Genotyping Methods Research Articles

Deciphering the genotypic diversity and epidemiology of scrub typhus genotypes in India: A systematic review and meta-analysis.

Scrub typhus is a significant cause of Acute Febrile Illness in India and presents a range of clinical manifestations influenced by the genotypes of the causative agent, Orientia tsutsugamushi. Understanding the distribution of these genotypes is crucial for public health planning, diagnostic accuracy, and vaccine development. In this systematic review, we analyzed 18 studies from an initial pool of 2013 records identified through database searches following PRISMA guidelines. The relevance of the findings to the genotype prevalence of Scrub typhus in India was taken into consideration throughout the selection process. Various genotyping methods, predominantly Polymerase Chain Reaction (PCR), were used in these studies to identify genotypes such as Karp, Gilliam, Kato, and others. The review revealed diverse prevalence rates of Scrub typhus genotypes across different Indian states. The Karp genotype is the most prevalent in India, with a pooled prevalence of 47%, followed by the Kato genotype at 40% and the Gilliam genotype at 28%. The Karp genotype showed significant variability, with prevalence rates ranging from 100% in Andhra Pradesh (2018) to 6.16% in Uttar Pradesh (2016-2017). The Kato genotype's presence varied significantly, with high prevalence in certain states like Karnataka and absence in others like Andhra Pradesh. The Gilliam genotype also displayed a unique distribution pattern, dominating in Uttar Pradesh with a prevalence of 93.8% in some years. Other genotypes like Boryong, IHS-II, Neimeng, Kuroki, TA763, and Kawasaki also showed varied prevalence across states. There was significant heterogeneity in the prevalence rates of different genotypes among studies, as indicated by I2 statistics. The diversity in Scrub typhus genotypes across India underscores the disease's complex epidemiology, presenting diagnostic challenges and opportunities for targeted public health interventions. For efficient disease management and control plans in India, it is imperative to understand the changing genetic landscape of scrub typhus through ongoing study and surveillance.

Anti-biofilm activity of carvacrol-thymoquinone nanocarriers on vulvovaginal candidiasis isolates.

Given the recurrent nature of vulvovaginal candidiasis (VVC), the restricted availability of effective antifungal agents, and the recent rise in drug resistance, this study sought to assess the antifungal efficacy of carvacrol-thymoquinone delivered via a nanocarrier on Candida isolates obtained from patients with VVC. Isolates were identified using phenotypic and genotypic methods. Nanocarriers were synthesized using the thin-film hydration method. The antifungal activity of carvacrol-thymoquinone was evaluated using the broth microdilution method (CLSIM27-A3). The impact of nanocarriers on the biofilm formation capabilities of Candida isolates was assessed using the MTT assay. Data were analyzed using the Mann-Whitney U test. The nanocarrier exhibited a spherical morphology with a diameter measuring 50 nm. The nano-formulated drug combination could inhibit biofilm formation in C. albicans at half the minimum inhibitory concentration and in C. glabrata at the minimum inhibitory concentration. Our results suggest that the carvacrol-thymoquinone nanocarrier can be studied further in vivo for potential use in the treatment of recurrent VVC.

Association of killer immunoglobulin-like receptor genotypes and haplotypes with acute lymphoblastic leukemia risk.

Killer immunoglobulin-like receptors (KIRs) are key molecules used by natural killer (NK) cells to interact with target cells. These receptors exhibit extensive genotypic polymorphism which has been associated with varying outcomes in immune responses against diseases. This study aimed to investigate the relationships between KIR genotypes and haplotypes with acute lymphoblastic leukemia (ALL) in Saudi patients. A total of 259 Saudi subjects including 145 cases of acute lymphoblastic leukemia (ALL) and 114 healthy controls living in Riyadh were genotyped for 16 KIR genes and the two HLA-C1 and -C2 allotypes using PCR-SSP genotyping method. A significant high frequency of the two inhibitory KIR genes; 2DL1 (OR = 2.4; p < 0.0001) and 3DL1(OR = 10.87; p = 0.0068) in ALL compared to healthy group was observed. In contrast, the activating 2DS4 gene was significantly higher in healthy controls (OR = 0.15, p < 0.0001) compared to ALL patients. Haplotype analysis shows that BX haplogroup was strongly associated with the occurrence of ALL (OR = 4.39; p < 0.0001). Further combinatory analysis of KIR genes with their HLA-C1 and -C2 ligands demonstrated strong statistically protective effect of the 2DS1-C2 combination from ALL (OR = 0.06; p = 0.0003). This study presents strong evidence supporting the connection between certain KIR genotypes, haplotypes, and KIR-HLA combinations with acute ALL in the Saudi population. The heightened occurrence of inhibitory KIR genes (2DL1 and 3DL1) and the BX haplotype in ALL patients indicates a possible involvement of these genetic variability with the dysfunctional of NK cells in the context of ALL disease.

Application of 2D polymerase chain reaction for single-tube detection of high-risk human papillomaviruses

PurposeThe persistent infection of high-risk HPV (HR-HPV) is intricately linked to the onset and progression of cervical cancer. This research endeavored to develop a high-throughput 2D PCR method for closed-tube genotyping of 11 HR-HPVs.MethodsBase-quenched probes were specifically designed for FAM, VIC, and CY5 channels. The 2D PCR system underwent optimization, with its detection performance assessed in terms of specificity and sensitivity. Plasmid mixtures was used to simulate multiple infections of HPV, providing preliminary insights into the detection efficacy and throughput of the 2D PCR technology. Ultimately, the detection capability of this method was assessed using clinical samples.ResultsThe sequenced tags, when paired with primers, could generate Tm differences exceeding 3°C. These were then integrated with a fluorescent channel and Tm to differentiate and identify target genes upon detection. The refined 2D PCR system was confirmed to be free from cross-reactions and exhibited high specificity, capable of detecting 12 target genes within a single tube. A total of 294 cervical exfoliated cell samples were tested using 2D PCR and flow fluorescence hybridization method. The overall concordance between the two detection methods was 96.17% (Kappa = 0.910).ConclusionThe 2D PCR method, which integrates asymmetric PCR amplification with melting curve analysis, has the capacity to detect 11 types of HR-HPVs across three channels. This closed-tube detection approach offers several benefits including high throughput, straightforward operation, and low detection cost. Consequently, it can be effectively utilized for early screening and prevention of cervical cancer.

Cost-Effective Detection of SNPs and Structural Variations in Full-Length Genes of Wheat and Sunflower Using Multiplex PCR and Rapid Nanopore Kit

The rapid identification of allele variants in target genes is crucial for accelerating marker-assisted selection (MAS) in plant breeding. Although current high-throughput genotyping methods are efficient in detecting known polymorphisms, they are limited when multiple variant sites are scattered along the gene. This study presents a target amplicon sequencing approach using Oxford Nanopore Technologies (ONT-TAS) to rapidly sequence full-length genes and identify allele variants in sunflower and wheat collections. This procedure combines multiplex PCR and a rapid sequencing kit, significantly reducing the time and cost compared to previous methods. The efficiency of the approach was demonstrated by sequencing four genes (Ahasl1, Ahasl2, Ahasl3, and FAD2) in 40 sunflower genotypes and three genes (Ppo, Wx, and Lox) in 30 wheat genotypes. The ONT-TAS revealed a complete picture of SNPs and InDels distributed over the individual alleles, enabling rapid (4.5 h for PCR and sequencing) characterization of the genetic diversity of the target genes in the germplasm collections. The results showed a significant diversity of the Ahasl1/Ahasl3 and Wx-A/Lox-B genes in the sunflower and wheat collections, respectively. This method offers a high-throughput, cost-effective (USD 3.4 per gene) solution for genotyping and identifying novel allele variants in plant breeding programs.

Evaluation of genomic and phenomic prediction for application in apple breeding

BackgroundApple breeding schemes can be improved by using genomic prediction models to forecast the performance of breeding material. The predictive ability of these models depends on factors like trait genetic architecture, training set size, relatedness of the selected material to the training set, and the validation method used. Alternative genotyping methods such as RADseq and complementary data from near-infrared spectroscopy could help improve the cost-effectiveness of genomic prediction. However, the impact of these factors and alternative approaches on predictive ability beyond experimental populations still need to be investigated. In this study, we evaluated 137 prediction scenarios varying the described factors and alternative approaches, offering recommendations for implementing genomic selection in apple breeding.ResultsOur results show that extending the training set with germplasm related to the predicted breeding material can improve average predictive ability across eleven studied traits by up to 0.08. The study emphasizes the usefulness of leave-one-family-out cross-validation, reflecting the application of genomic prediction to a new family, although it reduced average predictive ability across traits by up to 0.24 compared to 10-fold cross-validation. Similar average predictive abilities across traits indicate that imputed RADseq data could be a suitable genotyping alternative to SNP array datasets. The best-performing scenario using near-infrared spectroscopy data for phenomic prediction showed a 0.35 decrease in average predictive ability across traits compared to conventional genomic prediction, suggesting that the tested phenomic prediction approach is impractical.ConclusionsExtending the training set using germplasm related with the target breeding material is crucial to improve the predictive ability of genomic prediction in apple. RADseq is a viable alternative to SNP array genotyping, while phenomic prediction is impractical. These findings offer valuable guidance for applying genomic selection in apple breeding, ultimately leading to the development of breeding material with improved quality.

Isolation and Molecular Detection of Antimicrobial Resistance Genes in Escherichia coli and Staphylococcus Species from Joint Ill Cases of Calves in Puducherry

Background: Joint ill is a septicemic polyarthritis condition due to the localization of pathogenic bacteria within the joints of young calves. The emergence of antibiotic resistant pathogen in such cases may lead to further complication in treatment. Thus, the present study was aimed to isolate and identify the bacterial pathogens and its antimicrobial resistant genes from joint ill cases in calves. Methods: A total of 31 pus swabs were collected from joint ill cases from calves and subjected for bacterial isolation and identification by phenotypic and genotypic method followed by determination of its antimicrobial resistant genes (tet and mecA gene) and antibiogram. Result: Based on colony characters, microscopic observation, biochemical tests, 17/31 (54.83%) Escherichia coli and 14/31 (45.16%) Staphylococcus aureus were isolated and identified phenotypically. All the isolates were further confirmed by polymerase chain reaction using E. coli and S. aureus species specific primers. The antimicrobial resistant genes carrying E. coli and S. aureus isolates were also detected, in which 4/17 (23.5%) isolates were positive for tet gene and 4/14 (28.57%) were positive for the mecA gene. The antibiotic susceptibility test showed that the isolates were highly sensitive to Enrofloxacin (100%) and Gentamicin (77%), but were resistant to Amoxyclav (70%) and tetracycline (50%). On conclusion, E. coli and S. aureus are the most common bacterial pathogens identified from this study. The presence of antimicrobial resistant genes (tet and mecA) and antibiogram pattern of the bacterial isolates indicating the possible treatment failure and serious public health risks in future.

Nested Multiplex Endpoint Polymerase Chain Reaction: An Alternative Method for Human Papilloma Virus Genotyping in Resource-limited Settings

Nested Multiplex Endpoint Polymerase Chain Reaction: An Alternative Method for Human Papilloma Virus Genotyping in Resource-limited Settings

Neuroendocrine Tumors: Germline Genetics and Hereditary Syndromes.

The vast majority of neuroendocrine 'neoplasms (NENs) are sporadic, although recent evidence has indicated that a subset of these cancers may also originate as a result of genetic germline mutations. To date, 10% of these cancers can be linked to an inherited genetic syndrome. Genetic diagnosis is crucial for patients with a suspected hereditary NEN syndrome, as it recognizes patients carrying germline mutations and allows for personalized clinical follow-up, considering the higher risk of developing other tumours. The potential for early genetic detection has significant implications for the treatment of patients with hereditary NEN syndrome, as it may facilitate the delivery of precision therapy that differs from that typically provided to other patients. Thus, the integration of genotypic and phenotypic diagnostic methods help clinicians to provide more informed treatment and to extend appropriate prevention to family members.

A next-generation sequencing-based universal target panel and algorithm for one-stop detection of copy number alterations and single-nucleotide variations in the HBB gene cluster for rapid diagnosis of β-thalassemia.

This study aimed to develop and validate a targeted next-generation sequencing (NGS) panel along with a data analysis algorithm capable of detecting single-nucleotide variants (SNVs) and copy number variations (CNVs) within the beta-globin gene cluster. The aim was to reduce the turnaround time in conventional genotyping methods and provide a rapid and comprehensive solution for prenatal diagnosis, carrier screening, and genotyping of β-thalassemia patients. We devised a targeted NGS panel spanning an 80.4 kb region on chromosome 11, encompassing the beta-globin gene cluster and 5' locus control region. We also developed an advanced data analysis algorithm consisting of variant calling and depth plot analysis that facilitates simultaneous detection of SNVs and CNVs in a single run. The test panel and algorithm were validated with 14 in-house β-thalassemia carrier/patient samples and cross-checked against the HbVar database. We identified seven pathogenic SNVs and five CNVs within the beta-globin gene cluster in various combinations, such as heterozygous, homozygous, and compound heterozygous conditions. Additionally, the coordinates of 169 rare deletions and 11 fusion mutations reported in the HbVar database were checked to verify the theoretical ability of our developed gene panel to detect all the CNVs within the target region. The developed panel and NGS technology can detect both SNVs and CNVs in a single run and can also be utilized for prenatal diagnosis and carrier screening for hemoglobinopathies, underscoring its versatility and clinical utility.

A large geno-spatial cluster of multi-drug resistant tuberculosis outbreak in a western district of Thailand.

The growing issue of drug resistance, particularly multidrug-resistant TB (MDR-TB), has exacerbated this problem. The rise of drug resistance TB is a severe global health concern. In Thailand, a persistent community outbreak of primary MDR-TB has been confirmed in the Tha Maka district of Kanchanaburi province, with an increasing prevalence of MDR-TB among newly diagnosed pulmonary tuberculosis cases. It was the first site in Thailand where a cluster of MDR-TB, caused by the Asian African 3 Modern Beijing strain, and XDR-TB, caused by L2.1, outbreaks were reported. This study aims to assess the MDR-TB outbreak in detail by characterizing the genomic profiles of the prevalent MDR-TB strains and examining their geographical distribution within the affected district. Through whole-genome sequencing (WGS) and bioinformatic analysis of 188 MTB isolates, the study identified three major phylogenetic lineages: the East Asian lineage (L2, 92 %), the Indo-Oceanic lineage (L1, 5.9 %), and the Euro-American lineage (L4, 2.1 %). The detailed sub-lineage distribution offers valuable insights into the predominant genetic clusters of M. tuberculosis within the sampled population. Notably, Lineage 2, specifically the L2.2.M3 sub-lineage, stood out as the dominant strain of MDR-TB, accounting for 77.7 % of the isolates. This finding underscores the significant prevalence of the L2.2.M3 sub-lineage and its potential role in the local transmission dynamics of tuberculosis. The high proportion and genetic homogeneity of the L2.2.M3 cluster among MDR-TB patients may indicate the strain's adaptation for more effective transmission within the Thai population. The increasing prevalence of this pathogenic strain could significantly impact tuberculosis control programs. Early diagnosis and contact tracing with chemotherapeutic preventive therapy for MDR-TB will be essential in inhibiting the spread and reactivation of these strains. Additionally, further studies are needed to prospectively identify transmission routes through contact tracing and real-time genotypic methods. It will also be crucial to ensure that future vaccines and/or recommended chemoprophylaxis therapy for MDR-TB will provide protection against these emerging strains.

Fast and accurate imputation of genotypes from noisy low-coverage sequencing data in bi-parental populations.

Genotyping of bi-parental populations can be performed with low-coverage next-generation sequencing (LC-NGS). This allows the creation of highly saturated genetic maps at reasonable cost, precisely localized recombination breakpoints (i.e., the crossovers), and minimized mapping intervals for quantitative-trait locus analysis. The main issues with these low-coverage genotyping methods are (1) poor performance at heterozygous loci, (2) high percentage of missing data, (3) local errors due to erroneous mapping of sequencing reads and reference genome mistakes, and (4) global, technical errors inherent to NGS itself. Recent methods like Tassel-FSFHap or LB-Impute are excellent at addressing issues 1 and 2, but nonetheless perform poorly when issues 3 and 4 are persistent in a dataset (i.e., "noisy" data). Here, we present a new algorithm for imputation of LC-NGS data that eliminates the need of complex pre-filtering of noisy data, accurately types heterozygous chromosomal regions, precisely estimates crossover positions, corrects erroneous data, and imputes missing data. The imputation of genotypes and recombination breakpoints is based on maximum-likelihood estimation. We compare its performance with Tassel-FSFHap and LB-Impute using simulated data and two real datasets. NOISYmputer is consistently more efficient than the two other software tested and reaches average breakpoint precision of 99.9% and average recall of 99.6% on illumina simulated dataset. NOISYmputer consistently provides precise map size estimations when applied to real datasets while alternative tools may exhibit errors ranging from 3 to 1845 times the real size of the chromosomes in centimorgans. Furthermore, the algorithm is not only highly effective in terms of precision and recall but is also particularly economical in its use of RAM and computation time, being much faster than Hidden Markov Model methods. NOISYmputer and its source code are available as a multiplatform (Linux, macOS, Windows) Java executable at the URL https://gitlab.cirad.fr/noisymputer/noisymputerstandalone/-/tree/1.0.0-RELEASE?reftype=tags.

Enhancing Thalassemia Diagnosis: Advantages of Third-Generation Sequencing.

This study aimed to evaluate the efficacy of third-generation sequencing (TGS) and a thalassemia (Thal) gene diagnostic kit in identifying Thal gene mutations. Blood samples (n = 119) with positive hematology screening results were tested using polymerase chain reaction (PCR)-based methods and TGS on the PacBio-Sequel-II-platform, respectively. Out of the 119 cases, 106 cases showed fully consistent results between the two methods, with TGS identified HBA1/2 and HBB gene mutations in 82 individuals. Notably, TGS exhibited a 5.04% higher detection rate compared to PCR-based methods (68.91% vs. 63.87%). For HBA1/2 mutations, TGS accurately detected three types of rare HBA1/2 mutations (--THAI, HBA2:c.34A>C, and HBA1:c.354_355insATC), two types of rare HBA compound mutations (ɑWSɑ/ɑCap+23(C>G)ɑ and -ɑ3.7/ɑIVS-Ⅱ-34ɑ), and three rare triplicates of α-globin variants (ɑɑ/ɑɑɑanti3.7, --SEA/HKɑɑ, and ɑɑ/ɑɑɑanti4.2). For the HBB gene, TGS detected two rare HBB mutations, namely HBB:c.316-45G>C and HBB:c.170G>A. For these 13 cases of rare thalassemia gene mutations, most patients exhibited varying degrees of microcytic hypochromia. However, patients with mutation in HBA2:c.34A>C, HBA1:c.354_355insATC, and HBB:c.170G>A did not exhibit typical results in blood routine tests but had abnormal hemoglobin composition. Additionally, TGS confirmed the cis/trans configuration of 2 allelic gene mutations in one step. Compared to traditional genotyping methods, TGS increased the detection rate of positive HB gene mutations and identified rare Thal cases with variable phenotypes. For Thal screening, it is recommended to perform both blood routine tests and hemoglobin electrophoresis, combined with TGS, to minimize the risk of missed or incorrect diagnoses in clinical practice. Although TGS is currently more expensive than other methods, it pro-vides a comprehensive approach for Thal screening and clinical diagnosis, particularly for rare Thal variants. As sequencing throughput increases and costs decrease, TGS can be widely applied in the screening of genetic diseases.

Carbapenem-Resistant Klebsiella pneumoniae: A comprehensive review of phenotypic and genotypic methods for detection

Carbapenem-Resistant Klebsiella pneumoniae: A comprehensive review of phenotypic and genotypic methods for detection

Genomic Selection and WssGWAS of Sheep Body Weight and Milk Yield: Imputing Low-Coverage Sequencing Data with Similar Genetic Background Panels.

Low-coverage whole-genome sequencing (LcWGS), a cost-effective genotyping method, offers greater flexibility in variant detection than does single-nucleotide polymorphism (SNP) chips. However, to our knowledge, no studies have explored the application of LcWGS in sheep. This study aimed to evaluate the feasibility of implementing LcWGS and genotype imputation and assess their applicability in genomic studies of body weight and milk yield in sheep. A total of 45,787 birth weight (BW), 31,135 weaning daily gain (WDG), 8,928 milk yield (MY), and 4,918 milk yield per unit of metabolic body weight (MWMY) data were analyzed. Among these, 2,366 sheep had imputed high-density genotypes. Simulated sequencing depths from 0.1 × to 3 × were imputed using the reference panels of 100-600 individuals. Genotype concordance with true data improved from 0.8875 to 0.9852 as the sequencing depth and panel size increased. The ssGBLUP method applied to the imputed data yielded higher accuracy for BW, WDG, MY, and MWMY than traditional ABLUP, notably increased MY accuracy from 0.61 to 0.66. Furthermore, a weighted single-step genome-wide association study identified key genes associated with BW (ANKS1B, OPRM1, CSMD1), WDG (TKDP5, GRP, RAX, IGFBP7), MY (CCSER1, FGGY, HOOK1), and MWMY (NDUFA10, ZNF385D, NWD1), revealing the importance of multiple pathways in sheep growth and milk production. This is the first study to assess the feasibility of combining LcWGS with genotype imputation for sheep genomic selection, balancing economic costs and imputation efficiency. Furthermore, we demonstrate an effective approach for identifying genetic variants linked to body weight and milk production, offering a cost-effective strategy for dairy sheep breeding.

Multiplex allele-specific PCR and DNA chromatography to identify genotypes of Salmonella enterica serovar Typhimurium and its monophasic variants.

We constructed a simple genotyping method combining multiplex allele-specific PCR and DNA chromatography for Salmonella enterica subsp. enterica serovar Typhimurium and its monophasic variant. The developed method can be used to estimate the genetic background of isolates, and it facilitates easy identification of several epidemic clades among these serotypes.

How I use noninvasive prenatal testing for red blood cell and platelet antigens.

Alloimmunization during pregnancy occurs when a mother produces antibodies against fetal antigens, leading to complications like hemolytic disease of the fetus and newborn (HDFN) and fetal and neonatal alloimmune thrombocytopenia (FNAIT). HDFN involves destruction of fetal red blood cells, potentially causing severe anemia, hydrops fetalis, and fetal death. FNAIT affects fetal platelets and possibly endothelial cells, resulting in risk of intracranial hemorrhage and brain damage. Traditional invasive methods for fetal antigen genotyping, like amniocentesis, carried miscarriage risks. The discovery of cell-free fetal DNA (cff-DNA) in maternal plasma enabled safe, non-invasive prenatal testing (NIPT). Initially used for RhD blood group typing, NIPT now covers various blood group antigens. Advances in technology have further enhanced NIPT's accuracy. Despite challenges such as low cff-DNA fractions and complex genetic variations, NIPT has become essential in managing alloimmunized pregnancies. In NIPT it is important to prevent both false positive results and false negative results. Particularly in the coming decades, more possibilities for personalized antenatal treatment for HDFN and FNAIT cases will become apparent and accurate NIPT blood group antigen typing results are crucial for guiding clinical decisions. In this paper we describe this journey and provide practical tools for the clinic.

Exploring phenotypic and genotypic diversity among methicillin-resistant, vancomycin-resistant, and sensitive Staphylococcus aureus

Background: Methicillin-Resistant Staphylococcus aureus (MRSA) is a global concern owing to the increasing prevalence of multidrug-resistant (MDR) strains. Vancomycin has been the primary treatment for MRSA; however, Vancomycin-resistant strains are being increasingly reported worldwide. Therefore, comparative studies are essential to support antimicrobial stewardship and improving clinical management. Ultimately, the findings from this study are expected to inform treatment strategies and guide public health interventions effectively. Material and methods: This study investigated the prevalence, antimicrobial resistance, and virulence characteristics of Vancomycin-sensitive S. aureus (VSSA) and Vancomycin-resistant S. aureus (VRSA) within MRSA strains. By employing a combination of phenotypic methods, such as antimicrobial susceptibility testing, and genotypic techniques, including molecular typing and identification of virulence genes, we obtained comprehensive insights into VRSA and VSSA profiles. Results: Of 250 clinical samples, 62 (24.8%) were S. aureus and 27 (43.5%) were identified as MRSA. All MRSA isolates exhibited MDR patterns. Most MRSA strains were VSSA (20/27, 74.1%), while 7 (25.9%) were VRSA. The VRSA isolates showed more antimicrobial resistance than VSSA isolates; however, the VRSA isolates had less virulence than VSSA isolates. Linezolid was the most effective treatment, with a 3.7% resistance rate. A higher percentage of biofilm-producing MRSA (96.3%) was confirmed by both phenotypic and genotypic methods. All isolates, except one VRSA, showed multi-virulence patterns (harbored more than 3 virulence genes). High diversity and low clonality (D-value = 0.99) were found in both VSSA and VRSA. Based on our correlation findings, the emergence of vancomycin resistance could modify the association between antimicrobial resistance and virulence, potentially affecting the pathogenic profile of these strains. The study also revealed complex interactions among host factors (including age and gender), sample origin, antimicrobial resistance, biofilm production, and virulence genes. Conclusion: This study highlights the alarming spread of MRSA and VRSA, which show significant resistance and virulence.

Combined role of PEAR1 and GP genes in the mechanism of aspirin resistance and its clinical significance

Exploring the mechanism of aspirin resistance helps to improve the prevention and treatment of ischemic stroke diseases and reduce the interference of resistance mechanism on the therapeutic effect of aspirin. In this paper, the Affiliated Hospital of Beihua university, together with two other hospitals, was chosen as the research unit, from which 260 AIS patients were selected for aspirin resistance study. The joint role of PEAR1 gene and GP gene in the mechanism of aspirin resistance was discussed by genotyping, TEG assay, univariate analysis, and binary logistic analysis methods. Among the GP1BA genes in AIS patients, the frequency of CC-type genes was as high as 92.03%, and the risk of aspirin resistance was significantly increased in CC-type relative to TT-type. The incidence of rs5924 allele T was significantly higher in the AR group than in the AS group (OR = 1.632, p = 0.006), and there was a significant effect of diabetic conditions on aspirin resistance at the 1% level. Diabetes significantly inhibits plasma proteins in AIS patients and makes aspirin less inhibitory to platelets. Therefore, clarifying the influence of PEAR1 and GP genes in the mechanism of aspirin resistance can be used to obtain the precise loci from the gene therapy point of view to enhance the disease treatment effect of aspirin.

A Cost-effective, High-throughput, Highly Accurate Genotyping Method for Outbred Populations.

Affordable sequencing and genotyping methods are essential for large scale genome-wide association studies. While genotyping microarrays and reference panels for imputation are available for human subjects, non-human model systems often lack such options. Our lab previously demonstrated an efficient and cost-effective method to genotype heterogeneous stock rats using double-digest genotyping-by-sequencing. However, low-coverage whole-genome sequencing offers an alternative method that has several advantages. Here, we describe a cost-effective, high-throughput, high-accuracy genotyping method for N/NIH heterogeneous stock rats that can use a combination of sequencing data previously generated by double-digest genotyping-by-sequencing and more recently generated by low-coverage whole-genome-sequencing data. Using double-digest genotyping-by-sequencing data from 5,745 heterogeneous stock rats (mean 0.21x coverage) and low-coverage whole-genome-sequencing data from 8,760 heterogeneous stock rats (mean 0.27x coverage), we can impute 7.32 million bi-allelic single-nucleotide polymorphisms with a concordance rate >99.76% compared to high-coverage (mean 33.26x coverage) whole-genome sequencing data for a subset of the same individuals. Our results demonstrate the feasibility of using sequencing data from double-digest genotyping-by-sequencing or low-coverage whole-genome-sequencing for accurate genotyping, and demonstrate techniques that may also be useful for other genetic studies in non-human subjects.