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July 7, 2019

A novel aerobic degradation pathway of thiobencarb is initiated by a two-component FMN-dependent monooxygenase system TmoAB in Acidovorax sp. T1.

Thiobencarb is a thiocarbamate herbicide used in rice paddies worldwide. Microbial degradation plays a crucial role in the dissipation of thiobencarb in the environment. However, the physiological and genetic mechanisms underlying thiobencarb degradation remain unknown. In this study, a novel thiobencarb degradation pathway was proposed in Acidovorax sp. T1. Thiobencarb was oxidized and cleaved at the C-S bond, generating diethylcarbamothioic S-acid and 4-chlorobenzaldehyde (4CDA). 4CDA was then oxidized to 4-chlorobenzoic acid (4CBA) and hydrolytically dechlorinated to 4-hydroxybenzoic acid (4HBA). The identification of catabolic genes suggested further hydroxylation to protocatechuic acid (PCA) and finally degradation through the protocatechuate 4,5-dioxygenase pathway. A novel two-component monooxygenase system identified in this strain, TmoAB, was responsible for the initial catabolic reaction. TmoA shared 28-32% identities with the oxygenase components of pyrimidine monooxygenase from Agrobacterium fabrum, alkanesulfonate monooxygenase from Pseudomonas savastanoi and dibenzothiophene monooxygenase from Rhodococcus sp.. TmoB shared 25-37% identities with reported flavin reductases and oxidized NADH but not NADPH. TmoAB was an FMN-dependent monooxygenase and catalyzed the C-S bond cleavage of thiobencarb. Introduction of tmoAB into cells of the thiobencarb degradation-deficient mutant T1m restored its ability to degrade and utilize thiobencarb. A dehydrogenase gene, tmoC, was located 7129 bp downstream of tmoAB, and its transcription was clearly induced by thiobencarb. The purified TmoC catalyzed the dehydrogenation of 4CDA to 4CBA using NAD(+) as a cofactor. A gene cluster responsible for complete 4CBA metabolic pathway was also cloned, and its involvement in thiobencarb degradation was preliminarily verified by transcriptional analysis.IMPORTANCE Microbial degradation is the main factor of thiobencarb dissipation in soil. In previous reports, thiobencarb was degraded initially via N-deethylation, sulfoxidation, hydroxylation and dechlorination. However, enzymes and genes involved in microbial degradation of thiobencarb have not been studied. This study revealed a new thiobencarb degradation pathway in strain Acidovorax sp. T1 and identified a novel two-component FMN-dependent monooxygenase system TmoAB. Under TmoAB-mediated catalysis, thiobencarb was cleaved at the C-S bond, producing diethylcarbamothioic S-acid and 4CDA. Furthermore, the downstream degradation pathway of thiobencarb was proposed. Our study provides the physiological, biochemical and genetic foundation of thiobencarb degradation in this microorganism. Copyright © 2017 American Society for Microbiology.

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July 7, 2019

Genome sequence of the white-rot fungus Irpex lacteus F17, a type strain of lignin degrader fungus.

Irpex lacteus, a cosmopolitan white-rot fungus, degrades lignin and lignin-derived aromatic compounds. In this study, we report the high-quality draft genome sequence of I. lacteus F17, isolated from a decaying hardwood tree in the vicinity of Hefei, China. The genome is 44,362,654 bp, with a GC content of 49.64% and a total of 10,391 predicted protein-coding genes. In addition, a total of 18 snRNA, 842 tRNA, 15 rRNA operons and 11,710 repetitive sequences were also identified. The genomic data provides insights into the mechanisms of the efficient lignin decomposition of this strain.

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July 7, 2019

Complete genome of Arthrobacter alpinus strain R3.8, bioremediation potential unraveled with genomic analysis.

Arthrobacter alpinus R3.8 is a psychrotolerant bacterial strain isolated from a soil sample obtained at Rothera Point, Adelaide Island, close to the Antarctic Peninsula. Strain R3.8 was sequenced in order to help discover potential cold active enzymes with biotechnological applications. Genome analysis identified various cold adaptation genes including some coding for anti-freeze proteins and cold-shock proteins, genes involved in bioremediation of xenobiotic compounds including naphthalene, and genes with chitinolytic and N-acetylglucosamine utilization properties and also plant-growth-influencing properties. In this genome report, we present a complete genome sequence of A. alpinus strain R3.8 and its annotation data, which will facilitate exploitation of potential novel cold-active enzymes.

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July 7, 2019

Evolution of sphingomonad gene clusters related to pesticide catabolism revealed by genome sequence and mobilomics of Sphingobium herbicidovorans MH.

Bacterial degraders of chlorophenoxy herbicides have been isolated from various ecosystems, including pristine environments. Among these degraders, the sphingomonads constitute a prominent group that displays versatile xenobiotic-degradation capabilities. Four separate sequencing strategies were required to provide the complete sequence of the complex and plastic genome of the canonical chlorophenoxy herbicide-degrading Sphingobium herbicidovorans MH. The genome has an intricate organization of the chlorophenoxy-herbicide catabolic genes sdpA, rdpA, and cadABCD that encode the (R)- and (S)-enantiomer-specific 2,4-dichlorophenoxypropionate dioxygenases and four subunits of a Rieske non-heme iron oxygenase involved in 2-methyl-chlorophenoxyacetic acid degradation, respectively. Several major genomic rearrangements are proposed to help understand the evolution and mobility of these important genes and their genetic context. Single-strain mobilomic sequence analysis uncovered plasmids and insertion sequence-associated circular intermediates in this environmentally important bacterium and enabled the description of evolutionary models for pesticide degradation in strain MH and related organisms. The mobilome presented a complex mosaic of mobile genetic elements including four plasmids and several circular intermediate DNA molecules of insertion-sequence elements and transposons that are central to the evolution of xenobiotics degradation. Furthermore, two individual chromosomally integrated prophages were shown to excise and form free circular DNA molecules. This approach holds great potential for improving the understanding of genome plasticity, evolution, and microbial ecology.© The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

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July 7, 2019

Adaptation of surface-associated bacteria to the open ocean: A genomically distinct subpopulation of Phaeobacter gallaeciensis Ccolonizes Pacific mesozooplankton.

The marine Roseobacter group encompasses numerous species which occupy a large variety of ecological niches. However, members of the genus Phaeobacter are specifically adapted to a surface-associated lifestyle and have so far been found nearly exclusively in disjunct, man-made environments including shellfish and fish aquacultures, as well as harbors. Therefore, the possible natural habitats, dispersal and evolution of Phaeobacter spp. have largely remained obscure. Applying a high-throughput cultivation strategy along a longitudinal Pacific transect, the present study revealed for the first time a widespread natural occurrence of Phaeobacter in the marine pelagial. These bacteria were found to be specifically associated to mesoplankton where they constitute a small but detectable proportion of the bacterial community. The 16S rRNA gene sequences of 18 isolated strains were identical to that of Phaeobacter gallaeciensis DSM26640(T) but sequences of internal transcribed spacer and selected genomes revealed that the strains form a distinct clade within P. gallaeciensis. The genomes of the Pacific and the aquaculture strains were highly conserved and had a fraction of the core genome of 89.6%, 80 synteny breakpoints, and differed 2.2% in their nucleotide sequences. Diversification likely occurred through neutral mutations. However, the Pacific strains exclusively contained two active Type I restriction modification systems which is commensurate with a reduced acquisition of mobile elements in the Pacific clade. The Pacific clade of P. gallaeciensis also acquired a second, homolog phosphonate transport system compared to all other P. gallaeciensis. Our data indicate that a previously unknown, distinct clade of P. gallaeciensis acquired a limited number of clade-specific genes that were relevant for its association with mesozooplankton and for colonization of the marine pelagial. The divergence of the Pacific clade most likely was driven by the adaptation to this novel ecological niche rather than by geographic isolation.

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July 7, 2019

Bioinformatics analysis and characterization of highly efficient polyvinyl alcohol (PVA)-degrading enzymes from the novel PVA degrader Stenotrophomonas rhizophila QL-P4.

Polyvinyl alcohol (PVA) is used widely in industry, and associated environmental pollution is a serious problem. Herein, we report a novel, efficient PVA degrader, Stenotrophomonas rhizophila QL-P4, isolated from fallen leaves from virgin forest in the Qinling Mountains. The complete genome was obtained using single-molecule real-time (SMRT) technology and corrected using Illumina sequencing. Bioinformatics analysis revealed eight PVA/OVA (vinyl alcohol oligomer)-degrading genes. Of these, seven genes were predicted to be involved in the classical intracellular PVA/OVA degradation pathway, and one (BAY15_3292) was identified as a novel PVA oxidase. Five PVA/OVA-degrading enzymes were purified and characterised. Among which, BAY15_1712, a PVA dehydrogenase (PVADH), displayed high catalytic efficiency towards PVA and OVA substrate. All reported PVADHs only have PVA-degrading ability. Most importantly, we discovered a novel PVA oxidase (BAY15_3292) that exhibited highest PVA-degrading efficiency than the reported PVADHs. Further investigation indicated that BAY15_3292 plays a crucial role in PVA degradation in S. rhizophila QL-P4. Knocking out BAY15_3292 resulted in a significant decline in PVA-degrading activity in S. rhizophila QL-P4. Interestingly, we found that BAY15_3292 possesses exocrine activity, which distinguishes it from classical PVADHs. Transparent circle experiments further proved that BAY15_3292 greatly affects extracellular PVA degradation in S. rhizophila QL-P4. The exocrine characteristics of BAY15_3292 facilitate its potential application to PVA bioremediation. In addition, we report three new efficient secondary alcohol dehydrogenases (SADHs) with OVA-degrading ability in S. rhizophila QL-P4, compared with only one OVA-degrading SADH as reported previously.Importance With the widespread application of PVA in industry, PVA-related environmental pollution is an increasingly serious issue. Because PVA is difficult to degrade, it accumulates in aquatic environments and causes chronic toxicity to aquatic organisms. Biodegradation of PVA, as an economical and environment-friendly method, has attracted much interest. To date, effective and applicable PVA-degrading bacteria/enzymes have not been reported. Herein, we report a new efficient PVA degrader (S. rhizophila QL-P4) that has five PVA/OVA-degrading enzymes with high catalytic efficiency, among which BAY15_1712 is the only reported PVADH with both PVA- and OVA-degrading abilities. Importantly, we discovered a novel PVA oxidase (BAY15_3292) that is not only more efficient than other reported PVA-degrading PVADHs, but also has exocrine activity. Overall, our findings provide new insight into PVA-degrading pathways in microorganisms, and suggest S. rhizophila QL-P4 and its enzymes have potential for application to PVA bioremediation to reduce or eliminate PVA-related environmental pollution. Copyright © 2017 American Society for Microbiology.

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July 7, 2019

Comparative and population genomic landscape of Phellinus noxius: A hypervariable fungus causing root rot in trees.

The order Hymenochaetales of white rot fungi contain some of the most aggressive wood decayers causing tree deaths around the world. Despite their ecological importance and the impact of diseases they cause, little is known about the evolution and transmission patterns of these pathogens. Here, we sequenced and undertook comparative genomic analyses of Hymenochaetales genomes using brown root rot fungus Phellinus noxius, wood-decomposing fungus Phellinus lamaensis, laminated root rot fungus Phellinus sulphurascens and trunk pathogen Porodaedalea pini. Many gene families of lignin-degrading enzymes were identified from these fungi, reflecting their ability as white rot fungi. Comparing against distant fungi highlighted the expansion of 1,3-beta-glucan synthases in P. noxius, which may account for its fast-growing attribute. We identified 13 linkage groups conserved within Agaricomycetes, suggesting the evolution of stable karyotypes. We determined that P. noxius has a bipolar heterothallic mating system, with unusual highly expanded ~60 kb A locus as a result of accumulating gene transposition. We investigated the population genomics of 60 P. noxius isolates across multiple islands of the Asia Pacific region. Whole-genome sequencing showed this multinucleate species contains abundant poly-allelic single nucleotide polymorphisms with atypical allele frequencies. Different patterns of intra-isolate polymorphism reflect mono-/heterokaryotic states which are both prevalent in nature. We have shown two genetically separated lineages with one spanning across many islands despite the geographical barriers. Both populations possess extraordinary genetic diversity and show contrasting evolutionary scenarios. These results provide a framework to further investigate the genetic basis underlying the fitness and virulence of white rot fungi.© 2017 John Wiley & Sons Ltd.

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July 7, 2019

Complete genome sequence analysis of Enterobacter sp. SA187, a plant multi-stress tolerance promoting endophytic bacterium

Enterobacter sp. SA187 is an endophytic bacterium that has been isolated from root nodules of the indigenous desert plant Indigofera argentea. SA187 could survive in the rhizosphere as well as in association with different plant species, and was able to provide abiotic stress tolerance to Arabidopsis thaliana. The genome sequence of SA187 was obtained by using Pacific BioScience (PacBio) single-molecule sequencing technology, with average coverage of 275X. The genome of SA187 consists of one single 4,429,597 bp chromosome, with an average 56% GC content and 4,347 predicted protein coding DNA sequences (CDS), 153 ncRNA, 7 rRNA, and 84 tRNA. Functional analysis of the SA187 genome revealed a large number of genes involved in uptake and exchange of nutrients, chemotaxis, mobilization and plant colonization. A high number of genes were also found to be involved in survival, defense against oxidative stress and production of antimicrobial compounds and toxins. Moreover, different metabolic pathways were identified that potentially contribute to plant growth promotion. The information encoded in the genome of SA187 reveals the characteristics of a dualistic lifestyle of a bacterium that can adapt to different environments and promote the growth of plants. This information provides a better understanding of the mechanisms involved in plant-microbe interaction and could be further exploited to develop SA187 as a biological agent to improve agricultural practices in marginal and arid lands.

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July 7, 2019

Isolation and complete genome sequence of Halorientalis hydrocarbonoclasticus sp. nov., a hydrocarbon-degrading haloarchaeon.

Bioremediation in hypersaline environments is particularly challenging since the microbes that tolerate such harsh environments and degrade pollutants are quite scarce. Haloarchaea, however, due to their inherent ability to grow at high salt concentrations, hold great promise for remediating the contaminated hypersaline sites. This study aimed to isolate and characterize novel haloarchaeal strains with potentials in hydrocarbon degradation. A haloarchaeal strain IM1011 was isolated from Changlu Tanggu saltern near Da Gang Oilfield in Tianjin (China) by enrichment culture in hypersaline medium containing hexadecane. It could degrade 57 ± 5.2% hexadecane (5 g/L) in the presence of 3.6 M NaCl at 37 °C within 24 days. To get further insights into the mechanisms of petroleum hydrocarbon degradation in haloarchaea, complete genome (3,778,989 bp) of IM1011 was sequenced. Phylogenetic analysis of 16S rRNA gene, RNA polymerase beta-subunit (rpoB’) gene and of the complete genome suggested IM1011 to be a new species in Halorientalis genus, and the name Halorientalis hydrocarbonoclasticus sp. nov., is proposed. Notably, with insights from the IM1011 genome sequence, the involvement of diverse alkane hydroxylase enzymes and an intact ß-oxidation pathway in hexadecane biodegradation was predicted. This is the first hexadecane-degrading strain from Halorientalis genus, of which the genome sequence information would be helpful for further dissecting the hydrocarbon degradation by haloarchaea and for their application in bioremediation of oil-polluted hypersaline environments.

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July 7, 2019

Scallop genome reveals molecular adaptations to semi-sessile life and neurotoxins.

Bivalve molluscs are descendants of an early-Cambrian lineage superbly adapted to benthic filter feeding. Adaptations in form and behavior are well recognized, but the underlying molecular mechanisms are largely unknown. Here, we investigate the genome, various transcriptomes, and proteomes of the scallop Chlamys farreri, a semi-sessile bivalve with well-developed adductor muscle, sophisticated eyes, and remarkable neurotoxin resistance. The scallop’s large striated muscle is energy-dynamic but not fully differentiated from smooth muscle. Its eyes are supported by highly diverse, intronless opsins expanded by retroposition for broadened spectral sensitivity. Rapid byssal secretion is enabled by a specialized foot and multiple proteins including expanded tyrosinases. The scallop uses hepatopancreas to accumulate neurotoxins and kidney to transform to high-toxicity forms through expanded sulfotransferases, probably as deterrence against predation, while it achieves neurotoxin resistance through point mutations in sodium channels. These findings suggest that expansion and mutation of those genes may have profound effects on scallop’s phenotype and adaptation.

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July 7, 2019

Comparative genomics reveals specific genetic architectures in nicotine metabolism of Pseudomonassp. JY-Q.

Microbial degradation of nicotine is an important process to control nicotine residues in the aqueous environment. In this study, a high active nicotine degradation strain namedPseudomonassp. JY-Q was isolated from tobacco waste extract (TWE). This strain could completely degrade 5.0 g l-1nicotine in 24 h under optimal culture conditions, and it showed some tolerance even at higher concentrations (10.0 g l-1) of nicotine. The complete genome of JY-Q was sequenced to understand the mechanism by which JY-Q could degrade nicotine and tolerate such high nicotine concentrations. Comparative genomic analysis indicated that JY-Q degrades nicotine through putative novel mechanisms. Two candidate gene cluster duplications located separately at distant loci were predicted to be responsible for nicotine degradation. These two nicotine (Nic) degradation-related loci (AA098_21325-AA098_21340, AA098_03885-AA098_03900) exhibit nearly completely consistent gene organization and component synteny. The nicotinic acid(NA)degradation gene cluster (AA098_17770-AA098_17790) andNic-like clusters were both predicted to be flanked by mobile genetic elements (MGE). Furthermore, we analyzed the regions of genomic plasticity (RGP) in the JY-Q strain and found a dynamic genome carrying a type VI secretion system (T6SS) that promotes nicotine metabolism and tolerance based on transcriptomics and usedin silicomethods to identify the T6SS effector protein. Thus, a novel nicotine degradation mechanism was elucidated forPseudomonassp. JY-Q, suggesting its potential application in the bioremediation of nicotine-contaminated environments, such as TWEs.

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July 7, 2019

Genetic and functional characterization of an extracellular modular GH6 endo-ß-1,4-glucanase from an earthworm symbiont, Cellulosimicrobium funkei HY-13.

The gene (1608-bp) encoding a GH6 endo-ß-1,4-glucanase (CelL) from the earthworm-symbiotic bacterium Cellulosimicrobium funkei HY-13 was cloned from its whole genome sequence, expressed recombinantly, and biochemically characterized. CelL (56.0 kDa) is a modular enzyme consisting of an N-terminal catalytic GH6 domain (from Val57 to Pro396), which is 71 % identical to a GH6 protein (accession no.: WP_034662937) from Cellulomonas sp. KRMCY2, together with a C-terminal CBM 2 domain (from Cys429 to Cys532). The highest catalytic activity of CelL toward carboxymethylcellulose (CMC) was observed at 50 °C and pH 5.0, and was relatively stable at a broad pH range of 4.0-10.0. The enzyme was capable of efficiently hydrolyzing the cellulosic polymers in the order of barley ß-1,3-1,4-D-glucan > CMC > lichenan > Avicel > konjac glucomannan. However, cellobiose, cellotriose, p-nitrophenyl derivatives of mono- and disaccharides, or structurally unrelated carbohydrate polymers including ß-1,3-D-glucan, ß-1,4-D-galactomannan, and ß-1,4-D-xylan were not susceptible to CelL. The enzymatic hydrolysis of cellopentaose resulted in the production of a mixture of 68.6 % cellobiose and 31.4 % cellotriose but barley ß-1,3-1,4-D-glucan was 100 % degraded to cellotriose by CelL. The enzyme strongly bound to Avicel, ivory nut mannan, and chitin but showed relatively weak binding affinity to lichenan, lignin, or poly(3-hydroxybutyrate) granules.

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July 7, 2019

Identification and resolution of microdiversity through metagenomic sequencing of parallel consortia.

To gain a predictive understanding of the interspecies interactions within microbial communities that govern community function, the genomic complement of every member population must be determined. Although metagenomic sequencing has enabled the de novo reconstruction of some microbial genomes from environmental communities, microdiversity confounds current genome reconstruction techniques. To overcome this issue, we performed short-read metagenomic sequencing on parallel consortia, defined as consortia cultivated under the same conditions from the same natural community with overlapping species composition. The differences in species abundance between the two consortia allowed reconstruction of near-complete (at an estimated >85% of gene complement) genome sequences for 17 of the 20 detected member species. Two Halomonas spp. indistinguishable by amplicon analysis were found to be present within the community. In addition, comparison of metagenomic reads against the consensus scaffolds revealed within-species variation for one of the Halomonas populations, one of the Rhodobacteraceae populations, and the Rhizobiales population. Genomic comparison of these representative instances of inter- and intraspecies microdiversity suggests differences in functional potential that may result in the expression of distinct roles in the community. In addition, isolation and complete genome sequence determination of six member species allowed an investigation into the sensitivity and specificity of genome reconstruction processes, demonstrating robustness across a wide range of sequence coverage (9× to 2,700×) within the metagenomic data set. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

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