Serological Evaluation of Mycobacterium ulcerans Antigens Identified by Comparative Genomics

A specific and sensitive serodiagnostic test for Mycobacterium ulcerans infection would greatly assist the diagnosis of Buruli ulcer and would also facilitate seroepidemiological surveys. By comparative genomics, we identified 45 potential M. ulcerans specific proteins, of which we were able to express and purify 33 in E. coli. Sera from 30 confirmed Buruli ulcer patients, 24 healthy controls from the same endemic region and 30 healthy controls from a non-endemic region in Benin were screened for antibody responses to these specific proteins by ELISA. Serum IgG responses of Buruli ulcer patients were highly variable, however, seven proteins (MUP045, MUP057, MUL_0513, Hsp65, and the polyketide synthase domains ER, AT propionate, and KR A) showed a significant difference between patient and non-endemic control antibody responses. However, when sera from the healthy control subjects living in the same Buruli ulcer endemic area as the patients were examined, none of the proteins were able to discriminate between these two groups. Nevertheless, six of the seven proteins showed an ability to distinguish people living in an endemic area from those in a non-endemic area with an average sensitivity of 69% and specificity of 88%, suggesting exposure to M. ulcerans. Further validation of these six proteins is now underway to assess their suitability for use in Buruli ulcer seroepidemiological studies. Such studies are urgently needed to assist efforts to uncover environmental reservoirs and understand transmission pathways of the M. ulcerans

Comparative Genomics of Cell Envelope Components in Mycobacteria

Mycobacterial cell envelope components have been a major focus of research due to their unique features that confer intrinsic resistance to antibiotics and chemicals apart from serving as a low-permeability barrier. The complex lipids secreted by Mycobacteria are known to evoke/repress host-immune response and thus contribute to its pathogenicity. This study focuses on the comparative genomics of the biosynthetic machinery of cell wall components across 21-mycobacterial genomes available in GenBank release 179.0. An insight into survival in varied environments could be attributed to its variation in the biosynthetic machinery. Gene-specific motifs like ‘DLLAQPTPAW’ of ufaA1 gene, novel functional linkages such as involvement of Rv0227c in mycolate biosynthesis; Rv2613c in LAM biosynthesis and Rv1209 in arabinogalactan peptidoglycan biosynthesis were detected in this study. These predictions correlate well with the available mutant and coexpression data from TBDB. It also helped to arrive at a minimal functional gene set for these biosynthetic pathways that complements findings using TraSH

Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)1.

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field

Meat and Nicotinamide:A Causal Role in Human Evolution, History, and Demographics

Hunting for meat was a critical step in all animal and human evolution. A key brain-trophic element in meat is vitamin B 3 /nicotinamide. The supply of meat and nicotinamide steadily increased from the Cambrian origin of animal predators ratcheting ever larger brains. This culminated in the 3-million-year evolution of Homo sapiens and our overall demographic success. We view human evolution, recent history, and agricultural and demographic transitions in the light of meat and nicotinamide intake. A biochemical and immunological switch is highlighted that affects fertility in the ‘de novo’ tryptophan-to-kynurenine-nicotinamide ‘immune tolerance’ pathway. Longevity relates to nicotinamide adenine dinucleotide consumer pathways. High meat intake correlates with moderate fertility, high intelligence, good health, and longevity with consequent population stability, whereas low meat/high cereal intake (short of starvation) correlates with high fertility, disease, and population booms and busts. Too high a meat intake and fertility falls below replacement levels. Reducing variances in meat consumption might help stabilise population growth and improve human capital

Comparative analysis of Mycobacterium tuberculosis pe and ppe genes reveals high sequence variation and an apparent absence of selective constraints.

Contains fulltext : 110619.pdf (publisher's version ) (Open Access)Mycobacterium tuberculosis complex (MTBC) genomes contain 2 large gene families termed pe and ppe. The function of pe/ppe proteins remains enigmatic but studies suggest that they are secreted or cell surface associated and are involved in bacterial virulence. Previous studies have also shown that some pe/ppe genes are polymorphic, a finding that suggests involvement in antigenic variation. Using comparative sequence analysis of 18 publicly available MTBC whole genome sequences, we have performed alignments of 33 pe (excluding pe_pgrs) and 66 ppe genes in order to detect the frequency and nature of genetic variation. This work has been supplemented by whole gene sequencing of 14 pe/ppe (including 5 pe_pgrs) genes in a cohort of 40 diverse and well defined clinical isolates covering all the main lineages of the M. tuberculosis phylogenetic tree. We show that nsSNP's in pe (excluding pgrs) and ppe genes are 3.0 and 3.3 times higher than in non-pe/ppe genes respectively and that numerous other mutation types are also present at a high frequency. It has previously been shown that non-pe/ppe M. tuberculosis genes display a remarkably low level of purifying selection. Here, we also show that compared to these genes those of the pe/ppe families show a further reduction of selection pressure that suggests neutral evolution. This is inconsistent with the positive selection pressure of "classical" antigenic variation. Finally, by analyzing such a large number of genes we were able to detect large differences in mutation type and frequency between both individual genes and gene sub-families. The high variation rates and absence of selective constraints provides valuable insights into potential pe/ppe function. Since pe/ppe proteins are highly antigenic and have been studied as potential vaccine components these results should also prove informative for aspects of M. tuberculosis vaccine design