PURPOSE:The aim of this study was to compare the performance of the MeltPro Myco assay with that of MALDI-TOF mass spectrometry for detecting nontuberculous mycobacteria (NTMs) from clinical isolates and samples. METHOD:A total of 308 suspected NTM isolates from sputum or BALF samples of patients with pulmonary diseases were retrospectively analyzed. DNA sequencing was used as the gold standard method. An additional 164 clinical samples from patients with pulmonary diseases were used to assess the suitability of the MeltPro Myco assay for clinical use. RESULT:The results indicated that for NTM identification, the MeltPro Myco assay and MALDI-TOF had sensitivities of 99.3% and 97.2% and specificities of 90.9% and 100%.The MeltPro Myco assay achievies a sensitivity and specificity of 79.4% and 95.2% for the detection of NTM in clinical samples. CONCLUSION:Given its rapidity, straightforward operation, high accuracy, cost-effectiveness, and good reproducibility, the MeltPro Myco assay can be a valuable tool for timely screening of lung disease caused by NTM and has the potential to improve the diagnosis and prevention of NTM pulmonary disease in highly endemic areas.

The drug-resistant tuberculosis (TB) is a major public health problem worldwide. Capreomycin (Cm) has been used as an effective drug to treat multidrug-resistant tuberculosis (MDR-TB) since 1960s. Although Cm and amikacin (Am) are historically grouped as second-line injectable drugs (SLIDs) with shared resistance mechanisms (e.g., rrs mutations), recent clinical and genetic data suggests divergent evolutionary pathways. It remains unclear how Mtb evolves from sensitive to resistant phenotypes under Cm pressure. In this study, we generate in vitro drug-resistant Mtb models to systematically study the evolutionary mechanisms of Cm and Am cross-resistance. By integrating transcriptome/proteome/metabolome analyses, we identify both genetic mutations and non-genetic factors contributing to Cm resistance. We show that Cm- and Am-selective pressures trigger independent cross-resistance mechanisms revealed by distinct genetic mutations. Gly232AsptlyA and Trp120fstlyA mutations are directly associated with low level resistance to Cm, whereas Ala48ValmmaA2, Gln19ArgrpmA and variant at the promoter of eis (which encodes an N-acetyltransferase; c.-14c>t) mediate cross-resistance pathways specific to Cm selection. In addition, dysregulations of non-genetic factors including metal ions transportations and lipids metabolism also lead to drug-resistance. In summary, our findings systematically characterize the molecular mechanisms of Cm resistance, demonstrating that the ''genetic factors'' (DNA mutations) are often the drivers that induce the stable ''non-genetic factors'' (dysregulated pathways, such as eis overexpression, metal ion transport, and metabolic reprogramming), which ultimately mediate the resistance phenotype. These insights uncover the evolutionary trajectory of Cm and Am cross-resistance and provide potential strategies for optimizing Cm use to treat MDR-TB. Importance: Tuberculosis, caused by Mycobacterium tuberculosis, is responsible for the highest mortality rate worldwide among single pathogen infections. The diagnosis and treatment of drug-resistant tuberculosis pose a global challenge. Capreomycin (Cm) is one of the drugs utilized in the management of multidrug-resistant tuberculosis (MDR-TB) and extensively drug-resistant tuberculosis (XDR-TB). Currently, the clinical detection of mutations in rrs gene is employed to identify Cm resistance. However, the sensitivity and specificity of this method are suboptimal. To further investigate the mechanisms underlying Cm resistance, this study established a microevolution model in vitro, thereby simulating the progression of MTB from susceptible strains to drug-resistant strains. The genomes of each generation were sequenced throughout the evolutionary process. In addition, the transcriptome-proteome-lipid metabolome analysis of representative strains were performed. Our findings reveal that Cm resistance is a combinatory effect of both genetic and non-genetic changes, providing potential optimization strategies to the molecular drug sensitivity testing used in clinical practice.

Background: Rifampicin (RIF) and multidrug-resistant tuberculosis (TB) are major public health threats. As conventional phenotypic drug susceptibility testing requires two-eight weeks, molecular diagnostic assays are widely used to determine drug resistance.Methods: Clinical Mycobacterium tuberculosis isolates with consistent drug susceptibility results, tested using microbroth dilution and proportion methods in Lo center dot wenstein-Jensen medium from patients with TB in Guangdong province were utilized to evaluate MeltPro TB and whole-genome sequencing (WGS) assays in detecting resistance to RIF, isoniazid (INH), ethambutol (EMB), fluoroquinolones (FQ), and streptomycin (SM). Solid phenotypic drug susceptibility testing was used as the gold standard to evaluate the detection capacity of MeltPro TB on clinical sputum samples of patients with TB.Results: Similar to WGS, MeltPro TB successfully detected RIF, INH, and SM resistance with sensitivities of 86.3, 84.8, and 86.6 %, respectively. However, the resistant isolate detection rates were only 58.1 and 69.6 % for EMB and FQ-resistant strains. For clinical specimens, MeltPro TB still showed good detectable rates of RIF and INH resistance, with sensitivities of 82.4 % and 95.2 %, respectively. Detectable rates of FQ and EMB resistance were low: 77.8 % and 35.3 %, respectively.Conclusions: MeltPro TB can detect known DNA mutations associated with drug resistance in Mycobacterium tuberculosis strains with comparable efficacy to WGS. For FQ and EMB resistance testing, MeltPro TB requires optimization and is unsuitable for general use. MeltPro TB can be used for diagnosis of RIF and multidrugresistant tuberculosis to rapidly initiate appropriate anti-TB drug therapy.

Streptomycin-resistant (SM-resistant) Mycobacterium tuberculosis (M. tuberculosis) is a major concern in tuberculosis (TB) treatment. However, the mechanisms underlying streptomycin resistance remain unclear. This study primarily aimed to perform preliminary screening of genes associated with streptomycin resistance through conjoint analysis of multiple genomics. Genome-wide methylation, transcriptome, and proteome analyses were used to elucidate the associations between specific genes and streptomycin resistance in M. tuberculosis H37Rv. Methylation analysis revealed that 188 genes were differentially methylated between the SM-resistant and normal groups, with 89 and 99 genes being hypermethylated and hypomethylated, respectively. Furthermore, functional analysis revealed that these 188 differentially methylated genes were enriched in 74 pathways, with most of them being enriched in metabolic pathways. Transcriptome analysis revealed that 516 genes were differentially expressed between the drug-resistant and normal groups, with 263 and 253 genes being significantly upregulated and downregulated, respectively. KEGG analysis indicated that these 516 genes were enriched in 79 pathways, with most of them being enriched in histidine metabolism. The methylation level was negatively related to mRNA abundance. Proteome analysis revealed 56 differentially expressed proteins, including 14 upregulated and 42 downregulated proteins. Moreover, three hub genes (coaE, fadE5, and mprA) were obtained using synthetic analysis. The findings of this study suggest that an integrated DNA methylation, transcriptome, and proteome analysis can provide important resources for epigenetic studies in SM-resistant M. tuberculosis H37Rv.

Guangdong Province, China’s largest economy, has a high incidence of tuberculosis (TB). At present, there are few reports on the distribution, transmission and drug resistance of Mycobacterium tuberculosis (Mtb) strains in this region. In this study, we performed minimum inhibitory concentration testing for 14 anti-TB drugs and whole-genome sequencing of 713 clinical Mtb isolates from 20,662 sputum culture-positive tuberculosis patients registered at 31 tuberculosis drug resistance surveillance sites covering 20 cities in Guangdong Province from 2016 to 2018. Moreover, we evaluated genome-wide associations between mutations and drug resistance, and further investigated the differences in the MICs of mutations. The epidemiology, drug-resistant phenotypes and whole genome sequencing data of 713 clinical Mtb isolates were analyzed, revealing the lineage distribution and drug-resistant gene profiles in Guangdong Province. WGS combined with quantitative MIC measurements identified several novel loci associated with resistance, of which 16 loci were found to be related to resistance to more than one drug. This study analyzed the lineage distribution, prevalence characteristics and resistance-corresponding gene profiles of Mtb isolates in Guangdong province, and provided a theoretical basis for the formulation of tuberculosis prevention and control policy in the province.

目的:通过对实验室诱导产生的对氨基水杨酸(para-aminosalicylic acid,PAS)耐药结核分枝杆菌(Mycobacterium tuberculosis,MTB)和临床分离株进行全基因组测序,探索PAS的潜在作用机制和耐药性.方法:通过体外药物浓度梯度诱导法,将MTB标准菌株H37Rv诱导成标准耐PAS菌株及高水平耐药菌株,收集并保存每一代诱导菌株.用液体微孔板法检测上述诱导菌株的最低抑菌浓度(minimum inhibitory concentration,MIC)及交叉耐药情况,筛选耐药水平相似的临床分离株.通过对PAS耐药的实验室突变株和临床分离株进行全基因组测序,从基因水平上探索MTB对PAS的耐药机制.结果:本研究在体外建立了耐PAS的MTB耐药动态变化模型.全基因组测序结果显示,PAS耐药可能与3个位点突变直接相关,分别是plcC(Q462R)、folC(S150R)和1个thyA上游位点(3074495G→A).MTB对高水平的PAS耐药的产生除了基因突变因素以外,尚存在其他的调控机制.对PAS敏感和耐药的MTB临床分离株测序发现某些folC、thyA和thyX突变并非由PAS引起.结论:本研究构建的PAS耐药MTB菌株模型,为PAS耐药结核病研究和临床实践提供了理想的生物模型.全基因组测序分析在DNA水平上阐明了 MTB对PAS的耐药机制,尚存在其他的调控机制,仍需进一步研究.所得的与PAS耐药相关的基因突变也需要进一步的实验验证.

Antibiotic resistance of Mycobacterium tuberculosis (Mtb) is a major public health concern worldwide. Therefore, it is of great significance to characterize the mutational pathways by which susceptible Mtb evolves into drug resistance. In this study, we used laboratory evolution to explore the mutational pathways of aminoglycoside resistance. The level of resistance in amikacin inducing Mtb was also associated with changes in susceptibility to other anti-tuberculosis drugs such as isoniazid, levofloxacin and capreomycin. Whole-genome sequencing (WGS) revealed that the induced resistant Mtb strains had accumulated diverse mutations. We found that rrs A1401G was the predominant mutation in aminoglycoside-resistant clinical Mtb isolates from Guangdong. In addition, this study provided global insight into the characteristics of the transcriptome in four representative induced strains and revealed that rrs mutated and unmutated aminoglycoside-resistant Mtb strains have different transcriptional profiles. WGS analysis and transcriptional profiling of Mtb strains during evolution revealed that Mtb strains harbouring rrs A1401G have an evolutionary advantage over other drug-resistant strains under the pressure of aminoglycosides because of their ultra-high resistance level and low physiological impact on the strain. The results of this study should advance our understanding of aminoglycoside resistance mechanisms.

目的 比较核酸适配体荧光探针法和液基夹层杯法检测结核分枝杆菌(MTB)的临床应用价值.方法 采集175例肺结核患者和80名健康体检者的痰液样本,分别采用液体培养法、固体培养法、抗酸染色镜检法、液基夹层杯法、核酸适配体荧光探针法检测MTB.以液体培养法结果为标准,评价核酸适配体荧光探针法、液基夹层杯法与液体培养法结果的一致性.结果 与液体培养法结果比较,液基夹层杯法的敏感性为78.08％、特异性为98.04％;核酸适配体荧光探针法的敏感性为54.79％、特异性为50.98％.结论 液基夹层杯法与液体培养法一致性较好,具有较好的临床诊断价值;核酸适配体荧光探针法性能尚需优化.

目的 分析结核病患者诊断延误情况及其影响因素,为区域性结核病控制措施的持续改进提供科学依据.方法 从"中国疾病预防控制信息系统"子系统"结核病管理信息系统"中导出广东省英德市2010-2020年新发肺结核患者网报资料,回顾性分析该地区11年的肺结核患者诊断延误(从症状发生到肺结核确诊时间>28d者)总体情况、年间变化趋势以及性别、年龄、居住地域及就诊医疗机构等因素的可能影响.结果 (1)英德市11年间共报告新发肺结核病11973例,其中3189例患者(26.63%)存在诊断延误;以第29天开始计算诊断延误时间,其诊断延误时间中位数为17 d.(2)11年间诊断延误率除2010、2011和2020年高达约30%外,其他年份均在22.35%～26.65%的较窄范围内波动;2010-2014年诊断延误时间中位数呈逐年上升趋势,2014年达33d峰值,其后的6年呈现总体下降趋势,2018年为9d最低值.(4)≥60岁患者组诊断延误率为22.61%,低于其他年龄组(P = 0.000);慢性病防治院就诊患者诊断延误率为62.89%,远高于其他就诊医疗单位(P = 0.000);市区、郊区、农村等不同居住地域患者诊断延误率依次升高,且差异显著(P = 0.000).男女性别间则差异无统计学意义(P = 0.462).(4)不同分类组别的诊断延误时间中位数分别为:男、女患者17、16 d;<18岁、18～44岁、45～59岁和≥60岁各年龄组患者10、13、18、20 d;郊区、农村、市区患者19、17、16 d;综合医疗机构、镇卫生院、监狱医院、慢性病防治院报告患者20、15、12、8 d.结论 近年来该地区结核病患者诊断延误总体情况不断向好,但患者年龄、居住地域、就诊医疗机构为诊断延误重要影响因素,必须相宜施策.

目的 探讨2014-2020年广东省老年人结核病(≥60岁)分离株的耐药情况和不同来源的结核分枝杆菌(mycobacterium tuberculosis,MTB)耐药特征.方法 收集2014年1月至2020年12月从32个广东省结核病耐药监测哨点分离的老年人MTB菌株.按照比例法对一线和二线抗结核药物[包括异烟肼(isoniazid,INH)、乙胺丁醇(ethambutol,EMB)、利福平(rifampicin,RFP)、链霉素(streptomycin,Sm)和卡那霉素(kanamycin,Km)、卷曲霉素(capreomycin,Cm)、丙硫异烟胺(protionamide,Pto)、氧氟沙星(ofloxacin,Ofx)、对氨基水杨酸钠(sodium p-aminosalicylate,PAS)]进行药物敏感性试验,对MTB的耐药情况进行分析.结果 2014-2020年共分离9590株老年人MTB菌株,占总MTB检出数的24.6%(9590/39 046),年度间的MTB检出率存在显著差异(P趋势＜0.001),MTB的耐药率、单耐药率、多耐药率、耐多药率和广泛耐药率分别为26.7%、13.0%、5.0%、5.0%和2.1%,单耐药率、耐多药率和广泛耐药率年度间的分布存在差异性(均P＜0.05).老年女性结核病耐多药率和广泛耐药率显著高于男性(均P＜0.05),珠三角地区老年人结核分枝杆菌多耐药率、耐多药率和广泛耐药率均显著高于粤东西北地区(均P＜0.05).MTB对一线和二线抗结核药物的耐药率从高到低依次为Sm(14.9%)＞INH(11.6%)＞EMB(7.5%)＞RFP(7.1%)＞Ofx(5.8%)＞PAS(5.0%)＞Pto(4.6%)＞Cm(4.0%)＞Km(3.7%),除EMB和Pto外,上述9种一线和二线抗结核药物的耐药率年度趋势间差异有统计学意义(均P趋势＜0.05).复治患者MTB的广泛耐药率显著高于初治患者(P=0.006).结论 广东省老年人结核病的耐药率较高,珠三角地区居住、老年女性及复治结核病患者MTB耐药情况应引起临床工作者的重视.

目的:分析2014-2020年广东省儿童肺结核患者结核分枝杆菌(Mycobacerium tuberculosis,MTB)分离株耐药特征.方法:从广东省32个肺结核耐药监测点收集2014年1月1日至2020年12月30日39 046例肺结核患者耐药监测资料,依据入组标准从中选取1274例(3.3％)18岁及以下儿童患者耐药监测资料,分析MTB分离株对9种一线和二线抗结核药物[包括异烟肼(INH)、链霉素(Sm)、乙胺丁醇(EMB)、利福平(RFP)、卡那霉素(Km)、丙硫异烟胺(Pto)、卷曲霉素(Cm)、氧氟沙星(Ofx)和对氨基水杨酸钠(PAS)]的药物敏感性试验结果,并总结其耐药特征.结果:儿童肺结核患者构成比从2014年的4.3％(123/2856)下降至2020年的3.1％(177/5760),呈逐年下降趋势(x2趋势=25.864,P＜0.001).药物敏感性试验结果显示,1274株菌株对Sm、INH、RFP、EMB、Ofx、PAS、Pto、Km 和 Cm 的耐药率依次为 14.8％(189 株)、7.7％(98 株)、6.3％(80 株)、6.0％(77 株)、3.5％(44 株)、3.5％(44株)、2.9％(37株)、2.5％(32株)和2.2％(28株),总耐药率、单耐药率、多耐药率、耐多药率和广泛耐药率分别为26.0％(331株)、15.4％(196株)、4.2％(54株)、3.1％(39株)和0.8％(10株).其中,0～12岁年龄组和男性儿童组的MTB分离株广泛耐药率[3.0％(3/100)和1.3％(10/749)]分别明显高于13～18岁年龄组和女性儿童组[0.6％(7/1174)和0.0％(0/525)],珠江三角洲地区儿童组的MTB分离株单耐药率[28.1％(213/758)]明显高于粤东西北地区[22.9％(118/516)],差异均有统计学意义(x2=6.837,P=0.038;x2=10.679,P=0.007;x2=4.370,P=0.037).结论:广东省儿童耐药肺结核流行形势严峻,应对珠江三角洲地区和0～12岁男性儿童肺结核的耐药情况予以足够的重视.

Whole-genome sequencing (WGS) for prediction of drug resistance in Mycobacterium tuberculosis clinical isolates might contribute to the timely diagnosis and correct treatment of all tuberculosis cases. In 2021, the first catalogue of resistance-associated mutations in clinical M tuberculosis complex isolates was published by WHO,1 which provided a global standard for resistance interpretation. We read with interest the Article by Finci and colleagues2 reporting a good correlation between resistance genotype with phenotypic antimicrobial susceptibility testing (AST) using a large, longitudinal survey of M tuberculosis isolates from five countries.

目的 探索结核分枝杆菌感染及常规抗结核化疗过程中肠道菌群的变化差异,为结核病的临床治疗提供科学依据.方法 提取89份粪便样本的基因组DNA,利用宏基因组测序技术及生物信息学方法分析结核分枝杆菌感染及常规抗结核化疗过程中肠道微生物菌群的变化.结果 结核分枝杆菌感染后,结核病患者的肠道菌群结构组成发生显著改变(P=0.0134).随着常规抗结核化疗时间延长,Shannon指数下降,结核病患者肠道菌群多样性明显降低(P=0.0088),结核病患者治愈后菌群结构组成较治疗前出现显著变化(P=0.0021).结核分枝杆菌感染及常规抗结核化疗过程中,拟杆菌属和韦永氏球菌属的相对丰度逐渐增加.经抗结核化疗后结核病患者肠道内微生物群中部分功能途径发生变化.结论 结核分枝杆菌感染及常规抗结核化疗可以导致肠道菌群紊乱,对肠道菌群的多样性、结构组成和代谢功能等产生影响.

目的 系统梳理2010-2020年国内外结核病负担的研究文献,并挖掘研究热点.方法 检索中国知网(CNKI)和Web of Science(WOS)数据库2010-2020年国内外结核病负担的研究文献,利用CiteSpace5.7 R2软件进行可视化分析.结果 CNKI纳入45篇文献,频次最高的研究者是白丽琼和肖水源;热点关键词是患病代价、结核、疾病经济负担等.WOS纳入96篇文献,发文最多的国家是美国;频次最高的研究者是Garcia-Basteiro,.和Ogbo;热点关键词是impact、service、HIV等.结论 结核病负担的研究处于上升趋势,研究热点和前沿集中在经济负担.

目的:了解2016-2020年广东省结核病患者耐药情况及趋势,为全省耐药结核病的防控提供科学依据.方法:收集2016年1月1日至2020年12月31日广东省32个耐药监测点所有活动性肺结核患者作为研究对象,共纳入92 851例.收集研究对象临床信息,对研究对象的痰标本进行涂片和分离培养,培阳菌株进行菌群鉴定,共获得结核分枝杆菌(MTB)分离株30 362株.采用比例法对MTB分离株开展9种抗结核药品[异烟肼(INH)、利福平(RFP)、乙胺丁醇(EMB)、链霉素(Sm)、卡那霉素(Km)、氧氟沙星(Ofx)、卷曲霉素(Cm)、丙硫异烟胺(Pto)、对氨基水杨酸钠(PAS)]的药物敏感性试验(简称"药敏试验"),并分析菌株的耐药情况.结果:30 362株菌株的总耐药率为 26.75％(8121/30 362),耐多药率为 3.34％(1014/30 362),单耐药率为 15.88％(4820/30 362),多耐药率为7.53％(2286/30 362).菌株对9种抗结核药品的任一耐药率由高至低依次为Sm(14.59％,4430/30 362)＞INH(9.25％,2810/30 362)＞RFP(6.22％,1887/30 362)＞EMB(5.72％,1737/30 362)＞Ofx(3.03％,920/30 362)＞Pto(3.00％,912/30 362)＞PAS(2.25％,684/30 362)＞Cm(2.00％,606/30 362)＞Km(1.63％,494/30 362),差异有统计学意义(x2=8889.800,P＜0.01).菌株中27 783株分离自初治患者,2579株分离自复治患者,复治患者的耐药率[35.32％(911/2579)]和耐多药率[10.35％(267/2579)]均明显高于初治患者[耐药率为25.95％(7210/27 783),耐多药率为2.69％(747/27 783)],差异均有统计学意义(x2=105.800,P＜0.01;x2=265.700,P＜0.01).5年间初治患者的耐多药率呈上升趋势[2016年为2.76％(152/5502),2020年为3.19％(166/5203)],复治患者的耐药率呈下降趋势[2016 年为 38.85％(169/435),2020 年为 35.02％(173/494)],差异均有统计学意义(x2趋势值分别为6.584和6.334,P值分别为0.010和0.012).珠三角地区来源菌株的耐药率和耐多药率[分别为27.35％(4540/16 597)和3.59％(596/16 597)]均高于粤东西北地区来源菌株[耐药率为26.02％(3581/13 765);耐多药率为3.04％(418/13 765)],差异均有统计学意义(x2值分别为6.886和3.879,P值分别为0.009和0.049).结论:广东省耐药结核病流行形势依然严峻,利福平耐药、初治患者耐药及流动人口结核病耐药等问题需重点关注.

目的:研究广东省耐多药结核病(multidrug-resistant tuberculosis,MDR-TB)患者灾难性支出发生率现状及其影响因素,为降低MDR-TB患者经济负担提供基础数据和参考依据.方法:以广东省2018-2019年9个地级市11家定点医疗机构和确诊的MDR-TB患者为研究对象,通过问卷收集医疗机构、MDR-TB患者和地市的基本情况,以及患病就诊情况、经济负担和医疗保障政策,分析影响广东省MDR-TB患者发生灾难性支出的因素.结果:研究共纳入广东省MDR-TB患者398例,直接医疗费用为5.11(2.26,9.82)万元,直接非医疗费用为1.46(0.71,3.33)万元,间接费用为0.51(0.31,0.73)万元,总费用为8.43(4.29,14.76)万元,人均经济负担为5.84(2.97,11.18)万元.259例患者发生了灾难性支出,发生率为65.08％.两水平logistic回归模型显示,有债务患者家庭发生灾难性支出的风险是无债务患者的2.57倍(95％CI:1.43～4.62);随着住院时间的增加,患者家庭灾难性支出发生风险逐渐上升,住院1个月的患者发生灾难性支出的风险是未住院患者的4.38倍(95％CI:2.38～8.06),住院≥2个月的患者发生灾难性支出的风险是未住院患者的31.37倍(95％CI:11.09～88.74).结论:广东省MDR-TB患者灾难性支出发生率较高,有债务、住院时间长是导致患者灾难性支出发生风险升高的因素,应多措并举降低MDR-TB患者灾难性支出的发生率.

Objective:To explore the main influencing factors of tuberculosis students analyzing visit delay and diagnosis delay of students with pulmonary tuberculosis.Methods:The relevant information including symptom onset times, initial visit times, and disease diagnosis times of the hospitalized students with pulmonary tuberculosis from 2019 to 2021 was obtained, which were abstracted from the electronic medical records of the "Hospital Statistical Medical Record System" searching the terms "discharge time", "occupation", and "international classification code of main diagnoses of diseases". The intervals between symptom onset time and initial visit time and between the interval between initial visit time and disease diagnosis time were the visit time and the diagnosis time, respectively; the times ≥ 15 days were defined as visit delay and diagnosis delay accordingly. The two kinds of delays of different groups were described, inferred, and analyzed statistically; they were grouped according to gender (male / female), student levels (primary school student / junior high schools tudent / high school student / undergraduate / postgraduate), case finding methods (seeking medical attention in tuberculosis control agency / referral from general hospital / recommendation by medical staff of primary health care institutions / physical examination), and symptom onset month (from January to February / from March to June / from July to August / from September to December). The χ2, Mann-Whitney U, and Kruskal-Wallis rank sum tests were applied. Results:There were 723 hospitalized students with pulmonary tuberculosis during the three years; their visit time was 0-92 days, and 406 cases (56.16%) had visit delay of 15-92 days (median 25 days); their diagnosis time was 1-95 days ,and 503 cases (69.57%) had diagnosis delay of 15-95 days (median 24 days). There was a statistical difference in the incidence of visit delay between the groups of case finding method ( χ2=33.288, P?0.001); there were statistical differences in the incidence of diagnosis delay between the groups of gender and symptom onset month ( χ2=10.029, P=0.002; χ2=18.187, P?0.001) by χ2 test. There were statistical differences in visit delay time betweeb the groups of student level and symptom onset month ( χ2=33.000, P?0.001; χ2=12.750, P=0.005) and was in diagnosis delay time between the groups of symptom onset month ( χ2=15.987, P=0.001) by Mann- Whitney U test (comparison between two groups) or Kruskal-Wallis rank-sum test (comparison between ≥3 groups). Conclusions:There are high incidences of visit delay and diagnosis delay in students with pulmonary tuberculosis. And the degree of delay is closely related to gender, student level, case finding methods, and symptom onset month.

Currently, there are no particularly effective biomarkers to distinguish between latent tuberculosis infection (LTBI) and active pulmonary tuberculosis (PTB) and evaluate the outcome of TB treatment. In this study, we have characterized the changes in the serum metabolic profiles caused by Mycobacterium tuberculosis (Mtb) infection and standard anti-TB treatment with isoniazid-rifampin-pyrazinamide-ethambutol (HRZE) using GC-MS and LC-MS/MS. Seven metabolites, including 3-oxopalmitic acid, akeboside ste, sulfolithocholic acid, 2-decylfuran (4,8,8-trimethyldecahydro-1,4-methanoazulen-9-yl)methanol, d-(+)-camphor, and 2-methylaminoadenosine, were identified to have significantly higher levels in LTBI and untreated PTB patients (T0) than those in uninfected healthy controls (Un). Among them, akeboside Ste and sulfolithocholic acid were significantly decreased in PTB patients with 2-month HRZE (T2) and cured PTB patients with 2-month HRZE followed by 4-month isoniazid-rifampin (HR) (T6). Receiver operator characteristic curve analysis revealed that the combined diagnostic model showed excellent performance for distinguishing LT from T0 and Un. By analyzing the biochemical and disease-related pathways, we observed that the differential metabolites in the serum of LTBI or TB patients, compared to healthy controls, were mainly involved in glutathione metabolism, ascorbate and aldarate metabolism, and porphyrin and chlorophyll metabolism. The metabolites with significant differences between the T0 group and the T6 group were mainly enriched in niacin and nicotinamide metabolism. Our study provided more detailed experimental data for developing laboratory standards for evaluating LTBI and cured PTB.

Ethambutol (EMB) is a first-line antituberculosis drug currently being used clinically to treat tuberculosis. Mutations in the embCAB operon are responsible for EMB resistance. However, the discrepancies between genotypic and phenotypic EMB resistance have attracted much attention. We induced EMB resistance in Mycobacterium tuberculosis in vitro and used an integrated genome-methylome-transcriptome-proteome approach to study the microevolutionary mechanism of EMB resistance. We identified 509 aberrantly methylated genes (313 hypermethylated genes and 196 hypomethylated genes). Moreover, some hypermethylated and hypomethylated genes were identified using RNA-seq profiling. Correlation analysis revealed that the differential methylation of genes was negatively correlated with transcription levels in EMB-resistant strains. Additionally, two hypermethylated candidate genes (mbtD and celA1) were screened by iTRAQ-based quantitative proteomics analysis, verified by qPCR, and corresponded with DNA methylation differences. This is the first report that identifies EMB resistance-related genes in laboratory-induced mono-EMB-resistant M. tuberculosis using multi-omics profiling. Understanding the epigenetic features associated with EMB resistance may provide new insights into the underlying molecular mechanisms.

目的 探索结核菌素试验(PPD试验)在学校结核病疫情处置中对于界定密切接触者筛查范围的作用,为学校肺结核防治工作提供参考依据.方法 回顾性分析比较2019年8月广东省某中学发生肺结核疫情的高二年级与同期该校高一入学新生体检PPD试验结果,共1553名学生,并评估高二年级学生PPD试验流行病学分布与肺结核病例分布之间的潜在关联.结果 高二年级结核病疫情共确诊学生患者29例,其中7例病原学阳性,且菌株MIRU-VNTR基因分型相同.高一年级学生人群中PPD强阳性率为0.1％,PPD阳性率为34.3％;高二年级学生人群PPD强阳性率、阳性率分别为5.9％,52.0％,高二年级均高于高一年级,差异均有统计学意义(x2值分别为45.71,49.90,P值均＜0.01).在教学楼分布方面,疫情发源、病例集中的教学楼一层学生PPD强阳性率为19.4％,远高于教学楼其他楼层,PPD强阳性率逐楼层持续降低,差异有统计学意义(x2=89.81,P＜0.01);各楼层的PPD强阳人数与学生病例数量呈正相关(r=0.99,P＜0.01).在宿舍楼层分布方面,病例所在的宿舍楼层学生人群PPD强阳性率、阳性率明显大于无病例的楼层,差异有统计学意义(x2值分别为49.4,64.9,P值均＜0.01);各宿舍楼层PPD强阳人数、阳性人数与各层肺结核病例数之间呈正相关(r值分别为0.84,0.56,P值均＜0.05).结论 PPD强阳性率可以良好地反映学校肺结核密切接触人群结核杆菌感染和疫情扩散的情况,建议作为考量扩大密切接触者筛查范围的因素之一.