MTRR Gene Research Articles

The impact of single nucleotide polymorphisms in MTHFR and MTRR genes on disease activity and the presence of methotrexate treatment side effects in children with juvenile idiopathic arthritis.

To determine the impact of single nucleotide polymorphisms (SNPs) in MTHFR and MTRR genes on disease activity and the presence of MTX therapy adverse events in Polish children with juvenile idiopathic arthritis (JIA). SNP genotyping was performed using genomic DNA isolated from peripheral blood samples. Patients with MTHFR rs1801133 CT/TT variant had higher values of inflammatory markers, number of joints with active arthritis, and JADAS-71 value at the baseline of MTX treatment. Children with MTRR rs1801394 AG/AA variant presented higher inflammatory marker values at JIA diagnosis. MTHFR rs1801133 and MTRR rs1801394 polymorphisms are associated with higher disease activity at the moment of JIA diagnosis.

Genetic Polymorphisms of Gene Methionine Synthase Reductase (MTRR) and Risk of Urinary Bladder Cancer.

Methionine synthase reductase (MTRR) gene involved in the signaling for production of enzyme called methionine synthase reductase that use for the synthesis of methionine, which further used in DNA replication and repair. Genetic variation in MTRR gene may alter the susceptibility of developing urinary bladder cancer. The present study undertaken to identify the contribution of genetic polymorphisms in the MTRR gene on the selected polymorphic sites including c.66A>G and c.524C>T towards urinary bladder cancer risk. Direct-DNA sequencing method was applied for the observation of genotyping distribution of MTRR c.66A>G and c.524C>T polymorphisms in 232 histopathological confirmed cases of transitional cell carcinoma (TCC) of urinary bladder cancer and 250 age-, sex- and ethnicity-matched cancer free controls. With significant difference (p = 0.05) of genotype analysis further corresponding Odds ratio (OR) and 95% confidence interval (CI) were calculated. Multivariable logistic regression analysis was applied for adjusting significant confounder variables. Haploview software (version 4.2) was used to perform pairwise Linkage Disequilibrium (LD) analysis. Age (p = 0.01), Habit of smoking (p = 0.05), tobacco consumption (p = 0.001) and diet (p = 0.02) were significantly differed between cases and controls. Both the MTRR substitution showed higher risk of developing urinary bladder cancer (p = <0.001), although this effect alters in multivariable logistic regression analysis in a protective association for both the substitution. No LD observed between the c.66A>G and c.524C>T substitutions. In conclusion, MTRR c.66A>G and c.524C>T substitutions showed a joint effect with the other associated risk factors. Further studies with a greater number of subjects of different ethnicity and polymorphisms are recommended for the better understanding urinary bladder cancer etiology and to screen the population who are at higher risk of developing urinary bladder cancer.

Azithromycin Susceptibility Testing and Molecular Investigation of Neisseria gonorrhoeae Isolates Collected in Russia, 2020-2021.

The aim of this work was to study the resistance to macrolides (azithromycin) in the modern Russian population of N. gonorrhoeae with the analysis of genetic resistance determinants. Azithromycin is not used to treat gonococcal infection in Russia. However, among 162 isolates collected in 2020-2021, 22 isolates (13.6%) were phenotypically resistant to azithromycin. Mutations in 23S rRNA genes were found only in two isolates; erm and mefA genes were absent. Azithromycin resistance was shown to be predominantly associated with mutations in the mtrR and mtrD genes of the MtrCDE efflux pump and their mosaic alleles which may have formed due to a horizontal transfer from N. meningitidis. A total of 30 types of mtrR alleles and 10 types of mtrD alleles were identified including mosaic variants. Matching between the mtrR and mtrD alleles was revealed to indicate the cooperative molecular evolution of these genes. A link between the mtrR and mtrD alleles and NG-MAST types was found only for NG-MAST 228 and 807, typical of N. gonorrhoeae in Russia. The high level of resistance to azithromycin in Russia may be related to the spread of multiple transferable resistance to antimicrobials regardless of their use in the treatment of gonococcal infection.

Association of MTR and MTRR genes and oral health-related quality of life in children with dental caries.

This study aimed to assess whether genetic polymorphisms in MTR and MTRR are potential biomarkers of oral health-related quality of life (OHRQoL) in children with caries. A cross-sectional study was designed wherein pairs of parents/caregivers and children (aged two-five years) were selected. Clinical examination was used to detect dental caries, which were classified as low-severity and high-severity caries. The Early Childhood Oral Health Impact Scale (ECOHIS) questionnaire was used to assess OHRQoL. Genomic DNA extracted from the saliva was used to analyze two missense genetic polymorphisms: MTR (rs1805087) and MTRR (rs1801394). Mann-Whitney non-parametric test was used to analyze candidate genes with OHRQoL scale and domain, with a significance level of p≤0.05. MTR (rs1805087) was found associated (p = 0.05) with children's OHRQoL subscale scores in the dominant model (GG + AG). Genetic polymorphisms in MTR may increase the risk of poor OHRQoL in children with caries. Further studies are needed to investigate genetics, molecular factors, and OHRQoL.

MTHFR、MTRR基因多态性与不明原因复发性流产相关性的Meta分析

MTHFR、MTRR基因多态性与不明原因复发性流产相关性的Meta分析

325. Molecular Characterization of Antimicrobial Resistance Genes in Neisseria gonorrhoeae in Southeast Michigan

Abstract Background Antimicrobial resistance (AMR) in Neisseria gonorrhoeae (NG) is a public health crisis. Diagnosis of infection is primarily established using nucleic acid amplification technologies (NAATs) without susceptibility testing; thus rapid identification of resistance determinants is crucial. Whole-genome sequencing (WGS) is a promising alternative to current susceptibility methods. The purpose of this study was to describe AMR genes in a subset of isolates using WGS from Southeast Michigan. Methods Isolates, demographic data, and minimum inhibitory concentrations (MIC) via E-test (ceftriaxone/CRO) and broth microdilution (azithromycin/AZM) were obtained from the Michigan Department of Health and Human Services. Libraries from isolates were generated using QIAseq FX DNA library kit before sequencing on NovaSeq 6000. Reads were trimmed [Trimmomatic] and aligned to the NG genome [Burroughs-Wheeler Aligner] before processing with Samtools suite for variant detection and consensus WGS determination. AMRFinderPlus was used to categorize AMR genes and resistance-associated point mutations. Results Total of 38 isolates were analyzed. The majority were from males (63%) and Blacks (44.7%) living in Detroit City proper (47.3%) with median age 25 (range 13-56), unknown HIV status (55.2%) and sexual orientation (47.3%) (Table 1). Urine (47.3%) was the most common specimen source, followed by cervix/vagina (28.9%). More than a third had prior NG and other sexually transmitted infections. Eight resistance genes were found among the 38 isolates (Figure 1). The penA gene associated with cephalosporin resistance was found in all isolates; however, all isolates were susceptible to CRO (CLSI susceptible breakpoint MIC &amp;lt; 0.25). The mtrR gene associated with macrolide resistance was noted in 86.8% isolates. Among 33 isolates where AZM susceptibility was available, resistance was noted in 9 (27%) (CLSI resistance breakpoint MIC &amp;gt; 1). Conclusion We describe a potential utility for WGS to identify AMR genes in NG. Although numerous β-lactam resistance genes were detected in all isolates, all remained susceptible to CRO. Further research is needed to determine the extent to which genetic diversity affects phenotypic susceptibilities in NG. Disclosures All Authors: No reported disclosures.

Association of CBS 844ins68, MTR A2756G and MTRR A66G gene polymorphisms with depression: A population-based study from North India

BackgroundDepression is one of the major global health problem currently affecting about 350 million people worldwide. Its cause is a multi-factorial which involves a complex mechanism of social, physiological, and biological factors. Mutations of genes in one-carbon metabolic pathway such as cystathionine-β-synthase (CBS 844ins68), methyltransferase (MTR A2756G) and methyltransferase reductase (MTRR A66G) have been implicated in various neurological disorders. ObjectiveIn the present study, we attempted to understand the association of these gene polymorphisms with depression among a North Indian population. MethodsA total of 808 participants aged between 30‐ and 70 years of either sex were recruited through door-to-door household survey. Depression status was identified by administering BDI-II. Genotyping of the selected gene polymorphisms were performed following previously reported protocols. ResultsThe result revealed that only MTR A2756G gene polymorphism was significantly associated with mild depression [OR 1.65 (95 % CI: 0.05–2.62) p = 0.03] but not with moderate and severe depression among the studied population. Since CBS, MTR and MTRR genes are found in OCMP, MTR A2756G gene polymorphism was found to be associated with mild depression but not with CBS 844in68 and MTRR A66G gene polymorphisms in the present study. ConclusionIdentification of susceptible genes involved in depressive disorder and early screening for mild depression may create better opportunity for timely intervention and further improve the quality of life.

Identification of SNPs associated with methotrexate treatment outcomes in patients with early rheumatoid arthritis.

Methotrexate is one of the cornerstones of rheumatoid arthritis (RA) therapy. Genetic factors or single nucleotide polymorphisms (SNPs) are responsible for 15%-30% of the variation in drug response. Identification of clinically effective SNP biomarkers for predicting methotrexate (MTX) sensitivity has been a challenge. The aim of this study was to explore the association between the disease related outcome of MTX treatment and 23 SNPs in 8 genes of the MTX pathway, as well as one pro-inflammatory related gene in RA patients naïve to MTX. Categorical outcomes such as Disease Activity Score (DAS)-based European Alliance of Associations for Rheumatology (EULAR) non-response at 4 months, The American College of Rheumatology and EULAR (ACR/EULAR) non-remission at 6months, and failure to sustain MTX monotherapy from 12 to 24months were assessed, together with continuous outcomes of disease activity, joint pain and fatigue. We found that the SNPs rs1801394 in the MTRR gene, rs408626 in DHFR gene, and rs2259571 in AIF-1 gene were significantly associated with disease activity relevant continuous outcomes. Additionally, SNP rs1801133 in the MTHFR gene was identified to be associated with improved fatigue. Moreover, associations with p values at uncorrected significance level were found in SNPs and different categorical outcomes: 1) rs1476413 in the MTHFR gene and rs3784864 in ABCC1 gene are associated with ACR/EULAR non-remission; 2) rs1801133 in the MTHFR gene is associated with EULAR response; 3) rs246240 in the ABCC1 gene, rs2259571 in the AIF-1 gene, rs2274808 in the SLC19A1 gene and rs1476413 in the MTHFR gene are associated with failure to MTX monotherapy after 12-24months. The results suggest that SNPs in genes associated with MTX activity may be used to predict MTX relevant-clinical outcomes in patients with RA.

Experimental study of A66G-single nucleotide polymorphism in the MTRR gene and acute myeloid leukemia

Acute Myeloid leukemia (AML) is the most common leukemia among adult population which begins with the bone marrow, but it frequently spreads to the blood stream as well. One of the common polymorphisms in AML is A66G in Methionine Synthase Reductase (MTRR), which converts isoleucine into methionine residue in the protein chain, the methionine/homocysteine cycle is disrupted. Limited studies were documented between A66G polymorphism in MTRR gene with AML. This study aimed to investigate the A66G polymorphism in MTRR gene with AML patients in the Saudi population. Peripheral blood was collected from 100 AML patients and 100 controls and DNA was isolated. Using A66G primers, polymerase chain reaction was performed followed by restriction fragment length polymorphism analysis. The mean age of both cases and controls was found to be an average of 39 years. Allele (G vs A: OR-3.41 [95 %CI: 1.87–11.24; p = 0.0001) and genotype analysis (GG vs AA: OR-3.43 [95 %CI: 1.45–8.11]; p = 0.0004) has shown the association. In conclusion, A66G polymorphism has a strong genetic role in the AML patients in Saudi population.

Effects of genetic polymorphisms on methotrexate levels and toxicity in Chinese patients with acute lymphoblastic leukemia.

Methotrexate (MTX) has an antitumor effect when used for the treatment of acute lymphoblastic leukemia (ALL). This study aims at evaluating the associations between 14 polymorphisms of six genes involved in MTX metabolism with serum MTX concentration and toxicity accompanying high-dose MTX. Polymorphisms in 183 Chinese patients with ALL were analyzed using TaqMan single nucleotide polymorphism genotyping assay. The serum MTX concentration was determined using homogeneous enzyme immunoassay. MTX-related toxicities were also evaluated. Renal toxicity was significantly associated with higher serum MTX concentrations at 24, 48, and 72 hours, and MTX elimination delay (P = 0.001, P < 0.001, P < 0.001, and P < 0.001, respectively), whereas SLCO1B1 rs4149056 was associated with serum MTX concentrations at 48 and 72 hours, and MTX elimination delay in candidate polymorphisms (P = 0.014, P = 0.019, and P = 0.007, respectively). SLC19A1 rs2838958 and rs3788200 were associated with serum MTX concentrations at 24 hours (P = 0.016, P = 0.043, respectively). MTRR rs1801394 was associated with serum MTX concentrations at 72 hours (P = 0.045). Neutropenia was related to SLC19A1 rs4149056 (odds ratio [OR]: 3.172, 95% confidence interval [CI]: 1.310-7.681, P = 0.011). Hepatotoxicity was associated with ABCC2 rs2273697 (OR: 3.494, 95% CI: 1.236-9.873, P = 0.018) and MTRR rs1801394 (OR: 0.231, 95% CI: 0.084-0.632, P = 0.004). Polymorphisms of SLCO1B1, SLC19A1, ABCC2, and MTRR genes help predict higher risk of increased MTX levels or MTX-related toxicities in adult ALL patients.

The Roles of MTRR and MTHFR Gene Polymorphisms in Colorectal Cancer Survival.

Paradoxically epidemiological data illustrate a negative relationship between dietary folate intake and colorectal cancer (CRC) risk. The occurrence and progression of CRC may be influenced by variants in some key enzyme coding genes in the folate metabolic pathway. We investigated the correlation between genetic variants in methionine synthase reductase (MTRR) and methylenetetrahydrofolate reductase (MTHFR) and CRC survival. This study used data collected from the Newfoundland Familial Colorectal Cancer Study. A total of 532 patients diagnosed with CRC for the first time from 1999 to 2003 were enrolled, and their mortality were tracked until April 2010. DNA samples were genotyped by Illumina's integrated quantum 1 million chip. Cox models were established to assess 33 tag single-nucleotide polymorphisms in MTRR and MTHFR in relation to overall survival (OS), disease-free survival (DFS) and CRC-specific survival. The MTRR and MTHFR genes were associated with DFS and CRC-specific survival in CRC patients at the gene level. After multiple comparison adjustment, MTRR rs1801394 A (vs. G) allele was associated with increased DFS (p = 0.024), while MTHRT rs3737966 (G vs. A), rs4846049 (T vs. G), rs1476413 (A vs. G), rs1801131 (C vs. A), rs12121543 (A vs. C), rs1801133 (C vs. T), rs4846052 (T vs. C), rs2066471 (A vs. G) and rs7533315 (T vs. C) were related to worse CRC-specific survival. Additionally, significant interactions were seen among pre-diagnostic alcohol consumption with MTRR rs1801394, rs3776467, rs326124, rs162040, and rs3776455, with superior OS associated with those protective variant alleles limited to patients with alcohol consumption under the median. The MTHFR rs3737966 (G vs. A) allele seemed to be detrimental to CRC survival only among subjects with fruit intake below the median. Polymorphic variants in MTRR and MTHFR genes that code for key enzymes for folate metabolism may be associated with survival in patients with CRC. The gene-CRC outcome association seems modulated by alcohol drinking and fruit intake.

Association of affective disorders and MTHFR, MTR, and MTRR gene polymorphisms: preliminary results of a family study

Genetic polymorphisms associated with impaired one-carbon metabolism (1-CM) can be a risk factor not only for somatic and neurological diseases, but also for affective disorders (AD).Objective: to compare the frequency of genetic polymorphisms MTHFR, MTR, MTRR associated with 1-CM disorders among patients with AD, their blood relatives and healthy individuals.Patients and methods. This cross-sectional study of the frequency of genetic polymorphisms (MTHFR, MTR, MTRR) associated with 1-CM included patients with AD (n=24), their blood relatives (n=40), as well as a group of healthy individuals (n=35). All study participants underwent a structured diagnostic interview, as well as genetic analysis using real-time polymerase chain reaction.Results and discussion. Patients with AD were statistically more likely to carry the minor allele C of the 1298A&gt;C polymorphism of the MTHFR gene and the minor allele G of the 2756A&gt;G polymorphism of the MTR gene compared to the group of healthy individuals. The minor allele T of the 677C&gt;T polymorphism of the MTHFR gene was associated with longer depressive episodes, as well as with the presence of concomitant cardiovascular diseases in blood relatives of patients with AD.Conclusion. Genetic polymorphisms associated with 1-CM may contribute to familial aggregation of AD and somatic diseases. Further highquality family studies using molecular genetic methods are needed.

Molecular Mechanisms of Drug Resistance and Epidemiology of Multidrug-Resistant Variants of Neisseria gonorrhoeae.

The paper presents various issues related to the increasing drug resistance of Neisseria gonorrhoeae and the occurrence and spread of multidrug-resistant clones. One of the most important is the incidence and evolution of resistance mechanisms of N. gonorrhoeae to beta-lactam antibiotics. Chromosomal resistance to penicillins and oxyimino-cephalosporins and plasmid resistance to penicillins are discussed. Chromosomal resistance is associated with the presence of mutations in the PBP2 protein, containing mosaic variants and nonmosaic amino acid substitutions in the transpeptidase domain, and their correlation with mutations in the mtrR gene and its promoter regions (the MtrCDE membrane pump repressor) and in several other genes, which together determine reduced sensitivity or resistance to ceftriaxone and cefixime. Plasmid resistance to penicillins results from the production of beta-lactamases. There are different types of beta-lactamases as well as penicillinase plasmids. In addition to resistance to beta-lactam antibiotics, the paper covers the mechanisms and occurrence of resistance to macrolides (azithromycin), fluoroquinolones and some other antibiotics. Moreover, the most important epidemiological types of multidrug-resistant N. gonorrhoeae, prevalent in specific years and regions, are discussed. Epidemiological types are defined as sequence types, clonal complexes and genogroups obtained by various typing systems such as NG-STAR, NG-MAST and MLST. New perspectives on the treatment of N. gonorrhoeae infections are also presented, including new drugs active against multidrug-resistant strains.

Associations of variants of folate cycle genes with features of the clinical course of severe intraventricular hemorrhages in premature infants.

Summary data indicate that it has increased attention to the study of the role of the folate cycle and the genes encoding its key components in the complicated course of the neonatal period in premature infants. Therefore, the aim of our study was to investigate the relationship of folate cycle gene variants with the features of the neonatal course in premature infants with severe intraventricular hemorrhages (IVH). The study included 24 preterm infants with with IVHs of 3d and 4th degree that received standard clinical, laboratory and instrumental examination. Apgar scores at 1 and 5 minutes were significantly lower in patients with AA genotype according to variant A1298C of the MTHFR gene. The concentration of total protein on 6th day after birth was negatively correlated with the A66G variant of the MTRR gene. The mean concentration of ionized calcium in the first day after birth was higher in the subgroup of patients with the AA genotype (according to variant A1298C of the MTHFR gene). In the subgroup of patients requiring mechanical ventilation, the frequency of AA genotype according to variant A2756G of the MTR gene was significantly increased. The presence of respiratory disorders and oxygen dependence was negatively correlated with variant A1298C MTHFR. The day of surfactant administration was positively correlated with variant A1298C of the MTHFR gene. The results of this study indicate that gene variants MTHFR (C677T, A1298C), MTRR (A66G), MTR (A2756G), RFC1 (G80A) may affect the neonatal course in premature infants with severe IVH.

Different effects of maternal homocysteine concentration, MTHFR and MTRR genetic polymorphisms on the occurrence of fetal aneuploidy

Research questionDo maternal homocysteine (Hcy) concentrations, MTHFR and MTRR genes have effects on the occurrence of fetal aneuploidy? DesignA total of 619 aneuploidy mothers and 192 control mothers were recruited in this study. Differences in distributions of maternal MTHFR 677C>T, MTHFR 1298A>C and MTRR 66A>G genetic polymorphisms and maternal Hcy concentrations between aneuploidy mothers and control mothers were analysed. ResultsThe maternal MTHFR 677C>T polymorphism was found to be a risk factor for the occurrence of many fetal non-mosaic aneuploidies studied here, including trisomies 13, 15, 16, 18, 21, 22, TRA and TS. The maternal MTHFR 1298A>C polymorphism was found to be a risk factor specifically associated with the occurrence of fetal trisomy 15 and fetal TS. The maternal MTRR 66A>G polymorphism was found to be a risk factor only specifically associated with the occurrence of fetal trisomy 21. The Hcy concentrations of mothers of trisomies 22, 21, 18, 16, 15 and TS fetuses were significantly higher than the Hcy concentrations of control mothers. ConclusionsOverall, data suggested an association between these maternal polymorphisms and the susceptibility of fetal non-mosaic trisomy and Turner syndrome. However, these three maternal polymorphisms had different associations with the susceptibility of different fetal aneuploidies, and the elevated maternal Hcy concentration appeared to be a likely risk factor for fetal Turner syndrome and fetal trisomies.

Azithromycin resistance and its molecular characteristics in Neisseria gonorrhoeae isolates from a tertiary care centre in North India

Treatment guidelines for management of uncomplicated gonorrhoeae have been recently modified owing to alarming upsurge in azithromycin resistance. This study investigated the prevalence and genetic determinants of gonococcal azithromycin resistance in India. Four (5.7%) of 70 gonococcal isolates were resistant to azithromycin. Of 16 isolates investigated for molecular mechanisms of resistance, 13 (81.3%) and 6 (37.5%) isolates exhibited mutations in coding and promoter regions of mtrR gene, respectively. However, ermA, ermB and ermC genes or mutations in rrl gene were absent in all isolates. Azithromycin resistance is low in India posing no immediate threat to use of dual-therapy for syndromic management.

Evidence of Association between MTRR and TNF-α Gene Polymorphisms and Oral Health-Related Quality of Life in Children with Anterior Open Bite.

To assess the impact of AOB on OHRQoL in children and to evaluate whether MTR (rs1805087), MTRR (rs1801394), TGFβ1 (rs1800469) and TNF-α (rs1799964, rs1799724 and rs1800629) genes are potential biomarkers for OHRQoL in children with AOB. A cross-sectional study was performed with 173 children aged between 2-6 years. The Brazilian version of Early Childhood Oral Health Impact Scale (ECOHIS) was applied. Genetic polymorphisms were analyzed using real-time PCR. Mann-Whitney U-test and Chi-square were used. The overall mean ECOHIS scores were 5.49 (SD= 5.72) and 3.45 (SD = 4.49) (p < 0.01) in the AOB and control groups, respectively. Children with the CC genotype of TNF-α (rs1799724) had a significantly higher psychological QoL level. The MTRR AA genotype group showed a lower QoL level in the child subscale (p = 0.006), function (p = 0.017), and psychological (p = 0.006) domains. There was no significant difference between OHRQoL and the genetic polymorphisms in MTR and TGFβ1. Genetic polymorphisms in TNF-α and MTRR are associated with the impact on the OHRQoL in children with AOB.

Whole exome sequencing analysis of a patient with 45,X/46,XY mosaicism and autism spectrum disorder

To carry out genetic testing for a patient with 45,X/46,XY mosaicism and autism spectrum disorder (ASD). Peripheral blood samples of the patient and his parents were collected for the extraction of genomic DNA. Trio-based whole exome sequencing and Sanger sequencing were carried out thereafter. The proband and his father were found to harbor a heterozygous c.4781G>A (p.Arg1594Gln) variant of the CACNA1I gene. In addition, the proband was also found to harbor a de novo c.268C>T (p.Arg90Trp) missense variant of the MTRR gene. Based on guidelines of the American College of Medical Genetics and Genomics (ACMG), the c.4781G>A (p.Arg1594Gln) variant of the CACNA1I gene was predicted to be pathogenic (PVS1, PM1, PM2, PP3), while the c.268C>T (p.Arg90Trp) variant of the MTRR gene was predicted to be of uncertain significance. Variants of the CACNA1I and MTRR genes, together with the chromosomal mosaicism, may have predisposed to the susceptibility to the ASD in this patient.

Association of the DNA Methyltransferase and Folate Cycle Enzymes' Gene Polymorphisms with Coronary Restenosis.

Background: In recent years, the interest in genetic predisposition studies for coronary artery disease and restenosis has increased. Studies show that polymorphisms of genes encoding folate cycle and homocysteine metabolism enzymes significantly contribute to atherogenesis and endothelial dysfunction. The purpose of this study was to examine some SNPs of genes coding for folate cycle enzymes and DNA methyltransferases as risk factors for in-stent restenosis. Methods: The study included 113 patients after stent implantation and 62 patients without signs of coronary artery disease at coronary angiography as the control group. Real-time PCR and RFLP-PCR were applied to genotype all participants for MTHFR rs1801133, MTHFR rs1801131, MTR rs1805087, MTRR rs1801394, DNMT1 rs8101626, DNMT3B rs1569686, and DNMT3B rs2424913 gene polymorphisms. Statistical data processing was carried out using the R language and the SPSS Statistics 20 software. Results: Statistically significant differences in the DNMT3B gene polymorphisms were found between patients with and without in-stent restenosis. An association of TT rs1569686 and TT rs2424913 genotypes with the development of restenosis was revealed. The TT rs1569686 genotype was more frequent in the patients under the age of 65 years and in the subgroup of patients with post-12-month restenosis, as was the minor GG genotype for MTR rs1805087. The homozygous TT genotype for MTHFR rs1801133 was significantly more frequent in the subgroup over 65 years old. The frequencies of the heterozygous genotype for the MTRR gene and the minor GG homozygotes for the DNMT1 gene were significantly higher in the subgroup with in-stent restenosis under 65 years old. Conclusions: The results of this study could be used for a comprehensive risk assessment of ISR development, determining the optimal tactics and an individual approach in the treatment of patients with coronary artery disease before or after percutaneous coronary interventions, including homocysteine-lowering treatment in patients with hyperhomocysteinemia and a high risk of in-stent restenosis.

Hydrogel Droplet Microarray for Genotyping Antimicrobial Resistance Determinants in Neisseria gonorrhoeae Isolates.

A multiplex assay based on a low-density hydrogel microarray was developed to identify genomic substitutions in N. gonorrhoeae that determine resistance to the currently recommended treatment agents ceftriaxone and azithromycin and the previously used drugs penicillin, tetracycline, and ciprofloxacin. The microarray identifies 74 drug resistance determinants in the N. gonorrhoeae penA, ponA, porB, gyrA, parC, rpsJ, mtrR, blaTEM, tetM, and 23S rRNA genes. The hydrogel elements were formed by automated dispensing of nanoliter-volume droplets followed by UV-induced copolymerization of NH2-containing oligonucleotides with gel-forming monomers. Polybutylene terephthalate plates without special modifications were used as microarray substrates. Sequences and concentrations of immobilized oligonucleotides, gel composition, and hybridization conditions were carefully selected, and the median discrimination ratio ranged from 2.8 to 29.4, allowing unambiguous identification of single-nucleotide substitutions. The mutation identification results in a control sample of 180 N. gonorrhoeae isolates were completely consistent with the Sanger sequencing results. In total, 648 clinical N. gonorrhoeae isolates obtained in Russia during the last 5 years were analyzed and genotyped using these microarrays. The results allowed us to draw conclusions about the present situation with antimicrobial susceptibility of N. gonorrhoeae in Russia and demonstrated the possibility of using hydrogel microarrays to control the spread of antibiotic resistance.