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1.
FEMS Microbiol Ecol ; 98(4)2022 Apr 21.
Artigo em Inglês | MEDLINE | ID: mdl-35348674

RESUMO

Perchloroethene (PCE) is a widely used chlorinated solvent. PCE is toxic to humans and has been identified as an environmental contaminant at thousands of sites worldwide. Several Dehalococcoides mccartyi strains can transform PCE to ethene, and thus contribute to bioremediation of contaminated sites. Humic acids (HA) are ubiquitous redox-active compounds of natural aquatic and soil systems and have been intensively studied because of their effect in electron transfer. In this study, we observed the dechlorination of PCE was accelerated by HA in mixed cultures containing Dehalococcoides strains. Anthraquinone-2,6-disulfonic acid (AQDS), a humic acid analogue, inhibited PCE dechlorination in our cultures and thus induced an opposite effect on PCE dehalogenation than HA. We observed the same effect on PCE dechlorination with the pure culture of Dehalococcoides mccartyi strain CBDB1. Not only in mixed cultures but also in pure cultures, growth of Dehalococcoides was not influenced by HA but inhibited by AQDS. Enzymatic activity tests confirmed the dehalogenating activity of strain CBDB1 was increased by HA, especially when using hydrogen as electron donor. We conclude that HA enhanced PCE dechlorination by increasing the reaction speed between hydrogen and the dehalogenase enzyme rather than acting as electron shuttle through its quinone moieties.


Assuntos
Chloroflexi , Biodegradação Ambiental , Chloroflexi/química , Chloroflexi/metabolismo , Dehalococcoides/química , Dehalococcoides/metabolismo , Humanos , Substâncias Húmicas , Hidrogênio
2.
mBio ; 13(2): e0028722, 2022 Apr 26.
Artigo em Inglês | MEDLINE | ID: mdl-35229635

RESUMO

"Candidatus Thermofonsia" represents a novel class within the phylum Chloroflexi. Metagenomic analysis reveals "Ca. Thermofonsia" harbors phototrophs outside the classically phototrophic Chloroflexia class. Unfortunately, the paucity of pure cultures limits further insights into their potential phototrophy. Here, we report the successful isolation of a "Ca. Thermofonsia" representative (Phototrophicus methaneseepsis ZRK33) from a deep-sea cold seep. Using combined physiological, genomic, and transcriptomic methods, we further show the long-wavelength light (e.g., red and infrared light) could promote the growth of strain ZRK33 and upregulate the expression of genes associated with phototrophy. In particular, strain ZRK33 has a typical phototrophic lifestyle under both laboratory and deep-sea conditions. Strain ZRK33 also possesses the ability to fix inorganic carbon through the 3-hydroxypropionate bicycle in both laboratory and deep-sea in situ environments, and the combined autotrophic, phototrophic, and heterotrophic capabilities endow strain ZRK33 with a photomixotrophic lifestyle. Notably, the predicted genes associated with phototrophy broadly exist in the metagenomes of 27 deep-sea Chloroflexi members, strongly suggesting diverse phototrophic Chloroflexi members are distributed in various unexplored deep biospheres. IMPORTANCE The deep ocean microbiota represents the unexplored majority of global ocean waters. The phylum Chloroflexi is abundant and broadly distributed in various deep-sea ecosystems. It was reported that some members of "Candidatus Thermofonsia" clade 2 might possess phototrophs; however, the absence of cultured representatives is a significant bottleneck toward understanding their phototrophic characteristics. In the present study, we successfully isolated a representative of the novel class "Ca. Thermofonsia" from a deep-sea cold seep by using an enrichment medium constantly supplemented with rifampicin, allowing researchers to isolate more Chloroflexi members in the future. Importantly, outside the classically phototrophic Chloroflexia class, we discover a novel phototrophic clade within the phylum Chloroflexi and demonstrate the existence of phototrophic lifestyles in the deep sea. Thus, this study expands the range of phototrophic Chloroflexi and provides a good model to study the mechanism of phototrophy performed in the deep biosphere.


Assuntos
Chloroflexi , Carbono , Chloroflexi/genética , Ecossistema , Metagenoma , Metagenômica
3.
Environ Sci Technol ; 56(7): 4039-4049, 2022 04 05.
Artigo em Inglês | MEDLINE | ID: mdl-35298122

RESUMO

Polybrominated diphenyl ethers (PBDEs) and polychlorinated biphenyls (PCBs) are notorious persistent organic pollutants. However, few organohalide-respiring bacteria that harbor reductive dehalogenases (RDases) capable of dehalogenating these pollutants have been identified. Here, we report reductive dehalogenation of penta-BDEs and PCBs byDehalococcoides mccartyi strain MB. The PCE-pregrown cultures of strain MB debrominated 86.6 ± 7.4% penta-BDEs to di- to tetra-BDEs within 5 days. Similarly, extensive dechlorination of Aroclor1260 and Aroclor1254 was observed in the PCE-pregrown cultures of strain MB, with the average chlorine per PCB decreasing from 6.40 ± 0.02 and 5.40 ± 0.03 to 5.98 ± 0.11 and 5.19 ± 0.07 within 14 days, respectively; para-substituents were preferentially dechlorinated from PCBs. Moreover, strain MB showed distinct enantioselective dechlorination of different chiral PCB congeners. Dehalogenation activity and cell growth were maintained during the successive transfer of cultures when amended with penta-BDEs as the sole electron acceptors but not when amended with only PCBs, suggesting metabolic and co-metabolic dehalogenation of these compounds, respectively. Transcriptional analysis, proteomic profiling, and in vitro activity assays indicated that MbrA was involved in dehalogenating PCE, PCBs, and PBDEs. Interestingly, resequencing of mbrA in strain MB identified three nonsynonymous mutations within the nucleotide sequence, although the consequences of which remain unknown. The substrate versatility of MbrA enabled strain MB to dechlorinate PCBs in the presence of either penta-BDEs or PCE, suggesting that co-metabolic dehalogenation initiated by multifunctional RDases may contribute to PCB attenuation at sites contaminated with multiple organohalide pollutants.


Assuntos
Chloroflexi , Bifenil Polibromatos , Bifenilos Policlorados , Biodegradação Ambiental , Catálise , Chloroflexi/genética , Chloroflexi/metabolismo , Dehalococcoides , Éteres Difenil Halogenados/metabolismo , Bifenil Polibromatos/metabolismo , Bifenilos Policlorados/metabolismo , Proteômica
4.
Water Sci Technol ; 85(5): 1335-1350, 2022 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-35290215

RESUMO

Complete dechlorination of trichloroethene (TCE) by Dehalococcoides mccartyi is catalyzed by reductive dehalogenases (RDases), which possess cobalamin as the crucial cofactor. However, virtually all D. mccartyi isolated thus far are corrinoid auxotrophs. The exogenous addition of commercially available cobalamin for TCE-contaminated site decontamination is costly. In this study, TCE reduction by a D. mccartyi-containing microbial consortium utilizing biosynthetic cobalamin generated by interior corrinoid-producing organisms within this microbial consortium was studied. The results confirmed that subcultures without exogenous cobalamin in the medium were apparently unaffected and were able to successively metabolize TCE to nonchlorinated ethene. The 2-bromoethanesulfonate and ampicillin resistance tests results suggested that ampicillin-sensitive bacteria rather than methanogenic archaea within this microbial consortium were responsible for biosynthesizing cobalamin. Moreover, relatively stable carbon isotopic enrichment factor (ɛ-carbon) values of TCE were obtained regardless of whether exogenous cobalamin or selective inhibitors existed in the medium, indicating that the cobalamin biosynthesized by these organisms was absorbed and utilized by D. mccartyi for RDase synthesis and eventually participated in TCE reduction. Finally, the Illumina MiSeq sequencing analysis indicated that Desulfitobacterium and Acetobacterium in this microbial consortium were responsible for the de novo cobalamin biosynthesis to fulfill the requirements of D. mccartyi for TCE metabolism.


Assuntos
Chloroflexi , Tricloroetileno , Biodegradação Ambiental , Chloroflexi/genética , Chloroflexi/metabolismo , Dehalococcoides , Vitamina B 12/metabolismo
5.
Environ Sci Technol ; 56(6): 3430-3440, 2022 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-35239320

RESUMO

Chlorinated ethanes, including 1,2-dichloroethane (1,2-DCA) and 1,1,2-trichloroethane (1,1,2-TCA), are widespread groundwater contaminants. Enrichment cultures XRDCA and XRTCA derived from river sediment dihaloeliminated 1,2-DCA to ethene and 1,1,2-TCA to vinyl chloride (VC), respectively. The XRTCA culture subsequently converted VC to ethene via hydrogenolysis. Microbial community profiling demonstrated the enrichment of Geobacter 16S rRNA gene sequences in both the XRDCA and XRTCA cultures, and Dehalococcoides mccartyi (Dhc) sequences were only detected in the ethene-producing XRTCA culture. The presence of a novel Geobacter population, designated as Geobacter sp. strain IAE, was identified by the 16S rRNA gene-targeted polymerase chain reaction and Sanger sequencing. Time-resolved population dynamics attributed the dihaloelimination activity to strain IAE, which attained the growth yields of 0.93 ± 0.06 × 107 and 1.18 ± 0.14 × 107 cells per µmol Cl- released with 1,2-DCA and 1,1,2-TCA as electron acceptors, respectively. In contrast, Dhc growth only occurred during VC-to-ethene hydrogenolysis. Our findings discover a Geobacter sp. strain capable of respiring multiple chlorinated ethanes and demonstrate the involvement of a broader diversity of organohalide-respiring bacteria in the detoxification of 1,2-DCA and 1,1,2-TCA.


Assuntos
Chloroflexi , Geobacter , Cloreto de Vinil , Poluentes Químicos da Água , Biodegradação Ambiental , Chloroflexi/genética , Dicloretos de Etileno , RNA Ribossômico 16S/genética , Tricloroetanos
6.
J Biol Chem ; 298(3): 101656, 2022 03.
Artigo em Inglês | MEDLINE | ID: mdl-35124004

RESUMO

N-demethylases have been reported to remove the methyl groups on primary or secondary amines, which could further affect the properties and functions of biomacromolecules or chemical compounds; however, the substrate scope and the robustness of N-demethylases have not been systematically investigated. Here we report the recreation of natural evolution in key microdomains of the Thermomicrobium roseum sarcosine oxidase (TrSOX), an N-demethylase with marked stability (melting temperature over 100 °C) and enantioselectivity, for enhanced substrate scope and catalytic efficiency on -C-N- bonds. We obtained the structure of TrSOX by crystallization and X-ray diffraction (XRD) for the initial framework. The natural evolution in the nonconserved residues of key microdomains-including the catalytic loop, coenzyme pocket, substrate pocket, and entrance site-was then identified using ancestral sequence reconstruction (ASR), and the substitutions that accrued during natural evolution were recreated by site-directed mutagenesis. The single and double substitution variants catalyzed the N-demethylation of N-methyl-L-amino acids up to 1800- and 6000-fold faster than the wild type, respectively. Additionally, these single substitution variants catalyzed the terminal N-demethylation of non-amino-acid compounds and the oxidation of the main chain -C-N- bond to a -C=N- bond in the nitrogen-containing heterocycle. Notably, these variants retained the enantioselectivity and stability of the initial framework. We conclude that the variants of TrSOX are of great potential use in N-methyl enantiomer resolution, main-chain Schiff base synthesis, and alkaloid modification or degradation.


Assuntos
Chloroflexi , Oxirredutases N-Desmetilantes , Aminoácidos/química , Aminoácidos/genética , Aminoácidos/metabolismo , Catálise , Chloroflexi/enzimologia , Chloroflexi/genética , Mutagênese Sítio-Dirigida , Oxirredutases N-Desmetilantes/genética , Oxirredutases N-Desmetilantes/metabolismo , Engenharia de Proteínas , Especificidade por Substrato
7.
Microbiologyopen ; 11(1): e1258, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-35212484

RESUMO

Denitrification plays a central role in the global nitrogen cycle, reducing and removing nitrogen from marine and terrestrial ecosystems. The flux of nitrogen species through this pathway has a widespread impact, affecting ecological carrying capacity, agriculture, and climate. Nitrite reductase (Nir) and nitric oxide reductase (NOR) are the two central enzymes in this pathway. Here we present a previously unreported Nir domain architecture in members of phylum Chloroflexi. Phylogenetic analyses of protein domains within Nir indicate that an ancestral horizontal transfer and fusion event produced this chimeric domain architecture. We also identify an expanded genomic diversity of a rarely reported NOR subtype, eNOR. Together, these results suggest a greater diversity of denitrification enzyme arrangements exist than have been previously reported.


Assuntos
Chloroflexi/metabolismo , Nitrito Redutases/química , Oxirredutases/química , Chloroflexi/classificação , Chloroflexi/enzimologia , Chloroflexi/genética , Desnitrificação , Variação Genética , Nitrito Redutases/genética , Nitrito Redutases/metabolismo , Oxirredutases/genética , Oxirredutases/metabolismo , Filogenia
8.
Enzyme Microb Technol ; 156: 109989, 2022 May.
Artigo em Inglês | MEDLINE | ID: mdl-35134708

RESUMO

Thermomicrobium roseum DSM 5159 lipase (TrLip) is an enzyme with marked thermostability and excellent solvent resistance. However, TrLip reveals relatively high catalytic efficiency on short-chain substrates but poor activity against mid-long or long-chain fatty acids, which would limit its industrial application. In this study, ancestral sequence reconstruction (ASR), a common engineering tool for the evolutionary history of protein families, was employed to identify the natural evolutionary trends within 5 Å around the catalytic center. Two mutation libraries were constructed, one for the catalytic center and the other for the pocket flexibility. A total of 69 mutants were expressed and purified in the Escherichia coli expression system to determine the kinetic parameters, and W219G could significantly enhance the catalytic efficiency against substrates with 12-, 16- and 18-carbon side chains. In addition, the double mutant W219G/F265M could further catalyze the breakdown of the above three substrates up to 6.34-, 4.21- and 4.86-folds compared to the wild-type TrLip, while the initial pH and thermostability were maintained. Through bioinformatics analysis, the significantly enhanced catalytic efficiency against longer-side chain substrates should be associated with the reduction of steric hindrance. With the outstanding stability and the promoted activity, TrLip should be of great potential in chemical and food industry.


Assuntos
Chloroflexi , Lipase , Catálise , Chloroflexi/enzimologia , Chloroflexi/genética , Estabilidade Enzimática , Lipase/metabolismo , Especificidade por Substrato
9.
Environ Res ; 209: 112888, 2022 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-35143804

RESUMO

Chloroflexi members are ubiquitous and have been extensively studied; however, the evolution and metabolic pathways of Chloroflexi members have long been debated. In the present study, the evolution and the metabolic potentials of 17 newly obtained Chloroflexi metagenome-assembled genomes (MAGs) were evaluated using genome and horizontal gene transfer (HGT) analysis. Taxonomic analysis suggests that the MAGs of the present study might be novel. One MAG encodes genes for anoxygenic phototrophy. The HGT analysis suggest that genes responsible for anoxygenic phototrophy in the MAG might have been transferred from Proteobacteria/Chlorobi. The evolution of anaerobic photosynthesis, which has long been questioned, has now been shown to be the result of HGT events. An incomplete Wood-Ljungdahl pathway (with missing genes metF, acsE, fdh, and acsA) was reported in Dehalococcoidetes members. In the present study, MAGs that were not the Dehalococcoidetes members encode genes acsA, acsB, metF and acsE. The genes responsible for sulfate reduction (sat, cysC and sir), dissimilatory sulfite reductase (dsrA and dsrB), and aerobic and anaerobic carbon monoxide oxidation (coxSML and cooSF) were detected in the present study MAGs. The present study expands our knowledge of the possible metabolic potentials of the phylum Chloroflexi and clarifies the evolution of anaerobic photosynthesis.


Assuntos
Chloroflexi , Chloroflexi/genética , Chloroflexi/metabolismo , Redes e Vias Metabólicas , Metagenoma , Metagenômica , Filogenia
10.
J Hazard Mater ; 430: 128355, 2022 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-35149497

RESUMO

Chloroethenes are common groundwater pollutants, and have been classified as toxic and carcinogenic to humans. The metabolites of chloroethenes, cis-dichloroethene (cis-DCE) and vinyl chloride (VC) commonly accumulate in groundwater due to their recalcitrant reductive dechlorination under anaerobic conditions. Dehalococcoides mccartyi (Dhc) is the key anaerobic bacteria for complete dechlorination of chloroethene, and Clostridium butyricum (C. butyricum) can provide hydrogen for supporting the growth of Dhc. In this study, we co-immobilized Dhc strain BAV1 and C. butyricum in a silica gel to determine the ability of the complete dechlorination of cis-DCE. Our results showed that our immobilized system could protect BAV1 from a high concentration (8 mM) of cis-DCE to carry out complete dechlorination. After the long-term use of our immobilized system, the activity of complete dechlorination was maintained for more than 180 consecutive days. Furthermore, we applied the immobilized system to remediate contaminated groundwater and uncovered the complete dechlorination of cis-DCE into ethene, a non-toxic product, within 28 days. Therefore, this novel co-immobilized system could serve a solution for bioremediation at chloroethene-contaminated sites.


Assuntos
Chloroflexi , Clostridium butyricum , Tricloroetileno , Biodegradação Ambiental , Chloroflexi/metabolismo , Clostridium butyricum/metabolismo , Dehalococcoides , Etilenos , Humanos , Sílica Gel , Tricloroetileno/metabolismo
11.
Environ Sci Technol ; 56(5): 3065-3075, 2022 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-35187933

RESUMO

Marine sediments are a major sink of organohalide pollutants, but the potential for offshore marine microbiota to transform these pollutants remains underexplored. Here, we report dehalogenation of diverse organohalide pollutants by offshore marine microbiota. Dechlorination of polychlorinated biphenyls (PCBs) was observed in four marine sediment microcosms, which was positively correlated with in situ PCB contamination. Three distinct enrichment cultures were enriched from these PCB-dechlorinating microcosms using tetrachloroethene (PCE) as the sole organohalide. All enrichment cultures also dehalogenated polybrominated diphenyl ethers (PBDEs), tetrabromobisphenol A (TBBPA), and 2,4,6-trichlorophenol (2,4,6-TCP). Particularly, two enrichments completely debrominated penta-BDEs, the first observation of complete debromination of penta-BDEs in marine cultures. Multiple Dehalococcoides and uncultivated Dehalococcoidia were identified in the initial sediment microcosms, but only Dehalococcoides was dominant in all enrichments. Transcription of a gene encoding a PcbA5-like reductive dehalogenase (RDase) was observed during dehalogenation of different organohalides in each enrichment culture. When induced by a single organohalide substrate, the PcbA5-like RDase dehalogenated all tested organohalides (PCE, PCBs, PBDEs, TBBPA, and 2,4,6-TCP) in in vitro tests, suggesting its involvement in dehalogenation of structurally distinct organohalides. Our results demonstrate the versatile dehalogenation capacity of marine Dehalococcoidia and contribute to a better understanding of the fate of these pollutants in marine systems.


Assuntos
Chloroflexi , Poluentes Ambientais , Microbiota , Bifenilos Policlorados , Biodegradação Ambiental , Sedimentos Geológicos , Éteres Difenil Halogenados
12.
Environ Res ; 209: 112801, 2022 06.
Artigo em Inglês | MEDLINE | ID: mdl-35093309

RESUMO

Bioelectrochemical dechlorination using organohalide-respiring bacteria (ORBs) is a promising technique for remediating contaminated groundwater. Generally, a longer enrichment period is required for selecting the ORB consortia to achieve bioelectrochemical dechlorination. However, the full dechloriantion is difficult to be achieved due to the absence of functional species (e.g. Dehalococcoides) in previously used enrich cultures. To overcome these challenges, bioelectrochemical dechlorination using a culture enriched with the pre-augmented Dehalococcoides was performed for the first time in this study. A two-chamber bioelectrochemical system (BES) inoculated with a pure Dehalococcoides culture and paddy soil with an applied voltage of -0.3 V (versus a standard hydrogen electrode) as the sole electron donor was used to achieve dechlorination. The ethene formation rate was 10-100 times higher than that in previous studies, indicating that inoculating the system with a pure Dehalococcoides culture and soil microorganisms gave effective full dechlorination performance. Microbial community analysis and bioelectrochemical analysis indicated that Desulfosporosinus species may have facilitated dechlorination through syntrophic interactions with Dehalococcoides. The results indicated that adding Dehalococcoides cells before operating a bioelectrochemical system is an effective way of achieving full dechlorination.


Assuntos
Chloroflexi , Tricloroetileno , Biodegradação Ambiental , Dehalococcoides , Eletrodos , Etilenos , Solo , Tricloroetileno/química
13.
Environ Sci Technol ; 56(2): 907-916, 2022 01 18.
Artigo em Inglês | MEDLINE | ID: mdl-34978445

RESUMO

The microbial transformation potential of 6:2 chlorinated polyfluorooctane ether sulfonate (6:2 Cl-PFESA) was explored in anaerobic microbial systems. Microbial communities from anaerobic wastewater sludge, an anaerobic digester, and anaerobic dechlorinating cultures enriched from aquifer materials reductively dechlorinated 6:2 Cl-PFESA to 6:2 hydrogen-substituted polyfluorooctane ether sulfonate (6:2 H-PFESA), which was identified as the sole metabolite by non-target analysis. Rapid and complete reductive dechlorination of 6:2 Cl-PFESA was achieved by the anaerobic dechlorinating cultures. The microbial community of the anaerobic dechlorinating cultures was impacted by 6:2 Cl-PFESA exposure. Organohalide-respiring bacteria originally present in the anaerobic dechlorinating cultures, including Geobacter, Dehalobacter, and Dehalococcoides, decreased in relative abundance over time. As the relative abundance of organohalide-respiring bacteria decreased, the rates of 6:2 Cl-PFESA dechlorination decreased, suggesting that the most likely mechanism for reductive dechlorination of 6:2 Cl-PFESA was co-metabolism rather than organohalide respiration. Reductive defluorination of 6:2 Cl-PFESA was not observed. Furthermore, 6:2 H-PFESA exhibited 5.5 times lower sorption affinity to the suspended biosolids than 6:2 Cl-PFESA, with the prospect of increased mobility in the environment. These results show the susceptibility of 6:2 Cl-PFESA to microbially mediated reductive dechlorination and the likely persistence of the product, 6:2 H-PFESA, in anaerobic environments.


Assuntos
Chloroflexi , Anaerobiose , Biodegradação Ambiental , Chloroflexi/metabolismo , Éter/metabolismo , Éteres/metabolismo
14.
Environ Microbiol ; 24(1): 110-121, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34984789

RESUMO

The recently discovered rhodopsin family of heliorhodopsins (HeRs) is abundant in diverse microbial environments. So far, the functional and biological roles of HeRs remain unknown. To tackle this issue, we combined experimental and computational screens to gain some novel insights. Here, 10 readily expressed HeR genes were found using functional metagenomics on samples from two freshwater environments. These HeRs originated from diverse prokaryotic groups: Actinobacteria, Chloroflexi and Archaea. Heterologously expressed HeRs absorbed light in the green and yellow wavelengths (543-562 nm) and their photocycles exhibited diverse kinetic characteristics. To approach the physiological function of the HeRs, we used our environmental clones along with thousands of microbial genomes to analyze genes neighbouring HeRs. The strongest association was found with the DegV family involved in activation of fatty acids, which allowed us to hypothesize that HeRs might be involved in light-induced membrane lipid modifications.


Assuntos
Actinobacteria , Chloroflexi , Actinobacteria/genética , Archaea/genética , Água Doce , Metagenômica , Rodopsinas Microbianas
15.
FEMS Microbiol Ecol ; 97(12)2022 01 11.
Artigo em Inglês | MEDLINE | ID: mdl-34978329

RESUMO

Core samples from bioretention cell media as well as surface stormwater sediment samples from seven urban areas were collected to assess the potential for biotransformation activity of polychlorinated biphenyls (PCBs). The presence of putative organohalide-respiring bacteria in these samples was studied. Based on extracted DNA, Dehalobacter, Dehalogenimonas and Dehalococcoides were detected. Other organohalide-respiring bacteria like Desulfitobacterium and Sulfurospirillum were not studied. Bacteria containing the genes encoding for biphenyl 2,3-dioxygenase (bphA) or 2,3-dihydroxybiphenyl 1,2-dioxygenase (bphC) were detected in 29 of the 32 samples. These genes are key factors in PCB aerobic degradation. Transcribed bacterial genes from putative organohalide-respiring bacteria as well as genes encoding for bphA and bphC were obtained from the microbial community, thus showing the potential of organohalide respiration of PCBs and aerobic PCB degradation under both aerobic and anaerobic conditions in the surface samples collected at the bioretention site. Presence and concentrations of 209 PCB congeners in the bioretention media were also assessed. The total PCB concentration ranged from 38.4 ± 2.3 ng/g at the top layer of the inlet to 11.6 ± 1.2 ng/g at 20-30 cm at 3 m from the inlet. These results provide documentation that bacteria capable of PCB transformation, including both anaerobic dechlorination and aerobic degradation, were present and active in the bioretention.


Assuntos
Chloroflexi , Bifenilos Policlorados , Bactérias/genética , Biodegradação Ambiental , Biotransformação , Chloroflexi/genética , Sedimentos Geológicos , RNA Ribossômico 16S
16.
Environ Res ; 207: 112150, 2022 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-34619124

RESUMO

This study presents the isolation of a novel strain of Dehalococcoides mccartyi, NIT01, which can completely dechlorinate up to 4.0 mM of trichloroethene to ethene via 1,2-cis-dichroroethene and vinyl chloride within 25 days. Strain NIT01 dechlorinated chloroethenes (CEs) at a temperature range of 25-32 °C and pH range of 6.5-7.8. The activity of the strain was inhibited by salt at more than 1.3% and inactivated by 1 h exposure to 2.0% air or 0.5 ppm hypochlorous acid. The genome of NIT01 was highly similar to that of the Dehalococcoides strains DCMB5, GT, 11a5, CBDB1, and CG5, and all included identical 16S rRNA genes. Moreover, NIT01 had 19 rdhA genes including NIT01-rdhA7 and rdhA13, which are almost identical to vcrA and pceA that encode known dehalogenases for tetrachloroethene and vinyl chloride, respectively. We also extracted RdhAs from the membrane fraction of NIT01 using 0.5% n-dodecyl-ß-d-maltoside and separated them by anion exchange chromatography to identify those involved in CE dechlorination. LC/MS identification of the LDS-PAGE bands and RdhA activities in the fractions indicated cellular expression of six RdhAs. NIT01-RdhA7 (VcrA) and NIT01-RdhA15 were highly detected and NIT01-RdhA6 was the third-most detected. Among these three RdhAs, NIT01-RdhA15 and NIT01-RdhA6 had no biochemically identified relatives and were suggested to be novel functional dehalogenases for CEs. The expression of multiple dehalogenases may support bacterial tolerance to high concentrations of CEs.


Assuntos
Chloroflexi , Tricloroetileno , Cloreto de Vinil , Biodegradação Ambiental , Chloroflexi/genética , Chloroflexi/metabolismo , Dehalococcoides , RNA Ribossômico 16S/genética , Tricloroetileno/metabolismo , Cloreto de Vinil/química , Cloreto de Vinil/metabolismo
17.
J Environ Manage ; 303: 114145, 2022 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-34844052

RESUMO

Hydrogen-releasing substrates can stimulate the reductive dechlorination of trichloroethene (TCE) mediated by organohalide-respiring bacteria (OHRB) at contaminated sites. However, how the substrate affects microbiome assembly and the accompanying influences on the growth of OHRB and reductive TCE dechlorination remains unclear. We evaluated the effects of microbial community structures and potential functions on the reductive dechlorination of TCE in three anaerobic reactors with acetate, soybean oil, or molasses as the substrate and no cobalamin or amino acid supplementation. The molasses-fed reactor exhibited superior performance and dechlorination of TCE loadings to ethene, and the oil-fed reactor exhibited a high growth rate of the key OHRB, Dehalococcoides. This finding suggests an effect of the substrate on reductive dechlorination and the growth of Dehalococcoides. The three reactors developed distinct microbial community structures and the predicted metagenomes were distinguished on the basis of vitamin and amino acid metabolisms as well as fermentation pathways. In addition to the diversified hydrogen-producing pathways, the molasses-induced microbiome exhibited high potential to synthesize the cobalamin, which may account for its high Dehalococcoides activity and thus effective dechlorination performance. The substrate dependence of microbiomes may provide insight into strategies of exogenous amino acid supplementation to benefit Dehalococcoides growth. This study adds novel insight into the interplay of hydrogen-releasing substrates and OHRB. The results may contribute to the development of tailored and cost-effective management for the reductive dechlorination of chlorinated solvents in bioremediation.


Assuntos
Chloroflexi , Microbiota , Tricloroetileno , Biodegradação Ambiental , Chloroflexi/genética , Fermentação
18.
Appl Environ Microbiol ; 88(2): e0190621, 2022 01 25.
Artigo em Inglês | MEDLINE | ID: mdl-34788060

RESUMO

Dehalococcoides mccartyi (Dhc) and Dehalogenimonas spp. (Dhgm) are members of the class Dehalococcoidia, phylum Chloroflexi, characterized by streamlined genomes and a strict requirement for organohalogens as electron acceptors. Here, we used cryo-electron tomography to reveal morphological and ultrastructural features of Dhc strain BAV1 and "Candidatus Dehalogenimonas etheniformans" strain GP cells at unprecedented resolution. Dhc cells were irregularly shaped discs (890 ± 110 nm long, 630 ± 110 nm wide, and 130 ± 15 nm thick) with curved and straight sides that intersected at acute angles, whereas Dhgm cells appeared as slightly flattened cocci (760 ± 85 nm). The cell envelopes were composed of a cytoplasmic membrane (CM), a paracrystalline surface layer (S-layer) with hexagonal symmetry and ∼22-nm spacing between repeating units, and a layer of unknown composition separating the CM and the S-layer. Cell surface appendages were only detected in Dhc cells, whereas both cell types had bundled cytoskeletal filaments. Repetitive globular structures, ∼5 nm in diameter and ∼9 nm apart, were observed associated with the outer leaflet of the CM. We hypothesized that those represent organohalide respiration (OHR) complexes and estimated ∼30,000 copies per cell. In Dhgm cultures, extracellular lipid vesicles (20 to 110 nm in diameter) decorated with putative OHR complexes but lacking an S-layer were observed. The new findings expand our understanding of the unique cellular ultrastructure and biology of organohalide-respiring Dehalococcoidia. IMPORTANCE Dehalococcoidia respire organohalogen compounds and play relevant roles in bioremediation of groundwater, sediments, and soils impacted with toxic chlorinated pollutants. Using advanced imaging tools, we have obtained three-dimensional images at macromolecular resolution of whole Dehalococcoidia cells, revealing their unique structural components. Our data detail the overall cellular shape, cell envelope architecture, cytoskeletal filaments, the likely localization of enzymatic complexes involved in reductive dehalogenation, and the structure of extracellular vesicles. The new findings expand our understanding of the cell structure-function relationship in Dehalococcoidia with implications for Dehalococcoidia biology and bioremediation.


Assuntos
Chloroflexi , Água Subterrânea , Biodegradação Ambiental , Chloroflexi/metabolismo , Tomografia com Microscopia Eletrônica
19.
Appl Environ Microbiol ; 88(4): e0218121, 2022 02 22.
Artigo em Inglês | MEDLINE | ID: mdl-34910572

RESUMO

As a group, the genus Dehalococcoides dehalogenates a wide range of organohalide pollutants, but the range of organohalide compounds that can be utilized for reductive dehalogenation differs among Dehalococcoides strains. Dehalococcoides lineages cannot be reliably disambiguated in mixed communities using typical phylogenetic markers, which often confounds bioremediation efforts. Here, we describe a computational approach to identify Dehalococcoides genetic markers with improved discriminatory resolution. Screening core genes from the Dehalococcoides pangenome for degree of similarity and frequency of 100% identity found a candidate genetic marker encoding a bacterial neuraminidase repeat (BNR)-containing protein of unknown function. This gene exhibits the fewest completely identical amino acid sequences and has among the lowest average amino acid sequence identity in the core pangenome. Primers targeting BNR could effectively discriminate between 40 available BNR sequences (in silico) and 10 different Dehalococcoides isolates (in vitro). Amplicon sequencing of BNR fragments generated from 22 subsurface soil samples revealed a total of 109 amplicon sequence variants, suggesting a high diversity of Dehalococcoides distributed in the environment. Therefore, the BNR gene can serve as an alternative genetic marker to differentiate strains of Dehalococcoides in complicated microbial communities. IMPORTANCE The challenge of discriminating between phylogenetically similar but functionally distinct bacterial lineages is particularly relevant to the development of technologies seeking to exploit the metabolic or physiological characteristics of specific members of bacterial genera. A computational approach was developed to expedite screening of potential genetic markers among phylogenetically affiliated bacteria. Using this approach, a gene encoding a bacterial neuraminidase repeat (BNR)-containing protein of unknown function was selected and evaluated as a genetic marker to differentiate strains of Dehalococcoides, an environmentally relevant genus of bacteria whose members can transform and detoxify a range of halogenated organic solvents and persistent organic pollutants, in complex microbial communities to demonstrate the validity of the approach. Moreover, many apparently phylogenetically distinct, currently uncharacterized Dehalococcoides were detected in environmental samples derived from contaminated sites.


Assuntos
Chloroflexi , Biodegradação Ambiental , Chloroflexi/metabolismo , Dehalococcoides , Marcadores Genéticos , Filogenia
20.
PLoS One ; 16(11): e0259515, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34762689

RESUMO

Soil salinity is a serious environmental issue in arid China. Halophytes show extreme salt tolerance and are grow in saline-alkaline environments. There rhizosphere have complex bacterial communities, which mediate a variety of interactions between plants and soil. High-throughput sequencing was used to investigated rhizosphere bacterial community changes under the typical halophyte species in arid China. Three typical halophytes were Leymus chinensis (LC), Puccinellia tenuiflora (PT), Suaeda glauca (SG). The dominant phyla were Proteobacteria, Actinobacteria, Chloroflexi, Gemmatimonadetes, Acidobacteria and Bacteroidetes, Suaeda glauca rhizosphere has stronger enrichment of Nitrospirae and Cyanobacteria. The Ace, Chao and Shannon indices were significantly higher in soils under LC and SG (P<0.05). Functional predictions, based on 16S rRNA gene by PICRUSt, indicated that Energy metabolism, Amino acid metabolism, Carbohydrate metabolism and Fatty acid metabolism are dominant bacterial functions in three halophytes rhizosphere soil. Carbon metabolism, Oxidative phosphorylation, Methane metabolism, Sulfur metabolism and Nitrogen metabolism in SG were significantly higher than that in LC and PT. Regression analysis revealed that rhizosphere soil bacterial community structure is influenced by soil organic matter (SOM) and soil water content (SWC), while soil bacterial community diversity is affected by soil pH. This study contributes to our understanding of the distribution characteristics and metabolic functions under different halophyte rhizosphere bacterial communities, and will provide references for the use of rhizosphere bacteria to regulate the growth of halophytes and ecological restoration of saline soil.


Assuntos
Plantas Tolerantes a Sal/metabolismo , Solo/química , Acidobacteria/metabolismo , Actinobacteria/metabolismo , Bacteroidetes/metabolismo , Chenopodiaceae/metabolismo , China , Chloroflexi/metabolismo , Cianobactérias/metabolismo , Sequenciamento de Nucleotídeos em Larga Escala , Poaceae/metabolismo , Proteobactérias/metabolismo , Rizosfera , Salinidade , Tolerância ao Sal , Plantas Tolerantes a Sal/genética
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