RNA Sequencing and Bioinformatics Analysis to Reveal Potential Biomarkers in Patients with Combined Allergic Rhinitis and Asthma Syndrome.

Introduction: Combined allergic rhinitis and asthma syndrome (CARAS) is a concurrent clinical or subclinical allergic symptom of diseases of the upper and lower respiratory tract. This study is the first to explore the expression profiles of mRNA, lncRNA, and circRNA in CARAS using RNA sequencing, which may provide insight into the mechanisms underlying CARAS. Material and Methods: Whole blood samples from nine participants (three CARAS patients, three AR patients, and three normal control participants) were subjected to perform RNA sequencing, followed by identification of differentially expressed lncRNAs (DElncRNAs), circRNAs (DEcircRNAs) and mRNAs (DEmRNAs). Then, lncRNA/circRNA-mRNA regulatory pairs were constructed, followed by functional analysis, immune infiltration analysis, drug prediction, and expression validation with RT-qPCR and ELISA. Results: The results showed that 61 DEmRNAs, 23 DElncRNAs and 3 DEcircRNAs may be related to the occurrence and development of CARAS. KRT8 may be implicated in the development of AR into CARAS. Three immunity-related mRNAs (IDO1, CYSLTR2, and TEC) and two hypoxia-related mRNAs (TKTL1 and VLDLR) were associated with the occurrence and development of CARAS. TEC may be considered a drug target for Dasatinib in treating CARAS. Several lncRNA/circRNA-mRNA regulatory pairs were identified in CARAS, including LINC00452/MIR4280HG/hsa_circ_0007272/hsa_circ_0070934-CLC, HEATR6-DT/LINC00639/LINC01783/hsa_circ_0008903-TEC, RP11-71L14.3-IDO1/SMPD3, RP11-178F10.2-IDO1/HRH4, and hsa_circ_0008903-CYSLTR2, which may indicate potential regulatory effects of lncRNAs/circRNAs in CARAS. Dysregulated levels of immune cell infiltration may be closely related to CARAS. Conclusion: The regulating effect of lncRNA/circRNA-immunity/hypoxia-related mRNA regulatory pairs may be involved in the occurrence and development of CARAS.

rpoB Mutations are Associated with Variable Levels of Rifampin and Rifabutin Resistance in Mycobacterium tuberculosis.

Objective: To assess the relationship between the variant rpoB mutations and the degree of rifampin (RIF)/rifabutin (RFB) resistance in Mycobacterium tuberculosis (M. tuberculosis). Methods: We analyzed the whole rpoB gene in 177 M. tuberculosis clinical isolates and quantified their minimum inhibitory concentrations (MICs) using microplate-based assays. Results: The results revealed that of the 177 isolates, 116 were resistant to both RIF and RFB. There were 38 mutated patterns within the sequenced whole rpoB gene of the 120 isolates. Statistical analysis indicated that mutations, S450L, H445D, H445Y, and H445R, were associated with RIF and RFB resistance. Of these mutations, S450L, H445D, and H445Y were associated with high-level RIF and RFB MIC. H445R was associated with high-level RIF MIC, but not high-level RFB MIC. D435V and L452P were associated with only RIF, but not RFB resistance. Q432K and Q432L were associated with high-level RFB MIC. Several single mutations without statistical association with rifamycin resistance, such as V170F, occurred exclusively in low-level RIF but high-level RFB resistant isolates. Additionally, although cross-resistance to RIF and RFB is common, 21 RIF-resistant/RFB-susceptible isolates were identified. Conclusion: This study highlighted the complexity of rifamycin resistance. Identification of the rpoB polymorphism will be helpful to diagnose the RIF-resistant tuberculosis that has the potential to benefit from a treatment regimen including RFB.

rpoB Mutations and Effects on Rifampin Resistance in Mycobacterium tuberculosis.

OBJECTIVE: To investigate the mutations within the whole rpoB gene of Mycobacterium tuberculosis and analyze their effects on rifampin (RIF) resistance based on crystal structure. METHODS: We sequenced the entire rpoB gene in 175 tuberculosis isolates and quantified their minimum inhibitory concentrations using microplate-based assays. Additionally, the structural interactions between wild-type/mutant RpoB and RIF were also analyzed. RESULTS: Results revealed that a total of 34 mutations distributed across 17 different sites within the whole rpoB gene were identified. Of the 34 mutations, 25 could alter the structural interaction between RpoB and RIF and contribute to RIF resistance. Statistical analysis showed that S450L, H445D, H445Y and H445R mutations were associated with high-level RIF resistance, while D435V was associated with moderate-level RIF resistance. CONCLUSION: Some mutations within the rpoB gene could affect the interaction between RpoB and RIF and thus are associated with RIF resistance. These findings could be helpful to design new antibiotics and develop novel diagnostic tools for drug resistance in TB.

Clinical metagenomic sequencing for rapid diagnosis of pneumonia and meningitis caused by Chlamydia psittaci.

BACKGROUND: Chlamydia psittaci (C. psittaci) is a gram-negative intracellular parasitic pathogenic bacterium that can infect avian and mammalian hosts, including humans. The detection of C. psittaci infections typically relies on traditional antigen-based immunoassays or serological testing that often lack sensitivity and/or specificity. Metagenomic next generation sequencing (mNGS) is an emerging tool for diagnosis. AIM: To demonstrate that mNGS represents a valuable tool for rapid, sensitive, and accurate pathogen detection including C. psittaci infections. METHODS: Four cases of psittacosis pneumonia and one case of pediatric psittacosis meningitis were diagnosed between December 2019 and May 2020 using mNGS at Changzhou Second People's Hospital affiliated to Nanjing Medical University. Patients' clinical characteristics, manifestations, and treatment histories were retrospectively evaluated. RESULTS: All five patients had a history of exposure to wild (psittacine or other birds) or domesticated birds (chickens). All patients had a high fever (> 39℃) and three of them (60%) experienced organ insufficiency during the disease. The laboratory data showed normal to slightly increased leucocyte and neutrophil counts, and elevated procalcitonin levels in all five cases, and very high C-reactive protein levels in psittacosis pneumonia patients. mNGS identified a potential pathogen, C. psittaci, in patients' bronchoalveolar lavage fluid or cerebrospinal fluid. Computed tomography revealed lung air-space consolidation, pleural thickening, and effusion fluid buildup in psittacosis pneumonia cases, and an arachnoid cyst in the right temporal lobe of the pediatric psittacosis meningitis patient. All patients experienced complete recovery following the administration of targeted anti-chlamydia therapy. CONCLUSION: This study not only demonstrated that mNGS represents a valuable tool for rapid, sensitive, and accurate pathogen detection, but also raised public health concerns over C. psittaci infections.

Detecting Mycobacterium tuberculosis complex and rifampicin resistance via a new rapid multienzyme isothermal point mutation assay.

Simple, rapid, and accurate detection of the Mycobacterium tuberculosis complex (MTBC) and drug resistance is critical for improving patient care and decreasing the spread of tuberculosis. To this end, we have developed a new simple and rapid molecular method, which combines multienzyme isothermal rapid amplification and a lateral flow strip, to detect MTBC and simultaneously detect rifampin (RIF) resistance. Our findings showed that it has sufficient sensitivity and specificity for discriminating 118 MTBC strains from 51 non-tuberculosis mycobacteria strains and 11 of the most common respiratory tract bacteria. Further, compared to drug susceptibility testing, the assay has a sensitivity, specificity, and accuracy of 54.1%, 100.0%, and 75.2%, respectively, for detection of RIF resistance. Some of the advantages of this assay are that no special instrumentation is required, a constant low temperature of 39 °C is sufficient for the reaction, the turnaround time is less than 20 min from the start of the reaction to read out and the result can be seen with the naked eye and does not require specialized training. These characteristics of the new assay make it particularly useful for detecting MTBC and RIF resistance in resource-limited settings.

LncRNA NEAT1 activates MyD88/NF-κB pathway in bronchopneumonia through targeting miR-155-5p.

BACKGROUND: Bronchopneumonia is a disease of the respiratory tract. It leads to other complications and endangers life and health. Long non-coding RNA (lncRNA) participates in the occurrence and development of bronchopneumonia. Nuclear paraspeckle assembly transcript 1 (NEAT1) plays a key role in inflammatory diseases, but the function of NEAT1 in bronchopneumonia remains unclear. METHODS: RT-qPCR and Western blotting were performed to determine genes and proteins expressions. MTT was applied to test cell viability. Cell apoptosis was detected by flow cytometry. RIP was used to investigate the correlation between NEAT1 and miR-155-5p. The interaction between miR-155-5p and NEAT1 or MyD88 was evaluated by the dual-luciferase reporter gene. RESULTS: NEAT1 and MyD88 were upregulated in BEAS-2B cells by LPS, while miR-155-5p was downregulated. Knockdown of NEAT1 inhibited LPS-induced BEAS-2B cells growth inhibition by inhibiting the apoptosis. In addition, NEAT1 silencing suppressed LPS-induced inflammatory responses in BEAS-2B cells via suppression of TNF-α, IL-1ß, IL-6, and IL-18. Meanwhile, NEAT1 is directly bound to miR-155-5p to regulate MyD88/NF-κB axis, and overexpression of miR-155-5p increased cell proliferation and suppressed inflammatory factors expression levels and cell apoptosis. Furthermore, sh-NEAT1-induced inhibition of BEAS-2B cells injury was partially reversed by miR-155-5p inhibitor or MyD88 overexpression. CONCLUSION: NEAT1 silencing suppressed LPS-induced BEAS-2B cells injury and inflammation by the mediation of miR-155-5p/MyD88/NF-κB axis. Thus, our study might shed new light on exploring the new strategies for the treatment of bronchopneumonia.

Detecting Ethambutol Resistance in Mycobacterium tuberculosis Isolates in China: A Comparison Between Phenotypic Drug Susceptibility Testing Methods and DNA Sequencing of embAB.

With the increasing incidence of drug-resistant tuberculosis (DR-TB), determining a rapid and accurate drug susceptibility testing (DST) method to identify ethambutol (EMB) resistance in Mycobacterium tuberculosis has become essential for patient management in China. Herein, we evaluated the correlation between three phenotypic DST methods, namely, proportion method (PM), MGIT 960 system, and microplate alamar Blue assay (MABA), and DNA sequencing of embAB in 118 M. tuberculosis isolates from China. When the results of the phenotypic DST methods were compared with those of DNA sequencing, the overall agreement and kappa values of the PM, MGIT 960 system, and MABA were 81.4% and 0.61, 77.1% and 0.55, and 84.7% and 0.67, respectively. The agreement for EMB resistance between MABA and PM was significantly higher than that between the MGIT 960 system and PM (P = 0.02). Moreover, among the isolates with detectable embAB mutations, 97.2% (70/72 isolates) harbored mutations in embB. The analysis of embB mutations predicted EMB resistance with 81.3% sensitivity, 86.8% specificity, and 83.1% accuracy. Thus, MABA may be a better phenotypic DST method for detecting EMB resistance. DNA sequencing of embB may be useful for the early identification of EMB resistance and the consequent optimization of the treatment regimen.

Accuracy of a reverse dot blot hybridization assay for simultaneous detection of the resistance of four anti-tuberculosis drugs in Mycobacterium tuberculosis isolated from China.

BACKGROUND: Drug resistant tuberculosis poses a great challenge for tuberculosis control worldwide. Timely determination of drug resistance and effective individual treatment are essential for blocking the transmission of drug resistant Mycobacterium tuberculosis. We aimed to establish and evaluate the accuracy of a reverse dot blot hybridization (RDBH) assay to simultaneously detect the resistance of four anti-tuberculosis drugs in M. tuberculosis isolated in China. METHODS: In this study, we applied a RDBH assay to simultaneously detect the resistance of rifampicin (RIF), isoniazid (INH), streptomycin (SM) and ethambutol (EMB) in 320 clinical M. tuberculosis isolates and compared the results to that from phenotypic drug susceptibility testing (DST) and sequencing. The RDBH assay was designed to test up to 42 samples at a time. Pearson's chi-square test was used to compute the statistical measures of the RDBH assay using the phenotypic DST or sequencing as the gold standard method, and Kappa identity test was used to determine the consistency between the RDBH assay and the phenotypic DST or sequencing. RESULTS: The results showed that the concordances between phenotypic DST and RDBH assay were 95% for RIF, 92.8% for INH, 84.7% for SM, 77.2% for EMB and the concordances between sequencing and RDBH assay were 97.8% for RIF, 98.8% for INH, 99.1% for SM, 93.4% for EMB. Compared to the phenotypic DST results, the sensitivity and specificity of the RDBH assay for resistance detection were 92.4 and 98.5% for RIF, 90.3 and 97.3% for INH, 77.4 and 91.5% for SM, 61.4 and 85.7% for EMB, respectively; compared to sequencing, the sensitivity and specificity of the RDBH assay were 97.7 and 97.9% for RIF, 97.9 and 100.0% for INH, 97.8 and 100.0% for SM, 82.6 and 99.1% for EMB, respectively. The turnaround time of the RDBH assay was 7 h for testing 42 samples. CONCLUSIONS: Our data suggested that the RDBH assay could serve as a rapid and efficient method for testing the resistance of M. tuberculosis against RIF, INH, SM and EMB, enabling early administration of appropriate treatment regimens to the affected drug resistant tuberculosis patients.

[Effects of temperature and phosphorus application on wheat growth at seedling stage and phosphorus form in calcareous fluvo-aquic soil]. / 温度和施磷对石灰性潮土小麦苗期生长及磷形态的影响.

The effects of temperature and phosphate fertilizer application on wheat seedling growth and soil inorganic phosphorus transformation in calcareous fluvo-aquic soil were examined in a pot experiment. The results showed that temperature and phosphorus were important factors affecting wheat growth but without significant interaction. The effect of temperature on wheat growth was greater than that of phosphate fertilizer, and 15 ℃ was the suitable temperature for wheat seedling. Compared with the treatment without P application (-P) at 5 ℃, phosphate fertilizer treatment (+P) promoted the growth of wheat seedling. Shoot and root biomass increased by 18.2% and 33.3%, the accumulation of phosphorus in shoot and root were increased by 30.6% and 13.3%, and the root-shoot ratio, plant height, tillering number and root activity were increased by 3.5%, 10.0%, 10.5% and 70.3%, respectively. At 15 ℃, the effect of phosphorus fertilizer application did not affect wheat biomass and tiller, increased P accumulation in shoot and root of wheat by 32.3% and 23.8%, and increased the ratio of root to shoot, plant height and root activity by 15.6%, 2.5% and 32.8% respectively. There were no significant promoting effects on wheat growth between different phosphate applications at 25 ℃. At three temperatures, phosphate application significantly increased the contents of Olsen-P, Ca2-P, Ca8-P, Al-P and Fe-P. When treated with -P and +P, temperature had no significant effect on Ca2-P content, but had significant effect on the Olsen-P, Ca8-P, Al-P, Fe-P contents. The contents of Ca8-P and Fe-P were 5 ℃＞15 ℃＞25 ℃; Al-P content was 25 ℃＞15 ℃＞5 ℃. Wheat could absorb and utilize Ca2-P, Ca8-P, Al-P, Fe-P at seedling stage. The availability of Al-P, Fe-P to wheat was significantly lower than that of Ca2-P, Ca8-P. There was no significant difference of soil pH, O-P and Ca10-P across all treatments. In conclusion, temperature mainly affected the absorption of phosphorus by affecting wheat growth, and the application of phosphorus fertilizer at low temperature could significantly promote the growth of wheat. High temperature could accelerate the fixation of available phosphorus in calcareous soil, a process could be alleviated by phosphorus application.

Prevalence, risk and genetic characteristics of drug-resistant tuberculosis in a tertiary care tuberculosis hospital in China.

OBJECTIVES: To explore the prevalence, risk and genetic characteristics of drug-resistant tuberculosis (TB) from a tertiary care TB hospital in China. PATIENTS AND METHODS: We carried out a retrospective study including isolates from 189 patients with pulmonary TB at Fuzhou Pulmonary Hospital. All isolates from these patients were subjected to drug susceptibility testing and genotyping. For drug-resistant isolates, DNA sequencing was used to investigate mutations in 12 loci, including katG, inhA, oxyR-ahpC, rpoB, rpsL, rrs 1 (nucleotides 388-1084 of rrs), embB, tlyA, eis, rrs 2 (nucleotides 1158-1674 of rrs), gyrA and gyrB. RESULTS: Among 189 isolates, 28.6% were resistant to at least one of the seven anti-TB drugs, including isoniazid (INH), rifampin (RIF), streptomycin (STR), ethambutol (EMB), capreomycin (CAP), kanzmycin (KAN) and ofloxacin (OFX). The proportion of multidrug-resistant TB and extensively drug-resistant TB isolates was 9.5% and 1.1%, respectively. Patients in rural areas as well as previously treated patients showed a significantly increased risk of developing drug resistance. In addition, among these isolates, 111 (58.7%) were Beijing genotype strains, 84 (75.7%) of which belonged to modern Beijing sublineage. There was no association between genotype and drug resistance. The most common mutations were katG315, rpo B531 rpsL43, embB306, rrs1401 and gyrA94. CONCLUSION: These findings provided additional information of drug-resistant TB in China. Previously treated patients and patients in rural areas should receive greater attention owing to their higher risk of developing drug resistance.

Beijing genotype of Mycobacterium tuberculosis is less associated with drug resistance in south China.

Mycobacterium tuberculosis Beijing genotype strains are widespread globally. However, there has been no systematic study on the association between Beijing genotype and the characteristics of drug resistance. In this study, 359 M. tuberculosis isolates from south China were collected and their background information, genotype diversity and drug resistance was investigated. The results revealed that 66.0% of strains (237/359) were categorised as Beijing genotype. There was no statistical difference between Beijing and non-Beijing genotype strains in terms of patient sex, age, place of residence and treatment history. Drug resistance testing showed that 34.8% (125/359) of isolates were resistant to at least one of the seven drugs tested. The proportions of multidrug-resistant tuberculosis and extensively drug-resistant tuberculosis were 17.0% and 1.4%, respectively. Previously treated patients presented a significantly higher risk of developing drug resistance than new cases. Although the prevalence of drug resistance was higher in Beijing genotype than in non-Beijing genotype strains, there was no significant difference between these two genotypes in the multivariate analysis. Even in re-treated patients, the association of Beijing genotype with drug resistance was not significant. This study provides an insight into genotype diversity and demonstrates the characteristics of drug resistance in Beijing genotype strains, which will be useful in generating efficient tuberculosis prevention and control strategies in China.

Management of Persistent Air Leaks Using Endobronchial Autologous Blood Patch and Spigot Occlusion: A Multicentre Randomized Controlled Trial in China.

BACKGROUND: Optimal management of persistent air leaks (PALs) in patients with secondary spontaneous pneumothorax (SSP) remains controversial. OBJECTIVE: To evaluate the efficacy and safety of endobronchial autologous blood plus thrombin patch (ABP) and bronchial occlusion using silicone spigots (BOS) in patients with SSP accompanied by alveolar-pleural fistula (APF) and PALs. METHODS: This prospective multicentre randomized controlled trial compared chest tube-attached water-seal drainage (CTD), ABP, and BOS that were performed between February 2015 and June 2017 in one of six tertiary care hospitals in China. Patients diagnosed with APF experiencing PALs (despite 7 days of CTD) and inoperable patients were included. Outcome measures included success rate of pneumothorax resolution at the end of the observation period (further 14 days), duration of air leak stop, lung expansion, hospital stay, and complications. RESULTS: In total, 150 subjects were analysed in three groups (CTD, ABP, BOS) of 50 each. At 14 days, 60, 82, and 84% of CTD, ABP, and BOS subjects, respectively, experienced full resolution of pneumothorax (p = 0.008). All duration outcome measures were significantly better in the ABP and BOS groups than in the CTD group (p < 0.016 for all). The incidence of adverse events, including chest pain, cough, and fever, was not significantly different. All subjects in the ABP and BOS groups experienced temporary haemoptysis. Spigot displacement occurred in 8% of BOS subjects. CONCLUSION: ABP and BOS resulted in clinically meaningful outcomes, including higher success rate, duration of air leak stop, lung expansion, and hospital stay, with an acceptable safety profile.

Comprehensive analysis of miRNA-mRNA-lncRNA networks in severe asthma.

AIM: This study aimed to explore the molecular mechanism of severe asthma. MATERIALS & METHODS: The shared and divergent differentially expressed mRNAs (DEmRNAs), miRNAs (DEmiRNAs) and lncRNAs (DElncRNAs) in asthma and severe asthma were identified by RNA-sequencing. Severe asthma-specific and shared DEmiRNA-DEmRNA-DElncRNA interaction networks were performed. RESULTS: Compared with normal control, 1328 DEmRNAs, 608 DElncRNAs and 63 DEmiRNAs were identified in severe asthma. Compared with asthma, 95 DEmRNAs, 143 DElncRNAs and 96 DEmiRNAs were identified in severe asthma. MiR-133a-3p-EFHD2/CNN2-AC144831.1 interactions and miR-3613-3p-CD44/BCL11B-LINC00158/CTA-217C2.1/AC010976.2/RP11-641A6.2 interactions were speculated to involve with the development of severe asthma. The results of GSE69683 validation were generally consistent with our RNA-sequencing results. CONCLUSION: This study provides clues for understanding the mechanism of severe asthma.

Comprehensive Analysis of miRNA-mRNA-lncRNA Networks in Non-Smoking and Smoking Patients with Chronic Obstructive Pulmonary Disease.

BACKGROUND/AIMS: Chronic obstructive pulmonary disease (COPD) is a major cause of morbidity and mortality worldwide. This study aimed to identify overlapping or diverging dysregulated genes, lncRNAs, miRNAs and signaling pathways in smoking and non-smoking chronic obstructive pulmonary disease (COPD). METHODS: Compared to normal controls, we identified the shared and divergent differentially expressed mRNAs (DEmRNAs), miRNAs (DEmiRNAs) and lncRNAs (DElncRNAs) in smoking and non-smoking COPD by RNA-sequencing and bioinformatics analysis. Functional annotation of DEmRNAs were performed. Both cis and trans-target DEmRNAs of DElncRNAs were identified. The target DEmRNAs of DEmiRNAs were identified as well. The DEmiRNA-DEmRNA-DElncRNA interaction network was constructed. QRT-PCR was performed to validat the selected DEmiRNAs, DEmRNA and DElncRNAs in COPD. RESULTS: Compared to normal control, 1234 DEmRNAs, 96 DElncRNAs and 151 DEmiRNAs were identified in non-smoking patients with COPD; 670 DEmRNAs, 44 DElncRNAs and 63 DEmiRNAs were identified in smoking patients with COPD. Leukocyte transendothelial migration and pathways in cancer were significantly enriched pathways in non-smoking and smoking COPD, respectively. MiR-122-5p-A2M-LINC00987/A2M-AS1/ linc0061 interactions might play key roles in COPD irrespective with the smoking status. Let-7-ADRB1-HLA-DQB1-AS1 might play a key role in the pathogenesis of smoking COPD while miR-218-5p/miR15a-RORA-LOC101928100/LINC00861 and miR-218-5p/miR15a-TGFß3-RORA-AS1 interactions might involve with non-smoking COPD. CONCLUSION: We identified the shared and diverging genes, lncRNAs, miRNAs and their interactions and pathways in smoking and non-smoking COPD which provided clues for understanding the mechanism and developing novel diagnostic and therapeutic strategies for COPD.

Mutations within embCAB Are Associated with Variable Level of Ethambutol Resistance in Mycobacterium tuberculosis Isolates from China.

The EmbCAB proteins have been considered a target for ethambutol (EMB). Mutations in embCAB are known to confer most EMB resistance. However, the knowledge about the effects of embCAB mutations on the EMB resistance level and about the role of mutation-mutation interactions is limited in China. Here, we sequenced embCAB among 125 Mycobacterium tuberculosis isolates from China and quantified their EMB MICs by testing growth at 10 concentrations. Furthermore, a multivariate regression model was established to assess the effects of both individual mutations and multiple mutations. Our results revealed that in China, 82.6% of EMB-resistant isolates (71/86 isolates) harbored at least one mutation within embCAB Most of the mutations were located in the embB and embA upstream region. Several individual mutations and multiple mutations within this region contributed to the different levels of EMB resistance. Their effects were statistically significant. Additionally, there was an association between high-level EMB resistance and multiple mutations.

Evolutionary History and Ongoing Transmission of Phylogenetic Sublineages of Mycobacterium tuberculosis Beijing Genotype in China.

Mycobacterium tuberculosis Beijing genotype originated in China and has undergone a dramatic population growth and global spread in the last century. Here, a collection of M. tuberculosis Beijing family isolates from different provinces across all China was genotyped by high-resolution (24-MIRU-VNTR) and low-resolution, high-rank (modern and ancient sublineages) markers. The molecular profiles and global and local phylogenies were compared to the strain phenotype and patient data. The phylogeographic patterns observed in the studied collection demonstrate that large-scale (but not middle/small-scale) distance remains one of the decisive factors of the genetic divergence of M. tuberculosis populations. Analysis of diversity and network topology of the local collections appears to corroborate a recent intriguing hypothesis about Beijing genotype originating in South China. Placing our results within the Eurasian context suggested that important Russian B0/W148 and Asian/Russian A0/94-32 epidemic clones of the Beijing genotype could trace their origins to the northeastern and northwestern regions of China, respectively. The higher clustering of the modern isolates in children and lack of increased MDR rate in any sublineage suggest that not association with drug resistance but other (e.g., speculatively, virulence-related) properties underlie an enhanced dissemination of the evolutionarily recent, modern sublineage of the Beijing genotype in China.

Molecular Typing Characteristic and Drug Susceptibility Analysis of Mycobacterium tuberculosis Isolates from Zigong, China.

China is one of the 22 countries with high TB burden worldwide, and Sichuan contained the second-largest number of TB cases among all of the Chinese provinces. But the characteristics of Mycobacterium tuberculosis circulated in Zigong, Sichuan, were still unknown. To investigate the character and drug resistance profile, 265 clinical isolates were cultured from tuberculosis patient's sputum samples in the year of 2010, of which the genetic profile was determined by using Spoligotyping and MIRU-VNTR typing methods, and the drug sensibility testing to the four first-line and four second-line antituberculosis (anti-TB) drugs was performed by using proportion method on Lowenstein-Jensen (L-J) media. The major Spoligotype was Beijing family (143/265, 53.96%), followed by T (80/265, 30.19%) and H (9/265, 3.40%) genotypes; the total Hunter-Gaston discrimination index (HGDI) of the 24 loci MIRU-VNTR was 0.9995. About 27.17% (72/265) of the isolates were resistant to at least one of the eight tested anti-TB drugs, and for Beijing and non-Beijing family isolates the proportion of drug resistance was 28.47% (41/144) and 25.62% (31/121), respectively. That is, the most prevalent genotype here was Beijing family, and the 24 loci VNTR analysis could supply a high resolution for genotyping, and Beijing and non-Beijing isolates had no difference (p > 0.05) for drug resistance.

Prevalence of mutations conferring resistance among multi- and extensively drug-resistant Mycobacterium tuberculosis isolates in China.

To identify the mutations in multi- and extensively drug-resistant tuberculosis isolates and to evaluate the use of molecular markers of resistance, we analyzed 257 multi- and extensively drug-resistant isolates and 64 pan-sensitive isolates from 23 provinces in China. Seven loci associated with drug resistance, including rpoB for rifampin (RIF), katG, inhA and oxyR-ahpC for isoniazid (INH), gyrA and gyrB for ofloxacin (OFX), and rrs for kanmycin (KAN), were examined by DNA sequencing. Compared with the phenotypic data, the sensitivity and specificity for DNA sequencing were 91.1% and 98.4% for RIF, 80.2% and 98.4% for INH, 72.2% and 98.3% for OFX and 40% and 98.2% for KAN, respectively. The most common mutations found in RIF, INH, OFX and KAN resistance were Ser531Leu (48.2%) in rpoB, Ser315Thr (49.8%) in katG, C(-15)T (10.5%) in inhA, Asp94Gly (20.3%), Asp94Ala (12.7%) and Ala90Val (21.5%) in gyrA, and A1401G (40%) in rrs. This molecular information will be helpful to establish new molecular biology-based methods for diagnosing multi- and extensively drug-resistant tuberculosis in China.

[Serum miRNAs as new biomarkers for esophageal squamous cell carcinoma].

Esophageal squamous cell carcinoma (ESCC) is one of the most common types of malignancies in China. Most ESCC patients are diagnosed at middle to late stages with poor prognosis due to the lack of an effective method for early diagnosis. MicroRNAs (miRNAs) are a family of endogenous small non-coding RNAs that can regulate ESCC development and progression by repressing their specific target genes' expression. Compared to traditional biomarkers (e.g., mRNAs and proteins), miRNAs are more stable and can be readily screened and accurately quantitated and analyzed, making them ideal new-generation of biomarkers for early cancer detection and prognostic evaluation. Recent studies have shown that the changes of the expression levels of some serum miRNAs from ESCC patients significantly correlate with their diagnostic and prognostic outcome. In this review, we summarize the trend of the expression changes of miRNAs in ESCC patients' serum and discuss the possibility of detecting these miRNAs' expression changes as a novel method for ESCC early diagnosis and prognostic evaluation. Notably, the results of serum miRNAs from different detection methods are not completely consistent. Thus, we also discuss several possible reasons for such inconsistency.

Prevalence of drug resistant Mycobacterium tuberculosis among children in China.

The available data on the epidemic of drug resistant tuberculosis (TB) among children in China is limited. This study attempted to clarify the drug resistance profiles of clinical strains isolated from children and estimate risk factors related to acquisition of drug resistance. All Mycobacterium tuberculosis strains from children (age <15 years) and adolescent (age 15-18 years) TB patients received in the strain library of Chinese Center for Disease Control and Prevention between January 2005 and December 2012 were included in the study. A study collection included 450 clinical isolates (100 from children, 159 from adolescents, and 191 from adults) from all over China. Drug susceptibility testing was performed by a proportion method. As a result, the drug resistance and multi-drug resistance (MDR) rates in children were 55% (55/100) and 22% (22/100), respectively. In children with MDR-TB, new cases accounted for 40.9% (9/22). Compared with adults, the drug resistance rates were similar in all subgroups (new cases, previously treated cases and all cases) of children (P > 0.05), except for the lower resistance rate to isoniazid in total cases of children (P = 0.011). Patient related information was included in the MDR-TB association analysis. The treatment history was found to be strongly associated with MDR-TB in all three age groups (P < 0.05). Our results demonstrate that the prevalence of drug resistant TB in children in China is alarmingly high and similar to that seen in adults. In contrast, in adolescents, the drug resistance rate to most tested drugs was lower than in adults. Primary transmission and inadequate treatment are two equally important factors for the high MDR-TB rate in children. Thus, major efforts in the TB control in children should focus on decreasing the transmission of drug resistant TB and early testing of drug resistance.