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High microbial diversity promotes soil ecosystem functioning.
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High microbial diversity promotes soil ecosystem functioning.

Appl Environ Microbiol. 2018 Feb 16;:

Authors: Maron PA, Sarr A, Kaisermann A, Lévêque J, Mathieu O, Guigue J, Karimi B, Bernard L, Dequiedt S, Terrat S, Chabbi A, Ranjard L

Abstract
In soil, the link between microbial diversity and carbon transformations is challenged by the concept of functional redundancy. Here, we hypothesized that functional redundancy may decrease with increasing carbon source recalcitrance, and that coupling of diversity with C-cycling may change accordingly. We manipulated microbial diversity to examine how diversity decrease affects the decomposition of easily degradable (i.e. allochthonous plant residues) vs. recalcitrant (i.e. autochthonous organic matter) C-sources. We found that a decrease in microbial diversity (i) affected the decomposition of both autochthonous and allochthonous carbon sources hence reducing global CO2 emission by up to 40%, and (ii) shaped the source of CO2 emission towards preferential decomposition of most degradable C-sources. Our results also revealed that the significance of the "diversity effect" increases with nutrient availability. Altogether, these findings show that C-cycling in soil may be more vulnerable to microbial diversity changes than expected from previous studies, particularly in ecosystems exposed to nutrient inputs. Thus concern about the preservation microbial diversity may be highly relevant in the current 'global changes' context assumed to impact soil biodiversity and the pulse inputs of plant residues and rhizodeposits into the soil.IMPORTANCE With hundreds of thousands of taxa per gram of soil, microbial diversity dominates soil biodiversity. While numerous studies have established that microbial communities respond rapidly to environmental changes, the relationship between microbial diversity and soil functioning remains controversial. Using a well-controlled laboratory approach, we provide empirical evidence that microbial diversity may be of high significance for organic matter decomposition, a major process on which rely many of the ecosystem services provided by the soil ecosystem. These new findings should be taken into account in future studies aimed to understand and predict the functional consequences of changes in microbial diversity on soil ecosystem services and carbon storage in soil.

PMID: 29453268 [PubMed - as supplied by publisher]




Plant-microbe and abiotic factors influencing Salmonella survival and growth on alfalfa sprouts and Swiss chard microgreens.
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Plant-microbe and abiotic factors influencing Salmonella survival and growth on alfalfa sprouts and Swiss chard microgreens.

Appl Environ Microbiol. 2018 Feb 16;:

Authors: Reed E, Ferreira CM, Bell R, Brown EW, Zheng J

Abstract
Microgreens, like sprouts, are relatively fast growing, and generally consumed raw. Moreover, as observed in sprouts, microbial contamination from preharvest sources could also be present in the production of microgreens. In this study, two Salmonella enterica serovars (Hartford and Cubana) applied, at multiple inoculation levels, were evaluated for survival and growth on alfalfa sprouts and Swiss chard microgreens using the most probable number (MPN) method. Various abiotic factors were also examined for their effects on Salmonella survival and growth on sprouts and microgreens. Community level physiological profiles (CLPP) of sprout/microgreen rhizospheres with different levels of S. enterica inoculation at different growth stage were characterized with Biolog EcoPlates. In the seed contamination group, the ability of S. enterica to grow on sprouting alfalfa seeds was affected by both seed storage time and inoculation level, but not serovar. However, the growth of S. enterica on Swiss chard microgreens was affected by serovar, and inoculation level. Seed storage time had little effect on the average level of Salmonella population in microgreens. In the irrigation water contamination group, the growth of Salmonella in both alfalfa sprouts and microgreens was largely affected by inoculation level. Surprisingly, growing medium was found to play an important role in Salmonella survival and growth in microgreens. CLPP analysis showed significant changes in the microbial community metabolic diversity during sprouting for alfalfa sprouts, but few temporal changes were seen in microgreens. The data suggest that the change in rhizosphere bacterial functional diversity was host-dependent but independent of Salmonella contamination.IMPORTANCE: Sprouts and microgreens are considered "functional foods", containing health promoting or disease preventing properties in addition to normal nutritional values. However, the microbial risk associated with microgreens has not been well-studied. Compared with sprouts, this study evaluated Salmonella survival and growth on microgreens, and other abiotic factors that could affect Salmonella survival and growth on microgreens. This work provides baseline data to risk assessment of microbial contamination in sprouts and microgreens. Understanding the risks of Salmonella contamination and its effects on rhizosphere microbial communities enables a better understanding of host-pathogen dynamics in sprouts and microgreens. The data also contributes to innovative preventive control strategies for Salmonella contamination in sprouts and microgreens.

PMID: 29453267 [PubMed - as supplied by publisher]




The gills of reef fish support a distinct microbiome influenced by host-specific factors.
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The gills of reef fish support a distinct microbiome influenced by host-specific factors.

Appl Environ Microbiol. 2018 Feb 16;:

Authors: Pratte ZA, Besson M, Hollman RD, Stewart FJ

Abstract
Teleost fish represent the most diverse of the vertebrate groups and play important roles in food webs, as ecosystem engineers, and as vectors for microorganisms. However, the microbial ecology of fishes remains underexplored for most host taxa, and for certain niches on the fish body. This is particularly true for the gills, the key sites for respiration and waste exchange in fishes. Here, we provide a comprehensive analysis of the gill microbiome. We focus on ecologically diverse taxa from coral reefs around Moorea, sampling the gill and intestines of adults and juveniles representing 15 families. Gill microbiome composition differed significantly from that of the gut in both adults and juveniles, with fish-associated niches having lower alpha diversity and higher beta diversity compared to seawater, sediment, and algae-associated microbiomes. Of ∼45,000 operational taxonomic units (OTUs) detected across all samples, 11% and 13% were detected only in the gill and intestine, respectively. OTUs most enriched in the gill included members of the gammaproteobacterial genus Shewanella and family Endozoicimonaceae. In adult fish, both gill and intestinal microbiomes varied significantly among host species grouped by diet category. Gill and intestinal microbiomes from the same individual were more similar to one another compared to gill and intestinal microbiomes from different individuals. These results demonstrate that distinct body sites are jointly influenced by host-specific organizing factors operating at the level of the host individual. The results also identify taxonomic signatures unique to the gill and intestine, confirming fish-associated niches as distinct reservoirs of marine microbial diversity.Importance Fish breath and excrete waste through their gills. The gills are also potential sites of pathogen invasion and colonization by other microbes. However, we know little about the microbial communities that live on the gill and the factors shaping their diversity. Focusing on ecologically distinct types of coral reef fish, we provide a comprehensive analysis of the fish gill microbiome. By comparison to microbiomes of the gut and the surrounding environment, we identify microbes unique to the gill niche. These microbes may be targets for further studies to determine the contribution of the microbiome to waste exchange or host immunity. We also show that, despite exhibiting a unique taxonomic signature, the gill microbiome is influenced by factors that also influence the gut microbiome. These factors include the specific identity of the host individual. These results suggest basic principles describing how association with fishes structures the composition of microbial communities.

PMID: 29453266 [PubMed - as supplied by publisher]




Integrated Metabolomics and Morphogenesis Reveals Volatile Signaling of the Nematode-Trapping Fungus Arthrobotrys oligospora.
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Integrated Metabolomics and Morphogenesis Reveals Volatile Signaling of the Nematode-Trapping Fungus Arthrobotrys oligospora.

Appl Environ Microbiol. 2018 Feb 16;:

Authors: Wang BL, Chen YH, He JN, Xue HX, Yan N, Zeng ZJ, Bennett JW, Zhang KQ, Niu XM

Abstract
The adjustment of metabolic patterns is fundamental to fungal biology and plays vital roles in adaption to diverse ecological challenges. Nematode trapping fungi can switch lifestyles from saprophytic to pathogenic by developing specific trapping devices induced by nematodes to infect their prey as a response to nutrient depletion in nature. However, the chemical identity of the specific fungal metabolites used during the switch remains poorly understood. We hypothesized that these important signal molecules might be volatile in nature. GC-MS was used to carry out comparative analysis of fungal metabolomics during saprophytic and pathogenic lifestyles of the model species Arthrobotrys oligospora Two media commonly used in research on this species, corn meal agar (CMA) and potato dextrose agar (PDA), were chosen in this study. The fungus produced a small group of volatile furanone and pyrone metabolites that were associated with the switch from saprophytic to pathogenic stages. A. oligospora grown on CMA tended to produce more traps and employ attractive furanones to improve utilization of traps, while fungus grown on PDA developed fewer traps and used nematodetoxic furanone metabolites to compensate for insufficient traps. Another volatile pyrone metabolite, maltol, was identified as a morphological regulator for enhancing trap formation. Deletion of gene AOL_s00079g496 in A. oligospora led to increased furanone attractant (2 folds) in mutants and enhanced attractive activity (1.5 fold) of the fungus, while resulted in decreased trap formation. This investigation provides new insights regarding the comprehensive tactics of fungal adaptation to environmental stress, integrating both morphological and metabolomic mechanisms.Importance Nematode-trapping fungi are a unique group of soil-living fungi that can switch from saprophytic to pathogenic lifestyle once in contact with nematodes as a response to nutrient depletion. In this study, we investigated the metabolic response during the switch and the key types of metabolites involved in the interaction between fungi and nematodes. Our findings indicated that A. oligospora develop multiple and flexible metabolic tactics corresponding to different morphological responses to nematodes. A. oligospora can use similar volatile furanone and pyrone metabolites with different ecological functions to help capture nematodes in the fungal switch from saprophytic to pathogenic lifestyles. Furthermore, A. oligospora mutants with increased furanone and pyrone metabolites confirmed the results. This investigation reveals the importance of volatile signaling in the comprehensive tactics used by nematode trapping fungi, integrating both morphological and metabolomic mechanisms.

PMID: 29453265 [PubMed - as supplied by publisher]




Bacterial Community Shift and Coexisting/Coexcluding Patterns Revealed by Network Analysis in A Bioreduced Uranium Contaminated Site after Reoxidation.
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Bacterial Community Shift and Coexisting/Coexcluding Patterns Revealed by Network Analysis in A Bioreduced Uranium Contaminated Site after Reoxidation.

Appl Environ Microbiol. 2018 Feb 16;:

Authors: Li B, Wu WM, Watson DB, Cardenas E, Chao Y, Phillips DH, Mehlhorn T, Lowe K, Kelly SD, Li P, Tao H, Tiedje JM, Criddle CS, Zhang T

Abstract
A site in Oak Ridge, Tennessee, USA, has sediments that contain >3% iron oxides and is contaminated with uranium (U). The U(VI) was bioreduced to U(IV) and immobilized in situ through intermittent injections of ethanol. Then it was allowed to reoxidize via the invasion of low-pH (3.6-4.0), high-nitrate (up to 200 mM) groundwater back into the reduced zone for 1,383 days. To examine the biogeochemical response, high-throughput sequencing and network analysis were applied to characterize bacterial population shifts, as well as co-occurrence and co-exclusion patterns among microbial communities. Paired t-test indicated no significant changes of α-diversity for the bioactive wells. However, both nonmetric multidimensional scaling and analysis of similarity confirmed a significant distinction in the overall composition of the bacterial communities between the bioreduced and the reoxidized sediments. The top 20 major genera accounted for >70% of the cumulative contribution to the dissimilarity in the bacterial communities before and after the groundwater invasion. Castellaniella had the largest dissimilarity contribution (17.7%). For the bioactive wells, the abundance of the U(VI)-reducing genera Geothrix, Desulfovibrio, Ferribacterium, and Geobacter decreased significantly, whereas the denitrifying Acidovorax abundance increased significantly after groundwater invasion. Additionally, seven genera, i.e., Castellaniella, Ignavibacterium, Simplicispira, Rhizomicrobium, Acidobacteria Gp1, Acidobacteria Gp14 and Acidobacteria Gp23 were significant indicators of bioactive wells under reoxidation stage. Canonical correspondence analysis indicated that nitrate, manganese and pH affected mostly the U(VI)-reducing genera and indicator genera. Co-occurrence patterns among microbial taxa suggested the presence of taxa sharing similar ecological niches or mutualism/commensalism/synergism interactions.IMPORTANCE: High-throughput sequencing technology in combination with a network analysis approach were used to investigate the stabilization of uranium and the corresponding dynamics of bacterial communities at field conditions with regard to the heterogeneity and complexity of the subsurface over the long term. The study also examined diversity and microbial community composition shift, the common genera and indicator genera before and after long-term contaminated groundwater invasion, and the relationship between the target functional community structure and environmental factors. Additionally, deciphering co-occurrence and co-exclusion patterns among microbial taxa and environmental parameters could help predict potential biotic interactions (cooperation/competition), shared physiologies, or habitat affinities, thus, improving our understanding of ecological niches occupied by certain specific species. The above findings offer new insights into compositions of and associations among bacterial communities and serve as a foundation for future bioreduction implementation and monitoring efforts applied to uranium-contaminated sites.

PMID: 29453264 [PubMed - as supplied by publisher]




Enzyme activities of two recombinant heme-including peroxidases TvDyP1 and TvVP2 identified from the secretome of Trametes versicolor.
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Enzyme activities of two recombinant heme-including peroxidases TvDyP1 and TvVP2 identified from the secretome of Trametes versicolor.

Appl Environ Microbiol. 2018 Feb 16;:

Authors: Amara S, Perrot T, Navarro D, Deroy A, Benkhelfallah A, Chalak A, Daou M, Chevret D, Faulds CB, Berrin JG, Morel-Rouhier M, Gelhaye E, Record E

Abstract
Trametes versicolor is a wood inhabiting Agaricomycete known for its ability to cause strong white rot decay on hardwood and for its high tolerance toward phenolic compounds. The goal of the present work was to give insights on the molecular biology and biochemistry of heme-including class-II and dye-decolorizing peroxidases secreted from this fungus. Proteomic analysis of the secretome of T. versicolor BRFM1218 grown on oak wood revealed a set of 200 secreted proteins among which were a dye-decolorizing peroxidase TvDyP1 and a versatile peroxidase TvVP2. Both peroxidases were heterologously produced in E. coli, and were biochemically characterized and tested for their capacity to oxidize complex substrates. Both peroxidases were found to be active against several substrates in acidic conditions, and TvDyP1 was very stable in a relatively large range of pH (pH 2.0 to 6.0) while TvVP2 was more stable at pH 5.0-6.0 only. Thermostability of both enzymes was also tested and TvDyP1 was globally found to be more stable than TvVP2. After 180 min of incubation at T°C ranging from 30°C to 50°C, activities of TvVP2 drastically decreased retaining 10% to 30% of the its initial activity. In the same conditions, TvDyP1 retained 20% to 80% of enzyme activity. The two proteins were catalytically characterized and TvVP2 was shown to accept a wider range of reducing substrates than TvDyP1. Furthermore, both enzymes were found to be active against two flavonoids, quercetin and catechin, found in oak wood, TvVP2 displaying a more rapid oxidation of the two compounds. They were tested for their potential interest in dye decolourization of five industrial dyes and TvVP2 presented a wider oxidation and decolourization capacity towards the dye substrates than TvDyP1.IMPORTANCETrametes versicolor is a wood inhabiting Agaricomycete known for its ability to cause strong white rot decay on hardwood and for its high tolerance toward phenolic compounds. Among white-rot fungi, the basidiomycete T. versicolor has been extensively studied for its efficiency to degrade wood, and specifically lignin, thanks to an extracellular oxidative enzymatic system. The corresponding oxidative system was previously studied in several works for classical lignin and manganese peroxidases, and in this study, two new components of the oxidative system of T. versicolor, one dye-decoloririzing peroxidase and one versatile peroxidase were biochemically characterized in depth and compare to other fungal peroxidases.

PMID: 29453263 [PubMed - as supplied by publisher]




Insights into the fundamental physiology of the uncultured Fe-oxidizing bacterium Leptothrix ochracea.
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Insights into the fundamental physiology of the uncultured Fe-oxidizing bacterium Leptothrix ochracea.

Appl Environ Microbiol. 2018 Feb 16;:

Authors: Fleming EJ, Woyke T, Donatello AR, Kuypers MMM, Sczyrba A, Littmann S, Emerson D

Abstract
Leptothrix ochracea is known for producing large volumes of iron-oxyhydroxide sheaths that alter wetland biogeochemistry. For over a century, these delicate structures have fascinated microbiologists and geoscientists. Because L. ochracea still resists long-term in vitro culture, the debate regarding its metabolic classification dates back to 1885. We developed a novel culturing technique for L. ochracea using in situ natural waters, and coupled this with single cell genomics and nanoscale secondary ion mass spectrophotometry (nanoSIMS) to probe L. ochracea's physiology. In micro-slide cultures L. ochracea doubled every 5.7 hrs, had an absolute growth requirement for ferrous iron, had the genomic capacity for iron-oxidation, and a branched electron transport chain with cytochromes putatively involved in lithotrophic iron-oxidation. Additionally, its genome encoded several electron transport chain proteins including, a molybdopterin ACIII complex, a cytochrome bd oxidase reductase, and several terminal oxidase genes. L. ochracea contained two key autotrophic proteins in the Calvin Benson Bassham cycle, a Form II ribulose-bis-phosphate carboxylase and a phosphoribulose kinase. L. ochracea also assimilated bicarbonate, although calculations suggest bicarbonate assimilation is a small fraction of its total carbon assimilation. Finally, L. ochracea's fundamental physiology is a hybrid of the chemolithotrophic Gallionella-type iron-oxidizing bacteria and the sheathed, heterotrophic filamentous metal-oxidizing bacteria of the Leptothrix-Sphaerotilus genera. This allows L. ochracea to inhabit a unique niche within the neutrophilic iron seeps.ImportanceLeptothrix ochracea was one of three groups of organisms Sergei Winogradsky used in the 1880s to develop the hypothesis on chemolithotrophy. L. ochracea continues to resist cultivation and appears to have an absolute requirement for organic rich waters suggesting its true physiology remains unknown. Further, L. ochracea is an ecological engineer, a few L. ochracea cells can generate prodigious volumes of iron-oxyhydroxides changing the ecosystem geochemistry and ecology. Therefore, to determine Lochracea's basic physiology, we employed new single-cell techniques to demonstrate: L. ochracea oxidizes iron to generate energy and, despite predicted genes for autotrophic growth, L. ochacea assimilates a fraction of the total CO2 that autotrophs do. Although not a true chemolithoautotroph, L. ochracea's physiological strategy allows it to be flexible, extensively colonize iron-rich wetlands.

PMID: 29453262 [PubMed - as supplied by publisher]




Molecular and functional study of a branching sucrase-like glucansucrase reveals an evolutionary intermediate between two subfamilies of the GH70 enzymes.
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Molecular and functional study of a branching sucrase-like glucansucrase reveals an evolutionary intermediate between two subfamilies of the GH70 enzymes.

Appl Environ Microbiol. 2018 Feb 16;:

Authors: Yan M, Wang BH, Xu X, Chang P, Hang F, Wu Z, You C, Liu Z

Abstract
Family GH70 glucansucrases (GSs) catalyze the synthesis of α-glucans from sucrose, a reaction widely seen in lactic acid bacteria (LAB). These enzymes have been implicated in many aspects of the microbial life. Products of GSs have great commercial value as food supplements and medical materials, therefore these enzymes have attracted much attention from both fields of science and industry. However, certain issues concerning the origin and evolution of GS are still to be addressed, although an increasing number of GH70 enzymes have been characterized. This study described a GS enzyme with the appearance of a branching sucrase (BrS). Structure analysis indicated that this GS enzyme produces a type of glucan composed of α-(1→6) glucosidic backbone and α-(1→4) branches, as well as a considerable amount of α-(1→3) branches, distinguishing it from the GSs identified so far. Moreover, sequence-based analysis of the catalytic core of this enzyme suggested that it might be an evolutionary intermediate between the BrS and GS subgroups. These results provide an evolutionary link between these subgroups of GH70 enzymes and shed new light on the origination of GSs.IMPORTANCEFamily GH70 glucansucrases (GSs) catalyze the synthesis of α-glucans from sucrose, a reaction widely seen in lactic acid bacteria (LAB). Products of these enzymes have great commercial values as food supplements and medical materials. Moreover, these enzymes have attracted much attention from scientists as they have potentials in tailored synthesis of α-glucans with desired structures and properties. Although more and more GSs have been characterized, the origin and evolution of these enzymes has not been well addressed yet. This study described a glucansucrase with the appearance of a branching sucrase (BrS) (i.e. high similarity with BrS in sequence analysis). Further analysis indicated that this enzyme synthesized a type of insoluble glucan composed of α-(1→6) glucosidic backbone and a lot of α-(1→4) and α-(1→3) linked branches, the linkage composition of which has rarely been reported in the literature. This BrS-like GS enzyme might be an evolutionary intermediate between BrS and GS enzymes.

PMID: 29453261 [PubMed - as supplied by publisher]




A new cell extraction method reveals hemolymph microbiome of three aquatic invertebrates.
Related Articles A new cell extraction method reveals hemolymph microbiome of three aquatic invertebrates. Appl Environ Microbiol. 2018 Feb 16;: Authors: Zhang X, Sun Z, Zhang X, Zhang M, Li S Abstract Symbiotic microorganisms have been found in the hemolymph (blood) of many aquatic invertebrates, such as crabs, shrimps and oysters. Hemolymph is a critical site in host immune response. Currently, studies on hemolymph microorganisms are mostly performed with culture-dependent strategies using selective media (e.g., TCBS, 2216E, and LB) for enumerating and isolating microbial cells. However, doubts remain about the "true" representation of the microbial abundance and diversity of symbiotic microorganisms in hemolymph, particularly for uncultivable microorganisms which are believed to be more abundant than the cultured. To explore this, we developed a culture-independent cell extraction method for separating microbial cells from the hemolymph of three aquatic invertebrates (Scylla paramamosain, Litopenaeus vannamei, and Crassostrea angulata) involving filtration through a 5-μm mesh filter membrane (the Filtration Method). A combination of the Filtration Method with fluorescence microscopy and high-throughput sequencing technique provides insight into the abundances and diversity of the total microbiota in the hemolymph of these three invertebrates. More than 2.6 × 104 cells/mL of microbial cells dominated by Escherichia-Shigella and Halomonas, Photobacterium and Escherichia-Shigella, and Pseudoalteromonas and Arcobacter, were detected in the hemolymph of Scylla paramamosain, Litopenaeus vannamei, and Crassostrea angulata, respectively. A parallel study for investigating the hemolymph microbiome by comparing the Filtration Method and a culture-dependent method (the Plate Count Method), showed significantly higher microbial abundances (between 26 and 369-folds difference; P<0.05) and less biased community structures of the Filtration Method than those of the Plate Count Method. Furthermore, hemolymph of the three invertebrates harbored many potential pathogens, including Photobacterium, Arcobacter, and Vibrio Finally, the Filtration Method provides a solution that improves understanding of the metabolic functions of uncultivable hemolymph microorganisms (e.g., metagenomics) devoid of host hemocytes contamination.Importance Microorganisms are found in invertebrates' hemolymph, a critical site in host immune response. Currently, studies on hemolymph microorganisms are mostly performed with culture-dependent strategies. However, doubts remain about the "true" representation of hemolymph microbiome. This study developed a culture-independent cell extraction method that could separate microbial cells from the hemolymph of three aquatic invertebrates (S. paramamosain, L. vannamei, and C. angulata) based on filtration through a 5-μm mesh filter membrane (the Filtration Method). A combination of the Filtration Method with fluorescence microscopy and high-throughput sequencing technique provides insight into the abundances and diversity of the total microbiota in the hemolymph of these three invertebrates. Our results demonstrate that the hemolymph of aquatic invertebrates harbors a much higher microbial abundance and distinct microbial community composition than previously estimated. Furthermore, this work provides a less biased solution for studying the metabolic functions of uncultivable hemolymph microbiota devoid of host hemocytes contamination. PMID: 29453260 [PubMed - as supplied by publisher] [...]



Common hydraulic fracturing fluid additives alter the structure and function of anaerobic microbial communities.
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Common hydraulic fracturing fluid additives alter the structure and function of anaerobic microbial communities.

Appl Environ Microbiol. 2018 Feb 16;:

Authors: Mumford AC, Akob DM, Klinges JG, Cozzarelli IM

Abstract
The development of unconventional oil and gas (UOG) resources results in production of large volumes of wastewater containing a complex mixture of hydraulic fracturing chemical additives and components from the formation. Release of these wastewaters into the environment poses potential risks that are poorly understood. Microbial communities in stream sediments form the base of the food chain, and may serve as sentinels for changes in stream health. Iron reducing organisms have been shown to play a role in the biodegradation of a wide range of organic compounds, and to evaluate their response to UOG wastewater, we enriched anaerobic microbial communities from sediments collected upstream (background) and downstream (impacted) of an UOG wastewater injection disposal facility in the presence of hydraulic fracturing fluid (HFF) additives: guar gum, ethylene glycol, and two biocides, DBNPA and Bronopol. Iron reduction was significantly inhibited early in the incubations with the addition of biocides, whereas amendment with guar gum and ethylene glycol stimulated iron reduction relative to unamended controls. Changes in the microbial community structure were observed across all treatments, indicating the potential for even small amounts of UOG wastewater components to influence natural microbial processes. Microbial community structure differed between enrichments with background and impacted sediments, suggesting that impacted sediments may have been pre-conditioned by exposure to wastewater. These experiments demonstrated the potential for biocides to significantly decrease iron reduction rates immediately following a spill, and demonstrated how microbial communities previously exposed to UOG wastewater may be more resilient to additional spills.Importance:Organic components of UOG wastewater can alter microbial communities and biogeochemical processes, which could alter the rates of essential natural attenuation processes. These findings provide new insights into microbial responses following a release of UOG wastewaters, and are critical for identifying strategies for remediation and natural attenuation of impacted environments.

PMID: 29453259 [PubMed - as supplied by publisher]




Anthropogenic N deposition alters the composition of expressed class II fungal peroxidases.
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Anthropogenic N deposition alters the composition of expressed class II fungal peroxidases.

Appl Environ Microbiol. 2018 Feb 16;:

Authors: Entwistle EM, Romanowicz KJ, Argiroff WA, Freedman ZB, Morris JJ, Zak DR

Abstract
Here, we present evidence that ca. 20 yrs of experimental N deposition altered the composition of lignin-decaying class II peroxidases expressed by forest floor fungi, a response which has occurred concurrently with reductions in plant-litter decomposition and a rapid accumulation of soil organic matter. This finding suggests that anthropogenic N deposition has induced changes in the biological mediation of lignin-decay, the rate limiting step in plant litter decomposition. Thus, altered composition of transcripts for a critical gene that is associated with terrestrial C-cycling may explain increased soil C storage under long-term increases in anthropogenic N deposition.IMPORTANCEFungal class II peroxidases are enzymes that mediate the rate-limiting step in the decomposition of plant material, which involves the oxidation of lignin and other polyphenols. In field experiments, anthropogenic N deposition has increased soil C storage in forests, a result which could potentially arise from anthropogenic N-induced changes in the composition of class II peroxidases expressed by the fungal community. In this study, we have gained unique insight into how anthropogenic N deposition, a widespread agent of global change, affects the expression of a functional gene encoding an enzyme that plays a critical role in a biologically-mediated ecosystem process.

PMID: 29453258 [PubMed - as supplied by publisher]




A facile Arsenazo III based assay for monitoring rare earth element depletion from cultivation media of methanotrophic and methylotrophic bacteria.
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A facile Arsenazo III based assay for monitoring rare earth element depletion from cultivation media of methanotrophic and methylotrophic bacteria.

Appl Environ Microbiol. 2018 Feb 16;:

Authors: Hogendoorn C, Roszczenko-Jasińska P, Martinez-Gomez NC, de Graaff J, Grassl P, Pol A, Op den Camp HJM, Daumann LJ

Abstract
Recently, methanotrophic and methylotrophic bacteria were found to utilize rare earth elements (REE). To monitor the REE-content in culture media of these bacteria we have developed a rapid screening method using the Arsenazo III (AS III) dye for spectrophotometric REE-detection in the low μM (0.1-10 μM) range. We designed this assay to follow LaIII and EuIII depletion from the culture medium by the acidophilic verrucomicrobial methanotroph Methylacidiphilum fumariolicum SolV. The assay can also be modified to screen the uptake of other REE such as PrIII or to monitor the depletion of LaIII from growth media in neutrophilic methylotrophs such as Methylobacterium extorquens AM1. The AS III assay presents a convenient and fast detection method for REE levels in culture media and is a sensitive alternative to inductively coupled plasma mass spectrometry (ICP-MS) or atomic absorption spectroscopy (AAS).Importance REE-dependent bacterial metabolism is a quickly emerging field and while the importance of REE for both methanotropic and methylotrophic bacteria is now firmly established, many important questions, such as how these insoluble elements are taken up into cells, are still unanswered. Here, an Arsenazo III dye based assay has been developed for fast, specific and sensitive determination of REE content in different culture media. This assay presents a useful tool for optimizing cultivation protocols as well as for routine REE monitoring during bacterial growth without the need for specialized analytical instrumentation. Furthermore, this assay has the potential to promote the discovery of other REE-dependent microorganisms and can help to elucidate the mechanisms for acquisition of REE by methanotrophic and methylotrophic bacteria.

PMID: 29453257 [PubMed - as supplied by publisher]




Structural characterization and directed evolution of a novel acetyl xylan esterase reveals thermostability determinants of the Carbohydrate Esterase 7 family.
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Structural characterization and directed evolution of a novel acetyl xylan esterase reveals thermostability determinants of the Carbohydrate Esterase 7 family.

Appl Environ Microbiol. 2018 Feb 16;:

Authors: Adesioye FA, Makhalanyane TP, Vikram S, Sewell BT, Schubert WD, Cowan DA

Abstract
A hot desert hypolith metagenomic DNA sequence dataset was screened in-silico for genes annotated as acetyl xylan esterases (AcXEs). One of the genes identified encoded a ∼36 kDa protein (Axe1NaM1). The synthesised gene was cloned, expressed and the resulting protein, purified. NaM1 was optimally active at pH 8.5 and 30°C, and functionally stable at salt concentrations up to 5 M. The specific activity and catalytic efficiency were 488.9 Umg-1 and 3.26x106 M-1s-1, respectively. The crystal structure of wild type NaM1 was solved at a resolution of 2.03 Å and a comparison with the structures and models of more thermostable carbohydrate esterase (CE) 7 family enzymes and variants of NaM1 from a directed evolution experiment, respectively, suggest that reduced side chain volume of protein core residues is relevant to the thermal stability of NaM1. Surprisingly, a single point mutation (N96S), not only resulted in a simultaneous improvement in thermal stability and catalytic efficiency, but also increased the acyl moiety substrate range of NaM1.IMPORTANCE Acetyl xylan esterases (AcXEs) belong to nine carbohydrate esterase families (CE 1-7, 12 and 16), of which CE7 enzymes possess a unique and narrow specificity for acetylated substrates. All structurally characterised members of this family are moderately to highly thermostable. The crystal structure of a novel, mesophilic CE7 AcXE (Axe1NaM1), from a soil metagenome, provides a basis for comparisons with thermostable CE7 enzymes. Using error-prone polymerase chain reaction (PCR) and site-directed mutagenesis, we enhanced both the stability and activity of the mesophilic AcXE. With comparative structural analyses, we have also identified possible thermal stability determinants. These are valuable for understanding the thermal stability of enzymes within this family and as a guide for future protein engineering of CE7 and other α/β hydrolase enzymes.

PMID: 29453256 [PubMed - as supplied by publisher]




Endophyte-mediated modulation of defense-responsive genes and systemic resistance in Withania somnifera (L.) Dunal under Alternaria alternata stress.
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Endophyte-mediated modulation of defense-responsive genes and systemic resistance in Withania somnifera (L.) Dunal under Alternaria alternata stress.

Appl Environ Microbiol. 2018 Feb 16;:

Authors: Mishra A, Singh SP, Mahfooz S, Singh SP, Bhattacharya A, Mishra N, Nautiyal CS

Abstract
Endophytes have been explored to perform an important role in plant health. However, their effects on the host physiological function and disease management remains elusive. The present study was aimed to assess the potential of endophytes singly as well as in combination on various physiological parameters and systemic defense mechanism against Alternaria alternata in Withania somnifera (L.) Dunal. Seed priming with endophytic Bacillus amyloliquefaciens and Pseudomonas fluorescens individually and in combination demonstrated enhanced vigor index and germination rate. Interestingly, plants treated with the microbial combination showed remarkably reduced plant mortality (71.40%) under A. alternata stress. Physiological profiling of treated plants showed improved photosynthesis, respiration, transpiration and stomatal conductance under pathogenic stress. Additionally, these endophytes not only augmented defense enzymes and antioxidant activity in treated plants but also able to enhance the expression of salicylic-and jasmonic acid-responsive genes in the stressed plants. Reduction in reactive oxygen and nitrogen species along with enhanced callose deposition in host plant leaves corroborated well with the above findings. Altogether, the study provides novel insights into the underlying mechanisms behind the tripartite interaction of endophyte- A. alternata- W. somnifera and underscores their ability to boost up plant health under pathogen stress.ImportanceW. somnifera is well known for producing several medicinally important secondary metabolites. These secondary metabolites are required by various pharmaceuticals sectors to produce life saving drugs. However, the cultivation of W. somnifera faces severe damage owing to leaf spot disease caused by A. alternata To keep pace with the rising demand of this plant and considering its ability to cultivate under field conditions, the present study was undertaken to develop approaches to enhance production of W. somnifera through endophytes intervention. Application of bacterial endophytes not only suppresses the pathogenicity of A. alternata but also able to mitigate excessive ROS/RNS generation via enhanced physiological processes and antioxidant machinery. Plant defense related genes expression profiling further validates the efficacy of bacterial endophytes against leaf spot disease.

PMID: 29453255 [PubMed - as supplied by publisher]




Development of a versatile Cas9-driven subpopulation-selection toolbox in Lactococcus lactis.
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Development of a versatile Cas9-driven subpopulation-selection toolbox in Lactococcus lactis.

Appl Environ Microbiol. 2018 Feb 16;:

Authors: van der Els S, James JK, Kleerebezem M, Bron PA

Abstract
CRISPR-Cas9 technology has been exploited for the removal or replacement of genetic elements in a wide range of prokaryotes and eukaryotes. Here we describe the extension of the Cas9 application toolbox to the industrially important dairy species Lactococcus lactis The Cas9 expression vector pLABTarget was constructed, encoding the Streptocccus pyogenes Cas9 under control of a constitutive promoter, and allowing plug and play introduction of sgRNA sequences to target specific genetic loci. Introduction of a pepN-targeting derivative of pLABTarget into L. lactis MG1363 led to a strong reduction in the number of transformants obtained, which did not occur in a pepN deletion derivative of the same strain, demonstrating the specificity and lethality of the Cas9 mediated double strand breaks in the lactococcal chromosome. Moreover, the same pLABTarget derivative allowed the selection of a pepN deletion subpopulation from its corresponding single crossover plasmid integrant precursor, accelerating the construction and selection of gene-specific deletion derivatives in L. lactis Finally, pLABTarget containing sgRNAs designed to target mobile genetic elements, allowed the effective curing of plasmids, prophages and integrative conjugative elements (ICE). These results establish that pLABTarget enables the effective exploitation of Cas9 targeting in L. lactis, while the broad host range vector used suggests that this toolbox could be readily expanded to other Gram-positive bacteria.Significance statement Mobile genetic elements in Lactococcus lactis and other lactic acid bacteria play an important role in dairy fermentation, having both positive and detrimental effects during the production of fermented dairy products. The pLABTarget vector offers an efficient cloning platform for Cas9 application in lactic acid bacteria. Targeting Cas9 towards mobile genetic elements enabled their effective curing, which is of particular interest in the context of potentially problematic prophages present in a strain. Moreover, Cas9 targeting to other mobile genetic elements enables the deciphering of their contribution to dairy fermentation processes and further establishing their importance for product characteristics.

PMID: 29453254 [PubMed - as supplied by publisher]




Assembly of synthetic functional cellulosomal structures onto the Lactobacillus plantarum cell surface - a potent member of the gut microbiome.
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Assembly of synthetic functional cellulosomal structures onto the Lactobacillus plantarum cell surface - a potent member of the gut microbiome.

Appl Environ Microbiol. 2018 Feb 16;:

Authors: Stern J, Moraïs S, Ben-David Y, Salama R, Shamshoum M, Lamed R, Shoham Y, Bayer EA, Mizrahi I

Abstract
Heterologous display of enzymes on microbial cell surfaces is an extremely desirable approach, since it enables the engineered microbe to interact directly with the plant-wall extracellular polysaccharide matrix. In recent years, attempts have been made to endow non-cellulolytic microbes with genetically engineered cellulolytic capabilities for improved hydrolysis of lignocellulosic biomass and for advanced probiotics. Thus far, however, owing to the hurdles of secreting and assembling large, intricate complexes on the bacterial cell wall, only free cellulases or relatively simple cellulosome assemblies have been introduced into live bacteria. Here, we employed the "adaptor scaffoldin" strategy to overcome the low levels of protein displayed on the bacterial cell surface. The approach mimics natural cellulosome elaborated architectures, thus exploiting the exponential features of their Lego-like combinatorics. Using this approach, we produced several bacterial consortia of Lactobacillus plantarum, a potent gut microbe which provides a very robust genetic framework for lignocellulosic degradation. We successfully engineered surface display of large, fully active self-assembling cellulosomal complexes containing an unprecedented number of catalytic subunits all produced in vivo by the cell-consortia. Our results demonstrate superior enzyme stability and performance of the cellulosomal machinery, compared to the equivalent secreted free enzyme system and high cellulase-to-xylanase ratios proved beneficial for efficient degradation of wheat straw.Importance The multiple benefits of lactic acid bacteria are well established in health and industry. Here we present an approach to extensively increase the cell-surface display of proteins via successive assembly of interactive components. Our findings present a stepping stone towards proficient engineering of Lactobacillus plantarum, a widespread, environmentally important bacterium and potent microbiome member, for improved degradation of lignocellulosic biomass and advanced probiotics.

PMID: 29453253 [PubMed - as supplied by publisher]




Ecology and biotechnological potential of bacteria belonging to the Pseudovibrio genus.
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Ecology and biotechnological potential of bacteria belonging to the Pseudovibrio genus.

Appl Environ Microbiol. 2018 Feb 16;:

Authors: Romano S

Abstract
Bacteria belonging to the genus Pseudovibrio have been isolated worldwide from a great variety of marines sources as both free living and host associated. So far, the available data depict a group of Alphaproteobacteria characterized by a versatile metabolism, which allows them to use a variety of substrates to meet their carbon, nitrogen, sulfur, and phosphorous requirements. Additionally, Pseudovibrio-related bacteria have been shown to proliferate under extreme oligotrophic conditions, tolerate high heavy metal concentrations, and metabolize potentially toxic compounds. Considering this versatility, it is not surprising that they have been detected from temperate to tropical regions, and are often the most abundant isolates obtained from marine invertebrates. Such association is particularly recurrent with marine sponges and corals, animals that play a key role in benthic marine systems. The data so far available indicate that these bacteria are mainly beneficial to the host, and besides being involved in major nutrient cycles, they could provide the host with both vitamins/cofactors and protection from potential pathogens via the synthesis of antimicrobial secondary metabolites. In fact, the biosynthetic abilities of Pseudovibrio have been emerging in recent years, and both genomic and analytic studies underlined how these organisms promise novel natural products of biotechnological value.

PMID: 29453252 [PubMed - as supplied by publisher]