Deletion of region of difference 181 in Mycobacterium tuberculosis Beijing strains

AbstractObjectives/backgroundThe region of differences (RDs) polymorphisms is a potential molecular epidemiology method to distinguish origins of Mycobacterium tuberculosis. To date, 68 RDs have been identified in M. tuberculosis. This study was designed to determine the frequency of RD deletions in M. tuberculosis strains that were isolated from patients with pulmonary tuberculosis who were referred to the National Research Institute of Tuberculosis and Lung Disease for diagnosis and treatment. Therefore, highly polymorphic regions (RD1, RD150, and RD181) among M. tuberculosis strains isolates were investigated.MethodsA total of 250 M. tuberculosis isolates were identified by conventional and molecular methods. Subsequently, spoligotyping and RD typing (RD1, RD150 and RD181) were performed to genotype these strains.ResultsThe most frequent spoligotype belonged to Haarlem (n=85, 34.0%) followed by CAS (n=54, 21.6%), T1 (n=27, 10.8%), and Beijing (n=28, 11.2%) lineages. Deletion in RD181 was identified only among the Beijing lineage (Fig. 1).ConclusionAs we found a deletion in RD181 in the Beijing strains only, we propose to use this marker as an identification tool for genotyping of the Beijing strain

A 3D tailored monolithic glass chip for stimulating and recording zebrafish neuronal activity with a commercial light sheet microscope

Microfluidic technology is unrivaled in its ability to apply soluble chemical stimuli with high spatiotemporal precision. Analogous, light–sheet microscopy is unmatched in its ability of low phototoxic but fast volumetric in vivo imaging with single cell resolution. Due to their optical translucency during the larval stages, zebrafish (Danio rerio) are an ideal model to combine both techniques; yet, thus far this required light–sheet microscopes, which were in most cases custom–built and adapted to the available softlithographic chip technology. Our aim was to use a commercial light–sheet microscope to illuminate a microfluidic chip from two opposite lateral directions and to record images with the detection objective placed orthogonally above the chip. Deep tissue penetration can be achieved by superimposing beams from opposite directions to form a single light sheet. But a microfluidic chip that allows a) targeted stimulus application in a closed microenvironment, b) interference–free incoupling of excitation light from two directions and c) outcoupling of fluorescence in the perpendicular direction through an optically perfect cover glass was not known until now. Here, we present a monolithic glass chip with the required plane-parallel sidewalls and cover slide closure at the top, constructed by advanced femtosecond laser ablation, thermal bonding and surface smoothing processes. In addition, the 3D shape of a fish fixator unit was tailored to match the body shape of a zebrafish larva to ensure stable positioning during whole–brain recording. With hydrodynamic focusing a targeted partial exposure of the larva’s head to chemical stimuli and fast position switching (in less than 10 s) was possible. With the capabilities of this unique monolithic glass chip and its up–scalable wafer–level fabrication process, the new NeuroExaminer is prone to become an excellent addition to neurobiology laboratories already equipped with high–quality commercial light sheet microscopes

Study of Mycobacterium bovis genotypes in human and bovine isolates using spoligotyping, MIRUVNTR and RFLP-PCR

Aims and objectives: The aim of this study is to investigate and detect the prevalence of Mycobacterium bovis subtypes (Mycobacterium bovis subtype bovis and Mycobacterium bovis subtype caprae) in humans and compare the genetic diversity of Mycobacterium bovis in humans and cattle with spoligotyping methods, as well as pyrazinamide susceptibility study of subtypes. Methods: Examining these subtypes with molecular epidemiology techniques is particularly important due to different treatment of M. bovis diverse subtypes in humans. Culture tests were performed on clinical samples that were isolated from Lowenstein-Jensen culture medium. Identification tests were performed to differentiate Mycobacterium bovis from Mycobacterium tuberculosis. DNA was extracted and spoligotyping (spacer oligonucleotide typing) was performed using the DRb and DRa primers. Results: The results were analyzed with the SPOLDB4 site. PCR-RFLP method of pncA gene was used to evaluate the resistance to pyrazinamide and pncA gene polymorphism. Conclusions: Mycobacterium bovis subtype bovis in the Iranian population was reported with a frequency of 0.6% which was below the average of the previous reviews. In this study, all the strains were M. bovis subtype bovis resistant to pyrazinamide. No Mycobacterium bovis subtype caprae was detected. The only shared ST between humans and cattle was ST694. Mycobacterium bovis subtype bovis with ST 595 was reported as human bovis index