Two approaches for calculating female fetal DNA fraction in noninvasive prenatal testing based on size analysis of maternal DNA fragments

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.

Colorectal cancer screening:The value of early detection and modern challenges

The screening of colorectal cancer(CRC)is pivotal for both the prevention and treatment of this disease,significantly improving early-stage tumor detection rates.This advancement not only boosts survival rates and quality of life for patients but also reduces the costs associated with treatment.However,the adoption of CRC screening methods faces numerous challenges,including the technical limitations of both noninvasive and invasive methods in terms of sensitivity and specificity.Moreover,socioeconomic factors such as regional disparities,economic conditions,and varying levels of awareness affect screening uptake.The coronavirus disease 2019 pandemic further intensified these challenges,leading to reduced screening participation and increased waiting periods.Additionally,the growing prevalence of early-onset CRC necessitates innovative screening approaches.In response,research into new methodologies,including artificial intelligence-based systems,aims to improve the precision and accessibility of screening.Proactive measures by governments and health organizations to enhance CRC screening efforts are underway,including increased advocacy,improved service delivery,and international cooperation.The role of technological innovation and global health collaboration in advancing CRC screening is undeniable.Technologies such as artificial intelligence and gene sequencing are set to revolutionize CRC screening,making a significant impact on the fight against this disease.Given the rise in early-onset CRC,it is crucial for screening strategies to continually evolve,ensuring their effectiveness and applicability.

Multiparametric ultrasound as a new concept of assessment of liver tissue damage

Liver disease accounts for approximately 2 million deaths per year worldwide.All chronic liver diseases(CLDs),whether of toxic,genetic,autoimmune,or infectious origin,undergo typical histological changes in the structure of the tissue.These changes may include the accumulation of extracellular matrix material,fats,triglycerides,or tissue scarring.Noninvasive methods for diagnosing CLD,such as conventional B-mode ultrasound(US),play a significant role in diagnosis.Doppler US,when coupled with B-mode US,can be helpful in evaluating the hemodynamics of hepatic vessels and detecting US findings associated with hepatic decompensation.US elastography can assess liver stiffness,serving as a surrogate marker for liver fibrosis.It is important to note that interpreting these values should not rely solely on a histological classification.Contrast-enhanced US(CEUS)provides valuable information on tissue perfusion and enables excellent differentiation between benign and malignant focal liver lesions.Clinical evaluation,the etiology of liver disease,and the patient current comorbidities all influence the interpretation of liver stiffness measurements.These measurements are most clinically relevant when interpreted as a probability of compensated advanced CLD.B-mode US offers a subjective estimation of fatty infiltration and has limited sensitivity for mild steatosis.The controlled attenuation parameter requires a dedicated device,and cutoff values are not clearly defined.Quan-titative US parameters for liver fat estimation include the attenuation coefficient,backscatter coefficient,and speed of sound.These parameters offer the advantage of providing fat quantification alongside B-mode evaluation and other US parameters.Multiparametric US(MPUS)of the liver introduces a new concept for complete noninvasive diagnosis.It encourages examiners to utilize the latest features of an US machine,including conventional B-mode,liver stiffness evaluation,fat quantification,dispersion imaging,Doppler US,and CEUS for focal liver lesion characterization.This comprehensive approach allows for diagnosis in a single examination,providing clinicians worldwide with a broader perspective and becoming a cornerstone in their diagnostic arsenal.MPUS,in the hands of skilled clinicians,becomes an invaluable predictive tool for diagnosing,staging,and monitoring CLD.

Noninvasive diagnosis of hepatic fibrosis in chronic hepatitis C

Assessment of hepatic fibrosis is important for determining prognosis,guiding management decisions,and monitoring disease. Histological evaluation of liver biopsy specimens is currently considered the reference test for staging hepatic fibrosis. Since liver biopsy carries a small but significant risk,noninvasive tests to assess hepatic fibrosis are desirable. This editorial gives an overview on noninvasive methods currently available to determine hepatic fibrosis and their diagnostic accuracy for predicting significant fibrosis and cirrhosis in chronic hepatitis C. Based on available data,the performance of simple tests derived from routine laboratory parameters appears to be similar to that of more complex and expensive fibrosis panels. Transient elastography seems more accurate than blood tests for diagnosing cirrhosis.

Noninvasive molecular analysis of Helicobacter pylori : Is it time for tailored first-line therapy?

The main problem of Helicobacter pylori(H. pylori) infection management is linked to antibiotic resistances. This phenomenon has grown in the last decade, inducing a dramatic decline in conventional regimen effectiveness. The causes of resistance are point mutations in bacterial DNA, which interfere with antibiotic mechanism of action, especially clarithromycin and levofloxacin. Therefore, international guidelines have recently discouraged their use in areas with a relevant resistance percentage, suggesting first-line schedules with expected high eradication rates, i.e., bismuth containing or non-bismuth quadruple therapies. These regimens require the daily assumption of a large number of tablets. Consequently, a complete adherence is expected only in subjects who may be motivated by the presence of major disorders. However, an incomplete adherence to antibiotic therapies may lead to resistance onset, since sub-inhibitory concentrations could stimulate the selection of resistant mutants. Of note, a recent meta-analysis suggests that susceptibility tests may be more useful for the choice of first than second-line or rescue treatment. Additionally, susceptibility guided therapy has been demonstrated to be highly effective and superior to empiric treatments by both meta-analyses and recent clinical studies. Conventional susceptibility test is represented by culture and antibiogram. However, the method is not available everywhere mainly for methodology-related factors and fails to detect hetero-resistances. Polymerase chain reaction(PCR)-based, culture-free techniques on gastric biopsy samples are accurate in finding even minimal traces of genotypic resistant strains and hetero-resistant status by the identification of specific point mutations. The need for an invasive endoscopic procedure has been the most important limit to their spread. A further step has, moreover, been the detection of point mutations in bacterial DNA fecal samples. Few studies on clarithromycin susceptibility have shown an overall high sensitivity and specificity when compared with culture or PCR on gastric biopsies. On these bases, two commercial tests are now available although they have shown some controversial findings. A novel PCR method showed a full concordance between tissue and stool results in a preliminary experience. In conclusion, despite poor validation, there is increasing evidence of a potential availability of noninvasive investigations able to detect H. pylori resistances to antibiotics. These kinds of analysis are currently at a very early phase of development and caution should be paid about their clinical application. Only further studies aimed to evaluate their sensitivity and specificity will afford novel data for solid considerations. Nevertheless, noninvasive molecular tests may improve patient compliance, time/cost of infection management and therapeutic outcome. Moreover, the potential risk of a future increase of resistance to quadruple regimens as a consequence of their use on large scale and incomplete patient adherence could be avoided.

Serum 1,3-beta-D-glucan as a noninvasive test to predict histologic activity in patients with inflammatory bowel disease

BACKGROUND 1,3-beta-D-glucan(BG)is a ubiquitous cell wall component of gut microorganisms.We hypothesized that the serum levels of BG could reflect active intestinal inflammation in patients with inflammatory bowel disease.AIM To determine whether the serum BG concentrations correlate with intestinal inflammation.METHODS A prospective observational study was performed in a tertiary referral center,from 2016 to 2019,in which serum BG was determined in 115 patients with Crohn’s disease(CD),51 with ulcerative colitis(UC),and 82 controls using a photometric detection kit.Inflammatory activity was determined by ileocolonoscopy,histopathology,magnetic resonance enterography,and biomarkers,including fecal calprotectin(FC),C-reactive protein,and a panel of cytokines.The ability of BG to detect active vs inactive disease was assessed using the area under the receiver operating characteristic curve.In subgroup analysis,serial BG was used to assess the response to therapeutic interventions.RESULTS The serum BG levels were higher in CD patients than in controls(P=0.0001).The BG levels paralleled the endoscopic activity in CD patients and histologic activity and combined endoscopic and histologic activity in both CD and UC patients.The area under the curve(AUC)in receiver operating characteristic analysis to predict endoscopic activity was 0.694[95%confidence interval(CI):0.60-0.79;P=0.001]in CD,and 0.662(95%CI:0.51-0.81;P=0.066)in UC patients.The AUC in receiver operating characteristic analysis to predict histologic activity was 0.860(95%CI:0.77-0.95;P<0.001)in CD,and 0.786(95%CI:0.57-0.99;P=0.015)in UC patients.The cut-off values of BG for both endoscopic and histologic activity were 60μg/mL in CD,and 40μg/mL in UC patients.Performance analysis showed that the results based on BG of 40 and 60μg/mL were more specific for predicting endoscopic activity(71.8%and 87.2%for CD;and 87.5%and 87.5%for UC,respectively)than FC(53.3%and 66.7%for CD;and 20%and 80%for UC,respectively);and also histologic activity(60.5%and 76.3%for CD;and 90.0%and 95.0%for UC,respectively)than FC(41.7%and 50.0%for CD;and 25%and 50%for UC,respectively).Regarding the clinical,endoscopic,and histologic activities,the BG levels were reduced following therapeutic intervention in patients with CD(P<0.0001)and UC(P=0.003).Compared with endoscopic(AUC:0.693;P=0.002)and histologic(AUC:0.868;P<0.001)activity,no significant correlation was found between serum BG and transmural healing based on magnetic resonance enterography(AUC:0.576;P=0.192).Positive correlations were detected between BG and IL-17 in the CD(r:0.737;P=0.001)and the UC group(r:0.574;P=0.005),and between BG and interferon-gamma in the CD group(r:0.597;P=0.015).CONCLUSION Serum BG may represent an important novel noninvasive approach for detecting mucosal inflammation and therapeutically monitoring inflammatory bowel diseases,particularly in CD.

Routine blood tests to predict liver fibrosis in chronic hepatitis C

AIM: To verify the usefulness of FibroQ for predicting fi brosis in patients with chronic hepatitis C, compared with other noninvasive tests. METHODS: This retrospective cohort study included 237 consecutive patients with chronic hepatitis C who had undergone percutaneous liver biopsy before treatment. FibroQ, aspartate aminotransferase (AST)/alanine aminotransferase ratio (AAR), AST to platelet ratioindex, cirrhosis discriminant score, age-platelet index (API), Pohl score, FIB-4 index, and Lok's model were calculated and compared. RESULTS: FibroQ, FIB-4, AAR, API and Lok's model results increased significantly as fibrosis advanced (analysis of variance test: P < 0.001). FibroQ trended to be superior in predicting signifi cant fi brosis score in chronic hepatitis C compared with other noninvasive tests. CONCLUSION: FibroQ is a simple and useful test for predicting signifi cant fi brosis in patients with chronic hepatitis C.

Noninvasive Prenatal Testing for Fetal XXY Aneuploidies Among Pregnancies in Beijing of China

Objective:To evaluate the screening performance of noninvasive prenatal testing(NIPT)based on high-throughput massively parallel sequencing technology for the fetal XXY aneuploidies among pregnancies in Beijing of China.Methods:The study enrolled 26913 consecutive pregnancies,20-50 years old,who attended the Peking Union Medical College Hospital,Beijing,China,for prenatal screening from January 1,2016 to December 31,2019.Cell-free DNA was extracted from maternal peripheral blood to have a high-throughput massively parallel sequencing procedure.Cases with high-risk of fetal XXY were suggested to take invasive prenatal diagnosis(IPD)for confirmation.Maternal DNA sequencing was performed,if necessary,to find other potential factors that may lead to high-risk results of XXY by NIPT.Results:Among a cohort of 26913 pregnant women,34 were high-risk for fetal XXY,among which 30 accepted IPD while 4 declined.In those who accepted IPD,19 cases were confirmed fetal XXY by chromosome karyotyping analysis while 11 were verified as false positive.Among the 19 confirmed fetal XXY cases,14 elected pregnancy termination.For all the 34 high-risk cases,two were verified maternal sex chromosome aneuploidy.The calculated detection rate,positive predictive value,and false-positive rate of NIPT for fetal XXY in this cohort was 100.00%(19/19),63.33%(19/30),and 0.04%(11/26890),respectively.And the percentage of pregnancy termination was 73.68%(14/19).Conclusion:NIPT could be used as a potential method for fetal XXY screening,although the accuracy needs to be improved.As NIPT is not diagnostic,IPD is strongly recommended for those with high-risk results.For cases with discordance between NIPT and fetal karyotyping,maternal DNA sequencing would help to identify the cause of false-positive/false-negative results.

Reproductive management through integration of PGD and MPS-based noninvasive prenatal screening/diagnosis for a family with GJB2-associated hearing impairment

A couple with a proband child of GJB2 （encoding the gap junction protein connexin 26）-associated hearing impairment and a previous pregnancy miscarriage sought for a reproductive solution to bear a healthy child. Our study aimed to develop a cus- tomized preconception-to-neonate care trajectory to fulfill this clinical demand by integrating preimplantation genetic diagno- sis （PGD）, noninvasive prenatal testing （NIPT）, and noninvasive prenatal diagnosis （N1PD） into the strategy. Auditory and ge- netic diagnosis of the proband child was carried out to identify the disease causative mutations. The couple then received in-vitro-fertilization treatment, and eight embryos were obtained for day 5 biopsy. PGD was performed by short-tandem-repeat linkage analysis and Sanger sequencing of GJB2 gene. Transfer of a GJB2c.235delC heterozygous embryo resulted in a sin- gleton pregnancy. At the 13th week of gestation, genomic DNA （gDNA） from the trio family and cell-free DNA （cfDNA） from maternal plasma were obtained for assessment of fetal chromosomal aneuploidy and GJB2 mutations. NIPT and NIPD showed the absence of chromosomal aneuploidy and GJB2-associated disease in the fetus, which was later confirmed by inva- sire procedures and postnatal genetic/auditory diagnosis. This strategy successfully prevented the transmission of hearing im- pairment in the newborn, thus providing a valuable experience in reproductive management of similar cases and potentially other monogenic disorders.

Prenatal Testing or Screening?

Over the past 50 years,the scope and extent of prenatal diagnosis and screening for genetic disorders have improved geometrically.There has been a pendulum like swing from testing to screening back and forth as new technologies emerge.The concurrent developments of cell free fetal DNA analysis of maternal blood has dramatically changed patient’s choices towards screening.However,with the use of array comparative genomic hybridization of fetal DNA that requires diagnostic procedures(Chorionic villus sampling and amniocentesis),much more extensive diagnosis can be obtained.Until noninvasive methods can replicate what can be done with diagnostic procedures there still will be a"price to be paid"for opting for the non-invasive methods.

A Cross-sectional Real-life Study of the Prevalence,Severity,and Determinants of Metabolic Dysfunction-associated Fatty Liver Disease in Type 2 Diabetes Patients

Background and Aims:Most data on liver assessment in type 2 diabetes mellitus(T2DM)patients are from retrospective cohorts with selection bias.We aimed at appraising the feasibility,results,and benefits of an outpatient systematic noninvasive screening for metabolic dysfunction-associated fatty liver disease(MAFLD)severity and determinants in T2DM patients.Methods:We conducted a 50-week crosssectional study enrolling adult T2DM outpatients from a diabetes clinic.An algorithm based on guidelines was applied using simple bioclinical scores and,if applicable,ultrasound and/or elastometry.Results:Two hundred and thirteen patients were included.Mean age and body mass index were 62 years and 31 kg/m^(2) and 29% of patients had abnormal transaminase levels.The acceptance rate of additional liver examinations was 92%.The prevalence of MAFLD,advanced fibrosis and cirrhosis was 87%,11%,and 4%,respectively.More than half of the cases of advanced fibrosis had not been suspected and were detected by this screening.MAFLD was associated with poor glycemic control,elevated transaminases,low HDL-C and the absence of peripheral arterial disease.Advanced fibrosis was linked to high waist circumference and excessive alcohol consumption,which should be interpreted with caution owing to the small number of patients reporting excessive consumption.Conclusions:Simple bioclinical tools allowed routine triage of T2DM patients for MAFLD severity,with high adherence of high-risk patients to subsequent noninvasive exams.