Typical investigations into the biological consequences of suspected xenobiotics or nutrients introduced in watersheds include analytical chemistry screens of environmental samples—such as periphyton responses or studies of fish condition—which are all costly in terms of equipment, reagents, time, and human resources. An alternative is to assess pollutant effects on waterborne bacteria. A flow cytometric method was developed to yield rapid, same-day results that could be used to proactively screen for suspected chemical inputs into watersheds using water sampling methods that are identical to those in standard use. The analytical methods are microbe cultivation-independent, for use with waterborne bacteria that are typically viable but not culturable. The procedure is quick and inexpensive, generating measures of bacterial esterase that reflect metabolic activity and are sensitive and statistically robust. After phosphate-EDTA incubation to increase cell wall permeability, staining was performed with 5(6) carboxyfluorescein diacetate (enzyme activity) and propidium iodide (cell viability) with three bacterial species in exponential phase growth having been incubated with organic wastewater compounds (atrazine, pharmaceuticals [17α-ethynylestradiol and trenbolone], and antimicrobials [tylosin and butylparaben]). This method successfully detected metabolic changes in all bacterial species, with atrazine inducing the greatest change. Additional fluorescent stains can target specific microbial structures or functions of interest in a particular watershed. This biotechnology can inform analytical chemistry and study of biota at sites of interest and has the potential to be automated.

In Great Bay Estuary, New Hampshire, USA, Haplosporidium nelsoni and Perkinsus marinus are 2 active pathogens of the eastern oyster Crassostrea virginica (Gmelin), that cause MSX (multinucleated sphere with unknown affinity ‘X’) and dermo mortalities, respectively. Whereas studies have quantified infection intensities in oyster populations and determined whether these parasites exist in certain planktonic organisms, no studies thus far have examined both infectious agents simultaneously in water associated with areas that do and do not have oyster populations. As in other estuaries, both organisms are present in estuarine waters throughout the Bay, especially during June through November, when oysters are most active. Waters associated with oyster habitats had higher, more variable DNA concentrations from these pathogenic organisms than waters at a non-oyster site. This finding allows for enhanced understanding of disease-causing organisms in New England estuaries, where oyster restoration is a priority.

Ecosystems in coastal waters of Gulf of Maine (GOM) are undergoing environmental challenges in response to climate change and anthropogenic stressors. eDNA metabarcoding, a powerful tool for assessing the fish community structure, was used to identify fish communities in three types of GOM aquatic environments (sand, macroalgae, and eelgrass) in Maine and New Hampshire, USA. The available 12S rRNA fish universal primer analysis system (MiFish and 12S-V5) was modified using nested polymerase chain reaction (PCR) to improve targeting of fish products and reduce non-target products. The nested PCR strategy allowed successful amplification of 12S rRNA genes in fishes without production of non-target products and identified 28 fish groups at the genus level. Presence/Absence data and Relative Abundance showed significant differences among locales but not among habitats. Myoxocephalus sp. were found at all sampling sites. Relative Abundance data revealed that Menidia menidia and Brevoortia sp. were statistical indicator species in Goosefare, Maine, and New castle, New Hampshire, respectively. Although beta diversity indicated that fish communities were not different across habitats, statistical analysis found that Pholis sp. and Ammodytes sp. were dominant species in macroalgae and sand, respectively. To our knowledge, this is the first metabarcoding study to assess fish communities in the Western Atlantic region using the MiFish primer set, and the study suggests that metabarcoding is useful for mapping geographic and temporal marine fish diversity.

There is preliminary evidence that the anticonvulsant medication Zonisamide (ZON) may be an effective, well-tolerated treatment for alcohol use disorder (AUD). However, further evaluation of its efficacy for treating patients with AUD is needed, and much remains unknown about ZON’s therapeutic mechanisms. The present study aimed to evaluate the efficacy and tolerability of ZON in a double-blind, placebo-controlled, randomized trial. Eighty-one adults (ages 21-65) diagnosed with AUD were randomly assigned to receive either ZON (at a target dose of 400 mg/d) or a pill placebo over 12 weeks, followed by a two-week taper. All participants also received a computerized alcohol reduction program, Take Control (TC). Primary drinking outcomes were average daily drinks, percentage drinking days, and percentage heavy drinking days. Further, we evaluated changes in AUD clinical severity and performance on neuropsychological measures. For both groups, drinking outcomes generally decreased, as did AUD clinical severity, though group differences were not statistically significant. Neuropsychological testing performance was similar for both groups at baseline; however, at post-treatment, participants in the ZON group demonstrated poorer working memory and lower performance on verbal fluency tests compared to the placebo group, and these differences were statistically significant with moderate-large effect sizes. One serious adverse event was reported among individuals receiving ZON. Study findings indicate that ZON combined with TC does not demonstrate superior effectiveness for reducing average daily drinks in this clinical sample with principal AUD compared to placebo and TC, and treatment with ZON may be associated with reduced neurocognitive performance over time.

Nitrification inhibitors, such as nitrapyrin (NI), are increasingly co-applied with nitrogen (N) fertilizers as part of sustainable agricultural practice. Several studies in temperate regions have documented the effectiveness of NI in retaining soil ammonium (NH4+), minimizing N loss and increasing crop yields. However, less is known about the effects of NI in Mediterranean regions, where agricultural production is challenging and requires intensive irrigation and fertilization. We investigated the short-term impact of the nitrification inhibitor nitrapyrin (2-chloro-6-(trichloromethyl)pyridine) in a two-factor mesocosm experiment, using a typical Mediterranean soil, where NI was co-applied with a selection of urea-based fertilizers: urea (U), U with urease inhibitors (U + UI), methylene urea (MU) and zeolite-coated urea (ZU). NI co-applied with urea fertilizers resulted in higher availability of soil NH4+ and a concurrent increase in NH3 volatilization. Net cumulative soil NH4+ availability was 1.5-3.3 fold greater when NI was applied. Concurrently, net cumulative nitrate (NO3-) and nitrite (NO2-) availability was reduced by 10%-60%; this was found for all the tested fertilizer types except MU fertilizer, where the net cumulative soil NO3- and NO2- doubled. Nitrous oxide (N2O) emissions from urea fertilization were reduced by 40% with UI, 50% with NI and 66% with NI + UI. Interestingly, after 28 d, the composition of soil microbial communities was distinctly different, due to NI application. Specifically, NI application dramatically reduced the abundance of ammonia-oxidizing and denitrifying bacterial functional groups. NI was effective in reducing N2O emissions in this calcareous soil; however, NH3 emissions were remarkably enhanced. These findings have important implications for the large-scale adoption of inhibitor technologies in Mediterranean agroecosystems and for the reduction of greenhouse gas emissions.

The yellow pitcher plant, Sarracenia flava, is an insectivorous perennial distributed extensively in southeastern North America. In Virginia, it is restricted to a few wetland ecosystems, with only one natural site known to remain. To uncover whether there were microbial differences in the rhizospheres across natural and reintroduced sites of pitcher plant restoration, shotgun metagenome sequencing was undertaken to characterize the microbiomes of the healthy rhizosphere in the last remaining natural stand in Virginia compared to rhizospheres sampled in two restored habitats where pitcher plants were reintroduced and a nearby control habitat without pitcher plants. Statistical analysis showed no significant differences in rhizobiome communities among the natural, reintroduced, and control sites. Comparison of test rhizobiomes with those of other soil types revealed no significant difference in S. flava habitats versus wildland soil types but significant difference from agricultural soils. Indicator species analysis found Pseudomonas was a significantly more abundant genus in the S. flava habitats. The control site was enriched with iron-reducing bacteria compared to the rest of the sites. Further studies based on gene expression could better facilitate an understanding of the role of Pseudomonas in S. flava rhizosphere specific to habitats, which will provide better knowledge for local conservation of this plant.

Eastern oysters, Crassostrea virginica, are historically a keystone species in many of the estuaries in which they reside, providing critical ecosystem services. Because oyster populations have been on the decline, restoration initiatives currently are underway in many estuarine systems including Great Bay Estuary (GBE), New Hampshire. Results of prior studies of eastern oyster population genomics cannot be applied directly to GBE, as it is a well-mixed estuarine system that is relatively contained, and the sources of recruits are split among cultivated and native. This study aimed to identify the population genomic structure of eastern oysters in GBE, to facilitate determination of effective population size and estimation of genetic differentiation among subpopulations. Results showed moderate genomic differentiation among native, cultivated, and restoration C. virginica subpopulations in the Bay. A small number of breeders (Ne=163 – 276) was found in all subpopulations except the Lamprey River site (Ne=995). This research provides a contemporary snapshot of eastern oyster subpopulation structure at the genomic level in GBE that will facilitate restoration and enhanced management.

Past studies have combined passive acoustic monitoring with environmental genetic methods to detect target organisms, however, no studies to date have employed metagenomics concurrent with passive acoustic monitoring of soundscapes for comparison of marine habitats. The present study used both approaches simultaneously for holistic observation of marine habitats to reveal information beyond using either technique independently. Water samples for metabarcoding (four primer sets) were collected during periods of passive acoustic monitoring from three different marine habitats, each at four different geographic locations along the New Hampshire/Maine coastlines. Multivariate analyses compared discrimination among habitat types and geographic locations by analyzing acoustic metrics generated using the Soundscape Code and metagenomic taxonomic assignments. Passive acoustic monitoring provided insight into environmental features that were unobservable with metagenomics, especially anthropogenic activity and geophysical processes, whereas metagenomics provided a more complete picture of the biological composition of habitats through detection of organisms that were not actively producing sound. This enables simultaneous evaluation of biological and functional connectivity of marine habitats by detecting what organisms are present and their contributions to the soundscape. In future, genetic and acoustic indicators will be used for prediction of substrate characteristics and sound sources to model acoustic propagation environments.

Eastern oysters, Crassostrea virginica, are historically a keystone species in many of the estuaries in which they reside, providing critical ecosystem services. Because oyster populations have been on the decline for the past century, restoration initiatives currently are underway in many estuarine systems, including Great Bay Estuary (GBE), New Hampshire. Results of prior studies of eastern oyster population genomics cannot be applied directly to GBE, as it is a well-mixed estuarine system that is relatively contained, and the sources of recruits are split among cultivated and native. This study aimed to identify the population genomic structure of eastern oysters in GBE to facilitate determination of effective population size and estimation of genetic differentiation among subpopulations. Results showed moderate genomic differentiation among native, cultivated, and restoration C. virginica subpopulations in the Bay. A small number of breeders (Ne=163-276) was found in all subpopulations except the Lamprey River site (Ne=995). This research provides a contemporary snapshot of eastern oyster subpopulation structure at the genomic level in GBE that will facilitate restoration and enhanced management.

Great Bay Estuary (GBE), within the rapidly warming Gulf of Maine, has experienced significant ecological shifts this century due to naturalization of invasive species. The range expansion of the American blue crab (Callinectes sapidus) currently underway from the mid-Atlantic northward brings the possibility of similar ecological shifts. This study accounts recent trapping and diet analysis of C. sapidus in GBE. Diet is an important component of understanding how the blue crab range expansion may affect GBE ecosystem functions. Across all sites and trap types, 27 blue crabs were captured. Metagenomic analysis of shotgun sequencing techniques were used on the gut contents of blue crabs captured. Most specimens had > 50

ABSTRACTGreat Bay Estuary (GBE) is a complex estuarine system at the convergence of seven rivers, ocean and land. The GBE ecosystem supports typical estuarine species but, like many estuaries, has been a target of both invasive and range‐expanding species. The goal of this study was to compare abundance, sex and size distribution of green crabs between oyster farm sites and non‐farmed sites within GBE in 2021 and 2022. Overall catch was higher in 2021 compared to 2022. The highest population of green crabs was estimated to be in the farm area around Cedar Point, and natural/restoration area near Moody Point had the smallest European green crab population. In both years, the majority of green crabs captured (98%) were adult males, and few were juveniles captured. At the last sampling date of 2021 and 2022, most crabs were large (68 ± 0.5 mm and 63 ± 2 mm carapace width, respectively). Shellfish growers, restoration and management teams could use these data in mitigation strategies to sustain industries and enhance GBE health. This information is important for oyster restoration initiatives and oyster growers as they may want to: consider putting traps around their restoration reefs and farms to help reduce green crab infestation/predation in their growing areas; make decisions regarding best management practices and optimal growing locations when applying for permits; and work with management teams with gear improvement, such as floating gear, to reduce green crab intrusion into cages. High‐priority locations for trapping should be areas such as oyster farms, oyster reefs and restoration sites.

The Buffalo National River (BNR), on karst terrain in Arkansas, is considered an extraordinary water resource. Water collected in Spring 2017 along BNR was metagenomically analyzed using 16S rDNA, and for 17 months (5/2017-11/2018), bacterial responses were measured in relation to nutrients sampled along a stretch of BNR near a concentrated animal feed operation (CAFO) on Big Creek. Because cell count and esterase activity can increase proportionally with organic enrichment, they were hypothesized to be elevated near the CAFO. Counts (colony forming units; CFUs) were different among sites for 73 % of the months; Big Creek generated highest CFUs 27 % of the time, with the closest downstream site at 13.3 %. Esterase activity was different among sites 94 % of the time, with Big Creek exhibiting lowest activity 71 % of the time. Over the months, activity was similar across sites at similar to 70 % active, except at Big Creek (56 %). The alpha-diversity of BNR microbial consortia near a wastewater treatment plant (WWTP) and the CAFO was related to distance from the WWTP and CAFO. The inverse relationship between high CFUs and low esterase activity at Big Creek (r = -0.71) actuated in vitro exposures of bacteria to organic wastewater contaminants (OWC) previously identified in the watershed. Exponential-phase Escherichia coli (stock strain), Streptococcus suis (avirulent, from swine), and S. dysgalactiae (virulent, from silver carp, Hypophthalmichthys molitrix) were incubated with atrazine, pharmaceuticals (17 alpha-ethynylestradiol and trenbolone), and antimicrobials (tylosin and butylparaben). Bacteria were differentially responsive. Activity varied with exposure time and OWC type, but not concentration; atrazine decreased it most. Taken together - the metagenomic taxonomic similarities along BNR, slightly higher bacterial growth and lower bacterial esterase at the CAFO, and the lab exposures of bacterial strains showing that OWC altered metabolism - the results indicated that bioactive OWC entering the watershed can strongly influence microbial processes in the aquatic ecosystem.

Ecological connectivity among coastal marine habitats—linkage in the movement of organisms and natural processes across habitat boundaries—has significant implications for the health and resilience of commercially important or threatened species in the Gulf of Maine (GOM), off the coast of the northeastern United States. Methods designed to efficiently assess connectivity are vital for identifying and managing critical habitats (Perry et al., 2018). Paired use of passive acoustic monitoring (PAM) and metabarcoding seawater samples (MSS) for observing biological and functional connectivity at various spatiotemporal scales in the marine environment is largely unexplored and may provide an efficient alternative or supplement to existing strategies.

Identifying similarities and differences in soundscape properties among coastal marine habitats is valuable for determining indicators of habitat composition, assessing functional connectivity among habitats, and informing management decisions regarding the soundscapes of these habitats. The “Soundscape Code,” proposed and developed by Dylan Wilford, enables rapid calculation of values for four salient soundscape properties: amplitude, impulsivity, periodicity, and dissimilarity. This enables multivariate statistical analyses to quantitatively compare soundscapes in different habitat types and geographic regions. The objective of the current work was to determine whether geographic region or habitat type accounts for more variability in coastal soundscape properties. Passive acoustic recordings were acquired in three different habitat types (sand, macroalgae, and eelgrass dominated substrates), in each of four different geographic locations along the New Hampshire/Maine coastline, to compare the soundscapes of habitats with varied biological and geophysical substrate composition. Results indicate that geographic location accounted for more variability in the soundscapes than habitat type, suggesting that habitats’ local connectivity outweighs acoustic and biological uniformity of the same habitat type over broader spatial scales. Future analyses will incorporate metagenomic data for predictive modeling of habitat composition through the combined use of passive acoustic monitoring and metabarcoding of seawater samples.

On 2 September and 7 October 2022, we captured post-copulatory pairs of Callinectes sapidus (Blue Crab) in Green Crab traps in Great Bay Estuary, NH. On the first occasion, the 2 crabs included a mature post-ecdysis (12-24 hour) female and a mature male, and the female's shed exoskeleton also remained in the trap. Both seminal receptacles had pink sperm plugs and were turgid with spermatophores, indicative of recent successful insemination. Details of the second capture mirrored the first with the exception that the exuvia was not in the trap. This is the first scientific reporting of Blue Crabs mating in Great Bay Estuary, NH, documenting an ongoing range expansion of a species with the potential to have major ecological and economic impacts.

To better understand differential sensitivities among fish species to the piscicidal compound Antimycin-A (ANT-A), we hypothesized that variations in amino acids at the ANT-A binding site may reflect toxicity differences. Protein sequences for six motifs comprising the ANT-A binding site were obtained and compared for invasive carp species (N = 515) and seven non-target species (N = 277); a consensus was delineated from each species. The carp species, Common Carp (Cyprinus carpio), Silver Carp (Hypophthalmichthys molitrix), Bighead Carp (Hypophthalmichthys nobilis), Grass Carp (Ctenopharyngodon idella), and Black Carp (Mylopharyngodon piceus), showed the same amino acids at the site; thus, it was termed the carp consensus motif sequence (CCM). Channel Catfish (Ictalurus punctatus) showed the most amino acid polymorphisms, with three motifs 96–100% different from CCM. Within a species, Bluegill (Lepomis macrochirus) and Fathead Minnow (Pimephales promelas) variation per motif was most dissimilar (46.7% and 21.6%, respectively). Organismal mortality data from the literature indicated Yellow Perch (Perca flavescens), Walleye (Sander vitreus), and American Gizzard Shad (Dorosoma cepedianum) to be most sensitive to the piscicide, Catfish least sensitive, and all others intermediate. The protein sequence variations of the binding site appeared to be in accord with organismal sensitivity categories when they differed from the CCM; the motifs in Gizzard Shad and Walleye were the same as in CCM. The physical/chemical nature of ANT-A is important to consider in organismal response comparisons. This cellular approach of studying ANT-A binding at its target enzyme is a non-destructive way to predict piscicidal efficacy of ANT-A against fishes of interest, informs management decisions in control efforts for invasives, and can be used to forecast effects on sympatric species.

OBJECTIVE:The current study examined associations of symptoms of posttraumatic stress disorder [PTSD], depression, and generalized anxiety disorder [GAD] with alcohol consumption and drinking to cope in a sample of 310 nurses during the first six months of the COVID-19 pandemic. METHOD:Using a cross-sectional design, nurses completed online surveys. RESULTS:Over 50% of the sample reported alcohol misuse and 12.2% reported drinking to cope. Further, 38.7% reported elevated symptoms of PTSD, 29.7% moderate-to-high symptoms of depression, and 56.8% elevated symptoms of GAD symptoms. Hierarchical regression analyses were conducted to examine how mental health symptoms were associated with alcohol outcomes, controlling for age, gender pronouns, education, and race. No significant predictors emerged for alcohol consumption. Significant associations of symptoms of PTSD and depression were found for drinking to cope, such that higher levels of mental health symptoms were associated with greater endorsement of drinking to cope. CONCLUSION:Results are discussed in light of increasing prevention and support services for nurses.

INTRODUCTION In eukaryotic cells, the selective bidirectional transport of macromolecules between the nucleus and cytoplasm occurs through the nuclear pore complex (NPC). Embedded in nuclear envelope pores, the ~110-MDa human NPC is an ~1200-Å-wide and ~750-Å-tall assembly of ~1000 proteins, collectively termed nucleoporins. Because of the NPC’s eightfold rotational symmetry along the nucleocytoplasmic axis, each of the ~34 different nucleoporins occurs in multiples of eight. Architecturally, the NPC’s symmetric core is composed of an inner ring encircling the central transport channel and two outer rings anchored on both sides of the nuclear envelope. Because of its central role in the flow of genetic information from DNA to RNA to protein, the NPC is commonly targeted in viral infections and its nucleoporin constituents are associated with a plethora of diseases. RATIONALE Although the arrangement of most scaffold nucleoporins in the NPC’s symmetric core was determined by quantitative docking of crystal structures into cryo–electron tomographic (cryo-ET) maps of intact NPCs, the topology and molecular details of their cohesion by multivalent linker nucleoporins have remained elusive. Recently, in situ cryo-ET reconstructions of NPCs from various species have indicated that the NPC’s inner ring is capable of reversible constriction and dilation in response to variations in nuclear envelope membrane tension, thereby modulating the diameter of the central transport channel by ~200 Å. We combined biochemical reconstitution, high-resolution crystal and single-particle cryo–electron microscopy (cryo-EM) structure determination, docking into cryo-ET maps, and physiological validation to elucidate the molecular architecture of the linker-scaffold interaction network that not only is essential for the NPC’s integrity but also confers the plasticity and robustness necessary to allow and withstand such large-scale conformational changes. RESULTS By biochemically mapping scaffold-binding regions of all fungal and human linker nucleoporins and determining crystal and single-particle cryo-EM structures of linker-scaffold complexes, we completed the characterization of the biochemically tractable linker-scaffold network and established its evolutionary conservation, despite considerable sequence divergence. We determined a series of crystal and single-particle cryo-EM structures of the intact Nup188 and Nup192 scaffold hubs bound to their Nic96, Nup145N, and Nup53 linker nucleoporin binding regions, revealing that both proteins form distinct question mark–shaped keystones of two evolutionarily conserved hetero‑octameric inner ring complexes. Linkers bind to scaffold surface pockets through short defined motifs, with flanking regions commonly forming additional disperse interactions that reinforce the binding. Using a structure‑guided functional analysis in Saccharomyces cerevisiae , we confirmed the robustness of linker‑scaffold interactions and established the physiological relevance of our biochemical and structural findings. The near-atomic composite structures resulting from quantitative docking of experimental structures into human and S. cerevisiae cryo-ET maps of constricted and dilated NPCs structurally disambiguated the positioning of the Nup188 and Nup192 hubs in the intact fungal and human NPC and revealed the topology of the linker-scaffold network. The linker-scaffold gives rise to eight relatively rigid inner ring spokes that are flexibly interconnected to allow for the formation of lateral channels. Unexpectedly, we uncovered that linker‑scaffold interactions play an opposing role in the outer rings by forming tight cross-link staples between the eight nuclear and cytoplasmic outer ring spokes, thereby limiting the dilatory movements to the inner ring. CONCLUSION We have substantially advanced the structural and biochemical characterization of the symmetric core of the S. cerevisiae and human NPCs and determined near-atomic composite structures. The composite structures uncover the molecular mechanism by which the evolutionarily conserved linker‑scaffold establishes the NPC’s integrity while simultaneously allowing for the observed plasticity of the central transport channel. The composite structures are roadmaps for the mechanistic dissection of NPC assembly and disassembly, the etiology of NPC‑associated diseases, the role of NPC dilation in nucleocytoplasmic transport of soluble and integral membrane protein cargos, and the anchoring of asymmetric nucleoporins. Linker-scaffold architecture in the human NPC’s symmetric core. Near‑atomic composite structure of the NPC’s symmetric core obtained by quantitative docking of high-resolution crystal and single-particle cryo-EM structures into a cryo-ET reconstruction of the intact human NPC. Schematic representations of the intricate linker-scaffold topology of the cytoplasmic outer ring, inner ring, and nuclear outer ring (clockwise from top) are depicted for the boxed regions. C, C terminus; N, N terminus.