Synchrotron microtomography applied to the volumetric analysis of internal structures of Thoropa miliaris tadpoles.

Amphibians are models for studying applied ecological issues such as habitat loss, pollution, disease, and global climate change due to their sensitivity and vulnerability to changes in the environment. Developmental series of amphibians are informative about their biology, and X-ray based 3D reconstruction holds promise for quantifying morphological changes during growth-some with a direct impact on the possibility of an experimental investigation on several of the ecological topics listed above. However, 3D resolution and discrimination of their soft tissues have been difficult with traditional X-ray computed tomography, without time-consuming contrast staining. Tomographic data were initially performed (pre-processing and reconstruction) using the open-source software tool SYRMEP Tomo Project. Data processing and analysis of the reconstructed tomography volumes were conducted using the segmentation semi-automatic settings of the software Avizo Fire 8, which provide information about each investigated tissues, organs or bone elements. Hence, volumetric analyses were carried out to quantify the development of structures in different tadpole developmental stages. Our work shows that synchrotron X-ray microtomography using phase-contrast mode resolves the edges of the internal tissues (as well as overall tadpole morphology), facilitating the segmentation of the investigated tissues. Reconstruction algorithms and segmentation software played an important role in the qualitative and quantitative analysis of each target structure of the Thoropa miliaris tadpole at different stages of development, providing information on volume, shape and length. The use of the synchrotron X-ray microtomography setup of the SYRMEP beamline of Elettra Synchrotron, in phase-contrast mode, allows access to volumetric data for bone formation, eye development, nervous system and notochordal changes during the development (ontogeny) of tadpoles of a cycloramphid frog Thoropa miliaris. As key elements in the normal development of these and any other frog tadpole, the application of such a comparative ontogenetic study, may hold interest to researchers in experimental and environmental disciplines.

Molecular characterisation of multidrug-resistant Mycobacterium tuberculosis isolates from a high-burden tuberculosis state in Brazil.

Tuberculosis (TB) is the leading cause of death among infectious diseases worldwide. Among the estimated cases of drug-resistant TB, approximately 60% occur in the BRICS countries (Brazil, Russia, India, China and South Africa). Among Brazilian states, primary and acquired multidrug-resistant TB (MDR-TB) rates were the highest in Rio Grande do Sul (RS). This study aimed to perform molecular characterisation of MDR-TB in the State of RS, a high-burden Brazilian state. We performed molecular characterisation of MDR-TB cases in RS, defined by drug susceptibility testing, using 131 Mycobacterium tuberculosis (M.tb) DNA samples from the Central Laboratory. We carried out MIRU-VNTR 24loci, spoligotyping, sequencing of the katG, inhA and rpoB genes and RDRio sublineage identification. The most frequent families found were LAM (65.6%) and Haarlem (22.1%). RDRio deletion was observed in 42 (32%) of the M.tb isolates. Among MDR-TB cases, eight (6.1%) did not present mutations in the studied genes. In 116 (88.5%) M.tb isolates, we found mutations associated with rifampicin (RIF) resistance in rpoB gene, and in 112 isolates (85.5%), we observed mutations related to isoniazid resistance in katG and inhA genes. An insertion of 12 nucleotides (CCAGAACAACCC) at the 516 codon in the rpoB gene, possibly responsible for a decreased interaction of RIF and RNA polymerase, was found in 19/131 of the isolates, belonging mostly to LAM and Haarlem families. These results enable a better understanding of the dynamics of transmission and evolution of MDR-TB in the region.

Genomic characterization of MDR/XDR-TB in Kazakhstan by a combination of high-throughput methods predominantly shows the ongoing transmission of L2/Beijing 94-32 central Asian/Russian clusters.

BACKGROUND: Kazakhstan remains a high-burden TB prevalence country with a concomitent high-burden of multi-drug resistant tuberculosis. For this reason, we performed an in depth genetic diversity and population structure characterization of Mycobacterium tuberculosis complex (MTC) genetic diversity in Kazakhstan with both patient and community benefit. METHODS: A convenience sample of 700 MTC DNA cultures extracts from 630 tuberculosis patients recruited from 12 out of 14 regions in Kazakhstan, between 2010 and 2015, was independently studied by high-throughput hybridization-based methods, TB-SPRINT (59-Plex, n = 700), TB-SNPID (50-Plex, n = 543). DNA from 391 clinical isolates was successfully typed by two methods. To resolve the population structure of drug-resistant clades in more detail two complementary assays were run on the L2 isolates: an IS6110-NTF insertion site typing assay and a SigE SNP polymorphism assay. RESULTS: Strains belonged to L2/Beijing and L4/Euro-American sublineages; L2/Beijing prevalence totaled almost 80%. 50% of all samples were resistant to RIF and to INH., Subtyping showed that: (1) all L2/Beijing were "modern" Beijing and (2) most of these belonged to the previously described 94-32 sublineage (Central Asian/Russian), (3) at least two populations of the Central Asian/Russian sublineages are circulating in Kazakhstan, with different evolutionary dynamics. CONCLUSIONS: For the first time, the global genetic diversity and population structure of M. tuberculosis genotypes circulating in Kazakhstan was obtained and compared to previous local studies. Results suggest a region-specific spread of a very limited number of L2/Beijing clonal complexes in Kazakhstan many strongly associated with an MDR phenotype.

The efficacy of intraoperative autologous transfusion in major orthopedic surgery: a regression analysis.

Perioperative blood loss associated with 89 cases of major orthopedic surgery was compared with that of a control group of 89 to determine the effectiveness of intraoperative autologous transfusion. Volume of banked blood transfused and hematocrit change were used to determine total blood loss. The orthopedic cases consisted of cemented "virgin" total hip replacement, cemented virgin tricompartmental knee replacement, and spine fusion. Use of an autotransfusion device (Cell Saver) intraoperatively was associated with significantly smaller volumes of transfused banked blood and significantly smaller hematocrit drops in the groups of patients who underwent total hip replacement or spine fusion, but not in the group of patients who underwent total knee replacement. One potential source of bias in the study stems from the fact that four days were allotted for equilibrium from perioperative blood loss in the hip and knee replacement groups, while, for reasons of data availability, equilibrium time in the spine fusion groups was two days.