sirtuin, rapid alkalization factor, and nuclear pore anchor triggered quantitative resistance, supported with enriched organelles such as mitochondrion and pathways such as proteasome and cutin, suberine, and wax biosynthesis. Tio Joe maintained its genome integrity and inhibited viral replication by tightly controlling gene expression and preventing reactive oxygen species accumulation.

Warm growing season activates microbial nutrient cycling to promote fertilizer nitrogen uptake by maize

The influence of nitrogen (N) inputs on soil microbial communities and N uptake by plants is well-documented. Seasonal variations further impact these microbial communities and their nutrient-cycling functions, particularly within multiple cropping systems. Nevertheless, the combined effects of N fertilization and growing seasons on soil microbial communities and plant N uptake remain ambiguous, thereby constraining our comprehension of the optimal growing season for maximizing crop production. In this study, we employed 15N isotope labeling, high-throughput sequencing, and quantitative polymerase chain reaction (qPCR) techniques to investigate the effects of two distinct growing seasons on microbial communities and maize 15N uptake ratios (15NUR). Our results showed that the warm growing season (26.6 °C) increased microbial diversity, reduced network complexity but enhanced stability, decreased microbial associations, and increased modularization compared to the cool growing season (23.1 °C). Additionally, the warm growing season favored oligotrophic species and increased the abundance of microbial guilds and functional genes related to N, phosphorus, and sulfur cycling. Furthermore, alterations in the characteristics of soil microbial keystone taxa were closely linked to variations in maize 15NUR. Overall, our findings demonstrate significant seasonal variations in soil microbial diversity and functioning, with maize exhibiting higher 15NUR during the warm growing season of the double cropping system.

WNT1/ROR2 pathway enhances the Triple-Negative breast cancer invasion, migration, and Epithelial-Mesenchymal transition.

It has been evidenced that ROR2 influences the growth of many tumors, including non-small cell lung cancer, osteosarcoma, and breast cancer. This research examined the effect of the WNT1/ROR2 signaling pathway on the progression of triple-negative breast cancer (TNBC). Bioinformatics analysis results demonstrated that ROR2 had a higher messenger RNA (mRNA) expression level in TNBC tissues and was positively correlated with poor patient prognosis. Western blot analysis (WB) and quantitative reverse transcription polymerase chain reaction demonstrated that TNBC cells had relatively higher ROR2 mRNA and protein levels than normal cell lines. Transwell and Cell Counting Kit-8 (CCK-8) assay further proved that downregulating ROR2 expression dramatically slowed the MDA-MB-231 cell progression. WB detection of epithelial-mesenchymal transition (EMT)-related proteins suggested that knocking down ROR2 could alleviate the EMT tendency of cancer cells. The WNT1/ROR2 signaling pathway could be inhibited by the WNT inhibitor pyrvinium pamoate (PP). Experiments on in vitro cell functional recovery have demonstrated that PP could restore malignant phenotypes caused by ROR2 overexpression. Finally, the mouse model experiments further validated the anticancer effect of PP on TNBC. Generally speaking, the malignant progression of TNBC could be stimulated by the WNT1/ROR2 signaling pathway which can be inhibited by PP, suggesting the potential value of PP in controlling TNBC.

PCR GeneScan Analysis of Rearranged Immunoglobulin or T-Cell Receptor Genes for Clonality Diagnostics in Suspect Lymphoproliferations.

Assessment of the presence of clonal lymphoproliferations via polymerase chain reaction (PCR)-based analysis of rearranged immunoglobulin (IG) or T-cell receptor (TR) genes is a valuable method in the diagnosis of suspect lymphoproliferative disorders. Additionally, this methodology can be used for evaluating dissemination of lymphoma cells and for studying the clonal relationship between multiple (different locations) and consecutive (over time) lymphomas. Here we describe an integrated approach to assess clonality via analysis of Ig heavy chain (IGH), Ig kappa (IGK), TCR beta (TRB), and TCR gamma (TRG) gene rearrangements, based on the standardized multiplex PCRs as originally developed by the European BIOMED-2 consortium (currently named EuroClonality). The described protocol covers the pre-analytical phase of DNA isolation (from formalin-fixed paraffin-embedded and fresh tissues, body fluids, peripheral blood, and bone marrow), the analytical phase of PCR GeneScan analysis, and the post-analytical interpretation of the obtained profiles, following established guidelines.

Identification of pathogenic bacteria from eggshell of leatherback sea turtle (Dermochelys coriacea) in Lampuuk Beach, Aceh Besar using 16S rRNA gene

Currently, the leatherback turtle population in Indonesia tends to decline. One of the factors that cause turtles' existence is the turtle eggs that fail to hatch because bacteria contaminate them, 80% of the reasons turtle eggs fail to hatch are due to infection by microorganisms found on the eggshell. Turtle eggs have a soft structure and there are pores that function for gas exchange and water absorption. Microorganisms in sand can infect turtle eggs through the pores and cause hatching failure. This study aims to isolate and identify pathogenic bacteria found in leatherback turtle eggshell (Dermochelys coriacea) from Lampuuk Beach, Aceh Besar, based on the 16S rRNA gene analysis. Eight eggshell samples from leatherback turtle eggs that failed and hatched were cultured on Nutrient Agar (NA) medium. Then, the samples were inoculated on a blood agar medium for haemolysis test. Molecular identification of the 16S rRNA gene was also carried out to determine bacterial species. A total of five bacterial colonies from leatherback turtle eggshells were successfully isolated consisting of two Gram-negative bacteria (Penyu Belimbing Aceh (PBA) = PBA-1 and PBA-2 and three Gram-positive bacteria (PBA-3, PBA-4, and PBA-5). Based on the haemolysis test, the five bacterial isolates were unable to haemolyze blood. Bacterial DNA of PBA-1 and PBA-2 were successfully isolated and amplified using polymerase chain reaction (PCR) with a DNA target of ~1500 bp. Analysis using BLASTN and phylogenetic tree construction showed that PBA-1 isolate had 98.64% similarity with Acinetobacter baumannii, while PBA-2 isolate had 97.82% similarity with Enterobacter kobei. Both bacteria are members of the Enterobacteriaceae family. It can be concluded that, there were two pathogenic bacteria can potentially be opportunistic pathogens found in leatherback turtle eggshells that failed or succeeded in hatching, namely A. baumannii and E. Kobei

Characterization of Bread Wheat Genotypes Using SSR Markers for Terminal Heat Tolerance

Wheat is a major global food crop, but its productivity is increasingly threatened by terminal heat stress due to climate change. This study aimed to characterize the genetic diversity and heat tolerance of widely grown bread wheat genotypes using SSR markers to identify heat-tolerant cultivars adaptable to various regions in Bangladesh. A total of 15 genotypes were screened, and 13 polymorphic SSR (Simple Sequence Repeats) markers were used to determine the genetic similarity and categorize genotypes based on their heat tolerance. The molecular analysis revealed 13 polymorphic SSR markers that produced distinct PCR (Polymerase Chain Reaction) bands across the different genotypes. By the conclusion of the study, bread wheat genotypes were categorized as either tolerant or sensitive to heat, and the genetic similarity among the varieties was assessed using molecular markers. The genetic similarity coefficients obtained from the SSR primer screening ranged from 0.00 to 0.925. The lowest genetic distance (0.000) was found in Nadi 2 vs BAW1147 variety pair indicating that they are genetically similar to each other. Comparatively higher genetic distance (0. 925) was observed between BAW 1290 vs BARI Gom 28. The dendrogram divided the fifteen genotypes into two main groups, A and B, both formed at a similar coefficient of 0.05. Group A included seven genotypes and was further divided into two clusters, while Group B comprised eight genotypes, which were also divided into two clusters. The categorization of the genotypes was based on the average Heat Susceptibility Index (HSI), Thousand Grain Weight (TGW), grain yield, and the relative reduction in TGW and grain yield under stress conditions compared to timely sown conditions, aligning with the molecular data. Among the fifteen genotypes, BARI Gom 25, BARI Gom 26, BARI Gom 27, BARI Gom 28, BARI Gom 29, BARI Gom 30, and BARI Gom 31 demonstrated their adaptability to late sown conditions. The heat tolerance observed in these genotypes, as indicated by their SSR marker scores, is anticipated to provide valuable insights for molecular breeding studies focused on heat resistance.

Study of the relationships among known virulence genes, coccoid transformation and cytotoxicity of Helicobacter pylori in different clinical diseases

ABSTRACT Background Helicobacter pylori (H. pylori) has infected approximately 4.4 billion individuals worldwide. The known virulence genes and the existing H. pylori typing methods have not been shown to have a recognized correlation with its infectivity. The aim of this study was to elucidate the relationships among known important virulence genes, coccoid transformation, and cytotoxicity of H. pylori isolated from individuals with different clinical diseases to provide guidance for the development of new virulence typing methods for H. pylori. Methods The known important virulence genes of 35 H. pylori strains were identified by whole-gene next-generation sequencing (WGS) and polymerase chain reaction (PCR). The chronological changes in the proportion of coccoid forms of H. pylori and their ultramicroscopic structures were observed chronologically using transmission electron microscopy. Human gastric mucosal epithelial cells (GES-1) w Results There were no significant correlations among the known important virulence genes, coccoid transformation and cytotoxicity of H. pylori isolated from patients with different clinical diseases. We developed a new virulence classification based on the defensive and offensive abilities of H. pylori. Conclusions Coccoid transformation and virulence are two independent characteristics of H. pylori that reflect its defensive and offensive abilities, respectively. These two abilities work synergistically, warranting the construction of a new virulence typing method for H. pylori. However, the correlation between the new virulence classification and pathogenic ability still needs to be further ver

The Role of Quantitative PCR in Evaluating the Clinical Significance of Human Bocavirus Detection in Children.

Human bocavirus (HBoV) has emerged as a significant pathogen primarily associated with respiratory infections in children. This study aimed to evaluate the clinical relevance of HBoV infection by quantifying viral loads in nasopharyngeal swabs from hospitalized children with acute respiratory infections (ARIs) and investigating correlations with clinical outcomes. A total of 957 children were tested, with HBoV detected in 73 cases (7.6%), either as a sole infection or co-infection with other respiratory viruses. Quantitative polymerase chain reaction (qPCR) was employed to measure viral load, and a threshold of 104 copies/mL was used to differentiate high and low viral concentrations. Results have shown that children with lower respiratory tract infections (LRTIs) had significantly higher viral loads, most notably in cases where HBoV was the sole pathogen. Additionally, children with pre-existing health conditions were more likely to have elevated HBoV concentrations compared to those who were previously healthy. Children with higher viral loads were more likely to require oxygen supplementation and receive empirical antibiotic therapy, indicating a more severe clinical course. This study underscores the importance of considering HBoV viral load in clinical diagnostics, as higher concentrations were associated with more severe presentations, including the need for oxygen support. Future research should focus on refining diagnostic thresholds and exploring HBoV's role in co-infections to enhance patient care strategies.

Association between total functional tooth unit score and hemoglobin A1c levels in Japanese community-dwelling individuals: the Nagasaki Islands study.

It is widely recognized that periodontal disease is associated with diabetes mellitus. Periodontal disease is accompanied by inflammation of the periodontal tissue, impaired masticatory function, and the presence of periodontopathic bacteria, all of which may affect glycemic control. However, the exact relationship between these factors and glycemic control has not yet been established. In this study, we aimed to investigate the relationship between periodontal disease-related factors and glycemic control in Japanese community-dwelling individuals. We conducted a cross-sectional study involving 671 participants aged 29-92 (65.3 ± 12.1) years, using data from the Nagasaki Islands Study. Participants underwent routine medical examinations, including body mass index (BMI) and hemoglobin A1c (HbA1c) levels. Information on the participants' demographics (age and sex) and whether they were on diabetes medications, had an exercise habit, consumed alcohol, engaged in late-night eating, had regular dental checkups, and smoked was obtained using a self-administered questionnaire. Dental examinations were performed to examine dentition status, probing pocket depth, clinical attachment level (CAL), and bleeding on probing. Functional tooth units (FTUs), defined as pairs of occluding posterior teeth, were used as an indicator of occlusal support area. Saliva samples were collected and levels of two species of periodontopathic bacteria (Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans) were determined using real-time polymerase chain reaction. We analyzed the association between HbA1c levels and variables related to periodontal status, masticatory function, and salivary levels of periodontopathic bacteria. Bivariate analysis showed that HbA1c levels were significantly associated with age, sex, exercise habit, BMI, diabetes medications, CAL, salivary P. gingivalis level, number of teeth, and three FTU subcategories. In the multiple regression analysis, age, BMI, diabetes medications, and total FTU score (i.e., including natural teeth, implant-supported artificial teeth, fixed prostheses, and removable dentures) remained associated with HbA1c levels (B = 0.23, 0.14, 0.52, and - 0.12; p < 0.001, p < 0.001, p < 0.001, and p = 0.008, respectively). In this community-based cross-sectional study, total FTU was significantly associated with HbA1c levels, independent of other risk factors. This suggests that reconstructed occlusal support areas, including dentures, are associated with glycemic control in the older population.

Multiple bloodmeals enhance dissemination of arboviruses in three medically relevant mosquito genera

BackgroundMosquitoes in nature may acquire multiple bloodmeals (BMs) over the course of their lifetime; however, incorporation of frequent feeding behavior in laboratory vector competence studies is rarely done. We have previously shown that acquisition of a second non-infectious BM can enhance early dissemination of Zika virus (ZIKV), dengue virus, and chikungunya virus in Aedes aegypti and ZIKV in Aedes albopictus mosquitoes, yet it is unknown if other taxonomically-diverse virus-vector pairings show a similar trend under a sequential feeding regimen.MethodsTo test this, we evaluated the impact of a second noninfectious BM on the vector competence of Aedes aegypti and Anopheles quadrimaculatus for Mayaro virus, Culex quinquefasciatus for West Nile virus, Aedes triseriatus for La Crosse virus, and Aedes aegypti for Oropouche virus (OROV). Female mosquitoes were fed BMs containing these viruses and half of them were given a second noninfectious BM at 3 or 4-days post infection. Mosquitoes were harvested at various time points and assayed for virus infection in bodies and disseminated infection in legs by performing reverse transcription-quantitative polymerase chain reaction (RT-qPCR) assays.ResultsWe found that a second noninfectious BM had no impact on midgut infection rates but increased virus dissemination for all but one of the virus-vector pairings- Ae. aegypti and OROV. Unlike the other arboviruses under consideration, which are strictly mosquito-borne, biting midges (Culicoides spp.) serve as the main vector of OROV and this virus rarely disseminated to the mosquito leg tissue in our study.ConclusionsTaken together, our findings show that sequential blood feeding enhances virus dissemination across diverse arbovirus-vector pairings, representing three mosquito genera and virus families, but a second BM was insufficient to overcome a strong midgut virus escape barrier in a nonnatural virus–vector pairing.Graphical

EFNA5 suppresses cell proliferation and tumor metastasis in hepatoma via epithelial-to-mesenchymal transition

BackgroundEphrinA5 belongs to a subclass of ephrin ligands. Abnormal signal transduction of EFNA5 shows a relationship to the development of various tumors. In this study, we explored the level of EFNA5 in hepatoma cells and the influence of up regulation of EFNA5 expression level on the proliferation, invasion, and migration of HepG2 and LM3 cells. Additionally, this work focused on examining its possible mechanism of action, and future impacts on clinical practice.MethodsImmunohistochemistry was utilized to explore the connection between EFNA5 and hepatoma. Real-time quantitative polymerase chain reaction was used for determining the expression levels of EFNA5 in several hepatoma cell lines and normal hepatocytes. Cells were transfected with a pCMV3-EFNA5-flag plasmid and an EFNA5 plasmid. The expression efficiency of EFNA5 was identified through qRT-PCR. For the purpose of further identifying cell proliferation, the Cell Counting Kit-8 assay was applied. To identify changes of cell migration and invasion ability, Transwell and Boyden tests were utilized. Western blot was employed to identify the expressions mof EFNA5 and possible downstream molecules.ResultsData acquired from The Cancer Genome Atlas demonstrated that the level of EFNA5 in hepatoma was significantly downregulated in relative to the normal hepatocytes (P < 0.05). Upregulation of EFNA5 expression in hepatoma cells hindered the proliferative, invasive, and migratory ability of cells (P < 0.05). Additionally, EFNA5 downregulated the level of epithelial–mesenchymal transition-related molecules and EGFR.ConclusionsThe expression of EFNA5 was low in hepatoma cells. An increase in EFNA5 levels hinders the proliferation, invasion, and migration of hepatoma cells. These effects may occur through inhibition of hepatoma epithelial–mesenchymal transition by EFNA5. Moreover, the study on the mechanisms of proliferation, invasion and metastasis of hepatoma provides a novel theoretical basis, and may influence the clinical practice of tumor treatment in the future.

Potential role of heteroplasmic mitochondrial DNA mutations in modulating the subtype-specific adaptation of oral squamous cell carcinoma to cisplatin therapy

Cancer cells are constantly evolving to adapt to environmental changes, particularly during exposure to drug treatment. In this work, we aimed to characterize genetic and epigenetic changes in mitochondrial DNA (mtDNA) that may increase the resistance of oral squamous cell carcinoma (OSCC) to cisplatin. We first derived drug-resistant cells from two human OSCC cell lines, namely SAS and H103, by continual cisplatin treatments for about 4 months. To determine mtDNA changes induced by cisplatin, we performed nanopore sequencing and quantitative polymerase chain reaction analysis of mtDNA extracted from the cells pre- and post-treatment. We also assessed the mitochondrial functions of the cells and their capacity to generate intracellular reactive oxygen species (ROS). We found that in the cisplatin-resistant cells derived from SAS, there was a reduction in mtDNA content and significant enrichment of a m.3910G > C mutation in the MT-ND1 gene. However, such changes were not detected in cisplatin-resistant H103 cells. The cisplatin treatment also altered methylation patterns in both SAS and H103 cells and decreased their sensitivity to ROS-induced cytotoxicity. We suggest that the sequence alterations and epigenetic changes in mtDNA and the reduction in mtDNA content could be key drivers of cisplatin resistance in OSCC. These mtDNA alterations may participate in cellular adaptation that serves as a response to adverse changes in the environment, particularly exposure to cytotoxic agents. Importantly, the observed mtDNA changes may be influenced by the distinct genetic landscapes of various cancer subtypes. Overall, this study reveals significant insights into cisplatin resistance driven by complex mtDNA dynamics, particularly in OSCC. This underscores the need for targeted therapies tailored to the genetic profiles of individual OSCC patients to improve disease prognosis.

Macrophage-Derived Exosomes Promoted the Development and Stemness of Inflammatory Bowel Disease-Related Colorectal Cancer via nuclear paraspeckle assembly transcript 1-Mediated miRNA-34a-5p/phosphoprotein enriched in astrocytes 15 Axis.

Inflammatory bowel disease (IBD) is closely associated with the development of colorectal cancer (CRC) due to the chronic inflammatory response. Macrophages play critical roles in regulating the microenvironment to facilitate tumor progression. Exosomes are key modulators for the communication between macrophages and tumor cells. The mechanism of macrophage-derived exosomes in IBD-related CRC development remains unclear. The macrophages were isolated using fluorescence activating cell sorter (FACS). The RNA and protein expressions in exosomes and CRC cells were examined by quantitative real-time polymerase chain reaction and western blot assays, respectively. CRC cell development was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, BrdU staining, Transwell assay, and spheroid formation assay. The level of stemness was determined by detecting the proportion of leucine-rich repeat-containing G-protein-coupled receptor 5 (LGR5)-positive CRC cells and the expression of LGR5, CD133, and CD44. Molecular interaction experiments were done using luciferase reporter assay and RNA immunoprecipitation assay. Xenograft tumor model in vivo and immunohistochemistry were used to observe the pathological changes. Macrophage-derived exosomes from IBD-related CRC tissues were enriched with nuclear paraspeckle assembly transcript 1 (NEAT1) and able to promote the progression and stemness of CRC both in vitro and in vivo. The exosomal NEAT1 could sponge miR-34a-5p, leading to the restoration of PEA15 expression in CRC cells and promoting the development of CRC. Inhibition of NEAT1 in exosomes could effectivity inhibit the tumor growth in the CRC xenograft model. These findings provide novel insights into how macrophages affect CRC development and highlight exosomal NEAT1 as a therapeutic target for CRC treatment.

Safety and efficacy of sofosbuvir/ledipasvir combination in treatment of chronic hepatitis C infection in adolescents aged 12–17 years old

BackgroundChronic hepatitis C virus (CHC) infection represents a crucial health problem, especially among children and adolescents. The ledipasvir (LDV)/sofosbuvir (SOF) regimen has been approved to treat adolescents (aged 12 to 17 years old) infected with hepatitis C virus (HCV) genotypes 1, 4, 5, and 6 and then extended to include children above or equal to 3 years old. The current study aims to evaluate the safety and efficacy of SOF/LDV combination in treating CHC-infected 12- to 17-year-old adolescents.Patients and methodsThis retrospective cohort study was performed on 147 Egyptian adolescents with CHC. The patients were treated with SOF 400 mg/LDV 90 mg combination once daily for 12 weeks. Possible side effects and laboratory data including HCV ribonucleic acid polymerase chain reaction (RNA PCR), complete blood count (CBC), and liver tests were recorded at baseline and week 12 after the end of treatment (EOT).ResultsSustained virological response 12 weeks (SVR12) after end of treatment (EOT) was observed in 146 out of 147 patients (99.3%). The treatment regimen was efficiently tolerated with no reported cases of discontinuation caused by intolerability. Moreover, the side effects were minimal; 71.4% of the patients did not report any side effects related to the treatment. However, the rest mentioned fatigue, headache, or both of them. Fatigue was the main side effect reported in 16.3% of the patients. Furthermore, ALT and AST levels were normalized after treatment. FIB-4 and APRI scores were statistically significantly decreased 2 years post-SVR, in comparison to their levels before treatment, from 0.34 and 0.36 to 0.25 and 0.17, respectively.ConclusionThe LDV/SOF regimen is one of the safe regimens used to treat adolescent patients with CHC infection.

Relationship between the AGT M235T genetic variant and the characteristics and prognosis of coronary atherosclerosis in patients with acute myocardial infarction.

Along with environmental components, genetic factors play an essential role in the pathophysiology and progression of acute myocardial infarction (AMI). There is limited and conflicting data on the influence of the AGT M235T genetic variant on coronary atherosclerosis and death in AMI patients. We carried out a prospective cohort study among 504 Vietnamese AMI patients selected between January 2020 and May 2021. All patients underwent invasive coronary angiography, had AGT M235T genetic variant genotyped using the polymerase chain reaction method, and were followed up for 12-month all-cause mortality. The proportions of the MM, MT, and TT genotypes were 0.4%, 20.8%, and 78.8%, respectively. There was no significant difference between the TT genotype and the MM + MT genotype groups regarding the position and number of stenosed coronary artery branches and the Gensini score. The AGT M235T genetic variant did not affect 12-month mortality (hazard ratio of TT vs. MM + MT: 1.185; 95% confidence interval: 0.596-2.354; P = 0.629). Subgroup analyses by age, sex, hypertension, diabetes mellitus, dyslipidemia, obesity, smoking, and angiotensin-converting enzyme inhibitor or angiotensin II receptor blocker therapy also did not reveal an association between the AGT M235T variant and all-cause mortality. In summary, the AGT M235T genetic variant was not found to be associated with coronary atherosclerosis characteristics and 12-month mortality in Vietnamese patients with AMI. Further multicenter studies with larger sample sizes and extended follow-up periods are needed to investigate this issue.

Cloning, bioinformatics analysis and expression of the cysteine dioxygenase type 1 (CDO1) gene in domestic yak

IntroductionThe CDO1 gene is an important gene in the taurine synthesis pathway and has been observed to have high expression in ovaries of female mammals. This study aims to explore the conservation of CDO1 gene in domestic yaks, as well as to examine the fundamental characteristics of CDO1 gene and its expression in female yaks.MethodsOvarian samples were collected from yaks in the follicular phase, luteal phase and gestation period in this experiment, and their total RNA and protein were extracted. Then Polymerase Chain Reaction (PCR) and bioinformatics online software were used to clone and analyze the CDO1 gene. The relative expression of CDO1 in yak ovaries was detected by Quantitative Real-time PCR (RT-qPCR) and Western blotting. The distribution and localization of CDO1 protein in ovary were detected by immunohistochemistry.ResultsWe have successfully cloned the coding region of CDO1 gene in yak. The results showed that the CDS region of CDO1 gene was 603 bp, encoding 200 amino acids, and was a relatively stable hydrophilic protein. CDO1 is relatively conservative in species evolution. The protein encoded by CDO1 gene does not have a signaling peptide or a transmembrane structure. It is a protein that is not involved in transmembrane transport and is mainly located in the cytoplasm. The secondary structure of the protein is dominated by the random coil. CDO1 is estimated to interact with 10 proteins. The results of RT-qPCR and Western blotting showed that the CDO1 gene exhibited the highest expression in the ovary during the luteal phase and the lowest expression in the ovary during the follicular phase (P &amp;lt; 0.01). The results of immunohistochemistry showed that CDO1 was mainly expressed in granular cells, theca cells and lutein cells of ovarian tissue.ConclusionThese results suggest that the CDO1 gene has undergone minimal evolutionary changes during the course of animal evolution. The results provide a reference for further investigation of the function of CDO1 gene in reproduction and production in yaks.

Artificial intelligence can be trained to predict c-KIT-11 mutational status of canine mast cell tumors from hematoxylin and eosin-stained histological slides.

Numerous prognostic factors are currently assessed histologically and immunohistochemically in canine mast cell tumors (MCTs) to evaluate clinical behavior. In addition, polymerase chain reaction (PCR) is often performed to detect internal tandem duplication (ITD) mutations in exon 11 of the c-KIT gene (c-KIT-11-ITD) to predict the therapeutic response to tyrosine kinase inhibitors. This project aimed at training deep learning models (DLMs) to identify MCTs with c-KIT-11-ITD solely based on morphology. Hematoxylin and eosin (HE) stained slides of 368 cutaneous, subcutaneous, and mucocutaneous MCTs (195 with ITD and 173 without) were stained consecutively in 2 different laboratories and scanned with 3 different slide scanners. This resulted in 6 data sets (stain-scanner variations representing diagnostic institutions) of whole-slide images. DLMs were trained with single and mixed data sets and their performances were assessed under stain-scanner variations (domain shifts). The DLM correctly classified HE slides according to their c-KIT-11-ITD status in up to 87% of cases with a 0.90 sensitivity and a 0.83 specificity. A relevant performance drop could be observed when the stain-scanner combination of training and test data set differed. Multi-institutional data sets improved the average accuracy but did not reach the maximum accuracy of algorithms trained and tested on the same stain-scanner variant (ie, intra-institutional). In summary, DLM-based morphological examination can predict c-KIT-11-ITD with high accuracy in canine MCTs in HE slides. However, staining protocol and scanner type influence accuracy. Larger data sets of scans from different laboratories and scanners may lead to more robust DLMs to identify c-KIT mutations in HE slides.

Retinal detachment and mortality in patients with cytomegalovirus retinitis: A multicenter study in taiwan.

To characterize patients with cytomegalovirus (CMV) retinitis and identify risk factors for retinal detachment (RD) and mortality in this Taiwanese patient population. This retrospective study included patients diagnosed with CMV retinitis between 2007 and 2019. The diagnosis was confirmed through aqueous polymerase chain reaction (PCR). Relevant data were collected from the Chang Gung Research Database. Univariate Cox regression was performed to identify the associations of RD and mortality risks with various patient characteristics, including demographic features, comorbidities, laboratory results, and medication use patterns. In total, 32 patients with CMV retinitis were included. Among these patients, 78.1% had an immunocompromised status, including 56.3% with high-dose systemic steroid use, 21.9% with HIV infection, 12.5% with hematologic malignancy, and 9.4% with renal transplantation. Approximately 21.9% of patients had RD 2.4 ± 2.1 months after CMV retinitis diagnosis, and 34.4% died within 6.2 [4.2, 38.2] months after diagnosis. Patients with RD had a statistically significant, but likely not clinically significant, later initiation of anti-CMV medications compared to their non-RD counterparts (8 [5, 23] days vs. 2 [1,11] days, p = 0.039). Mortality was significantly associated with older age (hazard ratio [HR]: 1.06; 95% confidence interval [CI]: 1.02-1.10), hematologic malignancy (HR: 5.92; 95% CI: 1.44-24.37), and positivity for CMV on blood PCR (HR: 4.93; 95% CI: 1.49-16.35). Our study suggests that older age, hematologic malignancy, and positivity for CMV on blood PCR are risk factors for mortality in patients with CMV retinitis. What is known Cytomegalovirus (CMV) retinitis is the predominant sight-threatening opportunistic ocular infection in patients with acquired immunodeficiency syndrome (AIDS). What is new In the era of highly active antiretroviral therapy for AIDS, the majority of CMV retinitis patients are those receiving immunomodulatory therapy for underlying diseases. Older age, hematologic malignancy, and positive blood polymerase chain reaction for CMV are potential risk factors for mortality in patients with CMV retinitis.

Onychomycosis in the US Pediatric Population-AnEmphasis on Fusarium Onychomycosis.

Onychomycosis is a common nail disease that is often difficult to treat with a high risk of recurrence. To update our current understanding of the etiologic profile in pediatric patients with onychomycosis utilizing molecular diagnosis by polymerase chain reaction (PCR) combined with histopathologic examination. Records of 19,770 unique pediatric patients were retrieved from a single diagnostic laboratory in the United States spanning over a 9-year period (March 2015 to April 2024). This cohort represents patients clinically suspected of onychomycosis seen by dermatologists and podiatrists. Dermatophytes, nondermatophyte molds (NDMs), and yeasts were identified by multiplex real-time PCR corroborated by the demonstration of fungal invasion on histopathology. An average of 37.0% of all patients sampled were mycology-confirmed to have onychomycosis. Most patients were between ages 11 and 16 years, and the rate of mycologically confirmed onychomycosis was significantly higher among the 6- to 8-year (47.2%) and 9- to 11-year (42.7%) age groups compared to the 0- to 5-year (33.1%), 12- to 14-year (33.2%), and 15- to 17-year (36.7%) age groups. The majority of infections were caused dermatophytes (74.7%) followed by NDMs (17.4%). The Trichophyton rubrum complex represents the dominant pathogen with higher detection rates in the 6- to 11-year-olds. Fusarium was the most commonly isolated NDM with an increasing prevalence with age. Elementary school-aged children have a higher risk of contracting onychomycosis which may be attributed to the onset of hyperhidrosis at puberty, use of occlusive footwear, nail unit trauma, and walking barefoot. Fusarium onychomycosis may be more prevalent than expected, and this may merit consideration of management strategies.

Haemoprotozoan and haemorickettsial carrier status in pet and community owned dogs of south India

The increasing population of dogs and changes in the climatic conditions have resulted in the emergence and re-emergence of vector-borne diseases in canines. These vectors borne diseases in canines pose a diagnostic challenge to the field veterinarians because of co-infections with several pathogens. Comprehensive data on the prevalence of haemoparasites and haemorickettsiales in pet and community owned dogs from south India are scant. Hence, the present study aims to find and compare the prevalence of these infections in the pet and the community owned dogs of Kerala, a south Indian state. Two hundred and seventy-two pets and 150 community owned dogs were examined by microscopy and polymerase chain reaction (PCR) for infections with different heamoparasites and haemorickettsials from January 2018–November 2020 in the state of Kerala. A high prevalence of Babesia gibsoni infection (42.2–60.0 %) in pet and community owned dogs, followed by Babesia vogeli (5.8–39.3 %), Hepatozoon canis (0.7–28.0 %), Trypanosoma evansi (0.0–27.3 %), Ehrlichia canis (0.3–0.6 %) and Anaplasma platys (0.0–0.6 %) was observed in the present study. Eighty-eight per cent (132/150) of the community owned dogs and 49.2 % (134/272) of the pet dogs were positive for at least one pathogen. Phylogenetic analysis based on the partial nucleotide sequences of 18S rRNA and TRAP gene of B. gibsoni, 18S rRNA genes of B. vogeli and H. canis and RoTat, 1.2 virB9 and 16S rRNA genes of T. evansi, E. canis and A. platys, respectively was carried out. B. vogeli, H. canis, E. canis and A. platys revealed genetic relatedness between the Indian isolates and the isolates from other countries. However, B. gibsoni isolates from the Indian sub-continent were genetically unique compared to other Asian isolates. The clustering of T. evansi isolates from India in two clades viz., livestock origin (cattle, buffalo) and others indicated their genetic variability. The present study summarizes the prevalence of some of the haemoparasites and haemorickettsials in the dog populations of Kerala (south India) and also determined their genetic relationship with the isolates prevalent in dogs in other countries.