Haplosporidium nelsoni and Perkinsus marinus occurrence in waters of Great Bay Estuary, New Hampshire.

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.

Understanding the role of COVID-19-related workplace stress and institutional betrayal on mental health in nurses: Some heroes wear scrubs.

This study examined the association of three specific COVID-19-related workplace stressors (percentage of nursing work with COVID-positive [COVID+] patients, number of COVID-19-related patient deaths witnessed, and living separately from family for safety) and their associations with posttraumatic stress symptoms (PTSS) and symptoms of major depressive disorder (MDD) and generalized anxiety disorder (GAD) among 391 nurses (93.6% White, 93.4% utilize she/her pronouns). Cross-sectional data were collected via an online survey. Institutional betrayal (i.e., the perception that an institution failed to protect a member who depends on and trusts it) was examined as a moderator of these associations. Although institutional betrayal was not a significant moderator in the three individual models, it held small-to-medium-sized positive main effects with PTSS and symptoms of GAD and MDD in both the individual and combined models. In the individual models, the percentage of nursing work with COVID+ patients was significantly positively associated with all three mental health conditions, f2 = .019-.195, whereas it only showed a significant effect with PTSS in the combined model, f2 = .138. Living separately from family was significantly positively associated with PTSS and MDD symptoms in both the individual, f2 = .037 and .015, respectively, and combined models, f2 = .025 and .013, respectively. Number of patient deaths held a significant positive association with PTSS alone, f2 = .022, in the individual model only. The findings are discussed in light of ways in which health care settings can better support and prioritize mental health among nursing staff.

Alcohol Use and Mental Health Symptoms in Nurses during the Early Months of COVID-19.

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.

Juvenile African Clawed Frogs (Xenopus laevis) Express Growth, Metamorphosis, Mortality, Gene Expression, and Metabolic Changes When Exposed to Thiamethoxam and Clothianidin.

Neonicotinoids (NEO) represent the main class of insecticides currently in use, with thiamethoxam (THX) and clothianidin (CLO) primarily applied agriculturally. With few comprehensive studies having been performed with non-target amphibians, the aim was to investigate potential biomarker responses along an adverse outcome pathway of NEO exposure, whereby data were collected on multiple biological hierarchies. Juvenile African clawed frogs, Xenopus laevis, were exposed to commercial formulations of THX and CLO at high (100 ppm) and low (20 ppm) concentrations of the active ingredient. Mortality, growth, development, liver metabolic enzyme activity, and gene expression endpoints were quantified. Tadpoles (n > 1000) from NF 47 through tail resorption stage (NF 66) were exposed to NEO or to NEO-free media treatments. Liver cell reductase activity and cytotoxicity were quantified by flow cytometry. Compared to control reference gene expressions, levels of expression for NEO receptor subunits, cell structure, function, and decontamination processes were measured by RT-qPCR by using liver and brain. Mortality in THX high was 21.5% compared to the control (9.1%); the metabolic conversion of THX to CLO may explain these results. The NF 57 control tadpoles were heavier, longer, and more developed than the others. The progression of development from NF 57-66 was reduced by THX low, and weight gain was impaired. Liver reductases were highest in the control (84.1%), with low NEO exhibiting the greatest reductions; the greatest cytotoxicity was seen with THX high. More transcriptional activity was noted in brains than in livers. Results affirm the utility of a study approach that considers multiple complexities in ecotoxicological studies with non-target amphibians, underscoring the need for simultaneously considering NEO concentration-response relationships with both whole-organism and biomarker endpoints.

Psychobiological Responses Reveal Audiovisual Noise Differentially Challenges Speech Recognition.

OBJECTIVES: In noisy environments, listeners benefit from both hearing and seeing a talker, demonstrating audiovisual (AV) cues enhance speech-in-noise (SIN) recognition. Here, we examined the relative contribution of auditory and visual cues to SIN perception and the strategies used by listeners to decipher speech in noise interference(s). DESIGN: Normal-hearing listeners (n = 22) performed an open-set speech recognition task while viewing audiovisual TIMIT sentences presented under different combinations of signal degradation including visual (AVn), audio (AnV), or multimodal (AnVn) noise. Acoustic and visual noises were matched in physical signal-to-noise ratio. Eyetracking monitored participants' gaze to different parts of a talker's face during SIN perception. RESULTS: As expected, behavioral performance for clean sentence recognition was better for A-only and AV compared to V-only speech. Similarly, with noise in the auditory channel (AnV and AnVn speech), performance was aided by the addition of visual cues of the talker regardless of whether the visual channel contained noise, confirming a multimodal benefit to SIN recognition. The addition of visual noise (AVn) obscuring the talker's face had little effect on speech recognition by itself. Listeners' eye gaze fixations were biased toward the eyes (decreased at the mouth) whenever the auditory channel was compromised. Fixating on the eyes was negatively associated with SIN recognition performance. Eye gazes on the mouth versus eyes of the face also depended on the gender of the talker. CONCLUSIONS: Collectively, results suggest listeners (1) depend heavily on the auditory over visual channel when seeing and hearing speech and (2) alter their visual strategy from viewing the mouth to viewing the eyes of a talker with signal degradations, which negatively affects speech perception.

Adjustment among children with relatives who participated in the manhunt following the Boston Marathon attack.

BACKGROUND: Following the Boston Marathon attack, the extraordinary interagency manhunt and shelter-in-place made for a truly unprecedented experience for area families. Although research on Boston youth has found robust associations between manhunt-related experiences and post-attack functioning, such work does little to identify the specific needs of a particularly vulnerable population--i.e., children with a relative who participated in the manhunt. Understanding the adjustment of these youth is critical for informing clinical efforts. METHODS: Survey of Boston-area parents/caretakers (N = 460) reporting on their child's attack/manhunt-related experiences, as well as psychosocial functioning in the first six post-attack months; analyses compared youth with and without a relative in law enforcement or the armed services who participated in the manhunt. RESULTS: The proportion of youth with likely PTSD was 5.7 times higher among youth with relatives in the manhunt than among youth without. After accounting for child demographics, blast exposure, and children's own exposure to manhunt events (e.g., hearing/seeing gunfire/explosions, having officers enter/search home), having a relative in the manhunt significantly predicted child PTSD symptoms, emotional symptoms, and hyperactivity/inattention. Fear during the manhunt that a loved one could be hurt mediated relationships between having a relative in the manhunt and clinical outcomes; living within the zone of greatest manhunt activity did not moderate observed relationships. CONCLUSIONS: Children with relatives called upon to participate in the unprecedented interagency manhunt following the Boston Marathon attack carried a particularly heavy mental health burden. Continued research is needed to clarify the clinical needs of youth with relatives in high-risk occupations.

Flow cytometric assessments of metabolic activity in bacterial assemblages provide insight into ecosystem condition along the Buffalo National River, Arkansas.

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 ~70 % active, except at Big Creek (56 %). The α-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 α-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.

Distinct function of Chlamydomonas CTRA-CTR transporters in Cu assimilation and intracellular mobilization.

Successful acclimation to copper (Cu) deficiency involves a fine balance between Cu import and export. In the green alga Chlamydomonas reinhardtii, Cu import is dependent on a transcription factor, Copper Response Regulator 1 (CRR1), responsible for activating genes in Cu-deficient cells. Among CRR1 target genes are two Cu transporters belonging to the CTR/COPT gene family (CTR1 and CTR2) and a related soluble protein (CTR3). The ancestor of these green algal proteins was likely acquired from an ancient chytrid and contained conserved cysteine-rich domains (named the CTR-associated domains, CTRA) that are predicted to be involved in Cu acquisition. We show by reverse genetics that Chlamydomonas CTR1 and CTR2 are canonical Cu importers albeit with distinct affinities, while loss of CTR3 did not result in an observable phenotype under the conditions tested. Mutation of CTR1, but not CTR2, recapitulates the poor growth of crr1 in Cu-deficient medium, consistent with a dominant role for CTR1 in high-affinity Cu(I) uptake. On the other hand, the overaccumulation of Cu(I) (20 times the quota) in zinc (Zn) deficiency depends on CRR1 and both CTR1 and CTR2. CRR1-dependent activation of CTR gene expression needed for Cu over-accumulation can be bypassed by the provision of excess Cu in the growth medium. Over-accumulated Cu is sequestered into the acidocalcisome but can become remobilized by restoring Zn nutrition. This mobilization is also CRR1-dependent, and requires activation of CTR2 expression, again distinguishing CTR2 from CTR1 and consistent with the lower substrate affinity of CTR2. ONE SENTENCE SUMMARY: Regulation of Cu uptake and sequestration by members of the CTR family of proteins in Chlamydomonas.

Distinct function of Chlamydomonas CTRA-CTR transporters in Cu assimilation and intracellular mobilization.

Successful acclimation to copper (Cu) deficiency involves a fine balance between Cu import and export. In the unicellular green alga Chlamydomonas reinhardtii , Cu import is dependent on C opper R esponse R egulator 1 (CRR1), the master regulator of Cu homeostasis. Among CRR1 target genes are two Cu transporters belonging to the CTR/COPT gene family ( CTR1 and CTR2 ) and a related soluble cysteine-rich protein (CTR3). The ancestor of these green algal proteins was likely acquired from an ancient chytrid and contained conserved cysteine-rich domains (named the CTR-associated domains, CTRA) that are predicted to be involved in Cu acquisition. We show by reverse genetics that Chlamydomonas CTR1 and CTR2 are canonical Cu importers albeit with distinct affinities, while loss of CTR3 did not result in an observable phenotype under the conditions tested. Mutation of CTR1 , but not CTR2 , recapitulate the poor growth of crr1 in Cu-deficient medium, consistent with a dominant role for CTR1 in high affinity Cu(I) uptake. Notably, the over-accumulation of Cu(I) in Zinc (Zn)-deficiency (20 times the quota) depends on CRR1 and both CTR1 and CTR2. CRR1-dependent activation of CTR gene expression needed for Cu over-accumulation can be bypassed by the provision of excess Cu in the growth medium. Over-accumulated Cu is sequestered into the acidocalcisome but can become remobilized by restoring Zn nutrition. This mobilization is also CRR1-dependent, and requires activation of CTR2 expression, again distinguishing CTR2 from CTR1 and is consistent with the lower substrate affinity of CTR2.

Architecture of the linker-scaffold in the nuclear pore.

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 hetero­octameric 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 structure­guided functional analysis in Saccharomyces cerevisiae, we confirmed the robustness of linker­scaffold 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 linker­scaffold 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 linker­scaffold 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 NPC­associated diseases, the role of NPC dilation in nucleocytoplasmic transport of soluble and integral membrane protein cargos, and the anchoring of asymmetric nucleoporins. [Figure: see text].

Architecture of the cytoplasmic face of the nuclear pore.

INTRODUCTION The subcellular compartmentalization of eukaryotic cells requires selective transport of folded proteins and protein-nucleic acid complexes. Embedded in nuclear envelope pores, which are generated by the circumscribed fusion of the inner and outer nuclear membranes, nuclear pore complexes (NPCs) are the sole bidirectional gateways for nucleocytoplasmic transport. The ~110-MDa human NPC is an ~1000-protein assembly that comprises multiple copies of ~34 different proteins, collectively termed nucleoporins. The symmetric core of the NPC is composed of an inner ring encircling the central transport channel and outer rings formed by Y­shaped coat nucleoporin complexes (CNCs) anchored atop both sides of the nuclear envelope. The outer rings are decorated with compartment­specific asymmetric nuclear basket and cytoplasmic filament nucleoporins, which establish transport directionality and provide docking sites for transport factors and the small guanosine triphosphatase Ran. The cytoplasmic filament nucleoporins also play an essential role in the irreversible remodeling of messenger ribonucleoprotein particles (mRNPs) as they exit the central transport channel. Unsurprisingly, the NPC's cytoplasmic face represents a hotspot for disease­associated mutations and is commonly targeted by viral virulence factors. RATIONALE Previous studies established a near-atomic composite structure of the human NPC's symmetric core by combining (i) biochemical reconstitution to elucidate the interaction network between symmetric nucleoporins, (ii) crystal and single-particle cryo-electron microscopy structure determination of nucleoporins and nucleoporin complexes to reveal their three-dimensional shape and the molecular details of their interactions, (iii) quantitative docking in cryo-electron tomography (cryo-ET) maps of the intact human NPC to uncover nucleoporin stoichiometry and positioning, and (iv) cell­based assays to validate the physiological relevance of the biochemical and structural findings. In this work, we extended our approach to the cytoplasmic filament nucleoporins to reveal the near-atomic architecture of the cytoplasmic face of the human NPC. RESULTS Using biochemical reconstitution, we elucidated the protein-protein and protein-RNA interaction networks of the human and Chaetomium thermophilum cytoplasmic filament nucleoporins, establishing an evolutionarily conserved heterohexameric cytoplasmic filament nucleoporin complex (CFNC) held together by a central heterotrimeric coiled­coil hub that tethers two separate mRNP­remodeling complexes. Further biochemical analysis and determination of a series of crystal structures revealed that the metazoan­specific cytoplasmic filament nucleoporin NUP358 is composed of 16 distinct domains, including an N­terminal S­shaped α­helical solenoid followed by a coiled­coil oligomerization element, numerous Ran­interacting domains, an E3 ligase domain, and a C­terminal prolyl­isomerase domain. Physiologically validated quantitative docking into cryo-ET maps of the intact human NPC revealed that pentameric NUP358 bundles, conjoined by the oligomerization element, are anchored through their N­terminal domains to the central stalk regions of the CNC, projecting flexibly attached domains as far as ~600 Å into the cytoplasm. Using cell­based assays, we demonstrated that NUP358 is dispensable for the architectural integrity of the assembled interphase NPC and RNA export but is required for efficient translation. After NUP358 assignment, the remaining 4-shaped cryo­ET density matched the dimensions of the CFNC coiled­coil hub, in close proximity to an outer-ring NUP93. Whereas the N-terminal NUP93 assembly sensor motif anchors the properly assembled related coiled­coil channel nucleoporin heterotrimer to the inner ring, biochemical reconstitution confirmed that the NUP93 assembly sensor is reused in anchoring the CFNC to the cytoplasmic face of the human NPC. By contrast, two C. thermophilum CFNCs are anchored by a divergent mechanism that involves assembly sensors located in unstructured portions of two CNC nucleoporins. Whereas unassigned cryo­ET density occupies the NUP358 and CFNC binding sites on the nuclear face, docking of the nuclear basket component ELYS established that the equivalent position on the cytoplasmic face is unoccupied, suggesting that mechanisms other than steric competition promote asymmetric distribution of nucleoporins. CONCLUSION We have substantially advanced the biochemical and structural characterization of the asymmetric nucleoporins' architecture and attachment at the cytoplasmic and nuclear faces of the NPC. Our near­atomic composite structure of the human NPC's cytoplasmic face provides a biochemical and structural framework for elucidating the molecular basis of mRNP remodeling, viral virulence factor interference with NPC function, and the underlying mechanisms of nucleoporin diseases at the cytoplasmic face of the NPC. [Figure: see text].

Effects of age and composition of field-produced biofilms on oyster larval setting.

Lack of success in restoring the native Eastern oyster, Crassostrea virginica, to Chesapeake Bay has been linked to the low occurrence of oyster larval setting in tributaries to the Bay. Among the many potential factors that could affect efforts to produce oysters through aquaculture or supplementation of shell beds is substratum condition. The present study examined larval setting on field-produced biofilms from Little Wicomico River (Virginia, USA) to assess whether bacterial community structure (examined by terminal restriction fragment length polymorphism, T-RFLP) or other characteristics of contemporary biofilms in this tributary (biofilm age and mass, algal abundance, and percentage organic matter) inhibited setting of larval oysters. The structure of the natural and heterogenous bacterial community in the biofilms and the success of oyster set were correlated, suggesting that specific microbial species may play a role in oyster setting. Larval set increased with biofilm age and mass, suggesting that established field-produced biofilms have no inhibitory effect. In contrast, the percentage of organic matter was negatively correlated with oyster set, whereas chlorophyll a concentration had no observed effect. The study expands prior knowledge by providing a more realistic timeframe for biofilm development (weeks as opposed to days), recounting effects of biofilms that are more representative of the natural dynamic and disturbance processes that would be expected to occur on submerged structures, and by incorporating seasonal and spatial variation. An important negative effect observed during the study period was heavy predation by Stylochus ellipticus on newly set oysters. Overall, the results of this study, which is the first assessment of the effects of biofilms produced naturally within a Chesapeake Bay tributary, suggest that the absence of large numbers of oysters in Little Wicomico River is not related to microbes or other specific characteristics of biofilms that develop on suitable setting substrata, but rather to heavy predation of newly set larvae.

Nutrient bioassimilation capacity of aquacultured oysters: quantification of an ecosystem service.

Like many coastal zones and estuaries, the Chesapeake Bay has been severely degraded by cultural eutrophication. Rising implementation costs and difficulty achieving nutrient reduction goals associated with point and nonpoint sources suggests that approaches supplemental to source reductions may prove useful in the future. Enhanced oyster aquaculture has been suggested as one potential policy initiative to help rid the Bay waters of excess nutrients via harvest of bioassimilated nutrients. To assess this potential, total nitrogen (TN), total phosphorous (TP), and total carbon (TC) content were measured in oyster tissue and shell at two floating-raft cultivation sites in the Chesapeake Bay. Models were developed based on the common market measurement of total length (TL) for aquacultured oysters, which was strongly correlated to the TN (R2 = 0.76), TP (R2 = 0.78), and TC (R2 = 0.76) content per oyster tissue and shell. These models provide resource managers with a tool to quantify net nutrient removal. Based on model estimates, 10(6) harvest-sized oysters (76 mm TL) remove 132 kg TN, 19 kg TP, and 3823 kg TC. In terms of nutrients removed per unit area, oyster harvest is an effective means of nutrient removal compared with other nonpoint source reduction strategies. At a density of 286 oysters m(-2), assuming no mortality, harvest size nutrient removal rates can be as high as 378 kg TN ha(-1), 54 kg TP ha(-1), and 10,934 kg TC ha(-1) for 76-mm oysters. Removing 1 t N from the Bay would require harvesting 7.7 million 76-mm TL cultivated oysters.

Vesicular monoamine transporter-1 (VMAT-1) mRNA and immunoreactive proteins in mouse brain.

OBJECTIVE: Vesicular monoamine transporter 1 (VMAT-1) mRNA and protein were examined (1) to determine whether adult mouse brain expresses full-length VMAT-1 mRNA that can be translated to functional transporter protein and (2) to compare immunoreactive VMAT-1 proteins in brain and adrenal. METHODS: VMAT-1 mRNA was detected in mouse brain with RT-PCR. The cDNA was sequenced, cloned into an expression vector, transfected into COS-1 cells, and cell protein was assayed for VMAT-1 activity. Immunoreactive proteins were examined on western blots probed with four different antibodies to VMAT-1. RESULTS: Sequencing confirmed identity of the entire coding sequences of VMAT-1 cDNA from mouse medulla oblongata/pons and adrenal to a Gen-Bank reference sequence. Transfection of the brain cDNA into COS-1 cells resulted in transporter activity that was blocked by the VMAT inhibitor reserpine and a proton ionophore, but not by tetrabenazine, which has a high affinity for VMAT-2. Antibodies to either the C- or N- terminus of VMAT-1 detected two proteins (73 and 55 kD) in transfected COS-1 cells. The C-terminal antibodies detected both proteins in extracts of mouse medulla/pons, cortex, hypothalamus, and cerebellum but only the 73 kD protein and higher molecular weight immunoreactive proteins in mouse adrenal and rat PC12 cells, which are positive controls for rodent VMAT-1. CONCLUSIONS: These findings demonstrate that a functional VMAT-1 mRNA coding sequence is expressed in mouse brain and suggest processing of VMAT-1 protein differs in mouse adrenal and brain.

Direct-Care Staff Perceptions of Patient Engagement and Treatment Planning in Detox.

As the prevalence of substance use disorders and drug-related deaths continue to rise, addiction treatment facilities are charged with providing effective and efficient services to curb the national substance use crisis. Direct-care staff in treatment service facilities play a crucial role in whether or not evidence-based practices are incorporated. Without their understanding and utilization of patient engagement best practices, an organization risks maintaining the status quo rather than actively pursuing improved outcomes through empirically supported approaches. Through in-depth interviews (N=13) with nurses, counselors, and behavioral health technicians in an inpatient detoxification facility, this study evaluates the perspectives and experiences of direct-care staff through a lens of patient engagement in treatment planning. The findings from these interviews elucidate how participants' personal characteristics and values, perspectives of patient engagement, understanding of treatment planning, and organizational culture and operations facilitate or inhibit the integration of patient engagement for treatment planning in detox.

A baseline for microplastic particle occurrence and distribution in Great Bay Estuary.

We extracted and analyzed microplastics (MP) in archived sediment cores from Great Bay Estuary (GBE) in the Gulf of Maine region of North America. Results indicated that MP are distributed in GBE sediments, 0-30 cm, at an average occurrence of 116 ± 21 particles g-1 and that morphology varies by site and depth. Analysis by sediment depth and age class indicated that MP accumulation increased over several decades but recently (5-10 years) has likely begun to decrease. Hydrodynamic and particle transport modeling indicated that bed characteristics are a more controlling factor in MP distribution than typical MP properties and that the highest accumulation likely occurs in regions with weaker hydrodynamic flows and lower bed shear stress, e.g., eelgrass meadows and along fringes of the Bay. These results provide a baseline and predictive understanding of the occurrence, morphology, and sedimentation of MP in the estuary.

Use of Middleware Data to Dissect and Optimize Hematology Autoverification.

BACKGROUND: Hematology analysis comprises some of the highest volume tests run in clinical laboratories. Autoverification of hematology results using computer-based rules reduces turnaround time for many specimens, while strategically targeting specimen review by technologist or pathologist. METHODS: Autoverification rules had been developed over a decade at an 800-bed tertiary/quarternary care academic medical central laboratory serving both adult and pediatric populations. In the process of migrating to newer hematology instruments, we analyzed the rates of the autoverification rules/flags most commonly associated with triggering manual review. We were particularly interested in rules that on their own often led to manual review in the absence of other flags. Prior to the study, autoverification rates were 87.8% (out of 16,073 orders) for complete blood count (CBC) if ordered as a panel and 85.8% (out of 1,940 orders) for CBC components ordered individually (not as the panel). RESULTS: Detailed analysis of rules/flags that frequently triggered indicated that the immature granulocyte (IG) flag (an instrument parameter) and rules that reflexed platelet by impedance method (PLT-I) to platelet by fluorescent method (PLT-F) represented the two biggest opportunities to increase autoverification. The IG flag threshold had previously been validated at 2%, a setting that resulted in this flag alone preventing autoverification in 6.0% of all samples. The IG flag threshold was raised to 5% after detailed chart review; this was also the instrument vendor's default recommendation for the newer hematology analyzers. Analysis also supported switching to PLT-F for all platelet analysis. Autoverification rates increased to 93.5% (out of 91,692 orders) for CBC as a panel and 89.8% (out of 11,982 orders) for individual components after changes in rules and laboratory practice. CONCLUSIONS: Detailed analysis of autoverification of hematology testing at an academic medical center clinical laboratory that had been using a set of autoverification rules for over a decade revealed opportunities to optimize the parameters. The data analysis was challenging and time-consuming, highlighting opportunities for improvement in software tools that allow for more rapid and routine evaluation of autoverification parameters.

Developing an emergency department order set to treat acute pain in sickle cell disease.

STUDY OBJECTIVE: Patients with sickle cell disease (SCD) have many emergency department visits because of painful vaso-occlusive episodes (VOE). Guidelines recommend treatment within 30 minutes of triage, but this is rarely achieved in clinical practice. Our goal was to develop an order set that is being implemented in the ED to facilitate and standardize emergency care for SCD patients in acute pain from VOEs presenting to the emergency department (ED) in New York City (NYC). METHODS: Using a RAND/University of California, Los Angeles modified Delphi panel, we convened a multidisciplinary panel and reviewed evidence on how to best manage SCD pain in the ED. Panelists collaboratively developed then rated 202 items that could be included in an ED order set. RESULTS: A consensus order set, a practical how-to guide for managing SCD pain in the ED, was developed based on items that received high median ratings. CONCLUSIONS: The management of acute pain experienced during VOEs is critical to patients with SCD; ED order sets, such as this one, can help standardize pain management, including at triage, evaluation, discharge, and follow-up care. After implementation in NYC EDs, studies to examine changes in quality care metrics (eg, wait times, readmissions) are planned.

Trace Metal Availability Affects Greenhouse Gas Emissions and Microbial Functional Group Abundance in Freshwater Wetland Sediments.

We investigated the effects of trace metal additions on microbial nitrogen (N) and carbon (C) cycling using freshwater wetland sediment microcosms amended with micromolar concentrations of copper (Cu), molybdenum (Mo), iron (Fe), and all combinations thereof. In addition to monitoring inorganic N transformations (NO3 -, NO2 -, N2O, NH4 +) and carbon mineralization (CO2, CH4), we tracked changes in functional gene abundance associated with denitrification (nirS, nirK, nosZ), dissimilatory nitrate reduction to ammonium (DNRA; nrfA), and methanogenesis (mcrA). With regards to N cycling, greater availability of Cu led to more complete denitrification (i.e., less N2O accumulation) and a higher abundance of the nirK and nosZ genes, which encode for Cu-dependent reductases. In contrast, we found sparse biochemical evidence of DNRA activity and no consistent effect of the trace metal additions on nrfA gene abundance. With regards to C mineralization, CO2 production was unaffected, but the amendments stimulated net CH4 production and Mo additions led to increased mcrA gene abundance. These findings demonstrate that trace metal effects on sediment microbial physiology can impact community-level function. We observed direct and indirect effects on both N and C biogeochemistry that resulted in increased production of greenhouse gasses, which may have been mediated through the documented changes in microbial community composition and shifts in functional group abundance. Overall, this work supports a more nuanced consideration of metal effects on environmental microbial communities that recognizes the key role that metal limitation plays in microbial physiology.

Metagenomic analysis of planktonic riverine microbial consortia using nanopore sequencing reveals insight into river microbe taxonomy and function.

BACKGROUND: Riverine ecosystems are biogeochemical powerhouses driven largely by microbial communities that inhabit water columns and sediments. Because rivers are used extensively for anthropogenic purposes (drinking water, recreation, agriculture, and industry), it is essential to understand how these activities affect the composition of river microbial consortia. Recent studies have shown that river metagenomes vary considerably, suggesting that microbial community data should be included in broad-scale river ecosystem models. But such ecogenomic studies have not been applied on a broad "aquascape" scale, and few if any have applied the newest nanopore technology. RESULTS: We investigated the metagenomes of 11 rivers across 3 continents using MinION nanopore sequencing, a portable platform that could be useful for future global river monitoring. Up to 10 Gb of data per run were generated with average read lengths of 3.4 kb. Diversity and diagnosis of river function potential was accomplished with 0.5-1.0 ⋅ 106 long reads. Our observations for 7 of the 11 rivers conformed to other river-omic findings, and we exposed previously unrecognized microbial biodiversity in the other 4 rivers. CONCLUSIONS: Deeper understanding that emerged is that river microbial consortia and the ecological functions they fulfil did not align with geographic location but instead implicated ecological responses of microbes to urban and other anthropogenic effects, and that changes in taxa manifested over a very short geographic space.