2020 W.M. Keck Science Department Publications and Grants

*Indicates student co-author

Tombak, Kaia J., Sarah A. Budischak, Stephanie Hauck, Lindsay A. Martinez, and Daniel I. Rubenstein. “The Non-Invasive Measurement of Faecal Immunoglobulin in African Equids.” International Journal for Parasitology: Parasites and Wildlife, vol 12, 2020, pp. 105-112.

Abstract: Eco-immunological research is encumbered by a lack of basic research in a wild context and by the availability of few non-invasive tools to measure the internal state of wild animals. The recent development of an enzyme-linked immunosorbent assay for measuring immunoglobulins in faecal samples from Soay sheep prompted us to optimize such an assay to measure immunoglobulin A (IgA: an antibody associated with parasitic nematode fecundity) in faecal samples from equids. We measured total IgA in domestic donkeys, wild plains zebras, and wild Grevy's zebras sharing the same landscape in central Kenya over two field seasons. Attempts to measure anti-nematode IgA more specifically, using a homogenized extract from a mixture of excreted nematodes, failed to clear background. However, we found that total IgA positively correlated with strongyle nematode faecal egg counts (FECs) in donkeys sampled during the wetter field season - a time when the donkeys were in good condition. Further, this relationship appeared among donkeys with high body condition but not among those with low body condition. Time lags of 1–4 days introduced between IgA and FEC measurements in repeatedly sampled donkeys did not yield correlations, suggesting that IgA and FEC roughly tracked one another without much delay in the wet field season. Such a direct IgA-FEC relationship did not appear for zebras in either the wet or dry field season, possibly due to higher interindividual variation in body condition among the free-roaming zebras than in the donkeys. However, Grevy's zebras had higher overall levels of IgA than either plains zebras or donkeys, potentially associated with their reportedly lower FECs at the population level. Our results suggest that equids may mount an IgA response to nematode egg production when the host is in good condition and that equid species may differ in baseline levels of mucosal IgA.

Sakhrani, Varun V., Eduardo Hilario, Bethany G. Caulkins, Mary E. Hatcher-Skeers, Li Fan, Michael F. Dunn, and Leonard J. Mueller. “Backbone Assignments and Conformational Dynamics in the S. typhimurium Tryptophan Synthase α-Subunit from Solution-State NMR.” Journal of Biomolecular NMR, vol. 74, 2020, pp. 341-354.

Abstract: Backbone assignments for the isolated α-subunit of Salmonella typhimurium tryptophan synthase (TS) are reported based on triple resonance solution-state NMR experiments on a uniformly 2H,13C,15N-labeled sample. From the backbone chemical shifts, secondary structure and random coil index order parameters (RCI-S2) are predicted. Titration with the 3-indole-d-glycerol 3′-phosphate analog, N-(4′-trifluoromethoxybenzenesulfonyl)-2-aminoethyl phosphate (F9), leads to chemical shift perturbations indicative of conformational changes from which an estimate of the dissociation constant is obtained. Comparisons of the backbone chemical-shifts, RCI-S2 values, and site-specific relaxation times with and without F9 reveal allosteric changes including modulation in secondary structures and loop rigidity induced upon ligand binding. A comparison is made to the X-ray crystal structure of the α-subunit in the full TS αββα bi-enzyme complex and to two new X-ray crystal structures of the isolated TS α-subunit reported in this work.

Dalla Benetta, Elena, Igor Antoshechkin, Ting Yang, Hoa Quang My Nguyen, Patrick M. Ferree, and Omar S. Akbari. “Genome Elimination Mediated by Gene Expression from a Selfish Chromosome.” Science Advances, vol. 6, no. 14, 2020, eaaz9808.

Abstract: Numerous plants and animals harbor selfish B chromosomes that “drive” or transmit themselves at super-Mendelian frequencies, despite long-term fitness costs to the organism. Currently, it is unknown how B chromosome drive is mediated, and whether B-gene expression plays a role. We used modern sequencing technologies to analyze the fine-scale sequence composition and expression of paternal sex ratio (PSR), a B chromosome in the jewel wasp Nasonia vitripennis. PSR causes female-to-male conversion by destroying the sperm’s hereditary material in young embryos to drive. Using RNA interference, we demonstrate that testis-specific expression of a PSR-linked gene, named haploidizer, facilitates this genome elimination–and–sex conversion effect. haploidizer encodes a putative protein with a DNA binding domain, suggesting a functional link with the sperm-derived chromatin.

Chaverra-Rodgriguez, D., E. Dalla Benetta, C.C. Heu, J.L. Rasgon, P.M. Ferree, and Omar S. Akbari. “Germline Mutagenesis of Nasonia vitripennis through Ovarian Delivery of CRISPR-Cas9 Ribonucleoprotein.” Insect Molecular Biology, vol. 29, issue 6, 2020, pp. 569-577.

Abstract: CRISPR/Cas9 gene editing is a powerful technology to study the genetics of rising model organisms, such as the jewel wasp Nasonia vitripennis. However, current methods involving embryonic microinjection of CRISPR reagents are challenging. Delivery of Cas9 ribonucleoprotein into female ovaries is an alternative that has only been explored in a small handful of insects, such as mosquitoes, whiteflies and beetles. Here, we developed a simple protocol for germline gene editing by injecting Cas9 ribonucleoprotein in adult N. vitripennis females using either ReMOT control (Receptor‐Mediated Ovary Transduction of Cargo) or BAPC (Branched Amphiphilic Peptide Capsules) as ovary delivery methods. For ReMOT Control we used the Drosophila melanogaster‐derived peptide ‘P2C’ fused to EGFP to visualize the ovary delivery, and fused to Cas9 protein for gene editing of the cinnabar gene using saponin as an endosomal escape reagent. For BAPC we optimized the concentrations of protein, sgRNA and the transfection reagent. We demonstrate delivery of protein cargo such as EGFP and Cas9 into developing oocytes via P2C peptide and BAPC. Additionally, somatic and germline gene editing were demonstrated. This approach will greatly facilitate CRISPR‐applied genetic manipulation in this and other rising model organisms.

Eastin, Katherine J., Austin P. Huang, and Patrick M. Ferree. “A Novel Pattern of Germ Cell Divisions in the Production of Hymenopteran Insect Eggs.” Biology Letters, vol. 16, issue 5, 2020, 20200137.

Abstract: Egg development is a defining process of reproduction in higher eukaryotes. In the fruit fly, Drosophila melanogaster, this process begins with four mitotic divisions starting from a single germ cell, producing a cyst of 16 cystocytes; one of these cells will become the oocyte and the others supporting nurse cells. These mitotic divisions are exceptional because cytokinesis is incomplete, resulting in the formation of cytoplasmic bridges known as ring canals that interconnect the cystocytes. This organization allows all cystocytes to divide synchronously during each mitotic round, resulting in a final, power-of-2 number of germ cells. Given that numerous insects obey this power-of-2 rule, we investigated if strict cell doubling is a universal, underlying cause. Using confocal microscopy, we found striking departures from this paradigm in three different power-of-2 insects belonging to the Apocrita suborder (ants, bees and wasps). In these insects, the earliest-formed cystocytes cease to divide during the latter mitotic cycles while their descendants undergo further division, thereby producing a ‘radial’ direction of division activity. Such cystocyte division patterns that depart from strict cell doubling may be ‘fine-tuned’ in order to maintain a final, power-of-2 germ cell number.

Rangaswami, Xenia L.*, Gordon T. Ober, and Sarah E. Gilman. “D-lactate Production in the Acorn Barnacle Balanus Glandula (Darwin, 1854) (Cirripedia: Balanidae) Under Emersion Stress.” Journal of Crustacean Biology, vol. 40, issue 6, 2020, pp. 739-735.

Abstract: Anaerobic metabolism is an important response to stress in many organisms. Intertidal species often face heat stress during low tide. Balanus glandula (Darwin, 1854) is a high-shore intertidal barnacle common to the Pacific that experiences prolonged low-tide air exposure. It is not known whether B. glandula uses anaerobic metabolism during emersion, or if its use varies by latitude. We measured low tide D-lactate production in two US west coast populations of B. glandula separated by 14 degrees of latitude. We exposed barnacles to seven low-tide air temperatures (10, 15, 20, 25, 30, 35, and 38 °C) for which aerobic respiration has been previously measured. Our northern population of B. glandula increased D-lactate production at high air temperatures where aerobic metabolic depression is known to occur, indicating sublethal stress. In contrast, our southern population showed little increase in D-lactate over the same temperature range, coincident with high aerobic respiration across those temperatures. In a second experiment, we quantified D-lactate at 1, 2, and 6 hours post-emersion for northern B. glandula exposed to either a 10 or 38 °C low tide, to measure their potential lactate usage. While D-lactate was elevated at 38 °C compared to the 10 °C control immediately following low tide exposure, it dropped to control levels, and was likely excreted, within 1 hour of re-immersion. Our results suggest that the low latitude population of B. glandula may be more resilient to climate change than its high latitude counterpart in the absence of adaptation, which has strong implications for species distribution.

Sakhrani, Varun V., Eduardo Hilario, Bethany G. Caulkins, Mary E. Hatcher-Skeers, Li Fan, Michael F. Dunn, and Leonard J. Mueller. “Backbone Assignments and Conformational Dynamics in the S. typhimurium Tryptophan Synthase α-Subunit from Solution-State NMR.” Journal of Biomolecular NMR, vol. 74, 2020, pp. 341-354.

Abstract: Backbone assignments for the isolated α-subunit of Salmonella typhimurium tryptophan synthase (TS) are reported based on triple resonance solution-state NMR experiments on a uniformly 2H,13C,15N-labeled sample. From the backbone chemical shifts, secondary structure and random coil index order parameters (RCI-S2) are predicted. Titration with the 3-indole-d-glycerol 3′-phosphate analog, N-(4′-trifluoromethoxybenzenesulfonyl)-2-aminoethyl phosphate (F9), leads to chemical shift perturbations indicative of conformational changes from which an estimate of the dissociation constant is obtained. Comparisons of the backbone chemical-shifts, RCI-S2 values, and site-specific relaxation times with and without F9 reveal allosteric changes including modulation in secondary structures and loop rigidity induced upon ligand binding. A comparison is made to the X-ray crystal structure of the α-subunit in the full TS αββα bi-enzyme complex and to two new X-ray crystal structures of the isolated TS α-subunit reported in this work.

Friedman, Eric, John Friedman, Simon Johnson, and Adam Landsberg. “Transitioning Out of the Coronavirus Lockdown: A Framework for Evaluating Zone-Based Social Distancing." Frontiers in Public Health, vol. 8, 2020, article 266.

Abstract: In the face of elevated pandemic risk, canonical epidemiological models imply the need for extreme social distancing over a prolonged period. Alternatively, people could be organized into zones, with more interactions inside their zone than across zones. Zones can deliver significantly lower infection rates, with less social distancing, particularly if combined with simple quarantine rules and contact tracing. This paper provides a framework for understanding and evaluating the implications of zones, quarantines, and other complementary policies.

*LaBerge, Nicholas, Aria Chaderjian*, Victor Ginelli*, Margrethe Jebsen*, and Adam Landsberg. “Modeling Cultural Dissemination and Divergence Between Rural and Urban Regions.” Journal of Artificial Societies and Social Simulation, vol. 23, issue 4, 2020, 3.

Abstract: The process by which beliefs, opinions, and other individual, socially malleable attributes spread across a society, known as "cultural dissemination," is a broadly recognized concept among sociologists and political scientists. Yet fundamental aspects of how this process can ultimately lead to cultural divergences between rural and urban segments of society are currently poorly understood. This article uses an agent-based model to isolate and analyze one very basic yet essential facet of this issue, namely, the question of how the intrinsic differences in urban and rural population densities influence the levels of cultural homogeneity/heterogeneity that emerge within each region. Because urban and rural cultures do not develop in isolation from one another, the dynamical interplay between the two is of particular import in their evolution. It is found that, in urban areas, the relatively high number of local neighbors with whom one can interact tends to promote cultural homogeneity in both urban and rural regions. Moreover, and rather surprisingly, the higher frequency of potential interactions with neighbors within urban regions promotes homogeneity in urban regions but tends to drive rural regions towards greater levels of heterogeneity.

*Thompson, Arianna S., Susanna E. Barrett*, Aurora G. Weiden*, Ananya Venkatesh*, Madison K. C. Seto*, Simone Z. P. Gottlieb*, and Aaron M. Leconte. “Accurate and Efficient One-Pot Reverse Transcription and Amplification of 2' Fluoro-Modified Nucleic Acids by Commercial DNA Polymerases.” Biochemistry, vol. 59, issue 31, 2020, pp. 2833-2841.

Abstract: DNA is a foundational tool in biotechnology and synthetic biology but is limited by sensitivity to DNA-modifying enzymes. Recently, researchers have identified DNA polymerases that can enzymatically synthesize long oligonucleotides of modified DNA (M-DNA) that are resistant to DNA-modifying enzymes. Most applications require M-DNA to be reverse transcribed, typically using a RNA reverse transcriptase, back into natural DNA for sequence analysis or further manipulation. Here, we tested commercially available DNA-dependent DNA polymerases for their ability to reverse transcribe and amplify M-DNA in a one-pot reaction. Three of the six polymerases chosen (Phusion, Q5, and Deep Vent) could reverse transcribe and amplify synthetic 2'F M-DNA in a single reaction with <5 × 10-3 error per base pair. We further used Q5 DNA polymerase to reverse transcribe and amplify M-DNA synthesized by two candidate M-DNA polymerases (SFP1 and SFM4-6), allowing for quantification of the frequency, types, and locations of errors made during M-DNA synthesis. From these studies, we identify SFP1 as one of the most accurate M-DNA polymerases identified to date. Collectively, these studies establish a simple, robust method for the conversion of 2'F M-DNA to DNA in <1 h using commercially available materials, significantly improving the ease of use of M-DNA.

External Grant: Leconte, Aaron, Co-Investigator. “Moving the Dial: A Network for Systemic Change.” Cottrell Scholar Collaborative Project, Research Corporation for Science Advancement, 2020-2022, $25,000.

Abstract: Improvements in diversity, equity, and inclusion (DEI) in higher education have been incremental and minor gains are often touted as major improvements. This Collaborative will fuel substantial and lasting change by providing implementable strategies at a variety of levels, supporting networks of change agents, and laying the ground work for measurable implementations of best practices.

*Hsu, Alexander, and Sarah Marzen. “Time Cells Might be Optimized for Predictive Capacity, but Not for Redundancy Reduction or Memory Capacity.” Physical Review E, vol. 102, issue 6, 2020, 062404.

Abstract: Recently, researchers have found time cells in the hippocampus that appear to contain information about the timing of past events. Some researchers have argued that time cells are taking a Laplace transform of their input in order to reconstruct the past stimulus. We argue that stimulus prediction, not stimulus reconstruction or redundancy reduction, is in better agreement with observed responses of time cells. In the process, we introduce new analyses of nonlinear, continuous-time reservoirs that model these time cells.

Marzen, Sarah, and James P. Crutchfield. “Prediction and Dissipation in Nonequilibrium Molecular Sensors: Conditionally Markovian Channels Driven by Memoryful Environments.” Bulletin of Mathematical Biology, vol. 82, 2020, article 25.

Abstract: Biological sensors must often predict their input while operating under metabolic constraints. However, determining whether or not a particular sensor is evolved or designed to be accurate and efficient is challenging. This arises partly from the functional constraints being at cross purposes and partly since quantifying the prediction performance of even in silico sensors can require prohibitively long simulations, especially when highly complex environments drive sensors out of equilibrium. To circumvent these difficulties, we develop new expressions for the prediction accuracy and thermodynamic costs of the broad class of conditionally Markovian sensors subject to complex, correlated (unifilar hidden semi-Markov) environmental inputs in nonequilibrium steady state. Predictive metrics include the instantaneous memory and the total predictable information (the mutual information between present sensor state and input future), while dissipation metrics include power extracted from the environment and the nonpredictive information rate. Success in deriving these formulae relies on identifying the environment's causal states, the input's minimal sufficient statistics for prediction. Using these formulae, we study large random channels and the simplest nontrivial biological sensor model--that of a Hill molecule, characterized by the number of ligands that bind simultaneously—the sensor's cooperativity. We find that the seemingly impoverished Hill molecule can capture an order of magnitude more predictable information than large random channels.

Razo-Mejia, Manuel, Sarah Marzen, Griffin Chure, Rachel Taubman*, Muir Morrison, and Rob Phillips. “First-Principles Prediction of the Information Processing Capacity of a Simple Genetic Circuit.” Physical Review E, vol. 102, 2020, 022404.

Abstract: Given the stochastic nature of gene expression, genetically identical cells exposed to the same environmental inputs will produce different outputs. This heterogeneity has been hypothesized to have consequences for how cells are able to survive in changing environments. Recent work has explored the use of information theory as a framework to understand the accuracy with which cells can ascertain the state of their surroundings. Yet the predictive power of these approaches is limited and has not been rigorously tested using precision measurements. To that end, we generate a minimal model for a simple genetic circuit in which all parameter values for the model come from independently published data sets. We then predict the information processing capacity of the genetic circuit for a suite of biophysical parameters such as protein copy number and protein-DNA affinity. We compare these parameter-free predictions with an experimental determination of protein expression distributions and the resulting information processing capacity of E. coli cells. We find that our minimal model captures the scaling of the cell-to-cell variability in the data and the inferred information processing capacity of our simple genetic circuit up to a systematic deviation.

*Uppal, Abhinuv, Vanessa Ferdinand, and Sarah Marzen. “Inferring an Observer's Strategy in Sequence Learning Experiments.” Entropy, vol. 22, issue 8, 2020, 896.

Abstract: Cognitive systems exhibit astounding prediction capabilities that allow them to reap rewards from regularities in their environment. How do organisms predict environmental input and how well do they do it? As a prerequisite to answering that question, we first address the limits on prediction strategy inference, given a series of inputs and predictions from an observer. We study the special case of Bayesian observers, allowing for a probability that the observer randomly ignores data when building her model. We demonstrate that an observer's prediction model can be correctly inferred for binary stimuli generated from a finite-order Markov model. However, we can not necessarily infer the model's parameter values unless we have access to several "clones" of the observer. As stimuli become increasingly complicated, correct inference requires exponentially more data points, computational power, and computational time. These factors place a practical limit on how well we are able to infer an observer's prediction strategy in an experimental or observational setting.

Marx, Andrew, and Donald McFarlane. “Combining Unmanned Aerial Systems and Satellite Data to Monitor Phenological Changes in Tropical Forests: A Case Study from Costa Rica.” Case Studies in the Environment, vol. 3, issue 1, 2019, pp. 1-11. [Published in 2019 but not available until 2020]

Abstract: The migration of vegetation under the influence of climate change is of great interest to ecologists, but can be difficult to quantify—especially in less accessible landscapes. Monitoring land cover change through remote sensing has become the best solution, especially with the use of unmanned aerial systems (UASs; drones) as low-cost remote sensing platforms are able to collect data at high spatial and spectral resolutions. Unfortunately, in the context of climate change studies, the lack of comparative UAS data sets over decadal timescales has been limiting. Here, we describe a technique for the integration of historical, low spatial resolution satellite-based Normalized Difference Vegetation Index (NDVI) data with short-term high-resolution multispectral UAS data to track the vegetation changes in a Costa Rican rainforest over a 33-year time frame. The study reveals the transition of a mixed forest from strongly deciduous to weakly deciduous phenology in the Hacienda Barú National Wildlife Refuge (HBNWR), southwestern Costa Rica. This case study presents an approach for researchers and forest managers to study and track vegetation changes over time in locations that lack detailed historical vegetation data. Vegetation migration due to climate change is not well documented and difficult to monitor, especially in remote or inaccessible locations. This case study presents researchers, students, and forest managers an approach for leveraging freely available satellite imagery and UASs to track these changes over time.

Buza, Gergely, John Milton, Laszlo Bencsik, and Tamas Insperger. “Establishing Metrics and Control Laws for the Learning Process: Ball and Beam Balancing.” Biological Cybernetics, vol. 114, 2020, pp. 83-93.

Abstract: Understanding how dexterity improves with practice is a fundamental challenge of motor control and neurorehabilitation. Here we investigate a ball and beam implementation of a dexterity puzzle in which subjects stabilize a ball at the mid-point of a beam by manipulating the angular position of the beam. Stabilizability analysis of different biomechanical models for the ball and beam task with time-delayed proportional-derivative feedback identified the angular position of the beam as the manipulated variable. Consequently, we monitored the changes in the dynamics with learning by measuring changes in the control parameters. Two types of stable motion are possible: node type (nonoscillatory) and spiral type (oscillatory). Both types of motion are observed experimentally and correspond to well-defined regions in the parameter space of the control gains. With practice the control gains for each subject move close to or on the portion of the boundary which separates the node-type and spiral-type solutions and which is associated with the rightmost characteristic exponent of smallest real part. These observations suggest that with learning the control gains for ball and beam balancing change in such a way that minimizes overshoot and the settling time. This study provides an example of how mathematical analysis together with careful experimental observations can shed light onto the early stages of skill acquisition. Since the difficulty of this task depends on the length of the beam, ball and beam balancing tasks may be useful for the rehabilitation of children with dyspraxia and those recovering from a stroke.

Nishikawa, Kiisa, Samrat Dutta, Michael DuVall, Brent Nelson, Matthew J. Gage, and Jenna A. Monroy. “Calcium-Dependent Titin-Thin Filament Interactions in Muscle: Observations and Theory.” Journal of Muscle Research and Cell Motility, vol. 41, 2020, pp. 125-139.

Abstract: Gaps in our understanding of muscle mechanics demonstrate that the current model is incomplete. Increasingly, it appears that a role for titin in active muscle contraction might help to fill these gaps. While such a role for titin is increasingly accepted, the underlying molecular mechanisms remain unclear. The goals of this paper are to review recent studies demonstrating Ca2+-dependent interactions between N2A titin and actin in vitro, to explore theoretical predictions of muscle behavior based on this interaction, and to review experimental data related to the predictions. In a recent study, we demonstrated that Ca2+ increases the association constant between N2A titin and F-actin; that Ca2+ increases rupture forces between N2A titin and F-actin; and that Ca2+ and N2A titin reduce sliding velocity of F-actin and reconstituted thin filaments in motility assays. Preliminary data support a role for Ig83, but other Ig domains in the N2A region may also be involved. Two mechanical consequences are inescapable if N2A titin binds to thin filaments in active muscle sarcomeres: (1) the length of titin’s freely extensible I-band should decrease upon muscle activation; and (2) binding between N2A titin and thin filaments should increase titin stiffness in active muscle. Experimental observations demonstrate that these properties characterize wild type muscles, but not muscles from mdm mice with a small deletion in N2A titin, including part of Ig83. Given the new in vitro evidence for Ca2+-dependent binding between N2A titin and actin, it is time for skepticism to give way to further investigation.

Tahir, Uzma, Jenna A. Monroy, Nicole A. Rice, and Kiisa C. Nishikawa. “Effects of Titin Mutation on Force Enhancement and Force Depression in Mouse Soleus Muscles.” Journal of Experimental Biology, vol. 223, 2020, 197038.

Abstract: The active isometric force produced by muscles varies with muscle length in accordance with the force–length relationship. Compared with isometric contractions at the same final length, force increases after active lengthening (force enhancement) and decreases after active shortening (force depression). In addition to cross-bridges, titin has been suggested to contribute to force enhancement and depression. Although titin is too compliant in passive muscles to contribute to active tension at short sarcomere lengths on the ascending limb and plateau of the force–length relationship, recent evidence suggests that activation increases titin stiffness. To test the hypothesis that titin plays a role in force enhancement and depression, we investigated isovelocity stretching and shortening in active and passive wild-type and mdm (muscular dystrophy with myositis) soleus muscles. Skeletal muscles from mdm mice have a small deletion in the N2A region of titin and show no increase in titin stiffness during active stretch. We found that: (1) force enhancement and depression were reduced in mdm soleus compared with wild-type muscles relative to passive force after stretch or shortening to the same final length; (2) force enhancement and force depression increased with amplitude of stretch across all activation levels in wild-type muscles; and (3) maximum shortening velocity of wild-type and mdm muscles estimated from isovelocity experiments was similar, although active stress was reduced in mdm compared with wild-type muscles. The results of this study suggest a role for titin in force enhancement and depression, which contribute importantly to muscle force during natural movements.

Fahlman, Bradley D., Kathleen L. Purvis-Roberts, John S. Kirk, Patrick L. Daubenmire, and Resa Kelly,. Chemistry in Context: Applying Chemistry to Society, 10th Edition. McGraw-Hill, 2020.

Abstract: Chemistry in Context is a successful, issues-based curriculum developed by the American Chemical Society for non-science majors at the college level.

Fu, Franck, Kathleen L. Purvis-Roberts, and Branwen Williams. "Impact of Covid-19 Pandemic Lockdown on PM, Ozone, SO2, NO2 and CO in 20 Major Cities around the World.” Atmosphere, vol. 11, 2020, 1189.

Abstract: In order to fight against the spread of COVID-19, the most hard-hit countries in the spring of 2020 implemented different lockdown strategies. To assess the impact of the COVID-19 pandemic lockdown on air quality worldwide, Air Quality Index (AQI) data was used to estimate the change in air quality in 20 major cities on six continents. Our results show significant declines of AQI in NO2, SO2, CO, PM2.5 and PM10 in most cities, mainly due to the reduction of transportation, industry and commercial activities during lockdown. This work shows the reduction of primary pollutants, especially NO2, is mainly due to lockdown policies. However, preexisting local environmental policy regulations also contributed to declining NO2, SO2 and PM2.5 emissions, especially in Asian countries. In addition, higher rainfall during the lockdown period could cause decline of PM2.5, especially in Johannesburg. By contrast, the changes of AQI in ground-level O3 were not significant in most of cities, as meteorological variability and ratio of VOC/NOx are key factors in ground-level O3 formation.

Sanii, Babak. “Creating Augmented Reality USDZ Files to Visualize 3D Objects on Student Phones in the Classroom.” Journal of Chemical Education, vol. 97, issue 1, 2020, pp. 253-257.

Abstract: Augmented reality (AR) is a means of superimposing artificial 3D objects over the real world via a mobile device. An AR standard file format has been recently implemented on mobile devices that current students commonly own. Here we describe three relatively nontechnical methods to produce 3D AR objects for chemistry courses and demonstrate their use as both quick in-lecture activities and as part of an extended laboratory. Preliminary in-course student assessments (32 responses out of 42 students) indicated that this technology was successfully implemented on students’ devices (100%), that it generated interest/excitement (78%), and that it helped students visualize 3D objects (81%).

Wan, Kirsty Y., Sylvia K. Hürlimann, Aidan M. Fenix, Rebecca M. McGillivary, Tatyana Makushok, Evan Burns, Janet Y. Sheung, and Wallace F. Marshall. “Reorganization of Complex Ciliary Flows around Regenerating Stentor coeruleus.” Philosophical Transactions of the Royal Society B, vol. 375, issue 1792, 2020, 20190167.

Abstract: The phenomenon of ciliary coordination has garnered increasing attention in recent decades and multiple theories have been proposed to explain its occurrence in different biological systems. While hydrodynamic interactions are thought to dictate the large-scale coordinated activity of epithelial cilia for fluid transport, it is rather basal coupling that accounts for synchronous swimming gaits in model microeukaryotes such as Chlamydomonas. Unicellular ciliates present a fascinating yet understudied context in which coordination is found to persist in ciliary arrays positioned across millimetre scales on the same cell. Here, we focus on the ciliate Stentor coeruleus, chosen for its large size, complex ciliary organization, and capacity for cellular regeneration. These large protists exhibit ciliary differentiation between cortical rows of short body cilia used for swimming, and an anterior ring of longer, fused cilia called the membranellar band (MB). The oral cilia in the MB beat metachronously to produce strong feeding currents. Remarkably, upon injury, the MB can be shed and regenerated de novo. Here, we follow and track this developmental sequence in its entirety to elucidate the emergence of coordinated ciliary beating: from band formation, elongation, curling and final migration towards the cell anterior. We reveal a complex interplay between hydrodynamics and ciliary restructuring in Stentor, and highlight for the first time the importance of a ring-like topology for achieving long-range metachronism in ciliated structures.

Wu, Mei, Gang Feng, Buyu Zhang, Kaikun Xu, Zhen Wang, Sen Cheng, Cheng Chang, Aditi Vyas, Zhaohua Tang, and Xiaoyun Liu. “Phosphoproteomics Reveals Novel Targets and Phosphoprotein Networks in Cell Cycle Mediated by Dsk1 Kinase.” Journal of Proteome Research, vol. 19, issue 4, 2020, pp. 1776-1787.

Abstract: As the ortholog of human SR protein kinase 1 in fission yeast Schizosaccharomyces pombe, Dsk1 specifically phosphorylates SR proteins (serine/arginine-rich proteins) and promotes splicing of nonconsensus introns. The SRPK (SR protein-specific kinase) family performs highly conserved functions in eukaryotic cells including cell proliferation, differentiation, development, and apoptosis. Although Dsk1 was originally identified as a mitotic regulator, its specific targets involved in cell cycle have yet been unexplored. In this study, using a phosphoproteomics approach, we examined differential protein phosphorylation between wild-type cells and dsk1-deletion mutants. We found reduced phosphorylation of 149 peptides corresponding to 133 proteins in the dsk1-null cells. These proteins are involved in various cellular processes, including cytoskeleton organization and signal transduction, and specifically enriched in multiple steps of cell cycle control. Further, targeted MS analyses and in vitro biochemical assays established Cdr2 protein kinase and kinesin motor Klp9 as novel substrates of Dsk1, which function in cell size control for mitotic entry and in chromosome segregation for mitotic exit, respectively. The phosphoprotein networks mediated by Dsk1 reveal, for the first time, the molecular links connecting Dsk1 to mitotic phase transition, sister-chromatid segregation, and cytokinesis, providing further evidence of Dsk1’s diverse influence on cell cycle progression and regulation.

Funk, Jennifer L., Ingrid M. Parker, Erik T. Aschehoug, Wayne Dawson, S. Luke Flory, Carla M. D'Antonio, Virginia Matzek, Diane M. Thomson, and Justin Valliere. “Keys to Enhancing the Value of Invasion Ecology Research for Management.” Biological Invasions, vol. 22, 2020, pp. 2431-2445.

Abstract: Invasion ecology has grown to include scientists with diverse skill sets who focus on a range of taxa and biomes. These researchers have the capacity to contribute to practical management solutions while also answering fundamental biological questions; however, scientific endeavors often fail to meet the perceived needs of practitioners involved in on-the ground invasive plant management. One way that researchers have sought to bridge the gap between research and practice is by surveying managers to identify areas of study that are underexplored in invasion ecology. In this paper, we build on these efforts by reviewing the current state of knowledge and suggesting new directions for research in seven areas of plant invasion ecology that are highly relevant to management: seedbanks, dispersal and spread, life history, impacts, climate change, distribution, and succession. These topics were previously identified as urgent research priorities by land managers and are underrepresented in the invasion ecology literature. In addition to highlighting key knowledge gaps for these seven areas of research, we propose steps that academics can take to cultivate academic-practitioner relationships and remove barriers to conducting management-focused research, such as co-producing research questions with managers, addressing issues of working at management-appropriate spatial and temporal scales, and considering non-traditional funding and labor sources for long-term monitoring. Greater communication and collaborative selection of basic research questions will ensure that the goals of management and invasive species research remain aligned.

Thomson, Diane, Adin D. Bonapart*, Rachel A. King*, Emily L. Schultz*, and Charlotte R. Startin*. “Long-Term Monitoring of a Highly Invaded Annual Grassland Community through Drought, Before and After an Unintentional Fire.” Journal of Vegetation Science, vol. 31, issue 2, 2020, pp. 307-318.

Abstract: Questions: 1) How did seedling numbers and species composition change in the first year after a wildfire during drought, relative to pre‐fire variation? 2) Has the community returned to pre‐fire composition after five years? 3) Has the degree of dominance by exotic annual grasses changed? 4) Is there any evidence that drought conditions affected community cover, before or after fire? Location: Exotic‐dominated annual grassland in southern California, U.S.A. Methods: We monitored community cover and native annual forb densities for four years before and four (cover) to five (densities) years after an unintentional fire (fall 2013) coinciding with the spring 2012‐2019 California drought. We also measured seedling emergence both before and during the first year post‐fire. We assessed post‐fire changes in cover and density relative to pre‐fire variation, and tested correlations between community cover and annual rainfall measures. Results: Seedling emergence declined strongly after fire for exotic grasses, but remained stable for exotic forbs. Seedling densities of the most common native forbs declined, but several previously‐rare natives increased. Community cover initially shifted towards the exotic forbs Erodium spp., then returned to higher exotic grass densities. Yet the previously‐dominant Bromus diandrus declined steeply, even as other exotic grasses and some native forbs increased. Up to five years after fire, relative cover and abundance of the most common exotic and native species still differed from pre‐fire composition. Common species were uncorrelated with annual precipitation, but several may have responded to shorter growing seasons. Conclusions: Immediate post‐fire conditions favored exotic and native forbs over grasses, as predicted. Yet in contrast to many previous studies, the community did not return quickly to pre‐fire composition but showed persistent changes that favored neither natives nor exotics. Our results suggest post‐fire recovery in this habitat may be contingent on abiotic conditions, with drought one potential explanation for changes.

Thomson, Diane M. and Maureen L. Page. 2020. “The Importance of Competition Between Insect Pollinators in the Anthropocene.” Current Opinion in Insect Science, vol. 36, 2020, pp. 55-62.

Abstract: Resource competition likely plays an important role in some insect pollinator declines and in shaping effects of environmental change on pollination services. Past research supports that competition for floral resources affects bee foragers, but mostly with observational evidence and rarely linking foraging with population change. An increasing number of studies ask whether resources limit pollinator populations, using field measurements of reproductive success, time series and models. Findings generally support positive effects of floral resources, but also highlight the potential importance of nest site availability and parasitism. In parallel, recent experiments strengthen evidence that competition reduces access to floral resources. Developing common currencies for quantifying floral resources and integrating analyses of multiple limiting factors will further strengthen our understanding of competitive interactions and their effects in the Anthropocene.

Chang, Preston T., Krithika Rao, Lauren O. Longo, Elisabeth S. Lawton, Georgia Scherer, and Ethan B. Van Arnam. “Thiopeptide Defense by an Ant's Bacterial Symbiont.” Journal of Natural Products, vol. 83, issue 3, 2020, pp. 725-729.

Abstract: Fungus-growing ants and their microbial symbionts have emerged as a model system for understanding antibiotic deployment in an ecological context. Here we establish that bacterial symbionts of the ant Trachymyrmex septentrionalis antagonize their most likely competitors, other strains of ant-associated bacteria, using the thiopeptide antibiotic GE37468. Genomic analysis suggests that these symbionts acquired the GE37468 gene cluster from soil bacteria. This antibiotic, with known activity against human pathogens, was previously identified in a biochemical screen but had no known ecological role. GE37468's host-associated defense role in this insect niche intriguingly parallels the function of similar thiopeptides in the human microbiome.

King, Bradley H.*, Michelle L. Wang*, Kate A. Jesse*, Guneet Kaur*, Brianna Tran*, Ryan Walser-Kuntz, Robert G. Iafe, and Anna G. Wenzel. “Silver-Catalyzed, N-Formation of Amines Using Glycol Ethers.” Journal of Organic Chemistry, vol. 85, issue 20, 2020, pp. 13256-13263.

Abstract: A silver-catalyzed protocol was found to afford the N-formylation of amines in moderate-to-good yields. Ethylene glycol-derived, oligomeric ethers were found to function as the formylating agent, with 1,4-dioxane affording the best results. This reaction does not require the use of stoichiometric activating reagents, and avoids the use of explosive reagents or toxic gases, such as CO, as the C1 synthon. Mechanistic studies indicate a single-electron transfer-based pathway. This work highlights the ability of silver to participate in unexpected reaction pathways.

Wenzel, Anna G., Steven Casper, Cooper J. Galvin*, and Grace E. Beck*. "Science and Business of Medicinal Chemistry: a "Bench-to-Bedside" Course for Non-Majors." Journal of  Chemical Education, vol. 97, issue 2, 2020, pp. 414-420.

Abstract: "The Science and Business of Medicinal Chemistry" was developed to provide nonscience majors with an in-depth perspective into the pharmaceutical industry. As a lecture course with a laboratory component, topics discussed include an introduction to the basic concepts of medicinal chemistry, such as drug discovery, development, and commercialization; a discussion of chemical bonding and the organic functional groups found in drug molecules; and an examination of the physiochemical properties related to drug action. Student scientific literacy of the public policy debates surrounding the pharmaceutical industry and the commercialization of science was enhanced through class discussions and group-based projects. The details of this course, its implementation, and assessment are discussed.

Choy*, Emma, Kelly Watanabe*, Branwen Williams, Robert Stone, Peter Etnoyer, Ellen Druffel, Thomas Lorenson, and Mary Knaak. “Understanding Growth and Age of Red Tree Corals (Primnoa pacifica) in the North Pacific Ocean.” PLOS One, vol. 15, issue 12, 2020, e0241692.

Abstract: Massive, long-lived deep-sea red tree corals (Primnoa pacifica) form a solid, layered axis comprised of calcite and gorgonin skeleton. They are abundant on the outer continental shelf and upper slope of the Northeast Pacific, providing habitat for fish and invertebrates. Yet, their large size and arborescent morphology makes them susceptible to disturbance from fishing activities. A better understanding of their growth patterns will facilitate in-situ estimates of population age structure and biomass. Here, we evaluated relationships between ages, growth rates, gross morphological characteristics, and banding patterns in 11 colonies collected from depths of ~141–335 m off the Alaskan coast. These corals ranged in age from 12 to 80 years old. They grew faster radially (0.33–0.74 mm year-1) and axially (2.41–6.39 cm year-1) than in previously measured older colonies, suggesting that growth in P. pacifica declines slowly with age, and that basal diameter and axial height eventually plateau. However, since coral morphology correlated with age in younger colonies (< century), we developed an in-situ age estimation technique for corals from the Northeast Pacific Ocean providing a non-invasive method for evaluating coral age without removing colonies from the population. Furthermore, we determined that annual bands provided the most accurate means for determining coral age in live-collected corals, relative to radiometric dating. Taken together, this work provides insight into P. pacifica growth patterns to inform coastal managers about the demographics of this ecologically important species. With this new ability to estimate the age of red tree corals in-situ, we can readily determine the age-class structure and consequently, the maturity status of thickets, using non-invasive video survey techniques when coupled with mensuration systems such as lasers or stereo-cameras. Enhanced surveys could identify which populations are most vulnerable to disturbance from human activities, and which should be highlighted for protection.

Fu, Franck, Kathleen L. Purvis-Roberts, and Branwen Williams. "Impact of Covid-19 Pandemic Lockdown on PM, Ozone, SO2, NO2 and CO in 20 Major Cities around the World.” Atmosphere, vol. 11, 2020, 1189.

Abstract: In order to fight against the spread of COVID-19, the most hard-hit countries in the spring of 2020 implemented different lockdown strategies. To assess the impact of the COVID-19 pandemic lockdown on air quality worldwide, Air Quality Index (AQI) data was used to estimate the change in air quality in 20 major cities on six continents. Our results show significant declines of AQI in NO2, SO2, CO, PM2.5 and PM10 in most cities, mainly due to the reduction of transportation, industry and commercial activities during lockdown. This work shows the reduction of primary pollutants, especially NO2, is mainly due to lockdown policies. However, preexisting local environmental policy regulations also contributed to declining NO2, SO2 and PM2.5 emissions, especially in Asian countries. In addition, higher rainfall during the lockdown period could cause decline of PM2.5, especially in Johannesburg. By contrast, the changes of AQI in ground-level O3 were not significant in most of cities, as meteorological variability and ratio of VOC/NOx are key factors in ground-level O3 formation.

Hargan, Kathryn E., Branwen Williams, Bunlung Nuangsaeng, Sarawut Siriwong, Pisut Tassawad, Chatdanai Chaiharn, Brian McAdoo, and Marc Los Huertos. “Understanding the Fate of Shrimp Aquaculture Effluent in a Mangrove Ecosystem: Aiding Management for Coastal Conservation.” Journal of Applied Ecology, vol. 57, issue 4, 2020, pp. 754-765.

Abstract: Areas dedicated to shrimp aquaculture have increased dramatically over the last 50 years. Resultant land‐use changes directly threaten the extent of mangroves and yield conflicts on the discharge location of aquaculture effluent. Khung Krabaen Bay (KBB), Thailand, is reforesting mangroves while increasing the efficiency of shrimp aquaculture for local farmers. In this coupled shrimp farm–mangrove system, effective management requires understanding the fate of aquaculture organic matter (OM) in the coastal environment. We examined carbon and nitrogen stable isotope ratios (δ13C, δ15N) in primary producers and pools of particulate and sediment OM (POM, SOM) from the KKB mangrove and marine ecosystem to determine how shrimp aquaculture OM contributes to the coastal environment. Here, soy‐based shrimp feed resulted in low shrimp δ15N, similar to marine POM, and thus we focus on the use of δ13C in tracking shrimp pond effluent in the environment. δ13C signatures of SOM varied significantly along a land‐to‐ocean gradient (–29.1‰ to –23.9‰). We found consistently depleted mangrove SOM δ13C signatures (–29.4‰ to –28.2‰) indicating that mangrove leaf litter is the primary source of OM to mangrove sediments, and there is little evidence that marine and shrimp pond OM contributes to the mangrove habitat. In contrast, relatively low δ13C values for marine SOM (–25.7‰ to –23.9‰) overlap with the δ13C of shrimp feed (–25.3‰) and Bayesian mixing models indicate that shrimp aquaculture feed and mangrove vegetation contribute the greatest OM to the marine ecosystem.  Compared to 20 years ago, marine SOM δ13C signatures are depleted by ~10‰ and similar throughout KKB, indicating a homogenization of marine SOM carbon sources from 1998 to 2018. Synthesis and applications. The doubling of shrimp aquaculture in Khung Krabaen Bay (KKB) since 1998 led to increased discharge to the bay, swamping organic matter (OM) contributions from 13C‐enriched seagrasses and marine plankton. Because of this increase in effluent release to KKB, the chemical impact is likely to be greater for the marine ecosystem than the mangrove and should also be a focus of conservation efforts. Continued technological improvements (e.g. closed systems, better feed efficiency) and support to local aquaculture farmers will help reduce OM discharge to coastal ecosystems and promote sustainable farming practices.

Hong, Alison H.*, Kathryn E. Hargan, Branwen Williams, Bunlung Nuangsaeng, Sarawut Siriwong, Pisut Tassawad, Chatdanai Chaiharn, and Marc Los Huertos. “Examining Molluscs as Bioindicators of Shrimp Aquaculture Effluent Contamination in a Southeast Asian Mangrove.” Ecological Indicators, vol. 115, 2020, 106365.

Abstract: This study evaluates the presence of metal(loid) contamination from shrimp aquaculture effluent in a nearby mangrove ecosystem in Khung Krabaen Bay (KKB), Thailand. Our objectives were to: 1) examine how sediment metal(loid) concentrations change spatially in KKB relative to the aquaculture ponds; (2) compare mollusc trophic dynamics of elements associated with shrimp aquaculture; and (3) determine if certain mangrove mollusc species present better ecological indicators of aquaculture elemental contamination. We analyzed targeted elemental concentrations (As, Mn, Cu, Zn, Cr, Pb) in sediments sampled at increasing distance from intensive shrimp aquaculture and within the tissues of five species of molluscs. Differences in mollusc diet were studied using carbon (ẟ13C) and nitrogen (ẟ15N) stable isotopes, and biomagnification and biodilution of metal(loid)s were examined using ẟ15N values to infer trophic position. Elemental concentrations were low in farmed shrimp and pellet food relative to mangrove molluscs, but high in the sediments of drainage ponds filtering pond effluent. Declining elemental concentrations from the aquaculture drainage ponds to the mangrove sediments closest to shrimp farms suggested that the management settling ponds are effective at attenuating metal(loid) contamination. Large differences in mollusc mean ẟ13C (–25.8 to –20.1‰) and ẟ15N (1.9 to 8.3‰) values indicate variable diets across species, and trophically-elevated molluscs had greater marine-based diets resulting in a significant, positive correlation between mollusc ẟ13C and ẟ15N values. KKB molluscs biomagnify As, Zn, and Se, whereas declines in Mn and Cu with increasing trophic position indicate biodilution. The metals Pb, Fe, Al, Cr were elevated in the shrimp drainage pond sediments and in the tissues of the gastropods Ellobium aurisjudae, Cerithidea obtusa (both mud grazing), and Nerita lineata (algal grazing at low tide), identifying these gastropod species as probable bioindicators of aquaculture contamination in southeast Asian mangroves. Although we find biomagnification of some elements, mangrove mollusc metal(loid) concentrations are not likely to be of toxicological concern, and in fact, were lower than many mangroves elsewhere impacted by industrial and sewage contamination.

Stewart, Joseph A., Laura F. Robinson, Russell D. Day, Ivo Strawson, Andrea Burke, James W.B. Rae, Peter T. Spooner, Ana Samperiz, Peter J. Etnoyer, Branwen Williams, Adina Paytan, Melanie J. Leng, Vreni Häussermann, Leslie N. Wickes, Rachael Bratt, and Helena Pryer. “Refining Trace Metal Temperature Proxies in Cold-Water Scleractinian and Stylasterid Corals.” Earth and Planetary Science Letters, vol. 545, 2020, 116412.

Abstract: The Li/Mg, Sr/Ca and oxygen isotopic (O) compositions of many marine biogenic carbonates are sensitive to seawater temperature. Corals, as cosmopolitan marine taxa with carbonate skeletons that can be precisely dated, represent ideal hosts for these geochemical proxies. However, efforts to calibrate and refine temperature proxies in cold-water corals (<20 °C) remain limited. Here we present skeletal Li/Mg, Sr/Ca, O and carbon isotope (C) data from live-collected specimens of aragonitic scleractinian corals (Balanophyllia, Caryophyllia, Desmophyllum, Enallopsammia, Flabellum, Lophelia, and Vaughanella), both aragonitic and high-Mg calcitic stylasterid genera (Stylaster and Errina), and shallow-water high-Mg calcite crustose coralline algae (Lithophyllum, Hydrolithon, and Neogoniolithon). We interpret these data in conjunction with results from previously explored taxa including aragonitic zooxanthellate scleractinia and foraminifera, and high-Mg calcite octocorals. We show that Li/Mg ratios covary most strongly with seawater temperature, both for aragonitic and high-Mg calcitic taxa, making for reliable and universal seawater temperature proxies. Combining all of our biogenic aragonitic Li/Mg data with previous calibration efforts we report a refined relationship to temperature: Li/MgAll Aragonite = (). This calibration now permits paleo-temperature reconstruction to better than ±3.4 °C (95% prediction intervals) across biogenic aragonites, regardless of taxon, from 0 to 30 °C. For taxa in this study, aragonitic stylasterid Li/Mg offers the most robust temperature proxy (Li/MgStylasterid (Arag) = ()) with a reproducibility of ±2.3 °C. For the first time, we show that high-Mg calcites have a similar exponential relationship with temperature, but with a lower intercept value (Li/Mg = ()). This calibration opens the possibility of temperature reconstruction using high-Mg calcite corals and coralline algae. The commonality in the relationship between Li/Mg and temperature transcends phylogeny and suggests a similar abiogenic trace metal incorporation mechanism.

Williams, Branwen. “Proteinaceous Corals as Proxy Archives of Paleo-Environmental Change.” Earth Science Reviews, vol. 209, 2020, pp. 103326.

Abstract: High-resolution marine data are critical to evaluating anthropogenic impacts on our environment. Considering the spatial and temporal limitations of direct instrumental measurements, proxy data extracted from marine environmental archives are necessary to separate anthropogenic changes from those that would occur naturally. The majority of late-Holocene, high resolution environmental reconstructions are derived from marine carbonates including tropical scleractinian corals, bivalves, crustose coralline algae, and sclerosponges. However, these archives are restricted to the top hundreds of meters of the water column, and only preserve environmental variability that is captured by calcium carbonate skeletons. At a very few select locations, marine sediments archive annual environmental variability in organic and inorganic materials. Recently, additional types of corals are proving to yield information complimentary to these traditional marine archives, including cold water scleractinian corals and proteinaceous corals. A taxonomically-diverse group, the proteinaceous corals are broadly defined as those having a branching gross-morphology with skeleton comprised at least partly of protein-rich organic material. They encode characteristics of their food and ambient environment into the chemical and physical composition of their skeleton. This environmental-encoding combined with their banded skeleton and significant longevity means that proteinaceous corals hold information in their skeleton that helps fill the spatial and temporal gaps in our knowledge of past and present ocean conditions.

External Grant: Williams, Branwen, Principal Investigator. “Collaborative Research: Using Multi-Proxy Paleo Data to Constrain Natural and Anthropogenic Hydrographic Variability in the Gulf of Maine System over the Last 250 years.” National Science Foundation, NSF OCE #2028219, 2020-2023, $157,736.00.

Abstract: The Gulf of Maine is located in the Northwestern Atlantic Ocean. Its waters serve as home to the economically important Atlantic Cod and shellfish industry. But recent changes to this environment pose a threat to the ecosystem and the commercial fishing industry. However, only a few long-term records of the environment exist in this area making it difficult to tell exactly when these changes started and how much is related to human activities. In this project, a team of researchers from three universities will construct a 250-year history of the regional environment by measuring yearly growth bands in local clams and algae. Chemical fingerprints in these growth bands will be used to determine the water temperature, salinity, biological productivity and pH conditions. This allow scientists to study how natural changes have affected the environment and compare these to changes caused by greenhouse global warming. This information will inform policy decisions of the newly-formed Maine Climate Council on how to better manage this area. The proposed work will also support training for early career scientists, broaden participation in science programs, and support undergraduate research opportunities. Specifically, the team will use a multi-pronged approach using geochemical proxies in two types of high-resolution marine climate archives (clams and crustose coralline algae) to reconstruct past changes in oceanographic conditions, including near surface seawater temperatures, salinity, and pH in the Gulf of Maine through the last 250 years. Geochemical fingerprints (or proxies) preserved in the annual growth bands of clam shells and skeletons of long-lived marine calcifiers have been used extensively as archives of past ocean conditions. Discrete calcium carbonate layers can be sampled from the growth bands of clams and crustose coralline algae for geochemical analysis, generating proxy archive records. These reconstructions of past environmental conditions fill data gaps prior to instrumental records. Hence, the proposed research will potentially yield several multi-centennial, annually-resolved, absolutely-dated datasets of oceanographic variability in the Northwestern Atlantic, including the Gulf of Maine, prior to the Industrial Revolution, which will be used to evaluate the role of Atlantic meridional overturning circulation, and other climate forcings, on observed changes. By combining new data derived from traditional and novel proxies with existing hydrographic records, the relative contributions of source waters into the Gulf of Maine through time will be characterized to better understand the drivers of variability within the Gulf of Maine. Understanding past oceanographic variability in the Gulf of Maine is therefore critical for predicting the likely extent and magnitude of future change, and for planning to safeguard ecosystems and fisheries. Results will be widely disseminated to science and non-science audiences through publications, conference presentations, mentoring programs, coursework, and outreach activities at area schools, museums, and science centers.