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Genome-resolved analysis of Serratia marcescens strain SMTT infers niche specialization as a hydrocarbon-degrader

Abstract Bacteria that are chronically exposed to high levels of pollutants demonstrate genomic and corresponding metabolic diversity that complement their strategies for adaptation to hydrocarbon-rich environments. Whole genome sequencing was carried out to infer functional traits of Serratia marcescens strain SMTT recovered from soil contaminated with crude oil. The genome size (Mb) was 5,013,981 with a total gene count of 4,842. Comparative analyses with carefully selected S. marcescens strains, 2 of which are associated with contaminated soil, show conservation of central metabolic pathways in addition to intra-specific genetic diversity and metabolic flexibility. Genome comparisons also indicated an enrichment of genes associated with multidrug resistance and efflux pumps for SMTT. The SMTT genome contained genes that enable the catabolism of aromatic compounds via the protocatechuate para-degradation pathway, in addition to meta-cleavage of catechol (meta-cleavage pathway II); gene enrichment for aromatic compound degradation was markedly higher for SMTT compared to the other S. marcescens strains analysed. Our data presents a valuable genetic inventory for future studies on strains of S. marcescens and provides insights into those genomic features of SMTT with industrial potential.

Genetics & Heredity

Xylanolytic metabolism is regulated by coordination of transcription factors XynR and XylR in extremely thermophilic Caldicellulosiruptorales

ABSTRACT Global transcription factors (TFs) control metabolic processes in bacteria to efficiently utilize available carbon. The orderCaldicellulosiruptoraleshas drawn interest due to the ability of its members to degrade components of lignocellulosic biomass. Regulatory reconstruction ofAnaerocellum (f. Caldicellulosiruptor) besciiidentified two major global transcription factors for xylan utilization, XynR and XylR, and the corresponding putative transcription factor binding sites. Recombinant versions of XynR (LacI family) and XylR (ROK family) were subjected to fluorescence polarization (FP) and biolayer interferometry (BLI) analysis to confirm the predicted binding sites. Four XynR sites and two XylR sites were validated, accounting for 20 of 26 genes regulated by XynR and six of seven genes regulated by XylR. Bioinformatic analysis of the individual genes controlled by the two regulators showed an inter-dependent scheme for xylan conversion; the transport of xylooligosaccharides (XOS) is dependent on XylR, while enzymes responsible for hydrolysis are controlled by both regulators. For xylose catabolism by the xylose isomerase-xylulose kinase pathway, regulation is also split, with XylR controlling xylose isomerase and XynR controlling xylokinase. The XynR/XylR regulator pair withinA. besciiis conserved in all sequenced species ofCaldicellulosiruptorales, suggesting similarities in regulating linear xylan conversion. In other xylanolytic thermophiles, XylR homologs control xylan degradation, compared to just 6 out of 26 genes forA. bescii. These results show that two separate regulatory schemes (dual repression) are coordinated byA. besciito effectively regulate the hemicellulose inventory and xylan catabolism. IMPORTANCE To take full advantage of extreme thermophiles as platform metabolic engineering microorganisms, the tools for genetic manipulation must be further developed, and strategies that exploit a better understanding of metabolic regulation need to be discerned.Anaerocellum bescii, the most studied of the extremely thermophilic fermentative anaerobic bacteria that can utilize microcrystalline cellulose, can degrade microcrystalline cellulose and hemicellulose and has been metabolically engineered to convert the resulting sugars to products such as ethanol and acetone. For xylan, in particular, two major global transcription factors (TFs), XynR and XylR, play a role in sugar metabolism, although their predicted regulatory interdependence from bioinformatics analysis has not been elucidated experimentally. Here, fluorescence polarization (FP) and biolayer interferometry (BLI) were used to explore this issue to support metabolic engineering efforts aimed at improving carbohydrate processing to industrial chemicals.

Biotechnology & Applied Microbiology

Implementation of fuel management multi-cycle optimization capabilities in RAVEN optimization framework

Optimization in nuclear fuel-management assists the core reload engineer with finding optimal out-of-core and in-core strategies. RAVEN is INL’s open source software that is equipped with fuel-management optimization capabilities including single-cycle, single- and multi-objective optimization of pressurized water reactors (PWRs) loading patterns (LP) of a fresh core using genetic algorithm (GA) and non-dominated sorting genetic algorithm (NSGA-II). In practice, however, medium and long term planning of fuel-management needs a multi-cycle approach, where the history and availability of fuel assemblies is considered in the optimization process. In this paper, we present a description of an initial expansion of RAVEN fuel-management optimization capabilities for a multi-cycle optimization framework. N-th cycle optimization capabilities that account for the unique history of recycled fuel assembly in the core were added. The multi-cycle optimization approach taken is formulated as a cycle-wise optimization problem where out-of-core decisions are used to onset each cycle optimization. Out-of-core decisions are managed externally to the in-core optimization by a fuel inventory management module. A proof-of-concept optimization problem is also presented.

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