Secondary metabolites play essential roles in ecological interactions and nutrient acquisition, and are of interest for their potential uses in medicine and biotechnology. Genome mining for ...biosynthetic gene clusters (BGCs) can be used for the discovery of new compounds. Here, we use metagenomics and metatranscriptomics to analyze BGCs in free-living and particle-associated microbial communities through the stratified water column of the Cariaco Basin, Venezuela. We recovered 565 bacterial and archaeal metagenome-assembled genomes (MAGs) and identified 1154 diverse BGCs. We show that differences in water redox potential and microbial lifestyle (particle-associated vs. free-living) are associated with variations in the predicted composition and production of secondary metabolites. Our results indicate that microbes, including understudied clades such as Planctomycetota, potentially produce a wide range of secondary metabolites in these anoxic/euxinic waters.
Geochemical cycles of all nonconservative elements are mediated by microorganisms over nanometer spatial scales. The pelagic seascape is known to possess microstructure imposed by heterogeneous ...distributions of particles, polymeric gels, biologically important chemicals, and microbes. While indispensable, most traditional oceanographic observational approaches overlook this heterogeneity and ignore subtleties, such as activity hot spots, symbioses, niche partitioning, and intrapopulation phenotypic variations, that can provide a deeper mechanistic understanding of planktonic ecosystem function. As part of the movement toward cultivation-independent tools in microbial oceanography, techniques to examine the ecophysiology of individual populations and their role in chemical transformations at spatial scales relevant to microorganisms have been developed. This review presents technologies that enable geochemical and microbiological interrogations at spatial scales ranging from 0.02 to a few hundred micrometers, particularly focusing on atomic force microscopy, nanoscale secondary ion mass spectrometry, and confocal Raman microspectroscopy and introducing promising approaches for future applications in marine sciences.
Comprehensive methodologies for monitoring microplastics (MPs) in the ocean are critical for accurately assessing abundances across a broad size spectrum, and to document distributions, sources, ...sinks, temporal trends, and exposure risks for organisms. Discrete 0.5-L water samples from the northeastern-coast of Venezuela (NECV), Pacific-Arctic Ocean (PAO), and Gulf Stream Current (GSC) were analyzed by Raman microspectroscopy to detect MPs not captured by net-tow surveys. Equivalent spherical diameters (ESD) of most MPs were <5 μm, accounting for 68, 83, 86 % of total inventories in NECV, GSC, PAO samples. We did not observe a single MP particle >53 μm ESD. Abundances of MPs in the 0.5–200 μm size fraction were 5–6 orders of magnitude higher than previous surveys that were almost exclusively based on net tow collections of MPs > 300 μm ESD. Abundances of MPs in NECV samples were ~10-fold higher than those from PAO and GSC. The most abundant polymers were polypropylene (PP), polystyrene (PS) and polyethylene terephthalate (PET), consistent with composition of plastic waste generated globally.
•Most abundant microplastics were between 1 and 14 μm equivalent spherical diameters.•60 % of microplastics were <5 μm and none were larger than 53 μm.•Microplastics <53 μm were 6 orders of magnitude higher than surveys based on net tow.•Microplastics were more abundant in Venezuelan waters relative to the other sites.•Specific polymer contributions match plastic waste composition reported in the ocean.
The suitability of stable isotope probing (SIP) and Raman microspectroscopy to measure growth rates of heterotrophic bacteria at the single-cell level was evaluated. Label assimilation into ...Escherichia coli biomass during growth on a complex
C-labeled carbon source was monitored in time course experiments.
C incorporation into various biomolecules was measured by spectral "red shifts" of Raman-scattered emissions. The
C- and
C-isotopologues of the amino acid phenylalanine (Phe) proved to be quantitatively accurate reporter molecules of cellular isotopic fractional abundances (
). Values of
determined by Raman microspectroscopy and independently by isotope ratio mass spectrometry (IRMS) over a range of isotopic enrichments were statistically indistinguishable. Progressive labeling of Phe in E. coli cells among a range of
C/
C organic substrate admixtures occurred predictably through time. The relative isotopologue abundances of Phe determined by Raman spectral analysis enabled the accurate calculation of bacterial growth rates as confirmed independently by optical density (OD) measurements. The results demonstrate that combining SIP and Raman microspectroscopy can be a powerful tool for studying bacterial growth at the single-cell level on defined or complex organic
C carbon sources, even in mixed microbial assemblages.
Population growth dynamics and individual cell growth rates are the ultimate expressions of a microorganism's fitness under its environmental conditions, whether natural or engineered. Natural habitats and many industrial settings harbor complex microbial assemblages. Their heterogeneity in growth responses to existing and changing conditions is often difficult to grasp by standard methodologies. In this proof-of-concept study, we tested whether Raman microspectroscopy can reliably quantify the assimilation of isotopically labeled nutrients into E. coli cells and enable the determination of individual growth rates among heterotrophic bacteria. Raman-derived growth rate estimates were statistically indistinguishable from those derived by standard optical density measurements of the same cultures. Raman microspectroscopy can also be combined with methods for phylogenetic identification. We report the development of Raman-based techniques that enable researchers to directly link genetic identity to functional traits and rate measurements of single cells within mixed microbial assemblages, currently a major technical challenge in microbiological research.
Acceptance and commitment therapy (ACT) is a developing approach for chronic pain. The current study was designed to pilot test a brief, widely inclusive, local access format of ACT in a UK primary ...care setting. Seventy-three participants (68.5% women) were randomized to either ACT or treatment as usual (TAU). Many of the participants were aged 65 years or older (27.6%), were diagnosed with fibromyalgia (30.2%) and depression (40.3%), and had longstanding pain (median = 10 years). Standard clinical outcome measures included disability, depression, physical functioning, emotional functioning, and rated improvement. Process measures included pain-related and general psychological acceptance. The recruitment target was met within 6 months, and 72.9% of those allocated to ACT completed treatment. Immediately post treatment, relative to TAU, participants in ACT demonstrated lower depression and higher ratings of overall improvement. At a 3-month follow-up, again relative to TAU, those in ACT demonstrated lower disability, less depression, and significantly higher pain acceptance; d = .58, .59, and .64, respectively. Analyses based on intention-to-treat and on treatment “completers,” perhaps predictably, revealed more sobering and more encouraging results, respectively. A larger trial of ACT delivered in primary care, in the format employed here, appears feasible with some recommended adjustments in the methods used here (Trial registration: ISRCTN49827391).
This article presents a pilot randomized controlled trial of ACT for chronic pain in a primary care setting in the United Kingdom. Both positive clinical outcomes and ways to improve future trials are reported.
Summary
Genetic markers and geochemical assays of microbial nitrogen cycling processes, including autotrophic and heterotrophic denitrification, anammox, ammonia oxidation, and nitrite oxidation, ...were examined across the oxycline, suboxic, and anoxic zones of the Cariaco Basin, Venezuela. Ammonia and nitrite oxidation genes were expressed through the entire gradient. Transcripts associated with autotrophic and heterotrophic denitrifiers were mostly confined to the suboxic zone and below but were also present in particles in the oxycline. Anammox genes and transcripts were detected over a narrow depth range near the bottom of the suboxic zone and coincided with secondary NO2− maxima and available NH4+. Dissolved inorganic nitrogen (DIN) amendment incubations and comparisons between our sampling campaigns suggested that denitrifier activity may be closely coupled with NO3− availability. Expression of denitrification genes at depths of high rates of chemoautotrophic carbon fixation and phylogenetic analyses of nitrogen cycling genes and transcripts indicated a diverse array of denitrifiers, including chemoautotrophs capable of using NO3− to oxidize reduced sulfur species. Thus, results suggest that the Cariaco Basin nitrogen cycle is influenced by autotrophic carbon cycling in addition to organic matter oxidation and anammox.
Summary
Using the anoxic Cariaco Basin as a natural laboratory, particle association of bacterial and archaeal taxa was assessed by iTag sequencing and qPCR gene assays of samples spanning an ...oxic–anoxic–euxinic gradient. A total of 10%–12% of all bacterial and archaeal cells were found in the particle‐associated (PA) fraction, operationally defined as prokaryotes captured on 2.7 µm membranes. Both redox condition and size fraction segregated bacterial taxa. Archaeal taxa varied according to redox conditions, but were similar between size fractions. Taxa putatively associated with chemoautotrophic sulfur oxidation and nitrification dominated the free‐living (FL) fraction throughout the oxycline (< 1–120 µM O2) and upper anoxic layer. Bacteria in the oxycline's PA fraction included taxa known to be aerobic and anaerobic chemoorganotrophs. At shallow anoxic depths, PA taxa were primarily affiliated with anaerobic sulfate (
SO42–)‐reducing lineages. PA fractions in the most sulfidic samples were dominated by taxa affiliated with CH4 oxidizing, fermenting and
SO42– reducing lineages. Prevalence of particle‐associated
SO42–‐reducing taxa and abundant sulfur‐oxidizing taxa in both size fractions across the oxic–anoxic interface is consistent with the cryptic sulfur cycling concept. Bacterial assemblage diversity in the PA fraction always exceeded the FL fraction except in the most oxic samples, whereas Archaeal diversity was not consistently different between size fractions. Our results suggest that these particle‐associated and free‐living bacterial assemblages are functionally different and that the interplay between particle microhabitats and surrounding geochemical regimes is a strong selective force shaping microbial communities throughout the water column.
Plastic substrates introduced to the environment during the Anthropocene have introduced new pathways for microbial selection and dispersal. Some plastic‐colonising microorganisms have adapted ...phenotypes for plastic degradation (selection), while the spatial transport (dispersal) potential of plastic colonisers remains controlled by polymer‐specific density, hydrography and currents. Plastic‐degrading enzyme abundances have recently been correlated with concentrations of plastic debris in open ocean environments, making it critical to better understand colonisation of hydrocarbon degraders with plastic degradation potential in urbanised watersheds where plastic pollution often originates. We found that microbial colonisation by reputed hydrocarbon degraders on microplastics (MPs) correlated with a spatial contaminant gradient (New York City/Long Island waterways), polymer types, temporal scales, microbial domains and putative cell activity (DNA vs. RNA). Hydrocarbon‐degrading taxa enriched on polyethylene and polyvinyl chloride substrates relative to other polymers and were more commonly recovered in samples proximal to New York City. These differences in MP colonisation could indicate phenotypic adaptation processes resulting from increased exposure to urban plastic runoff as well as differences in carbon bioavailability across polymer types. Shifts in MP community potential across urban coastal contaminant gradients and polymer types improve our understanding of environmental plastic discharge impacts toward biogeochemical cycling across the global ocean.
Plastisphere communities varied significantly along a contaminant gradient within an urbanised watershed (New York City/Long Island waterways), across polymer types (polyethylene, polystyrene, polypropylene, polycarbonate, polyethylene terephthalate and polyvinyl chloride), temporal scales, microbial domains (bacteria, archaea and fungi) and putative cell activity (DNA vs. RNA).
Microbial diversity and distribution are topics of intensive research. In two companion papers in this issue, we describe the results of the Cariaco Microbial Observatory (Caribbean Sea, Venezuela). ...The Basin contains the largest body of marine anoxic water, and presents an opportunity to study protistan communities across biogeochemical gradients. In the first paper, we survey 18S ribosomal RNA (rRNA) gene sequence diversity using both Sanger- and pyrosequencing-based approaches, employing multiple PCR primers, and state-of-the-art statistical analyses to estimate microbial richness missed by the survey. Sampling the Basin at three stations, in two seasons, and at four depths with distinct biogeochemical regimes, we obtained the largest, and arguably the least biased collection of over 6000 nearly full-length protistan rRNA gene sequences from a given oceanographic regime to date, and over 80,000 pyrosequencing tags. These represent all major and many minor protistan taxa, at frequencies globally similar between the two sequence collections. This large data set provided, via the recently developed parametric modeling, the first statistically sound prediction of the total size of protistan richness in a large and varied environment, such as the Cariaco Basin: over 36,000 species, defined as almost full-length 18S rRNA gene sequence clusters sharing over 99% sequence homology. This richness is a small fraction of the grand total of known protists (over 100,000-500,000 species), suggesting a degree of protistan endemism.
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