Single Nucleotide Polymorphism Typing Research Articles

Whole-Genome Sequencing Analysis of Salmonella Enterica Serotype Enteritidis Isolated from Poultry Sources in South Korea, 2010-2017.

Salmonella enterica subsp. enterica serotype Enteritidis (SE) is recognized as a major cause of human salmonellosis worldwide, and most human salmonellosis is due to the consumption of contaminated poultry meats and poultry byproducts. Whole-genome sequencing (data were obtained from 96 SE isolates from poultry sources, including an integrated broiler supply chain, farms, slaughterhouses, chicken transporting trucks, and retail chicken meats in South Korea during 2010–2017. Antimicrobial resistance and virulence genes were investigated using WGS data, and the phylogenetic relationship of the isolates was analyzed using single-nucleotide polymorphism (SNP) typing and core genome multilocus sequence typing (cgMLST). All isolates carried aminoglycoside resistance genes, aac(6’)-Iaa, and 56 isolates carried multiple antimicrobial resistance genes. The most frequent virulence gene profile, pef-fim-sop-inv.-org-sip-spa-sif-fli-flg-hil-ssa-sse-prg-pag-spv, was found in 90 isolates. The SNP analysis provided a higher resolution than the cgMLST analysis, but the cgMLST analysis was highly congruent with the SNP analysis. The phylogenetic results suggested the presence of resident SE within the facility of processing plants, environments of slaughterhouses, and the integrated broiler supply chain, and the phylogenetically related isolates were found in retail meats. In addition, the SE isolates from different origins showed close genetic relationships indicating that these strains may have originated from a common source. This study could be valuable reference data for future traceback investigations in South Korea.

Značaj polimorfizama regulatornih i katalitičkih antioksidativnih proteina u hroničnoj bubrežnoj slabosti

Both excessive production of reactive oxygen species (ROS) and impaired antioxidant function are found in patients with chronic kidney disease (CKD). Therefore, individual susceptibility towards CKD can be induced by functional variations of genes encoding antioxidant regulatory (nuclear factor erythroid 2 - related factor 2 (Nrf2)) and catalytic (superoxide dismutase (SOD2) and glutathione peroxidase (GPX1)) proteins. Several types of single nucleotide polymorphisms (SNPs) have been found within the genes encoding these proteins, with Nrf2 (-617C/A), SOD2 (Ala16Val) and GPX1 (Pro198Leu) conferring impaired catalytic activity. The most unexplored gene polymorphism in CKD susceptibility, progression and survival, with only two original studies published, is the Nrf2 (-617C/A) polymorphism. The results of these studies showed that there was no individual impact of this polymorphism on the susceptibility towards end stage renal disease (ESRD) development, oxidative phenotype and mortality. However, Nrf2 had a significant role in ESRD risk and survival, when combined with other antioxidant genes. The results regarding the impact of SOD2 (Ala16Val) and GPX1 (Pro198Leu) polymorphisms on either CKD or ESRD are still inconclusive. Namely, some studies showed that patients having variant SOD2 (Val) or GPX1 (Leu) allele were at increased risk of CKD development and progression, while other studies reported only weak or no association between these polymorphisms and CKD. Surprisingly, the only study that reported an association of GPX1 polymorphism with overall/cardiovascular survival in ESRD patients showed a significant impact of low activity GPX1 (Leu/Leu) genotype on better survival. In this review, we comprehensively and critically appraise the literature on these polymorphisms related to oxidative stress in CKD patients, in order to identify gaps and provide recommendations for further clinical research and translation. New developments in the field of antioxidant polymorphisms in CKD patients could lead to better stratification of CKD patients, based on a prognostic antioxidant gene panel, and provide a more personalised medicine approach for the need of antioxidant therapy in these patients.

Obesity in Children with Leptin Receptor Gene Polymorphisms.

The study of single nucleotide polymorphisms (SNPs) of the leptin receptor gene (LEPR) based on next generation genomic sequencing (NGS) data is becoming an increasingly important aspect of diagnosis, treatment and prevention of both metabolically healthy (MHO) and metabolically unhealthy obesity (MUO) phenotypes. 35 obese children 6-18 years old were examined by the NGS method with bioinformatic analysis. The main group (n = 18) was formed by children with MUO, according to the recommendations of the expert group of the National Heart, Lung, and Blood Institute. The control group (n = 17) was represented by children with MHO. Statistical methods were used: analysis of variance, Wald's sequential analysis, Spearman's correlation analysis, analysis of nominal data and multiple discriminant analysis. 10 types of non-synonymous SNPs (rs3790435, rs1137100, rs2186248, rs70940803, rs79639154, rs1359482195, rs1137101, rs1805094, rs13306520, rs13306522) of the LEPR gene in obese children have been identified. Multiple discriminant analysis demonstrated that the following LEPR SNPs are of greatest importance in the development of MUO: rs3790435, rs13306522, rs13306520. Analysis of nominal data revealed significant differences in the groups for Copy number variation (CNV) rs3790435 of the LEPR gene. Wald's analysis allowed us to identify 6 important predictors of MUO (І ≥ 0.5): 2 CNV rs3790435 (Relative Risk, RR = 2, Prognostic coefficient, PC = +2.76); male gender of the child (RR = 1.3, PC = +1.35); rs3790435 (RR = 1.9, PC = +2.76); hyperleptinemia more than 40.56 ng/ml (RR = 2, PC = +3); CNV rs1359482195 ≥ 3 (RR = 1.9, PC = +5.8); SNP of the LEPR gene ≥4 (RR = 3.8, PC = +5.8). Children with the genotype rs3790435 gene LEPR had signs of MUO more often.

Resizing reaction volumes for the ForenSeq™ DNA Signature Prep kit library preparation.

The introduction of next generation sequencing (NGS; also known as massively parallel sequencing) technology in the field of forensic genetics has been welcomed by the scientific community, above all because it complements the weaknesses of capillary electrophoresis (CE) in the analysis of genetic markers, such as single nucleotide polymorphism (SNP) typing. However, one of the main obstacles to its adoption does not seem to be the cost of the instrumentation, but rather the cost of the NGS library preparation kits. With the aim of reducing the cost of library preparation without compromising the quality of the results, we tried to scale down reaction volumes for the first two polymerase chain reactions in the amplification and enrichment phases of the targeted loci of library preparation using the ForenSeq™ DNA Signature Prep kit. We used 1 µL templated DNA input to a concentration of 1 ng/µL, instead of the 5 µL at 0.2 ng/µL recommended by the manufacturer. Our findings indicate that reduction of the library preparation volume using the ForenSeq™ DNA Signature Prep kit did not interfere with the quality and reproducibility of the DNA profiles obtained and can help lower the overall cost of NGS.

Association Study between Proinflammatory Cytokine Gene Polymorphism and Risk of Recurrent Spontaneous Abortion

To investigate the correlation between inflammatory cytokine gene polymorphisms and the risk of recurrent miscarriage. A total of 120 patients with recurrent spontaneous abortion in our hospital between January 2019 and December 2020 were selected as the recurrent spontaneous abortion group and 120 healthy pregnant women with early pregnancy who visited the hospital during the same period were used as the control group and the genotype of interleukin-18 was detected by single nucleotide polymorphism typing technique and the validation method was Hardy Weinberg’s law of genetic equilibrium was used to verify the genotype of interleukin-18 and the relationship between interleukin-18 genotype and the risk of recurrent spontaneous abortion was analyzed by logistic method. The results showed that the level of interleukin-18 in the recurrent spontaneous abortion group was significantly higher than that in the control group (p 0.05); the interleukin-18 (rs1946519) and interleukin- 18(rs360717) in the control group The actual genotype frequencies of interleukin-18(rs1946519) and interleukin-18 (rs360717) in the control group were also not significantly different from the theoretical values (test values were 0.474 and 0.153, p>0.05); the genotypes of the rs1946519 and rs360717 loci showed significant differences and the recurrent spontaneous abortion group had significantly more A alleles than the control group (p<0.05); rs1946519 and rs360717 locus mutant cases showed significant differences in serum interleukin-17 levels, with AA and TT genotypes showing higher concentrations than other genotypes, respectively (p<0.05); the relative risk of recurrent spontaneous abortion risk in rs1946519 AG and AA gene carriers was 2.468 and 2.468 times higher than that of the GG allele, respectively. The relative risk of recurrent spontaneous abortion in rs360717 TC and TT gene carriers was 1.973 and 1.545 times higher than that of CC allele, respectively and all of them had higher risk of recurrent spontaneous abortion (p<0.05). The rs1946519 polymorphic allele G and rs360717 allele C mutations of interleukin-18 affect the serum hormone levels of patients and lead to an increased risk of recurrent spontaneous abortion in pregnant women. Therefore, interleukin-18 gene polymorphisms may be an important basis for the pathogenesis of recurrent spontaneous abortion and its treatment.

Genotyping of Francisella tularensis subsp. holarctica from Hares in Germany.

Francisella tularensis is the causative agent of the zoonotic disease tularemia. In Germany, most human infections are caused by contact with infected hares. The aim of this study was to characterize Francisella tularensis subsp. holarctica strains isolated from hares in Germany and to develop bioinformatics tools to analyze their genetic relatedness. In total, 257 German isolates—obtained mainly from hares (n = 233), other vertebrate animals, and ticks, but also from humans (n = 3)—were analyzed within this study. Publically available sequence data from 49 isolates were used to put our isolates into an epidemiological context and to compare isolates from natural foci and humans. Whole-genome sequences were analyzed using core-genome Multi-Locus-Sequence-Typing, canonical Single Nucleotide Polymorphism (SNP) typing and whole-genome SNP typing. An overall conformity of genotype clustering between the typing methods was found, albeit with a lower resolution for canonical single SNP typing. The subclade distribution, both on local and national levels, among strains from humans and hares was similar, suggesting circulation of the same genotypes both in animals and humans. Whilst close to identical isolates of the same subclade were found distributed over large areas, small geographical foci often harbored members of different subclades. In conclusion, although genomic high-resolution typing was shown to be robust, reproducible and allowed the identification of highly closely related strains, genetic profiling alone is not always conclusive for epidemiological linkage of F. tularensis strains.

Estimation of Variance Components and Genomic Prediction for Individual Birth Weight Using Three Different Genome-Wide SNP Platforms in Yorkshire Pigs.

Simple SummaryThe individual birth weight (IBW) of pigs is an important trait regarding its relevance to mortality at weaning, sow prolificacy, and growth performance. This study investigates the variance component estimation, informative window regions, and the efficiency of genomic predictions associated with IBW traits in Yorkshire pigs. The low heritability (0.13) is estimated on the basis of a full model including individual genetic, sow genetic, and common environmental effects. Two common window regions of the genome are identified under three different genotyping platforms found within the ARAP2 and TSN genes concerning the IBW trait. The genomic prediction ability is improved using deregressed estimated breeding values including parental information as a response variable despite Bayesian methods and genotyping platforms for the IBW trait in Korean Yorkshire pigs.This study estimates the individual birth weight (IBW) trait heritability and investigates the genomic prediction efficiency using three types of high-density single nucleotide polymorphism (SNP) genotyping panels in Korean Yorkshire pigs. We use 38,864 IBW phenotypic records to identify a suitable model for statistical genetics, where 698 genotypes match our phenotypic records. During our genomic analysis, the deregressed estimated breeding values (DEBVs) and their reliabilities are used as derived response variables from the estimated breeding values (EBVs). Bayesian methods identify the informative regions and perform the genomic prediction using the IBW trait, in which two common significant window regions (SSC8 27 Mb and SSC15 29 Mb) are identified using the three genotyping platforms. Higher prediction ability is observed using the DEBV-including parent average as a response variable, regardless of the SNP genotyping panels and the Bayesian methods, relative to the DEBV-excluding parent average. Hence, we suggest that fine-mapping studies targeting the identified informative regions in this study are necessary to find the causal mutations to improve the IBW trait’s prediction ability. Furthermore, studying the IBW trait using a genomic prediction model with a larger genomic dataset may improve the genomic prediction accuracy in Korean Yorkshire pigs.

Chickens productivity selection affects immune system genes.

The quantitative trait loci associated with the immune properties of chickens are of interest from the point of view of obtaining animals resistant to infectious agents using marker-assisted selection. In the process of selecting markers for genomic selection in broiler-type chickens, a non-standard genotype frequency of the RACK1 gene allele (SNP Gga_rs15788101) in the B5 line of broiler-type chicken cross Smena 8 was identified and it was suggested that this gene was involved in selection. Therefore, it was decided to investigate the available polymorphisms in the three genes responsible for the IgY titer (DMA, RACK1 and CD1B). Molecular typing of single nucleotide polymorphisms of three loci revealed an approach to fixation of the unfavorable allele of the DMA gene (SNP Gga_rs15788237), an approach to fixation of the unfavorable allele of the RACK1 gene and the prevalence of the favorable CD1B gene allele (SNP Gga_rs16057130). Analysis of the haplotypes revealed a strong linkage disequilibrium of these genes. This suggests that these genes experience selection pressure. Analysis of the protein-coding sequences of the CD1B and DMA genes of various breeds of chickens revealed a negative selection of these genes. In order to understand whether the fixation of the studied alleles is the result of artificial selection of the B5 line of the cross Smena 8, an analysis of similar loci in layer chickens Hisex White was carried out. The frequencies of the alleles at the loci of the CD1B gene (Gga_rs16057130) and the RACK1 gene (Gga_rs15788101) in the Hisex White chicken genome differ from the frequencies of the alleles obtained for chickens of the B5 line of the cross Smena 8. It can be assumed that the fixation of the allele in the DMA gene (SNP Gga_rs15723) is associated with artificial or natural selection, consistent in broilers and layers. Changes in the loci Gga_rs16057130 and Gga_rs15788101 in the B5 line of the Smena 8 chickens are most likely associated with artificial selection of broiler productivity traits, which can subsequently lead to fixation of alleles at these loci. Artificial breeding of chickens leads to degradation of the variability of genes encoding elements of the immune system, which can cause a decrease in resistance to various diseases. The study of the negative impact of selection of economic traits on immunity should provide means to mitigate negative consequences and help find ways to obtain disease-resistant animals.

In Silico Tools and Approaches for the Prediction of Functional and Structural Effects of Single-Nucleotide Polymorphisms on Proteins: An Expert Review

Single-nucleotide polymorphisms (SNPs) are single-base variants that contribute to human biological variation and pathogenesis of many human diseases. Among all SNP types, nonsynonymous single-nucleotide polymorphisms (nsSNPs) can alter many structural, biochemical, and functional features of a protein such as folding characteristics, charge distribution, stability, dynamics, and interactions with other proteins/nucleotides. These modifications in the protein structure can lead nsSNPs to be closely associated with many multifactorial diseases such as cancer, diabetes, and neurodegenerative diseases. Predicting structural and functional effects of nsSNPs with experimental approaches can be time-consuming and costly; hence, computational prediction tools and algorithms are being widely and increasingly utilized in biology and medical research. This expert review examines the in silico tools and algorithms for the prediction of functional or structural effects of SNP variants, in addition to the description of the phenotypic effects of nsSNPs on protein structure, association between pathogenicity of variants, and functional or structural features of disease-associated variants. Finally, case studies investigating the functional and structural effects of nsSNPs on selected protein structures are highlighted. We conclude that creating a consistent workflow with a combination of in silico approaches or tools should be considered to increase the performance, accuracy, and precision of the biological and clinical predictions made in silico.

Insights on SNP types, detection methods and their utilization in Brassica species: Recent progress and future perspectives

The genus Brassica, family Brassicaceae (Cruciferae), comprises many important species of oil crops, vegetables and medicinal plants including B. rapa, B. oleracea, B. nigra, B. napus, B. juncea, B. carinata. Genomic researches in Brassica species is constrained by polyploidization, mainly due to its complicated genomic structure. However, rapid development of methods for detecting single nucleotide polymorphisms (SNP), such as next generation sequencing and SNP microarray, has accelerated release of reference Brassica species genomes as well as discovery of large numbers and genome-wide SNPs, thus intensifying forward genetics in this genus. In this review, we summarize biological characteristics, classification and various methods for detecting SNPs, focusing on high-throughput techniques. Moreover, we describe the pivotal roles of SNPs in genetic diversity, linkage map construction and QTL mapping, comparative genomics, linkage disequilibrium and genome-wide association studies. These insights are expected to deepen our understanding and guide further advancements in Brassica species research.

Mining Listeria monocytogenes single nucleotide polymorphism sites to identify the major serotypes using allele-specific multiplex PCR

Worldwide, Listeria monocytogenes is a common foodborne pathogen and serotyping is necessary for traceability and surveillance. Due to high accuracy and speed, PCR is commonly used for serotyping by targeting specific genes, such as lmo1118 and ORF2110 for 1/2c-3c and 4b-4d-4e, respectively. Single nucleotide polymorphisms (SNPs) are single base alterations at specific loci which are associated with various phenomena. In this study, a method was explored to mine specific SNPs from 253 L. monocytogenes genomes with known serotypes by writing Python programming language script. After blasting all the CDS, seventeen candidate genes with specific SNPs were obtained and these genes contained multiple types of SNPs, including lineages I, II, III, 1/2a-3a, 1/2b-3b-7, and 1/2c-3c specific SNPs. The sensitivity and specificity of these candidate SNP sites are higher than 85% in the genomes examined. Combining lineage I-specific, lineage II-specific, 1/2b-3b-7-specific, and 1/2c-3c-specific SNP sites together, using allele-specific multiplex PCR, we could serotype major L. monocytogenes serotypes. This method was used for 60 L. monocytogenes strains isolated from various food samples and the serotyping results were 100% identical to those of the currently accepted method with the reduced primers from ten to eight. Our results indicate that allele-specific multiplex PCR after mining specific SNPs from common genome database can be used for bacterial typing.

Utilization of Clustered Regularly Interspaced Short Palindromic Repeats to Genotype Escherichia coli Serogroup O80.

The hypervariable nature of clustered regularly interspaced short palindromic repeats (CRISPRs) makes them valuable biomarkers for subtyping and epidemiological investigation of Escherichia coli. Shiga toxin-producing E. coli (STEC) serogroup O80 is one hybrid pathotype that is emerging recently in Europe and is involved in hemolytic uremic syndrome with bacteremia. However, whether STEC O80 strains can be genotyped using CRISPR has not been evaluated. In this study, we aimed to characterize the genetic diversity of 81 E. coli serogroup O80 isolates deposited in the National Center for Biotechnology Information databases using CRISPR typing and to explore the association between virulence potential and CRISPR types (CTs). A total of 21 CTs were identified in 80 O80 strains. CRISRP typing provided discrimination with variants of a single serotype, which suggested a stronger discriminatory power. Based on CRISPR spacer profiles, 70 O80:H2 isolates were further divided into four lineages (lineage LI, LII, LIII, and LIV), which correlated well with whole-genome single nucleotide polymorphisms typing and virulence gene profiles. Moreover, the association between CRISPR lineages and virulence gene profiles hinted that STEC O80:H2 strains may originate from O80:H19 or O80:H26 and that lineage LI may have been evolved from lineage LII. CT2 and CT13 were shared by human and cattle isolates, suggesting that there might be the potential transmission between cattle and human. Collectively, CRISPR typing is one technology that can be used to monitor the transmission of STEC O80 strains and provide new insights into microevolution of serogroup O80.

Molecular epidemiology of resistance to antimalarial drugs in the Greater Mekong subregion: an observational study

SummaryBackgroundThe Greater Mekong subregion is a recurrent source of antimalarial drug resistance in Plasmodium falciparum malaria. This study aimed to characterise the extent and spread of resistance across this entire region between 2007 and 2018.MethodsP falciparum isolates from Myanmar, Thailand, Laos, and Cambodia were obtained from clinical trials and epidemiological studies done between Jan 1, 2007, and Dec 31, 2018, and were genotyped for molecular markers (pfkelch, pfcrt, pfplasmepsin2, and pfmdr1) of antimalarial drug resistance. Genetic relatedness was assessed using microsatellite and single nucleotide polymorphism typing of flanking sequences around target genes.Findings10 632 isolates were genotyped. A single long pfkelch Cys580Tyr haplotype (from −50 kb to +31·5 kb) conferring artemisinin resistance (PfPailin) now dominates across the eastern Greater Mekong subregion. Piperaquine resistance associated with pfplasmepsin2 gene amplification and mutations in pfcrt downstream of the Lys76Thr chloroquine resistance locus has also developed. On the Thailand–Myanmar border a different pfkelch Cys580Tyr lineage rose to high frequencies before it was eliminated. Elsewhere in Myanmar the Cys580Tyr allele remains widespread at low allele frequencies. Meanwhile a single artemisinin-resistant pfkelch Phe446Ile haplotype has spread across Myanmar. Despite intense use of dihydroartemisinin–piperaquine in Kayin state, eastern Myanmar, both in treatment and mass drug administrations, no selection of piperaquine resistance markers was observed. pfmdr1 amplification, a marker of resistance to mefloquine, remains at low prevalence across the entire region.InterpretationArtemisinin resistance in P falciparum is now prevalent across the Greater Mekong subregion. In the eastern Greater Mekong subregion a multidrug resistant P falciparum lineage (PfPailin) dominates. In Myanmar a long pfkelch Phe446Ile haplotype has spread widely but, by contrast with the eastern Greater Mekong subregion, there is no indication of artemisinin combination therapy (ACT) partner drug resistance from genotyping known markers, and no evidence of spread of ACT resistant P falciparum from the east to the west. There is still a window of opportunity to prevent global spread of ACT resistance.FundingThailand Science Research and Innovation, Initiative 5%, Expertise France, Wellcome Trust.

A quasimetagenomics method for concerted detection and subtyping of Salmonella enterica and E. coli O157:H7 from romaine lettuce

Quasimetagenomics refers to the sequencing of a modified food microbiome to facilitate combined detection and subtyping of targeted pathogens in a single workflow. Through quasimetagenomic sequencing, pathogens are detected and subtyped in a shortened time frame compared to traditional culture enrichment and whole genome sequencing-based analyses. While this method was previously used to detect and subtype Salmonella enterica from chicken, iceberg lettuce, and black pepper, it has not been applied to investigate multiple pathogens in one workflow. A quasimetagenomic method to concertedly detect and subtype Salmonella enterica and Escherichia coli O157:H7 from artificially contaminated romaine lettuce in a single workflow was developed. All quasimetagenomic samples with initial target pathogen inoculum levels of ~1 CFU/g were detected and serotyped after co-enrichment of the two pathogens for 12 h. Single nucleotide polymorphism typing was achievable for some initial pathogen inoculum levels as low as ~0.1 CFU/g. Our results suggest that this method can be used for concerted detection and subtyping of multiple bacterial pathogens from romaine lettuce even at low contamination levels.

Socru: typing of genome-level order and orientation around ribosomal operons in bacteria.

Rearrangements of large genome fragments occur in bacteria between repeat sequences and can impact on growth and gene expression. Homologous recombination resulting in inversion between indirect repeats and excision/translocation between direct repeats enables these structural changes. One form of rearrangement occurs around ribosomal operons, found in multiple copies across many bacteria, but identification of these rearrangements by sequencing requires reads of several thousand bases to span the ribosomal operons. With long-read sequencing aiding the routine generation of complete bacterial assemblies, we have developed socru, a typing method for the order and orientation of genome fragments between ribosomal operons. It allows for a single identifier to convey the order and orientation of genome-level structure and we have successfully applied this typing to 433 of the most common bacterial species. In a focused analysis, we observed the presence of multiple structural genotypes in nine bacterial pathogens, underscoring the importance of routinely assessing this form of variation alongside traditional single-nucleotide polymorphism (SNP) typing.

Association of gene polymorphisms of CD55 with susceptibility to and severity of hand, foot, and mouth disease caused by enterovirus 71 in the Han Chinese population.

Hand, foot, and mouth disease (HFMD) caused by enterovirus 71 (EV71) can lead to high morbidity and mortality, and genetic background plays an important role during the disease process. We investigated the association between the single-nucleotide polymorphism (SNP) rs2564978 of the CD55 gene and susceptibility to and severity of HFMD using the SNPs can multiple SNP typing methods. Soluble CD55 (sCD55) expression was significantly lower in the EV71 HFMD group than in the control group and lower in severe cases than in mild cases (P < .001). Moreover, CD55 rs2564978 (C vs T OR = 1.300, 95% CI, 1.120-1.509) was associated with the risk of EV71 infection, and genotype TC was related to the severity of the infection (TC vs TT OR = 4.523, 95% CI, 2.033-10.066). Our results suggest that sCD55 expression and the CD55 polymorphism rs2564978 may influence the susceptibility to and severity of EV71 infection.

Genetic Polymorphisms of Cytokines Might Affect Postoperative Sufentanil Dosage for Analgesia in Patients.

ObjectiveTo explore the effect of genetic polymorphisms of cytokines on the dosage of sufentanil for patient-controlled intravenous analgesia (PCIA) after radical lung cancer surgery.MethodsA total of 100 patients, aged 18 years and above, with ASA grade Ⅰ-Ⅱ and body mass index (BMI) 18.5 to 30, and who were scheduled for radical lung cancer surgery under total intravenous anaesthesia with PCIA of sufentanil from September 2015 to March 2016, were selected. DNA was collected from peripheral blood samples before surgery, and the iMLDRTM multiple single-nucleotide polymorphism typing kit was used to detect 16 related single-nucleotide polymorphism (SNP) sites of interleukin-1A (IL-1A), interleukin-1β (IL-1β), interleukin-1RN (IL-1RN), interleukin-6 (IL-6), C-X-C motif chemokine ligand 8 (CXCL8), interleukin-10 (IL-10), tumour necrosis factor (TNF), nuclear factor kappa-B1 (NFκB1), REL (REL proto-oncogene, NF-kB subunit), and nuclear factor kappa-B inhibitor alpha (NFκBIA). The general characteristics of patients, surgery and anaesthesia data, postoperative resting VAS pain scores, postoperative opioid dosages of sufentanil for PCIA and opioid-related adverse events were recorded. The effects of the examined genetic polymorphisms of the cytokines on the dosage of sufentanil were analysed.ResultsEight of 100 patients withdrew for various reasons, and, eventually, 92 patients were included. The patients’ resting visual analogue scale (VAS) scores at 24 h, 48 h, and 72 h after surgery were 2.3 ± 1.2, 2.0 ± 0.9, and 1.9 ± 1.0, respectively. The total amounts of sufentanil used were 34.7 ± 10.5 μg, 65.2 ± 13.7 μg, and 94.7 ± 11.6 μg, respectively. We found that the TT genotype of NFκBIA rs696 had higher PCIA sufentanil dosages than the CC genotype and the CT genotype at 48–72 h postoperation (p=0.023, p=0.025, respectively).ConclusionThe genetic polymorphisms of the cytokine NFκBIA rs696 might affect the dosage of sufentanil for PCIA after radical lung cancer surgery. The specific mechanism needs further study.

Population Genomics of Mycobacterium leprae Reveals a New Genotype in Madagascar and the Comoros.

Human settlement of Madagascar traces back to the beginning of the first millennium with the arrival of Austronesians from Southeast Asia, followed by migrations from Africa and the Middle East. Remains of these different cultural, genetic, and linguistic legacies are still present in Madagascar and other islands of the Indian Ocean. The close relationship between human migration and the introduction and spread of infectious diseases, a well-documented phenomenon, is particularly evident for the causative agent of leprosy, Mycobacterium leprae. In this study, we used whole-genome sequencing (WGS) and molecular dating to characterize the genetic background and retrace the origin of the M. leprae strains circulating in Madagascar (n = 30) and the Comoros (n = 3), two islands where leprosy is still considered a public health problem and monitored as part of a drug resistance surveillance program. Most M. leprae strains (97%) from Madagascar and Comoros belonged to a new genotype as part of branch 1, closely related to single nucleotide polymorphism (SNP) type 1D, named 1D-Malagasy. Other strains belonged to the genotype 1A (3%). We sequenced 39 strains from nine other countries, which, together with previously published genomes, amounted to 242 genomes that were used for molecular dating. Specific SNP markers for the new 1D-Malagasy genotype were used to screen samples from 11 countries and revealed this genotype to be restricted to Madagascar, with the sole exception being a strain from Malawi. The overall analysis thus ruled out a possible introduction of leprosy by the Austronesian settlers and suggests a later origin from East Africa, the Middle East, or South Asia.

Exploring a multiplex DNA methylation-based SNP typing method for body fluids identification: As a preliminary report

In forensic investigation, identification of the cellular origin from body fluid can be essential in the crime scene reconstruction. Recently, DNA methylation could potentially be used as a novel marker for body fluid identification. The simultaneous analysis of CpGs and neighboring single nucleotide polymorphisms (SNPs) has been proposed as an efficient assay for body fluids identification. In this study, a multiplex DNA methylation-based SNP typing system was developed. The specificity, sensitivity and detectability in mixtures and degraded samples were explored in our study. As results, four DNA methylation-based semen-specific SNP (SE1-4) showed good specificity, but two markers associative with saliva (SA1) and vaginal fluid (VA3) was observed cross-reactivity sporadically. Interesting, VA3 were found only presented in the female which may be useful for sexual identification. Moreover, this multiplex system successfully amplification in mixtures and aged samples which proves it be used as a valuable protocol in the identification of actual forensic samples. The strategy indicated that the approach was suitable and reliable for the body fluids analysis in mix stains in Han Chinese for forensic purposes.

Large-scale identification of human bone remains via SNP microarray analysis with reference SNP database

Single nucleotide polymorphisms (SNPs) are valuable markers complementary to conventional forensic short tandem repeat (STR) markers in genetic typing, with potential advantages in challenging forensic casework. With the advent of high-throughput technologies, such as microarrays and massively parallel sequencing, the use of SNP typing has now expanded to large-scale forensic applications. Herein, a forensic case is presented to demonstrate the usefulness of SNP typing in identifying large-scale human bone remains with reference database construction. A total of 402 bone remains were recovered from an island in the Jeju Province of Korea where a massive disaster occurred in 1948. The first phase of the identification process was accomplished via conventional DNA typing methods including autosomal and Y-chromosomal STR typing, and mitochondrial DNA sequencing, which resulted in the identification of 74 of 402 remains. The second phase of the identification involved the remaining 327 unidentified remains using SNP typing as a supplementary tool based on Affymetrix resequencing array. The SNP markers of 782 family members were also analyzed and a reference database was constructed for comparison. An additional 51 bone remains were identified in the second phase. SNP data obtained from the supplementary genotyping yielded additional genetic information as well as contributed to kinship testing to determine the second degrees of relationship. In addition SNPs are useful in discriminating ambiguous relationship when only STR data are available. A software program developed for SNP typing system enabled efficient kinship analysis for large-scale forensic identification. The results and the casework are described and discussed.