Sulphate-reducing bacterial community structure from produced water of the Periquito and Galo de Campina onshore oilfields in Brazil.

Sulphate-reducing bacteria (SRB) cause fouling, souring, corrosion and produce H2S during oil and gas production. Produced water obtained from Periquito (PQO) and Galo de Campina (GC) onshore oilfields in Brazil was investigated for SRB. Produced water with Postgate B, Postgate C and Baars media was incubated anaerobically for 20 days. DNA was extracted, 16S rDNA PCR amplified and fragments were sequenced using Illumina TruSeq. 4.2 million sequence reads were analysed and deposited at NCBI SAR accession number SRP149784. No significant differences in microbial community composition could be attributed to the different media but significant differences in the SRB were observed between the two oil fields. The dominant bacterial orders detected from both oilfields were Desulfovibrionales, Pseudomonadales and Enterobacteriales. The genus Pseudomonas was found predominantly in the GC oilfield and Pleomorphominas and Shewanella were features of the PQO oilfield. 11% and 7.6% of the sequences at GC and PQO were not classified at the genus level but could be partially identified at the order level. Relative abundances changed for Desulfovibrio from 29.8% at PQO to 16.1% at GC. Clostridium varied from 2.8% at PQO and 2.4% at GC. These data provide the first description of SRB from onshore produced water in Brazil and reinforce the importance of Desulfovibrionales, Pseudomonadales, and Enterobacteriales in produced water globally. Identifying potentially harmful microbes is an important first step in developing microbial solutions that prevent their proliferation.

Bacterioplankton response to nitrogen and dissolved organic matter produced from salmon mucus.

Aquaculture releases organic matter to the water column through excretion, fecal pellets, and uneaten food, but also by the continuous release of fish epithelium mucus. The effect of the latter on natural bacterial assemblages was determined using ammonium amended experiments at Puyuhuapi fjord in Chilean Patagonia. Mucus was added to seawater coming from 2 and 100 m depth and ammonium, nitrite and nitrate, dissolved organic carbon (DOC), picoplankton abundance, and active composition (i-tag 16S rRNA) were followed for 24 h. The results showed a significant response from the microbial community but only at surface depth after 2 and 6 h of incubation. A reduction of DOC and ammonium concentration and accumulation of nitrite and nitrate over time was observed, mainly at 100 m. Changes in the composition of active bacteria between treatments were observed at different taxonomic levels, associated with Alphaproteobacteria (Clade SAR11), Bacteroidetes (Polaribacter) and Gammaproteobacteria (Colwellia, Oceaniserpentilla) and other bacteria such as Nitrospina sp, a nitrite-oxidizing bacteria at some hours during the incubation. Fish pathogens, such as Vibrio and Piscirickettsia were rare (<0.02%). Overall, our study suggests that fish mucus can cause rapid modifications in microbial assemblages and stimulate organic matter and nutrient cycling, including heterotrophic and autotrophic (nitrification) in areas influenced by aquaculture.

Antibiotic Resistance in Vibrio cholerae: Mechanistic Insights from IncC Plasmid-Mediated Dissemination of a Novel Family of Genomic Islands Inserted at trmE.

Cholera remains a formidable disease, and reports of multidrug-resistant strains of the causative agent Vibrio cholerae have become common during the last 3 decades. The pervasiveness of resistance determinants has largely been ascribed to mobile genetic elements, including SXT/R391 integrative conjugative elements, IncC plasmids, and genomic islands (GIs). Conjugative transfer of IncC plasmids is activated by the master activator AcaCD whose regulatory network extends to chromosomally integrated GIs. MGIVchHai6 is a multidrug resistance GI integrated at the 3' end of trmE (mnmE or thdF) in chromosome 1 of non-O1/non-O139 V. cholerae clinical isolates from the 2010 Haitian cholera outbreak. In the presence of an IncC plasmid expressing AcaCD, MGIVchHai6 excises from the chromosome and transfers at high frequency. Herein, the mechanism of mobilization of MGIVchHai6 GIs by IncC plasmids was dissected. Our results show that AcaCD drives expression of GI-borne genes, including xis and mobIM , involved in excision and mobilization. A 49-bp fragment upstream of mobIM was found to serve as the minimal origin of transfer (oriT) of MGIVchHai6. The direction of transfer initiated at oriT was determined using IncC plasmid-driven mobilization of chromosomal markers via MGIVchHai6. In addition, IncC plasmid-encoded factors, including the relaxase TraI, were found to be required for GI transfer. Finally, in silico exploration of Gammaproteobacteria genomes identified 47 novel related and potentially AcaCD-responsive GIs in 13 different genera. Despite sharing conserved features, these GIs integrate at trmE, yicC, or dusA and carry a diverse cargo of genes involved in phage resistance.IMPORTANCE The increasing association of the etiological agent of cholera, Vibrio cholerae serogroup O1 and O139, with multiple antibiotic resistance threatens to deprive health practitioners of this effective tool. Drug resistance in cholera results mainly from acquisition of mobile genetic elements. Genomic islands conferring multidrug resistance and mobilizable by IncC conjugative plasmids were reported to circulate in non-O1/non-O139 V. cholerae clinical strains isolated from the 2010 Haitian cholera outbreak. As these genomic islands can be transmitted to pandemic V. cholerae serogroups, their mechanism of transmission needed to be investigated. Our research revealed plasmid- and genomic island-encoded factors required for the resistance island excision, mobilization, and integration, as well as regulation of these functions. The discovery of related genomic islands carrying diverse phage resistance genes but lacking antibiotic resistance-conferring genes in a wide range of marine dwelling bacteria suggests that these elements are ancient and recently acquired drug resistance genes.

ShadowCaster: Compositional Methods under the Shadow of Phylogenetic Models to Detect Horizontal Gene Transfers in Prokaryotes.

Horizontal gene transfer (HGT) plays an important role for evolutionary innovations within prokaryotic communities and is a crucial event for their survival. Several computational approaches have arisen to identify HGT events in recipient genomes. However, this has been proven to be a complex task due to the generation of a great number of false positives and the prediction disagreement among the existing methods. Phylogenetic reconstruction methods turned out to be the most reliable ones, but they are not extensible to all genes/species and are computationally demanding when dealing with large datasets. In contrast, the so-called surrogate methods that use heuristic solutions either based on nucleotide composition patterns or phyletic distribution of BLAST hits can be applied easily to the genomic scale, but they fail in identifying common HGT events. Here, we present ShadowCaster, a hybrid approach that sequentially combines nucleotide composition-based predictions by support vector machines (SVMs) under the shadow of phylogenetic models independent of tree reconstruction, to improve the detection of HGT events in prokaryotes. ShadowCaster successfully predicted close and distant HGT events in both artificial and bacterial genomes. ShadowCaster detected HGT related to heavy metal resistance in the genome of Rhodanobacter denitrificans with higher accuracy than the most popular state-of-the-art computational approaches, encompassing most of the predicted cases made by other methods. ShadowCaster is released at the GitHub platform as an open-source software under the GPLv3 license.

A novel bacterial carboxylesterase identified in a metagenome derived-clone from Brazilian mangrove sediments.

A functional screening of 1152 clones from a plasmid library constructed with DNA extracted from Brazilian mangrove sediments revealed 3 positive clones for ester-hydrolyzing enzymes, or about one lipolytic gene per 1.2 Mb DNA, which corroborates the idea that oil-contaminated mangroves are a good source of novel microbial lipases/esterases. The partial sequence of the clone LipG7 (1179 bp) showed 30.2% of predicted structure identity with a known esterase, confirming LipG7 as a new member of family VIII esterases. LigG7 shared 80% sequence identity with 1,4-butanediol diacrylate esterase from the Gammaprotebacteria Porticoccus hydrocarbonoclasticus, suggesting it belongs to the Porticoccaceae family. LipG7 was heterologously expressed in Escherichia coli Rosetta-Gami DE3; the purified recombinant enzyme exhibited a predicted molecular weight of 45.2 kDa and exceptional activity towards 4-nitrophenyl butyrate, compared with other recombinant esterases, highlighting its enormous potential for biological applications.

Bacterial diversity in deep-sea sediments under influence of asphalt seep at the São Paulo Plateau.

Here we investigated the diversity of bacterial communities from deep-sea surface sediments under influence of asphalt seeps at the Sao Paulo Plateau using next-generation sequencing method. Sampling was performed at North São Paulo Plateau using the human occupied vehicle Shinkai 6500 and her support vessel Yokosuka. The microbial diversity was studied at two surficial sediment layers (0-1 and 1-4 cm) of five samples collected in cores in water depths ranging from 2456 to 2728 m. Bacterial communities were studied through sequencing of 16S rRNA gene on the Ion Torrent platform and clustered in operational taxonomic units. We observed high diversity of bacterial sediment communities as previously described by other studies. When we considered community composition, the most abundant classes were Alphaproteobacteria (27.7%), Acidimicrobiia (20%), Gammaproteobacteria (11.3%) and Deltaproteobacteria (6.6%). Most abundant OTUs at family level were from two uncultured bacteria from Actinomarinales (5.95%) and Kiloniellaceae (3.17%). The unexpected high abundance of Alphaproteobacteria and Acidimicrobiia in our deep-sea microbial communities may be related to the presence of asphalt seep at North São Paulo Plateau, since these bacterial classes contain bacteria that possess the capability of metabolizing hydrocarbon compounds.

The gill-associated microbiome is the main source of wood plant polysaccharide hydrolases and secondary metabolite gene clusters in the mangrove shipworm Neoteredo reynei.

Teredinidae are a family of highly adapted wood-feeding and wood-boring bivalves, commonly known as shipworms, whose evolution is linked to the acquisition of cellulolytic gammaproteobacterial symbionts harbored in bacteriocytes within the gills. In the present work we applied metagenomics to characterize microbiomes of the gills and digestive tract of Neoteredo reynei, a mangrove-adapted shipworm species found over a large range of the Brazilian coast. Comparative metagenomics grouped the gill symbiont community of different N. reynei specimens, indicating closely related bacterial types are shared. Similarly, the intestine and digestive gland communities were related, yet were more diverse than and showed no overlap with the gill community. Annotation of assembled metagenomic contigs revealed that the gill symbiotic community of N. reynei encodes a plethora of plant cell wall polysaccharides degrading glycoside hydrolase encoding genes, and Biosynthetic Gene Clusters (BGCs). In contrast, the digestive tract microbiomes seem to play little role in wood digestion and secondary metabolites biosynthesis. Metagenome binning recovered the nearly complete genome sequences of two symbiotic Teredinibacter strains from the gills, a representative of Teredinibacter turnerae "clade I" strain, and a yet to be cultivated Teredinibacter sp. type. These Teredinibacter genomes, as well as un-binned gill-derived gammaproteobacteria contigs, also include an endo-ß-1,4-xylanase/acetylxylan esterase multi-catalytic carbohydrate-active enzyme, and a trans-acyltransferase polyketide synthase (trans-AT PKS) gene cluster with the gene cassette for generating ß-branching on complex polyketides. Finally, we use multivariate analyses to show that the secondary metabolome from the genomes of Teredinibacter representatives, including genomes binned from N. reynei gills' metagenomes presented herein, stands out within the Cellvibrionaceae family by size, and enrichments for polyketide, nonribosomal peptide and hybrid BGCs. Results presented here add to the growing characterization of shipworm symbiotic microbiomes and indicate that the N. reynei gill gammaproteobacterial community is a prolific source of biotechnologically relevant enzymes for wood-digestion and bioactive compounds production.

Distribution and phylogenetics of whiteflies and their endosymbiont relationships after the Mediterranean species invasion in Brazil.

The Bemisia tabaci is a polyphagous insect and a successful vector of plant viruses. B. tabaci is a species complex and in Brazil native species from the New World (NW) group, as well as the invasive species, Middle East-Asia Minor 1 (MEAM1) and Mediterranean (MED) were reported. For better understanding the distribution of the different species four years after the Mediterranean species invasion in Brazil, whiteflies were collected from 237 locations throughout the country between the years of 2013 and 2017, species were identified and the facultative endosymbionts detected. The survey revealed that MEAM1 was the prevalent species found on major crops across Brazil. It is the only species present in North, Northwestern and Central Brazil and was associated with virus-infected plants. MED was found in five States from Southeast to South regions, infesting mainly ornamental plants and was not associated with virus-infected plants. The prevalent endosymbionts identified in MEAM1 were Hamiltonella and Rickettsia; and the mtCOI analysis revealed low genetic diversity for MEAM1. In contrast, several different endosymbionts were identified in MED including Hamiltonella, Rickettsia, Wolbachia and Arsenophonus; and two distinct genetic groups were found based on the mtCOI analysis. Monitoring the distribution of the whiteflies species in Brazil is essential for proper management of this pest.

Insights into the biology of acidophilic members of the Acidiferrobacteraceae family derived from comparative genomic analyses.

The family Acidiferrobacteraceae (order Acidiferrobacterales) currently contains Gram negative, neutrophilic sulfur oxidizers such as Sulfuricaulis and Sulfurifustis, as well as acidophilic iron and sulfur oxidizers belonging to the Acidiferrobacter genus. The diversity and taxonomy of the genus Acidiferrobacter has remained poorly explored. Although several metagenome and bioleaching studies have identified its presence worldwide, only two strains, namely Acidiferrobacter thiooxydans DSM 2932T, and Acidiferrobacter spp. SP3/III have been isolated and made publically available. Using 16S rRNA sequence data publically available for the Acidiferrobacteraceae, we herein shed light into the molecular taxonomy of this family. Results obtained support the presence of three clades Acidiferrobacter, Sulfuricaulis and Sulfurifustis. Genomic analyses of the genome sequences of A. thiooxydansT and Acidiferrobacter spp. SP3/III indicate that ANI relatedness between the SPIII/3 strain and A. thiooxydansT is below 95-96%, supporting the classification of strain SP3/III as a new species within this genus. In addition, approximately 70% of Acidiferrobacter sp. SPIII/3 predicted genes have a conserved ortholog in A. thiooxydans strains. A comparative analysis of iron, sulfur oxidation pathways, genome plasticity and cell-cell communication mechanisms of Acidiferrobacter spp. are also discussed.

Culturable symbionts associated with the reproductive and digestive tissues of the Neotropical brown stinkbug Euschistus heros.

Symbionts are widely distributed in eukaryotes, and potentially affect the physiology, ecology and evolution of their host. Most insects harbour free-living bacteria in their haemocoel and gut lumen, intracellular-living bacteria in a range of tissues or bacteria in host-derived specialized cells. Stinkbugs, as do many arthropods, harbour extracellular bacteria in the gut that may affect the fitness of their host. This study identified the culturable symbionts associated with the ovaries, spermatheca, seminal vesicle and posterior midgut region (V4) of males and females of Euschistus heros (F.) (Hemiptera: Pentatomidae). Several culture media were used to isolate the bacteria associated with these structures. The selected colonies (morphotypes) were cultured in liquid medium, subjected to genomic DNA extraction, 16S rRNA gene amplification, and restriction fragment length polymorphism (RFLP) analyses. Morphotypes with distinct RFLP patterns were purified and sequenced, and the sequences obtained were used for putative identification and phylogenetic analysis. Comparison of the sequences with those available in the EzTaxon-e database and the use of a matrix of paired distances grouped the isolates in phylotypes belonging to the Phylum Proteobacteria. Proteobacteria was represented by γ-Proteobacteria phylotypes belonging to Enterobacteriaceae, while Firmicutes had Bacilli phylotypes distributed in Enterococcaceae and Staphylococcaceae. Some of the phylotypes identified were associated exclusively with single structures, such as ovaries, spermatheca and the V4 midgut region of males and females. All culturable bacteria associated with the seminal vesicle were also associated with other tissues.

Filamentous Giant Beggiatoaceae from the Guaymas Basin Are Capable of both Denitrification and Dissimilatory Nitrate Reduction to Ammonium.

Filamentous large sulfur-oxidizing bacteria (FLSB) of the family Beggiatoaceae are globally distributed aquatic bacteria that can control geochemical fluxes from the sediment to the water column through their metabolic activity. FLSB mats from hydrothermal sediments of Guaymas Basin, Mexico, typically have a "fried-egg" appearance, with orange filaments dominating near the center and wider white filaments at the periphery, likely reflecting areas of higher and lower sulfide fluxes, respectively. These FLSB store large quantities of intracellular nitrate that they use to oxidize sulfide. By applying a combination of 15N-labeling techniques and genome sequence analysis, we demonstrate that the white FLSB filaments were capable of reducing their intracellular nitrate stores to both nitrogen gas and ammonium by denitrification and dissimilatory nitrate reduction to ammonium (DNRA), respectively. On the other hand, our combined results show that the orange filaments were primarily capable of DNRA. Microsensor profiles through a laboratory-incubated white FLSB mat revealed a 2- to 3-mm vertical separation between the oxic and sulfidic zones. Denitrification was most intense just below the oxic zone, as shown by the production of nitrous oxide following exposure to acetylene, which blocks nitrous oxide reduction to nitrogen gas. Below this zone, a local pH maximum coincided with sulfide oxidation, consistent with nitrate reduction by DNRA. The balance between internally and externally available electron acceptors (nitrate) and electron donors (reduced sulfur) likely controlled the end product of nitrate reduction both between orange and white FLSB mats and between different spatial and geochemical niches within the white FLSB mat.IMPORTANCE Whether large sulfur bacteria of the family Beggiatoaceae reduce NO3- to N2 via denitrification or to NH4+ via DNRA has been debated in the literature for more than 25 years. We resolve this debate by showing that certain members of the Beggiatoaceae use both metabolic pathways. This is important for the ecological role of these bacteria, as N2 production removes bioavailable nitrogen from the ecosystem, whereas NH4+ production retains it. For this reason, the topic of environmental controls on the competition for NO3- between N2-producing and NH4+-producing bacteria is of great scientific interest. Recent experiments on the competition between these two types of microorganisms have demonstrated that the balance between electron donor and electron acceptor availability strongly influences the end product of NO3- reduction. Our results suggest that this is also the case at the even more fundamental level of enzyme system regulation within a single organism.

Description of Sansalvadorimonas verongulae gen. nov., sp. nov., a gammaproteobacterium isolated from the marine sponge Verongula gigantea.

A Gram-stain-negative, strictly aerobic, motile, rod-shaped bacterium, designated strain RKSG058T, was isolated from the marine sponge Verongula gigantea, collected off the west coast of San Salvador, The Bahamas. Phylogenetic analyses based on 16S rRNA gene sequences revealed that RKSG058T formed a distinct lineage within the family Hahellaceae (order Oceanospirillales, class Gammaproteobacteria), and was most closely related to the genus Endozoicomonas, with sequence similarities to members of this genus ranging from 92.0 to 93.7 %. Optimal growth occurred at 30 °C, at pH 7 and in the presence of 2-3 % (w/v) NaCl. The predominant cellular fatty acids were summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c), summed feature 8 (C18 : 1ω7c and/or C18 : 1ω6c) and C16 : 0. The major and minor respiratory quinones were Q-9 and Q-8, respectively. The polar lipids comprised diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, three unidentified aminolipids, an unidentified phospholipid and five unidentified lipids. The DNA G+C content was 42.3 mol%. Biochemical, chemotaxonomic and phylogenetic analyses indicated that strain RKSG058T represents the first cultured isolate of a novel bacterial genus and species within the family Hahellaceae, for which the name Sansalvadorimonas verongulae gen. nov., sp. nov. is proposed. The type strain of Sansalvadorimonas verongulae is RKSG058T (=TSD-72T=LMG 29871T). An emended description of the genus Kistimonas is provided.

Metabolic potential and in situ transcriptomic profiles of previously uncharacterized key microbial groups involved in coupled carbon, nitrogen and sulfur cycling in anoxic marine zones.

Anoxic marine zones (AMZs) impact biogeochemical cycles at the global scale, particularly the nitrogen cycle. Key microbial players from AMZs have been identified, but the majority remains unrecognized or uncharacterized. Thirty-one single-cell amplified genomes (SAGs) from the eastern tropical North and South Pacific AMZs were sequenced to gain insight into the distribution, metabolic potential and contribution to the community transcriptional profile of these uncharacterized bacterial and archaeal groups. Detailed analyses focused on SAG-bins assigned to three of these groups that presented 79%-100% estimated genome completeness: the putative sulphur-oxidizing Gamaproteobacteria EOSA II clade, a Marinimicrobia member of the recently recognized PN262000N21 clade found to be abundant in AMZ anoxic cores, and a representative of the Marine Benthic Group A Thaumarchaeota. Community-based analyses revealed that these three groups are significantly more abundant and transcriptionally more active in the AMZ microbial communities than previously described phylogenetically related microbial groups. Collectively, these groups have the potential to link biogeochemically relevant processes by coupling the carbon, nitrogen and sulfur cycles. Together, these results increase our understanding of key microbial components inhabiting AMZs and other oxygen-deficient marine environments, enhancing our capacity to predict the impact of the expansion of these ecosystems due to climate change.

An Unbiased Genome-Wide View of the Mutation Rate and Spectrum of the Endosymbiotic Bacterium Teredinibacter turnerae.

Mutations contribute to genetic variation in all living systems. Thus, precise estimates of mutation rates and spectra across a diversity of organisms are required for a full comprehension of evolution. Here, a mutation-accumulation (MA) assay was carried out on the endosymbiotic bacterium Teredinibacter turnerae. After ∼3,025 generations, base-pair substitutions (BPSs) and insertion-deletion (indel) events were characterized by whole-genome sequencing analysis of 47 independent MA lines, yielding a BPS rate of 1.14 × 10-9 per site per generation and indel rate of 1.55 × 10-10 events per site per generation, which are among the highest within free-living and facultative intracellular bacteria. As in other endosymbionts, a significant bias of BPSs toward A/T and an excess of deletion mutations over insertion mutations are observed for these MA lines. However, even with a deletion bias, the genome remains relatively large (∼5.2 Mb) for an endosymbiotic bacterium. The estimate of the effective population size (Ne) in T. turnerae is quite high and comparable to free-living bacteria (∼4.5 × 107), suggesting that the heavy bottlenecking associated with many endosymbiotic relationships is not prevalent during the life of this endosymbiont. The efficiency of selection scales with increasing Ne and such strong selection may have been operating against the deletion bias, preventing genome erosion. The observed mutation rate in this endosymbiont is of the same order of magnitude of those with similar Ne, consistent with the idea that population size is a primary determinant of mutation-rate evolution within endosymbionts, and that not all endosymbionts have low Ne.

Comparative metagenomics of hydrocarbon and methane seeps of the Gulf of Mexico.

Oil and gas percolate profusely through the sediments of the Gulf of Mexico, leading to numerous seeps at the seafloor, where complex microbial, and sometimes animal communities flourish. Sediments from three areas (two cold seeps with contrasting hydrocarbon composition and a site outside any area of active seepage) of the Gulf of Mexico were investigated and compared. Consistent with the existence of a seep microbiome, a distinct microbial community was observed in seep areas compared to sediment from outside areas of active seepage. The microbial community from sediments without any influence from hydrocarbon seepage was characterized by Planctomycetes and the metabolic potential was consistent with detrital marine snow degradation. By contrast, in seep samples with methane as the principal hydrocarbon, methane oxidation by abundant members of ANME-1 was likely the predominant process. Seep samples characterized by fluids containing both methane and complex hydrocarbons, were characterized by abundant Chloroflexi (Anaerolinaceae) and deltaproteobacterial lineages and exhibited potential for complex hydrocarbon degradation. These different metabolic capacities suggested that microorganisms in cold seeps can potentially rely on other processes beyond methane oxidation and that the hydrocarbon composition of the seep fluids may be a critical factor structuring the seafloor microbial community composition and function.

Psychromonas aquatilis sp. nov., isolated from seawater samples obtained in the Chilean Antarctica.

A slightly beige-white pigmented, Gram-staining-negative, rod-shaped bacterium, strain M1A1T, was isolated from seawater samples obtained in Fildes Bay, Antarctica (62°12' S 58° 57' W). Phylogenetic analysis based on nearly full-length 16S rRNA gene sequences showed that the isolate shared 98.4 % 16S rRNA gene sequence identity to the type strain of Psychromonas arctica, but less than 97 % 16S rRNA gene sequence similarities to all other species of the genus Psychromonas. DNA-DNA hybridization with Psychromonas arctica DSM 14288T showed low values (21 %, reciprocal 27 %). The main cellular fatty acid of strain M1A1T was summed feature 3 fatty acids (C16 : 1ω7c/C16 : 1ω8c), followed by C16 : 0. Based on phylogenetic, chemotaxonomic, genomic and phenotypic analyses, we propose a novel species of the genus Psychromonas with the name Psychromonas aquatilis sp. nov. and the strain M1A1T (=CIP 111183T=CCM 8710T=LMG 29766T) as type strain.

Streptomyces effect on the bacterial microbiota associated to Crassostrea sikamea oyster.

AIM: To determine the composition and diversity of the microbiota associated to Crassostrea sikamea treated during 30 days with Streptomyces strains N7 and RL8. METHODS AND RESULTS: DNA was extracted from oysters followed by 16S rRNA gene amplification and pyrosequencing. The highest and lowest species diversity richness was observed in the initial and final control group, whereas Streptomyces-treated oysters exhibited intermediate values. Proteobacteria was the most abundant phylum (81·4-95·1%), followed by Bacteroidetes, Actinobacteria and Firmicutes. The genera Anderseniella, Oceanicola, Roseovarius, Ruegeria, Sulfitobacter, Granulosicoccus and Marinicella encompassed the core microbiota of all experimental groups. The genus Bacteriovorax was detected in all groups except in the final control and the depurated N7, whereas Vibrio remained undetected in all Streptomyces-treated groups. RL8 was the only group that harboured the genus Streptomyces in its microbiota. Principal component analysis showed that Streptomyces strains significantly changed oyster microbiota with respect to the initial and final control. CONCLUSIONS: Crassostrea sikamea treated with Streptomyces showed high species diversity and a microbiota composition shift, characterized by keeping the predator genus Bacteriovorax and decreasing the pathogenic Vibrio. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first culture-independent study showing the effect of Streptomyces over the oyster microbiota. It also sheds light about the potential use of Streptomyces to improve mollusc health and safety for consumers after the depuration process.

A glimpse of the endophytic bacterial diversity in roots of blackberry plants (Rubus fruticosus).

The aim of this study was to explore the diversity of culturable bacterial communities residing in blackberry plants (Rubus fruticosus). Bacterial endophytes were isolated from plant roots, and their 16S rDNA sequences were amplified and sequenced. Our results show that the roots of R. fruticosus exhibit low colony forming units of bacterial endophytes per gram of fresh tissue (6 x 102 ± 0.5 x 102). We identified 41 endophytic bacterial species in R. fruticosus by BLAST homology search and a subsequent phylogenetic analysis, belonging to the classes Actinobacteria, Bacilli, Alfaproteobacteria, Betaproteobacteria, and Gammaproteobacteria. Predominantly, genera belonging the Proteobacteria (Burkholderia, 29.4%; Herbaspirillum, 10.7%; Pseudomonas, 4.9%; and Dyella, 3.9%), Firmicutes (Bacillus, 42.1%), and Actinobacteria (two isolates showing high identity with the Streptomyces genus, 1.9%) divisions were identified. Fifty percent of the bacterial endophytes produced the phytohormone indole-acetic acid (IAA), eleven of which exhibited higher IAA production (>5.8 mg/mL) compared to the plant growth-promoting strain, Pseudomonas fluorescens UM270. Additionally, the endophytic isolates exhibited protease activity (22%), produced siderophores (26.4%), and demonstrated antagonistic action (>50% inhibition of mycelial growth) against the grey mold phytopathogen Botrytis cinerea (3.9%). These results suggested that field-grown R. fruticosus plants contain bacterial endophytes within their tissues with the potential to promote plant growth and display antagonism towards plant pathogens.

Biochemical Characterization and Low-Resolution SAXS Molecular Envelope of GH1 ß-Glycosidase from Saccharophagus degradans.

The marine bacteria Saccharophagus degradans (also known as Microbulbifer degradans), are rod-shaped and gram-negative motile γ-proteobacteria, capable of both degrading a variety of complex polysaccharides and fermenting monosaccharides into ethanol. In order to obtain insights into structure-function relationships of the enzymes, involved in these biochemical processes, we characterized a S. degradans ß-glycosidase from glycoside hydrolase family 1 (SdBgl1B). SdBgl1B has the optimum pH of 6.0 and a melting temperature T m of approximately 50 °C. The enzyme has high specificity toward short D-glucose saccharides with ß-linkages with the following preferences ß-1,3 > ß-1,4 â‰« ß-1,6. The enzyme kinetic parameters, obtained using artificial substrates p-ß-NPGlu and p-ß-NPFuc and also the disaccharides cellobiose, gentiobiose and laminaribiose, revealed SdBgl1B preference for p-ß-NPGlu and laminaribiose, which indicates its affinity for glucose and also preference for ß-1,3 linkages. To better understand structural basis of the enzyme activity its 3D model was built and analysed. The 3D model fits well into the experimentally retrieved low-resolution SAXS-based envelope of the enzyme, confirming monomeric state of SdBgl1B in solution.

Rhizobacterial Community Structures Associated with Native Plants Grown in Chilean Extreme Environments.

Chile is topographically and climatically diverse, with a wide array of diverse undisturbed ecosystems that include native plants that are highly adapted to local conditions. However, our understanding of the diversity, activity, and role of rhizobacteria associated with natural vegetation in undisturbed Chilean extreme ecosystems is very poor. In the present study, the combination of denaturing gradient gel electrophoresis and 454-pyrosequencing approaches was used to describe the rhizobacterial community structures of native plants grown in three representative Chilean extreme environments: Atacama Desert (ATA), Andes Mountains (AND), and Antarctic (ANT). Both molecular approaches revealed the presence of Proteobacteria, Bacteroidetes, and Actinobacteria as the dominant phyla in the rhizospheres of native plants. Lower numbers of operational taxonomic units (OTUs) were observed in rhizosphere soils from ATA compared with AND and ANT. Both approaches also showed differences in rhizobacterial community structures between extreme environments and between plant species. The differences among plant species grown in the same environment were attributed to the higher relative abundance of classes Gammaproteobacteria and Alphaproteobacteria. However, further studies are needed to determine which environmental factors regulate the structures of rhizobacterial communities, and how (or if) specific bacterial groups may contribute to the growth and survival of native plants in each Chilean extreme environments.