Objective To evaluate a rapid biochip system for the determination of muhidrugresistant tuberculosis (MDR-TB) in Mycobacterium tuberculosis isolates. MethodsA total of 1 186 clinical strains, including 800 rifampin (RFP) resistant isolates, 797 isoniozid (INH)resistant isolates, 791 MDR-TB and 380 susceptible strains, were selected from Beijing Chest Hospital, Shanghai Pulmonary Hospital and Guangzhou Chest Hospital respectively using stratified sampling method. Biochips were used to detect loci of rpoB 511 (T→C), 513 (A→C, C→A), 516 (G→T, A→T, A→G) , 526 (C→T, C→G, A→T, A→G), 531 (C→T, C→G), 533 (T→C), katG 315 ( G→C, G→A) and inhA -15 (C→T). Absolute concentration drug susceptibility test of RFP and INH were performed to serve as the gold standard to calculate susceptibility, specificity and overall concordance of biochip test. All polymerase chain reaction (PCR) products were sequenced to confirm the mutations. ResultsThe concordances between the biochip system and absolute concentration drug susceptibility test were 93.7％ ( 1 108/1 183 ) for RFP, 83. 8％(994/1 186) for INH and 82.4％ (975/1 183) for MDR-TB. Compared with absolute concentration drug susceptibility test, the biochip method displayed a sensitivity of 92. 0％ (733/797) and 77. 4％ (617/797)and a specificity of 97. 2％ (375/386) and 96. 9％ (377/389) for RIF and INH, respectively. For MDR-TB, the biochip system reached a sensitivity of 74. 6％ ( 588/788 ) and a specificity of 98.0％ ( 387/395 ).Among rpoB mutants, mutations were mostly detected at codon 531[64. 5％ (480/744)]. In stains with mutations in katG or inhA, 77.4％ ( 487/629 ) had mutation at codon 315 ( TCG ) of katG only. The sequencing results had a high concordance with that of the biochip method. There were slight differences in 5 strains, among which one strain was detected by biochip as katG 315(G→C) mutant, but was identified by sequencing as wild type, and mutation types other than those detected by the biochip were confirmed in the other 4 strains by sequencing. Conclusion This biochip system is adapted for extensive application in clinical diagnosis, as it allows fast and reliable detection of resistance to isoniazid and rifampin in tuberculosis clinical isolates.

BACKGROUND:Mouse osteogenic potential is regulated by the JAK-STAT signaling pathway,and interleukin-9 can regulate multiple cellular functions through the JAK-STAT pathway,which has the potential to be a novel cytokine that regulates osteogenic potential. OBJECTIVE:To investigate the effect of interleukin-9 deficiency on osteogenic potential in mice METHODS:The femurs collected from 2-month-old wild-type and interleukin-9 knockout mice were subjected to Micro-CT scanning to analyze the changes in bone mass.Then,hematoxylin-eosin staining,Masson staining,and immunohistochemical staining of type Ⅰ collagen were performed on the slices of the femurs of mice.Bone marrow cells from 2-month-old wild-type and interleukin-9 knockout mice were extracted for colony-forming assay and detection of osteogenic gene expression in bone marrow mesenchymal stem cells.To further verify whether interleukin-9 worked through the JAK-STAT pathway,the expression of STAT3 protein was detected by western blot. RESULTS AND CONCLUSION:Micro-CT results showed the bone mass of interleukin-9 knockout mice decreased significantly compared with that of wild-type mice.In addition,the bone mineral density,bone volume fraction,trabecular number significantly decreased and trabecular separation markedly escalated in interleukin-9 knockout mice.The findings of hematoxylin-eosin staining were consistent with Micro-CT results.Interleukin-9 knockout mice had lower bone trabecular density.Type I collagen immunohistochemistry staining and Masson staining indicated the number of type Ⅰ collagen positive osteoblasts was significantly reduced and the capacity of collagen formation was damaged in interleukin-9 knockout mice.The results of colony-forming assay indicated that the mineralization capacity of osteoblast in interleukin-9 knockout mice were significantly lower than that in wild-type mice.Western blot results showed that osteogenesis induction activated STAT3 signaling,and the pSTAT3 level in wild-type mice with osteogenic induction was significantly higher than that in interleukin-9 knockout mice with osteogenic induction.These findings suggest that interleukin-9 regulates osteogenesis through the JAK-STAT3 pathway and interleukin-9 deficiency inhibits osteoblast differentiation and function,which may lead to reduced bone mass in interleukin-9 knockout mice.