BACKGROUND Colorectal cancer(CRC)is a major global health burden.The current diagnostic tests have shortcomings of being invasive and low accuracy.AIM To explore the combination of intestinal microbiome composition an...BACKGROUND Colorectal cancer(CRC)is a major global health burden.The current diagnostic tests have shortcomings of being invasive and low accuracy.AIM To explore the combination of intestinal microbiome composition and multi-target stool DNA(MT-sDNA)test in the diagnosis of CRC.METHODS We assessed the performance of the MT-sDNA test based on a hospital clinical trial.The intestinal microbiota was tested using 16S rRNA gene sequencing.This case-control study enrolled 54 CRC patients and 51 healthy controls.We identified biomarkers of bacterial structure,analyzed the relationship between different tumor markers and the relative abundance of related flora components,and distinguished CRC patients from healthy subjects by the linear discriminant analysis effect size,redundancy analysis,and random forest analysis.RESULTS MT-sDNA was associated with Bacteroides.MT-sDNA and carcinoembryonic antigen(CEA)were positively correlated with the existence of Parabacteroides,and alpha-fetoprotein(AFP)was positively associated with Faecalibacterium and Megamonas.In the random forest model,the existence of Streptococcus,Escherichia,Chitinophaga,Parasutterella,Lachnospira,and Romboutsia can distinguish CRC from health controls.The diagnostic accuracy of MT-sDNA combined with the six genera and CEA in the diagnosis of CRC was 97.1%,with a sensitivity and specificity of 98.1%and 92.3%,respectively.CONCLUSION There is a positive correlation of MT-sDNA,CEA,and AFP with intestinal microbiome.Eight biomarkers including six genera of gut microbiota,MT-sDNA,and CEA showed a prominent sensitivity and specificity for CRC prediction,which could be used as a non-invasive method for improving the diagnostic accuracy for this malignancy.展开更多
Introduction: Anxiety disorders have a lifetime prevalence of 34% with a similar level of heritability (31%) yet lack objective markers that could differentiate patients with underlying conditions. Up to 60%-70% of pa...Introduction: Anxiety disorders have a lifetime prevalence of 34% with a similar level of heritability (31%) yet lack objective markers that could differentiate patients with underlying conditions. Up to 60%-70% of patients with Ehlers-Danlos syndrome have anxiety that meets criteria of generalized anxiety disorder, their clinical-DNA findings worth examining as biomarkers for patients with generalized anxiety. Method: Of the 1899 patients diagnosed with Ehlers-Danlos syndrome, 1261 were systematically evaluated for 80 history and 40 physical findings and separated into 826 who reported anxiety and 435 who did not. The most consistently reported or management-directing 60 of these clinical findings were, along with variations in genes relevant to these disorders, examined for association with anxiety. Results: Among the 30 anxiety- associated findings judged most predictive of Ehlers-Danlos syndrome in patients with anxiety were expected ones of adrenergic stimulation (difficulty concentrating-87% frequency and 1.26 anxiety/no anxiety ratio;chronic fatigue-84%, 1.17;sleep issues 69%, 1.52 that are criteria for generalized anxiety disorder) or of cholinergic suppression (e.g., frequent nausea 64%, 1.26). Less associated but more discriminating for underlying disease were those reflective of neuromuscular impact (e.g., chronic daily headaches 76%, 1.12);hypermobility (e.g., awareness of flexibility 72%, 1.03), or skin changes (e.g., elasticity around jaw 71%, 1.06). Anxiety-associated DNA variants included 54 of 88 in collagen type I/V/VII/IX genes, 14 of 16 in sodium channel SCN9A/10A/ 11A genes, 59 of 85 in POLG/MT-DNA genes, and 21 of 28 in profilaggrin- FLG genes that respectively impacted tissue laxity, sensory neural, autonomic-mitochondrial, and autonomic-inflammatory functions. Conclusion: Analysis of pathogenetic mechanisms in Ehlers-Danlos syndrome selected some 50 clinical-DNA findings useful for its diagnosis in those with generalized anxiety disorders.展开更多
AIM:To compare the clinical performance of a real-time PCR assay with the COBAS Amplicor Hepatitis B Virus (HBV) Monitor test for quantitation of HBV DNA in serum samples. METHODS: The reference sera of the Chinese Na...AIM:To compare the clinical performance of a real-time PCR assay with the COBAS Amplicor Hepatitis B Virus (HBV) Monitor test for quantitation of HBV DNA in serum samples. METHODS: The reference sera of the Chinese National Institute for the Control of Pharmaceutical and Biological Products and the National Center for Clinical Laboratories of China, and 158 clinical serum samples were used in this study. The linearity, accuracy, reproducibility, assay time, and costs of the real-time PCR were evaluated and compared with those of the Cobas Amplicor test. RESULTS: The intra-assay and inter-assay variations of the real-time PCR ranged from 0.3% to 3.8% and 1.4% to 8.1%, respectively. The HBV DNA levels measured by the real-time PCR correlated very well with those obtained with the COBAS Amplicor test (r = 0.948). The real-time PCR HBV DNA kit was much cheaper and had a wider dynamic range. CONCLUSION: The real-time PCR assay is an excellent tool for monitoring of HBV DNA levels in patients with chronic hepatitis B.展开更多
AIM:To explore patient interest in a potential multiorgan stool-DNA test(MUST)for pan-digestive cancer screening.METHODS:A questionnaire was designed and mailed to 1200 randomly-selected patients from the Mayo Clinic ...AIM:To explore patient interest in a potential multiorgan stool-DNA test(MUST)for pan-digestive cancer screening.METHODS:A questionnaire was designed and mailed to 1200 randomly-selected patients from the Mayo Clinic registry.The 29-item survey questionnaire included items related to demographics,knowledge of digestive cancers,personal and family history of cancer,personal concern of cancer,colorectal cancer(CRC)screening behavior,interest in MUST,importance of test features in a cancer screening tool,and comparison of MUST with available CRC screening tests.All responses were summarized descriptively.χ2 and Rank Sum Test were used for categorical and continuous variables,respectively.RESULTS:Completed surveys were returned by 434(29%aged 50-59,37%60-69,34%70-79,52%women).Most participants(98%)responded they would use MUST.In order of importance,respondents rated multicancer detection,absence of bowel preparation,safety and noninvasiveness as most attractive characteristics.For CRC screening,MUST was preferred over colorectal-only stool-DNA testing(53%),occult blood testing(75%),colonoscopy(84%),sigmoidoscopy(91%),and barium enema(95%),P<0.0001 for each.Among those not previously screened,most(96%)indicated they would use MUST if available.Respondents were confident in their ability to follow instructions to perform MUST(98%).Only 9%of respondents indicated that fear of finding cancer was a concern with MUST,and only 3%indicated unpleasantness of stool sampling as a potential barrier.CONCLUSION:Patients are receptive to the concept of MUST,preferred MUST over conventional CRC screening modalities and valued its potential feature of multi-cancer detection.展开更多
Objective: To improve the detecting accuracy of chromosomal aneuploidy of fetus by non-invasive prenatal testing (NIPT) using next generation sequencing data of pregnant women’s cell-free DNA. Methods: We proposed th...Objective: To improve the detecting accuracy of chromosomal aneuploidy of fetus by non-invasive prenatal testing (NIPT) using next generation sequencing data of pregnant women’s cell-free DNA. Methods: We proposed the multi-Z method which uses 21 z-scores for each autosomal chromosome to detect aneuploidy of the chromosome, while the conventional NIPT method uses only one z-score. To do this, mapped read numbers of a certain chromosome were normalized by those of the other 21 chromosomes. Average and standard deviation (SD), which are used for calculating z-score of each sample, were obtained with normalized values between all autosomal chromosomes of control samples. In this way, multiple z-scores can be calculated for 21 autosomal chromosomes except oneself. Results: Multi-Z method showed 100% sensitivity and specificity for 187 samples sequenced to 3 M reads while the conventional NIPT method showed 95.1% specificity. Similarly, for 216 samples sequenced to 1 M reads, Multi-Z method showed 100% sensitivity and 95.6% specificity and the conventional NIPT method showed a result of 75.1% specificity. Conclusion: Multi-Z method showed higher accuracy and robust results than the conventional method even at low coverage reads.展开更多
The concentration of cell-free fetal DNA fragments should be detected before noninvasive prenatal testing(NIPT).The fetal DNA molecules have significant clinical potential in determining the overall performance of NIP...The concentration of cell-free fetal DNA fragments should be detected before noninvasive prenatal testing(NIPT).The fetal DNA molecules have significant clinical potential in determining the overall performance of NIPT and clinical interpretation.It is important to measure fetal DNA fraction before NIPT.However,there is still little research on how to calculate the concentration of female fetuses.Two estimation approaches were proposed to calculate fetal DNA fraction,including the fragments size-based approach,aneuploid-based approach,which are all approaches based on chromosome segments.Based on high-throughput sequencing data,two approaches to calculate the DNA fraction of male fetuses were tested and obtained the experiment values,which were close to the actual values.The correlation coefficient of fragments size-based approach was 0.9243(P<0.0001)and the aneuploid-based approach reached 0.9339(P<0.0001).We calculated the concentration of female fetuses and obtained remarkable experimental results.We came up with two approaches for calculating the fetal DNA fraction of female fetuses.It provides an important theoretical basis for the detection of female fetal concentration in future clinical diagnosis.展开更多
Background:The analysis of chromatin integrity has become an important determinant of sperm quality.In frozenthawed bovine sperm,neither the sequence of post-thaw injury events nor the dynamics of different types of s...Background:The analysis of chromatin integrity has become an important determinant of sperm quality.In frozenthawed bovine sperm,neither the sequence of post-thaw injury events nor the dynamics of different types of sperm DNA breaks are well understood.The aim of the present work was to describe such sperm degradation aftermath focusing on DNA damage dynamics,and to assess if this parameter can predict pregnancy rates in cattle.Results:A total of 75 cryopreserved ejaculates from 25 Holstein bulls were evaluated at two post-thawing periods(0-2 h and 2-4 h),analyzing global and double-stranded DNA damage through alkaline and neutral Comet assays,chromatin deprotamination and decondensation,sperm motility,viability,acrosomal status,and intracellular levels of total ROS,superoxides and calcium.Insemination of 59,605 females was conducted using sperm from the same bulls,thus obtaining the non-return to estrus rates after 90 d(NRR).Results showed an increased rate of double-stranded breaks in the first period(0-2 h:1.29±1.01%/h vs.2-4 h:0.13±1.37%/h;P<0.01),whereas the rate of sperm with moderate+high single-stranded breaks was higher in the second period(0-2 h:3.52±7.77%/h vs.2-4h:21.06±11.69%/h;P<0.0001).Regarding sperm physiology,viability decrease rate was different between the two periods(0-2 h:-4.49±1.79%/h vs.2-4 h:-2.50±3.39%/h;P=0.032),but the progressive motility decrease rate was constant throughout post-thawing incubation(0-2 h:-4.70±3.42%/h vs.2-4 h:-1.89±2.97%/h;P>0.05).Finally,whereas no correlations between bull fertility and any dynamic parameter were found,there were correlations between the NRR and the basal percentage of highly-damaged sperm assessed with the alkaline Comet(Rs=-0.563,P=0.003),between NRR and basal progressive motility(Rs=0.511,P=0.009),and between NRR and sperm with high ROS at 4 h post-thaw(Rs=0.564,P=0.003).Conclusion:The statistically significant correlations found between intracellular ROS,sperm viability,sperm motility,DNA damage and chromatin deprotamination suggested a sequence of events all driven by oxidative stress,where viability and motility would be affected first and sperm chromatin would be altered at a later stage,thus suggesting that bovine sperm should be used for fertilization within 2 h post-thaw.Fertility correlations supported that the assessment of global DNA damage through the Comet assay may help predict bull fertility.展开更多
基金Supported by the Medical and Health Research Project of Zhejiang Province,No.2021KY1048 and 2022KY1142Ningbo Health Young Technical Backbone Talents Training Program,No.2020SWSQNGG-02the Key Science and Technology Project of Ningbo City,No.2021Z133.
文摘BACKGROUND Colorectal cancer(CRC)is a major global health burden.The current diagnostic tests have shortcomings of being invasive and low accuracy.AIM To explore the combination of intestinal microbiome composition and multi-target stool DNA(MT-sDNA)test in the diagnosis of CRC.METHODS We assessed the performance of the MT-sDNA test based on a hospital clinical trial.The intestinal microbiota was tested using 16S rRNA gene sequencing.This case-control study enrolled 54 CRC patients and 51 healthy controls.We identified biomarkers of bacterial structure,analyzed the relationship between different tumor markers and the relative abundance of related flora components,and distinguished CRC patients from healthy subjects by the linear discriminant analysis effect size,redundancy analysis,and random forest analysis.RESULTS MT-sDNA was associated with Bacteroides.MT-sDNA and carcinoembryonic antigen(CEA)were positively correlated with the existence of Parabacteroides,and alpha-fetoprotein(AFP)was positively associated with Faecalibacterium and Megamonas.In the random forest model,the existence of Streptococcus,Escherichia,Chitinophaga,Parasutterella,Lachnospira,and Romboutsia can distinguish CRC from health controls.The diagnostic accuracy of MT-sDNA combined with the six genera and CEA in the diagnosis of CRC was 97.1%,with a sensitivity and specificity of 98.1%and 92.3%,respectively.CONCLUSION There is a positive correlation of MT-sDNA,CEA,and AFP with intestinal microbiome.Eight biomarkers including six genera of gut microbiota,MT-sDNA,and CEA showed a prominent sensitivity and specificity for CRC prediction,which could be used as a non-invasive method for improving the diagnostic accuracy for this malignancy.
文摘Introduction: Anxiety disorders have a lifetime prevalence of 34% with a similar level of heritability (31%) yet lack objective markers that could differentiate patients with underlying conditions. Up to 60%-70% of patients with Ehlers-Danlos syndrome have anxiety that meets criteria of generalized anxiety disorder, their clinical-DNA findings worth examining as biomarkers for patients with generalized anxiety. Method: Of the 1899 patients diagnosed with Ehlers-Danlos syndrome, 1261 were systematically evaluated for 80 history and 40 physical findings and separated into 826 who reported anxiety and 435 who did not. The most consistently reported or management-directing 60 of these clinical findings were, along with variations in genes relevant to these disorders, examined for association with anxiety. Results: Among the 30 anxiety- associated findings judged most predictive of Ehlers-Danlos syndrome in patients with anxiety were expected ones of adrenergic stimulation (difficulty concentrating-87% frequency and 1.26 anxiety/no anxiety ratio;chronic fatigue-84%, 1.17;sleep issues 69%, 1.52 that are criteria for generalized anxiety disorder) or of cholinergic suppression (e.g., frequent nausea 64%, 1.26). Less associated but more discriminating for underlying disease were those reflective of neuromuscular impact (e.g., chronic daily headaches 76%, 1.12);hypermobility (e.g., awareness of flexibility 72%, 1.03), or skin changes (e.g., elasticity around jaw 71%, 1.06). Anxiety-associated DNA variants included 54 of 88 in collagen type I/V/VII/IX genes, 14 of 16 in sodium channel SCN9A/10A/ 11A genes, 59 of 85 in POLG/MT-DNA genes, and 21 of 28 in profilaggrin- FLG genes that respectively impacted tissue laxity, sensory neural, autonomic-mitochondrial, and autonomic-inflammatory functions. Conclusion: Analysis of pathogenetic mechanisms in Ehlers-Danlos syndrome selected some 50 clinical-DNA findings useful for its diagnosis in those with generalized anxiety disorders.
文摘AIM:To compare the clinical performance of a real-time PCR assay with the COBAS Amplicor Hepatitis B Virus (HBV) Monitor test for quantitation of HBV DNA in serum samples. METHODS: The reference sera of the Chinese National Institute for the Control of Pharmaceutical and Biological Products and the National Center for Clinical Laboratories of China, and 158 clinical serum samples were used in this study. The linearity, accuracy, reproducibility, assay time, and costs of the real-time PCR were evaluated and compared with those of the Cobas Amplicor test. RESULTS: The intra-assay and inter-assay variations of the real-time PCR ranged from 0.3% to 3.8% and 1.4% to 8.1%, respectively. The HBV DNA levels measured by the real-time PCR correlated very well with those obtained with the COBAS Amplicor test (r = 0.948). The real-time PCR HBV DNA kit was much cheaper and had a wider dynamic range. CONCLUSION: The real-time PCR assay is an excellent tool for monitoring of HBV DNA levels in patients with chronic hepatitis B.
基金Supported by Research grant from the Oswald Foundation
文摘AIM:To explore patient interest in a potential multiorgan stool-DNA test(MUST)for pan-digestive cancer screening.METHODS:A questionnaire was designed and mailed to 1200 randomly-selected patients from the Mayo Clinic registry.The 29-item survey questionnaire included items related to demographics,knowledge of digestive cancers,personal and family history of cancer,personal concern of cancer,colorectal cancer(CRC)screening behavior,interest in MUST,importance of test features in a cancer screening tool,and comparison of MUST with available CRC screening tests.All responses were summarized descriptively.χ2 and Rank Sum Test were used for categorical and continuous variables,respectively.RESULTS:Completed surveys were returned by 434(29%aged 50-59,37%60-69,34%70-79,52%women).Most participants(98%)responded they would use MUST.In order of importance,respondents rated multicancer detection,absence of bowel preparation,safety and noninvasiveness as most attractive characteristics.For CRC screening,MUST was preferred over colorectal-only stool-DNA testing(53%),occult blood testing(75%),colonoscopy(84%),sigmoidoscopy(91%),and barium enema(95%),P<0.0001 for each.Among those not previously screened,most(96%)indicated they would use MUST if available.Respondents were confident in their ability to follow instructions to perform MUST(98%).Only 9%of respondents indicated that fear of finding cancer was a concern with MUST,and only 3%indicated unpleasantness of stool sampling as a potential barrier.CONCLUSION:Patients are receptive to the concept of MUST,preferred MUST over conventional CRC screening modalities and valued its potential feature of multi-cancer detection.
文摘Objective: To improve the detecting accuracy of chromosomal aneuploidy of fetus by non-invasive prenatal testing (NIPT) using next generation sequencing data of pregnant women’s cell-free DNA. Methods: We proposed the multi-Z method which uses 21 z-scores for each autosomal chromosome to detect aneuploidy of the chromosome, while the conventional NIPT method uses only one z-score. To do this, mapped read numbers of a certain chromosome were normalized by those of the other 21 chromosomes. Average and standard deviation (SD), which are used for calculating z-score of each sample, were obtained with normalized values between all autosomal chromosomes of control samples. In this way, multiple z-scores can be calculated for 21 autosomal chromosomes except oneself. Results: Multi-Z method showed 100% sensitivity and specificity for 187 samples sequenced to 3 M reads while the conventional NIPT method showed 95.1% specificity. Similarly, for 216 samples sequenced to 1 M reads, Multi-Z method showed 100% sensitivity and 95.6% specificity and the conventional NIPT method showed a result of 75.1% specificity. Conclusion: Multi-Z method showed higher accuracy and robust results than the conventional method even at low coverage reads.
基金supported by the National Key Research and Development Program of China 2016YFC1000307The sub-project of National Key Research and Development Program of China 2016YFC1000307-10the Program of National Research Institute for Family Planning(2017GJM04,2018CNV).
文摘The concentration of cell-free fetal DNA fragments should be detected before noninvasive prenatal testing(NIPT).The fetal DNA molecules have significant clinical potential in determining the overall performance of NIPT and clinical interpretation.It is important to measure fetal DNA fraction before NIPT.However,there is still little research on how to calculate the concentration of female fetuses.Two estimation approaches were proposed to calculate fetal DNA fraction,including the fragments size-based approach,aneuploid-based approach,which are all approaches based on chromosome segments.Based on high-throughput sequencing data,two approaches to calculate the DNA fraction of male fetuses were tested and obtained the experiment values,which were close to the actual values.The correlation coefficient of fragments size-based approach was 0.9243(P<0.0001)and the aneuploid-based approach reached 0.9339(P<0.0001).We calculated the concentration of female fetuses and obtained remarkable experimental results.We came up with two approaches for calculating the fetal DNA fraction of female fetuses.It provides an important theoretical basis for the detection of female fetal concentration in future clinical diagnosis.
基金the European Union’s Horizon 2020 Research and Innovation scheme under the Marie Sklodowska-Curie grant agreement No.801342(Tecniospring INDUSTRY,TECSPR-19-1-0003)the Ministry of Science and Innovation,Spain(AGL2017-88329-R and PID2020-113320RBI00)+2 种基金the Catalan Agency for Management of University and Research Grants,Regional Government of Catalonia,Spain(2017-SGR-1229)the Catalan Institution for Research and Advanced Studies(ICREA)La Maratóde TV3 Foundation(214/857-202039)。
文摘Background:The analysis of chromatin integrity has become an important determinant of sperm quality.In frozenthawed bovine sperm,neither the sequence of post-thaw injury events nor the dynamics of different types of sperm DNA breaks are well understood.The aim of the present work was to describe such sperm degradation aftermath focusing on DNA damage dynamics,and to assess if this parameter can predict pregnancy rates in cattle.Results:A total of 75 cryopreserved ejaculates from 25 Holstein bulls were evaluated at two post-thawing periods(0-2 h and 2-4 h),analyzing global and double-stranded DNA damage through alkaline and neutral Comet assays,chromatin deprotamination and decondensation,sperm motility,viability,acrosomal status,and intracellular levels of total ROS,superoxides and calcium.Insemination of 59,605 females was conducted using sperm from the same bulls,thus obtaining the non-return to estrus rates after 90 d(NRR).Results showed an increased rate of double-stranded breaks in the first period(0-2 h:1.29±1.01%/h vs.2-4 h:0.13±1.37%/h;P<0.01),whereas the rate of sperm with moderate+high single-stranded breaks was higher in the second period(0-2 h:3.52±7.77%/h vs.2-4h:21.06±11.69%/h;P<0.0001).Regarding sperm physiology,viability decrease rate was different between the two periods(0-2 h:-4.49±1.79%/h vs.2-4 h:-2.50±3.39%/h;P=0.032),but the progressive motility decrease rate was constant throughout post-thawing incubation(0-2 h:-4.70±3.42%/h vs.2-4 h:-1.89±2.97%/h;P>0.05).Finally,whereas no correlations between bull fertility and any dynamic parameter were found,there were correlations between the NRR and the basal percentage of highly-damaged sperm assessed with the alkaline Comet(Rs=-0.563,P=0.003),between NRR and basal progressive motility(Rs=0.511,P=0.009),and between NRR and sperm with high ROS at 4 h post-thaw(Rs=0.564,P=0.003).Conclusion:The statistically significant correlations found between intracellular ROS,sperm viability,sperm motility,DNA damage and chromatin deprotamination suggested a sequence of events all driven by oxidative stress,where viability and motility would be affected first and sperm chromatin would be altered at a later stage,thus suggesting that bovine sperm should be used for fertilization within 2 h post-thaw.Fertility correlations supported that the assessment of global DNA damage through the Comet assay may help predict bull fertility.