Microbes and microplastics: Community shifts along an urban coastal contaminant gradient.

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.

Transport-limited reactions in microbial systems.

Predicting microbial metabolic rates and emergent biogeochemical fluxes remains challenging due to the many unknown population dynamical, physiological and reaction-kinetic parameters and uncertainties in species composition. Here, we show that the need for these parameters can be eliminated when population dynamics and reaction kinetics operate at much shorter time scales than physical mixing processes. Such scenarios are widespread in poorly mixed water columns and sediments. In this 'fast-reaction-transport' (FRT) limit, all that is required for predictions are chemical boundary conditions, the physical mixing processes and reaction stoichiometries, while no knowledge of species composition, physiology or population/reaction kinetic parameters is needed. Using time-series data spanning years 2001-2014 and depths 180-900 m across the permanently anoxic Cariaco Basin, we demonstrate that the FRT approach can accurately predict the dynamics of major electron donors and acceptors (Pearson r ≥ 0.9 in all cases). Hence, many microbial processes in this system are largely transport limited and thus predictable regardless of species composition, population dynamics and kinetics. Our approach enables predictions for many systems in which microbial community dynamics and kinetics are unknown. Our findings also reveal a mechanism for the frequently observed decoupling between function and taxonomy in microbial systems.

Circumventing kinetics in biogeochemical modeling.

Microbial metabolism drives biogeochemical fluxes in virtually every ecosystem. Modeling these fluxes is challenged by the incredible diversity of microorganisms, whose kinetic parameters are largely unknown. In poorly mixed systems, such as stagnant water columns or sediments, however, long-term bulk microbial metabolism may become limited by physical transport rates of substrates across space. Here we mathematically show that under these conditions, biogeochemical fluxes are largely predictable based on the system's transport properties, chemical boundary conditions, and the stoichiometry of metabolic pathways, regardless of the precise kinetics of the resident microorganisms. We formalize these considerations into a predictive modeling framework and demonstrate its use for the Cariaco Basin subeuphotic zone, one of the largest anoxic marine basins worldwide. Using chemical concentration data solely from the upper boundary (depth 180 m) and lower boundary (depth 900 m), but without a priori knowledge of metabolite fluxes, chemical depth profiles, kinetic parameters, or microbial species composition, we predict the concentrations and vertical fluxes of biologically important substances, including oxygen, nitrate, hydrogen sulfide, and ammonium, across the entire considered depth range (180-900 m). Our predictions largely agree with concentration measurements over a period of 14 years ([Formula: see text] = 0.78-0.92) and become particularly accurate during a period where the system was near biogeochemical steady state (years 2007-2009, [Formula: see text] = 0.86-0.95). Our work enables geobiological predictions for a large class of ecosystems without knowledge of kinetic parameters or geochemical depth profiles. Conceptually, our work provides a possible explanation for the decoupling between microbial species composition and bulk metabolic function, observed in various ecosystems.

Diverse nitrogen cycling pathways across a marine oxygen gradient indicate nitrogen loss coupled to chemoautotrophic activity.

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.

Using Stable Isotope Probing and Raman Microspectroscopy To Measure Growth Rates of Heterotrophic Bacteria.

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 13C-labeled carbon source was monitored in time course experiments. 13C incorporation into various biomolecules was measured by spectral "red shifts" of Raman-scattered emissions. The 13C- and 12C-isotopologues of the amino acid phenylalanine (Phe) proved to be quantitatively accurate reporter molecules of cellular isotopic fractional abundances (fcell). Values of fcell 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 13C/12C 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 13C carbon sources, even in mixed microbial assemblages. IMPORTANCE 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.

Free-living chemoautotrophic and particle-attached heterotrophic prokaryotes dominate microbial assemblages along a pelagic redox gradient.

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.

Discovery of a resting stage in the harmful, brown-tide-causing pelagophyte, Aureoumbra lagunensis: a mechanism potentially facilitating recurrent blooms and geographic expansion.

To date, the life stages of pelagophytes have been poorly described. This study describes the ability of Aureoumbra lagunensis to enter a resting stage in response to environmental stressors including high temperature, nutrient depletion, and darkness as well as their ability to revert from resting cells back to vegetative cells after exposure to optimal light, temperature, and nutrient conditions. Resting cells became round in shape and larger in size, filled with red accumulation bodies, had smaller and fewer plastids, more vacuolar space, contained lower concentrations of chl a and RNA, displayed reduced photosynthetic efficiency, and lower respiration rates relative to vegetative cells. Analysis of vegetative and resting cells using Raman microspectrometry indicated resting cells were enriched in sterols within red accumulation bodies and were depleted in pigments relative to vegetative cells. Upon reverting to vegetative cells, cells increased their chl a content, photosynthetic efficiency, respiration rate, and growth rate and lost accumulation bodies as they became smaller. The time required for resting cells to resume vegetative growth was proportional to both the duration and temperature of dark storage, possibly due to higher metabolic demands on stored energy (sterols) reserves during longer period of storage and/or storage at higher temperature (20°C vs. 10°C). Resting cells kept in the dark at 10°C for 7 months readily reverted back to vegetative cells when transferred to optimal conditions. Thus, the ability of Aureoumbra to form a resting stage likely enables them to form annual blooms within subtropic ecosystems, resist temperature extremes, and may facilitate geographic expansion via anthropogenic transport.

Ecosystem responses in the southern Caribbean Sea to global climate change.

Over the last few decades, rising greenhouse gas emissions have promoted poleward expansion of the large-scale atmospheric Hadley circulation that dominates the Tropics, thereby affecting behavior of the Intertropical Convergence Zone (ITCZ) and North Atlantic Oscillation (NAO). Expression of these changes in tropical marine ecosystems is poorly understood because of sparse observational datasets. We link contemporary ecological changes in the southern Caribbean Sea to global climate change indices. Monthly observations from the CARIACO Ocean Time-Series between 1996 and 2010 document significant decadal scale trends, including a net sea surface temperature (SST) rise of ∼1.0 ± 0.14 °C (±SE), intensified stratification, reduced delivery of upwelled nutrients to surface waters, and diminished phytoplankton bloom intensities evident as overall declines in chlorophyll a concentrations (ΔChla = -2.8 ± 0.5%⋅y(-1)) and net primary production (ΔNPP = -1.5 ± 0.3%⋅y(-1)). Additionally, phytoplankton taxon dominance shifted from diatoms, dinoflagellates, and coccolithophorids to smaller taxa after 2004, whereas mesozooplankton biomass increased and commercial landings of planktivorous sardines collapsed. Collectively, our results reveal an ecological state change in this planktonic system. The weakening trend in Trade Winds (-1.9 ± 0.3%⋅y(-1)) and dependent local variables are largely explained by trends in two climatic indices, namely the northward migration of the Azores High pressure center (descending branch of Hadley cell) by 1.12 ± 0.42°N latitude and the northeasterly progression of the ITCZ Atlantic centroid (ascending branch of Hadley cell), the March position of which shifted by about 800 km between 1996 and 2009.

From the Caribbean to the Arctic, the most abundant microplastic particles in the ocean have escaped detection.

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.

Plasmid-Borne Biosynthetic Gene Clusters within a Permanently Stratified Marine Water Column.

Plasmids are mobile genetic elements known to carry secondary metabolic genes that affect the fitness and survival of microbes in the environment. Well-studied cases of plasmid-encoded secondary metabolic genes in marine habitats include toxin/antitoxin and antibiotic biosynthesis/resistance genes. Here, we examine metagenome-assembled genomes (MAGs) from the permanently-stratified water column of the Cariaco Basin for integrated plasmids that encode biosynthetic gene clusters of secondary metabolites (smBGCs). We identify 16 plasmid-borne smBGCs in MAGs associated primarily with Planctomycetota and Pseudomonadota that encode terpene-synthesizing genes, and genes for production of ribosomal and non-ribosomal peptides. These identified genes encode for secondary metabolites that are mainly antimicrobial agents, and hence, their uptake via plasmids may increase the competitive advantage of those host taxa that acquire them. The ecological and evolutionary significance of smBGCs carried by prokaryotes in oxygen-depleted water columns is yet to be fully elucidated.

Preserved particulate organic carbon is likely derived from the subsurface sulfidic photic zone of the Proterozoic Ocean: evidence from a modern, oxygen-deficient lake.

Biological processes in the Proterozoic Ocean are often inferred from modern oxygen-deficient environments (MODEs) or from stable isotopes in preserved sediment. To date, few MODE studies have simultaneously quantified carbon fixation genes and attendant stable isotopic signatures. Consequently, how carbon isotope patterns reflect these pathways has not been thoroughly vetted. Addressing this, we profiled planktonic productivity and quantified carbon fixation pathway genes and associated organic carbon isotope values (δ13 CPOC ) of size-fractionated (0.2-2.7 and >2.7 µm) particulate matter from meromictic Fayetteville Green Lake, NY, USA. The high-O2 Calvin-Benson-Bassham (CBB) gene (cbbL) was most abundant in the <2.7 µm size fraction in shallow oxic and deep hypoxic waters, corresponding with cyanobacterial and eukaryote algal populations. The low-O2 CBB gene (cbbM) was most abundant near the lower oxycline boundary in the larger size fraction, coincident with purple sulfur bacteria populations. The reverse citric acid cycle gene (aclB) was equally abundant in both size fractions in the deepest photic zone, coinciding with green sulfur bacteria populations. Methane coenzyme reductase A (mcrA), of anaerobic methane cyclers, was most abundant at the lower oxycline boundary in both size fractions, coinciding with Methanoregula populations. δ13 CPOC values overlapped with the high-O2 CBB fixation range except for two negative excursions near the lower oxycline boundary, likely reflecting assimilation of isotopically-depleted groundwater-derived carbon by autotrophs and sulfate-reducers. Throughout aphotic waters, δ13 CPOC values of the large size fraction became 13 C-enriched, likely reflecting abundant purple sulfur bacterial aggregates. Eukaryote algae- or cyanobacteria-like isotopic signatures corresponded with increases in cbbL, cbbM, and aclB, and enrichment of exopolymer-rich prokaryotic photoautotrophs aggregates. Results suggest that δ13 CPOC values of preserved sediments from areas of the Proterozoic Ocean with sulfidic photic zones may reflect a mixture of alternate carbon-fixing populations exported from the deep photic zone, challenging the paradigm that sedimentary stable carbon isotope values predominantly reflect oxygenic photosynthesis from surface waters.

Diverse secondary metabolites are expressed in particle-associated and free-living microorganisms of the permanently anoxic Cariaco Basin.

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.

Class Cariacotrichea, a novel ciliate taxon from the anoxic Cariaco Basin, Venezuela.

The majority of environmental micro-organisms identified with the rRNA approach have never been visualized. Thus, their reliable classification and taxonomic assignment is often difficult or even impossible. In our preliminary 18S rRNA gene sequencing work from the world's largest anoxic marine environment, the Cariaco Basin (Caribbean Sea, Venezuela), we detected a ciliate clade, designated previously as CAR_H [Stoeck, S., Taylor, G. T. & Epstein, S. S. (2003). Appl Environ Microbiol 63, 5656-5663]. Here, we combine the traditional rRNA detection method of fluorescent in situ hybridization (FISH) with scanning electron microscopy (SEM) and confirm the phylogenetic separation of the CAR_H sequences from all other ciliate classes by showing an outstanding morphological feature of this group: a unique, archway-shaped kinety surrounding the oral apparatus and extending to the posterior body end in CAR_H cells. Based on this specific feature and the molecular phylogenies, we propose a novel ciliate class, Cariacotrichea nov. cl.

One Cell at a Time: Advances in Single-Cell Methods and Instrumentation for Discovery in Aquatic Microbiology.

Studying microbes from a single-cell perspective has become a major theme and interest within the field of aquatic microbiology. One emerging trend is the unfailing observation of heterogeneity in activity levels within microbial populations. Wherever researchers have looked, intra-population variability in biochemical composition, growth rates, and responses to varying environmental conditions has been evident and probably reflect coexisting genetically distinct strains of the same species. Such observations of heterogeneity require a shift away from bulk analytical approaches and development of new methods or adaptation of existing techniques, many of which were first pioneered in other, unrelated fields, e.g., material, physical, and biomedical sciences. Many co-opted approaches were initially optimized using model organisms. In a field with so few cultivable models, method development has been challenging but has also contributed tremendous insights, breakthroughs, and stimulated curiosity. In this perspective, we present a subset of methods that have been effectively applied to study aquatic microbes at the single-cell level. Opportunities and challenges for innovation are also discussed. We suggest future directions for aquatic microbiological research that will benefit from open access to sophisticated instruments and highly interdisciplinary collaborations.

Eukaryotic Parasites Are Integral to a Productive Microbial Food Web in Oxygen-Depleted Waters.

Oxygen-depleted water columns (ODWCs) host a diverse community of eukaryotic protists that change dramatically in composition over the oxic-anoxic gradient. In the permanently anoxic Cariaco Basin, peaks in eukaryotic diversity occurred in layers where dark microbial activity (chemoautotrophy and heterotrophy) were highest, suggesting a link between prokaryotic activity and trophic associations with protists. Using 18S rRNA gene sequencing, parasites and especially the obligate parasitic clade, Syndiniales, appear to be particularly abundant, suggesting parasitism is an important, but overlooked interaction in ODWC food webs. Syndiniales were also associated with certain prokaryotic groups that are often found in ODWCs, including Marinimicrobia and Marine Group II archaea, evocative of feedbacks between parasitic infection events, release of organic matter, and prokaryotic assimilative activity. In a network analysis that included all three domains of life, bacterial and archaeal taxa were putative bottleneck and hub species, while a large proportion of edges were connected to eukaryotic nodes. Inclusion of parasites resulted in a more complex network with longer path lengths between members. Together, these results suggest that protists, and especially protistan parasites, play an important role in maintaining microbial food web complexity, particularly in ODWCs, where protist diversity and microbial productivity are high, but energy resources are limited relative to euphotic waters.

Raman Microspectroscopy Goes Viral: Infection Dynamics in the Cosmopolitan Microalga, Emiliania huxleyi.

Emiliania huxleyi is a cosmopolitan member of the marine phytoplankton. This species' capacities for carbon sequestration and sulfur mobilization make it a key player in oceanic biogeochemical cycles that influence climate on a planetary scale. Seasonal E. huxleyi blooms are abruptly terminated by viral epidemics caused by a clade of large DNA viruses collectively known as coccolithoviruses (EhVs). EhVs thereby mediate a significant part of material and energy fluxes associated with E. huxleyi population dynamics. In this study, we use spontaneous Raman microspectroscopy to perform label-free and non-invasive measurements of the macromolecular composition of individual virions and E. huxleyi host cells. Our novel autofluorescence suppression protocol enabled spectroscopic visualization of evolving macromolecular redistributions in individual E. huxleyi cells at different stages of EhV infection. Material transfer from E. huxleyi hosts to single EhV-163 virions was confirmed by combining stable isotope probing (SIP) experiments with Raman microspectroscopy. Inheritance of the host cells' 13C-enriched isotopic signature was quantified based on red shifts of Raman peaks characteristic of phenylalanine's phenyl ring. Two-dimensional Raman mapping of EhV-infected E. huxleyi cells revealed that the compact region producing an intense Raman DNA signal (i.e., the nucleus) in healthy E. huxleyi cells becomes diffuse during the first hours of infection. Raman DNA emissions integrated throughout individual cells decreased during the infection cycle. Our observations are consistent with EhV-163 degrading the host's nuclear DNA, scavenging released nucleotides for its own genome replication, and shedding newly-produced virions prior to host lysis via budding.

Viral elements and their potential influence on microbial processes along the permanently stratified Cariaco Basin redoxcline.

Little is known about viruses in oxygen-deficient water columns (ODWCs). In surface ocean waters, viruses are known to act as gene vectors among susceptible hosts. Some of these genes may have metabolic functions and are thus termed auxiliary metabolic genes (AMGs). AMGs introduced to new hosts by viruses can enhance viral replication and/or potentially affect biogeochemical cycles by modulating key microbial pathways. Here we identify 748 viral populations that cluster into 94 genera along a vertical geochemical gradient in the Cariaco Basin, a permanently stratified and euxinic ocean basin. The viral communities in this ODWC appear to be relatively novel as 80 of these viral genera contained no reference viral sequences, likely due to the isolation and unique features of this system. We identify viral elements that encode AMGs implicated in distinctive processes, such as sulfur cycling, acetate fermentation, signal transduction, [Fe-S] formation, and N-glycosylation. These AMG-encoding viruses include two putative Mu-like viruses, and viral-like regions that may constitute degraded prophages that have been modified by transposable elements. Our results provide an insight into the ecological and biogeochemical impact of viruses oxygen-depleted and euxinic habitats.

Diagnostic Assessment of Lower Urinary Tract Symptoms in Men Considering Prostate Surgery: A Noninferiority Randomised Controlled Trial of Urodynamics in 26 Hospitals.

BACKGROUND: Prostate surgery can improve lower urinary tract symptoms (LUTS) by relieving bladder outlet obstruction (BOO). However, surgery is less effective without BOO, or if detrusor underactivity is present. Urodynamics (UDS) can identify BOO and measure detrusor activity, but evidence in clinical practice is lacking. OBJECTIVE: Urodynamics for Prostate Surgery Trial: Randomised Evaluation of Assessment Methods (UPSTREAM) aimed to evaluate whether a care pathway including UDS would reduce surgery without increasing urinary symptoms. DESIGN, SETTING, AND PARTICIPANTS: UPSTREAM is a pragmatic, noninferiority, randomised controlled trial in men with bothersome LUTS, in whom surgery was an option, in 26 hospitals in England (ISRCTN56164274). INTERVENTION: Participants were randomised (1:1) to routine care (RC) diagnostic tests, or RC plus UDS. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: The primary outcome was the International Prostate Symptom Score (IPSS; patient-reported outcome scale from 0 to 35 points) 18 mo after randomisation, with a noninferiority margin of 1 point. Urological surgery rates were a key secondary outcome. The primary outcome was compared between the arms using linear regression, analysed on an intention-to-treat basis. RESULTS AND LIMITATIONS: Between October 2014 and December 2016, 820 men (median age 68 yr) were randomised (393 and 427 in the RC and UDS arms, respectively). The UDS arm showed noninferiority of the mean IPSSs (UDS 12.6; RC 13.1; adjusted difference at 18 mo -0.33 [95% confidence interval {CI} -1.47, +0.80]). In the UDS arm, 153/408 (38%) received surgery compared with 138/384 (36%) in the RC arm (adjusted odds ratio 1.05; 95% CI 0.77, 1.43). A total of 428 adverse events (UDS 234; RC 194) were recorded, with related events similar in both arms and 11 unrelated deaths. CONCLUSIONS: In this population, the UDS randomised group was noninferior to RC for the IPSS but did not reduce surgical rates. This study shows that routine use of UDS in the evaluation of uncomplicated LUTS has a limited role and should be used selectively. PATIENT SUMMARY: For men with uncomplicated lower urinary tract symptoms, symptom improvements after treatment and the number of operations performed are similar, irrespective of whether or not urodynamic tests are conducted in addition to routine tests. Accordingly, routine use of urodynamics has a limited role in this population group.

Urodynamics tests for the diagnosis and management of bladder outlet obstruction in men: the UPSTREAM non-inferiority RCT.

BACKGROUND: Lower urinary tract symptoms (LUTS) in men may indicate bladder outlet obstruction (BOO) or weakness, known as detrusor underactivity (DU). Severe bothersome LUTS are a common indication for surgery. The diagnostic tests may include urodynamics (UDS) to confirm whether BOO or DU is the cause, potentially reducing the number of people receiving (inappropriate) surgery. OBJECTIVES: The primary objective was to determine whether a care pathway including UDS is no worse for symptom outcome than one in which it is omitted, at 18 months after randomisation. Rates of surgery was the key secondary outcome. DESIGN: This was a pragmatic, multicentre, two-arm (unblinded) randomised controlled trial, incorporating a health economic analysis and qualitative research. SETTING: Urology departments of 26 NHS hospitals in England. PARTICIPANTS: Men (aged ≥ 18 years) seeking further treatment, potentially including surgery, for bothersome LUTS. Exclusion criteria were as follows: unable to pass urine without a catheter, having a relevant neurological disease, currently undergoing treatment for prostate or bladder cancer, previously had prostate surgery, not medically fit for surgery and/or unwilling to be randomised. INTERVENTIONS: Men were randomised to a care pathway based on non-invasive routine tests (control) or routine care plus invasive UDS (intervention arm). MAIN OUTCOME MEASURES: The primary outcome was International Prostate Symptom Score (IPSS) at 18 months after randomisation and the key secondary outcome was rates of surgery. Additional secondary outcomes included adverse events (AEs), quality of life, urinary and sexual symptoms, UDS satisfaction, maximum urinary flow rate and cost-effectiveness. RESULTS: A total of 820 men were randomised (UDS, 427; routine care, 393). Sixty-seven men withdrew before 18 months and 11 died (unrelated to trial procedures). UDS was non-inferior to routine care for IPSS 18 months after randomisation, with a confidence interval (CI) within the margin of 1 point (-0.33, 95% CI -1.47 to 0.80). A lower surgery rate in the UDS arm was not found (38% and 36% for UDS and routine care, respectively), with overall rates lower than expected. AEs were similar between the arms at 43-44%. There were more cases of acute urinary retention in the routine care arm. Patient-reported outcomes for LUTS improved in both arms and satisfaction with UDS was high in men who received it. UDS was more expensive than routine care. From a secondary care perspective, UDS cost an additional £216 over an 18-month time horizon. Quality-adjusted life-years (QALYs) were similar, with a QALY difference of 0.006 in favour of UDS over 18 months. It was established that UDS was acceptable to patients, and valued by both patients and clinicians for its perceived additional insight into the cause and probable best treatment of LUTS. LIMITATIONS: The trial met its predefined recruitment target, but surgery rates were lower than anticipated. CONCLUSIONS: Inclusion of UDS in the diagnostic tests results in a symptom outcome that is non-inferior to a routine care pathway, but does not affect surgical rates for treating BOO. Results do not support the routine use of UDS in men undergoing investigation of LUTS. FUTURE WORK: Focus should be placed on indications for selective utilisation of UDS in individual cases and long-term outcomes of diagnosis and therapy. TRIAL REGISTRATION: Current Controlled Trials ISRCTN56164274. FUNDING: This project was funded by the National Institute for Health Research (NIHR) Health Technology Assessment programme and will be published in full in Health Technology Assessment; Vol. 24, No. 42. See the NIHR Journals Library website for further project information.

After hospital referral, men with bothersome lower urinary tract symptoms (LUTS) are assessed with standard tests. These include measurement of urine flow rate, bladder diaries and questionnaires, including the International Prostate Symptom Score (IPSS). UPSTREAM (Urodynamics for Prostate Surgery Trial; Randomised Evaluation of Assessment Methods) researched whether or not including an extra test, urodynamics (UDS), helps when considering treatment options. UDS is a more invasive test and measures pressure in the bladder to check whether or not the prostate is causing obstruction. It was presumed that, if there is no obstruction, surgery would not be offered, so that using UDS would reduce the number of prostate operations. Each man participating (820 in total) was assessed with the standard tests. Around half of them had no extra tests (the 'routine care' arm of the trial); the rest had the UDS tests (the 'UDS' arm). Men then went on to have treatment, which they chose having discussed their test results with a urologist. IPSS and other symptom scores were examined for each man 18 months after joining the trial. At 18 months, surgery outcomes were known for 792 men and IPSS was known for 669 men. We investigated if the two trial arms showed similar changes in the IPSS and if there were fewer operations done in the UDS arm. We identified similar reductions in the IPSS in both arms. However, UDS tests did not reduce the number of operations. Analysing all the costs, it was found that a pathway including UDS costs more than routine care. Interviews were conducted that showed that men found UDS acceptable, and that the additional information helped both the men and their doctors consider which treatment would be most appropriate. These results do not support the routine use of UDS in the assessment of every man considering prostate surgery for LUTS. Further exploration of the data may identify circumstances in which UDS could be helpful.

Windows into Microbial Seascapes: Advances in Nanoscale Imaging and Application to Marine Sciences.

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.