Xuxia Zhang

Comparison of gyrA gene mutations between laboratory-selected ofloxacin-resistant Mycobacterium tuberculosis strains and clinical isolates

To understand the relationship between mutations in the quinolone resistance-determining region (QRDR) of the gyrA gene and drug resistance to ofloxacin, 85 laboratory-selected ofloxacin-resistant Mycobacterium tuberculosis mutant strains and 110 M. tuberculosis clinical isolates, screened by denaturing high-performance liquid chromatography to contain mutations, were analysed for their mutation patterns by sequencing as well as their ofloxacin minimal inhibitory concentrations (MICs). All mutations detected occurred at the codons Ala74, Ala90, Ser91 and Asp94 in all strains. One of the five different forms of missense mutation in Asp94 occurred in 60% of the laboratory-selected strains and 78% of the clinical isolates. However, 53 clinical isolates (48%) and only 2 laboratory-selected strains (2.4%) harboured double point mutations. The mutation Ala74Ser occurred only in the clinical isolates and only in combination with the Asp94Gly mutation. The ofloxacin MIC for the clinical isolates ranged from 0.5microg/mL to 20microg/mL, whilst the MICs for the laboratory-selected strains were > or =10microg/mL. The differences in gyrA gene mutation patterns and MICs between the laboratory-selected resistant strains and clinically isolated resistant strains identified here might help to understand the mechanisms involved in fluoroquinolone resistance.

Identification of a novel IRGM promoter single nucleotide polymorphism associated with tuberculosis

BACKGROUND Human immunity-related GTPase M (IRGM) is found to play an important role in defense against intracellular pathogen Mycobacterium tuberculosis in vitro by regulating autophagy. To verify whether single nucleotide polymorphisms (SNPs) in the promoter region of IRGM gene are associated with tuberculosis (TB) 1.7 kb IRGM promoter region was sequenced and SNP analysis was conducted in TB patients and healthy controls. METHODS A simple and rapid procedure for extracting DNA from clotted-blood was developed in this study. A 1.7 kb IRGM promoter region was amplified and sequenced for nucleotide polymorphism search. Then, 3 SNPs were selected and analyzed in 216TB patients and 275 healthy subjects by ligase detection reaction technique. RESULTS DNA extracted by our method was of high quality and suitable for PCR, sequencing, and genotyping. We identified 29 polymorphisms in the 1.7 kb IRGM promoter region, including 11 novel polymorphisms not yet reported. Large population analysis showed that frequencies of -1208A allele (P=0.031), -1208AA genotype (P=0.042), and -1208A/-1161C/-947C (P=0.035) and -1208G/-1161C/-947C (P=0.030) haplotypes in cases were significantly different from those in controls. CONCLUSIONS In 1.7 kb IRGM promoter region, only -1208A/G polymorphism is associated with susceptibility to TB.

Characterization of Extensively Drug-Resistant Mycobacterium tuberculosis Clinical Isolates in China

ABSTRACT Thirteen extensively drug-resistant tuberculosis isolates which were highly resistant to a broad spectrum of antituberculosis drugs were identified from 1,926 clinical isolates in China. They had highly diverse mycobacterial interspersed repetitive-unit-variable-number tandem-repeat patterns. Most, but not all, of the drug target genes had mutations contributing to resistance to the corresponding drug.

Decreased serum 5-oxoproline in TB patients is associated with pathological damage of the lung

BACKGROUND Tuberculosis (TB) is a serious world-wide health problem, causing millions of deaths every year. Metabolomics is a relatively new approach to identify disease specific biomarkers. However, there is little information available on metabolite biomarkers in TB. In this study, we used gas chromatography/time-of-flight mass spectrometry (GC/TOF-MS) to identify serum metabolite biomarkers associated with the active state of TB. METHODS Potential biomarkers were selected by comparing serum metabolites in 10 healthy donors with 10TB patients, and in 6TB patients before and after treatment. Selected biomarkers were then validated using a larger population of samples from 120 healthy donors and 120TB patients derived from different clinical backgrounds RESULTS The 5-oxoproline level was consistently low in patients with active TB. Further validation in larger population of clinical samples showed that 5-oxoproline was associated with pathological damage of the lung but not with age, sex, or bacterial burden in TB patients. CONCLUSIONS Serum 5-oxoproline may be a useful biomarker for active TB and pathological damage of the lung.

Genotypic diversity analysis of Mycobacterium tuberculosis strains collected from Beijing in 2009, using spoligotyping and VNTR typing.

Background Tuberculosis (TB) is a serious problem in China. While there have been some studies on the nationwide genotyping of Mycobacterium tuberculosis (M. tuberculosis), there has been little detailed research in Beijing, the capital of China, which has a huge population. Here, M. tuberculosis clinical strains collected in Beijing during 2009 were genotyped by classical methods. Methodology/Principal Findings Our aim was to analyze the genetic diversity of M. tuberculosis strains within the Beijing metropolitan area. We characterized these strains using two standard methods, spoligotyping (n = 1585) and variable number of tandem repeat (VNTR) typing (n = 1053). We found that the most prominent genotype was Beijing family genotype. Other genotypes included the MANU, T and H families etc. Spoligotyping resulted in 137 type patterns, included 101 unclustered strains and 1484 strains clustered into 36 clusters. In VNTR typing analysis, we selected 12-locus (QUB-11b, MIRU10, Mtub21, MIRU 23, MIRU39, MIRU16, MIRU40, MIRU31, Mtub24, Mtub04, MIRU20, and QUB-4156c) and named it 12-locus (BJ) VNTR. VNTR resulted in 869 type patterns, included 796 unclustered strains and 257 strains clustered into 73 clusters. It has almost equal discriminatory power to the 24-locus VNTR. Conclusions/Significance Our study provides a detailed characterization of the genotypic diversity of M. tuberculosis in Beijing. Combining spoligotyping and VNTR typing to study the genotyping of M. tuberculosis gave superior results than when these techniques were used separately. Our results indicated that Beijing family strains were still the most prevalent M. tuberculosis in Beijing. Moreover, VNTR typing analyzing of M. tuberculosis strains in Beijing was successfully accomplished using 12-locus (BJ) VNTR. This method used for strains genotyping from the Beijing metropolitan area was comparable. This study will not only provide TB researchers with valuable information for related studies, but also provides guidance for the prevention and control of TB in Beijing.

Evaluation of Spoligotyping and MIRU-VNTR for Mycobacterium bovis in Xinjiang, China.

Mycobacterium bovis, the causative agent of bovine tuberculosis (bTB) in cattle, is also a pathogen for human and other mammals. In this study, 406 cows were screened for bTB by both single intradermal comparative cervical tuberculin (SICCT) test and IFN-γ assay. 135 M. bovis were isolated from 31 SICCT and IFN-γ double-positive cows in Xinjiang Uygur Autonomous Region of China. Spoligotyping and MIRU-VNTR were evaluated for genotyping, and 4 and 7 genotypes were identified, respectively. A new combination of nine MIRU-VNTR loci was most discriminative for M. bovis clones from Xinjiang. Interestingly, two new spoligotypes (SB1903 and SB1904) and special repeat numbers of three loci (ETR-D, QUB 1895 and QUB 3336) were discovered in this study. These results indicated a specific epidemic conservation in Xinjiang, China. M. bovis strains with the unique genotypes were isolated from the herds maintaining parent cows imported from the bTB-free countries, suggesting a possible transmission from the local breed of Xinjiang brown cattle.

Efficacy and Safety of Basiliximab Versus Daclizumab in Kidney Transplantation: A Meta-Analysis

BACKGROUND Interleukin-2 receptor antagonists (IL-2RAs) have been extensively used in kidney transplant patients to prevent the occurrence of acute rejection. The efficacy and safety of basiliximab and daclizumab, the 2 most commonly used IL-2RAs in clinics, have been compared in a number of randomized controlled trials, but no definite conclusions have been drawn. OBJECTIVE This meta-analysis aimed to compare the efficacy and safety of basiliximab and daclizumab in kidney transplant patients. METHODS We performed keyword searches in Pubmed, Embase, and the Cochrane library. In total, 6 randomized controlled trials with 509 patients were included in this meta-analysis. Data collected included patient survival, graft survival, acute rejection, infection, and cytomegalovirus infection. The outcome measure was the relative risk of basiliximab versus daclizumab. RESULTS Therapy with basiliximab and daclizumab resulted in similar outcomes regarding acute rejection (6-month 95% confidence interval [CI], 0.09-1.14; 12-month 95% CI, 0.53-1.91), patient survival (95% CI, 0.97-1.04), graft survival (95% CI, 0.98-1.08), infection (95% CI, 0.66-1.01), and cytomegalovirus infection (95% CI, 0.45-1.14) within the follow-up period. There were no significant differences in safety and efficacy between the 2 drugs. CONCLUSIONS The safety and efficacy of daclizumab and basiliximab are similar in kidney transplant recipients.

Potential role for Rv2026c- and Rv2421c- specific antibody responses in diagnosing active tuberculosis

The current diagnostic methods for tuberculosis (TB) have several limitations. Although commercial serological tests based on antibody detection are available, their variable accuracies limit their roles in the clinic. The aim of this study was to discover the improved biomarkers for TB disease by investigating the serum profiles of IgG and IgM antibodies against nearly all Mycobacterium tuberculosis (MTB) antigens in 36 active TB patients and 18 healthy controls (HCs) using proteome microarrays. Our results revealed that multiple antigens could induce stronger serum IgG or IgM responses in TB patients compared to HCs, among them, Rv2026c and Rv2421c were further validated by ELISA with sera from 221 samples and showed the moderate performance in diagnosing TB by receiver operating characteristic analysis. Moreover, logistic regression analysis was performed to establish a combined panel that provided better sensitivity and specificity at 82.5% and 88.12%, respectively, than single antigens in the diagnosis of active TB. Furthermore, the antibody reactivity against Rv2026c and Rv2421c was correlated with clinical backgrounds. These results suggest that the combination of different antigens and classes of antibodies could provide promise and encouragement in developing an efficient serological test for the diagnosis of active TB.

The study on the association between Beijing genotype family and drug susceptibility phenotypes of Mycobacterium tuberculosis in Beijing

The predominant prevalent Mycobacterium tuberculosis (M. tb) lineage was the Beijing genotype family in Beijing. There has been no systematic study on the association between drug resistance and Beijing genotype. Here we collected 268 M. tb strains, analyzed the background information and the bacteriological characteristics. The mean age of the cases was 40.12 years; male patients were almost three times than female patients. After genotyping analyzation, 81.7% (219/268) strains were categorized as Beijing genotype; no significant difference was observed between Beijing and non-Beijing genotype in gender, age and treatment history. Drug susceptibility testing (DST) analyzation demonstrated that 172 (64.2%) strains were fully sensitive to all drugs (Isoniazid, Rifampin, Streptomycin, and Ethambutol), while 96 (35.8%) strains were resistant to at least one of the drugs. Beijing genotype strains exhibited a significantly higher clustering rate. However, no significant association relationship was observed between drug resistance and Beijing genotype family. The study provided insights into the genotype diversity and revealed that the frequencies of drug-resistance of Beijing genotype strains.It would be helpful for the establishment of the efficient tuberculosis (TB) prevention and control strategy in Beijing.