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1.  Characterization of air contaminants formed by the interaction of lava and sea water. 
Environmental Health Perspectives  1994;102(5):478-482.
We made environmental measurements to characterize contaminants generated when basaltic lava from Hawaii's Kilauea volcano enters sea water. This interaction of lava with sea water produces large clouds of mist (LAZE). Island winds occasionally directed the LAZE toward the adjacent village of Kalapana and the Hawaii Volcanos National Park, creating health concerns. Environmental samples were taken to measure airborne concentrations of respirable dust, crystalline silica and other mineral compounds, fibers, trace metals, inorganic acids, and organic and inorganic gases. The LAZE contained quantifiable concentrations of hydrochloric acid (HCl) and hydrofluoric acid (HF); HCl was predominant. HCl and HF concentrations were highest in dense plumes of LAZE near the sea. The HCl concentration at this sampling location averaged 7.1 ppm; this exceeds the current occupational exposure ceiling of 5 ppm. HF was detected in nearly half the samples, but all concentrations were <1 ppm Sulfur dioxide was detected in one of four short-term indicator tube samples at approximately 1.5 ppm. Airborne particulates were composed largely of chloride salts (predominantly sodium chloride). Crystalline silica concentrations were below detectable limits, less than approximately 0.03 mg/m3 of air. Settled dust samples showed a predominance of glass flakes and glass fibers. Airborne fibers were detected at quantifiable levels in 1 of 11 samples. These fibers were composed largely of hydrated calcium sulfate. These findings suggest that individuals should avoid concentrated plumes of LAZE near its origin to prevent over exposure to inorganic acids, specifically HCl.
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PMCID: PMC1567144  PMID: 8593853
2.  Cultivation and Complete Genome Sequencing of Gloeobacter kilaueensis sp. nov., from a Lava Cave in Kīlauea Caldera, Hawai'i 
PLoS ONE  2013;8(10):e76376.
The ancestor of Gloeobacter violaceus PCC 7421T is believed to have diverged from that of all known cyanobacteria before the evolution of thylakoid membranes and plant plastids. The long and largely independent evolutionary history of G. violaceus presents an organism retaining ancestral features of early oxygenic photoautotrophs, and in whom cyanobacteria evolution can be investigated. No other Gloeobacter species has been described since the genus was established in 1974 (Rippka et al., Arch Microbiol 100:435). Gloeobacter affiliated ribosomal gene sequences have been reported in environmental DNA libraries, but only the type strain's genome has been sequenced. However, we report here the cultivation of a new Gloeobacter species, G. kilaueensis JS1T, from an epilithic biofilm in a lava cave in Kīlauea Caldera, Hawai'i. The strain's genome was sequenced from an enriched culture resembling a low-complexity metagenomic sample, using 9 kb paired-end 454 pyrosequences and 400 bp paired-end Illumina reads. The JS1T and G. violaceus PCC 7421T genomes have little gene synteny despite sharing 2842 orthologous genes; comparing the genomes shows they do not belong to the same species. Our results support establishing a new species to accommodate JS1T, for which we propose the name Gloeobacter kilaueensis sp. nov. Strain JS1T has been deposited in the American Type Culture Collection (BAA-2537), the Scottish Marine Institute's Culture Collection of Algae and Protozoa (CCAP 1431/1), and the Belgian Coordinated Collections of Microorganisms (ULC0316). The G. kilaueensis holotype has been deposited in the Algal Collection of the US National Herbarium (US# 217948). The JS1T genome sequence has been deposited in GenBank under accession number CP003587. The G+C content of the genome is 60.54 mol%. The complete genome sequence of G. kilaueensis JS1T may further understanding of cyanobacteria evolution, and the shift from anoxygenic to oxygenic photosynthesis.
doi:10.1371/journal.pone.0076376
PMCID: PMC3806779  PMID: 24194836
3.  Characterization of Phototrophic Purple Nonsulfur Bacteria Forming Colored Microbial Mats in a Swine Wastewater Ditch 
The community structure of pink-colored microbial mats naturally occurring in a swine wastewater ditch was studied by culture-independent biomarker and molecular methods as well as by conventional cultivation methods. The wastewater in the ditch contained acetate and propionate as the major carbon nutrients. Thin-section electron microscopy revealed that the microbial mats were dominated by rod-shaped cells containing intracytoplasmic membranes of the lamellar type. Smaller numbers of oval cells with vesicular internal membranes were also found. Spectroscopic analyses of the cell extract from the biomats showed the presence of bacteriochlorophyll a and carotenoids of the spirilloxanthin series. Ubiquinone-10 was detected as the major quinone. A clone library of the photosynthetic gene, pufM, constructed from the bulk DNA of the biomats showed that all of the clones were derived from members of the genera Rhodobacter and Rhodopseudomonas. The dominant phototrophic bacteria from the microbial mats were isolated by cultivation methods and identified as being of the genera Rhodobacter and Rhodopseudomonas by studying 16S rRNA and pufM gene sequence information. Experiments of oxygen uptake with lower fatty acids revealed that the freshly collected microbial mats and the Rhodopseudomonas isolates had a wider spectrum of carbon utilization and a higher affinity for acetate than did the Rhodobacter isolates. These results demonstrate that the microbial mats were dominated by the purple nonsulfur bacteria of the genera Rhodobacter and Rhodopseudomonas, and the bioavailability of lower fatty acids in wastewater is a key factor allowing the formation of visible microbial mats with these phototrophs.
doi:10.1128/AEM.00796-06
PMCID: PMC1563592  PMID: 16957249
4.  Deposition of Biogenic Iron Minerals in a Methane Oxidizing Microbial Mat 
Archaea  2013;2013:102972.
The syntrophic community between anaerobic methanotrophic archaea and sulfate reducing bacteria forms thick, black layers within multi-layered microbial mats in chimney-like carbonate concretions of methane seeps located in the Black Sea Crimean shelf. The microbial consortium conducts anaerobic oxidation of methane, which leads to the formation of mainly two biomineral by-products, calcium carbonates and iron sulfides, building up these chimneys. Iron sulfides are generated by the microbial reduction of oxidized sulfur compounds in the microbial mats. Here we show that sulfate reducing bacteria deposit biogenic iron sulfides extra- and intracellularly, the latter in magnetosome-like chains. These chains appear to be stable after cell lysis and tend to attach to cell debris within the microbial mat. The particles may be important nuclei for larger iron sulfide mineral aggregates.
doi:10.1155/2013/102972
PMCID: PMC3697272  PMID: 23843725
5.  An interlaboratory comparison of 16S rRNA gene-based terminal restriction fragment length polymorphism and sequencing methods for assessing microbial diversity of seafloor basalts 
Environmental Microbiology  2009;11(7):1728-1735.
We present an interlaboratory comparison between full-length 16S rRNA gene sequence analysis and terminal restriction fragment length polymorphism (TRFLP) for microbial communities hosted on seafloor basaltic lavas, with the goal of evaluating how similarly these two different DNA-based methods used in two independent labs would estimate the microbial diversity of the same basalt samples. Two samples were selected for these analyses based on differences detected in the overall levels of microbial diversity between them. Richness estimators indicate that TRFLP analysis significantly underestimates the richness of the relatively high-diversity seafloor basalt microbial community: at least 50% of species from the high-diversity site are missed by TRFLP. However, both methods reveal similar dominant species from the samples, and they predict similar levels of relative diversity between the two samples. Importantly, these results suggest that DNA-extraction or PCR-related bias between the two laboratories is minimal. We conclude that TRFLP may be useful for relative comparisons of diversity between basalt samples, for identifying dominant species, and for estimating the richness and evenness of low-diversity, skewed populations of seafloor basalt microbial communities, but that TRFLP may miss a majority of species in relatively highly diverse samples.
doi:10.1111/j.1462-2920.2009.01899.x
PMCID: PMC2784043  PMID: 19508561
6.  Olivine-Respiring Bacteria Isolated from the Rock-Ice Interface in a Lava-Tube Cave, a Mars Analog Environment 
Astrobiology  2012;12(1):9-18.
Abstract
The boundary between ice and basalt on Earth is an analogue for some near-surface environments of Mars. We investigated neutrophilic iron-oxidizing microorganisms from the basalt-ice interface in a lava tube from the Oregon Cascades with perennial ice. One of the isolates (Pseudomonas sp. HerB) can use ferrous iron Fe(II) from the igneous mineral olivine as an electron donor and O2 as an electron acceptor. The optimum growth temperature is ∼12–14°C, but growth also occurs at 5°C. Bicarbonate is a facultative source of carbon. Growth of Pseudomonas sp. HerB as a chemolithotrophic iron oxidizer with olivine as the source of energy is favored in low O2 conditions (e.g., 1.6% O2). Most likely, microbial oxidation of olivine near pH 7 requires low O2 to offset the abiotic oxidation of iron. The metabolic capabilities of this bacterium would allow it to live in near-surface, icy, volcanic environments of Mars in the present or recent geological past and make this type of physiology a prime candidate in the search for life on Mars. Key Words: Extremophiles—Mars—Olivine—Iron-oxidizing bacteria—Redox. Astrobiology 12, 9–18.
doi:10.1089/ast.2011.0639
PMCID: PMC3264960  PMID: 22165996
7.  Unexpected Diversity and Complexity of the Guerrero Negro Hypersaline Microbial Mat 
We applied nucleic acid-based molecular methods, combined with estimates of biomass (ATP), pigments, and microelectrode measurements of chemical gradients, to map microbial diversity vertically on a millimeter scale in a hypersaline microbial mat from Guerrero Negro, Baja California Sur, Mexico. To identify the constituents of the mat, small-subunit rRNA genes were amplified by PCR from community genomic DNA extracted from layers, cloned, and sequenced. Bacteria dominated the mat and displayed unexpected and unprecedented diversity. The majority (1,336) of the 1,586 bacterial 16S rRNA sequences generated were unique, representing 752 species (≥97% rRNA sequence identity) in 42 of the main bacterial phyla, including 15 novel candidate phyla. The diversity of the mat samples differentiated according to the chemical milieu defined by concentrations of O2 and H2S. Bacteria of the phylum Chloroflexi formed the majority of the biomass by percentage of bulk rRNA and of clones in rRNA gene libraries. This result contradicts the general belief that cyanobacteria dominate these communities. Although cyanobacteria constituted a large fraction of the biomass in the upper few millimeters (>80% of the total rRNA and photosynthetic pigments), Chloroflexi sequences were conspicuous throughout the mat. Filamentous Chloroflexi bacteria were identified by fluorescence in situ hybridization within the polysaccharide sheaths of the prominent cyanobacterium Microcoleus chthonoplastes, in addition to free living in the mat. The biological complexity of the mat far exceeds that observed in other polysaccharide-rich microbial ecosystems, such as the human and mouse distal guts, and suggests that positive feedbacks exist between chemical complexity and biological diversity.
doi:10.1128/AEM.72.5.3685-3695.2006
PMCID: PMC1472358  PMID: 16672518
8.  Comparative Metagenomics of Two Microbial Mats at Cuatro Ciénegas Basin I: Ancient Lessons on How to Cope with an Environment Under Severe Nutrient Stress 
Astrobiology  2012;12(7):648-658.
Abstract
The Cuatro Ciénegas Basin (CCB) is an oasis in the desert of Mexico characterized by low phosphorus availability and by its great diversity of microbial mats. We compared the metagenomes of two aquatic microbial mats from the CCB with different nutrient limitations. We observed that the red mat was P-limited and dominated by Pseudomonas, while the green mat was N-limited and had higher species richness, with Proteobacteria and Cyanobacteria as the most abundant phyla. From their gene content, we deduced that both mats were very metabolically diverse despite their use of different strategies to cope with their respective environments. The red mat was found to be mostly heterotrophic, while the green mat was more autotrophic. The red mat had a higher number of transporters in general, including transporters of cellobiose and osmoprotectants. We suggest that generalists with plastic genomes dominate the red mat, while specialists with minimal genomes dominate the green mat. Nutrient limitation was a common scenario on the early planet; despite this, biogeochemical cycles were performed, and as a result the planet changed. The metagenomes of microbial mats from the CCB show the different strategies a community can use to cope with oligotrophy and persist. Key Words: Microbial mats—Metagenomics—Metabolism. Astrobiology 12, 648–658.
doi:10.1089/ast.2011.0694
PMCID: PMC3426886  PMID: 22920515
9.  Local Extinction and Unintentional Rewilding of Bighorn Sheep (Ovis canadensis) on a Desert Island 
PLoS ONE  2014;9(3):e91358.
Bighorn sheep (Ovis canadensis) were not known to live on Tiburón Island, the largest island in the Gulf of California and Mexico, prior to the surprisingly successful introduction of 20 individuals as a conservation measure in 1975. Today, a stable island population of ∼500 sheep supports limited big game hunting and restocking of depleted areas on the Mexican mainland. We discovered fossil dung morphologically similar to that of bighorn sheep in a dung mat deposit from Mojet Cave, in the mountains of Tiburón Island. To determine the origin of this cave deposit we compared pellet shape to fecal pellets of other large mammals, and extracted DNA to sequence mitochondrial DNA fragments at the 12S ribosomal RNA and control regions. The fossil dung was 14C-dated to 1476–1632 calendar years before present and was confirmed as bighorn sheep by morphological and ancient DNA (aDNA) analysis. 12S sequences closely or exactly matched known bighorn sheep sequences; control region sequences exactly matched a haplotype described in desert bighorn sheep populations in southwest Arizona and southern California and showed subtle differentiation from the extant Tiburón population. Native desert bighorn sheep previously colonized this land-bridge island, most likely during the Pleistocene, when lower sea levels connected Tiburón to the mainland. They were extirpated sometime in the last ∼1500 years, probably due to inherent dynamics of isolated populations, prolonged drought, and (or) human overkill. The reintroduced population is vulnerable to similar extinction risks. The discovery presented here refutes conventional wisdom that bighorn sheep are not native to Tiburón Island, and establishes its recent introduction as an example of unintentional rewilding, defined here as the introduction of a species without knowledge that it was once native and has since gone locally extinct.
doi:10.1371/journal.pone.0091358
PMCID: PMC3960132  PMID: 24646515
10.  Episodic Eruptions of Volcanic Ash Trigger a Reversible Cascade of Nuisance Species Outbreaks in Pristine Coral Habitats 
PLoS ONE  2012;7(10):e46639.
Volcanically active islands abound in the tropical Pacific and harbor complex coral communities. Whereas lava streams and deep ash deposits are well-known to devastate coral communities through burial and smothering, little is known about the effect of moderate amounts of small particulate ash deposits on reef communities. Volcanic ash contains a diversity of chemical compounds that can induce nutrient enrichments triggering changes in benthic composition. Two independently collected data sets on the marine benthos of the pristine and remote reefs around Pagan Island, Northern Mariana Islands, reveal a sudden critical transition to cyanobacteria-dominated communities in 2009–2010, which coincides with a period of continuous volcanic ash eruptions. Concurrently, localized outbreaks of the coral-killing cyanobacteriosponge Terpios hoshinota displayed a remarkable symbiosis with filamentous cyanobacteria, which supported the rapid overgrowth of massive coral colonies and allowed the sponge to colonize substrate types from which it has not been documented before. The chemical composition of tephra from Pagan indicates that the outbreak of nuisance species on its reefs might represent an early succession stage of iron enrichment (a.k.a. “black reefs”) similar to that caused by anthropogenic debris like ship wrecks or natural events like particulate deposition from wildfire smoke plumes or desert dust storms. Once Pagan's volcanic activity ceased in 2011, the cyanobacterial bloom disappeared. Another group of well-known nuisance algae in the tropical Pacific, the pelagophytes, did not reach bloom densities during this period of ash eruptions but new species records for the Northern Mariana Islands were documented. These field observations indicate that the study of population dynamics of pristine coral communities can advance our understanding of the resilience of tropical reef systems to natural and anthropogenic disturbances.
doi:10.1371/journal.pone.0046639
PMCID: PMC3464252  PMID: 23056381
11.  Millimeter-Scale Patterns of Phylogenetic and Trait Diversity in a Salt Marsh Microbial Mat 
Intertidal microbial mats are comprised of distinctly colored millimeter-thick layers whose communities organize in response to environmental gradients such as light availability, oxygen/sulfur concentrations, and redox potential. Here, slight changes in depth correspond to sharp niche boundaries. We explore the patterns of biodiversity along this depth gradient as it relates to functional groups of bacteria, as well as trait-encoding genes. We used molecular techniques to determine how the mat’s layers differed from one another with respect to taxonomic, phylogenetic, and trait diversity, and used these metrics to assess potential drivers of community assembly. We used a range of null models to compute the degree of phylogenetic and functional dispersion for each layer. The SSU-rRNA reads were dominated by Cyanobacteria and Chromatiales, but contained a high taxonomic diversity. The composition of each mat core was significantly different for developmental stage, year, and layer. Phylogenetic richness and evenness positively covaried with depth, and trait richness tended to decrease with depth. We found evidence for significant phylogenetic clustering for all bacteria below the surface layer, supporting the role of habitat filtering in the assembly of mat layers. However, this signal disappeared when the phylogenetic dispersion of particular functional groups, such as oxygenic phototrophs, was measured. Overall, trait diversity measured by orthologous genes was also lower than would be expected by chance, except for genes related to photosynthesis in the topmost layer. Additionally, we show how the choice of taxa pools, null models, spatial scale, and phylogenies can impact our ability to test hypotheses pertaining to community assembly. Our results demonstrate that given the appropriate physiochemical conditions, strong phylogenetic, and trait variation, as well as habitat filtering, can occur at the millimeter-scale.
doi:10.3389/fmicb.2012.00293
PMCID: PMC3415947  PMID: 22908010
microbial mat; community assembly; biodiversity; phylogenetics; null models; metagenomics; salt marsh
12.  One-, two- and three-phase viscosity treatments for basaltic lava flows 
Lava flows comprise three-phase mixtures of melt, crystals, and bubbles. While existing one-phase treatments allow melt phase viscosity to be assessed on the basis of composition, water content, and/or temperature, two-phase treatments constrain the effects of crystallinity or vesicularity on mixture viscosity. However, three-phase treatments, allowing for the effects of coexisting crystallinity and vesicularity, are not well understood. We investigate existing one- and two-phase treatments using lava flow case studies from Mauna Loa (Hawaii) and Mount Etna (Italy) and compare these with a three-phase treatment that has not been applied previously to basaltic mixtures. At Etna, melt viscosities of 425 ± 30 Pa s are expected for well-degassed (0.1 w. % H2O), and 135 ± 10 Pa s for less well-degassed (0.4 wt % H2O), melt at 1080°C. Application of a three-phase model yields mixture viscosities (45% crystals, 25–35% vesicles) in the range 5600–12,500 Pa s. This compares with a measured value for Etnean lava of 9400 ± 1500 Pa s. At Mauna Loa, the three-phase treatment provides a fit with the full range of field measured viscosities, giving three-phase mixture viscosities, upon eruption, of 110–140 Pa s (5% crystals, no bubble effect due to sheared vesicles) to 850–1400 Pa s (25–30% crystals, 40–60% spherical vesicles). The ability of the three-phase treatment to characterize the full range of melt-crystal-bubble mixture viscosities in both settings indicates the potential of this method in characterizing basaltic lava mixture viscosity.
doi:10.1029/2007JB005035
PMCID: PMC3117617  PMID: 21691456
13.  Consumption of Methane and CO2 by Methanotrophic Microbial Mats from Gas Seeps of the Anoxic Black Sea▿ †  
The deep anoxic shelf of the northwestern Black Sea has numerous gas seeps, which are populated by methanotrophic microbial mats in and above the seafloor. Above the seafloor, the mats can form tall reef-like structures composed of porous carbonate and microbial biomass. Here, we investigated the spatial patterns of CH4 and CO2 assimilation in relation to the distribution of ANME groups and their associated bacteria in mat samples obtained from the surface of a large reef structure. A combination of different methods, including radiotracer incubation, beta microimaging, secondary ion mass spectrometry, and catalyzed reporter deposition fluorescence in situ hybridization, was applied to sections of mat obtained from the large reef structure to locate hot spots of methanotrophy and to identify the responsible microbial consortia. In addition, CO2 reduction to methane was investigated in the presence or absence of methane, sulfate, and hydrogen. The mat had an average δ13C carbon isotopic signature of −67.1‰, indicating that methane was the main carbon source. Regions dominated by ANME-1 had isotope signatures that were significantly heavier (−66.4‰ ± 3.9 ‰ [mean ± standard deviation; n = 7]) than those of the more central regions dominated by ANME-2 (−72.9‰ ± 2.2 ‰; n = 7). Incorporation of 14C from radiolabeled CH4 or CO2 revealed one hot spot for methanotrophy and CO2 fixation close to the surface of the mat and a low assimilation efficiency (1 to 2% of methane oxidized). Replicate incubations of the mat with 14CH4 or 14CO2 revealed that there was interconversion of CH4 and CO2. The level of CO2 reduction was about 10% of the level of anaerobic oxidation of methane. However, since considerable methane formation was observed only in the presence of methane and sulfate, the process appeared to be a rereaction of anaerobic oxidation of methane rather than net methanogenesis.
doi:10.1128/AEM.02685-06
PMCID: PMC1855681  PMID: 17277205
14.  Dissolution of Arsenic Minerals Mediated by Dissimilatory Arsenate Reducing Bacteria: Estimation of the Physiological Potential for Arsenic Mobilization 
BioMed Research International  2014;2014:841892.
The aim of this study was characterization of the isolated dissimilatory arsenate reducing bacteria in the context of their potential for arsenic removal from primary arsenic minerals through reductive dissolution. Four strains, Shewanella sp. OM1, Pseudomonas sp. OM2, Aeromonas sp. OM4, and Serratia sp. OM17, capable of anaerobic growth with As (V) reduction, were isolated from microbial mats from an ancient gold mine. All of the isolated strains: (i) produced siderophores that promote dissolution of minerals, (ii) were resistant to dissolved arsenic compounds, (iii) were able to use the dissolved arsenates as the terminal electron acceptor, and (iii) were able to use copper minerals containing arsenic minerals (e.g., enargite) as a respiratory substrate. Based on the results obtained in this study, we postulate that arsenic can be released from some As-bearing polymetallic minerals (such as copper ore concentrates or middlings) under reductive conditions by dissimilatory arsenate reducers in indirect processes.
doi:10.1155/2014/841892
PMCID: PMC3958731  PMID: 24724102
15.  The biosynthesis of cyanobacterial sunscreen scytonemin in intertidal microbial mat communities 
FEMS microbiology ecology  2011;77(2):322-332.
We have examined the biosynthesis and accumulation of cyanobacterial sunscreening pigment scytonemin within intertidal microbial mat communities using a combination of chemical, molecular, and phylogenetic approaches. Both laminated (layered) and non-laminated mats contained scytonemin, with morphologically distinct mats having different cyanobacterial community compositions. Within laminated microbial mats, regions with and without scytonemin had different dominant oxygenic phototrophs, with scytonemin-producing areas consisting primarily of Lyngbya aestuarii and scytonemin-deficient areas dominated by a eukaryotic alga. The non-laminated mat was populated by a diverse group of cyanobacteria and did not contain algae. The amplification and phylogenetic assignment of scytonemin biosynthetic gene scyC from laminated mat samples confirmed that the dominant cyanobacterium in these areas, L. aestuarii, is likely responsible for sunscreen production. This study is the first to utilize an understanding of the molecular basis of scytonemin assembly to explore its synthesis and function within natural microbial communities.
doi:10.1111/j.1574-6941.2011.01113.x
PMCID: PMC3134115  PMID: 21501195
cyanobacteria; microbial mat; scytonemin; secondary metabolite biosynthesis
16.  Phylogenetic stratigraphy in the Guerrero Negro hypersaline microbial mat 
The ISME Journal  2012;7(1):50-60.
The microbial mats of Guerrero Negro (GN), Baja California Sur, Mexico historically were considered a simple environment, dominated by cyanobacteria and sulfate-reducing bacteria. Culture-independent rRNA community profiling instead revealed these microbial mats as among the most phylogenetically diverse environments known. A preliminary molecular survey of the GN mat based on only ∼1500 small subunit rRNA gene sequences discovered several new phylum-level groups in the bacterial phylogenetic domain and many previously undetected lower-level taxa. We determined an additional ∼119 000 nearly full-length sequences and 28 000 >200 nucleotide 454 reads from a 10-layer depth profile of the GN mat. With this unprecedented coverage of long sequences from one environment, we confirm the mat is phylogenetically stratified, presumably corresponding to light and geochemical gradients throughout the depth of the mat. Previous shotgun metagenomic data from the same depth profile show the same stratified pattern and suggest that metagenome properties may be predictable from rRNA gene sequences. We verify previously identified novel lineages and identify new phylogenetic diversity at lower taxonomic levels, for example, thousands of operational taxonomic units at the family-genus levels differ considerably from known sequences. The new sequences populate parts of the bacterial phylogenetic tree that previously were poorly described, but indicate that any comprehensive survey of GN diversity has only begun. Finally, we show that taxonomic conclusions are generally congruent between Sanger and 454 sequencing technologies, with the taxonomic resolution achieved dependent on the abundance of reference sequences in the relevant region of the rRNA tree of life.
doi:10.1038/ismej.2012.79
PMCID: PMC3526174  PMID: 22832344
Guerrero Negro; rRNA phylogeny; microbial mats; microbial ecology; QIIME
17.  Morphological Survey of Microbial Mats Near Deep-Sea Thermal Vents † 
A microscopic survey is presented of the most commonly observed and morphologically conspicuous microorganisms found attached to natural surfaces or to artificial materials deposited in the immediate vicinity of thermal submarine vents at the Galapagos Rift ocean spreading zone at a depth of 2,550 meters. Of special interest were the following findings: (i) all surfaces intermittently exposed to H2S-containing hydrothermal fluid were covered by layers, ca. 5 to 10 μm thick, of procaryotic, gram-negative cells interspaced with amorphous metal (Mn-Fe) deposits; (ii) although some of the cells were encased by dense metal deposits, there was little apparent correlation between metal deposition and the occurrence of microbial mats, (iii) highly differentiated forms appeared to be analogues of certain cyanobacteria, (iv) isolates from massive mats of a prosthecate bacterium could be identified as Hyphomicrobium spp., (v) intracellular membrane systems similar to those found in methylotrophic and nitrifying bacteria were observed in approximately 20% of the cells composing the mats, (vi) thiosulfate enrichments made from mat material resulted in isolations of different types of sulfur-oxidizing bacteria including the obligately chemolithotrophic genus Thiomicrospira.
Images
PMCID: PMC243726  PMID: 16345722
18.  Understanding Cervical Cancer Prevention and Screening in Chuukese Women in Hawai‘i 
Hawaii Medical Journal  2010;69(6 Suppl 3):13-16.
Background
Cervical cancer is the primary cause of death due to cancer in women in Chuuk State, Federated States of Micronesia. The Chuukese population is the fastest growing segment of the Micronesian community in Hawai‘i. Little is known about the health beliefs or practices of this population in Hawai‘i. The purpose of this project was to describe the knowledge, attitudes, and beliefs of Chuukese women in Hawai‘i regarding cervical cancer prevention and screening.
Methods
Research assistants from the Chuukese community were recruited and trained as members of the research team. A culturally sensitive survey tool was developed and piloted by the research team and used to interview ten key informants from the Chuukese community in Honolulu, Hawai‘i.
Results
There is limited knowledge about cervical cancer, especially the association with human papillomavirus (HPV). This may be indicative of a lack of health information in general. Fear, privacy concerns, lack of awareness and cultural beliefs represent the main barriers mentioned when discussing cervical cancer. Education, done in a group setting with other women, is the most recommended method of informing this community and improving preventive and screening services for cervical cancer in these women.
PMCID: PMC3123140  PMID: 20539995
19.  Breastfeeding Patterns in the Rural Community of Hilo, Hawai‘i: An Exploration of Existing Data Sets 
Before any breastfeeding promotion effort, an understanding of the existing breastfeeding patterns is essential. Hawai‘i County is a rural, ethnically diverse, medically underserved community. The purpose of this study was to describe the breastfeeding patterns of women living in Hilo, Hawai‘i. Data from several existing national, state, and local data sets were accessed to identify and describe the breastfeeding patterns of women in this community. Available breastfeeding data about women in Hilo was obtained from the Hawai‘i WIC program and includes initiation, duration, exclusivity of breastfeeding, and reasons for not breastfeeding. These data were compared to data from published reports available at the county, state, and national level. The State of Hawai‘i and Hilo exceed national targets for breastfeeding initiation; however, rates soon drop following delivery, and mixed feedings of infants is common. The highest percentage of mothers weaned their infants within the first four weeks postpartum. The reasons the majority of the mothers gave for weaning were tied to breastfeeding situations that are amenable to skilled lactation support (eg, milk supply issues and latch or sucking problems). While available data sets offer valuable information on the breastfeeding patterns in this rural community, there are limitations to their usefulness, primarily due to the inconsistent operational definitions of infant feeding variables used in the surveys, and the lack of availability of community level data.
PMCID: PMC3602946  PMID: 23520565
20.  Genome sequence of the reddish-pigmented Rubellimicrobium thermophilum type strain (DSM 16684T), a member of the Roseobacter clade 
Standards in Genomic Sciences  2013;8(3):480-490.
Rubellimicrobium thermophilum Denner et al. 2006 is the type species of the genus Rubellimicrobium, a representative of the Roseobacter clade within the Rhodobacteraceae. Members of this clade were shown to be abundant especially in coastal and polar waters, but were also found in microbial mats and sediments. They are metabolically versatile and form a physiologically heterogeneous group within the Alphaproteobacteria. Strain C-Ivk-R2A-2T was isolated from colored deposits in a pulp dryer; however, its natural habitat is so far unknown. Here we describe the features of this organism, together with the draft genome sequence and annotation and novel aspects of its phenotype. The 3,161,245 bp long genome contains 3,243 protein-coding and 45 RNA genes.
doi:10.4056/sigs.4247911
PMCID: PMC3910695  PMID: 24501632
rod-shaped; reddish-pigmented; thermophile; chemoheterotrophic; prophage-like structures; Rhodobacteraceae; Roseobacter clade; Alphaproteobacteria
21.  Ultra-diffuse hydrothermal venting supports Fe-oxidizing bacteria and massive umber deposition at 5000 m off Hawaii 
The ISME Journal  2011;5(11):1748-1758.
A novel hydrothermal field has been discovered at the base of Lōihi Seamount, Hawaii, at 5000 mbsl. Geochemical analyses demonstrate that ‘FeMO Deep', while only 0.2 °C above ambient seawater temperature, derives from a distal, ultra-diffuse hydrothermal source. FeMO Deep is expressed as regional seafloor seepage of gelatinous iron- and silica-rich deposits, pooling between and over basalt pillows, in places over a meter thick. The system is capped by mm to cm thick hydrothermally derived iron-oxyhydroxide- and manganese-oxide-layered crusts. We use molecular analyses (16S rDNA-based) of extant communities combined with fluorescent in situ hybridizations to demonstrate that FeMO Deep deposits contain living iron-oxidizing Zetaproteobacteria related to the recently isolated strain Mariprofundus ferroxydans. Bioenergetic calculations, based on in-situ electrochemical measurements and cell counts, indicate that reactions between iron and oxygen are important in supporting chemosynthesis in the mats, which we infer forms a trophic base of the mat ecosystem. We suggest that the biogenic FeMO Deep hydrothermal deposit represents a modern analog for one class of geological iron deposits known as ‘umbers' (for example, Troodos ophilolites, Cyprus) because of striking similarities in size, setting and internal structures.
doi:10.1038/ismej.2011.48
PMCID: PMC3197161  PMID: 21544100
geomicrobiology; deep biosphere; hydrothermal; iron bacteria; iron oxidation
22.  Spatial Structure and Activity of Sedimentary Microbial Communities Underlying a Beggiatoa spp. Mat in a Gulf of Mexico Hydrocarbon Seep 
PLoS ONE  2010;5(1):e8738.
Background
Subsurface fluids from deep-sea hydrocarbon seeps undergo methane- and sulfur-cycling microbial transformations near the sediment surface. Hydrocarbon seep habitats are naturally patchy, with a mosaic of active seep sediments and non-seep sediments. Microbial community shifts and changing activity patterns on small spatial scales from seep to non-seep sediment remain to be examined in a comprehensive habitat study.
Methodology/Principal Findings
We conducted a transect of biogeochemical measurements and gene expression related to methane- and sulfur-cycling at different sediment depths across a broad Beggiatoa spp. mat at Mississippi Canyon 118 (MC118) in the Gulf of Mexico. High process rates within the mat (∼400 cm and ∼10 cm from the mat's edge) contrasted with sharply diminished activity at ∼50 cm outside the mat, as shown by sulfate and methane concentration profiles, radiotracer rates of sulfate reduction and methane oxidation, and stable carbon isotopes. Likewise, 16S ribosomal rRNA, dsrAB (dissimilatory sulfite reductase) and mcrA (methyl coenzyme M reductase) mRNA transcripts of sulfate-reducing bacteria (Desulfobacteraceae and Desulfobulbaceae) and methane-cycling archaea (ANME-1 and ANME-2) were prevalent at the sediment surface under the mat and at its edge. Outside the mat at the surface, 16S rRNA sequences indicated mostly aerobes commonly found in seawater. The seep-related communities persisted at 12–20 cm depth inside and outside the mat. 16S rRNA transcripts and V6-tags reveal that bacterial and archaeal diversity underneath the mat are similar to each other, in contrast to oxic or microoxic habitats that have higher bacterial diversity.
Conclusions/Significance
The visual patchiness of microbial mats reflects sharp discontinuities in microbial community structure and activity over sub-meter spatial scales; these discontinuities have to be taken into account in geochemical and microbiological inventories of seep environments. In contrast, 12–20 cm deep in the sediments microbial communities performing methane-cycling and sulfate reduction persist at lower metabolic rates regardless of mat cover, and may increase activity rapidly when subsurface flow changes.
doi:10.1371/journal.pone.0008738
PMCID: PMC2806916  PMID: 20090951
23.  Microstructural Characterization of Cyanobacterial Mats from the McMurdo Ice Shelf, Antarctica 
The three-dimensional structures of two types of cyanobacterium-dominated microbial mats from meltwater ponds on the McMurdo Ice Shelf were as determined by using a broad suite of complementary techniques, including optical and fluorescence microscopy, confocal scanning laser microscopy, scanning electron microscopy with back-scattered electron-imaging mode, low-temperature scanning electron microscopy, and microanalyitical X-ray energy dispersive spectroscopy. By using a combination of the different in situ microscopic techniques, the Antarctic microbial mats were found to be structures with vertical stratification of groups of cyanobacteria and mineral sediments, high contents of extracellular polymeric substances, and large void spaces occupied by water. In cyanobacterium-rich layers, heterocystous nostocalean and nonheterocystous oscillatorialean taxa were the most abundant taxa and appeared to be intermixed with fine-size deposits of epicellular silica and calcium carbonate. Most of the cyanobacterial filaments had similar orientations in zones without sediment particles, but thin filaments were tangled among thicker filaments. The combination of the microscopic techniques used showed the relative positions of biological and mineral entities within the microbial mats and enabled some speculation about their interactions.
doi:10.1128/AEM.70.1.569-580.2004
PMCID: PMC321274  PMID: 14711689
24.  Identification of Members of the Metabolically Active Microbial Populations Associated with Beggiatoa Species Mat Communities from Gulf of Mexico Cold-Seep Sediments 
In this study, the composition of the metabolically active fraction of the microbial community occurring in Gulf of Mexico marine sediments (water depth, 550 to 575 m) with overlying filamentous bacterial mats was determined. The mats were mainly composed of either orange- or white-pigmented Beggiatoa spp. Complementary 16S ribosomal DNA (crDNA) was obtained from rRNA extracted from three different sediment depths (0 to 2, 6 to 8, and 10 to 12 cm) that had been subjected to reverse transcription-PCR amplification. Domain-specific 16S PCR primers were used to construct 12 different 16S crDNA libraries containing 333 Archaea and 329 Bacteria clones. Analysis of the Archaea clones indicated that all sediment depths associated with overlying orange- and white-pigmented microbial mats were almost exclusively dominated by ANME-2 (95% of total Archaea clones), a lineage related to the methanogenic order Methanosarcinales. In contrast, bacterial diversity was considerably higher, with the dominant phylotype varying by sediment depth. An equivalent number of clones detected at 0 to 2 cm, representing a total of 93%, were related to the γ and δ classes of Proteobacteria, whereas clones related to δ-Proteobacteria dominated the metabolically active fraction of the bacterial community occurring at 6 to 8 cm (79%) and 10 to 12 cm (85%). This is the first phylogenetics-based evaluation of the presumptive metabolically active fraction of the Bacteria and Archaea community structure investigated along a sediment depth profile in the northern Gulf of Mexico, a hydrocarbon-rich cold-seep region.
doi:10.1128/AEM.70.9.5447-5458.2004
PMCID: PMC520919  PMID: 15345432
25.  Formation of Multilayered Photosynthetic Biofilms in an Alkaline Thermal Spring in Yellowstone National Park, Wyoming▿  
In this study, glass rods suspended at the air-water interface in the runoff channel of Fairy Geyser, Yellowstone National Park, WY, were used as a substratum to promote the development of biofilms that resembled multilayered mat communities in the splash zone at the geyser's source. This approach enabled the establishment of the temporal relationship between the appearance of Cyanobacteria, which ultimately formed the outer green layer, and the development of a red underlayer containing Roseiflexus-like Chloroflexi. This is the first study to define time-dependent successional events involved in the development of differently colored layers within microbial mats associated with many thermal features in Yellowstone National Park. Initial (1-month) biofilms were localized below the air-water interface (60 to 70°C), and the majority of retrieved bacterial sequence types were similar to Synechococcus and Thermus isolates. Biofilms then shifted, becoming established at and above the air-water interface after 3 months. During winter sampling (6 to 8 months), distinct reddish orange microcolonies were observed, consistent with the appearance of Roseiflexus-like sequences and bacteriochlorophyll a pigment signatures. Additionally, populations of Cyanobacteria diversified to include both unicellular and filamentous cell and sequence types. Distinct green and red layers were observed at 13 months. Planctomycetes-like sequences were also retrieved in high abundance from final biofilm layers and winter samples. Finally, biomass associated with geyser vent water contained Roseiflexus-like sequence types, in addition to other high-abundance sequence types retrieved from biofilm samples, supporting the idea that geothermal water serves as an inoculum for these habitats.
doi:10.1128/AEM.01802-08
PMCID: PMC2675224  PMID: 19218404

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