A soil bacterial consortium that was grown on diesel fuel and consisted of more than 10 members from different genera was maintained through repetitive subculturing and was utilized as a practical ...model to investigate a bacterial community that was continuously exposed to petroleum hydrocarbons. Through metagenomics analyses, consortium member isolation, growth assays, and metabolite identification which supported the linkage of genomic data and functionality, two pioneering genera,
and
, whose catabolic capabilities were differentiated, were found to be responsible for the creation of specialized ecological niches that were apparently occupied by other bacterial members for survival within the consortium. Coexisting genera
and
maintained their existence in the consortium through metabolic dependencies by utilizing hydrocarbon biotransformation products of pioneer metabolism, which was confirmed through growth tests and identification of biotransformation products of the isolated strains. Pioneering
and
s spp. utilized relatively water-insoluble hydrocarbon parent compounds and facilitated the development of a consortium community structure that resulted in the creation of niches in response to diesel fuel exposure which were created through the production of more-water-soluble biotransformation products available to cocolonizers. That these and other organisms were still present in the consortium after multiple transfers spanning 15 years provided evidence for these ecological niches. Member survival through occupation of these niches led to robustness of each group within the multispecies bacterial community. Overall, these results contribute to our understanding of the complex ecological relationships that may evolve during prokaryotic hydrocarbon pollutant biodegradation.
There are few metagenome studies that have explored soil consortia maintained on a complex hydrocarbon substrate after the community interrelationships were formed. A soil bacterial consortium maintained on diesel fuel was utilized as a practical model to investigate bacterial community relationships through metagenomics analyses, consortium member isolation, growth assays, and metabolite identification, which supported the linkage of genomic data and functionality. Two pioneering genera were responsible for the biodegradation of aromatics and alkanes by initiating biotransformation and thereby created specialized niches that were populated by other members. A model that represents these relationships was constructed, which contributes to our understanding of the complex ecological relationships that evolve during prokaryotic hydrocarbon pollutant biodegradation.
Interest in the biodegradation mechanisms and environmental fate of polycyclic aromatic hydrocarbons (PAHs) is prompted by their ubiquitous distribution and their potentially deleterious effects on ...human health. The bacterial biodegradation of PAHs with more than three rings is discussed.
In many bacteria, cyclic diguanosine monophosphate (c-di-GMP), synthesized by diguanylate cyclase (DGC), serves as a second messenger involved in the regulation of biofilm formation. Although studies ...have suggested that c-di-GMP also regulates the formation of electrochemically active biofilms (EABFs) by
MR-1, DGCs involved in this process remained to be identified. Here, we report that the SO_1646 gene, hereafter named
, is upregulated under medium flow conditions in electrochemical flow cells (EFCs), and its product (DgcS) functions as a major DGC in MR-1.
assays demonstrated that purified DgcS catalyzed the synthesis of c-di-GMP from GTP. Comparisons of intracellular c-di-GMP levels in the wild-type strain and a
deletion mutant (Δ
mutant) showed that production of c-di-GMP was markedly reduced in the Δ
mutant when cells were grown in batch cultures and on electrodes in EFCs. Cultivation of the Δ
mutant in EFCs also revealed that the loss of DgcS resulted in impaired biofilm formation and decreased current generation. These findings demonstrate that MR-1 uses DgcS to synthesize c-di-GMP under medium flow conditions, thereby activating biofilm formation on electrodes.
Bioelectrochemical systems (BESs) have attracted wide attention owing to their utility in sustainable biotechnology processes, such as microbial fuel cells and electrofermentation systems. In BESs, electrochemically active bacteria (EAB) form biofilms on electrode surfaces, thereby serving as effective catalysts for the interconversion between chemical and electric energy. It is therefore important to understand mechanisms for the formation of biofilm by EAB grown on electrodes. Here, we show that a model EAB,
MR-1, expresses DgcS as a major DGC, thereby activating the formation of biofilms on electrodes via c-di-GMP-dependent signal transduction cascades. The findings presented herein provide the molecular basis for improving electrochemical interactions between EAB and electrodes in BESs. The results also offer molecular insights into how
regulates biofilm formation on solid surfaces in the natural environment.
► Shewanella oneidensis MR-1 reduced tellurite to elemental tellurium. ► Crystal Te(0) nanorods accumulated intracellularly. ► Te(IV) caused changes in cell membrane rigidity by shifting the ...compositions of lipid components. ► Shewanella may provide an alternative method for Te(0) nanorod preparation.
The dissimilatory metal-reducing bacterium, Shewanella oneidensis MR-1, reduced tellurite (Te(IV), TeO32−) to elemental tellurium under anaerobic conditions resulting in the intracellular accumulation of needle shaped crystalline Te(0) nanorods. Fatty acid analyses showed that toxic Te(IV) increased the unsaturated fatty acid composition of the lipid components of the cell membrane, implying a deconstruction of the integrity of the cellular membrane structure. The current results suggest that dissimilatory metal reducing bacteria such as S. oneidensis MR-1 may play an important role in recycling toxic tellurium elements, and may be applied as a novel selective biological filter via the accumulation of industry-applicable rare materials, Te(0) nanorods, in the cell.
Understanding the biotransformation mechanisms of toxic sulfur-containing polycyclic aromatic hydrocarbon (PASH) pollutants such as benzothiophene (BT) is useful for predicting their environmental ...fates. In the natural environment, nondesulfurizing hydrocarbon-degrading bacteria are major active contributors to PASH biodegradation at petroleum-contaminated sites; however, BT biotransformation pathways by this group of bacteria are less explored when compared to desulfurizing organisms. When a model nondesulfurizing polycyclic aromatic hydrocarbon-degrading soil bacterium,
Sphingobium barthaii
KK22, was investigated for its ability to cometabolically biotransform BT by quantitative and qualitative methods, BT was depleted from culture media but was biotransformed into mostly high molar mass (HMM) hetero and homodimeric
ortho
-substituted diaryl disulfides (diaryl disulfanes). HMM diaryl disulfides have not been reported as biotransformation products of BT. Chemical structures were proposed for the diaryl disulfides by comprehensive mass spectrometry analyses of the chromatographically separated products and were supported by the identification of transient upstream BT biotransformation products, which included benzenethiols. Thiophenic acid products were also identified, and pathways that described BT biotransformation and novel HMM diaryl disulfide formation were constructed. This work shows that nondesulfurizing hydrocarbon-degrading organisms produce HMM diaryl disulfides from low molar mass polyaromatic sulfur heterocycles, and this may be taken into consideration when predicting the environmental fates of BT pollutants.
Quinones may be formed metabolically or abiotically from environmental pollutants and polycyclic aromatic hydrocarbons (PAHs); many are recognized as toxicological intermediates that cause a variety ...of deleterious cellular effects including mutagenicity. The PAH-o-quinone, 1,2-naphthoquinone (1,2-NQ), may exert its genotoxic effects through interactions with cellular nucleophiles such as DNA, however, the mechanisms of 1,2-NQ adduct formation are still under investigation. With the aim to further understand these mechanisms, the chemical structures of adducts formed from the reaction of 2′-deoxyguanosine (dG) with 1,2-NQ under physiological conditions were investigated by liquid chromatography electrospray ionization tandem mass spectrometry and 1H NMR analyses. Results showed that 1,2-NQ underwent non-enzymatic oxidation to form a 1,2-NQ-epoxide which in turn formed at least four bulky adducts with dG, and these adducts were more likely to be formed under physiological conditions. A mechanism was proposed whereby hydration of 1,2-NQ to form unstable naphthohydroquinones and 2-hydroxy-1,4-naphthoquinone resulted in formation of hydrogen peroxide that oxidized 1,2-NQ. These results suggest that the genotoxicity of 1,2-NQ may not only be caused through oxidative DNA damage and adduct formation through Michael addition but also through non-enzymatic oxidative transformation of 1,2-NQ itself to form an intermediate PAH-epoxide which covalently binds to DNA.
Aromatic amines are a class of chemical carcinogens that are activated by cytochrome P450 enzymes to form arylhydroxylamines that are conjugated to form N-acetoxyarylamines or ...N-sulfonyloxyarylamines. These conjugates undergo N–O bond cleavage to become reactive nitrenium ions that may form DNA adducts. Numerous studies in the past using N-acetoxyarylamines to investigate DNA adduct formation were conducted, however, less is known in regard to DNA adduct formation directly from arylhydroxylamines - especially under conditions that mimic the physiological conditions of cells such as weakly basic conditions. In this study, 2′-deoxyguanosine (dG) was exposed to N-(2,6-dimethylphenyl)hydroxylamine (2,6-DMPHA) and N-phenylhydroxylamine (PHA) at pH 7.4 without enzymes and analyzed by liquid chromatography high resolution mass spectrometry (LC-HRMS). 2,6-DMPHA exposure resulted in the production of relatively low amounts of adducts however the identities of at least six different adducts that were formed through reactions with carbon, nitrogen and oxygen of 2′-deoxyguanosine were proposed based upon different analytical approaches including HRMS CID fragmentation and NMR analyses. Contrastively, PHA exposure under identical conditions resulted in one adduct at the C8 position. It was concluded from these results and results of theoretical calculations that nitrenium ions produced from 2,6-DMPHA were relatively more stable resulting in longer nitrenium ion lifetimes which ultimately led to greater potential for 2,6-DMPHA nitrenium ions to react with multiple sites on dG.
Display omitted
•2′-deoxyguanosine (dG) was exposed to arylhydroxylamines under weakly basic conditions.•Multiple dG adducts were formed from N-(2,6-dimethylphenyl)hydroxylamine without conjugation.•2,6-DMPHA adducts were identified by comprehensive LC-HRMS and NMR.•Nitrenium ion relative stability contributed to production of more adduct types from 2,6-DMPHA.
Summary
Interest in understanding prokaryotic biotransformation of high‐molecular‐weight polycyclic aromatic hydrocarbons (HMW PAHs) has continued to grow and the scientific literature shows that ...studies in this field are originating from research groups from many different locations throughout the world. In the last 10 years, research in regard to HMW PAH biodegradation by bacteria has been further advanced through the documentation of new isolates that represent diverse bacterial types that have been isolated from different environments and that possess different metabolic capabilities. This has occurred in addition to the continuation of in‐depth comprehensive characterizations of previously isolated organisms, such as Mycobacterium vanbaalenii PYR‐1. New metabolites derived from prokaryotic biodegradation of four‐ and five‐ring PAHs have been characterized, our knowledge of the enzymes involved in these transformations has been advanced and HMW PAH biodegradation pathways have been further developed, expanded upon and refined. At the same time, investigation of prokaryotic consortia has furthered our understanding of the capabilities of microorganisms functioning as communities during HMW PAH biodegradation.
In the development of new chemical substances, genetic toxicity evaluations are a high priority for safety risk management. Evaluation of the possibility of compound carcinogenicity with accuracy and ...at reasonable cost in the early stages of development by
techniques is preferred. Currently, DNA damage-related
genotoxicity tests are widely-used screening tools after which next generation toxicity testing may be applied to confirm DNA damage. DNA adductomics may be used to evaluate DNA damage
, however confirmation of DNA adduct identities through comparison to authentic standards may be time-consuming and expensive processes. Considering this, a streamlined method for confirming putative DNA adducts that are detected by DNA adductomics may be useful. With this aim,
DNA adductome methods in conjunction with
RNA adductome methods may be proposed as a DNA adductome verification approach by which to eliminate false positive annotations. Such an approach was evaluated by conducting
assays whereby Hep G2 cell lines that were exposed to or not exposed to benzo
pyrene were digested to their respective 2'-deoxynucleosides or ribonucleosides and analyzed by liquid chromatography electrospray ionization tandem mass spectrometry (LC/ESI-MS/MS) by comparative DNA and RNA adductomics through neutral loss targeting of the M + H
> M + H - 116
or M + H
> M + H -132
transitions over predetermined ranges. Comparisons of DNA adductome maps revealed putative DNA adducts that were detected in exposed cells but not in unexposed cells. Similarly, comparisons of RNA adductome maps revealed putative RNA adducts in exposed cells but not in unexposed cells. Taken together these experiments revealed that analogous forms of putative damage had occurred in both DNA and RNA which supported that putative DNA adducts detected by DNA adductomics were DNA adducts. High resolution mass spectrometry (HRMS) was utilized to confirm that putative nucleic acid adducts detected in both DNA and RNA were derived from benzo
pyrene exposure and these putative adducts were identified as 7,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo
pyrene- (B
PDE)-type adducts. Overall, this study demonstrates the usefulness of utilizing DNA/RNA adductomics to screen for nucleic acid damage.
Chromids (secondary chromosomes) in bacterial genomes that are present in addition to the main chromosome appear to be evolutionarily conserved in some specific bacterial groups. In rare cases among ...these groups, a small number of strains from
and
were shown to possess a naturally fused single chromosome that was reported to have been generated through intragenomic homologous recombination between repeated sequences on the chromosome and chromid. Similar examples have never been reported in the family
, a well-documented group that conserves chromids. Here, an in-depth genomic characterization was performed on a
bacterium that was isolated from a soil bacterial consortium maintained on diesel fuel and mutagenic benzo
pyrene. This organism, Cupriavidus necator strain KK10, was revealed to carry a single chromosome with unexpectedly large size (>6.6 Mb), and results of comparative genomics with the genome of C. necator N-1
indicated that the single chromosome of KK10 was generated through fusion of the prototypical chromosome and chromid at the rRNA operons. This fusion hypothetically occurred through homologous recombination with a crossover between repeated rRNA operons on the chromosome and chromid. Some metabolic functions that were likely expressed from genes on the prototypical chromid region were indicated to be retained. If this phenomenon-the bacterial chromosome-chromid fusion across the rRNA operons through homologous recombination-occurs universally in prokaryotes, the multiple rRNA operons in bacterial genomes may not only contribute to the robustness of ribosome function, but also provide more opportunities for genomic rearrangements through frequent recombination.
A bacterial chromosome that was naturally fused with the secondary chromosome, or "chromid," and presented as an unexpectedly large single replicon was discovered in the genome of Cupriavidus necator strain KK10, a biotechnologically useful member of the family
. Although
is a well-documented group that conserves chromids in their genomes, this chromosomal fusion event has not been previously reported for this family. This fusion has hypothetically occurred through intragenomic homologous recombination between repeated rRNA operons and, if so, provides novel insight into the potential of multiple rRNA operons in bacterial genomes to lead to chromosome-chromid fusion. The harsh conditions under which strain KK10 was maintained-a genotoxic hydrocarbon-enriched milieu-may have provided this genotype with a niche in which to survive.
PDF
