Effects of microplastics/nanoplastics on Vallisneria natans roots and sediment: Size effect, enzymology, and microbial communities.

Microplastics/nanoplastics (MNPs) pollution in different environmental media and its adverse effects on organisms have received increasing attention from researchers. This paper compares the effects of natural concentrations of three different sizes (20 nm, 200 nm, and 2 µm) of MNPs on Vallisneria natans and sediments. MNPs with smaller sizes adhere more readily to V. natans roots, further promoting root elongation. In addition, the larger the particle size of MNPs, the higher the reactive oxygen species level in the roots, and the malondialdehyde level increased accordingly. In the sediment, 20 nm, and 200 nm MNPs increased the activity of related enzymes, including acid phosphatase, urease, and nitrate reductase. In addition, the dehydrogenase content in the treated sediments increased, and the content changes were positively correlated with the size of MNPs. Changes in microorganisms were only observed on the root surface. The addition of MNPs reduced the abundance of Proteobacteria and increased the abundance of Chloroflexi. In addition, at the class level of species composition on the root surface, the abundance of Gammaproteobacteria under the 20 nm, 200 nm, and 2 µm MNP treatments decreased by 21.19%, 16.14%, and 17.03%, respectively, compared with the control group, while the abundance of Anaerolineae increased by 44.63%, 26.31%, and 62.52%, respectively. These findings enhance the understanding of the size effects of MNPs on the roots of submerged plants and sediment.

Redox gradient shapes the abundance and diversity of mercury-methylating microorganisms along the water column of the Black Sea.

In the global context of seawater deoxygenation triggered by climate change and anthropogenic activities, changes in redox gradients impacting biogeochemical transformations of pollutants, such as mercury, become more likely. Being the largest anoxic basin worldwide, with high concentrations of the potent neurotoxic methylmercury (MeHg), the Black Sea is an ideal natural laboratory to provide new insights about the link between dissolved oxygen concentration and hgcAB gene-carrying (hgc+) microorganisms involved in the formation of MeHg. We combined geochemical and microbial approaches to assess the effect of vertical redox gradients on abundance, diversity, and metabolic potential of hgc+ microorganisms in the Black Sea water column. The abundance of hgcA genes [congruently estimated by quantitative PCR (qPCR) and metagenomics] correlated with MeHg concentration, both maximal in the upper part of the anoxic water. Besides the predominant Desulfobacterales, hgc+ microorganisms belonged to a unique assemblage of diverse-previously underappreciated-anaerobic fermenters from Anaerolineales, Phycisphaerae (characteristic of the anoxic and sulfidic zone), Kiritimatiellales, and Bacteroidales (characteristic of the suboxic zone). The metabolic versatility of Desulfobacterota differed from strict sulfate reduction in the anoxic water to reduction of various electron acceptors in the suboxic water. Linking microbial activity and contaminant concentration in environmental studies is rare due to the complexity of biological pathways. In this study, we disentangle the role of oxygen in shaping the distribution of Hg-methylating microorganisms consistently with MeHg concentration, and we highlight their taxonomic and metabolic niche partitioning across redox gradients, improving the prediction of the response of marine communities to the expansion of oxygen-deficient zones. IMPORTANCE Methylmercury (MeHg) is a neurotoxin detected at high concentrations in certain marine ecosystems, posing a threat to human health. MeHg production is mainly mediated by hgcAB gene-carrying (hgc+) microorganisms. Oxygen is one of the main factors controlling Hg methylation; however, its effect on the diversity and ecology of hgc+ microorganisms remains unknown. Under the current context of seawater deoxygenation, mercury cycling is expected to be disturbed. Here, we show the strong effect of oxygen gradients on the distribution of potential Hg methylators. In addition, we show for the first time the significant contribution of a unique assemblage of potential fermenters from Anaerolineales, Phycisphaerae, and Kiritimatiellales to Hg methylation, stratified in different redox niches along the Black Sea gradient. Our results considerably expand the known taxonomic diversity and ecological niches prone to the formation of MeHg and contribute to better apprehend the consequences of oxygen depletion in seawater.

Cultivation of marine bacteria of the SAR202 clade.

Bacteria of the SAR202 clade, within the phylum Chloroflexota, are ubiquitously distributed in the ocean but have not yet been cultivated in the lab. It has been proposed that ancient expansions of catabolic enzyme paralogs broadened the spectrum of organic compounds that SAR202 bacteria could oxidize, leading to transformations of the Earth's carbon cycle. Here, we report the successful cultivation of SAR202 bacteria from surface seawater using dilution-to-extinction culturing. The growth of these strains is very slow (0.18-0.24 day-1) and is inhibited by exposure to light. The genomes, of ca. 3.08 Mbp, encode archaella (archaeal motility structures) and multiple sets of enzyme paralogs, including 80 genes coding for enolase superfamily enzymes and 44 genes encoding NAD(P)-dependent dehydrogenases. We propose that these enzyme paralogs participate in multiple parallel pathways for non-phosphorylative catabolism of sugars and sugar acids. Indeed, we demonstrate that SAR202 strains can utilize several substrates that are metabolized through the predicted pathways, such as sugars ʟ-fucose and ʟ-rhamnose, as well as their lactone and acid forms.

New insights on the photocomplex of Roseiflexus castenholzii revealed from comparisons of native and carotenoid-depleted complexes.

In wild-type phototrophic organisms, carotenoids (Crts) are primarily packed into specific pigment-protein complexes along with (Bacterio)chlorophylls and play important roles in the photosynthesis. Diphenylamine (DPA) inhibits carotenogenesis but not phototrophic growth of anoxygenic phototrophs and eliminates virtually all Crts from photocomplexes. To investigate the effect of Crts on assembly of the reaction center-light-harvesting (RC-LH) complex from the filamentous anoxygenic phototroph Roseiflexus (Rfl.) castenholzii, we generated carotenoidless (Crt-less) RC-LH complexes by growing cells in the presence of DPA. Here, we present cryo-EM structures of the Rfl. castenholzii native and Crt-less RC-LH complexes with resolutions of 2.86 Å and 2.85 Å, respectively. From the high-quality map obtained, several important but previously unresolved details in the Rfl. castenholzii RC-LH structure were determined unambiguously including the assignment and likely function of three small polypeptides, and the content and spatial arrangement of Crts with bacteriochlorophyll molecules. The overall structures of Crt-containing and Crt-less complexes are similar. However, structural comparisons showed that only five Crts remain in complexes from DPA-treated cells and that the subunit X (TMx) flanked on the N-terminal helix of the Cyt-subunit is missing. Based on these results, the function of Crts in the assembly of the Rfl. castenholzii RC-LH complex and the molecular mechanism of quinone exchange is discussed. These structural details provide a fresh look at the photosynthetic apparatus of an evolutionary ancient phototroph as well as new insights into the importance of Crts for proper assembly and functioning of the RC-LH complex.

The effects of arsenic on dechlorination of trichloroethene by consortium DH: Microbial response and resistance.

Inorganic arsenic and organochlorines are frequently co-occurring contaminants in anoxic groundwater environments, and the bioremediation of their composite pollution has long been a rigorous predicament. Currently, the dechlorination behaviors and stress responses of microbial dechlorination consortia to arsenic are not yet fully understood. This study assessed the reductive dechlorination performance of a Dehalococcoides-bearing microcosm DH under gradient concentrations of arsenate [As(V)] or arsenite [As(III)] and investigated the response patterns of different functional microorganisms. Our results demonstrated that although the dechlorination rates declined with increasing arsenic concentrations in both As(III/V) scenarios, the inhibitory impact was more pronounced in As(III)-amended groups compared to As(V)-amended groups. Moreover, the vinyl chloride (VC)-to-ethene step was more susceptible to arsenic exposure compared to the trichloroethene (TCE)-to-dichloroethane (DCE) step, while high levels of arsenic exposure [e.g. As(III) > 75 µM] can induce significant accumulation of VC. Functional gene variations and microbial community analyses revealed that As(III/V) affected reductive dechlorination by directly inhibiting organohalide-respiring bacteria (OHRB) and indirectly inhibiting synergistic populations such as acetogens. Metagenomic results indicated that arsenic metabolic and efflux mechanisms were identical among different Dhc strains, and variations in arsenic uptake pathways were possibly responsible for their differential responses to arsenic exposures. By comparison, fermentative bacteria showed high potential for arsenic resistance due to their inherent advantages in arsenic detoxification and efflux mechanisms. Collectively, our findings expanded the understanding of the response patterns of different functional populations to arsenic stress in the dechlorinating consortium and provided insights into modifying bioremediation strategies at co-contaminated sites for furtherance.

Intermittent Oxygen Supply Facilitates Codegradation of Trichloroethene and Toluene by Anaerobic Consortia.

Biodegradation is commonly employed for remediating trichloroethene- or toluene-contaminated sites. However, remediation methods using either anaerobic or aerobic degradation are inefficient for dual pollutants. We developed an anaerobic sequencing batch reactor system with intermittent oxygen supply for the codegradation of trichloroethylene and toluene. Our results showed that oxygen inhibited anaerobic dechlorination of trichloroethene, but dechlorination rates remained comparable to that at dissolved oxygen levels of 0.2 mg/L. Intermittent oxygenation engendered reactor redox fluctuations (-146 to -475 mV) and facilitated rapid codegradation of targeting dual pollutants, with trichloroethene degradation constituting only 27.5% of the noninhibited dechlorination. Amplicon sequencing analysis revealed the predominance of Dehalogenimonas (16.0% ± 3.5%) over Dehalococcoides (0.3% ± 0.2%), with ten times higher transcriptomic activity in Dehalogenimonas. Shotgun metagenomics revealed numerous genes related to reductive dehalogenases and oxidative stress resistance in Dehalogenimonas and Dehalococcoides, as well as the enrichment of diversified facultative populations with functional genes related to trichloroethylene cometabolism and aerobic and anaerobic toluene degradation. These findings suggested that the codegradation of trichloroethylene and toluene may involve multiple biodegradation mechanisms. Overall results of this study demonstrate the effectiveness of intermittent micro-oxygenation in aiding trichloroethene-toluene degradation, suggesting the potential for the bioremediation of sites with similar organic pollutants.

Insight into the Mechanism Underlying Dehalococcoides mccartyi Strain CBDB1-Mediated B12-Dependent Aromatic Reductive Dehalogenation.

Anaerobic bacteria transform aromatic halides through reductive dehalogenation. This dehalorespiration is catalyzed by the supernucleophilic coenzyme vitamin B12, cob(I)alamin, in reductive dehalogenases. So far, the underlying inner-sphere electron transfer (ET) mechanism has been discussed controversially. In the present study, all 36 chloro-, bromo-, and fluorobenzenes and full-size cobalamin are analyzed at the quantum chemical density functional theory level with respect to a wide range of theoretically possible inner-sphere ET mechanisms. The calculated reaction free energies within the framework of CoI···X (X = F, Cl, and Br) attack rule out most of the inner-sphere pathways. The only route with feasible energetics is a proton-coupled two-ET mechanism that involves a B12 side-chain tyrosine (modeled by phenol) as a proton donor. For 12 chlorobenzenes and 9 bromobenzenes with experimental data from Dehalococcoides mccartyi strain CBDB1, the newly proposed PC-TET mechanism successfully discriminates 16 of 17 active from 4 inactive substrates and correctly predicts the observed regiospecificity to 100%. Moreover, fluorobenzenes are predicted to be recalcitrant in agreement with experimental findings. Conceptually, based on the Bell-Evans-Polanyi principle, the computational approach provides novel mechanistic insights and may serve as a tool for predicting the energetic feasibility of reductive aromatic dehalogenation.

Microbiome convergence and deterministic community assembly along successional biocrust gradients on potash salt heaps.

Potash mining, typically performed for agricultural fertilizer production, can create piles of residual salt waste that are ecologically detrimental and difficult to revegetate. Biological soil crusts (biocrusts) have been found growing on and around these heaps, suggesting resilience to the hypersaline environment. We set out to understand the community dynamics of biocrust formation by examining two succesionary salinity gradients at historical mining sites using a high throughput amplicon sequencing. Bare heaps were distinct, with little overlap between sites, and were characterized by high salinity, low nutrient availability, and specialized, low diversity microbial communities, dominated by Halobacteria, Chloroflexia, and Deinococci. 'Initial' stages of biocrust development were dominated by site-specific Cyanobacteria, with significant overlap between sites. Established biocrusts were the most diverse, with large proportions of Alphaproteobacteria, Anaerolineae, and Planctomycetacia. Along the salinity gradient at both sites, salinity decreased, pH decreased, and nutrients and Chlorophyll a increased. Microbiomes between sites converged during succession and community assembly process analysis revealed biocrusts at both sites were dominated by deterministic, niche-based processes; indicating a high degree of phylogenetic turnover. We posit early cyanobacterial colonization is essential for biocrust initiation, and facilitates later establishment of plant and other higher-level biota.

Exploring Prokaryotic Communities in the Guts and Mucus of Nudibranchs, and Their Similarity to Sediment and Seawater Microbiomes.

In the present study, we compared mucus and gut-associated prokaryotic communities from seven nudibranch species with sediment and seawater from Thai coral reefs using high-throughput 16S rRNA gene sequencing. The nudibranch species were identified as Doriprismatica atromarginata (family Chromodorididae), Jorunna funebris (family Discodorididae), Phyllidiella nigra, Phyllidiella pustulosa, Phyllidia carlsonhoffi, Phyllidia elegans, and Phyllidia picta (all family Phyllidiidae). The most abundant bacterial phyla in the dataset were Proteobacteria, Tenericutes, Chloroflexi, Thaumarchaeota, and Cyanobacteria. Mucus and gut-associated communities differed from one another and from sediment and seawater communities. Host phylogeny was, furthermore, a significant predictor of differences in mucus and gut-associated prokaryotic community composition. With respect to higher taxon abundance, the order Rhizobiales (Proteobacteria) was more abundant in Phyllidia species (mucus and gut), whereas the order Mycoplasmatales (Tenericutes) was more abundant in D. atromarginata and J. funebris. Mucus samples were, furthermore, associated with greater abundances of certain phyla including Chloroflexi, Poribacteria, and Gemmatimonadetes, taxa considered to be indicators for high microbial abundance (HMA) sponge species. Overall, our results indicated that nudibranch microbiomes consisted of a number of abundant prokaryotic members with high sequence similarities to organisms previously detected in sponges.

Structural basis of a bi-functional malonyl-CoA reductase (MCR) from the photosynthetic green non-sulfur bacterium Roseiflexus castenholzii.

Malonyl-CoA reductase (MCR) is a NADPH-dependent bi-functional enzyme that performs alcohol dehydrogenase and aldehyde dehydrogenase (CoA-acylating) activities in the N- and C-terminal fragments, respectively. It catalyzes the two-step reduction of malonyl-CoA to 3-hydroxypropionate (3-HP), a key reaction in the autotrophic CO2 fixation cycles of Chloroflexaceae green non-sulfur bacteria and the archaea Crenarchaeota. However, the structural basis underlying substrate selection, coordination, and the subsequent catalytic reactions of full-length MCR is largely unknown. For the first time, we here determined the structure of full-length MCR from the photosynthetic green non-sulfur bacterium Roseiflexus castenholzii (RfxMCR) at 3.35 Å resolution. Furthermore, we determined the crystal structures of the N- and C-terminal fragments bound with reaction intermediates NADP+ and malonate semialdehyde (MSA) at 2.0 Å and 2.3 Å, respectively, and elucidated the catalytic mechanisms using a combination of molecular dynamics simulations and enzymatic analyses. Full-length RfxMCR was a homodimer of two cross-interlocked subunits, each containing four tandemly arranged short-chain dehydrogenase/reductase (SDR) domains. Only the catalytic domains SDR1 and SDR3 incorporated additional secondary structures that changed with NADP+-MSA binding. The substrate, malonyl-CoA, was immobilized in the substrate-binding pocket of SDR3 through coordination with Arg1164 and Arg799 of SDR4 and the extra domain, respectively. Malonyl-CoA was successively reduced through protonation by the Tyr743-Arg746 pair in SDR3 and the catalytic triad (Thr165-Tyr178-Lys182) in SDR1 after nucleophilic attack from NADPH hydrides. IMPORTANCE The bi-functional MCR catalyzes NADPH-dependent reduction of malonyl-CoA to 3-HP, an important metabolic intermediate and platform chemical, from biomass. The individual MCR-N and MCR-C fragments, which contain the alcohol dehydrogenase and aldehyde dehydrogenase (CoA-acylating) activities, respectively, have previously been structurally investigated and reconstructed into a malonyl-CoA pathway for the biosynthetic production of 3-HP. However, no structural information for full-length MCR has been available to illustrate the catalytic mechanism of this enzyme, which greatly limits our capacity to increase the 3-HP yield of recombinant strains. Here, we report the cryo-electron microscopy structure of full-length MCR for the first time and elucidate the mechanisms underlying substrate selection, coordination, and catalysis in the bi-functional MCR. These findings provide a structural and mechanistic basis for enzyme engineering and biosynthetic applications of the 3-HP carbon fixation pathways.

Nano-Nd2O3 reduced soil bacterial community function by altering the relative abundance of rare and sensitive taxa.

Nanoparticulate-Nd2O3 (nano-Nd2O3) has been excessively utilized in agriculture, industry, and medicine. Hence, nano-Nd2O3 can have environmental implications. However, the impact of nano-Nd2O3 on alpha diversity, composition, and function of soil bacterial communities has not been thoroughly evaluated. We amended soil to achieve different concentrations of nano-Nd2O3 (0, 10, 50, and 100 mg kg-1 soil) and incubated the mesocosms for 60 days. On days 7 and 60 of the experiment, we measured the effect of nano-Nd2O3 on alpha diversity and composition of soil bacterial community. Further, the effect of nano-Nd2O3 on the function of soil bacterial community was assessed based on changes in the activities of the six potential enzymes that mediate the cycling of nutrients in the soil. Nano-Nd2O3 did not alter the alpha diversity and composition of the soil bacterial community; however, it negatively affected community function in a dose-dependent manner. Specifically, the activities of ß-1,4-glucosidase and ß-1,4-n-acetylglucosaminidase that mediate soil carbon and nitrogen cycling, respectively, were significantly affected on days 7 and 60 of the exposure. The effect of nano-Nd2O3 on the soil enzymes correlated with changes in relative abundances of the rare and sensitive taxa, viz., Isosphaerales, Isosphaeraceae, Ktedonobacteraceae, and Streptomyces. Overall, we provide information for the safe implementation of technological applications that use nano-Nd2O3.

Preparation and characterization of a laccase-like enzyme from Thermomicrobium roseum.

In this study, a laccase-like gene from Thermomicrobium roseum DSM 5159 (TrLac-like) (NCBI: WP_012642205.1) was recombinantly expressed in Bacillus subtilis WB600. The optimum temperature and pH for TrLac-like were 50 °C and 6.0, respectively. TrLac-like showed high tolerance to mixed systems of water and organic solvents, indicating its potential for large-scale application in various industries. It showed 36.81 % similarity with YlmD from Geobacillus stearothermophilus (PDB:6T1B) in sequence alignment; therefore, 6T1B was employed as the template for homology modeling. To improve catalytic efficiency, amino acid substitutions within 5 Å of the inosine ligand were simulated to reduce the binding energy and promote substrate affinity. Single and double substitutions (44 and 18, respectively) were prepared, and the catalytic efficiency of the mutant A248D was increased to approximately 110-fold that of the wild type, while the thermal stability was maintained. Bioinformatics analysis revealed that the significant improvement in catalytic efficiency could be attributed to the formation of new hydrogen bonds between the enzyme and substrate. With a further decrease in the binding energy, the catalytic efficiency of the multiple mutant H129N/A248D was approximately 14-fold higher than that of the wild type but lower than that of the single mutant A248D. This is possibly because kcat also decreased with the decrease of Km; consequently, the substrate could not be released in time owing to the enzyme with the combination mutation not being able to release the substrate at a high rate.

Dual effects of PFOA or PFOS on reductive dechlorination of trichloroethylene (TCE).

PFASs and chlorinated solvents are the common co-contaminants in soil and groundwater at firefighter training areas (FTAs). Although PFASs mixtures could have adverse impacts on bioremediation of trichloroethylene (TCE) by inhibiting Dehalococcoides (Dhc), little is known about the effect and contribution of PFOA or PFOS on dechlorination of TCE by non-Dhc organohalide-respiring bacteria (OHRB). To study this, PFOA and PFOS were amended to the growth medium of a non-Dhc OHRB-containing enrichment culture to determine the impact on dechlorination. This study demonstrated that high levels of PFOA or PFOS (100 mg L-1) inhibited TCE dechlorination in four non-Dhc OHRB-containing community including Geobacter, Desulfuromonas, Desulfitobacterium, and Dehalobacter, but low levels of PFOA or PFOS (≤10 mg L-1) enhanced TCE dechlorination. Four non-Dhc OHRB were less inhibited by PFOA than that by PFOS, and high level of PFOS killed Desulfitobacterium and Dehalobacter and decreased the biodiversity of bacterial community. Although most fermenters were killed by the presence of 100 mg L-1 PFOS, two important co-cultures (Desulfovibrio and Sedimentibacter) of OHRB were enriched, indicating that the syntrophic relationships between OHRB and co-cultures still remained, and PFOA or PFOS inhibited TCE dechlorination by directly repressing non-Dhc OHRB. Our results highlight that the bioattenuation of chloroethene contamination could be confounded by non-Dhc OHRB in high levels of PFOS contaminated subsurface environments at FTAs.

Microbial Reductive Dechlorination by a Commercially Available Dechlorinating Consortium Is Not Inhibited by Perfluoroalkyl Acids (PFAAs) at Field-Relevant Concentrations.

Perfluoroalkyl acids (PFAAs) have been shown to inhibit biodegradation (i.e., organohalide respiration) of chlorinated ethenes. The potential negative impacts of PFAAs on microbial species performing organohalide respiration, particularly Dehalococcoides mccartyi (Dhc), and the efficacy of in situ bioremediation are a critical concern for comingled PFAA-chlorinated ethene plumes. Batch reactor (no soil) and microcosm (with soil) experiments, containing a PFAA mixture and bioaugmented with KB-1, were completed to assess the impact of PFAAs on chlorinated ethene organohalide respiration. In batch reactors, PFAAs delayed complete biodegradation of cis-1,2-dichloroethene (cis-DCE) to ethene. Maximum substrate utilization rates (a metric for quantifying biodegradation rates) were fit to batch reactor experiments using a numerical model that accounted for chlorinated ethene losses to septa. Fitted values for cis-DCE and vinyl chloride biodegradation were significantly lower (p < 0.05) in batch reactors containing ≥50 mg/L PFAAs. Examination of reductive dehalogenase genes implicated in ethene formation revealed a PFAA-associated change in the Dhc community from cells harboring the vcrA gene to those harboring the bvcA gene. Organohalide respiration of chlorinated ethenes was not impaired in microcosm experiments with PFAA concentrations of 38.7 mg/L and less, suggesting that a microbial community containing multiple strains of Dhc is unlikely to be inhibited by PFAAs at lower, environmentally relevant concentrations.

Biogeographic distribution, ecotype partitioning and controlling factors of Chloroflexi in the sediments of six hadal trenches of the Pacific Ocean.

The hadal trenches are "hot spots" for mineralization of organic matter in the deep ocean. Chloroflexi are one of the most dominant and active taxa in trench sediments, serving as important drivers of carbon cycles in hadal trenches. However, current understanding on hadal Chloroflexi is largely restricted to individual trench. This study systematically analyzed the diversity, biogeographic distribution, ecotype partitioning as well as environmental drivers of Chloroflexi in the sediments of hadal trenches, by reanalyzing 16S rRNA gene libraries of 372 samples from 6 trenches around the Pacific Ocean. The results showed that Chloroflexi averagely account for 10.10 % and up to 59.95 % of total microbial communities in the trench sediments. Positive correlations between relative abundance of Chloroflexi and depths down the vertical sediment profiles were observed in all of the sediment cores analyzed, suggesting the increasing significance of Chloroflexi in deeper sediment layers. Overall, trench sediment Chloroflexi were mainly composed of the classes Dehalococcidia, Anaerolineae and JG30-KF-CM66, and four orders i.e. SAR202, Anaerolineales, norank JG30-KF-CM66 and S085, were identified as core taxa that were dominant and prevalent in the hadal trench sediments. A total of 22 subclusters were identified within these core orders, and distinct patterns of ecotype partitioning related with depths down the vertical sediment profiles were observed, suggesting the great diversification of metabolic potentials and environment preference of different Chloroflexi lineages. The spatial distribution of hadal Chloroflexi were found to be significantly related with multiple environmental factors, while depths down the vertical sediment profiles explained the highest proportion of variations. These results provide valuable information for further exploring the roles of Chloroflexi in biogeochemical cycle of the hadal zone, and lay the foundation for understanding the adaptive mechanisms and evolutionary characteristics of microorganisms in hadal trenches.

Thermophilic Dehalococcoidia with unusual traits shed light on an unexpected past.

Although the phylum Chloroflexota is ubiquitous, its biology and evolution are poorly understood due to limited cultivability. Here, we isolated two motile, thermophilic bacteria from hot spring sediments belonging to the genus Tepidiforma and class Dehalococcoidia within the phylum Chloroflexota. A combination of cryo-electron tomography, exometabolomics, and cultivation experiments using stable isotopes of carbon revealed three unusual traits: flagellar motility, a peptidoglycan-containing cell envelope, and heterotrophic activity on aromatics and plant-associated compounds. Outside of this genus, flagellar motility has not been observed in Chloroflexota, and peptidoglycan-containing cell envelopes have not been described in Dehalococcoidia. Although these traits are unusual among cultivated Chloroflexota and Dehalococcoidia, ancestral character state reconstructions showed flagellar motility and peptidoglycan-containing cell envelopes were ancestral within the Dehalococcoidia, and subsequently lost prior to a major adaptive radiation of Dehalococcoidia into marine environments. However, despite the predominantly vertical evolutionary histories of flagellar motility and peptidoglycan biosynthesis, the evolution of enzymes for degradation of aromatics and plant-associated compounds was predominantly horizontal and complex. Together, the presence of these unusual traits in Dehalococcoidia and their evolutionary histories raise new questions about the timing and selective forces driving their successful niche expansion into global oceans.

Sustained detoxification of 1,2-dichloroethane to ethylene by a symbiotic consortium containing Dehalococcoides species.

1,2-Dichloroethane (1,2-DCA) is a ubiquitous volatile halogenated organic pollutant in groundwater and soil, which poses a serious threat to the ecosystem and human health. Microbial reductive dechlorination has been recognized as an environmentally-friendly strategy for the remediation of sites contaminated with 1,2-DCA. In this study, we obtained an anaerobic microbiota derived from 1,2-DCA contaminated groundwater, which was able to sustainably convert 1,2-DCA into non-toxic ethylene with an average dechlorination rate of 30.70 ± 11.06 µM d-1 (N = 6). The microbial community profile demonstrated that the relative abundance of Dehalococcoides species increased from 0.53 ± 0.08% to 44.68 ± 3.61% in parallel with the dechlorination of 1,2-DCA. Quantitative PCR results showed that the Dehalococcoides species 16S rRNA gene increased from 2.40 ± 1.71 × 108 copies∙mL-1 culture to 4.07 ± 2.45 × 108 copies∙mL-1 culture after dechlorinating 110.69 ± 30.61 µmol of 1,2-DCA with a growth yield of 1.55 ± 0.93 × 108 cells per µmol Cl- released (N = 6), suggesting that Dehalococcoides species used 1,2-DCA for organohalide respiration to maintain cell growth. Notably, the relative abundances of Methanobacterium sp. (p = 0.0618) and Desulfovibrio sp. (p = 0.0001995) also increased significantly during the dechlorination of 1,2-DCA and were clustered in the same module with Dehalococcoides species in the co-occurrence network. These results hinted that Dehalococcoides species, the obligate organohalide-respiring bacterium, exhibited potential symbiotic relationships with Methanobacterium and Desulfovibrio species. This study illustrates the importance of microbial interactions within functional microbiota and provides a promising microbial resource for in situ bioremediation in sites contaminated with 1,2-DCA.

Decoupling Fe0 Application and Bioaugmentation in Space and Time Enables Microbial Reductive Dechlorination of Trichloroethene to Ethene: Evidence from Soil Columns.

Fe0 is a powerful chemical reductant with applications for remediation of chlorinated solvents, including tetrachloroethene and trichloroethene. Its utilization efficiency at contaminated sites is limited because most of the electrons from Fe0 are channeled to the reduction of water to H2 rather than to the reduction of the contaminants. Coupling Fe0 with H2-utilizing organohalide-respiring bacteria (i.e., Dehalococcoides mccartyi) could enhance trichloroethene conversion to ethene while maximizing Fe0 utilization efficiency. Columns packed with aquifer materials have been used to assess the efficacy of a treatment combining in space and time Fe0 and aD. mccartyi-containing culture (bioaugmentation). To date, most column studies documented only partial conversion of the solvents to chlorinated byproducts, calling into question the feasibility of Fe0 to promote complete microbial reductive dechlorination. In this study, we decoupled the application of Fe0 in space and time from the addition of organic substrates andD. mccartyi-containing cultures. We used a column containing soil and Fe0 (at 15 g L-1 in porewater) and fed it with groundwater as a proxy for an upstream Fe0 injection zone dominated by abiotic reactions and biostimulated/bioaugmented soil columns (Bio-columns) as proxies for downstream microbiological zones. Results showed that Bio-columns receiving reduced groundwater from the Fe0-column supported microbial reductive dechlorination, yielding up to 98% trichloroethene conversion to ethene. The microbial community in the Bio-columns established with Fe0-reduced groundwater also sustained trichloroethene reduction to ethene (up to 100%) when challenged with aerobic groundwater. This study supports a conceptual model where decoupling the application of Fe0 and biostimulation/bioaugmentation in space and/or time could augment microbial trichloroethene reductive dechlorination, particularly under oxic conditions.

Surface-layer protein is a public-good matrix exopolymer for microbial community organisation in environmental anammox biofilms.

Extracellular polymeric substances (EPS) are core biofilm components, yet how they mediate interactions within and contribute to the structuring of biofilms is largely unknown, particularly for non-culturable microbial communities that predominate in environmental habitats. To address this knowledge gap, we explored the role of EPS in an anaerobic ammonium oxidation (anammox) biofilm. An extracellular glycoprotein, BROSI_A1236, from an anammox bacterium, formed envelopes around the anammox cells, supporting its identification as a surface (S-) layer protein. However, the S-layer protein also appeared at the edge of the biofilm, in close proximity to the polysaccharide-coated filamentous Chloroflexi bacteria but distal to the anammox bacterial cells. The Chloroflexi bacteria assembled into a cross-linked network at the edge of the granules and surrounding anammox cell clusters, with the S-layer protein occupying the space around the Chloroflexi. The anammox S-layer protein was also abundant at junctions between Chloroflexi cells. Thus, the S-layer protein is likely transported through the matrix as an EPS and also acts as an adhesive to facilitate the assembly of filamentous Chloroflexi into a three-dimensional biofilm lattice. The spatial distribution of the S-layer protein within the mixed species biofilm suggests that it is a "public-good" EPS, which facilitates the assembly of other bacteria into a framework for the benefit of the biofilm community, and enables key syntrophic relationships, including anammox.

The majority of microorganisms in gas hydrate-bearing subseafloor sediments ferment macromolecules.

BACKGROUND: Gas hydrate-bearing subseafloor sediments harbor a large number of microorganisms. Within these sediments, organic matter and upward-migrating methane are important carbon and energy sources fueling a light-independent biosphere. However, the type of metabolism that dominates the deep subseafloor of the gas hydrate zone is poorly constrained. Here we studied the microbial communities in gas hydrate-rich sediments up to 49 m below the seafloor recovered by drilling in the South China Sea. We focused on distinct geochemical conditions and performed metagenomic and metatranscriptomic analyses to characterize microbial communities and their role in carbon mineralization. RESULTS: Comparative microbial community analysis revealed that samples above and in sulfate-methane interface (SMI) zones were clearly distinguished from those below the SMI. Chloroflexota were most abundant above the SMI, whereas Caldatribacteriota dominated below the SMI. Verrucomicrobiota, Bathyarchaeia, and Hadarchaeota were similarly present in both types of sediment. The genomic inventory and transcriptional activity suggest an important role in the fermentation of macromolecules. In contrast, sulfate reducers and methanogens that catalyze the consumption or production of commonly observed chemical compounds in sediments are rare. Methanotrophs and alkanotrophs that anaerobically grow on alkanes were also identified to be at low abundances. The ANME-1 group actively thrived in or slightly below the current SMI. Members from Heimdallarchaeia were found to encode the potential for anaerobic oxidation of short-chain hydrocarbons. CONCLUSIONS: These findings indicate that the fermentation of macromolecules is the predominant energy source for microorganisms in deep subseafloor sediments that are experiencing upward methane fluxes. Video Abstract.