Exocellular electron transfer in anaerobic microbial communities

Exocellular electron transfer plays an important role in anaerobic microbial communities that degrade organic matter. Interspecies hydrogen transfer between microorganisms is the driving force for complete biodegradation in methanogenic environments. Many organic compounds are degraded by obligatory syntrophic consortia of proton-reducing acetogenic bacteria and hydrogen-consuming methanogenic archaea. Anaerobic microorganisms that use insoluble electron acceptors for growth, such as iron- and manganese-oxide as well as inert graphite electrodes in microbial fuel cells, also transfer electrons exocellularly. Soluble compounds, like humic substances, quinones, phenazines and riboflavin, can function as exocellular electron mediators enhancing this type of anaerobic respiration. However, direct electron transfer by cell-cell contact is important as well. This review addresses the mechanisms of exocellular electron transfer in anaerobic microbial communities. There are fundamental differences but also similarities between electron transfer to another microorganism or to an insoluble electron acceptor. The physical separation of the electron donor and electron acceptor metabolism allows energy conservation in compounds as methane and hydrogen or as electricity. Furthermore, this separation is essential in the donation or acceptance of electrons in some environmental technological processes, e.g. soil remediation, wastewater purification and corrosion.     

Mixed aerobic and anaerobic microbial communities in benzene-contaminated groundwater

Aims:  To investigate the factors affecting benzene biodegradation and microbial community composition in a contaminated aquifer.
Methods and Results:  We identified the microbial community in groundwater samples from a benzene-contaminated aquifer situated below a petrochemical plant. Eleven out of twelve groundwater samples with in situ dissolved oxygen concentrations between 0 and 2·57 mg l⁻¹ showed benzene degradation in aerobic microcosm experiments, whereas no degradation in anaerobic microcosms was observed. The lack of aerobic degradation in the remaining microcosm could be attributed to a pH of 12·1. Three groundwaters, examined by 16S rRNA gene clone libraries, with low in situ oxygen concentrations and high benzene levels, each had a different dominant aerobic (or denitrifying) population, either Pseudomonas , Polaromonas or Acidovorax species. These groundwaters also had syntrophic organisms, and aceticlastic methanogens were detected in two samples. The alkaline groundwater was dominated by organisms closely related to Hydrogenophaga .
Conclusions:  Results show that pH 12·1 is inimical to benzene biodegradation, and that oxygen concentrations below 0·03 mg l⁻¹ can support aerobic benzene-degrading communities.
Significance and Impact of the Study:  These findings will help to guide the treatment of contaminated groundwaters, and raise questions about the extent to which aerobes and anaerobes may interact to effect benzene degradation.     

Transition of microbial communities during the adaption to anaerobic digestion of carrot waste

In this study a microbial community suitable for anaerobic digestion of carrot pomace was developed from inocula obtained from natural environmental sources. The changes along the process were monitored using pyrosequencing of the 16S rRNA gene. As the community adapted from a diverse natural community to a community with a definite function, diversity decreased drastically. Major bacterial groups remaining after enrichment were Bacilli (31-45.3%), Porphyromonadaceae (12.1-24.8%) and Spirochaetes (12.5-18.5%). The archaeal population was even less diverse and mainly represented by a single OTU that was 99.7% similar to Methanosarcina mazei. One enrichment which failed to produce large amounts of methane had shifts in the bacterial populations and loss of methanogenic archaea.     

Characterization of microbial communities in anaerobic bioreactors using molecular probes

The microbial community structure of twenty-one single-phase and one two-phase full-scale anaerobic sewage sludge digesters was evaluated using oligonucleotide probes complementary to conserved tracts of the 16S rRNAs of phylogenetically defined groups of methanogens and sulfate-reducing bacteria. These probe results were interpreted in combination with results from traditional chemical analyses and metabolic activity assays. It was determined that methanogens in healthy mesophilic, single-phase sewage sludge digesters accounted for approximately 8–12% of the total community and thatMethanosarcinales andMethanomicrobiales constituted the majority of the total methanogen population.Methanobacteriales andMethanococcales played a relatively minor role in the digesters. Phylogenetic groups of mesophilic, Gram-negative sulfate-reducing bacteria were consistently present at significant levels:Desulfovibrio andDesulfobulbus spp. were the dominant sulfate-reducing populations,Desulfobacter andDesulfobacterium spp. were present at lower levels, andDesulfosarcina, Desulfococcus, andDesulfobotulus spp. were absent. Sulfate reduction by one or more of these populations played a significant role in all digesters evaluated in this study. In addition, sulfate-reducing bacteria played a role in favoring methanogenesis by providing their substrates. The analysis of the two-phase digester indicated that true phase separation was not accomplished: significant levels of active methanogens were present in the first phase. It was determined that the dominant populations in the second phase were different from those in the single-phase digesters.     

Effect of adaptation to phenol on biodegradation of monosubstituted phenols by aquatic microbial communities.

The adaptation of a mixed aquatic microbial community to phenol was examined in microcosms receiving phenol as a sole carbon source. Extended exposure (adaptation) to phenol resulted in adaptation of the microbial community to the structurally related aromatic compounds m-cresol, m-aminophenol, and p-chlorophenol. The increased biodegradation potential of the phenol-adapted microbial community was accompanied by a concurrent increase in the number of microorganisms able to degrade the three test compounds. Thus, adaptation to the three test chemicals was likely a growth-related result of extended exposure to phenol. The results indicate that adaptation to a single chemical may increase the assimilative capacity of an aquatic environment for other related chemicals even in the absence of adaptation-inducing levels of those materials.     

Special traits of decomposition of azo dyes by anaerobic microbial communities

We present the results of an investigation into the special traits of conversion of azo dyes Acid Orange 6, Acid Orange 7, Methyl Orange, and Methyl Red under anaerobic conditions in comparison to aerobic conditions. In the presence of oxygen, only Methyl Red underwent decomposition, while under oxygen-free conditions, all remaining substances were fully decolourised under the action of a methanogenous consortium of microorganisms. The products of reduction of the azo bond are determined in the case of each dye. Introduction of additional acceptors of electrons (sulfate and nitrate) had a negative influence on the discoloration of azo dyes. Addition of ethanol as an available organic cosubstrate accelerated decomposition of azo dyes both under methanogenous and sulfate- and nitrate-reducing conditions. There is no direct correlation between the rates of conversion of azo dyes under anaerobic conditions or their toxicity to acetoclastic methanogens. Changes in the morphological composition of the community decolouring an azo dye depended on the duration of its impact on microorganisms. The mechanism of the reduction of the azo bond under the action of substances acting as mediators is explained. These substances are products of the metabolism of the microbial community in anaerobic conditions. It is shown that the supposed mediators NADH and sulfide efficiently decolourise azo dyes in a cell-free system, while riboflavin significantly increased the rate of conversion of substrates in recurrent cycles of discoloration only in the presence of an anaerobic microbial consortium.     

The pattern of recovery of disturbed microbial communities inhabiting hard substrates

The recovery of microbial communities removed from hard substrates was studied in the laboratory and on microepiphyton in the White Sea, in St. Petersburg, and in Karelija. The removal of these communities from hard substrates destroyed their spatial structure. Recovery was allowed to proceed from the suspension on a horizontal substrate in stagnant water at low temperature. The sequence of colonization in the laboratory was the same as that in the field. Laboratory recovery was complete in 12–24 h and did not depend on the age of the natural community. From the comparison between laboratory and field data on such mechanical disturbance, a pattern of recovery, termed ‘self-assembly’, is proposed. This revised version was published online in September 2006 with corrections to the Cover Date.     

Oxidation of sulfur-containing substrates by aboriginal and experimentally designed microbial communities

Aboriginal and experimental (constructed of pure microbial cultures) communities of acidophilic chemolithotrophs have been studied. The oxidation of elemental sulfur, sodium thiosulfate, and potassium tetrathionate as sole sources of energy has been monitored. The oxidation rate of the experimental community is higher as compared to the aboriginal community isolated from a flotation concentrate of pyrrhotine-containing pyrite-arsenopyrite gold-arsenic sulfide ore. The degree of oxidation of the mentioned S substrates amounts to 17.91, 68.30, and 93.94% for the experimental microbial community and to 10.71, 56.03, and 79.50% for the aboriginal community, respectively. The degree of oxidation of sulfur sulfide forms in the ore flotation concentrate is 59.15% by the aboriginal microbial community and 49.40% by the experimental microbial community. Despite a higher rate of oxidation of S substrates as a sole source of energy by the experimental microbial community, the aboriginal community oxidizes S substrates at a higher rate in the flotation concentrate of pyrrhotine-containing pyrite-arsenopyrite gold-arsenic sulfide ore, from which it was isolated. Bacterial-chemical oxidation of the flotation concentrate by the aboriginal microbial community allows for the extraction of an additional 32.3% of gold from sulfide minerals, which is by 5.7% larger compared to the yield obtained by the experimental microbial community.     

Environmental shaping of codon usage and functional adaptation across microbial communities

Microbial communities represent the largest portion of the Earth’s biomass. Metagenomics projects use high-throughput sequencing to survey these communities and shed light on genetic capabilities that enable microbes to inhabit every corner of the biosphere. Metagenome studies are generally based on (i) classifying and ranking functions of identified genes; and (ii) estimating the phyletic distribution of constituent microbial species. To understand microbial communities at the systems level, it is necessary to extend these studies beyond the species’ boundaries and capture higher levels of metabolic complexity. We evaluated 11 metagenome samples and demonstrated that microbes inhabiting the same ecological niche share common preferences for synonymous codons, regardless of their phylogeny. By exploring concepts of translational optimization through codon usage adaptation, we demonstrated that community-wide bias in codon usage can be used as a prediction tool for lifestyle-specific genes across the entire microbial community, effectively considering microbial communities as meta-genomes. These findings set up a ‘functional metagenomics’ platform for the identification of genes relevant for adaptations of entire microbial communities to environments. Our results provide valuable arguments in defining the concept of microbial species through the context of their interactions within the community.     

Different substrates within a lake harbour connected but specialised microbial communities

Hydrobiologia - Natural water bodies contain physically interconnected habitats suitable for microbes, such as different water layers and substrates for biofilms. Yet, little is known on the extent...     

Diversity of five anaerobic toluene-degrading microbial communities investigated using stable isotope probing

Time-series DNA-stable isotope probing (SIP) was used to identify the microbes assimilating carbon from [(13)C]toluene under nitrate- or sulfate-amended conditions in a range of inoculum sources, including uncontaminated and contaminated soil and wastewater treatment samples. In all, five different phylotypes were found to be responsible for toluene degradation, and these included previously identified toluene degraders as well as novel toluene-degrading microorganisms. In microcosms constructed from granular sludge and amended with nitrate, the putative toluene degraders were classified in the genus Thauera, whereas in nitrate-amended microcosms constructed from a different source (agricultural soil), microorganisms in the family Comamonadaceae (genus unclassified) were the key putative degraders. In one set of sulfate-amended microcosms (agricultural soil), the putative toluene degraders were identified as belonging to the class Clostridia (genus Desulfosporosinus), while in other sulfate-amended microcosms, the putative degraders were in the class Deltaproteobacteria, within the family Syntrophobacteraceae (digester sludge) or Desulfobulbaceae (contaminated soil) (genus unclassified for both). Partial benzylsuccinate synthase gene (bssA, the functional gene for anaerobic toluene degradation) sequences were obtained for some samples, and quantitative PCR targeting this gene, along with SIP, was further used to confirm anaerobic toluene degradation by the identified species. The study illustrates the diversity of toluene degraders across different environments and highlights the utility of ribosomal and functional gene-based SIP for linking function with identity in microbial communities.     

Local adaptation of microbial communities to heavy metal stress in polluted sediments of Lake Erie

Microbial communities must balance the assimilation of biologically necessary metals with resistance to toxic metal concentrations. To investigate the impact of heavy metal contaminants on microbial communities, we performed two experiments measuring extracellular enzyme activities (EEA) in polluted and unpolluted sediments of Lake Erie. In the first experiment, inoculations with moderate concentrations of copper and zinc appreciably diminished EEA from uncontaminated sites, whereas EEA from contaminated sediments increased or were only negligibly affected. In the second experiment, we compared the effects of three separate metals (i.e. copper, arsenic, and cadmium) on microbial community metabolism in polluted and unpolluted locations. Although copper and arsenic elicited differential effects by inhibiting EEA only in unpolluted sediments, cadmium inhibited EEA in both polluted and unpolluted sediments. Multivariate analyses of EEA from polluted sediments revealed direct associations among hydrolytic enzymes and inverse or absent associations between hydrolases and oxidases; these associations demonstrated resilience to heavy metal stress. In contrast, addition of heavy metals to unpolluted sediments appeared to have a higher impact on the multivariate pattern of EEA associations as revealed by an increase in the number of associations, more inverse relationships, and potential enzymatic trade-offs. The results of this study suggest community-level adaptations through the development of resistance mechanisms to the types and local levels of heavy metals in the environment.     

Molecular characterization of anaerobic microbial communities from benzene-degrading sediments under methanogenic conditions

Anaerobic benzene degradation was confirmed in microbial communities enriched from Baltimore Harbor (Baltimore, MD) sediments under methanogenic conditions. Molecular characterization based on 16S rDNA gene sequences revealed that the strains in the communities were diversely affiliated with such phylogenetic branches as the Bacteroidetes, Euryarchaeota, Firmicutes, and Thermotogae phyla. Of interest was that the majority of the microbial populations detected in these cultures were closely related to the members of dechlorinating microbial communities. Further, some of those species were previously found in naphthalene- or phenanthrene-degrading methanogenic communities. Finally, this result could be used to design targeted isolation strategies for anaerobic benzene-degrading strains under methanogenic conditions.     

Methanogenic degradation of (amino)aromatic compounds by anaerobic microbial communities

degradation of a range of aromatic substrates by anaerobic microbial communities was studied. Active methanogenic microbial communities decomposing aminoaromatic acids and azo dyes into CH₄ and CO₂ were isolated. Products of primary conversion were found to be 2-hydroxybenzyl and benzyl alcohols gradually transforming into benzoate. It was shown that isolated microbial communities are capable of converting the initial substrates—benzyl alcohol, benzoate, salicylic acid, and azo dye Acid Orange 6—into biogas without a lag-phase but with different velocities. Aromatic and linear intermediates of biodegradation of aromatic amines by obtained enrichment cultures were determined for the first time. Selective effect of aromatic substrates on a microbial community that was expressed in decrease in diversity and gradual change of dominant morphotypes was revealed.     

Methanogenic destruction of (amino)aromatic compounds by anaerobic microbial communities

Destruction of a number of aromatic substrates by anaerobic microbial communities was studied. Active methanogenic microbial communities decomposing aminoaromatic acids and azo dyes into CH4 and CO2 were isolated. Products of primary conversion were found to be 2-hydroxybenzyl and benzyl alcohols gradually transforming into benzoate. It was shown that isolated microbial communities are capable of converting the initial substrates--benzyl alcohol, benzoate, salicylic acid, and golden yellow azo dye--into biogas without a lag-phase but with different velocities. Aromatic and linear intermediates of biodestruction of aromatic amines by obtained enrichment cultures were determined for the first time. Selective effect of aromatic substrates on a microbial community that was expressed in decrease in diversity and gradual change of dominant morphotypes was revealed.     

Analysis of microbial communities developed on the fouling layers of a membrane-coupled anaerobic bioreactor applied to wastewater treatment

The structure of the biofouling layers formed on a pilot-scale membrane-coupled upflow anaerobic sludge blanket bioreactor (UASB) used to treat urban wastewater was analyzed by scanning electron microscopy and electron-dispersive X-ray microanalysis. For comparison, control samples of the membranes were fed either UASB effluent or raw wastewater in a laboratory-scale experiment. Microbial diversity in the fouling materials was analyzed by temperature gradient gel electrophoresis (TGGE) combined with sequence analysis of partial 16S rRNA. Significant differences in structure of the Bacteria communities were observed amongst the different fouling layers analyzed in the UASB membranes, particularly following a chemical cleaning step (NaClO), while the Archaea communities retained more similarity in all samples. The main Bacteria populations identified were evolutively close to Firmicutes (42.3%) and Alphaproteobacteria (30.8%), while Archaea were mostly affiliated to the Methanosarcinales and Methanospirillaceae. Sphingomonadaceae-related bacteria and methanogenic Archaea were persistently found as components of biofouling, regardless of chemical cleaning.     

Pristine soils mineralize 3-chlorobenzoate and 2,4-dichlorophenoxyacetate via different microbial populations.

Biodegradation of two chlorinated aromatic compounds was found to be a common capability of the microorganisms found in the soils of undisturbed, pristine ecosystems. We used 2,4-dichlorophenoxyacetate (2,4-D) and 3-chlorobenzoate (3CBA) as enrichment substrates to compare populations of degrading bacteria from six different regions making up two ecosystems. We collected soil samples from four Mediterranean (California, central Chile, the Cape region of South Africa, and southwestern Australia) and two boreal (northern Saskatchewan and northwestern Russia) ecosystems that had no direct exposure to pesticides or to human disturbance. Between 96 and 120 samples from each of the six regions were incubated with 50 ppm of [U-14C]2,4-D or [U-14C]3CBA. Soils from all regions samples mineralized both 2,4-D and 3CBA, but 3CBA was mineralized without a lag period, while 2,4-D was generally not mineralized until the second week. 3CBA degradative capabilities were more evenly distributed spatially than those for 2,4-D. The degradative capabilities of the soils were readily transferred to fresh liquid medium. 3CBA degraders were easily isolated from most soils. We recovered 610 strains that could release carbon dioxide from ring-labeled 3CBA. Of these, 144 strains released chloride and degraded over 80% of 1 mM 3CBA in 3 weeks or less. In contrast, only five 2,4-D degraders could be isolated, although a variety of methods were used in an attempt to culture the degraders. The differences in the distribution and culturability of the bacteria responsible for 3CBA and 2,4-D degradation in these ecosystems suggest that the two substrates are degraded by different populations. We also describe a 14C-based microtiter plate method that allows efficient screening of a large number of samples for biodegradation activity.     

Bacterial adaptation to sublethal antibiotic gradients can change the ecological properties of multitrophic microbial communities

Antibiotics leak constantly into environments due to widespread use in agriculture and human therapy. Although sublethal concentrations are well known to select for antibiotic-resistant bacteria, little is known about how bacterial evolution cascades through food webs, having indirect effect on species not directly affected by antibiotics (e.g. via population dynamics or pleiotropic effects). Here, we used an experimental evolution approach to test how temporal patterns of antibiotic stress, as well as migration within metapopulations, affect the evolution and ecology of microcosms containing one prey bacterium, one phage and two protist predators. We found that environmental variability, autocorrelation and migration had only subtle effects for population and evolutionary dynamics. However, unexpectedly, bacteria evolved greatest fitness increases to both antibiotics and enemies when the sublethal levels of antibiotics were highest, indicating positive pleiotropy. Crucially, bacterial adaptation cascaded through the food web leading to reduced predator-to-prey abundance ratio, lowered predator community diversity and increased instability of populations. Our results show that the presence of natural enemies can modify and even reverse the effects of antibiotics on bacteria, and that antibiotic selection can change the ecological properties of multitrophic microbial communities by having indirect effects on species not directly affected by antibiotics.