Single Nucleotide Polymorphism Analysis Research Articles

Whole-genome sequencing reveals genomic characterization of Listeria monocytogenes from food in China.

Listeria monocytogenes is a foodborne bacterium that could persist in food and food processing environments for a long time. Understanding the population structure and genomic characterization of foodborne L. monocytogenes is essential for the prevention and control of listeriosis. A total of 322 foodborne L. monocytogenes isolates from 13 geographical locations and four food sources in China between 2000 and 2018 were selected for whole-genome sequencing. In silico subtyping divided the 322 isolates into five serogroups, 35 sequence types (STs), 26 clonal complexes (CCs) and four lineages. Serogroup IIa was the most prevalent serogroup and ST9 was the most prevalent ST of foodborne L. monocytogenes strains isolated in China. The in-depth phylogenetic analysis on CC9 revealed that ST122 clone might be original from ST9 clone. Furthermore, 23 potentially relevant clusters were identified by pair-wised whole-genome single nucleotide polymorphism analysis, indicating that persistent- and/or cross-contamination had occurred in markets in China. ST8 and ST121 were the second and third top STs of L. monocytogenes in China, which had heterogeneity with that of L. monocytogenes isolates from other countries. The antibiotic resistance genes aacA4, tetM, tetS, dfrG carried by different mobile elements were found in L. monocytogenes strains. One lineage II strain carrying Listeria Pathogenicity Island 3 was first reported. In addition, a novel type of premature stop codon in inlA gene was identified in this study. These findings revealed the genomic characteristics and evolutionary relationship of foodborne L. monocytogenes in China on a scale larger than previous studies, which further confirmed that whole-genome sequencing analysis would be a helpful tool for routine surveillance and source-tracing investigation.

Recombination and Mutation in a New HP-PRRSV Strain (SD2020) from China.

A new HP-PRRSV strain (SD2020) was isolated from pigs with suspected highly pathogenic porcine reproductive and respiratory syndrome disease in a pig farm in Shandong Province, China, and its genome was sequenced. This pig farm has been using the VR-2332 vaccine strain to immunize pigs for a long time. The phylogenic and single nucleotide polymorphisms (SNPs) analysis of the viruses isolated from dead pigs showed that SD2020 was a natural recombinant virus of the VR-2332 vaccine strain and the JXA1 similar strain, and that two splicing fragments highly homologous to JXA1 in the virus genome were probably derived from the JXA1 wild strain and JXA1-R vaccine strain, respectively. Therefore, the possible recombination events of SD2020 and its mutation site might be related to high pathogenicity.

Clostridioides difficile in South American Camelids in Germany: First Insights into Molecular and Genetic Characteristics and Antimicrobial Resistance.

Little is known about zoonotic pathogens and their antimicrobial resistance in South American camelids (SAC) in Germany including Clostridioides (C.) difficile. The aim of this study was to investigate prevalence, molecular characteristics and antimicrobial resistance of C. difficile in SAC. Composite SAC faecal samples were collected in 43 husbandries in Central Germany and cultured for C. difficile. Toxinotyping and ribotyping was done by PCR. Whole genome sequencing was performed with Illumina® Miseq™. The genomes were screened for antimicrobial resistance determinants. Genetic relatedness of the isolates was investigated using core genome multi locus sequence typing (cgMLST) and single nucleotide polymorphism analysis. Antimicrobial susceptibility testing was done using the Etest® method. Eight C. difficile isolates were recovered from seven farms. The isolates belonged to different PCR ribotypes. All isolates were toxinogenic. cgMLST revealed a cluster containing isolates recovered from different farms. Seven isolates showed similar resistance gene patterns. Different phenotypic resistance patterns were found. Agreement between phenotypic and genotypic resistance was identified only in some cases. Consequently, SAC may act as a reservoir for C. difficile. Thus, SAC may pose a risk regarding zoonotic transmission of toxinogenic, potentially human-pathogenic and resistant C. difficile isolates.

Polymorphisms in TRIB2 and CAPRIN2 Genes Contribute to the Susceptibility to High Myopia-Induced Cataract in Han Chinese Population.

BACKGROUND Myopia has been shown to be associated with many pathological complications including cataracts, and previous evidence supported that high myopia facilitates the formation of cataracts. However, no studies have identified a link between the genetic susceptibility of high myopia-induced cataracts (HMC) and the underlying genetic mechanisms. Our study aimed to determine how the TRIB2 and CAPRIN2 genes correlate to the risk of HMC. MATERIAL AND METHODS In total, we successfully recruited 3162 participants, including 1026 participants with high myopia and cataracts and 2136 controls with high myopia only. For genotyping, 22 tag single nucleotide polymorphisms (SNPs) in TRIB2 and CAPRIN2 genes were chosen. Single marker association analysis and functional effects of significant SNPs were carried out. RESULTS Strong correlation signals were captured for SNP rs890069 (χ²=22.13, P=2.55×10-6) in TRIB2 and SNP rs17739338 (χ²=16.07, P=6.10×10-5) in CAPRIN2. In patients with high myopia, the C allele at SNP rs890069 was strongly linked to cataract risk (OR [95% CI]=1.36 [1.20-1.55]). In patients with high myopia, the T allele at SNP rs17739338 was significantly related to a lower risk of cataract (OR [95% CI]=0.54 [0.40-0.74]). In different types of human tissues, SNPs rs890069 and rs17739338 were found to be significantly correlated to the levels of TRIB2 and CAPRIN2 gene expression. CONCLUSIONS Our study indicated that both TRIB2 and CAPRIN2 genes conferred the susceptibility to cataract in patients with high myopia and Chinese Han ancestry. Future research remains necessary for fully understanding the pathogenic mechanisms and genetic characteristics of cataract.

Whole-genome sequencing for genetic diversity analysis of Iranian Brucella spp. isolated from humans and livestock

Brucellosis is one of the most common zoonoses in the Middle East. It is causing economic losses to the livestock industry and has a great public health concern. Little is known about the genetic diversity and distribution of brucellae in Iran. Therefore, forty Brucella spp. strains (B. abortus and B. melitensis) isolated from animals and humans were analyzed by whole genome sequencing (WGS) technology using single nucleotide polymorphism (SNP) analysis and core genome multilocus sequence typing (cgMLST). Brucella isolates were obtained from lymph nodes (cows and camels), milk (cows, camels and sheep), and aborted foetus samples (sheep and goats), as well as cerebrospinal fluid and blood of humans. The isolates were originating from thirteen provinces of Iran and isolated between 2015 and 2020. According to in-silico MLST, ST8 and ST2 were the most frequent sequence types in B. melitensis and B. abortus, respectively. Based on phylogeographic reconstruction using cgSNP analysis, the investigated Iranian B. melitensis strains belonged to the American and Mediterranean lineages of the B. melitensis phylogeny. Furthermore, cgSNP analysis revealed a similarity between Iranian B. abortus isolates and strains from Iraq and Egypt. Therefore, the origin of the Iranian strains can be suggested to be strains from neighboring and Middle East countries. Moreover, cgMLST analysis showed that the Iranian B. melitensis strains were closely relative to strains recovered from sheep and humans in Iraq, Afghanistan, Syria, Turkmenistan, and Pakistan. In the current panel of strains, cgMLST and cgSNP analysis provided an appropriate and accurate tool for effective traceback analyses for Brucella spp. from Iran. The results of cgSNP and cgMLST helped to understand the geographic distribution and interspecies transmission of Iranian strains and highlight the importance of specific brucellosis control measures in Iran with regard to the One-Health approach.

Genetic diversity of Rhododendron henanense subsp. lingbaoense revealed by whole-genome resequencing

Rhododendron henanense subsp. lingbaoense Fang (Rhl) is a subspecies endemic to China with important ornamental value. Recently, the Rhl population has been shrinking as a result of its limited distribution range, poor natural regeneration, and rising human exploitation. In this study, high-quality single nucleotide polymorphisms (SNPs) identified through whole-genome resequencing were examined in 50 individuals from four populations of Rhl with a depth of ~34× to evaluate genome-level diversity and population structure. Population structure profiling, principal component analysis, and phylogenetic analysis clustered the Rhl samples into two groups corresponding to their geographical distributions. Analysis of SNPs indicated that Rhl populations have high genetic diversity and little genetic differentiation. Additionally, demographic history indicated that all four populations of Rhl have experienced long-term population decline. The above findings showed that we should take action to protect this rare species.

Robust nontarget DNA-triggered catalytic hairpin assembly amplification strategy for the improved sensing of microRNA in complex biological matrices.

A simple but robust fluorescence strategy based on a nontarget DNA-triggered catalytic hairpin assembly (CHA) was constructed to probe microRNA-21 (miR-21). A short ssDNA rather than degradable target miRNA was employed as an initiator. Two molecular beacons needed to assist the CHA process were simplified to avoid unfavorable nonspecific interactions. In the presence of the target, the initiator was released from a partially duplex and triggered the cyclic CHA reaction, resulting in a significantly amplified optical readout. A wide linear range from 0.1 pM to 1000 pM for the sensing of miR-21 in buffer was achieved with a low detection limit of 0.76 pM. Fortunately, this strategy demonstrated an obviously improved performance for miR-21 detection in diluted serum. The fluorescence signals were enhanced remarkably and the sensitivity was further improved to 0.12 pM in 10% serum. The stability for miR-21 quantification and the capability for the analysis of single nucleotide polymorphisms (SNPs) were also improved greatly. More importantly, the biosensor could be applied to image miR-21 in different living tumor cells with high resolution, illustrating its promising potential for the assay of miRNAs in various complex situations for early-stage disease diagnosis and biological studies in cells.

Atypical diarrheagenic Escherichia coli in milk related to a large foodborne outbreak.

A foodborne outbreak related to milk cartons served in school lunches occurred in June 2021, which involved more than 1,800 cases from 25 schools. The major symptoms were abdominal pain, diarrhoea, vomiting, and fever. Although major foodborne toxins and pathogens were not detected, a specific Escherichia coli strain, serotype OUT (OgGp9):H18, was predominantly isolated from milk samples related to the outbreak and most patients tested. The strains from milk and patient stool samples were identified as the same clone by core genome multilocus sequence typing and single-nucleotide polymorphism analysis. The strain was detected in milk samples served for two days related to the foodborne outbreak at a rate of 69.6% and levels of less than ten most probable number/100mL but not on days unrelated to the outbreak. The acid tolerance of the strain for survival in the stomach was similar to that of enterohaemorrhagic E. coli O157:H7, and the same inserts in the chu gene cluster in the acid fitness island were genetically revealed. The pathogenicity of the strain was not clear; however, it was indicated that the causative pathogen was atypical diarrhoeagenic E. coli OUT (OgGp9):H18.

Comparative whole-genome analysis of diphtheria toxin-producing Corynebacterium ulcerans isolates from sheltered cats in Osaka, Japan.

Diphtheria toxin-producing Corynebacterium ulcerans is a zoonotic pathogen that causes human diphtheria-like symptoms. After performing whole-genome analysis of the five isolates from sheltered cats in Osaka, Japan, we compared them with genome sequences of 25 strains of C. ulcerans from a public database. The five isolates from cats harbored 14 genes encoding possible virulence factors in diphtheria-toxin-producing C. ulcerans. These isolates also had diphtheria toxin gene-encoding prophage in their chromosome, although differences were found in other prophages possession. Whole-genome single-nucleotide polymorphism analysis showed that cats' isolates belonged to ST337 branch, as were strains from Japanese human patients, with 41 or more single-nucleotide polymorphisms variations. High-resolution single-nucleotide polymorphism analysis of C. ulcerans was sufficient to distinguish cats' isolates clearly as not different by conventional genotyping methods.

In Silico Analysis of Genome Wide Non-Synonymous Single Nucleotide Polymorphisms in Indigenous Cattle Breeds of Pakistan

In Silico Analysis of Genome Wide Non-Synonymous Single Nucleotide Polymorphisms in Indigenous Cattle Breeds of Pakistan

Population genetics of marmosets in Asian primate research centers and loci associated with epileptic risk revealed by whole-genome sequencing.

The common marmoset ( Callithrix jacchus) has emerged as a valuable nonhuman primate model in biomedical research with the recent release of high-quality reference genome assemblies. Epileptic marmosets have been independently reported in two Asian primate research centers. Nevertheless, the population genetics within these primate centers and the specific genetic variants associated with epilepsy in marmosets have not yet been elucidated. Here, we characterized the genetic relationships and risk variants for epilepsy in 41 samples from two epileptic marmoset pedigrees using whole-genome sequencing. We identified 14 558 184 single nucleotide polymorphisms (SNPs) from the 41 samples and found higher chimerism levels in blood samples than in fingernail samples. Genetic analysis showed fourth-degree of relatedness among marmosets at the primate centers. In addition, SNP and copy number variation (CNV) analyses suggested that the WW domain-containing oxidoreductase ( WWOX) and Tyrosine-protein phosphatase nonreceptor type 21 ( PTPN21) genes may be associated with epilepsy in marmosets. Notably, KCTD18-like gene deletion was more common in epileptic marmosets than control marmosets. This study provides valuable population genomic resources for marmosets in two Asian primate centers. Genetic analyses identified a reasonable breeding strategy for genetic diversity maintenance in the two centers, while the case-control study revealed potential risk genes/variants associated with epilepsy in marmosets.

Population Genomic Analysis of Diploid-Autopolyploid Species.

This chapter outlines an empirical analysis of genome-wide single-nucleotide polymorphism (SNP) variation and its underlying drivers among multiple natural populations within a diploid-autopolyploid species. The aim is to reconstruct the genetic structure among natural populations of varying ploidy and infer footprints of selection in these populations, framed around specific questions that are typically encountered when analyzing a mixed-ploidy data set,e.g., addressing the relevance of natural whole-genome duplication for speciation and adaptation. We briefly review the options for the analysis of polyploid population genomic data involving variant calling, population structure, demographic history inference, and selection scanning approaches. Further, we provide suggestions for methods and associated software, possible caveats, and examples of their application to mixed-ploidy and autopolyploid data sets.

Genotypes Diversity of Acute Gastroenteritis Outbreaks Caused by Human Sapovirus - Beijing Municipality, China, 2015-2021.

Human sapovirus (HuSaV) is an enteric virus responsible for sporadic cases and outbreaks of acute gastroenteritis (AGE) globally. A seven-year active surveillance study was conducted to investigate the molecular epidemiology of HuSaVs associated with AGE outbreaks in Chaoyang District of Beijing Municipality, China from January 2015 to December 2021. Fecal and anal swab samples were obtained from patients experiencing AGE outbreaks. HuSaVs were identified through reverse transcription polymerase chain reaction (RT-PCR), and partial viral protein 1 (VP1) sequences (approximately 434 base pairs) were utilized for genotyping, single nucleotide polymorphism (SNP) analysis, and phylogenetic examination. HuSaVs were identified in 71 AGE outbreaks, demonstrating a detection rate of 10.5%, second only to norovirus. The primary demographic affected by HuSaV were children under the age of 5 in kindergarten settings. Infection rates tended to peak during two distinct periods: May to June and September to December. Upon genotyping, seven distinct genotypes emerged. GII.3 was the most prevalent, accounting for 54.9% of cases, followed by GI.1 (12.7%), GI.2 (9.9%), GII.5 (7.0%), GI.5 (2.8%), GI.6 (1.4%), GII.1 (1.4%), and untyped cases (9.9%). A phylogenetic analysis of GII.3 identified three distinct groups, with 15 notable SNPs observed. This study offers a comprehensive analysis of the persistent prevalence of HuSaV outbreaks in Chaoyang District, Beijing Municipality, China. Over time, the diversity of HuSaV subtypes has shifted, and it is now recognized as the second leading viral agent responsible for AGE outbreaks. This highlights the importance of ongoing surveillance in the future.

Radiation-Induced Chromosome Instability in WTK1 and TK6 Human Lymphoblastoid Cells

The effects of ionizing radiation on single nucleotide polymorphism (SNP) copy number variations between TK6 and WTK1 cell lines are described herein. Specifically, the integrity of the chromosomes for two WIL2-derived lymphoblastoid cell lines (TK6 and WTK1) was analyzed in the presence and absence of ionizing radiation. WTK1 cells contain a p53 mutation, whereas the TK6 cell line has the native p53 tumor-suppressor gene. Each cell line was isolated post-irradiation for SNP analysis, which showed significant, genome-wide impacts on both cell lines; for the mutant WTK1 sample, there were a total of 48 gene deletions and no gene amplifications, whereas for the wild-type TK6 sample, there were 217 gene deletions and 9 gene amplifications. It appears that both cell lines are affected in the areas of cell-cycle control, but that other affected areas differ significantly between the two.

Impact of cognition-related single nucleotide polymorphisms on brain imaging phenotype in Parkinson's disease.

Multiple single nucleotide polymorphisms may contribute to cognitive decline in Parkinson's disease. However, the mechanism by which these single nucleotide polymorphisms modify brain imaging phenotype remains unclear. The aim of this study was to investigate the potential effects of multiple single nucleotide polymorphisms on brain imaging phenotype in Parkinson's disease. Forty-eight Parkinson's disease patients and 39 matched healthy controls underwent genotyping and 7T magnetic resonance imaging. A cognitive-weighted polygenic risk score model was designed, in which the effect sizes were determined individually for 36 single nucleotide polymorphisms. The correlations between polygenic risk score, neuroimaging features, and clinical data were analyzed. Furthermore, individual single nucleotide polymorphism analysis was performed to explore the main effects of genotypes and their interactive effects with Parkinson's disease diagnosis. We found that, in Parkinson's disease, the polygenic risk score was correlated with the neural activity of the hippocampus, parahippocampus, and fusiform gyrus, and with hippocampal-prefrontal and fusiform-temporal connectivity, as well as with gray matter alterations in the orbitofrontal cortex. In addition, we found that single nucleotide polymorphisms in α-synuclein (SNCA) were associated with white matter microstructural changes in the superior corona radiata, corpus callosum, and external capsule. A single nucleotide polymorphism in catechol-O-methyltransferase was associated with the neural activities of the lingual, fusiform, and occipital gyri, which are involved in visual cognitive dysfunction. Furthermore, DRD3 was associated with frontal and temporal lobe function and structure. In conclusion, imaging genetics is useful for providing a better understanding of the genetic pathways involved in the pathophysiologic processes underlying Parkinson's disease. This study provides evidence of an association between genetic factors, cognitive functions, and multi-modality neuroimaging biomarkers in Parkinson's disease.

Systematic headwater sampling of white‐spotted charr reveals stream capture events across dynamic topography

AbstractAimRiver system rearrangements such as stream capture play a crucial role in determining range expansion and gene flow of stream organisms between different watersheds. However, it is challenging to clarify the role of stream capture in dynamic landscapes where frequent connections and disconnections occur in stream networks. We hypothesized that fine‐scale sampling with genome‐wide analysis could reveal the effect of stream capture on the range expansion of stream fish and detect unidentified river rearrangement of headwater streams in a highly heterogeneous landscape.LocationHonshu Island, Japan.Taxon Salvelinus leucomaenis. MethodsTo identify stream capture events from biological evidence, a sampling strategy was designed with fish collected from multiple pairs of adjacent headwater streams. This included a ‘typical site’ where the occurrence of a stream capture event has been confirmed with geological evidence. The genetic population structure was analysed using genome‐wide single nucleotide polymorphism (SNP) data using MIG‐seq, followed by a Bayesian clustering approach.ResultsEvidence was found for historical dispersal across watersheds, including a typical stream capture site. This cross‐drainage dispersal promoted gene flow among the three genetic groups of focal fish inhabiting western Honshu Island. The likelihood of past stream capture increased substantially when the inter‐headwater distance was less than 2000 m, or the altitudinal difference was less than 200 m in the current topography.Main ConclusionsIn highly dynamic areas, such as the study site, which is surrounded by active faults, stream capture may be frequent, and aids inter‐drainage dispersal of freshwater organisms, where large‐scale range expansions would otherwise be impossible. Systematic pairwise headwater sampling, together with genome‐wide SNP analysis, provides biological evidence for the history of water system rearrangements and enables the identification of geological events.

Strategies for activity analysis of single nucleotide polymorphisms associated with human diseases.

Genome-wide association studies (GWAS) have identified a large number of single nucleotide polymorphism (SNP) sites associated with human diseases. In the annotation of human diseases, especially cancers, SNPs, as an important component of genetic factors, have gained increasing attention. Given that most of the SNPs are located in non-coding regions, the functional verification of these SNPs is a great challenge. The key to functional annotation for risk SNPs is to screen SNPs with regulatory activity from thousands of disease associated-SNPs. In this review, we systematically recapitulate the characteristics and functional roles of SNP sites, discuss three parallel reporter screening strategies in detail based on barcode tag classification, and recommend the common in silico strategies to help supplement the annotation of SNP sites with epigenetic activity analysis, prediction of target genes and trans-acting factors. We hope that this review will contribute to this exuberant research field by providing robust activity analysis strategies that can facilitate the translation of GWAS results into personalized diagnosis and prevention measures for human diseases.

Global Analysis of Plasmodium falciparum Dihydropteroate Synthase Variants Associated with Sulfadoxine Resistance Reveals Variant Distribution and Mechanisms of Resistance: A Computational-Based Study.

The continual rise in sulfadoxine (SDX) resistance affects the therapeutic efficacy of sulfadoxine-pyrimethamine; therefore, careful monitoring will help guide its prolonged usage. Mutations in Plasmodium falciparum dihydropteroate synthase (Pfdhps) are being surveilled, based on their link with SDX resistance. However, there is a lack of continuous analyses and data on the potential effect of molecular markers on the Pfdhps structure and function. This study explored single-nucleotide polymorphisms (SNPs) in Pfdhps that were isolated in Africa and other countries, highlighting the regional distribution and its link with structure. In total, 6336 genomic sequences from 13 countries were subjected to SNPs, haplotypes, and structure-based analyses. The SNP analysis revealed that the key SDX resistance marker, A437G, was nearing fixation in all countries, peaking in Malawi. The mutation A613S was rare except in isolates from the Democratic Republic of Congo and Malawi. Molecular docking revealed a general loss of interactions when comparing mutant proteins to the wild-type protein. During MD simulations, SDX was released from the active site in mutants A581G and A613S before the end of run-time, whereas an unstable binding of SDX to mutant A613S and haplotype A437A/A581G/A613S was observed. Conformational changes in mutant A581G and the haplotypes A581G/A613S, A437G/A581G, and A437G/A581G/A613S were seen. The radius of gyration revealed an unfolding behavior for the A613S, K540E/A581G, and A437G/A581G systems. Overall, tracking such mutations by the continuous analysis of Pfdhps SNPs is encouraged. SNPs on the Pfdhps structure may cause protein-drug function loss, which could affect the applicability of SDX in preventing malaria in pregnant women and children.

Diversity and Prevalence of Clostridium innocuum in the Human Gut Microbiota.

Clostridia are a polyphyletic group of Gram-positive, spore-forming anaerobes in the Firmicutes phylum that significantly impact metabolism and functioning of the human gastrointestinal tract. Recently, Clostridia were divided into two separate classes, Clostridia and Erysipelotrichia, based on phenotypic and 16S rRNA gene-based differences. While Clostridia include many well-known pathogenic bacteria, Erysipelotrichia remain relatively uncharacterized, particularly regarding their role as a pathogen versus commensal. Despite wide recognition as a commensal, the erysipelotrichial species Clostridium innocuum has recently been associated with various disease states. To further understand the ecological and potential virulent role of C. innocuum, we conducted a genomic comparison across 38 C. innocuum isolates and 194 publicly available genomes. Based on colony morphology, we isolated multiple C. innocuum cultivars from the feces of healthy human volunteers (n = 5). Comparison of the 16S rRNA gene of our isolates against publicly available microbiota data sets in healthy individuals suggests a high prevalence of C. innocuum across the human population (>80%). Analysis of single nucleotide polymorphisms (SNPs) across core genes and average nucleotide identify (ANI) revealed the presence of four clades among all available genomes (n = 232 total). Investigation of carbohydrate and protein utilization pathways, including comparison against the carbohydrate-activating enzyme (CAZyme) database, demonstrated inter- and intraclade differences that were further substantiated in vitro. Collectively, these data indicate genetic variance within the C. innocuum species that may help clarify its role in human disease and health. IMPORTANCE Clostridia are a group of medically important anaerobes as both commensals and pathogens. Recently, a new class of Erysipelotrichia containing a number of reassigned clostridial species has emerged, including Clostridium innocuum. Recent studies have implicated C. innocuum as a potential causative agent of diarrhea in patients from whom Clostridioides difficile could not be isolated. Using genomic and in vitro comparison, this study sought to characterize C. innocuum in the healthy human gut. Our analyses suggest that C. innocuum is a highly prevalent and diverse species, demonstrating clade-specific differences in metabolism and potential virulence. Collectively, this study is the first investigation into a broader description of C. innocuum as a human gut inhabitant.

Application of conventional IVF during preimplantation genetic testing for aneuploidies: a feasibility study

Research questionIs it feasible to apply conventional IVF to couples undergoing preimplantation genetic testing for aneuploidies (PGT-A) with non-severe male infertility? DesignThe last wash fluid of biopsied trophectoderm (TE) cells was collected for whole genome amplification (WGA). A method was developed to determine parental contamination. Using single-nucleotide polymorphism (SNP) analysis, two standard curves were established; further mixtures were used for verification. Finally, 29 WGA products from couples undergoing conventional IVF were used to evaluate parental contamination. ResultsThe WGA results of the last wash fluid of biopsied TE cells revealed almost no free DNA. By adopting two strategies based on maternally and paternally biased SNP in the mixture, data from bioinformatics analysis were analysed to determine the relationship between maternal (Index M) and paternal (Index F) bias statistics. Two standard curves were successfully established based on these indices that allowed the prediction of maternal and parental contamination, which correlated well with actual ratios of known composition mixtures during validation. The average contamination level was 10.6% determined from 10 WGA products that featured maternal contamination, whereas that of the other 19 products that featured paternal contamination was less than 10%. ConclusionsThis study confirmed the feasibility of applying conventional IVF to couples undergoing PGT-A with non-severe male infertility.