Molecular characterisation of Mycobacterium tuberculosis isolates in the First National Survey of Anti-tuberculosis Drug Resistance from Venezuela

Background

Molecular typing of Mycobacterium tuberculosis strains has become a valuable tool in the epidemiology of tuberculosis (TB) by allowing detection of outbreaks, tracking of epidemics, identification of genotypes and transmission events among patients who would have remained undetected by conventional contact investigation. This is the first genetic biodiversity study of M. tuberculosis in Venezuela. Thus, we investigated the genetic patterns of strains isolated in the first survey of anti-tuberculosis drug-resistance realised as part of the Global Project of Anti-tuberculosis Drug Resistance Surveillance (WHO/IUATLD).

Results

Clinical isolates (670/873) were genotyped by spoligotyping. The results were compared with the international spoligotyping database (SpolDB4). Multidrug resistant (MDR) strains (14/18) were also analysed by IS6110-RFLP assays, and resistance to isoniazid and rifampicin was characterised. Spoligotyping grouped 82% (548/670) of the strains into 59 clusters. Twenty new spoligotypes (SITs) specific to Venezuela were identified. Eight new inter-regional clusters were created. The Beijing genotype was not found. The genetic network shows that the Latin American and Mediterranean family constitutes the backbone of the genetic TB population-structure in Venezuela, responsible of >60% of total TB cases studied. MDR was 0.5% in never treated patients and 13.5% in previously treated patients. Mutations in rpoB gene and katG genes were detected in 64% and 43% of the MDR strains, respectively. Two clusters were found to be identical by the four different analysis methods, presumably representing cases of recent transmission of MDR tuberculosis.

Conclusion

This study gives a first overview of the M. tuberculosis strains circulating in Venezuela during the first survey of anti-tuberculosis drug-resistance. It may aid in the creation of a national database that will be a valuable support for further studies.     

Interleukin-15 and interferon-gamma participate in the cross-talk between natural killer and monocytic cells required for tumour necrosis factor production

We have characterized the lymphocyte subset and the receptor molecules involved in inducing the secretion of TNF by monocytic cells in vitro. The TNF secreted by monocytic cells was measured when they were co-cultured with either resting or IL-15-stimulated lymphocytes, T cells, B cells or natural killer (NK) cells isolated from the peripheral blood of healthy subjects and from the synovial fluid from patients with inflammatory arthropathies. Co-culture with IL-15-activated peripheral blood or synovial fluid lymphocytes induced TNF production by monocytic cells within 24 hours, an effect that was mainly mediated by NK cells. In turn, monocytic cells induced CD69 expression and IFN-gamma production in NK cells, an effect that was mediated mainly by beta2 integrins and membrane-bound IL-15. Furthermore, IFN-gamma increased the production of membrane-bound IL-15 in monocytic cells. Blockade of beta2 integrins and membrane-bound IL-15 inhibited TNF production, whereas TNF synthesis increased in the presence of anti-CD48 and anti-CD244 (2B4) monoclonal antibodies. All these findings suggest that the cross-talk between NK cells and monocytes results in the sustained stimulation of TNF production. This phenomenon might be important in the pathogenesis of conditions such as rheumatoid arthritis in which the synthesis of TNF is enhanced.     

Anti-Trypanosoma activity of some natural stilbenoids and synthetic related heterocyclic compounds

We report the anti-Chagasic activity of the natural dihydrostilbenoid isonotholaenic acid and several simple derivatives, as well as that of some representative compounds of related synthetic series, with basic structures of benzalphthalides, dihydrostilbamides, isoindoles, phthalazin-1-ones, imidazo[2,1-a]isoindoles and pyrimido[2,1-a]isoindoles. The evaluation was performed in vitro on cultures of epimastigote and trypomastigote forms of Trypanosoma cruzi. Some of the tested compounds resulted to be as potent as benznidazole (epimastigotes), and others were shown to be more active than gentian violet (trypomastigotes), used as reference drugs.     

A distinct pathway remodels mitochondrial cristae and mobilizes cytochrome c during apoptosis.

The mechanism during apoptosis by which cytochrome c is rapidly and completely released in the absence of mitochondrial swelling is uncertain. Here, we show that two distinct pathways are involved. One mediates release of cytochrome c across the outer mitochondrial membrane, and another, characterized in this study, is responsible for the redistribution of cytochrome c stored in intramitochondrial cristae. We have found that the "BH3-only" molecule tBID induces a striking remodeling of mitochondrial structure with mobilization of the cytochrome c stores (approximately 85%) in cristae. This reorganization does not require tBID's BH3 domain and is independent of BAK, but is inhibited by CsA. During this process, individual cristae become fused and the junctions between the cristae and the intermembrane space are opened.     

Research resource: Diagnostic and therapeutic potential of nuclear receptor expression in lung cancer

Lung cancer is the leading cause of cancer-related death. Despite a number of studies that have provided prognostic biomarkers for lung cancer, a paucity of reliable markers and therapeutic targets exist to diagnose and treat this aggressive disease. In this study we investigated the potential of nuclear receptors (NRs), many of which are well-established drug targets, as therapeutic markers in lung cancer. Using quantitative real-time PCR, we analyzed the expression of the 48 members of the NR superfamily in a human panel of 55 normal and lung cancer cell lines. Unsupervised cluster analysis of the NR expression profile segregated normal from tumor cell lines and grouped lung cancers according to type (i.e. small vs. non-small cell lung cancers). Moreover, we found that the NR signature was 79% accurate in diagnosing lung cancer incidence in smokers (n = 129). Finally, the evaluation of a subset of NRs (androgen receptor, estrogen receptor, vitamin D receptor, and peroxisome proliferator-activated receptor-γ) demonstrated the therapeutic potential of using NR expression to predict ligand-dependent growth responses in individual lung cancer cells. Preclinical evaluation of one of these receptors (peroxisome proliferator activated receptor-γ) in mouse xenografts confirmed that ligand-dependent inhibitory growth responses in lung cancer can be predicted based on a tumor's receptor expression status. Taken together, this study establishes NRs as theragnostic markers for predicting lung cancer incidence and further strengthens their potential as therapeutic targets for individualized treatment.     

Research on arbuscular mycorrhizae in Mexico: an historical synthesis and future prospects

This review analyzes the historical development and advances of the research on arbuscular mycorrhizal fungi (AMF) in Mexico, as well as the prospects for future research. AMF-research has been focused on studying both diversity and functionality in several ecosystems of Mexico, but mainly in the tropical dry and rainy ecosystems, and the agricultural systems. In Mexico, 95 species of AMF have been recorded, representing 41% of the known species worldwide. The functional effects of AMF colonization have been examined in approximately 10% of the known host plants, but greenhouse studies continue to dominate over those conducted under field conditions. Even though research to date has been at the organismic level, further effort is needed due to the high plant diversity in Mexico. Studies on AMF biomass under field conditions and more taxonomic determination are required based on morphological features, biochemical determinations (fatty acids) and molecular tools. In addition, ecophysiological and ecological in situ studies would help in understanding the relationships among AMF, soil fauna, nutrients, and host plants. The contribution of AMF to ecosystemic processes is a priority line of research that requires an integrated approach (inter- and multidisciplinary) in order to define the role of AM symbioses for biogeochemical models. The creation of a Mexican mycorrhizal research network has and will help to identify the main challenges. Generating similar research protocols, and sharing databases and experience will assist mycorrhizologists working under the diverse financial and ecological contexts that is to be found in Mexico and Latin America.     

The extinction context enables extinction performance after a change in context

One experiment with human participants determined the extent to which recovery of extinguished responding with a context switch was due to a failure to retrieve contextually controlled learning, or some other process such as participants learning that context changes signal reversals in the meaning of stimulus-outcome relationships. In a video game, participants learned to suppress mouse clicking in the presence of a stimulus that predicted an attack. Then, that stimulus underwent extinction in a different context (environment within the game). Following extinction, suppression was recovered and then extinguished again during testing in the conditioning context. In a final test, participants that were tested in the context where extinction first took place showed less of a recovery than those tested in a neutral context, but they showed a recovery of suppression nevertheless. A change in context tended to cause a change in the meaning of the stimulus, leading to recovery in both the neutral and extinction contexts. The extinction context attenuated that recovery, perhaps by enabling retrieval of the learning that took place in extinction. Recovery outside an extinction context is due to a failure of the context to enable the learning acquired during extinction, but only in part.     

Non-invasive prenatal diagnosis of single-gene disorders from maternal blood

Prenatal diagnosis (PD) is available for pregnancies at risk of monogenic disorders. However, PD requires the use of invasive obstetric techniques for fetal-sample collection and therefore, involves a risk of fetal loss. Circulating fetal DNA in the maternal bloodstream is being used to perform non-invasive prenatal diagnosis (NIPD). NIPD is a challenging discipline because of the biological features of the maternal blood sample. Maternal blood is an unequal mixture of small (and fragmented) amounts of fetal DNA within a wide background of maternal DNA. For this reason, initial NIPD studies have been based on the analysis of specific paternally inherited fetal tracts not present in the maternal genome so as to ensure their fetal origin. Following this strategy, different NIPD studies have been carried out, such as fetal-sex assessment for pregnancies at risk of X-linked disorders, RhD determination, and analysis of single-gene disorders with a paternal origin. The study of the paternal mutation can be used for fetal diagnosis of dominant disorders or to more accurately assess the risk of an affected child in case of recessive diseases. Huntington's disease, cystic fibrosis, or achondroplasia are some examples of diseases studied using NIPD. New technologies are opening NIPD to the analysis of maternally inherited fetal tracts. NIPD of trisomy 21 is the latest study derived from the use of next-generation sequencing (NGS).