Diagnostic Value of Targeted Next-generation Sequencing in Pulmonary Mycobacterial Infections

Objective This study aimed to explore the diagnostic value of novel technique-targeted next-generation sequencing(tNGS)of bronchoalveolar lavage fluid(BALF)in pulmonary mycobacterial infections.Methods This retrospective study was conducted on patients who underwent bronchoscopy and tNGS,smear microscopy,and mycobacterial culture of BALF.Patients with positive Mycobacterium tuberculosis(MTB)culture or GeneXpert results were classified into the tuberculosis case group.Those diagnosed with nontuberculous mycobacteria(NTM)-pulmonary disease(NTM-PD)composed the case group of NTM-PD patients.The control group comprised patients without tuberculosis or NTM-PD.Sensitivity,specificity,and receiver operating characteristic(ROC)curves were used to evaluate the diagnostic performance.Results For tuberculosis patients with positive mycobacterial culture results,the areas under the ROC curves(AUCs)for tNGS,GeneXpert,and smear microscopy were 0.975(95%CI:0.935,1.000),0.925(95%CI:0.859,0.991),and 0.675(95%CI:0.563,0.787),respectively.For tuberculosis patients with positive GeneXpert results,the AUCs of tNGS,culture,and smear microscopy were 0.970(95%CI:0.931,1.000),0.850(95%CI:0.770,0.930),and 0.680(95%CI:0.579,0.781),respectively.For NTM-PD,the AUCs of tNGS,culture,and smear-positive but GeneXpert-negative results were 0.987(95%CI:0.967,1.000),0.750(95%CI:0.622,0.878),and 0.615(95%CI:0.479,0.752),respectively.The sensitivity and specificity of tNGS in NTM-PD patients were 100%and 97.5%,respectively.Conclusion tNGS demonstrated superior diagnostic efficacy in mycobacterial infection,indicating its potential for clinical application.

Construction of SNP genetic maps based on targeted next-generation sequencing and QTL mapping of vital agronomic traits in faba bean(Vicia faba L.)

Owing to the limitation of a large genome size(~13 Gb),the genetic and gene mapping studies on faba bean(Vicia faba L.)are lagging far behind those for other legumes.In this study,we selected three purified faba bean lines(Yundou 8137,H0003712,and H000572)as parents and constructed two F2 populations.These two F2 populations,namely 167 F2 plants in Pop1(Yundou 8137×H0003712)and 204 F2 plants in Pop2(H000572×Yundou 8137),were genotyped using a targeted next-generation sequencing(TNGS)genotyping platform,and two high-density single nucleotide polymorphisms(SNP)genetic linkage maps of faba bean were constructed.The map constructed from Pop1 contained 5103 SNPs with a length of 1333.31 cM and an average marker density of 0.26 cM.The map constructed from Pop2 contained 1904 SNPs with a greater length of 1610.61 cM.In these two F2 populations,QTL mapping identified 98 QTLs for 14 agronomic traits related to the flowers,pods,plant types and grains.The two maps were then merged into an integrated genetic linkage map containing 6895 SNPs,with a length of 3324.48 cM.These results not only lay the foundation for fine mapping and map-based cloning of related genes,but can also accelerate the molecular marker-assisted breeding of faba bean.

Targeted next-generation sequencing identifies a novel nonsense mutation in ANK1 for hereditary spherocytosis:A case report

BACKGROUND Hereditary spherocytosis(HS)is characterized by anemia,jaundice,splenomegaly,and cholelithiasis,and is caused by abnormal genes encoding red blood cell membrane components.The most common mutations found in HS are in the ANK1 gene.CASE SUMMARY A 4-mo-old girl was admitted to our hospital with pallor that had lasted for more than 2 mo.She presented with jaundice,anemia and splenomegaly.A heterozygous mutation of ANK1(exon23:c.G2467T:p.E823X)was identified,and the mutation was determined to be autosomal dominant.This mutation is linked to the relatively serious anemia she had after birth;this anemia improved with age.CONCLUSION The utilization of next-generation sequencing may assist with the accurate diagnosis of HS,especially in atypical cases.

Prevalence of gene mutations in a Chinese 46,XY disorders of sex development cohort detected by targeted next-generation sequencing

46,XY disorders of sex development(DSD)is characterized by incomplete masculinization genitalia,with gonadal dysplasia and with/without the presence of Mullerian structures.At least 30 genes related to 46,XY DSD have been found.However,the clinical phenotypes of patients with different gene mutations overlap,and accurate diagnosis relies on gene sequencing technology.Therefore,this study aims to determine the prevalence of pathogenic mutations in a Chinese cohort with 46,XY DSD by the targeted nextgeneration sequencing(NGS)technology.Eighty-seven 46,XY DSD patients were enrolled from the Peking Union Medical College Hospital(Beijing,China).A total of fifty-four rare variants were identified in 60 patients with 46,XY DSD.The incidence of these rare variants was approximately 69.0%(60/87).Twenty-five novel variants and 29 reported variants were identified.Based on the American College of Medical Genetics and Genomics(ACMG)guidelines,thirty-three variants were classified as pathogenic or likely pathogenic variants and 21 variants were assessed as variants of uncertain significance.The overall diagnostic rate was about 42.5%based on the pathogenic and likely pathogenic variants.Androgen receptor{AR),steroid 5-alpha-reductase 2(SRD5A2)and nuclear receptor subfamily 5 Group A member 1(NR5A1)gene variants were identified in 21,13 and 13 patients,respectively.The incidence of these three gene variants was about 78.3%(47/60)in patients with rare variants.It is concluded that targeted NGS is an effective method to detect pathogenic mutations in 46,XY DSD patients and AR,SRD5A2,and NR5A1 genes were the most common pathogenic genes in our cohort.

A genomic study of adult-onset idiopathic hypoparathyroidism in Chinese by targeted next-generation sequencing

Objective To screen gene mutation in adult-onset hypoparathyroidism in Chinese through the targeted nextgeneration sequencing(NGS).Methods We recruited 17patients with adult-onset hypoparathyroidism who were regularly followed or newly diagnosed at our centre during the past one year.Nine of them developed hypercalciuria during the treatment with calcium and vitamin D.

The Role of Quality Control in Targeted Next-generation Sequencing Library Preparation

Next-generation sequencing （NGS） is getting routinely used in the diagnosis of hereditary diseases, such as human cardiomyopathies. Hence. it is of utter importance to secure high quality sequencing data, enabling the identification of disease-relevant mutations or the conclusion of negative test results. During the process of sample preparation, each protocol for target enrichment library preparation has its own requirements for quality control （QC）; however, there is little evi- dence on the actual impact of these guidelines on resulting data quality. In this study, we analyzed the impact of QC during the diverse library preparation steps of Agilent SureSelect XT target enrichment and lllumina sequencing. We quantified the parameters for a cohort of around 600 samples, which include starting amount of DNA, amount of sheared DNA, smallest and largest fragment size of the starting DNA; amount of DNA after the pre-PCR, and smallest and largest fragment size of the resulting DNA; as well as the amount of the final library, the corresponding smallest and largest fragment size, and the number of detected variants. Intriguingly, there is a high tolerance for variations in all QC steps, meaning that within the boundaries proposed in the current study, a considerable variance at each step of QC can be well tolerated without compromising NGS quality.

Impact of next-generation sequencing on antimicrobial treatment in immunocompromised adults with suspected infections

BACKGROUND:Prompt pathogen identification can have a substantial impact on the optimization of antimicrobial treatment.The objective of the study was to assess the diagnostic value of next-generation sequencing(NGS)for identifying pathogen and its clinical impact on antimicrobial intervention in immunocompromised patients with suspected infections.METHODS:This was a retrospective study.Between January and August 2020,47 adult immunocompromised patients underwent NGS testing under the following clinical conditions:1)prolonged fever and negative conventional cultures;2)new-onset fever despite empiric antimicrobial treatment;and 3)afebrile with suspected infections on imaging.Clinical data,including conventional microbial test results and antimicrobial treatment before and after NGS,were collected.Data were analyzed according to documented changes in antimicrobial treatment(escalated,no change,or deescalated)after the NGS results.RESULTS:The median time from hospitalization to NGS sampling was 19 d.Clinically relevant pathogens were detected via NGS in 61.7% of patients(29/47),more than half of whom suffered from fungemia(n=17),resulting in an antimicrobial escalation in 53.2% of patients(25/47)and antimicrobial de-escalation in 0.2% of patients(1/47).Antimicrobial changes were mostly due to the identification of fastidious organisms such as Legionella,Pneumocystis jirovecii,and Candida.In the remaining three cases,NGS detected clinically relevant pathogens also detected by conventional cultures a few days later.The antimicrobial treatment was subsequently adjusted according to the susceptibility test results.Overall,NGS changed antimicrobial management in 55.3%(26/47)of patients,and conventional culture detected clinically relevant pathogens in 14.9% of the patients(7/47).CONCLUSION:With its rapid identification and high sensitivity,NGS could be a promising tool for identifying relevant pathogens and enabling rapid appropriate treatment in immunocompromised patients with suspected infections.

Bronchoalveolar lavage fluid metagenomic next-generation sequencing assay for identifying pathogens in lung cancer patients

Background:For patients with lung cancer,timely identification of new lung lesions as infectious or non-infectious,and accurate identification of pathogens is very important in improving OS of patients.As a new auxiliary examination,metagenomic next-generation sequencing(mNGS)is believed to be more accurate in diagnosing infectious diseases in patients without underlying diseases,compared with conventional microbial tests(CMTs).We designed this study tofind out whether mNGS has better performance in distinguishing infectious and non-infectious diseases in lung cancer patients using bronchoalveolar lavagefluid(BALF).Materials and Methods:This study was a real-world retrospective review based on electronic medical records of lung cancer patients with bronchoalveolar lavage(BAL)and BALF commercial mNGS testing as part of clinical care from 1 April 2019 through 30 April 2022 at The First Affiliated Hospital of Sun Yat-sen University.164 patients were included in this study.Patients were categorized into the pulmonary non-infectious disease(PNID)group(n=64)and the pulmonary infectious disease(PID)group(n=100)groups based onfinal diagnoses.Results:BALF mNGS increased the sensitivity rate by 60%compared to CMTs(81%vs.21%,p<0.05),whereas there was no significant difference in specificity(75%vs.98.4%,p>0.1).Among the patients with PID,bacteria were the most common cause of infection.Fungal infections occurred in 32%of patients,and Pneumocystis Yersini was most common.Patients with Tyrosine kinase inhibitors(TKIs)therapy possess longer overall survival(OS)than other anti-cancer agents,the difference between TKIs and immuno-checkpoint inhibitors(ICIs)was insignificant(median OS TKIs vs.ICIs vs.Anti-angiogenic vs.Chemo vs.Radiotherapy=76 vs.84 vs.61 vs.58 vs.60).Conclusions:our study indicates that BALF mNGS can add value by improving overall sensitivity in lung cancer patients with potential pulmonary infection,and was outstanding in identifying Pneumocystis infection.It could be able to help physicians adjust the follow-up treatment to avoid the abuse of antibiotics.

Brain abscess from oral microbiota approached by metagenomic next-generation sequencing: A case report and review of literature

BACKGROUND Brain abscess is a serious and potentially fatal disease caused primarily by microbial infection.Although progress has been made in the diagnosis and treatment of brain abscesses,the diagnostic timeliness of pathogens needs to be improved.CASE SUMMARY We report the case of a 54-year-old male with a brain abscess caused by oral bacteria.The patient recovered well after receiving a combination of metagenomic next-generation sequencing(mNGS)-assisted guided medication and surgery.CONCLUSION Therefore,mNGS may be widely applied to identify the pathogenic microor-ganisms of brain abscesses and guide precision medicine.

Metagenomic next-generation sequencing may assist diagnosis of osteomyelitis caused by Mycobacterium houstonense:A case report

BACKGROUND Mycobacterium houstonense(M.houstonense)belongs to the nontuberculous mycobacterium group.Infection caused by M.houstonense is prone to recurrence.CASE SUMMARY We present a patient who was diagnosed with osteomyelitis caused by M.houstonense and treated with a combination of cefoxitin,and amikacin combined with linezolid.CONCLUSION The emergence of metagenomic next-generation sequencing(NGS)has brought new hope for the diagnosis and treatment of listeria meningitis.NGS can analyze a large number of nucleic acid sequences in a short time and quickly determine the pathogen species in the sample.Compared with traditional cerebrospinal fluid culture,NGS can greatly shorten the diagnosis time and provide strong support for the timely treatment of patients.Regarding treatment,NGS can also play an important role.Rapid and accurate diagnosis can enable patients to start targeted treatment as soon as possible and improve the treatment effect.At the same time,by monitoring the changes in pathogen resistance,the treatment plan can be adjusted in time to avoid treatment failure.

Diagnosis of Acute Q Fever in an Elderly Patient Using Metagenomic Next-Generation Sequencing:A Case Report

Query fever(Q fever)is a globally spread zoonotic disease caused by Coxiella burnetii,commonly found in natural foci but rarely seen in Hebei Province.The clinical manifestations of Q fever are diverse and nonspecific,which often leads to missed or incorrect diagnoses in clinical practice.This article reports a case of acute Q fever diagnosed in an elderly patient using metagenomic next-generation sequencing.

To Analyze the Sensitivity of RT-PCR Assays Employing S Gene Target Failure with Whole Genome Sequencing Data during Third Wave by SARS-CoV-2 Omicron Variant

Introduction: Omicron is a highly divergent variant of concern (VOCs) of a severe acute respiratory syndrome SARS-CoV-2. It carries a high number of mutations in its spike protein hence;it is more transmissible in the community by immune evasion mechanisms. Due to mutation within S gene, most Omicron variants have reported S gene target failure (SGTF) with some commercially available PCR kits. Such diagnostic features can be used as markers to screen Omicron. However, Whole Genome Sequencing (WGS) is the only gold standard approach to confirm novel microorganisms at genetically level as similar mutations can also be found in other variants that are circulating at low frequencies worldwide. This Retrospective study is aimed to assess RT-PCR sensitivity in the detection of S gene target failure in comparison with whole genome sequencing to detect variants of Omicron. Methods: We have analysed retrospective data of SARS-CoV-2 positive RT-PCR samples for S gene target failure (SGTF) with TaqPath COVID-19 RT-PCR Combo Kit (ThermoFisher) and combined with sequencing technologies to study the emerged pattern of SARS-CoV-2 variants during third wave at the tertiary care centre, Surat. Results: From the first day of December 2021 till the end of February 2022, a total of 321,803 diagnostic RT-PCR tests for SARS-CoV-2 were performed, of which 20,566 positive cases were reported at our tertiary care centre with an average cumulative positivity of 6.39% over a period of three months. In the month of December 21 samples characterized by the SGTF (70/129) were suggestive of being infected by the Omicron variant and identified as Omicron (B.1.1.529 lineage) when sequence. In the month of January, we analysed a subset of samples (n = 618) with SGTF (24%) and without SGTF (76%) with Ct values Conclusions: During the COVID-19 pandemic, it took almost more than 15 days to diagnose infection and identify pathogen by sequencing technology. In contrast to that molecular assay provided quick identification with the help of SGTF phenomenon within 5 hours of duration. This strategy helps scientists and health policymakers for the quick isolation and identification of clusters. That ultimately results in a decreased transmission of pathogen among the community.

Application of next-generation sequencing technology to precision medicine in cancer: joint consensus of the Tumor Biomarker Committee of the Chinese Society of Clinical Oncology

Next-generation sequencing(NGS) technology is capable of sequencing millions or billions of DNA molecules simultaneously.Therefore, it represents a promising tool for the analysis of molecular targets for the initial diagnosis of disease, monitoring of disease progression, and identifying the mechanism of drug resistance. On behalf of the Tumor Biomarker Committee of the Chinese Society of Clinical Oncology(CSCO) and the China Actionable Genome Consortium(CAGC), the present expert group hereby proposes advisory guidelines on clinical applications of NGS technology for the analysis of cancer driver genes for precision cancer therapy. This group comprises an assembly of laboratory cancer geneticists, clinical oncologists, bioinformaticians,pathologists, and other professionals. After multiple rounds of discussions and revisions, the expert group has reached a preliminary consensus on the need of NGS in clinical diagnosis, its regulation, and compliance standards in clinical sample collection. Moreover, it has prepared NGS criteria, the sequencing standard operation procedure(SOP), data analysis, report, and NGS platform certification and validation.

Effects of viral infection and microbial diversity on patients with sepsis:A retrospective study based on metagenomic next-generation sequencing

BACKGROUND: The study aims to investigate the performance of a metagenomic next-generationsequencing (NGS)-based diagnostic technique for the identifi cation of potential bacterial and viral infectionsand eff ects of concomitant viral infection on the survival rate of intensive care unit (ICU) sepsis patients.METHODS: A total of 74 ICU patients with sepsis who were admitted to our institution from February1, 2018 to June 30, 2019 were enrolled. Separate blood samples were collected from patients for bloodcultures and metagenomic NGS when the patients’ body temperature was higher than 38 °C. Patients’demographic data, including gender, age, ICU duration, ICU scores, and laboratory results, were recorded.The correlations between pathogen types and sepsis severity and survival rate were evaluated.RESULTS: NGS produced higher positive results (105 of 118;88.98%) than blood cultures(18 of 118;15.25%) over the whole study period. Concomitant viral infection correlated closelywith sepsis severity and had the negative effect on the survival of patients with sepsis. However,correlation analysis indicated that the bacterial variety did not correlate with the severity of sepsis.CONCLUSIONS: Concurrent viral load correlates closely with the severity of sepsis and thesurvival rate of the ICU sepsis patients. This suggests that prophylactic administration of antiviraldrugs combined with antibiotics may be benefi cial to ICU sepsis patients.

Genomic characterization of esophageal squamous cellcarcinoma:insights from next-generation sequencing

Two major types of cancer occur in the esophagus: squamous cell carcinoma, which is associated with chronic smoking and alcohol consumption, and adenocarcinoma, which typically arises in gastric reflux-associated Barrett's esophagus. Although there is increasing incidence of esophageal adenocarcinoma in Western counties, esophageal squamous cell carcinoma(ESCC) accounts for most esophageal malignancies in East Asia, including China and Japan. Technological advances allowing for massively parallel, high-throughput next-generation sequencing(NGS) of DNA have enabled comprehensive characterization of somatic mutations in large numbers of tumor samples. Recently, several studies were published in which whole exome or whole genome sequencing was performed in ESCC tumors and compared with matched normal DNA. Mutations were validated in several genes, including in TP53, CDKN2 A, FAT1, NOTCH1, PIK3 CA, KMT2 D and NFE2L2, which had been previously implicated in ESCC. Several new recurrent alterations have also been identified in ESCC. Combining the clinicopathological characteristics of patients with information obtained from NGS studies may lead to the development of effective diagnostic and therapeutic approaches for ESCC. As this research becomes more prominent, it is important that gastroenterologist become familiar with the various NGS technologies and the results generated using these methods. In the present study, we describe recent research approaches using NGS in ESCC.

An updated review of gastric cancer in the next-generation sequencing era:Insights from bench to bedside and vice versa

Gastric cancer(GC)is one of the most common malignancies and remains the second leading cause of cancer-related death worldwide.There is an increasing understanding of the roles that genetic and epigenetic alterations play in GCs.Recent studies using nextgeneration sequencing(NGS)have revealed a number of potential cancer-driving genes in GC.Whole-exome sequencing of GC has identified recurrent somatic mutations in the chromatin remodeling gene ARID1A and alterations in the cell adhesion gene FAT4,a member of the cadherin gene family.Mutations in chromatin remodeling genes(ARID1A,MLL3 and MLL)have been found in 47%of GCs.Whole-genome sequencing and whole-transcriptome sequencing analyses have also discovered novel alterations in GC.Recent studies of cancer epigenetics have revealed widespread alterations in genes involved in the epigenetic machinery,such as DNA methylation,histone modifications,nucleosome positioning,noncoding RNAs and microRNAs.Recent advances in molecular research on GC have resulted in the introduction of new diagnostic and therapeutic strategies into clinical settings.The antihuman epidermal growth receptor 2(HER2)antibody trastuzumab has led to an era of personalized therapy in GC.In addition,ramucirumab,a monoclonal antibody targeting vascular endothelial growth factor receptor(VEGFR)-2,is the first biological treatment that showed survival benefits as a single-agent therapy in patients with advanced GC who progressed after firstline chemotherapy.Using NGS to systematically identify gene alterations in GC is a promising approach with remarkable potential for investigating the pathogenesis of GC and identifying novel therapeutic targets,as well as useful biomarkers.In this review,we will summarize the recent advances in the understanding of the molecular pathogenesis of GC,focusing on the potential use of these genetic and epigenetic alterations as diagnostic biomarkers and novel therapeutic targets.

Next-generation sequencing technology for diagnosis and efficacy evaluation of a patient with visceral leishmaniasis: A case report

BACKGROUND Visceral leishmaniasis(VL)is a parasitic disease caused by Leishmania and transmitted by infected sand flies.VL has a low incidence in China,and its clinical presentation is complex and atypical.This disease is easily misdiagnosed and can become life-threatening within a short period of time.Therefore,early,rapid and accurate diagnosis and treatment of the disease are essential.CASE SUMMARY A 25-year-old male patient presented with the clinical manifestations of irregular fever,hepatosplenomegaly,increased polyclonal globulin,and pancytopenia.The first bone marrow puncture biopsy did not provide a clear diagnosis.In order to relieve the pressure and discomfort of the organs caused by the enlarged spleen and to confirm the diagnosis,splenectomy was performed,and hemophagocytic syndrome was diagnosed by pathological examination of the spleen biopsy.Following bone marrow and spleen pathological re-diagnosis and metagenomic next-generation sequencing(mNGS)technology detection,the patient was finally diagnosed with VL.After treatment with liposomal amphotericin B,the body temperature quickly returned to normal and the hemocytes recovered gradually.Post-treatment re-examination of the bone marrow puncture and mNGS data showed that Leishmania was not detected.CONCLUSION As a fast and accurate detection method,mNGS can diagnose and evaluate the efficacy of treatment in suspicious cases of leishmaniasis.

Clinical applications of next-generation sequencing in colorectal cancers

Like other solid tumors,colorectal cancer(CRC) is a genomic disorder in which various types of genomic alterations,such as point mutations,genomic rearrangements,gene fusions,or chromosomal copy number alterations,can contribute to the initiation and progression of the disease. The advent of a new DNA sequencing technology known as next-generation sequencing(NGS) has revolutionized the speed and throughput of cataloguing such cancer-related genomic alterations. Now the challenge is how to exploit this advanced technology to better understand the underlying molecular mechanism of colorectal carcinogenesis and to identify clinically relevant genetic biomarkers for diagnosis and personalized therapeutics. In this review,we will introduce NGS-based cancer genomics studies focusing on those of CRC,including a recent large-scale report from the Cancer Genome Atlas. We will mainly discuss how NGS-based exome-,whole genome- and methylome-sequencing have extended our understanding of colorectal carcinogenesis. We will also introduce the unique genomic features of CRC discovered by NGS technologies,such as the relationship with bacterial pathogens and the massive genomic rearrangements of chromothripsis. Finally,we will discuss the necessary steps prior to development of a clinical application of NGS-related findings for the advanced management of patients with CRC.

Next-generation sequencing in clinical virology: Discovery of new viruses

Viruses are a cause of significant health problem world-wide, especially in the developing nations. Due to different anthropological activities, human populations are exposed to different viral pathogens, many of which emerge as outbreaks. In such situations, discovery of novel viruses is utmost important for deciding prevention and treatment strategies. Since last century, a number of different virus discovery methods, based on cell culture inoculation, sequence-independent PCR have been used for identification of a variety of viruses. However, the recent emergence and commercial availability of nextgeneration sequencers(NGS) has entirely changed the field of virus discovery. These massively parallel sequencing platforms can sequence a mixture of genetic materials from a very heterogeneous mix, with high sensitivity. Moreover, these platforms work in a sequenceindependent manner, making them ideal tools for virus discovery. However, for their application in clinics, sample preparation or enrichment is necessary to detect low abundance virus populations. A number of techniques have also been developed for enrichment or viral nucleic acids. In this manuscript, we review the evolution of sequencing; NGS technologies available today as well as widely used virus enrichment technologies. We also discuss the challenges associated with their applications in the clinical virus discovery.

Novel mutations in CRYBB1/CRYBB2 identified by targeted exome sequencing in Chinese families with congenital cataract

AIM：To summarize the phenotypes and identify the underlying genetic cause of the CRYBB1 and CRYBB2 gene responsible for congenital cataract in two Chinese families.METHODS：Detailed family histories and clinical data were collected from patients during an ophthalmologic examination. Of 523 inheritable genetic vision systemrelated genes were captured and sequenced by targeted next-generation sequencing,and the results were confirmed by Sanger sequencing. The possible functional impacts of an amino acid substitution were performed with Poly Phen-2 and SIFT predictions.RESULTS：The patients in the two families were affected with congenital cataract. Sixty-five （FAMILY-1） and sixty two （FAMILY-2） single-nucleotide polymorphisms and indels were selected by recommended filtering criteria.Segregation was then analyzed by applying Sanger sequencing with the family members. A heterozygous CRYBB1 mutation in exon 4 （c.347T〉C, p.L116P） was identified in sixteen patients in FAMILY-1. A heterozygous CRYBB2 mutation in exon 5 （c.355G〉A, p.G119R） was identified in three patients in FAMILY-2. Each mutation cosegregated with the affected individuals and did not exist in unaffected family members and 200 unrelated normal controls.The mutation was predicted to be highly conservative and to be deleterious by both PolyPhen-2 and SIFT.CONCLUSION：TheCRYBB1 mutation（c.347T〉C）and CRYBB2 mutation （c.355G〉A） are novel in patients with congenital cataract. We summarize the variable phenotypes among the patients, which expanded the phenotypic spectrum of congenital cataract in a different ethnic background.