Microbial and functional diversity of a subterrestrial high pH groundwater associated to serpentinization

Microbial and functional diversity were assessed, from a serpentinization‐driven subterrestrial alkaline aquifer – Cabeço de Vide Aquifer (CVA) in Portugal. DGGE analyses revealed the presence of a stable microbial community. By 16S rRNA gene libraries and pyrosequencing analyses, a diverse bacterial composition was determined, contrasting with low archaeal diversity. Within Bacteria the majority of the populations were related to organisms or sequences affiliated to class Clostridia, but members of classes Acidobacteria, Actinobacteria, Alphaproteobacteria, Betaproteobacteria, Deinococci, Gammaproteobacteria and of the phyla Bacteroidetes, Chloroflexi and Nitrospira were also detected. Domain Archaea encompassed mainly sequences affiliated to Euryarchaeota. Only form I RuBisCO – cbbL was detected. Autotrophic carbon fixation via the rTCA, 3‐HP and 3‐HP/4H‐B cycles could not be confirmed. The detected APS reductase alpha subunit – aprA sequences were phylogenetically related to sequences of sulfate‐reducing bacteria belonging to Clostridia, and also to sequences of chemolithoautothrophic sulfur‐oxidizing bacteria belonging to Betaproteobacteria. Sequences of methyl coenzyme M reductase – mcrA were phylogenetically affiliated to sequences belonging to Anaerobic Methanotroph group 1 (ANME‐1). The populations found and the functional key markers detected in CVA suggest that metabolisms related to H₂, methane and/or sulfur may be the major driving forces in this environment.     

Microbial characterization of toluene-degrading denitrifying consortia obtained from terrestrial and marine ecosystems

The degradation characteristics of toluene coupled to nitrate reduction were investigated in enrichment culture and the microbial communities of toluene-degrading denitrifying consortia were characterized by denaturing gradient gel electrophoresis (DGGE) technique. Anaerobic nitrate-reducing bacteria were enriched from oil-contaminated soil samples collected from terrestrial (rice field) and marine (tidal flat) ecosystems. Enriched consortia degraded toluene in the presence of nitrate as a terminal electron acceptor. The degradation rate of toluene was affected by the initial substrate concentration and co-existence of other hydrocarbons. The types of toluene-degrading denitrifying consortia depended on the type of ecosystem. The clone RS-7 obtained from the enriched consortium of the rice field was most closely related to a toluene-degrading and denitrifying bacterium, Azoarcus denitrificians (A. tolulyticus sp. nov.). The clone TS-11 detected in the tidal flat enriched consortium was affiliated to Thauera sp. strain S2 (T. aminoaromatica sp. nov.) that was able to degrade toluene under denitrifying conditions. This indicates that environmental factors greatly influence microbial communities obtained from terrestrial (rice field) and marine (tidal flat) ecosystems.     

Abundance,community structure and diversity of nitrifying bacterial enrichments from low and high saline brackishwater environments

The study reports diversity in nitrifying microbial enrichments from low (0·5–5‰) and high (18–35‰) saline ecosystems. Microbial community profiling of ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB) enrichments was analysed by sequencing 16S rRNA and was processed using Mothur pipeline. The α-diversity indices showed the richness of nitrifying bacterial consortia from the high saline environment and were clustering based on the source of the sample. AOB and NOB enrichments from both the environments showed diverse lineages of phyla distributed in both groups with 38 and 34 phyla from low saline and 53 and 40 phyla in high saline sources, respectively. At class level, α- and γ-proteobacteria were found to be more dominant in both the enrichments. AOBs and NOBs in enrichments from low saline environments were dominated by Nitrosomonadaceae, Gallionellaceae (Nitrotoga sp.) and Ectothiorhodospiraceae and Nitrospira, respectively. Though Chromatiaceae were present in both AOB and NOB enrichments, Nitrosoglobus and Nitrosococcus dominated the AOBs while NOBs were dominated by uncultured genera, whereas Rhizobiales were found in both the enrichments. AOBs and NOBs in enrichments from high saline environments were dominated by Nitrospira-like AOBs, Nitrosomonas and Nitrosococcus genera, whereas ammonia-oxidizing archaea (AOA) group included Nitrosopumilus and Nitrososphaera genera comprising and Nitrospirae, respectively. The majority of the genera obtained in both the salinities were found to be either uncultured or unclassified groups. Results of the study suggest that the AOB and NOB consortia have unique and diverse microbes in each of the enrichments, capable of functioning in aquaculture systems practised at different salinities (0–60 ppt).     

Novel multispecies microbial consortia involved in lignocellulose and 5-hydroxymethylfurfural bioconversion

To develop a targeted metagenomics approach for the analysis of novel multispecies microbial consortia involved in the bioconversion of lignocellulose and furanic compounds, we applied replicated sequential batch aerobic enrichment cultures with either pretreated or untreated wheat straw as the sources of carbon and energy. After each transfer, exponential growth of bacteria was detected using microscopic cell counts, indicating that the substrate was being utilized. In batch, the final bacterial abundances increased from an estimated 5 to 8.7–9.5 log 16S rRNA gene copy numbers/ml. The abundances of fungal propagules showed greater variation, i.e., between 5.4 and 8.0 log ITS1 copies/ml. Denaturing gradient gel electrophoresis analyses showed that the bacterial consortia in both treatments reached approximate structural stability after six transfers. Moreover, the structures of the fungal communities were strongly influenced by substrate treatment. A total of 124 bacterial strains were isolated from the two types of enrichment cultures. The most abundant strains were affiliated with the genera Raoultella/Klebsiella, Kluyvera, Citrobacter, Enterobacter, Pseudomonas, Acinetobacter, Flavobacterium and Arthrobacter. Totals of 43 and 11 strains obtained from the untreated and pretreated substrates, respectively, showed (hemi)cellulolytic activity (CMC-ase and xylanase), whereas 96 strains were capable of growth in 7.5 mM 5-hydroxymethylfurfural. About 50 % of the latter showed extracellular oxidoreductase activity as detected by a novel iodide oxidation method. Also, (hemi)cellulolytic fungal strains related to Coniochaeta, Plectosphaerella and Penicillium were isolated. One Trichosporon strain was isolated from pretreated wheat straw. The two novel bacterial–fungal consortia are starting points for lignocellulose degradation applications.     

Enumeration and Characterization of Iron(III)-Reducing Microbial Communities from Acidic Subsurface Sediments Contaminated with Uranium(VI)

Iron(III)-reducing bacteria have been demonstrated to rapidly catalyze the reduction and immobilization of uranium(VI) from contaminated subsurface sediments. Thus, these organisms may aid in the development of bioremediation strategies for uranium contamination, which is prevalent in acidic subsurface sediments at U.S. government facilities. Iron(III)-reducing enrichment cultures were initiated from pristine and contaminated (high in uranium, nitrate; low pH) subsurface sediments at pH 7 and pH 4 to 5. Enumeration of Fe(III)-reducing bacteria yielded cell counts of up to 240 cells ml⁻¹ for the contaminated and background sediments at both pHs with a range of different carbon sources (glycerol, acetate, lactate, and glucose). In enrichments where nitrate contamination was removed from the sediment by washing, MPN counts of Fe(III)-reducing bacteria increased substantially. Sediments of lower pH typically yielded lower counts of Fe(III)-reducing bacteria in lactate- and acetate-amended enrichments, but higher counts were observed when glucose was used as an electron donor in acidic enrichments. Phylogenetic analysis of 16S rRNA gene sequences extracted from the highest positive MPN dilutions revealed that the predominant members of Fe(III)-reducing consortia from background sediments were closely related to members of the Geobacteraceae family, whereas a recently characterized Fe(III) reducer (Anaeromyxobacter sp.) and organisms not previously shown to reduce Fe(III) (Paenibacillus and Brevibacillus spp.) predominated in the Fe(III)-reducing consortia of contaminated sediments. Analysis of enrichment cultures by terminal restriction fragment length polymorphism (T-RFLP) strongly supported the cloning and sequencing results. Dominant members of the Fe(III)-reducing consortia were observed to be stable over several enrichment culture transfers by T-RFLP in conjunction with measurements of Fe(III) reduction activity and carbon substrate utilization. Enrichment cultures from contaminated sites were also shown to rapidly reduce millimolar amounts of U(VI) in comparison to killed controls. With DNA extracted directly from subsurface sediments, quantitative analysis of 16S rRNA gene sequences with MPN-PCR indicated that Geobacteraceae sequences were more abundant in pristine compared to contaminated environments,whereas Anaeromyxobacter sequences were more abundant in contaminated sediments. Thus, results from a combination of cultivation-based and cultivation-independent approaches indicate that the abundance/community composition of Fe(III)-reducing consortia in subsurface sediments is dependent upon geochemical parameters (pH, nitrate concentration) and that microorganisms capable of producing spores (gram positive) or spore-like bodies (Anaeromyxobacter) were representative of acidic subsurface environments.     

A Novel Community of Acidophiles in an Acid Mine Drainage Sediment

A sediment sample (pH 2.5) was collected at an acid mine drainage site in Anhui, China. The present acidophilic microbial community in the sediment was studied with a 16S rRNA gene clone library. Small-subunit rRNA genes were PCR amplified, cloned and screened by amplified rDNA restriction analysis (ARDRA). Subsequently, 10 different clones were identified and they were affiliated with Acidobacteria, β/γ-Proteobacteria, δ-Proteobacteria, Nitrospira, Candidate division TM7, and Low G + C Gram-positives. Phylogenetic analysis of 16S rRNA gene sequences revealed a diversity of acidophiles in the sediment that were mostly novel. Unexpectedly, 16S rRNA gene sequences affiliated with δ-Proteobacteria were found to constitute more than 60% of clone library. To our knowledge, this is the first occasion that bacteria of δ-Proteobacteria have been found dominant in the acidic habitat. Anaerobic sulfate- or metal reduction is the predominant physiological trait of bacteria of this subdivision. The high sulfate, ferric iron and the presence of bioavailable carbon in the anaerobic microenvironment may result in the dominance of bacteria of δ-Proteobacteria.     

Diversity and abundance of aerobic and anaerobic ammonium-oxidizing bacteria in freshwater sediments of the Xinyi River (China)

Here we report on the biodiversity and abundance of aerobic and anaerobic ammonium-oxidizing bacteria in sediment samples from the Xinyi River, Jinagsu Province (China). The biodiversity of aerobic ammonium-oxidizing bacteria in the sediment was assessed using the amoA gene as functional marker. The retrieved amoA clones were affiliated to environmental sequences from freshwater habitats. The closest cultivated relative was Nitrosomonas urea. Anaerobic ammonium-oxidizing (anammox) bacteria were studied using anammox and planctomycete-specific 16S rRNA gene primers. The sediments contained 16S rRNA genes and bacterial cells closely related to the known anammox bacterium Candidatus'Brocadia anammoxidans'. Anaerobic continuous flow reactors were set up to enrich anammox organisms from the sediments. After an adaptation period of about 25 days the reactors started to consume ammonium and nitrite, indicating that the anammox reaction was occurring with a rate of 41-58 nmol cm(-3) h(-1). Community analysis of the enrichments by quantitative fluorescence in situ hybridization showed an increase in the abundance of anammox bacteria from < 1% to 6 +/- 2% of the total population. Analysis of the 16S rRNA genes showed that the enriched anammox organisms were related to the Candidatus'Scalindua' genus.     

Hydrolysis of lignocellulose by anaerobic fungi produces free sugars and organic acids for two-stage fine chemical production with Kluyveromyces marxianus

Development of the bioeconomy is driven by our ability to access the energy-rich carbon trapped in recalcitrant plant materials. Current strategies to release this carbon rely on expensive enzyme cocktails and physicochemical pretreatment, producing inhibitory compounds that hinder subsequent microbial bioproduction. Anaerobic fungi are an appealing solution as they hydrolyze crude, untreated biomass at ambient conditions into sugars that can be converted into value-added products by partner organisms. However, some carbon is lost to anaerobic fungal fermentation products. To improve efficiency and recapture this lost carbon, we built a two-stage bioprocessing system pairing the anaerobic fungus Piromyces indianae with the yeast Kluyveromyces marxianus, which grows on a wide range of sugars and fermentation products. In doing so we produce fine and commodity chemicals directly from untreated lignocellulose. P. indianae efficiently hydrolyzed substrates such as corn stover and poplar to generate sugars, fermentation acids, and ethanol, which K. marxianus consumed while producing 2.4 g/L ethyl acetate. An engineered strain of K. marxianus was also able to produce 550 mg/L 2-phenylethanol and 150 mg/L isoamyl alcohol from P. indianae hydrolyzed lignocellulosic biomass. Despite the use of crude untreated plant material, production yields were comparable to optimized rich yeast media due to the use of all available carbon including organic acids, which formed up to 97% of free carbon in the fungal hydrolysate. This work demonstrates that anaerobic fungal pretreatment of lignocellulose can sustain the production of fine chemicals at high efficiency by partnering organisms with broad substrate versatility.     

The nexus of syntrophy‐associated microbiota in anaerobic digestion revealed by long‐term enrichment and community survey

Anaerobic digestion (AD) processes are known to effectively convert organic waste to CO₂ and CH₄, but much of the microbial ecology remains unclear. Specifically, we have limited insights into symbiotic syntroph and methanogen (‘syntrophy’) acid degradation, although they are essential for preventing process deterioration. Also, we often observed many uncharacterized or uncultivated organisms, but poorly understood their role(s) in relation to syntrophy. To define syntrophy‐associated populations, this study enriched methanogenic communities with propionate, butyrate, benzoate, acetate, formate and H₂ from two different inocula over 3 years. 16S pyrotag analysis revealed core populations of known syntrophs (six clades) and methanogens (nine clades) associated with acid degradation, and evidence for substrate‐ and/or inoculum‐dependent specificity in syntrophic partnerships. Based on comprehensive re‐evaluation of publically available microbial community data for AD, the known syntrophs and methanogens identified were clearly representatives of the AD‐associated syntrophs and methanogens. In addition, uncultivated clades related to Bacteroidetes, Firmicutes, Actinobacteria and Chloroflexi were ubiquitously found in AD and enrichments. These organisms may be universally involved in AD syntrophic degradation, but only represented <23% of the yet‐to‐be‐cultivated organisms (89 of 390 clades). Thus, the contribution of these uncultured organisms in AD remains unclear and warrants further investigation.     

Enumeration and characterization of iron(III)-reducing microbial communities from acidic subsurface sediments contaminated with uranium(VI)

Iron(III)-reducing bacteria have been demonstrated to rapidly catalyze the reduction and immobilization of uranium(VI) from contaminated subsurface sediments. Thus, these organisms may aid in the development of bioremediation strategies for uranium contamination, which is prevalent in acidic subsurface sediments at U.S. government facilities. Iron(III)-reducing enrichment cultures were initiated from pristine and contaminated (high in uranium, nitrate; low pH) subsurface sediments at pH 7 and pH 4 to 5. Enumeration of Fe(III)-reducing bacteria yielded cell counts of up to 240 cells ml(-1) for the contaminated and background sediments at both pHs with a range of different carbon sources (glycerol, acetate, lactate, and glucose). In enrichments where nitrate contamination was removed from the sediment by washing, MPN counts of Fe(III)-reducing bacteria increased substantially. Sediments of lower pH typically yielded lower counts of Fe(III)-reducing bacteria in lactate- and acetate-amended enrichments, but higher counts were observed when glucose was used as an electron donor in acidic enrichments. Phylogenetic analysis of 16S rRNA gene sequences extracted from the highest positive MPN dilutions revealed that the predominant members of Fe(III)-reducing consortia from background sediments were closely related to members of the Geobacteraceae family, whereas a recently characterized Fe(III) reducer (Anaeromyxobacter sp.) and organisms not previously shown to reduce Fe(III) (Paenibacillus and Brevibacillus spp.) predominated in the Fe(III)-reducing consortia of contaminated sediments. Analysis of enrichment cultures by terminal restriction fragment length polymorphism (T-RFLP) strongly supported the cloning and sequencing results. Dominant members of the Fe(III)-reducing consortia were observed to be stable over several enrichment culture transfers by T-RFLP in conjunction with measurements of Fe(III) reduction activity and carbon substrate utilization. Enrichment cultures from contaminated sites were also shown to rapidly reduce millimolar amounts of U(VI) in comparison to killed controls. With DNA extracted directly from subsurface sediments, quantitative analysis of 16S rRNA gene sequences with MPN-PCR indicated that Geobacteraceae sequences were more abundant in pristine compared to contaminated environments,whereas Anaeromyxobacter sequences were more abundant in contaminated sediments. Thus, results from a combination of cultivation-based and cultivation-independent approaches indicate that the abundance/community composition of Fe(III)-reducing consortia in subsurface sediments is dependent upon geochemical parameters (pH, nitrate concentration) and that microorganisms capable of producing spores (gram positive) or spore-like bodies (Anaeromyxobacter) were representative of acidic subsurface environments.     

Isolation of sulfate‐reducing bacteria from deep sediment layers of the pacific ocean

Bacterial populations exist at great depths in marine sediments, but little is known about the type and characteristics of organisms in this unique bacterial environment. Cascadia Margin sediments from the Pacific Ocean have deep bacterial activity and bacterial populations, which are stimulated around a gas hydrate zone (215–225 m below sea floor [mbsf]). Bacterial sulfate reduction is the dominant anaerobic process within these sediments, and the depth distribution of sulfate‐reducing activity corresponds with distributions of viable sulfate‐reducing bacteria (SRB). Anaerobically stored sediments from this site were used to isolate sulfate‐reducing bacteria using a temperature‐gradient system, elevated pressure and temperatures, different media, and a range of growth substrates. A variety of enrichments on lactate were obtained from 0.5 and 222 mbsf, with surprisingly more rapid growth from the deeper sediments. The temperature range of enrichments producing strong growth from 222 mbsf was markedly wider than those from the near surface sediment (15–45°C and 9–19°C, respectively). This presumably reflects a temperature increase in deeper sediments. Only a few of these enrichments were successfully isolated due to very slow or no growth on subculture, despite the use of a wide range of different media and growth conditions. Psychrophilic and mesophilic sulfate‐reducing isolates were obtained from 0.5 m depth. As the minimum growth temperature of the mesophile (probably a Desulfotomaculum sp.) was above the in situ temperature of 3°C, it must have been present in the sediment as spores. A larger number of isolates (23) was obtained from 222 mbsf, and these barophilic SRB were closely related (based on 16S rRNA gene analysis), but not identical to, Desulfovibrio profundus, recently isolated from deep sediments from the Japan Sea. Bacteria related to D. profundus may be widespread in deep marine sediments.     

Identities of Epilithic Hydrocarbon-Utilizing Diazotrophic Bacteria from the Arabian Gulf Coasts, and Their Potential for Oil Bioremediation Without Nitrogen Supplementation

Gravel particles from four sites along the Arabian Gulf coast in autumn, winter, and spring were naturally colonized with microbial consortia containing between 7 and 400 × 10² cm⁻² of cultivable oil-utilizing bacteria. The 16S rRNA gene sequences of 70 representatives of oil-utilizing bacteria revealed that they were predominantly affiliated with the Gammaproteobacteria and the Actinobacteria. The Gammaproteobacteria comprised among others, the genera Pseudomonas, Pseudoalteromonas, Shewanella, Marinobacter, Psychrobacter, Idiomarina, Alcanivorax, Cobetia, and others. Actinobacteria comprised the genera Dietzia, Kocuria, Isoptericola, Rhodococcus, Microbacterium, and others. In autumn, Firmicutes members were isolated from bay and nonbay stations while Alphaproteobacteria were detected only during winter from Anjefa bay station. Fingerprinting by denaturing gradient gel electrophoresis of amplified 16S rRNA genes of whole microbial consortia confirmed the culture-based bacterial diversities in the various epilithons in various sites and seasons. Most of the representative oil-utilizing bacteria isolated from the epilithons were diazotrophic and could attenuate oil also in nitrogen-rich (7.9–62%) and nitrogen-free (4–54%) cultures, which, makes the microbial consortia suitable for oil bioremediation in situ, without need for nitrogen supplementation. This was confirmed in bench-scale experiments in which unfertilized oily seawater was bioremediated by epilithon-coated gravel particles.     

Ovine Ruminal Microbes Are Capable of Biotransforming Hexahydro-1,3,5-Trinitro-1,3,5-Triazine (RDX)

Bioremediation is of great interest in the detoxification of soil contaminated with residues from explosives such as hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX). Although there are numerous forms of in situ and ex situ bioremediation, ruminants would provide the option of an in situ bioreactor that could be transported to the site of contamination. Bovine rumen fluid has been previously shown to transform 2,4,6-trinitrotoluene (TNT), a similar compound, in 4 h. In this study, RDX incubated in whole ovine rumen fluid was nearly eliminated within 4 h. Whole ovine rumen fluid was then inoculated into five different types of media to select for archaeal and bacterial organisms capable of RDX biotransformation. Cultures containing 30 μg mL⁻¹ RDX were transferred each time the RDX concentration decreased to 5 μg mL⁻¹ or less. Time point samples were analyzed for RDX biotransformation by HPLC. The two fastest transforming enrichments were in methanogenic and low nitrogen basal media. After 21 days, DNA was extracted from all enrichments able to partially or completely transform RDX in 7 days or less. To understand microbial diversity, 16S rRNA-gene-targeted denaturing gradient gel electrophoresis (DGGE) fingerprinting was conducted. Cloning and sequencing of partial 16S rRNA fragments were performed on both low nitrogen basal and methanogenic media enrichments. Phylogenetic analysis revealed similar homologies to eight different bacterial and one archaeal genera classified under the phyla Firmicutes, Actinobacteria, and Euryarchaeota. After continuing enrichment for RDX degraders for 1 year, two consortia remained: one that transformed RDX in 4 days and one which had slowed after 2 months of transfers without RDX. DGGE comparison of the slower transforming consortium to the faster one showed identical banding patterns except one band. Homology matches to clones from the two consortia identified the same uncultured Clostridia genus in both; Sporanaerobacter acetigenes was identified only in the consortia able to completely transform RDX. This is the first study to examine the rumen as a potential bioremediation tool for soils contaminated with RDX, as well as to discover S. acetigenes in the rumen and its potential ability to metabolize this energetic compound.     

Iron Cycling Potentials of Arsenic Contaminated Groundwater in Bangladesh as Revealed by Enrichment Cultivation

The activities of iron-oxidizing and reducing microorganisms impact the fate of arsenic in groundwater. Phylogenetic information cannot exclusively be used to infer the potential for iron oxidation or reduction in aquifers. Therefore, we complemented a previous cultivation-independent microbial community survey covering 22 arsenic contaminated drinking water wells in Bangladesh, with the characterization of enrichments of microaerophilic iron oxidizers and anaerobic iron reducers, conducted on the same water samples. All investigated samples revealed a potential for microbial iron oxidation and reduction. Microbial communities were phylogenetically diverse within and between enrichments as was also observed in the previous cultivation-independent analysis of the water samples from which these enrichments were derived. Enrichment uncovered a larger diversity in iron-cycling microorganisms than previously indicated. The iron-reducing enrichments revealed the presence of several 16S ribosomal RNA (16S rRNA) gene sequences most closely related to Acetobacterium, Clostridium, Bacillus, Rhizobiales, Desulfovibrio, Bacteroides, and Spirochaetes, in addition to well-known dissimilatory iron-reducing Geobacter and Geothrix species. Although a large diversity of Geobacteraceae was observed, they comprised only a small part of the iron-reducing consortia. Iron-oxidizing gradient tube enrichments were dominated by Comamonadaceae and Rhodocyclaceae instead of Gallionellaceae. Forty-five percent of these enrichments also revealed the presence of the gene encoding arsenite oxidase, which converts arsenite to less toxic and less mobile arsenate. Their potential for ferric (oxyhydr)oxides precipitation and arsenic immobilization makes these iron-oxidizing enrichments of interest for rational bioaugmentation of arsenite contaminated groundwater.     

Interactions in Syntrophic Associations of Endospore-Forming, Butyrate-Degrading Bacteria and H2-Consuming Bacteria

Butyrate is an important intermediate in the anaerobic degradation of organic matter. In sulfate-depleted environments butyrate is oxidized to acetate and hydrogen by obligate proton reducers, in syntrophic association with hydrogen-consuming methanogens. This paper describes two enrichments of endospore-forming bacteria degrading butyrate in consortia with methanogens. The isolates are readily established in coculture with H₂-consuming, sulfate-reducing bacteria by pasteurizing the culture. The two original enrichments differed in that one grew to an optically dense culture while the second grew in clumps. Examination by scanning electron microscopy showed that clumping resulted from the production of large amounts of extracellular polymer. Several H₂-consuming methanogens were identified in the enrichments. Some of them grew closely associated to the butyrate degraders. This attachment to the hydrogen producer may permit some methanogens to compete for the growth substrate against other bacteria having higher substrate affinity.     

Synergistic growth in bacteria depends on substrate complexity

Both positive and negative interactions among bacteria take place in the environment. We hypothesize that the complexity of the substrate affects the way bacteria interact with greater cooperation in the presence of recalcitrant substrate. We isolated lignocellulolytic bacteria from salt marsh detritus and compared the growth, metabolic activity and enzyme production of pure cultures to those of three-species mixed cultures in lignocellulose and glucose media. Synergistic growth was common in lignocellulose medium containing carboxyl methyl cellulose, xylan and lignin but absent in glucose medium. Bacterial synergism promoted metabolic activity in synergistic mixed cultures but not the maximal growth rate (μ). Bacterial synergism also promoted the production of β-1,4-glucosidase but not the production of cellobiohydrolase or β-1,4-xylosidase. Our results suggest that the chemical complexity of the substrate affects the way bacteria interact. While a complex substrate such as lignocellulose promotes positive interactions and synergistic growth, a labile substrate such as glucose promotes negative interactions and competition. Synergistic interactions among indigenous bacteria are suggested to be important in promoting lignocellulose degradation in the environment.     

Dominant ectosymbiotic bacteria of cellulolytic protists in the termite gut also have the potential to digest lignocellulose

Wood‐feeding lower termites harbour symbiotic gut protists that support the termite nutritionally by degrading recalcitrant lignocellulose. These protists themselves host specific endo‐ and ectosymbiotic bacteria, functions of which remain largely unknown. Here, we present draft genomes of a dominant, uncultured ectosymbiont belonging to the order Bacteroidales, ‘Candidatus Symbiothrix dinenymphae’, which colonizes the cell surface of the cellulolytic gut protists Dinenympha spp. We analysed four single‐cell genomes of Ca. S. dinenymphae, the highest genome completeness was estimated to be 81.6–82.3% with a predicted genome size of 4.28–4.31 Mb. The genome retains genes encoding large parts of the amino acid, cofactor and nucleotide biosynthetic pathways. In addition, the genome contains genes encoding various glycoside hydrolases such as endoglucanases and hemicellulases. The genome indicates that Ca. S. dinenymphae ferments lignocellulose‐derived monosaccharides to acetate, a major carbon and energy source of the host termite. We suggest that the ectosymbiont digests lignocellulose and provides nutrients to the host termites, and hypothesize that the hydrolytic activity might also function as a pretreatment for the host protist to effectively decompose the crystalline cellulose components.     

Cellulolytic bacteria from soils in harsh environments

It is believed that the exposure of organisms to harsh climate conditions may select for differential enzymatic activities, making the surviving organisms a very promising source for bioprospecting. Soil bacteria play an important role in degradation of organic matter, which is mostly due to their ability to decompose cellulose-based materials. This work focuses on the isolation and identification of cellulolytic bacteria from soil found in two environments with stressful climate conditions (Antarctica and the Brazilian semi-arid caatinga). Cellulolytic bacteria were selected using enrichments at high and low temperatures (4 or 60°C) in liquid media (trypic soy broth—TSB and minimum salt medium—MM) supplemented with cellulose (1%). Many of the isolates (119 out of 254—46.9%) displayed the ability to degrade carboxymethyl-cellulose, indicating the presence of endoglucolytic activity, while only a minority of these isolates (23 out of 254—9.1%) showed exoglucolytic activity (degradation of avicel). The obtained isolates revealed a preferential endoglucolytic activity according to the temperature of enrichments. Also, the identification of some isolates by partial sequencing of the 16S rRNA gene indicated that the Bacteroidetes (e.g., Pedobacter, Chryseobacterium and Flavobacterium) were the main phylum of cellulolytic bacteria isolated from soil in Antarctica; the Firmicutes (e.g., Bacillus) were more commonly isolated from samples from the caatinga; and Actinobacteria were found in both types of soil (e.g., Microbacterium and Arthrobacter). In conclusion, this work reports the isolation of bacteria able to degrade cellulose-based material from soil at very low or very high temperatures, a finding that should be further explored in the search for cellulolytic enzymes to be used in the bioenergy industry.     

Onset and establishment of diazotrophs and other bacterial associates in the early life history stages of the coral Acropora millepora

Early establishment of coral–microbial symbioses is fundamental to the fitness of corals, but comparatively little is known about the onset and succession of bacterial communities in their early life history stages. In this study, bacterial associates of the coral Acropora millepora were characterized throughout the first year of life, from larvae and 1‐week‐old juveniles reared in laboratory conditions in the absence of the dinoflagellate endosymbiont Symbiodinium to field‐outplanted juveniles with established Symbiodinium symbioses, and sampled at 2 weeks and at 3, 6 and 12 months. Using an amplicon pyrosequencing approach, the diversity of both nitrogen‐fixing bacteria and of bacterial communities overall was assessed through analysis of nifH and 16S rRNA genes, respectively. The consistent presence of sequences affiliated with diazotrophs of the order Rhizobiales (23–58% of retrieved nifH sequences; 2–12% of 16S rRNA sequences), across all samples from larvae to 12‐month‐old coral juveniles, highlights the likely functional importance of this nitrogen‐fixing order to the coral holobiont. Dominance of Roseobacter‐affiliated sequences (>55% of retrieved 16S rRNA sequences) in larvae and 1‐week‐old juveniles, and the consistent presence of sequences related to Oceanospirillales and Altermonadales throughout all early life history stages, signifies their potential importance as coral associates. Increased diversity of bacterial communities once juveniles were transferred to the field, particularly of Cyanobacteria and Deltaproteobacteria, demonstrates horizontal (environmental) uptake of coral‐associated bacterial communities. Although overall bacterial communities were dynamic, bacteria with likely important functional roles remain stable throughout early life stages of Acropora millepora.