BacillOndex: An Integrated Data Resource for Systems and Synthetic Biology

BacillOndex is an extension of the Ondex data integration system, providing a semantically annotated, integrated knowledge base for the model Gram-positive bacterium Bacillus subtilis. This application allows a user to mine a variety of B. subtilis data sources, and analyse the resulting integrated dataset, which contains data about genes, gene products and their interactions. The data can be analysed either manually, by browsing using Ondex, or computationally via a Web services interface. We describe the process of creating a BacillOndex instance, and describe the use of the system for the analysis of single nucleotide polymorphisms in B. subtilis Marburg. The Marburg strain is the progenitor of the widely-used laboratory strain B. subtilis 168. We identified 27 SNPs with predictable phenotypic effects, including genetic traits for known phenotypes. We conclude that BacillOndex is a valuable tool for the systems-level investigation of, and hypothesis generation about, this important biotechnology workhorse. Such understanding contributes to our ability to construct synthetic genetic circuits in this organism

Systematic whole-genome sequencing reveals an unexpected diversity among actinomycetoma pathogens and provides insights into their antibacterial susceptibilities

Mycetoma is a neglected tropical chronic granulomatous inflammatory disease of the skin and subcutaneous tissues. More than 70 species with a broad taxonomic diversity have been implicated as agents of mycetoma. Understanding the full range of causative organisms and their antibiotic sensitivity profiles are essential for the appropriate treatment of infections. The present study focuses on the analysis of full genome sequences and antibiotic inhibitory concentration profiles of actinomycetoma strains from patients seen at the Mycetoma Research Centre in Sudan with a view to developing rapid diagnostic tests. Seventeen pathogenic isolates obtained by surgical biopsies were sequenced using MinION and Illumina methods, and their antibiotic inhibitory concentration profiles determined. The results highlight an unexpected diversity of actinomycetoma causing pathogens, including three Streptomyces isolates assigned to species not previously associated with human actinomycetoma and one new Streptomyces species. Thus, current approaches for clinical and histopathological classification of mycetoma may need to be updated. The standard treatment for actinomycetoma is a combination of sulfamethoxazole/trimethoprim and amoxicillin/clavulanic acid. Most tested isolates had a high IC (inhibitory concentration) to sulfamethoxazole/trimethoprim or to amoxicillin alone. However, the addition of the β-lactamase inhibitor clavulanic acid to amoxicillin increased susceptibility, particularly for Streptomyces somaliensis and Streptomyces sudanensis. Actinomadura madurae isolates appear to have a particularly high IC under laboratory conditions, suggesting that alternative agents, such as amikacin, could be considered for more effective treatment. The results obtained will inform future diagnostic methods for the identification of actinomycetoma and treatment

Mirubactin C rescues the lethal effect of cell wall biosynthesis mutations in Bacillus subtilis

Growth of most rod-shaped bacteria is accompanied by the insertion of new peptidoglycan into the cylindrical cell wall. This insertion, which helps maintain and determine the shape of the cell, is guided by a protein machine called the rod complex or elongasome. Although most of the proteins in this complex are essential under normal growth conditions, cell viability can be rescued, for reasons that are not understood, by the presence of a high (mM) Mg(2+) concentration. We screened for natural product compounds that could rescue the growth of mutants affected in rod-complex function. By screening > 2,000 extracts from a diverse collection of actinobacteria, we identified a compound, mirubactin C, related to the known iron siderophore mirubactin A, which rescued growth in the low micromolar range, and this activity was confirmed using synthetic mirubactin C. The compound also displayed toxicity at higher concentrations, and this effect appears related to iron homeostasis. However, several lines of evidence suggest that the mirubactin C rescuing activity is not due simply to iron sequestration. The results support an emerging view that the functions of bacterial siderophores extend well beyond simply iron binding and uptake

Diverse control of metabolism and other cellular processes in Streptomyces coelicolor by the PhoP transcription factor: genome-wide identification of in vivo targets.

Streptomycetes sense and respond to the stress of phosphate starvation via the two-component PhoR-PhoP signal transduction system. To identify the in vivo targets of PhoP we have undertaken a chromatin-immunoprecipitation-on-microarray analysis of wild-type and phoP mutant cultures and, in parallel, have quantified their transcriptomes. Most (ca. 80%) of the previously in vitro characterized PhoP targets were identified in this study among several hundred other putative novel PhoP targets. In addition to activating genes for phosphate scavenging systems PhoP was shown to target two gene clusters for cell wall/extracellular polymer biosynthesis. Furthermore PhoP was found to repress an unprecedented range of pathways upon entering phosphate limitation including nitrogen assimilation, oxidative phosphorylation, nucleotide biosynthesis and glycogen catabolism. Moreover, PhoP was shown to target many key genes involved in antibiotic production and morphological differentiation, including afsS, atrA, bldA, bldC, bldD, bldK, bldM, cdaR, cdgA, cdgB and scbR-scbA. Intriguingly, in the PhoP-dependent cpk polyketide gene cluster, PhoP accumulates substantially at three specific sites within the giant polyketide synthase-encoding genes. This study suggests that, following phosphate limitation, Streptomyces coelicolor PhoP functions as a 'master' regulator, suppressing central metabolism, secondary metabolism and developmental pathways until sufficient phosphate is salvaged to support further growth and, ultimately, morphological development