A new Multilocus Sequence Analysis Scheme for Mycobacterium tuberculosis

Objective Tuberculosis remains one of the most serious infectious diseases in the world. In this study, a scheme of Mycobacterium tuberculosis （M. tuberculosis） multilocus sequence analysis （MLSA） was established for the phylogenetic and epidemiology analysis. Methods To establish the scheme of M. tuberculosis MLSA, the genome of H37Rv, CCDC5079 and CCDC5180 were compared, and some variable genes were chosen to be the MLSA typing scheme. 44 M. tuberculosis clinical isolates were typed by MLSA, IS6110-RFLP, and soligotyping, to evaluate the MLSA methods. Results After comparison of the genome, seven high discrimination gene loci （recX, rpsL, rmlC, rpmG1, mprA, gcvH, ideR） were chosen to be the MLSA typing scheme finally. 11 variable SNP sites of those seven genes were found among the 44 M. tuberculosis isolate strains and 11 sequence types （STs） were identified. Based on the Hunter-Gaston Index （HGI）, MLSA typing was not as good for discrimination at the strain level as IS6110-RFLP, but the HGI was much better than that of spoligotyping. In addition, the MEGA analysis result of MLSA data was similar to spoligotyping/PGG lineage, showing a strong phylogenetic signal in the modern strains of M. tuberculosis. The MLSA data analysis by eBURST revealed that 4 sequence types （ST） came into a main cluster, showing the major clonal complexes in those 44 strains. Conclusion MLSA genotyping not only can be used for molecular typing, but also is an ideal method for the phylogenetic analysis for M. tuberculosis.

Detection of the mutations in katG315 and inhA-15 of Mycobacterium tuberculosis strains isolated from Chinese patients

The situation of tuberculosis and M. tuberculosis （MTB） drug resistance is severe in China.Among the MTB isolates, 27.8% were resistant to one primary antituberculosis drug at least. Isoniazid （INH） is one of the first-line anti-TB drugs. The rates of primary resistance and acquired resistance to INH were 11.0% and 31.0%, respectively. MTB strains resistance to INH is mainly caused by the alterations in several genes encoding the molecular targets as follows： catalase peroxidase （katG）,

Controlling strategy of dormant Mycobacterium tuberculosis

Objective This study aimed to review the available literatures on control of latent tuberculosis （TB） infection and propose a new control strategy to shorten the course of TB chemotherapy. Data sources The data used in this review were mainly obtained from articles listed in PubMed. The search terms were ＂therapy （treatment） of tuberculosis; ＂therapy （treatment） of latent TB infection; and ＂vaccine of TB.＂ Study selection Articles regarding treatment and vaccine of TB were selected and reviewed. Results The most crucial reason causing the prolonged course of TB chemotherapy is the dormant state of Mycobacterium tuberculosis （M. tuberculosis）. Nevertheless, there are, to date, no effective drugs that can directly kill the dormant cells of M. tuberculosis in clinical therapy. In accordance with the growth cycle of dormant M. tuberculosis in the body, the methods for controlling dormant M. tuberculosis include direct killing with drugs, prevention of dormant M. tuberculosis resuscitation with vaccines, and resuscitating dormant M. tuberculosis with preparations or drugs and then thoroughly killing these resuscitated M. tuberculosis by using anti-TB therapy. Conclusions The comprehensive analysis of the above three methods suggests that the drugs directly killing dormant cells are in clinical trials, TMC207 is the most beneficial for controlling TB. Because the side effect of vaccines is less and their action period is long, prevention of dormant cells resuscitation with vaccines is promising. The last control method makes it probable that when a huge number of active cells of M. tuberculosis have been killed and eradicated after 1-month short chemotherapy, only a strong short-term subsequent chemotherapy can completely kill and eradicate the remaining M. tuberculosis. This control strategy is expected to significantly shorten the course of TB chemotherapy and bring a new change and breakthrough in TB treatment.

Screening and Assessing 11 Mycobacterium tuberculosis Proteins as Potential Serodiagnostical Markers for Discriminating TB Patients from BCG Vaccinees

Purified protein derivative （PPD） skin tests often yield poor specificity, so that to develop new serological antigens for distinguishing between Mycobacterium tuberculosis infection and Bacille Calmette-Guerin （BCG） vaccination is a priority, especially for developing countries like China. We predicted the antigenicity for selected open reading frames （ORFs） based on the genome sequences of M. tuberculosis H37Rv and M. boris BCG, as well as their functions and differences of expression under different stimulus. The candidate ORFs were cloned from H37Rv sequences and expressed as recombinant proteins in Escherichia coli. We studied the serodiagnostic potential of 11 purified recombinants by using enzyme-linked immunosorbent assay （ELISA） and involving a cohort composed of 58 TB patients （34 males and 24 females）, 8 healthy volunteers and 50 PPD-negative individuals before and after BCG vaccination. For all the 11 antigens, the median OD values for the sera from TB patients were statistically significantly higher than those for PPD-negative individuals before or after BCG vaccination （P〈0.01）. They had at least 92% specificity in healthy controls and six seroantigens （Rv0251c, Rv1973, Rv2376c, Rv2537c, Rv2785c and Rv3873A） were never reported with seroantigenicities previously. Thus the approach combining comparative genomics, bioinformatics and ELISA techniques can be employed to identify new seroantigens distinguishing M. tuberculosis infection from BCG vaccination.

High-Speed Analyzing PCR Products of M.tuberculosis Genome Stained by Ethidium Bromide on Microchip Gel Electrophoresis

The technique of microchip gel electrophoresis (MCGE) was used to analyze the polymerase chain reaction (PCR) products of M. tuberculosis Genome stained by ethidium bromide. The electrophoretic process was completed within 3-4 min and the results show that the technique of microchip electrophoresis is a high speed and high sensitivity analyzing method.

Tuberculous Spondylitis: Clinical Features of 36 Patients

We report 36 cases of spinal tuberculosis who were evaluated at the Internal Medicine Department of Istanbul University between January 1990 and March 2016. Twelve cases were accompanied by active pulmonary tuberculosis while ten patients had a previous history of tuberculosis. Eight patients had diabetes mellitus;six patients were on chronic steroid treatment with a mean dose of 24 mg/day while two patients used azathioprine and methotrexate. The dominant symptom was back pain that was present in 64% of the subjects followed by low grade fever (42%), and malaise (38%). Lytic and destructive lesions were noted in various vertebrae in all of the cases while four patients had spinal compression, and two patients had iliopsoas abscess. The preliminary diagnosis was myeloproliferative disease with vertebral metastasis in eight patients. Tuberculin test was over 15 mm in 20 patients (58.8%) while ERS and Creactive protein were highly elevated in 78%, and 84% of the patients. Diagnosis of tuberculosis was confirmed by culture of aspirated material from intervertebral disc space, collection under CT guidance, sputum or bronchial lavage, and by retrospective anti-tuberculous treatment response. MRI revealed compatible findings with spinal tuberculosis in 30 (80%) patients. In endemic countries, tuberculous spondilitis should be considered in immunosuppressed patients with back pain. Current or past tuberculosis infection is not a reliable indicator for Pott’s disease in these patients. Laboratory findings were not usually useful to support the diagnosis. The low sensitivity of the tuberculin test may lead to an erronous diagnosis. Sensitivity of vertebral radiography and CT was low. MRI was the most useful radiologic investigation for the diagnosis of spinal tuberculosis. Tuberculosis of the spine is a diagnostic impasse notably in immunodeficient patients for the clinician.