Diagnostic value of circular free DNA for colorectal cancer detection

BACKGROUND Minimally invasive or noninvasive,sensitive and accurate detection of colorectal cancer(CRC)is urgently needed in clinical practice.AIM To identify a noninvasive,sensitive and accurate circular free DNA marker detected by digital polymerase chain reaction(dPCR)for the early diagnosis of clinical CRC.METHODS A total of 195 healthy control(HC)individuals and 101 CRC patients(38 in the early CRC group and 63 in the advanced CRC group)were enrolled to establish the diagnostic model.In addition,100 HC individuals and 62 patients with CRC(30 early CRC and 32 advanced CRC groups)were included separately to validate the model.CAMK1D was dPCR.Binary logistic regression analysis was used to establish a diagnostic model including CAMK1D and CEA.RESULTS To differentiate between the 195 HCs and 101 CRC patients(38 early CRC and 63 advanced CRC patients),the common biomarkers CEA and CAMK1D were used alone or in combination to evaluate their diagnostic value.The area under the curves(AUCs)of CEA and CAMK1D were 0.773(0.711,0.834)and 0.935(0.907,0.964),respectively.When CEA and CAMK1D were analyzed together,the AUC was 0.964(0.945,0.982).In differentiating between the HC and early CRC groups,the AUC was 0.978(0.960,0.995),and the sensitivity and specificity were 88.90%and 90.80%,respectively.In differentiating between the HC and advanced CRC groups,the AUC was 0.956(0.930,0.981),and the sensitivity and specificity were 81.30%and 95.90%,respectively.After building the diagnostic model containing CEA and CAMK1D,the AUC of the CEA and CAMK1D joint model was 0.906(0.858,0.954)for the validation group.In differentiating between the HC and early CRC groups,the AUC was 0.909(0.844,0.973),and the sensitivity and specificity were 93.00%and 83.30%,respectively.In differentiating between the HC and advanced CRC groups,the AUC was 0.904(0.849,0.959),and the sensitivity and specificity were 93.00%and 75.00%,respectively.CONCLUSION We built a diagnostic model including CEA and CAMK1D for differentiating between HC individuals and CRC patients.Compared with the common biomarker CEA alone,the diagnostic model exhibited significant improvement.

Revolutionizing Non-Invasive Biomarker Discoveries: The Power of Methylation Screening Analysis in Cell-Free DNA Liquid Biopsy

Epigenetic changes of DNA, including methylation, have long been recognized as key indicators of various diseases, including aging, cancer, and neurological disorders. Biomarker discoveries based on distinct methylation patterns for both hypermethylation and hypomethylation lead the way in discovery of novel diagnosis and treatment targets. Many different approaches are present to detect the level of methylation in whole genome (whole genome bisulfite sequencing, microarray) as well as at specific loci (methylation specific PCR). Cell-free DNA (cf-DNA) found in body fluids like blood provides information about DNA methylation and serves as a less invasive approach for genetic screening. Cell-free DNA and methylation screening technologies, when combined, have the potential to transform the way we approach genetic screening and personalized therapy. These technologies can help enhance disease diagnostic accuracy and inform the development of targeted therapeutics by providing a non-invasive way for acquiring genomic information and identifying disease-associated methylation patterns. We highlight the clinical benefits of using cell-free DNA (cf-DNA) liquid biopsy analysis and available methylation screening technologies that have been crucial in identifying biomarkers for disease from patients using a non-invasive way. Powering such biomarker discoveries are various methods of cf-DNA methylation analysis such as Bisulfite Sequencing and most recently, Methylation-Specific Restriction Enzyme (MSRE-seq) Analysis, paving the way for novel epigenetic biomarker discoveries for more robust diagnosis such as early disease detection, prognosis, monitoring of disease progression and treatment response as well as discovery of novel drug targets.

Quick recovery and characterization of cell-free DNA in seminal plasma of normozoospermia and azoospermia： implications for non-invasive genetic utilities

We established a quick and reliable method for recovering cell-free seminal DNA （cfsDNA）, by using the binding-washing-elution procedure on the DNA purification column. Low variations （below 15%） among the triplicate values of cfsDNA quantity verified the reproducibility of our cfsDNA recovery method. Similar cfsDNA yield and size distribution between seminal plasma acquired by filtration and centrifugation confirmed the presence of cfsDNA. To investigate the general characterization of cfsDNA, the quantitation and size distribution of cfsDNA from normozoospermic and azoospermic semen were analyzed by real-time PCR and electrophoresis, respectively. CfsDNA concentration in semen with normozoospermia （n = 11） was 1.34 ± 0.65 μg ·mL^-1, whereas a higher cfsDNA concentration was observed in azoospermia （2.56 ± 1.43 μg ·mL^-1, n = 9）. The continuous distribution of DNA fragments ranging from -1 kb to 15 kb and a spectrum of multiples of 180-bp fragments were observed in each normozoospermic and azoospermic sample. Distinct characteristic DNA ladder fragmentations in some azoospermic samples implicated that cfsDNA originate partly from apoptotic cells. CfsDNAs of 36 selected azoospermic patients with known information of Y chromosome microdeletion were subjected to the same microdeletion analysis by multiplex PCR and PCR amplification of sY114 （1 450 bp）. All multiplex PCR reactions with cfsDNA amplified successfully and provided the same result as leukocyte DNA. PCR amplification of sY114 gave a 1 450-bp amplicon as expected. Our data suggested the potential use of cfsDNA in search of biomarker or diagnostic procedures.

Value of dynamic plasma cell-free DNA monitoring in septic shock syndrome: A case report

BACKGROUND Mortality due to septic shock is relatively high.The dynamic monitoring of plasma cell-free DNA(cfDNA)can guide the treatment of septic shock.CASE SUMMARY Herein,we present a typical case of septic shock syndrome caused by the bacilli Acinetobacter baumannii and Pantoea.The patient complained of abdominal pain,fever and chills upon admission to the Emergency Department.Marked decreases in white blood cells and procalcitonin(PCT)were observed after the patient received continuous renal replacement and extracorporeal membrane oxygenation.Plasma cfDNA levels were consistently high,peaking at 1366.40 ng/mL,as measured by a duplex real-time PCR assay with an internal control,which was developed as a novel method for the accurate quantification of cfDNA.The patient died of septic shock on HD 8,suggesting that cfDNA could be used to monitor disease progression more effectively than PCT and the other inflammatory factors measured in this case.CONCLUSION CfDNA may be a promising marker that complements other inflammatory factors to monitor disease progression in patients with septic shock.

Value of circulating cell-free DNA in diagnosis of hepatocelluar carcinoma

AIM:To investigate the value of combined detection of circulating cell-free DNA(cfDNA),a-fetal protein(AFP) and a L-fucosidase(AFU) for diagnosis of hepatocellular carcinoma(HCC).METHODS:Serum samples from 39 HCC patients and 45 normal controls were collected.Branched DNA(bDNA) was used to detect the level of cfDNA,and a receiver operating characteristic curve was employed to evaluate the diagnostic sensitivity,specificity,accuracy,positive predictive value,negative predictive value,positive likelihood ratio,negative likelihood ratio and Youden index,and to assess the diagnostic efficiency and their correlations with the clinicopathological features.AFP and AFU were detected by chemiluminescence and colorimetry,respectively.The significance of combined detection of the three biomarkers was discussed.RESULTS:cfDNA level was increased in 22 of the 39 HCC samples and in 2 of the 45 normal controls.cfDNA level in HCC samples was significantly higher than that in normal controls(P < 0.05).There were significant differences in sex and extra-and intrahepatic metastasis(P < 0.05).There was no significant correlation between cfDNA,AFP and AFU in the detection of HCC.The sensitivity of combined detection of cfDNA with one marker(AFP or AFU) and cfDNA with two markers(AFP and AFU) was 71.8%,87.2% and 89.7% vs 56.4%,53.8% and 66.7% for cfDNA,AFP and AFU used alone,respectively,the difference being statistically significant(P < 0.05).CONCLUSION:Quantitative analysis of cfDNA is sensitive and feasible,and the combined detection of cfDNA with AFP or AFU or both could improve the diagnostic sensitivity for HCC.

Detection of fusion gene in cell-free DNA of a gastric synovial sarcoma

Synovial sarcoma(SS) is genetically characterized by chromosomal translocation, which generates SYT-SSX fusion transcripts. Although SS can occur in any body part, primary gastric SS is substantially rare. Here we describe a detection of the fusion gene sequence of gastric SS in plasma cell-free DNA(cf DNA). A gastric submucosal tumor was detected in the stomach of a 27-year-old woman and diagnosed as SS. Candidate intronic primers were designed to detect the intronic fusion breakpoint and this fusion sequence was confirmed in intron 10 of SYT and intron 5 of SSX2 by genomic polymerase chain reaction(PCR) and direct sequencing. A locked nucleic acid(LNA) probe specificto the fusion sequence was designed for detecting the fusion sequence in plasma and the fusion sequence was detected in preoperative plasma cfD NA, while not detected in postoperative plasma cfD NA. This technique will be useful for monitoring translocation-derived diseases such as SS.

Multiple z-Score Based Method for Noninvasive Prenatal Test Using Cell-Free DNA in Maternal Plasma

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.

The Prognostic Value of Cell-Free DNA in Advanced Non-Small-Cell Lung Cancer

Background: Cell-free DNA (cfDNA) holds promise as a tumor marker of clinical importance. We aimed to investigate the prognostic value of baseline cfDNA in non small-cell lung cancer (NSCLC). Material and Methods: During a three-year period, patients with newly diagnosed, previously untreated advanced NSCLC were included in a consecutive, prospective marker-trial. Plasma was isolated from a pre-treatment peripheral blood sample and the level of total cfDNA was measured by an in-house assay qPCR-method. The treatment comprised carboplatin (AUC 5) intravenously day 1), and vinorelbine (30 mg/m2 intravenously day 1 and 60 mg/m2 perorally day 8) q3w for a maximum of six cycles. The primary end-point was overall survival (OS). Secondary end-points were progression free survival (PFS) and overall response rate (ORR). Results: 245 patients were included and received a minimum of 1 cycle of chemotherapy (median 4). The median OS was 8.9 months, the median PFS by intention to treat 5.4 months and the ORR was 25%. The patients were divided into four groups based on quartiles of cfDNA and subsequently dichotomized by the 75th percentile revealing a significantly worse prognosis for patients in the upper 75th percentile (median OS 4.9 months) compared to patients with lower levels (10.0 months) (HR 2.1, 95%CI 1.4 - 3.1, p 0.0001). A multivariate analysis confirmed the independent prognostic value of cfDNA. A subgroup analysis of patients with high cfDNA and poor performance status (PS = 2) identified a group of patients with even worse prognosis (median OS 2.0 versus 9.1 months, HR 3.6, 95%CI 1.4 - 9.2, p 0.0001). Similar and significant results were found when comparing level of cfDNA and PFS. Conclusions: High pre-treatment level of cfDNA seems to have a strong prognostic impact in patients with newly diagnosed advanced NSCLC. Combined with PS it identifies a patient group with minimal or no benefit of chemotherapy.

Vangenes Cell-Free DNA采血管与 Streck Cell-Free DNA采血管在无创产前检测应用中的效果比较

目的:比较Vangenes■Cell-Free DNA采血管和Streck■Cell-Free DNA 采血管对血液样品的运输保存效果.方法:通过静脉抽血, 18名志愿者分别使用Vangenes■Cell-Free DNA 采血管与Streck■Cell-Free DNA采血管各采集一管10mL血液.经过约50h运输样本回到实验室,进行血浆分离、游离DNA提取、构建文库、上机测序并进行建库浓度、测序数据的unique read counts、 GC%以及duplication值等分析.结果:经比较, Vangenes ?Cell-Free DNA采血管保存样品在血浆颜色、建库浓度以及测序数据的unique read counts、 GC%以及duplication值等达到了Streck■Cell-Free DNA 采血管保存样品相似的结果,二者不存在显著性差异.结论:在血液样品运输保存过程中, Vangenes■Cell-Free DNA采血管可有效阻止有核细胞破裂导致的基因组DNA对于游离DNA的污染,对于利用胎儿游离DNA进行的无创产前检测技术具有重要意义.

血浆cfDNA甲基化联合检测在肝癌诊断中的价值

目的 探讨血浆循环游离DNA(cfDNA)中GNB4、TCF24、Riplet、ACP1和TSPYL5基因甲基化对肝癌诊断的临床价值。方法 利用滑动窗口技术在公共数据库中筛选肝癌和正常组织间的差异甲基化标志物并分析其甲基化水平。从郑州大学第一附属医院收集肝癌、肝硬化组织和健康人白细胞样本,Sanger测序检测筛选出的标志物的甲基化水平,选出敏感度和特异度较好的标志物。收集24例肝癌、23例肝硬化和99例健康人血浆,通过甲基化特异性PCR检测上述标志物单独和组合时对肝癌的诊断性能。结果 5个标志物(GNB4、TCF24、Riplet、ACP1和TSPYL5)在肝癌样本中显著高甲基化。组织测序结果显示除TCF24外,其他标志物在肝硬化组织中的甲基化阴性率均大于80.0%。在血浆样本验证中,GNB4单独诊断肝癌的曲线下面积(AUC)最大,为0.765,敏感度为66.7%,特异度为91.8%。在多基因组合中,GNB4+TSPYL5的诊断性能最佳,AUC值为0.893,敏感度为83.3%,特异度为90.2%。结论 GNB4、Riplet、ACP1和TSPYL5甲基化可用于肝癌诊断,且联合诊断性能优于单一基因。

血浆无细胞线粒体外线粒体DNA与牙周炎临床指标的相关性研究

目的血浆无细胞线粒体外线粒体DNA(cf-exmtDNA)具有促炎潜能,本文探讨血浆cf-exmtDNA与牙周炎临床指标的相关性。方法纳入18~45岁受试者78人,其中牙周健康者11人,牙龈炎患者11人,牙周炎患者56人。检查并记录基线牙周指标、年龄、性别、体质指数(BMI)和空腹血糖(FBG)。取4 mL抗凝静脉血,采用二次离心法提取其中cf-exmtDNA,使用实时荧光定量聚合酶链式反应检测cf-exmtDNA拷贝数。比较不同牙周炎症状态组血浆cf-exmtDNA水平,并对血浆cf-exmtDNA与平均探诊深度(mPD)、平均附着水平(mCAL)、平均出血指数(mBI)、平均菌斑指数(mPLI)、年龄、FBG、BMI等指标进行相关性分析以及多重线性回归分析。结果牙周炎组血浆cf-exmtDNA水平显著高于牙周健康组(P=0.042);样本整体血浆cf-exmtDNA水平与年龄(P=0.023)、mPD(P<0.001)、mCAL(P=0.006)、mBI(P=0.026)呈正相关关系;多重回归分析中,血浆cf-exmtDNA水平主要取决于mPD。结论在18~45岁人群中,牙周炎患者血浆cf-exmtDNA水平较牙周健康者显著升高,血浆cf-exmtDNA水平与年龄、mPD、mCAL、mBI呈正相关关系。

游离DNA在肝脏疾病中的应用进展

体液活检与传统侵入性检查相比具有许多独特的优势,尤其在产前诊断、器官移植后免疫排异反应、肿瘤早期诊断和肝脏疾病诊疗方面展现出巨大潜力。在肝脏疾病领域,游离DNA参与机体内在的炎性反应和凝血过程。基于表观遗传学的研究,游离DNA片段的甲基化分析对肝脏恶性肿瘤的早期诊断和治疗大有帮助。在器官移植后,游离DNA有望成为监测免疫排异反应的重要标志物,为关于免疫排异反应机制的研究提供了新角度。文章对游离DNA的来源、提取方法、检测方法,以及其在肝移植、肝癌、肝衰竭等疾病中的研究应用进展进行综述。

循环游离DNA在肾脏疾病中的研究进展

肾脏疾病是一类严重威胁人类健康的疾病,其发病率及死亡率高,寻求高灵敏度的生物学指标有助于早期诊断和改善预后。循环游离DNA是一类存在于血浆、尿液及其他体液中游离于细胞外的双链DNA片段。循环游离DNA参与机体内在的炎症反应和凝血过程,在肾脏疾病中显著升高,可作为肾脏疾病的潜在生物标志物,对疾病的诊断和预后有重大意义。本文对循环游离DNA的生物学特性、在肾脏疾病中的作用及应用进展进行综述,以期为肾脏疾病的早期诊断及预后提供参考。

基于甲基化特异性PCR的外周血循环游离DNA甲基化检测对乳腺癌诊断价值的荟萃分析

目的通过Meta分析系统评估基于甲基化特异性PCR(MSP)的外周血循环游离DNA(cfDNA)甲基化检测对乳腺癌的诊断价值。方法检索PubMed、Embase、Cochrane数据库,检索时间为2011年1月至2021年12月,以“Breast neoplasms”“Breast cancer”“Methylation”“Cell-free DNA”等为检索词,检索有关血液循环cfDNA甲基化用于乳腺癌诊断的文献,根据纳入及排除标准筛选文献。使用QUADAS-2对纳入文献进行质量评价,提取研究数据并使用Stata 16.0对各效应量进行合并,分析异质性及来源,以Deek’s法检验发表偏倚。结果共检索318篇文献,最终纳入12篇文献的27项研究进行Meta分析,患者2195例,纳入研究存在高度异质性(I^(2)>50.00%)。采用随机效应模型合并灵敏度为0.43[95%CI(0.31~0.56)],合并特异度为0.97[95%CI(0.94~0.99)],合并阳性似然比(LR+)为16.30[95%CI(7.00~37.80)],合并阴性似然比(LR-)为0.59[95%CI(0.47~0.73)],合并诊断比值比(DOR)为28.00[95%CI(11.00~70.00)],合并AUC为0.85[95%CI(0.82~0.88)]。结论基于MSP的外周血cfDNA甲基化检测对乳腺癌有较高的辅助诊断价值。

多发性骨髓瘤血浆循环游离DNA定量检测和完整性分析

目的:研究血浆循环游离DNA(cf-DNA)在初诊多发性骨髓瘤(MM)患者筛查诊断中的作用,并探讨经化疗后患者的cf-DNA含量和完整性的变化在疾病评估中的作用。方法:收集35例初诊MM患者和18例健康志愿者的外周血标本,另选取MM患者中完成规范诱导化疗3个疗程的13例患者作为随访组。以ALU247片段及ALU115片段为目的基因,用荧光定量PCR检测患者及健康志愿者血浆中的cf-DNA含量,并以ALU247/ALU115比值计算cf-DNA的完整性。结果:MM患者ALU247、ALU115片段浓度和cf-DNA完整性均显著高于健康志愿者(均P<0.05)。经3个疗程诱导化疗的患者治疗后的ALU247和ALU115片段浓度以及cf-DNA的完整性均较治疗前显著降低(均P<0.05),且治疗前后cf-DNA完整性与血清M蛋白含量及骨髓异常浆细胞比例之间均存在较强正相关关系(r_(M蛋白)=0.703,r_(骨髓浆细胞)=0.705)。结论:cf-DNA对于MM的筛查诊断具有一定的积极意义。cf-DNA或许可以协同或者替代血清M蛋白含量及骨髓异常浆细胞比例评估患者的病情、疗效及预后。

游离DNA最新研究进展及法医学应用展望

近年来,随着DNA提取和检测技术的不断进步,游离DNA(cell-free DNA,cfDNA)已经在生命科学领域得到了广泛应用,在法医学鉴定领域中的潜在应用价值也越来越明显。本文回顾了cfDNA概念、形成机制与分类等,并阐述了cfDNA在法医学现场接触检材的个体识别和无创产前亲缘关系鉴定应用中的最新研究进展,同时总结了cfDNA在损伤推断中的应用潜力,并探讨了常用cfDNA分析方法和技术的优缺点及应用展望,为cfDNA在法医学领域的广泛应用提供新思路。

游离DNA甲基化在非肿瘤性疾病中的应用研究进展

游离DNA已成为新兴的、非侵入性的重要临床样本类型,游离DNA甲基化具有组织特异性,可以反映来源细胞的DNA甲基化状态,在临床疾病诊疗中可能成为重要的潜在标志物。该文旨在综述近年常用的游离DNA甲基化检测技术,以及游离DNA甲基化在神经退行性疾病、精神性疾病、心血管疾病、糖尿病和出生缺陷等非肿瘤疾病中的应用研究进展。

尿游离DNA的液体活检有望成为泌尿系疾病诊治的重要方向

尿液由泌尿系统直接产生,其中的游离DNA(cfDNA)携带了来源于泌尿系的基因组DNA,并且尿液样本具有非侵入性、数量不受限及获取方便等优点,因此尿cfDNA是诊治泌尿系疾病的一种很有潜力的生物标志物,对尿cfDNA进行液体活检在泌尿系疾病的诊治中具有重要的临床应用价值。本文就尿cfDNA在泌尿系疾病临床诊治中的应用研究作一综述。

磁共振动态增强结合血循环肿瘤细胞、循环游离DNA对乳腺癌的临床诊断研究

目的探讨磁共振动态增强(DCE-MRI)结合血循环肿瘤细胞(CTCs)、循环游离DNA(cfDNA)对乳腺癌的诊断价值。方法选取沧州市人民医院2020年6月至2021年11月符合纳入标准的乳腺癌病人82例(乳腺癌组)。另选同期65例乳腺良性病变且未合并其他疾病者作为良性组。比较两组DCE-MRI定量参数、CTCs、cfDNA水平,对比MRI特征,分析DCE-MRI结合CTCs、cfDNA诊断乳腺癌的价值。结果乳腺癌组DCE-MRI定量参数速度常数(K^(trans))值、速率常数(K_(ep))值、Alu247/115水平及CTCs计数分别为(0.18±0.04)min、(1.32±0.27)min、(12.97±2.14)个/毫升、0.97±0.32,比良性组高(0.05±0.01)min、(0.51±0.11)min、(10.15±1.45)个/毫升、0.55±0.12(P<0.05)。良性组边缘多光滑,形状规则,内部强化以均匀为主,时间-信号强度曲线(TIC)以Ⅰ型多见,早期增强率<60%;乳腺癌组边缘毛刺征、形状不规则,内部不均匀强化为主,以TIC分型Ⅲ型多见,早期增强率<60%占比高。两组MRI形态表现比较差异有统计学意义(P<0.05)。淋巴结转移、Ⅲ~Ⅳ期者CTCs数量、Alu247/115水平明显高于无淋巴结转移、Ⅰ~Ⅱ期者(P<0.05)。受试者操作特征(ROC)曲线分析结果显示,三者联合(DCE-MRI+CTCs+cfDNA)诊断乳腺癌的曲线下面积(AUC)、灵敏度、特异度分别为0.86、0.93、0.84,诊断效果优于单一检测CTCs、cfDNA检测(P<0.05)。结论乳腺癌病人CTCs、cfDNA存在异常表达,可能与病人临床分期、淋巴结转移有关,相对单一实验室检测,DCE-MRI结合CTCs、cfDNA检测能获取更多可靠信息。

结核分枝杆菌游离DNA检测在肺外结核诊断中的价值

目的探讨结核分枝杆菌(MTB)游离DNA(cfDNA)检测技术在肺外结核诊断中的应用价值。方法选取2021年7月至2022年12月该院收治的高度疑似肺外结核患者266例作为研究对象,根据临床诊断结果,分为肺外结核组117例和非结核组149例。所有患者均采集脑脊液、胸腔积液、尿液及血液标本,并同时进行MTB cfDNA检测、结核分枝杆菌及利福平耐药快速检测(GeneXpert MTB/RIF)、涂片抗酸染色及血液标本结核感染T细胞γ干扰素释放试验(TB-IGRA)。通过分析4种检测方法的效能指标评估MTB cfDNA检测技术在肺外结核病诊断中的临床价值。结果MTB cfDNA检测法对脑脊液、胸腔积液、尿液中MTB的检出率均显著高于涂片抗酸染色(χ^(2)=5.714、9.289、9.226,P=0.017、0.002、0.002)。虽然MTB cfDNA检测法对脑脊液、尿液中MTB的检出率高于GeneXpert MTB/RIF,但差异无统计学意义(χ^(2)=0.143,P=0.705;χ^(2)=0.648,P=0.421);MTB cfDNA检测法对胸腔积液中MTB的检出率显著高于GeneXpert MTB/RIF(χ^(2)=4.222,P=0.040)。在肺外结核组,MTB cfDNA检测法的MTB检出率为26.50%(31/117),显著高于GeneXpert MTB/RIF[15.38%(18/117)]和涂片抗酸染色[3.42%(4/117)],差异均有统计学意义(χ^(2)=4.362,P=0.037;χ^(2)=24.492,P<0.05);而TB-IGRA的MTB检出率为72.65%(85/117),显著高于MTB cfDNA检测(χ^(2)=49.850,P<0.001)。MTB cfDNA检测法的灵敏度、特异度、阳性预测值、阴性预测值分别为26.50%、100.00%、100.00%、63.40%,其灵敏度显著高于GeneXpert MTB/RIF(15.38%)和涂片抗酸染色(3.42%),但低于TB-IGRA(72.65%),差异均有统计学意义(P<0.05)。结论MTB cfDNA检测在肺外结核诊断中具有较高的灵敏度及特异度,故MTB cfDNA检测在肺外结核诊断中具有较好的应用前景。