Epigenetic DNA Methylation Profiling with MSRE: A Quantitative NGS Approach Using a Parkinson's Disease Test Case.

Epigenetics is a rapidly developing field focused on deciphering chemical fingerprints that accumulate on human genomes over time. As the nascent idea of precision medicine expands to encompass epigenetic signatures of diagnostic and prognostic relevance, there is a need for methodologies that provide high-throughput DNA methylation profiling measurements. Here we report a novel quantification methodology for computationally reconstructing site-specific CpG methylation status from next generation sequencing (NGS) data using methyl-sensitive restriction endonucleases (MSRE).

Transcriptional Control in Marine Copiotrophic and Oligotrophic Bacteria with Streamlined Genomes.

Unlabelled: Bacteria often respond to environmental stimuli using transcriptional control, but this may not be the case for marine bacteria such as "Candidatus Pelagibacter ubique," a cultivated representative of the SAR11 clade, the most abundant organism in the ocean. This bacterium has a small, streamlined genome and an unusually low number of transcriptional regulators, suggesting that transcriptional control is low in Pelagibacter and limits its response to environmental conditions. Transcriptome sequencing during batch culture growth revealed that only 0.

Single-cell activity of freshwater aerobic anoxygenic phototrophic bacteria and their contribution to biomass production.

Aerobic anoxygenic phototrophic (AAP) bacteria are photoheterotrophs that despite their low abundances have been hypothesized to play an ecologically and biogeochemically important role in aquatic systems. Characterizing this role requires a better understanding of the in situ dynamics and activity of AAP bacteria. Here we provide the first assessment of the single-cell activity of freshwater AAP bacteria and their contribution to total bacterial production across lakes spanning a wide trophic gradient, and explore the role of light in regulating AAP activity.

Growth rates and rRNA content of four marine bacteria in pure cultures and in the Delaware estuary.

Interpretation of 16S ribosomal RNA (rRNA) to 16S rRNA gene ratios (rRNA:rDNA) is based on a limited number of studies with rapidly growing copiotrophic bacteria. The most abundant bacteria in the ocean are oligotrophs, which probably grow more slowly than those bacteria whose rRNA:rDNA versus growth rate relationships are known. To examine whether rRNA:rDNA varies differently in oligotrophic marine bacteria than in copiotrophic bacteria, we used quantitative PCR and reverse transcriptase quantitative PCR to measure rRNA:rDNA in two marine copiotrophs and in two marine oligotrophs, including Candidatus Pelagibacter ubique HTCC1062, a coastal isolate of SAR11, the most abundant bacterial clade in the ocean.

Patterns in Abundance, Cell Size and Pigment Content of Aerobic Anoxygenic Phototrophic Bacteria along Environmental Gradients in Northern Lakes.

There is now evidence that aerobic anoxygenic phototrophic (AAP) bacteria are widespread across aquatic systems, yet the factors that determine their abundance and activity are still not well understood, particularly in freshwaters. Here we describe the patterns in AAP abundance, cell size and pigment content across wide environmental gradients in 43 temperate and boreal lakes of Québec. AAP bacterial abundance varied from 1.

Leucine incorporation by aerobic anoxygenic phototrophic bacteria in the Delaware estuary.

Aerobic anoxygenic phototrophic (AAP) bacteria are well known to be abundant in estuaries, coastal regions and in the open ocean, but little is known about their activity in any aquatic ecosystem. To explore the activity of AAP bacteria in the Delaware estuary and coastal waters, single-cell (3)H-leucine incorporation by these bacteria was examined with a new approach that combines infrared epifluorescence microscopy and microautoradiography. The approach was used on samples from the Delaware coast from August through December and on transects through the Delaware estuary in August and November 2011.

Uptake of dissolved organic carbon by gammaproteobacterial subgroups in coastal waters of the West Antarctic Peninsula.

Heterotrophic bacteria are well known to be key players in the turnover of dissolved organic material (DOM) in the oceans, but the relationship between DOM uptake and bacterial clades is still not well understood. Here we explore the turnover and single-cell use of glucose, an amino acid mixture, N-acetylglucosamine (NAG), and protein by gammaproteobacterial clades in coastal waters of the West Antarctic Peninsula in summer and fall. More than 60% of the cells within two closely related gammaproteobacterial clades, Ant4D3 and Arctic96B-16, were active in using the amino acid mixture, protein, and NAG.

Growth activity of gammaproteobacterial subgroups in waters off the west Antarctic Peninsula in summer and fall.

Characterizing both growth and abundance is important in understanding the role of bacterial communities in biogeochemical cycling of global oceans. However, these two quantities are seldom measured together for specific bacterial clades. Our goal was to examine growth and abundance of three gammaproteobacterial subgroups, including SAR86, at the single-cell level by microautoradiography combined with fluorescence in situ hybridization (FISH) in coastal waters of the west Antarctic Peninsula region during two austral summers and one austral fall.

Summer community structure of aerobic anoxygenic phototrophic bacteria in the western Arctic Ocean.

Aerobic anoxygenic phototrophic (AAP) bacteria are found in a range of aquatic and terrestrial environments, potentially playing unique roles in biogeochemical cycles. Although known to occur in the Arctic Ocean, their ecology and the factors that govern their community structure and distribution in this extreme environment are poorly understood. Here, we examined summer AAP abundance and diversity in the North East Pacific and the Arctic Ocean with emphasis on the southern Beaufort Sea.

Arsenite modifies structure of soil microbial communities and arsenite oxidization potential.

The influence of arsenite [As(III)] on natural microbial communities and the capacity of exposed communities to oxidize As(III) has not been well explored. In this study, we conducted soil column experiments with a natural microbial community exposed to different carbon conditions and a continuous flow of As(III). We measured the oxidation rates of As(III) to As(V), and the composition of the bacterial community was monitored by 454 pyrosequencing of 16S rRNA genes.

Assimilation of marine extracellular polymeric substances by deep-sea prokaryotes in the NW Mediterranean Sea.

This study examined total uptake of extracellular polymeric substances (EPS) and glucose and the percentage of prokaryotic cells (Bacteria, Crenarchaea and Euryarchaea) consuming these compounds in the major water masses at the DYFAMED site (NW Mediterranean Sea). The potential assimilation rates of EPS at 10 m depth were higher but on the same order of magnitude as those at 2000 m depth (from 43.4 to 29.

Abundance, diversity, and activity of ammonia-oxidizing prokaryotes in the coastal Arctic ocean in summer and winter.

Ammonia oxidation, the first step in nitrification, is performed by certain Beta- and Gammaproteobacteria and Crenarchaea to generate metabolic energy. Ammonia monooxygenase (amoA) genes from both Bacteria and Crenarchaea have been found in a variety of marine ecosystems, but the relative importance of Bacteria versus Crenarchaea in ammonia oxidation is unresolved, and seasonal comparisons are rare. In this study, we compared the abundance of betaproteobacterial and crenarchaeal amoA genes in the coastal Arctic Ocean during summer and winter over 2 years.

Summer distribution and diversity of aerobic anoxygenic phototrophic bacteria in the Mediterranean Sea in relation to environmental variables.

Aerobic anoxygenic phototrophic bacteria (AAP) represent an important fraction of bacterioplankton assemblages in various oceanic regimes. Although their abundance and distribution have been explored recently in diverse oceanic regions, the environmental factors controlling the population structure and diversity of these photoheterotrophic bacteria remain poorly understood. Here, we investigate the horizontal and vertical distributions and the genetic diversity of AAP populations collected in late summer throughout the Mediterranean Sea using pufM-temporal temperature gel gradient electrophoresis (TTGE) and clone library analyses.

Primary production in a subtropical stratified coastal lagoon--contribution of anoxygenic phototrophic bacteria.

Anaerobic anoxygenic phototrophic bacteria can be found in the suboxic waters of shallow stratified coastal systems, and may play important roles in the total primary production of subtropical stratified coastal lagoons. We investigated the spatiotemporal variability of light CO(2) fixation and net oxygen production in the stratified Conceição Lagoon (Brazil) in summer and fall of 2007, as well as the contribution of bacteriochlorophyll a (BChl a)-containing bacteria to photosynthetically driven electron transfer. Both chlorophyll a (Chl a) and BChl a varied in space, while only BChl a varied in time (three-fold increase from summer to fall).

Bacteriochlorophyll and community structure of aerobic anoxygenic phototrophic bacteria in a particle-rich estuary.

Photoheterotrophic microbes use organic substrates and light energy to satisfy their demand for carbon and energy and seem to be well adapted to eutrophic estuarine and oligotrophic oceanic environments. One type of photoheterotroph, aerobic anoxygenic phototrophic (AAP) bacteria, is especially abundant in particle-rich, turbid estuaries. To explore questions regarding the controls of these photoheterotrophic bacteria, we examined their abundance by epifluorescence microscopy, concentrations of the light-harvesting pigment, bacteriochlorophyll a (BChl a) and the diversity of pufM and 16S ribosomal RNA (rRNA) genes in the Chesapeake Bay.

The structure of bacterial communities in the western Arctic Ocean as revealed by pyrosequencing of 16S rRNA genes.

Bacterial communities in the surface layer of the oceans consist of a few abundant phylotypes and many rare ones, most with unknown ecological functions and unclear roles in biogeochemical processes. To test hypotheses about relationships between abundant and rare phylotypes, we examined bacterial communities in the western Arctic Ocean using pyrosequence data of the V6 region of the 16S rRNA gene. Samples were collected from various locations in the Chukchi Sea, the Beaufort Sea and Franklin Bay in summer and winter.

The GAAS metagenomic tool and its estimations of viral and microbial average genome size in four major biomes.

Metagenomic studies characterize both the composition and diversity of uncultured viral and microbial communities. BLAST-based comparisons have typically been used for such analyses; however, sampling biases, high percentages of unknown sequences, and the use of arbitrary thresholds to find significant similarities can decrease the accuracy and validity of estimates. Here, we present Genome relative Abundance and Average Size (GAAS), a complete software package that provides improved estimates of community composition and average genome length for metagenomes in both textual and graphical formats.

Light-dependent growth and proteorhodopsin expression by Flavobacteria and SAR11 in experiments with Delaware coastal waters.

Proteorhodopsin (PR)-containing bacteria are hypothesized to use both light and organic compounds as energy sources. Recent studies have found that PR is common in marine microorganisms, but the impact of light on the growth of PR-containing organisms and PR transcription in the environment remains unclear. We examined the diversity of PR genes and transcripts by PCR amplification and sequencing in Delaware coastal waters.

Photoheterotrophic microbes in the Arctic Ocean in summer and winter.

Photoheterotrophic microbes, which are capable of utilizing dissolved organic materials and harvesting light energy, include coccoid cyanobacteria (Synechococcus and Prochlorococcus), aerobic anoxygenic phototrophic (AAP) bacteria, and proteorhodopsin (PR)-containing bacteria. Our knowledge of photoheterotrophic microbes is largely incomplete, especially for high-latitude waters such as the Arctic Ocean, where photoheterotrophs may have special ecological relationships and distinct biogeochemical impacts due to extremes in day length and seasonal ice cover. These microbes were examined by epifluorescence microscopy, flow cytometry, and quantitative PCR (QPCR) assays for PR and a gene diagnostic of AAP bacteria (pufM).

Geographic and phylogenetic variation in bacterial biovolume as revealed by protein and nucleic acid staining.

Biovolume is an important characteristic of cells that shapes the contribution of microbes to total biomass and biogeochemical cycling. Most studies of bacterial cell volumes use DAPI (4',6'-diamidino-2-phenylindole), which stains nucleic acids and therefore only a portion of the cell. We used SYPRO Ruby protein stain combined with fluorescence in situ hybridization to examine biovolumes of bacteria in the total community, as well in phylogenetic subgroups.

Abundant proteorhodopsin genes in the North Atlantic Ocean.

Proteorhodopsin (PR) is a light-driven proton pump that has been found in a variety of marine bacteria, including Pelagibacter ubique, a member of the ubiquitous SAR11 clade. The goals of this study were to explore the diversity of PR genes and to estimate their abundance in the North Atlantic Ocean using quantitative polymerase chain reaction (QPCR). We found that PR genes in the western portion of the Sargasso Sea could be grouped into 27 clusters, but five clades had the most sequences.

Diversity and abundance of glycosyl hydrolase family 5 in the North Atlantic Ocean.

The diversity and abundance of glycosyl hydrolase family 5 (GH5) were studied in the North Atlantic Ocean. This family was chosen because of the large number of available sequences from cultured bacteria, the variety of substrates it targets, and the high number of similar sequences in the Sargasso Sea environmental genome database. Three clone libraries of a GH5 subcluster were constructed from the Mid-Atlantic Bight and the eastern and western North Atlantic Ocean.

Light-stimulated bacterial production and amino acid assimilation by cyanobacteria and other microbes in the North Atlantic ocean.

We examined the contribution of photoheterotrophic microbes--those capable of light-mediated assimilation of organic compounds--to bacterial production and amino acid assimilation along a transect from Florida to Iceland from 28 May to 9 July 2005. Bacterial production (leucine incorporation at a 20 nM final concentration) was on average 30% higher in light than in dark-incubated samples, but the effect varied greatly (3% to 60%). To further characterize this light effect, we examined the abundance of potential photoheterotrophs and measured their contribution to bacterial production and amino acid assimilation (0.

High abundances of aerobic anoxygenic photosynthetic bacteria in the South Pacific Ocean.

Little is known about the abundance, distribution, and ecology of aerobic anoxygenic phototrophic (AAP) bacteria, particularly in oligotrophic environments, which represent 60% of the ocean. We investigated the abundance of AAP bacteria across the South Pacific Ocean, including the center of the gyre, the most oligotrophic water body of the world ocean. AAP bacteria, Prochlorococcus, and total prokaryotic abundances, as well as bacteriochlorophyll a (BChl a) and divinyl-chlorophyll a concentrations, were measured at several depths in the photic zone along a gradient of oligotrophic conditions.

Bacterial community structure of biofilms on artificial surfaces in an estuary.

This study examined bacterial community structure of biofilms on stainless steel and polycarbonate in seawater from the Delaware Bay. Free-living bacteria in the surrounding seawater were compared to the attached bacteria during the first few weeks of biofilm growth. Surfaces exposed to seawater were analyzed by using 16S rDNA libraries, fluorescence in situ hybridization (FISH), and denaturing gradient gel electrophoresis (DGGE).