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2017
Lan, Y, Sun J, Tian RM, Bartlett DH, Li RS, Wong YH, Zhang WP, Qiu JW, Xu T, He LS, Tabata HG, Qian PY.  2017.  Molecular adaptation in the world's deepest-living animal: Insights from transcriptome sequencing of the hadal amphipod Hirondellea gigas. Molecular Ecology. 26:3732-3743.   10.1111/mec.14149   AbstractWebsite

The Challenger Deep in the Mariana Trench is the deepest point in the oceans of our planet. Understanding how animals adapt to this harsh environment characterized by high hydrostatic pressure, food limitation, dark and cold is of great scientific interest. Of the animals dwelling in the Challenger Deep, amphipods have been captured using baited traps. In this study, we sequenced the transcriptome of the amphipod Hirondellea gigas collected at a depth of 10,929 m from the East Pond of the Challenger Deep. Assembly of these sequences resulted in 133,041 contigs and 22,046 translated proteins. Functional annotation of these contigs was made using the GO and KEGG databases. Comparison of these translated proteins with those of four shallow-water amphipods revealed 10,731 gene families, of which 5659 were single-copy orthologs. Base substitution analysis on these single-copy orthologs showed that 62 genes are positively selected in H. gigas, including genes related to beta-alanine biosynthesis, energy metabolism and genetic information processing. For multiple-copy orthologous genes, gene family expansion analysis revealed that cold-inducible proteins (i.e., transcription factors II A and transcription elongation factor 1) as well as zinc finger domains are expanded in H. gigas. Overall, our results indicate that genetic adaptation to the hadal environment by H. gigas may be mediated by both gene family expansion and amino acid substitutions of specific proteins.

Yang, Y, Xu GP, Liang JD, He Y, Xiong L, Li H, Bartlett D, Deng ZX, Wang ZJ, Xiao X.  2017.  DNA backbone sulfur-modification expands microbial growth range under multiple stresses by its antioxidation function. Scientific Reports. 7   10.1038/s41598-017-02445-1   AbstractWebsite

DNA phosphorothioate (PT) modification is a sulfur modification on the backbone of DNA introduced by the proteins DndA-E. It has been detected within many bacteria isolates and metagenomic datasets, including human pathogens, and is considered to be widely distributed in nature. However, little is known about the physiological function of this modification, and thus its evolutionary significance and application potential remains largely a mystery. In this study, we focused on the advantages of DNA PT modification to bacterial cells coping with environmental stresses. We show that the mesophile Escherichia coli and the extremophile Shewanella piezotolerans both expanded their growth ranges following exposure to extreme temperature, salinity, pH, pressure, UV, X-ray and heavy metals as a result of DNA phophorothioation. The phophorothioated DNA reacted to both H2O2 and hydroxyl radicals in vivo, and protected genomic DNA as well as sensitive enzymes from intracellular oxidative damage. We further demonstrate that this process has evolved separate from its associated role in DNA restriction and modification. These findings provide a physiological role for a covalent modification widespread in nature and suggest possible applications in biotechnology and biomedicine.

Kusube, M, Kyaw TS, Tanikawa K, Chastain RA, Hardy KM, Cameron J, Bartlett DH.  2017.  Colwellia marinimaniae sp nov., a hyperpiezophilic species isolated from an amphipod within the Challenger Deep, Mariana Trench. International Journal of Systematic and Evolutionary Microbiology. 67:824-831.   10.1099/ijsem.0.001671   AbstractWebsite

An obligately piezophilic strain was isolated from an amphipod crustacean obtained in the Challenger Deep region of the Mariana Trench during the DEEPSEA CHALLENGE expedition. The strain, MTCD1(T), grew at extremely high hydrostatic pressures, with a growth range of 80-140 MPa (optimum, 120 MPa) at 6 degrees C. Phylogenetic analyses based on the 16S rRNA gene sequence indicate that it is closely affiliated with the genus Colwellia. Comparative 16S rRNA gene sequence analyses revealed 95.7, 95.5 and 95.2% similarity to Colwellia maris ABE-1(T), Colwellia piezophila Y233G(T) and Colwellia psychrerythraea ATCC 27364(T), respectively. The major cellular fatty acids were C-16 : 1, C-16 : 0 and C-22 : 6 (docosahexaenoic acid), and the sole isoprenoid quinone produced was ubiqinone-8. DNA G+C content was 48.6 mol%. The strain was positive for oxidase and catalase activities. Based on the results from this study, strain MTCD1(T) is a novel Gram-negative species of the genus Colwellia, and the name Colwellia marinimaniae sp. nov. (type strain MTCD1(T) = ATCC TSD-5(T) = JCM 30270(T)) is proposed. It is the most piezophilic organism yet described.

2016
Lan, Y, Sun J, Bartlett DH, Rouse GW, Tabata HG, Qian PY.  2016.  The deepest mitochondrial genome sequenced from Mariana Trench Hirondellea gigas (Amphipoda). Mitochondrial DNA Part B-Resources. 1:802-803.   10.1080/23802359.2016.1214549   AbstractWebsite

Hirondellea gigas is an amphipod that is a dominant animal resident living in the Challenger Deep (similar to 11,000m depth) of the Mariana Trench, which is the world deepest point in the ocean. Here we report a nearly complete mitochondrial genome of H. gigas, the world deepest mitogenome. The genome consists of two contigs with lengths of 8,603 bp and 6,984 bp, respectively, and it includes 13 complete protein-coding genes, 2 rRNA genes, and 21 tRNA genes. The ends of both contigs are highly repetitive and AT rich. The gene order of H. gigas is similar to another amphipod (Onisimus nanseni) in the same superfamily, Lysianassoidea. Phylogenetic analysis showed that Lysianassoidea is grouped with Gammaroidea and Calliopioidea in the same clade. Our result also suggested that the H. gigas collected from Izu-Bonin Trench and Japan Trench are indeed the same species as those from the Mariana Trench. These results will contribute to a better understanding of the phylogeny of amphipod and other hadal species.

Tarn, J, Peoples LM, Hardy K, Cameron J, Bartlett DH.  2016.  Identification of free-living and particle-associated microbial communities present in hadal regions of the Mariana Trench. Frontiers in Microbiology. 7   10.3389/fmicb.2016.00665   AbstractWebsite

Relatively few studies have described the microbial populations present in ultra-deep hadal environments, largely as a result of difficulties associated with sampling. Here we report Illumina-tag V6 16S rRNA sequence-based analyses of the free-living and particle-associated microbial communities recovered from locations within two of the deepest hadal sites on Earth, the Challenger Deep (10,918 meters below surface-mbs) and the Sirena Deep (10,667 mbs) within the Mariana Trench, as well as one control site (Ulithi Atoll, 761 mbs). Seawater samples were collected using an autonomous lander positioned similar to 1 m above the seafloor. The bacterial populations within the Mariana Trench bottom water samples were dissimilar to other deep-sea microbial communities, though with overlap with those of diffuse flow hydrothermal vents and deep subsurface locations. Distinct particle-associated and free-living bacterial communities were found to exist. The hadal bacterial populations were also markedly different from one another, indicating the likelihood of different chemical conditions at the two sites. In contrast to the bacteria, the hadal archaeal communities were more similar to other less deep datasets and to each other due to an abundance of cosmopolitan deep-sea taxa. The hadal communities were enriched in 34 bacterial and 4 archaeal operational taxonomic units (OTUs) including members of the Gammaproteobacteria, Epsilonproteobacteria, Marinimicrobia, Cyanobacteria, Deltaproteobacteria, Gemmatimonadetes, Atribacteria, Spirochaetes, and Euryarchaeota. Sequences matching cultivated piezophiles were notably enriched in the Challenger Deep, especially within the particle-associated fraction, and were found in higher abundances than in other hadal studies, where they were either far less prevalent or missing. Our results indicate the importance of heterotrophy, sulfur-cycling, and methane and hydrogen utilization within the bottom waters of the deeper regions of the Mariana Trench, and highlight novel community features of these extreme habitats.

Huang, Q, Tran KN, Rodgers JM, Bartlett DH, Hemley RJ, Ichiye T.  2016.  A molecular perspective on the limits of life: Enzymes under pressure. Condensed Matter Physics. 19   10.5488/cmp.19.22801   AbstractWebsite

From a purely operational standpoint, the existence of microbes that can grow under extreme conditions, or "extremophiles", leads to the question of how the molecules making up these microbes can maintain both their structure and function. While microbes that live under extremes of temperature have been heavily studied, those that live under extremes of pressure have been neglected, in part due to the difficulty of collecting samples and performing experiments under the ambient conditions of the microbe. However, thermodynamic arguments imply that the effects of pressure might lead to different organismal solutions than from the effects of temperature. Observationally, some of these solutions might be in the condensed matter properties of the intracellular milieu in addition to genetic modifications of the macromolecules or repair mechanisms for the macromolecules. Here, the effects of pressure on enzymes, which are proteins essential for the growth and reproduction of an organism, and some adaptations against these effects are reviewed and amplified by the results from molecular dynamics simulations. The aim is to provide biological background for soft matter studies of these systems under pressure.

2015
Leon-Zayas, R, Novotny M, Podell S, Shepard CM, Berkenpas E, Nikolenko S, Pevzner P, Lasken RS, Bartlett DH.  2015.  Single cells within the Puerto Rico Trench suggest hadal adaptation of microbial lineages. Applied and Environmental Microbiology. 81:8265-8276.   10.1128/aem.01659-15   AbstractWebsite

Hadal ecosystems are found at a depth of 6,000 m below sea level and below, occupying less than 1% of the total area of the ocean. The microbial communities and metabolic potential in these ecosystems are largely uncharacterized. Here, we present four single amplified genomes (SAGs) obtained from 8,219 m below the sea surface within the hadal ecosystem of the Puerto Rico Trench (PRT). These SAGs are derived from members of deep-sea clades, including the Thaumarchaeota and SAR11 clade, and two are related to previously isolated piezophilic (high-pressure-adapted) microorganisms. In order to identify genes that might play a role in adaptation to deep-sea environments, comparative analyses were performed with genomes from closely related shallow-water microbes. The archaeal SAG possesses genes associated with mixotrophy, including lipoylation and the glycine cleavage pathway. The SAR11 SAG encodes glycolytic enzymes previously reported to be missing from this abundant and cosmopolitan group. The other SAGs, which are related to piezophilic isolates, possess genes that may supplement energy demands through the oxidation of hydrogen or the reduction of nitrous oxide. We found evidence for potential trench-specific gene distributions, as several SAG genes were observed only in a PRT metagenome and not in shallower deep-sea metagenomes. These results illustrate new ecotype features that might perform important roles in the adaptation of microorganisms to life in hadal environments.

Zhang, Y, Li XG, Bartlett DH, Xiao X.  2015.  Current developments in marine microbiology: high-pressure biotechnology and the genetic engineering of piezophiles. Current Opinion in Biotechnology. 33:157-164.   10.1016/j.copbio.2015.02.013   AbstractWebsite

A key aspect of marine environments is elevated pressure; for example, similar to 70% of the ocean is at a pressure of at least 38 MPa. Many types of Bacteria and Archaea reside under these high pressures, which drive oceanic biogeochemical cycles and catalyze reactions among rocks, sediments and fluids. Most marine prokaryotes are classified as piezotolerant or as (obligate)-piezophiles with few cultivated relatives. The biochemistry and physiology of these organisms are largely unknown. Recently, high-pressure cultivation technology has been combined with omics and DNA recombination methodologies to examine the physiology of piezophilic marine microorganisms. We are now beginning to understand the adaptive mechanisms of these organisms, along with their ecological functions and evolutionary processes. This knowledge is leading to the further development of high-pressure-based biotechnology.

Gallo, ND, Cameron J, Hardy K, Fryer P, Bartlett DH, Levin LA.  2015.  Submersible- and lander-observed community patterns in the Mariana and New Britain trenches: Influence of productivity and depth on epibenthic and scavenging communities. Deep-Sea Research Part I-Oceanographic Research Papers. 99:119-133.   10.1016/j.dsr.2014.12.012   AbstractWebsite

Deep-sea trenches remain one of the least explored ocean ecosystems due to the unique challenges of sampling at great depths. Five submersible dives conducted using the DEEPSEA CHALLENGER submersible generated video of undisturbed deep-sea communities at bathyal (994 m), abyssal (3755 m), and hadal (8228 m) depths in the New Britain Trench, bathyal depths near the Ulithi atoll (1192 m), and hadal depths in the Mariana Trench Challenger Deep (10908 m). The New Britain Trench is overlain by waters with higher net primary productivity (similar to 3-fold) than the Mariana Trench and nearby Ulithi, and receives substantially more allochthonous input from terrestrial sources, based on the presence of terrestrial debris in submersible video footage. Comparisons between trenches addressed how differences in productivity regime influence benthic and demersal deep-sea community structure. In addition, the scavenger community was studied using paired lander deployments to the New Britain (8233 m) and Mariana (10918 m) trenches. Differences in allochthonous input were reflected in epibenthic community abundance, biodiversity, and lifestyle representation. More productive locations were characterized by higher faunal abundances (similar to 2-fold) at both bathyal and hadal depths. In contrast, biodiversity trends showed a unimodal pattern with more food-rich areas exhibiting reduced bathyal diversity and elevated hadal diversity. Hadal scavenging communities exhibited similar higher abundance but also similar to 3-fold higher species richness in the more food-rich New Britain Trench compared to the Mariana Trench. High species- and phylum-level diversity observed in the New Britain Trench suggest that trench environments may foster higher megafaunal biodiversity than surrounding abyssal depths if food is not limiting. However, the absence of fish at our hadal sites suggests that certain groups do have physiological depth limits. Submersible video footage allowed novel in situ observation of holothurian orientation, jellyfish feeding behavior as well as lifestyle preferences for substrate, seafloor and overlying water. This study documents previously unreported species in the New Britain Trench, including an ulmariid scyphozoan (8233 m) and an acrocirrid polychaete (994 m), and reports the first observation of an abundant population of elpidiid holothurians in the Mariana Trench (10908 m). It also provides the first megafaunal community analysis of the world's deepest epibenthic community in the Mariana Trench Challenger Deep, which was composed of elpidiid holothurians, amphipods, and xenophyophores. (C) 2015 The Authors. Published by Elsevier Ltd.

Marietou, A, Nguyen ATT, Allen EE, Bartlett D.  2015.  Adaptive laboratory evolution of Escherichia coli K-12 MG1655 for growth at high hydrostatic pressure. Frontiers in Microbiology. 5   10.3389/fmicb.2014.00749   AbstractWebsite

Much of microbial life on Earth grows and reproduces under the elevated hydrostatic pressure conditions that exist in deep-ocean and deep-subsurface environments. In this study adaptive laboratory evolution (ALE) experiments were conducted to investigate the possible modification of the piezosensitive Escherichia coli for improved growth at high pressure. After approximately 500 generations of selection, a strain was isolated that acquired the ability to grow at pressure non-permissive for the parental strain. Remarkably, this strain displayed growth properties and changes in the proportion and regulation of unsaturated fatty acids that indicated the acquisition of multiple piezotolerant properties. These changes developed concomitantly with a change in the gene encoding the acyl carrier protein, which is required for fatty acid synthesis.

2014
Marietou, A, Bartlett DH.  2014.  Effects of high hydrostatic pressure on coastal bacterial community abundance and diversity. Applied and Environmental Microbiology. 80:5992-6003.   10.1128/aem.02109-14   AbstractWebsite

Hydrostatic pressure is an important parameter influencing the distribution of microbial life in the ocean. In this study, the response of marine bacterial populations from surface waters to pressures representative of those under deep-sea conditions was examined. Southern California coastal seawater collected 5 m below the sea surface was incubated in microcosms, using a range of temperatures (16 to 3 degrees C) and hydrostatic pressure conditions (0.1 to 80 MPa). Cell abundance decreased in response to pressure, while diversity increased. The morphology of the community also changed with pressurization to a predominant morphotype of small cocci. The pressure-induced community changes included an increase in the relative abundance of Alphaproteobacteria, Gammaproteobacteria, Actinobacteria, and Flavobacteria largely at the expense of Epsilonproteobacteria. Culturable high-pressure-surviving bacteria were obtained and found to be phylogenetically similar to isolates from cold and/or deep-sea environments. These results provide novel insights into the response of surface water bacteria to changes in hydrostatic pressure.

Lauro, FM, Eloe-Fadrosh EA, Richter TKS, Vitulo N, Ferriera S, Johnson JH, Bartlett DH.  2014.  Ecotype diversity and conversion in photobacterium profundum strains. Plos One. 9   10.1371/journal.pone.0096953   AbstractWebsite

Photobacterium profundum is a cosmopolitan marine bacterium capable of growth at low temperature and high hydrostatic pressure. Multiple strains of P. profundum have been isolated from different depths of the ocean and display remarkable differences in their physiological responses to pressure. The genome sequence of the deep-sea piezopsychrophilic strain Photobacterium profundum SS9 has provided some clues regarding the genetic features required for growth in the deep sea. The sequenced genome of Photobacterium profundum strain 3TCK, a non-piezophilic strain isolated from a shallow-water environment, is now available and its analysis expands the identification of unique genomic features that correlate to environmental differences and define the Hutchinsonian niche of each strain. These differences range from variations in gene content to specific gene sequences under positive selection. Genome plasticity between Photobacterium bathytypes was investigated when strain 3TCK-specific genes involved in photorepair were introduced to SS9, demonstrating that horizontal gene transfer can provide a mechanism for rapid colonisation of new environments.

Fang, JS, Li C, Zhang L, Davis T, Kato C, Bartlett DH.  2014.  Hydrogen isotope fractionation in lipid biosynthesis by the piezophilic bacterium Moritella japonica DSK1. Chemical Geology. 367:34-38.   10.1016/j.chemgeo.2013.12.018   AbstractWebsite

The delta D of fatty acids is emerging as an important marine biogeochemical proxy, but the microbiological and environmental factors controlling the variations of delta D of the lipids are not fully constrained. We report here the first measurement of D/H ratios of fatty acids in a piezophilic bacterium and show that hydrostatic pressure and the lipid biosynthetic pathway probably exerts dominant control over the delta D of fatty acids. Piezophilic bacterium Moritella japonica DSK1 was grown at a pressure of 30 MPa with glucose as substrate. Fatty acids in DSK1 showed vastly varied delta D, ranging from + 44.4 to - 171 parts per thousand. Short-chain fatty acids (SCFA), which are synthesized by the fatty acid synthase (FAS) pathway, had positive delta D (average + 3 parts per thousand), whereas long-chain polyunsaturated fatty acid (LC-PUFA) synthesized via the polyketide pathway exhibited much depleted delta D (- 171 parts per thousand). Our results suggest that the lipid biosynthetic pathways can exert first-order control on the hydrogen isotope signature of bacterial membrane lipids under elevated pressure. Our findings have important implications in marine biogeochemistry. D-depleted fatty acids in marine sediments and in the water column may be derived from piezophilic bacterial reworking and resynthesis of organic matter at high pressure condition. Thus, caution must be exercised in the interpretation of hydrogen isotope signatures of lipids in, e.g., deducing sources of organic matter and tracing microbial biogeochemical processes in the deep ocean and the deep biosphere. (C) 2014 Elsevier B.V. All rights reserved.

Cao, Y, Chastain RA, Eloe EA, Nogi Y, Kato C, Bartlett DH.  2014.  Novel Psychropiezophilic oceanospirillales species Profundimonas piezophila gen. nov., sp nov., isolated from the deep-sea environment of the Puerto Rico Trench. Applied and Environmental Microbiology. 80:54-60.   10.1128/aem.02288-13   AbstractWebsite

The diversity of deep-sea high-pressure-adapted (piezophilic) microbes in isolated monoculture remains low. In this study, a novel obligately psychropiezophilic bacterium was isolated from seawater collected from the Puerto Rico Trench at a depth of similar to 6,000 m. This isolate, designated YC-1, grew best in a nutrient-rich marine medium, with an optimal growth hydrostatic pressure of 50 MPa (range, 20 to 70 MPa) at 8 degrees C. Under these conditions, the maximum growth rate was extremely slow, 0.017 h(-1), and the maximum yield was 3.51 X 10(7) cells ml(-1). Cell size and shape changed with pressure, shifting from 4.0 to 5.0 mu m in length and 0.5 to 0.8 mu m in width at 60 MPa to 0.8-to 1.0-mu m diameter coccoid cells under 20 MPa, the minimal pressure required for growth. YC-1 is a Gram-negative, facultatively anaerobic heterotroph. Its predominant cellular fatty acids are the monounsaturated fatty acids (MUFAs) C-16:1 and C-18:1 Unlike many other psychropiezophiles, YC-1 does not synthesize any polyunsaturated fatty acids (PUFAs). Phylogenetic analysis placed YC-1 within the family of Oceanospirillaceae, closely related to the uncultured symbiont of the deep-sea whale bone-eating worms of the genus Osedax. In common with some other members of the Oceanospirillales, including those enriched during the Deepwater Horizon oil spill, YC-1 is capable of hydrocarbon utilization. On the basis of its characteristics, YC-1 appears to represent both a new genus and a new species, which we name Profundimonas piezophila gen. nov., sp. nov.

2013
Davydov, DR, Sineva EV, Davydova NY, Bartlett DH, Halpert JR.  2013.  CYP261 enzymes from deep sea bacteria: A clue to conformational heterogeneity in cytochromes P450. Biotechnology and Applied Biochemistry. 60:30-40.   10.1002/bab.1083   AbstractWebsite

We have explored the adaptation of the cytochromes P450 (P450) of deep-sea bacteria to high hydrostatic pressures. Strict conservation of the protein fold and functional importance of protein-bound water make P450 a unique subject for the studies of high-pressure adaptation. Earlier, we expressed and purified a fatty-acid binding P450 from the deep-sea bacteria Photobacterium profundum SS9 (CYP261C1). Here, we report purification and initial characterization of its mesophilic ortholog from the shallow-water P. profundum 3TCK (CYP261C2), as well as another piezophilic enzyme, CYP261D1, from deep-sea Moritella sp. PE36. Comparison of the three enzymes revealed a striking peculiarity of the piezophilic enzymes. Both CYP261C1 and CYP261D1 possess an apparent pressure-induced conformational toggle actuated at the pressures commensurate with the physiological pressure of habitation of the host bacteria. Furthermore, in contrast to CYP261C2, the piezophilic CYP261 enzymes may be chromatographically separated into two fractions with different properties, and different thermodynamic parameters of spin equilibrium in particular. According to our concept, the changes in the energy landscape that evolved in pressure-tolerant enzymes must stabilize the less-hydrated, closed conformers, which may be transient in the catalytic mechanisms of nonpiezophilic enzymes. The studies of enzymes of piezophiles should help unravel the mechanisms that control water access during the catalytic cycle.

Meersman, F, Daniel I, Bartlett DH, Winter R, Hazael R, McMillan PF.  2013.  High-Pressure Biochemistry and Biophysics. Carbon in Earth. 75( Hazen RM, Jones AP, Baross JA, Eds.).:607-648., Chantilly: Mineralogical Soc Amer   10.2138/rmg.2013.75.19   Abstract
n/a
2012
Campanaro, S, De Pascale F, Telatin A, Schiavon R, Bartlett DH, Valle G.  2012.  The transcriptional landscape of the deep-sea bacterium Photobacterium profundum in both a toxR mutant and its parental strain. BMC Genomics. 13   10.1186/1471-2164-13-567Rita vj, 1991, cell, v64, p29   AbstractWebsite

Background: The deep-sea bacterium Photobacterium profundum is an established model for studying high pressure adaptation. In this paper we analyse the parental strain DB110 and the toxR mutant TW30 by massively parallel cDNA sequencing (RNA-seq). ToxR is a transmembrane DNA-binding protein first discovered in Vibrio cholerae, where it regulates a considerable number of genes involved in environmental adaptation and virulence. In P. profundum the abundance and activity of this protein is influenced by hydrostatic pressure and its role is related to the regulation of genes in a pressure-dependent manner. Results: To better characterize the ToxR regulon, we compared the expression profiles of wt and toxR strains in response to pressure changes. Our results revealed a complex expression pattern with a group of 22 genes having expression profiles similar to OmpH that is an outer membrane protein transcribed in response to high hydrostatic pressure. Moreover, RNA-seq allowed a deep characterization of the transcriptional landscape that led to the identification of 460 putative small RNA genes and the detection of 298 protein-coding genes previously unknown. We were also able to perform a genome-wide prediction of operon structure, transcription start and termination sites, revealing an unexpected high number of genes (992) with large 5'-UTRs, long enough to harbour cis-regulatory RNA structures, suggesting a correlation between intergenic region size and UTR length. Conclusion: This work led to a better understanding of high-pressure response in P. profundum. Furthermore, the high-resolution RNA-seq analysis revealed several unexpected features about transcriptional landscape and general mechanisms of controlling bacterial gene expression.

Lucas, S, Han J, Lapidus A, Cheng JF, Goodwin LA, Pitluck S, Peters L, Mikhailova N, Teshima H, Detter JC, Han C, Tapia R, Land M, Hauser L, Kyrpides NC, Ivanova N, Pagani I, Vannier P, Oger P, Bartlett DH, Noll KM, Woyke T, Jebbar M.  2012.  Complete Genome Sequence of the Thermophilic, Piezophilic, Heterotrophic Bacterium Marinitoga piezophila KA3. Journal of Bacteriology. 194:5974-5975.   10.1128/JB.01430-12   Abstract

Marinitoga piezophila KA3 is a thermophilic, anaerobic, chemoorganotrophic, sulfur-reducing bacterium isolated from the Grandbonum deep-sea hydrothermal vent site at the East Pacific Rise (13 degrees N, 2,630-m depth). The genome of M. piezophila KA3 comprises a 2,231,407-bp circular chromosome and a 13,386-bp circular plasmid. This genome was sequenced within Department of Energy Joint Genome Institute CSP 2010.

2011
Eloe, EA, Malfatti F, Gutierrez J, Hardy K, Schmidt WE, Pogliano K, Pogliano J, Azam F, Bartlett DH.  2011.  Isolation and Characterization of a Psychropiezophilic Alphaproteobacterium. Applied and Environmental Microbiology. 77:8145-8153.   10.1128/aem.05204-11   AbstractWebsite

Cultivated psychropiezophilic (low-temperature- and high-pressure-adapted) bacteria are currently restricted to phylogenetically narrow groupings capable of growth under nutrient-replete conditions, limiting current knowledge of the extant functional attributes and evolutionary constraints of diverse microorganisms inhabiting the cold, deep ocean. This study documents the isolation of a deep-sea bacterium following dilution-to-extinction cultivation using a natural seawater medium at high hydrostatic pressure and low temperature. To our knowledge, this isolate, designated PRT1, is the slowest-growing (minimal doubling time, 36 h) and lowest cell density-producing (maximal densities of 5.0 x 10(6) cells ml(-1)) piezophile yet obtained. Optimal growth was at 80 MPa, correlating with the depth of capture (8,350 m), and 10 degrees C, with average cell sizes of 1.46 mu m in length and 0.59 mu m in width. Through detailed growth studies, we provide further evidence for the temperature-pressure dependence of the growth rate for deep-ocean bacteria. PRT1 was phylogenetically placed within the Roseobacter clade, a bacterial lineage known for widespread geographic distribution and assorted lifestyle strategies in the marine environment. Additionally, the gene transfer agent (GTA) g5 capsid protein gene was amplified from PRT1, indicating a potential mechanism for increased genetic diversification through horizontal gene transfer within the hadopelagic environment. This study provides a phylogenetically novel isolate for future investigations of high-pressure adaptation, expands the known physiological traits of cultivated members of the Roseobacter lineage, and demonstrates the feasibility of cultivating novel microbial members from the deep ocean using natural seawater.

Eloe, EA, Fadrosh DW, Novotny M, Allen LZ, Kim M, Lombardo MJ, Yee-Greenbaum J, Yooseph S, Allen EE, Lasken R, Williamson SJ, Bartlett DH.  2011.  Going Deeper: Metagenome of a Hadopelagic Microbial Community. Plos One. 6   10.1371/journal.pone.0020388   AbstractWebsite

The paucity of sequence data from pelagic deep-ocean microbial assemblages has severely restricted molecular exploration of the largest biome on Earth. In this study, an analysis is presented of a large-scale 454-pyrosequencing metagenomic dataset from a hadopelagic environment from 6,000 m depth within the Puerto Rico Trench (PRT). A total of 145 Mbp of assembled sequence data was generated and compared to two pelagic deep ocean metagenomes and two representative surface seawater datasets from the Sargasso Sea. In a number of instances, all three deep metagenomes displayed similar trends, but were most magnified in the PRT, including enrichment in functions for two-component signal transduction mechanisms and transcriptional regulation. Overrepresented transporters in the PRT metagenome included outer membrane porins, diverse cation transporters, and di- and tri-carboxylate transporters that matched well with the prevailing catabolic processes such as butanoate, glyoxylate and dicarboxylate metabolism. A surprisingly high abundance of sulfatases for the degradation of sulfated polysaccharides were also present in the PRT. The most dramatic adaptational feature of the PRT microbes appears to be heavy metal resistance, as reflected in the large numbers of transporters present for their removal. As a complement to the metagenome approach, single-cell genomic techniques were utilized to generate partial whole-genome sequence data from four uncultivated cells from members of the dominant phyla within the PRT, Alphaproteobacteria, Gammaproteobacteria, Bacteroidetes and Planctomycetes. The single-cell sequence data provided genomic context for many of the highly abundant functional attributes identified from the PRT metagenome, as well as recruiting heavily the PRT metagenomic sequence data compared to 172 available reference marine genomes. Through these multifaceted sequence approaches, new insights have been provided into the unique functional attributes present in microbes residing in a deeper layer of the ocean far removed from the more productive sun-drenched zones above.

Phillips, RS, Ghaffari R, Dinh P, Lima S, Bartlett D.  2011.  Properties of tryptophan indole-lyase from a piezophilic bacterium, Photobacterium profundum SS9. Archives of Biochemistry and Biophysics. 506:35-41.   10.1016/j.abb.2010.11.002   AbstractWebsite

Tryptophan indole-lyase (Trpase), PBPRA2532, from Photobacterium profundum SS9, a piezophilic marine bacterium, has been cloned, expressed in Escherichia coli, and purified. The P. profundum Trpase (PpTrpase) exhibits similar substrate specificity as the enzyme from E. coli (EcTrpase). PpTrpase has an optimum temperature for activity at about 30 degrees C, compared with 53 degrees C for EcTrpase, and loses activity rapidly (t(1/2) similar to 30 min) when incubated at 50 degrees C, while EcTrpase is stable up to 65 degrees C. PpTrpase retains complete activity when incubated more than 3 h at 0 degrees C, while EcTrpase has only about 20% remaining activity. Under hydrostatic pressure, PpTrpase remains fully active up to 100 MPa (986 atm), while EcTrpase exhibits only about 10% activity at 100 MPa. PpTrpase forms external aldimine and quinonoid intermediates in stopped-flow experiments with L-Trp, S-Et-L-Cys, S-benzyl-L-Cys, oxindolyl-L-Ala, L-Ala and L-Met, similar to EcTrpase. However, with L-Trp a gem-diamine is observed that decays to a quinonoid complex. An aminoacrylate is observed with L-Trp in the presence of benzimidazole, as was seen previously with EcTrpase [28] but not with S-Et-L-Cys. The results show that PpTrpase is adapted for optimal activity in the low temperature, high pressure marine environment. (C) 2010 Elsevier Inc. All rights reserved.

Oger, P, Sokolova TG, Kozhevnikova DA, Chernyh NA, Bartlett DH, Bonch-Osmolovskaya EA, Lebedinsky AV.  2011.  Complete genome sequence of the hyperthermophilic archaeon Thermococcus sp strain AM4, capable of organotrophic growth and growth at the expense of hydrogenogenic or sulfidogenic oxidation of carbon monoxide. Journal of Bacteriology. 193:7019-7020.   10.1128/jb.06259-11   AbstractWebsite

Analysis of the complete genome of Thermococcus sp. strain AM4, which was the first lithotrophic Thermococcales isolate described and the first archaeal isolate to exhibit a capacity for hydrogenogenic carboxydotrophy, reveals a proximity with Thermococcus gammatolerans, corresponding to close but distinct species that differ significantly in their lithotrophic capacities.

Eloe, EA, Shulse CN, Fadrosh DW, Williamson SJ, Allen EE, Bartlett DH.  2011.  Compositional differences in particle-associated and free-living microbial assemblages from an extreme deep-ocean environment. Environmental Microbiology Reports. 3:449-458.: Blackwell Publishing Ltd   10.1111/j.1758-2229.2010.00223.x   AbstractWebsite

Relatively little information is available for the composition of microbial communities present in hadal environments, the deepest marine locations. Here we present a description of the phylogenetic diversity of particle-associated (> 3 µm) and free-living (3–0.22 µm) microorganisms present in a pelagic trench environment. Small subunit ribosomal RNA gene sequences were recovered from members of the Bacteria, Archaea and Eukarya obtained from a depth of 6000 m in the Puerto Rico Trench (PRT). Species richness estimates for the bacterial particle-associated fraction were greater compared with the free-living fraction and demonstrated statistically significant compositional differences, while the archaeal fractions were not found to be significantly different. The particle-associated fraction contained more Rhodobacterales and unclassified Myxococcales along with Bacteroidetes, Planctomycetes and chloroplast sequences, whereas the free-living fraction contained more Caulobacterales, Xanthomonadales and Burkholderiales, along with Marine Group A and Gemmatimonadetes. The Eukarya contained a high abundance of Basidiomycota Fungi 18S rRNA genes, as well as representatives from the super-groups Rhizaria, Excavata and Chromalveolata. A diverse clade of diplonemid flagellates was also identified from the eukaryotic phylotypes recovered, which was distinct from previously identified deep-sea pelagic diplonemid groups. The significance of these results to considerations of deep-sea microbial life and particle colonization is discussed in comparison to the few other deep-ocean phylogenetic surveys available.

2010
Yoshioka, H, Maruyama A, Nakamura T, Higashi Y, Fuse H, Sakata S, Bartlett DH.  2010.  Activities and distribution of methanogenic and methane-oxidizing microbes in marine sediments from the Cascadia Margin. Geobiology. 8:223-233.   10.1111/j.1472-4669.2009.00231.x   AbstractWebsite

We investigated methane production and oxidation and the depth distribution and phylogenetic affiliation of a functional gene for methanogenesis, methyl coenzyme M reductase subunit A (mcrA), at two sites of the Integrated Ocean Drilling Program Expedition 311. These sites, U1327 and U1329, are respectively inside and outside the area of gas hydrate distribution on the Cascadia Margin. Radiotracer experiments using 14C-labelled substrates indicated high potential methane production rates in hydrate-bearing sediments [128-223 m below seafloor (mbsf)] at U1327 and in sediments between 70 and 140 mbsf at U1329. Tracer-free experiments indicated high cumulative methane production in sediments within and below the gas hydrate layer at U1327 and in sediments below 70 mbsf at U1329. Stable tracer experiments using 13C-labelled methane showed high potential methane oxidation rates in near-surface sediments and in sediments deeper than 100 mbsf at both sites. Results of polymerase chain reaction amplification of mcrA in DNA were mostly consistent with methane production: relatively strong mcrA amplification was detected in the gas hydrate-bearing sediments at U1327, whereas at U1329, it was mainly detected in sediments from around the bottom-simulating reflector (126 mbsf). Phylogenetic analysis of mcrA separated it into four phylotype clusters: two clusters of methanogens, Methanosarcinales and Methanobacteriales, and two clusters of anaerobic methanotrophic archaea, ANME-I and ANME-II groups, supporting the activity measurement results. These results reveal that in situ methanogenesis in deep sediments probably contributes to gas hydrate formation and are inconsistent with the geochemical model that microbial methane currently being generated in shallow sediments migrates downward and contributes to the hydrate formation. At Site U1327, gas hydrates occurred in turbidite sediments, which were absent at Site U1329, suggesting that a geological setting suitable for a gas hydrate reservoir is more important for the accumulation of gas hydrate than microbiological properties.

Purdy, AE, Balch D, Lizarraga-Partida ML, Islam MS, Martinez-Urtaza J, Huq A, Colwell RR, Bartlett DH.  2010.  Diversity and distribution of cholix toxin, a novel ADP-ribosylating factor from Vibrio cholerae. Environmental Microbiology Reports. 2:198-207.   10.1111/j.1758-2229.2010.00139.x   AbstractWebsite

Non-toxigenic non-O1, non-O139 Vibrio cholerae strains isolated from both environmental and clinical settings carry a suite of virulence factors aside from cholera toxin. Among V. cholerae strains isolated from coastal waters of southern California, this includes cholix toxin, an ADP-ribosylating factor that is capable of halting protein synthesis in eukaryotic cells. The prevalence of the gene encoding cholix toxin, chxA, was assessed among a collection of 155 diverse V. cholerae strains originating from both clinical and environmental settings in Bangladesh and Mexico and other countries around the globe. The chxA gene was present in 47% of 83 non-O1, non-O139 strains and 16% of 72 O1/O139 strains screened as part of this study. A total of 86 chxA gene sequences were obtained, and phylogenetic analysis revealed that they fall into two distinct clades. These two clades were also observed in the phylogenies of several housekeeping genes, suggesting that the divergence observed in chxA extends to other regions of the V. cholerae genome, and most likely has arisen from vertical descent rather than horizontal transfer. Our results clearly indicate that ChxA is a major toxin of V. cholerae with a worldwide distribution that is preferentially associated with nonpandemic strains.