Fetal alcohol spectrum disorder (FASD) is considered one of the most prevalent forms of preventable mental retardation resulting in life-long behavioral and intellectual impairments. Many of the neurobehavioral deficits may not be apparent until the educational years and are typically not recognized as a result of fetal alcohol exposure (FAE). The hippocampus has been shown to be particularly susceptible to the adverse consequences of FAE in both rodent and non-human primate models. In our model of naturalistic and moderate FAE in vervet monkeys (Chlorocebus sabeus) we have previously shown diminished stem cell proliferation in the dentate gyrus in neonatal subjects. Here we expand on previous studies to test the hypothesis that FAE diminishes the number of immature neurons in the early differentiation period. Pregnant vervets were allowed to drink the equivalent of 3-5 standard drinks four times a week during the third trimester and subsequently sacrificed at 3 months of age which corresponds to the end of the rapid post-natal neurogenesis period. Serial sections were immunostained with nestin, a putative marker for early neuronal differentiation. Preliminary analysis suggests a decrease in nestin-positive neurons in the dentate gyrus of FAE subjects. Although the immature neuronal population is susceptible to FAE, it may also be the key to therapeutic intervention aimed at reducing the impact of FAE-induced neurological impairment.

The President’s Council of Advisors on Science and Technology estimates that the United States will need to increase its number of science, technology, engineering, and mathematics (STEM) professionals by 34% over the next decade. This guidance seems contrary to the rising trend of STEM majors becoming increasingly selective of the students they admit. Competitive degrees with additional requirements for entry (ARE) are a universally accepted phenomenon here at UW. However, there are no studies characterizing their prevalence in higher education or investigating their impact on students. We are conducting an exploratory study to characterize the number of STEM degrees across the U.S. with ARE and the factors departments use to admit students. First, we compiled institutional data for the research universities (R1) and Master’s-granting institutions with the highest number of undergraduates in each state. Next, we determined whether or not they offered STEM degrees with ARE. Programs were identified through a combination of data-mining institutional websites and contacting individuals at each institution. Finally, we administered an online survey to each department to gain deeper insight into the factors used in their admission decisions. We hypothesized that variables such as high school GPA, university GPA, and SAT score will commonly be used in the decisions to admit students. SAT scores and GPA often have significant socioeconomic and demographic biases. Consequently, if we confirm our hypothesis, this could raise questions about the equity of the admission processes for selective degrees. Preliminary results revealed over 250 degrees with ARE. Most of those degrees are offered in biology, health science, or engineering departments at R1 institutions. Ultimately, our results will inform a future study that will take a closer look at the impacts of degrees with ARE on STEM retention and diversity from a representative sample of institutions.

Alzheimer’s Disease (AD) is one of the leading causes of death in the United States. Studies suggest that the root cause of AD is due to two proteins, Aβ1-42 and tau, which accumulate and aggregate into amyloid plaques and neurofibrillary tangles, respectively. Current studies on the heritability and genetic foundation for AD are not well resolved. In order to understand which genes are best able to resist the effects of Aβ1-42 and tau, we utilized an AD fruit fly model. More specifically, we used a mutant line of Drosophila melanogaster, which we call R32, in which human transgenes of Aβ1-42 and tau are expressed in the fly eye. We used this reagent to express AD proteins in the Drosophila Genome Reference Panel (DGRP), a set of 200 fully sequenced, inbred lines derived from a wild population, in order to simulate a model of natural genetic variation. Aβ1-42 and tau proteins lead to degradation in the eyes causing fusions and irregular shaping of ommatidia. Accordingly, we imaged fly eyes at 28 days of age. We found significant genetic variation in ommatidial circularity and area, two measures of eye quality. We performed a genome-wide association study to identify specific single nucleotide polymorphisms (SNPs) that act as markers signaling degradation in the ommatidia. Significant SNPs were found linked to genes associated with neurotransmitter receptors, neural signalling, and photoreceptor differentiation. Further studies will try to validate the effect of the identified genes and quantify the effects of the SNPs on the degenerative eye phenotype. Additionally, metabolomic studies as well as a model that expresses the AD-related proteins in the brain would be utilized to study the effects on lifespan, memory, and locomotion.

Wheat Take-All Disease (TAD) is caused by the effect of the soil-based fungus Gaeumannomyces graminis var. tritici on wheat crops. We as Pierce College Biology students in conjunction with other local institutions sequenced part of a strain of Pseudomonas fluorescens, L5.1-96 (P.f. L5.1-96), because of its known trait to supercolonize soil. Sequencing was initially achieved at Pierce College by linear sequencing reaction before being completed by Sanger Sequencing at Washington State University. Resulting data was then interpreted using FinchTV, and the NIH BLAST and tBLAST programs. P.f. L5.1-96 produces 2,4-diacetylphloroglucinol (DAPG) which is a key component in Take-All Decline, the natural suppression of the fungus after several generations, in soils. Using Escherichia coli bacteria to clone 1.7kb long plasmids of the P.f. L5.1-96 strain, the plasmids were amplified and purified in order to discover genes relevant to the empirical objective to better understand this bacteria. Our DNA insert, the portion of data analyzed from FinchTV by cross reference with BLAST, aligned with several important enzymes related to antibiotic resistance and toxin metabolism, both of which could be key to supercolonization of soil. In addition, a phylogenetic relationship has been established using BLAST between P. brassicacearum, P. chlororaphis, and our strain of Pseudomonas fluorescens L5.1-96. The findings of the research could provide greater insight into what constitutes an effective biological control agent against TAD in Washington, and possibly other affected areas of the world.

Synaptogenesis is the developmental process in which neurons form connections with their appropriate partners. However, the mechanistic bases for synaptic partner matching are largely unknown. To help address this gap in our knowledge about neural circuit formation, we are attempting to identify factors that drive specificity in synapse formation between nociceptive sensory neurons, known as Class IV dendritic arborization neurons (C4da), and their synaptic partners, A08n interneurons in Drosophila melanogaster. We hypothesize that C4da neurons express cell adhesion molecules that serve as specificity factors. Expressing such specificity factors in sensory neurons that do not normally synapse with A08n, for example C3da neurons, should drive spurious synapse formation with A08n. To test this, I used the split Gal4 system to develop transgenic tools for targeting gene expression specifically in C3da neurons. In this system the Gal4 transcription factor is split into its DNA binding domain (DBD) and activation domain (AD) which are independently expressed in overlapping cell populations and assemble to yield functional Gal4 only in cells that co-express both pieces. The complete Gal4 protein can then interact with UAS (upstream activating sequences) elements to drive gene expression. To develop a C3da-specific Gal4 driver I first engineered transgenes containing the Gal4-DBD fused to promoters predicted to be expressed in C3da neurons and other non-da neurons. I generated transgenic fly lines harboring these transgenes via direct injection of DNA into the fly germline. I then crossed these transgenic lines to a fly line that expresses the Gal4-AD element in C1-C4da sensory neurons. I expect to see functional Gal4 only in C3da neurons, which I can test by monitoring Gal4-dependent Green Fluorescent Protein expression. The resulting C3da neuron-specific driver will allow us to test whether ectopic expression of candidate specificity factors in C3da neurons is sufficient to drive synapse formation with A08n interneurons.

Across animals, embryos can hatch in response to environmental cues. Early hatching can improve embryo survival when eggs are in danger, but may come at a cost to the larval life-stage. Thus, understanding the adaptive value of hatching plasticity requires evaluating the costs and benefits that characterize selective trade-offs. Several species of glassfrogs (Centrolenidae) can hatch as early as 7 days in response to predators, dehydration, and parental abandonment, or delay hatching to 20 days under good conditions. We measured two fitness correlates, for survival and growth, to test the adaptive value of delayed hatching in five species: Cochranella granulosa, Hyalinobatrachium fleischmanni, Hyalinobatrachium colymbiphyllum, Teratohyla pulverata, and Teratohyla spinosa. Hatchlings fall from arboreal eggs into streams and must dive to the bottom to escape predatory fish. We measured diving performance for early and late hatchlings to infer predation risk. As tadpoles cannot benefit from external food resources until they reach feeding competence, we quantified the onset of feeding and compared gut-coil development in siblings of different hatching ages. Older hatchlings dove faster and began feeding sooner after hatching than did younger individuals, suggesting that delayed hatching is advantageous for escaping predators and reducing lag-time in exotrophic-based growth of larva. This study provides insight on the selective trade-offs that favor hatching plasticity in glassfrogs.

Pseudomonas fluorescens is a bacterium recognized for producing an antifungal against the debilitating wheat take-all disease. In this study I am investigating the Pseudomonas fluorescens L5.1-96 strain to identify the genes making it a supercolonizer of take-all disease resistant soil and how it fits phylogenetically with other bacteria. This project will contribute to the understanding of how biological agents may be used to combat wheat take-all disease. To investigate this, a plasmid of DNA from strain L5.1-96 was inserted into Escherichia coli to make a bacterial library. I separated the plasmid from the E. coli and amplified it by linear sequencing then sent the DNA to Bellevue College for Sanger sequencing. I used bioinformatics to analyze my unique insert which I found did not match unknown genes, rather it coded for proteins regarding nitrogen fixation. The sequences matched those of Pseudomonas brassicacearum. This provides evidence to renaming the bacterial strain. A match to an anaerobic enzyme in nitrogen fixation might provide insight to its supercolonizer abilities. As we are sequencing a minor portion of the strain L5.1-96 genome, there is need for further gene sequencing in order to complete its genome.

8-methoxypsoralen is a photoactivated DNA-intercalating agent which, after receiving two high energy UVA photons, covalently binds pyrimidine bases on both strands of DNA, forming an Interstrand Crosslink (ICL). These lesions represent a complete block to replication and transcription, and ICL-inducing agents have been used widely in chemotherapies, yet the mechanism by which these lesions are processed in the cell remains poorly understood. In 1985, Ahmed and Holland reported an Escherichia coli mutant that was hyper-resistant to ICL agents. The mutation was mapped to 57.2 minutes on the chromosome and potentially encoded a 55-kDa protein that was induced during stress as part of the SOS response. Although these genes were never identified, hscA and hscB genes map to this location, have a similar size, and are SOS-inducible. To determine if these genes are responsible for ICL resistance, two strains of E. coli were constructed. In the first strain, the genes of interest were deleted and replaced by genetic recombination with chloramphenicol resistance. In the second construct, a wild-type strain was stably transformed with an arabinose-inducible plasmid containing hscA and hscB, which allows them to be overexpressed when the sugar arabinose is added to media. Here I report the resistance of both of these constructs and discuss the likelihood that these are involved in the cellular response to ICLs. In a second approach to identify ICL-resistance genes, a screen was established in which wild type cultures were grown and exposed to UVA light and 8-methoxypsoralen. Surviving colonies were then recovered and subsequently re-exposed to the ICL-inducing agents over several iterations. The accumulated resistance of this strain will be documented, with genome sequencing of any hyper-resistant mutants being performed to identify alterations contributing to ICL-resistance.

Alzheimer’s Disease (AD) is a progressive brain disorder that results in middle to late life dementia. AD, the most common form of dementia, is the sixth leading cause of death in the United States, and the third leading cause of death for older people over 65 years of age. AD is pathologically characterized by the accumulation and aggregation of an alternatively spliced amyloid-beta (Aβ) and hyperphosphorylated tau, which lead to the formation of neuritic plaques and neurofibrillary tangles (NFT’s) in the brain and ultimately to decreased neural function and cell death. Previous studies indicate that the extent to which an individual is affected by the interaction of these proteins and the severity of AD is largely influenced by genetic variation. However, the specific genes involved, their regulation and expression remain largely unknown. Our ongoing project explores the genetic variation underlying Drosophila susceptibility to severe eye cell degeneration caused by human Aβ and tau. We express Aβ and tau in the fly eye through the UAS-GAL4 system to create our AD model Drosophila, and cross them with a panel of flies (the Drosophila Genome Reference Panel, DGRP) that allows us to map natural genetic variation for resistance to Aβ and tau. Our preliminary results show significant variation in neuronal eye cell degeneration among DGRP lines, implying that neurodegeneration would also differ between DGRP lines if Aβ and tau were expressed in the nervous system. Based on current progress, this new project will continue exploring variation in DGRP lines, but this time we will express them in the fly brain. I will measure how different genotypes shape the effect of brain-specific Aβ and tau on lifespan and protein aggregation, and will use Genome Wide Association Studies to identify single nucleotide polymorphisms associated with variation in expression of Aβ and tau.

Pseudomonas fluorescens strain L5.1-96 has been known to help combat the wheat take-all fungal pathogen, Gaeumannomyces graminis var. tritici (Ggt). Strain L5.1-96 is maintained in the soil longer than others, so it is known as a supercolonizer, which contributes to the phenomenon referred to as Take-All Decline (TAD). The strain L5.1-96 is a non-pathogenic bacterium which grows in the rhizosphere of plants and produces an antifungal agent known as 2.4-diacetylphloroglucinol (DAPG) causing TAD. The DNA fragment we amplified, sequenced, and analyzed was a perfect match to P. brassicacearum strain BS3663. Through bioinformatics, we also found our fragment matched the genetic code for pyridoxal 5'-phosphate (PLP)-dependent aminotransferase, a co-enzyme which catalyzes the transamination of amino acids to alpha-keto acids. We did not identify new proteins that are responsible for the advantages that this bacterium holds against wheat take-all; although, we did find the best phylogenetic fit for this strain. These findings contribute information to the broader study at Washington State University to investigate whether strain L5.1-96 can be used as a biocontrol agent against wheat take-all.​

Reports have shown the importance of student engagement and student-centered pedagogies in biology classes. A sense of community in a classroom can help students be less isolated from their peers and increase their willingness to answer questions, as well as their willingness to ask for help. This classroom sense of community can enhance students’ perception of learning and can be achieved online. Furthermore, a sense of community among underrepresented groups (URG), which have the highest attrition rates in STEM, can be a powerful tool for increasing student retention.  We hypothesize that blogging enhances a sense of community and connectivity among students. Our study analyzed 82 students’ interactions during a semester-long writing assignment in a biology class for non-biology majors at Emory University. Half of the students submitted their assignments as blog posts, the other half through a traditional educational content management system. Both groups had equivalent distributions of diverse ethnicities. Student connectivity was incentivized and monitored by having students turn in editor-sheets, reporting whose assignment they edited, and how. This student connectivity was subsequently analyzed via a Network Analysis.  Student surveys showed trends of higher perceived sense of community among bloggers relative to traditional writers. The network analysis revealed double the average connectivity among bloggers, with 2-folds higher average connections per student in the blogging group. Students also showed more evenly distributed connections among bloggers relative to traditional writers.  We conclude that the use of blogging among non-biology majors can facilitate student-student interactions and has the potential to create a sense of community, which has positive implications in the retention of under-represented groups in STEM fields.

In this study, we focused on identifying specific genes that make Pseudomonas fluorescens L5.1-96 a potential biocontrol agent to combat wheat take-all disease. This strain is a supercolonizer meaning it suppresses fungal pathogens in the soil and colonizes the rhizosphere of plants. This strain has not yet been fully sequenced; therefore, analyzing genomic data will contribute to its classification. Our first step was increasing the amount of L5.1-96 DNA through culturing E. coli that contained an insert of L5.1-96 DNA. Next we extracted L5.1-96 DNA using the Qiagen miniprep kit, amplified the DNA insert using linear sequencing, and sent our cleaned DNA samples to Bellevue College for Sanger sequencing. We ran bioinformatics software (NCBI BLAST) to compare our DNA insert with previously sequenced genomic data. We found our DNA sample resulted in a 98% match with genes from Pseudomonas brassicacearum strain BS3663; therefore this indicates a phylogenetic relationship between the two Pseudomonas species. While we did not discover any new genes or novel proteins, our sample coded for tRNA preQ1(34)S-adenosylmethionine ribosyltransferase-isomerase QueA which is a nutrient protein found in both eukaryotes and bacteria. We have contributed to the genomic analysis of strain L5.1-96 and with this information it is possible one day all of the data together will lead researchers to fully evaluate strain L5.1-96 and its ability to suppress wheat take-all disease.

In educational and research settings, Tetrahymena thermophila is a fresh water protozoan that serve as an excellent model organism for elucidating phagocytosis and ciliary motion. As a step towards determining how Tetrahymena thermophila adapt to environmental changes, we developed new, practical, inexpensive approaches that label organelle changes in fixed and living cells. To address our research objective, we pursued the following experimental approaches: One, a novel heat fixation method was used to fix Tetrahymena thermophila prior to doing a Wright stain and different conditions of Wright stain were performed and observed with a light compound microscope; two, to observe cells that have loss of plasma membrane integrity, we pre-incubated 20µM of Sytox green dye for three hours and observed for green fluorescence in the macronucleus (MAC) and micronucleus (MIC). We used a new portable battery-operated inverted fluorescence microscope designed by Dr. Rebecca Richards-Kortum’s research group and Cellcams for acquiring images and time-lapse movies. We found that heat fixed Tetrahymena thermophila followed by a Wright stain labeled organelles at different stages of their life cycle. We conclude that using Wright stain and Sytox green fluorescence dye in combination with heat fixation are novel low-cost applications that allow phenotypic analysis in Tetrahymena thermophila. In the future, these useful approaches can be applied broadly into many lab inquiry settings, such as toxicology and molecular genetics.

According to the Center for Disease Control, more than one third of the U.S population is obese. Preliminary data in our laboratory has shown that obesity reduces the number of T cells in the intestines of mice. This reduction of T cells leads to impaired tissue repair in dextran sulfate sodium (DSS)-induced colitis models and compromises the protective barrier against pathogens and exposure from the outside environment. Thus, our goal was to identify how to restore the number of T cells to improve intestinal homeostasis. We investigated whether T cell numbers in the intestine can be fully restored through diet-induced weight loss. Briefly, 6 week old male mice were placed in one out of three cohorts: one cohort was placed on a normal chow diet for 14 weeks, the second cohort was placed on a high fat diet for 14 weeks, and the third cohort was placed a high fat diet for 7 weeks and switched to a normal chow diet for the remaining 7 weeks. While mice placed on a high fat diet exhibited a significant reduction in T cell number, mice that underwent diet-induced weight loss exhibited a full restoration of T cell number in the small intestine. In addition, the dieted mice also showed improved survival when challenged with DSS-induced colitis. These studies are now being followed up with RNA-sequencing experiments to investigate how gene expression is changing within the intestine and better understand the mechanisms of T cell loss in obesity.

Flowering plants have coevolved with insect pollinators in a mutualistic relationship; plants provide nectar to insects and insects in turn cross pollinate flowers. Plants have employed a range of colors, scents and shapes to attract their insect pollinators. Vision, olfaction, and mechanosensation all have roles in the identification and interactions with flowers. While the roles of vision and odor sensing have dominated the literature, recent work has shown that mechanosensation plays as much a role as vision in ensuring that moths stay engaged with a moving flower. Moths use their straw-like proboscis to probe a flower, using the overall shape of the flower as well as tiny grooves in the petals to guide the insect’s mouthparts toward the nectary. Using the feeding behavior of the nocturnal hawkmoth, Manduca sexta, we ask how moths use mechanosensory information to find the nectary opening in flowers. We combine 3D printing technology--to generate different 3D flower curvatures, with micro-sensing technology--that allows us to detect the proboscis tip inside nectary, and computer vision techniques--to detect the motion of the hovering moth, to test the relationship between flower shape and efficiency in locating the nectary. We find that flower exploration depends on its 3D curvature. Moreover, over repeated trials, moths take less time to find the nectary across all curvatures suggesting that moths use tactile exploration to learn flower shape. Mechanosensation is one of many inputs used by moths in the wild to interact with their environment. In future studies we wish to explore the integration of mechanosensation with other sensory information, and the salience of each modality.

In the population of bottlenose dolphins (Tursiops aduncus) in Shark Bay, Australia, groups of two or three adult males form long-term alliances that sequester, harass, and intimidate adult females, presumably to increase their chances of mating success. Costs that males inflict on females include physical injury, changes in home range and habitat use, reduced foraging, and changes in calf care. Although altered female behavior is evident during association with males, the direct impact males have on female behavior is not entirely clear. That is, male behavior and female responses to such behavior during consortships have not been explicitly documented. A long-term study has collected detailed behavioral data on the Shark Bay population since 1988. Using this dataset, this project will quantify the rate of male aggression directed at females and how female activity budgets and foraging tactics change in response. Preliminary work indicated that rates of received aggression were higher for cycling females than non-cycling females, and higher when females were in the presence of more than one male, potentially confirming that males use aggression to coerce reproductively viable females. Females also spent less time foraging when in the presence of more than one male, hinting at potential costs to the female owing to loss of food. Because females have highly specialized foraging tactics, changes in foraging behavior and home range during prolonged or repeated consortships could impact female condition.

Giardia, a protozoan parasite which resides in human intestine, can cause Giardiasis, a common parasitic disease associated with severe diarrhea. It possesses a simple life cycle including the motile trophozoite stage and infectious cyst stage. Once a human consumes unsanitary drinking water or food with Giardia cysts, the low pH in stomach drives excystation to release the trophozoites. When they perceive stimuli from the high bile and alkalinized pH environment, they will initiate encystation by turning on the Cyst Wall Protein gene to form cysts. Our purpose is to understand the spatiotemporal encystation process and the encystation rate of Giardia in a physiological environment (murine intestine) via observing the ratio metric change of encysting and non-encystation cells in the Dual-color Bioluminescence Imaging (BLI) System. BLI is essential to monitor noninvasive biological processes in living organisms via detecting photon emissions that requires the enzyme luciferase to catalyze a substrate, usually luciferin, with oxygen, ATP, and magnesium. To test if dual-color BLI can report the ratio metric change of encysting and non-encysting cells in vitro, we have generated the in vitro encystation assay. The promoter of Glutamate Dehydrogenase, a constantly expressed gene at all stages of Giardia, is combined with a luciferase gene to represent non-encysting cells when transcribed in normal media. The promoter of cyst specific gene, Cyst Wall Protein 1, is combined with another luciferase gene to represent encysting cells when expressed only in encystation media. By successfully observing the ratio metric change of luciferase expressions in vitro, we can then proceed to future mouse experiments. Once the technique has been confirmed feasible in the mouse model, it can be applied to monitor the life cycle of all cyst forming parasites and understand when and where they are at infectious stage, thereby improving the accuracy for alternative drug screening.

Seasonal stratification of the water column likely influences food availability to mussels grown at certain depths on aquaculture lines, as well as the environmental conditions experienced by the mussels. Fatty acids are important structural molecules that reflect the diet of the organism, such that fatty acid composition provides information on how the condition of mussels grown in different environments responds to food changes. Food sources vary based on a number of environmental conditions, including water temperature and turbulence, which differ between stratified water layers. Fatty acids in aquaculture mussels grown at 1m depth were compared to those at 7m in depth in the summer and fall of 2016 to test effects of varying environmental conditions between the depth layers. The results showed significant variability in the fatty acid composition of mussels grown at different depths in the summer months, but none in the fall. The difference between depths was contingent on the season. The variability in the summer months is likely due to stratification from increased surface temperatures, which decreases mixing and nutrient supply to mussels at lower depths. Stratification is then reduced in the fall and may explain homogenization of mussel fatty acid signatures from that period. In 2017, monthly sampling at the two depths was repeated and another experiment analyzing mussel plasticity was conducted in which mussels growing at 1m and 7m depth were switched during the summer and monitored at the new depth. Fatty acid signatures of switched mussels are predicted to adjust to become consistent with signatures of mussels established at the new depth by the end of the five-month sampling period. The results of this study will demonstrate the adaptability of mussels to new feeding environments and the effects of environmental changes on mussels as variable water conditions impact their algal food sources and overall health.

Skeletal reduction is a common feature among deep-sea fishes that have diversified from shallow-water relatives, such as snailfishes. These skeletal reductions may be an adaptation to environmental conditions of high pressures, low temperatures, declining luminosity and limited food availability. Snailfishes (family Liparidae) are found across a large bathymetric range (0 –>8,000 m), with intertidal ancestors giving rise to a large clade of deep-sea species. We used microcomputed tomography (micro-CT) to estimate average bone mineral density and examine jaw, pectoral girdle, and neurocranium morphology. Our results suggest at least three mechanisms of skeletal reduction: (1) reduction of bone size, (2) reduction of bone density, and (3) loss of skeletal elements. First, using phylogenetic generalized least squares (PGLS) analysis, we found that the change in cranial dimensions with depth was not uniform. While the size of the maxilla, dentary, and pectoral girdle decreased with greater depth, length of the upper premaxilla and the neurocranium did not vary with collection depth. Second, average density of the lower jaw decreased with increasing depth. Lastly, the ventral suction disc has been lost multiple times within the deep sea lineage. While all three methods are seen in snailfishes, other groups may use some or all of these mechanisms to different extents. Some mechanisms of skeletal reduction may be more advantageous than others. The extent to which a structure is retained in deep-dwelling fishes may indicate its functional importance. Variable skeletal reduction in the family Liparidae provides insights into the physiological adaptations that allow fishes to survive in deep-water environments. We conclude that some skeletal elements are maintained at the expense of others as fishes balance the functional demands of life in the deep sea.

Despite the large number of gametes that clams release, very few offspring survive to reproductive age and harvestable sizes. There are several ways that aquaculturists try to protect young settlers, including adding gravel on top of fine sediment. We tested whether the addition of gravel helped decrease the mortality rate from small crabs in lab experiments. We tested predation rates on juvenile manila clams, Ruditapes philippinarum by the hairy shore crab, Hemigrapsus oregonensis in different sediments. Clams were placed in tubs with no sediment (NS), fine sand (FS), or sand with gravel on top (SG). After allowing time for the clams to burrow, crabs were added to each tub and given 24 hours to consume the clams. Crabs consumed some clams in all treatments although there was high variation among individuals. Gravel addition did not reduce predation rates compared to fine sand alone, but there was a trend for more predation in no sediment than with sediment. We also tested the ability to consume small clams of different predators that live in the same sediment habitat as R. philippinarum. The worms, Nereis brandti, Paranemertes peregrina and Hemipodus borealis all showed no predation on clams within a 24-hour period. The snail, Alia also showed no predation on juvenile clams. By understanding multiple factors that lead to juvenile clam mortality, fisheries can improve upon their methods to increase harvestable populations.

Post-secondary education is often seen as critical to the ultimate goal of progressing up the socio-economic ladder. However, social inequality can impact academic success. Social inequality can be expressed through classism, obstacles and interruptions in the academic journey of low-income college students, racial isolation in schools, residential segregation, negative academic experiences, and differences in child development. This type of inequality establishes divisions of privilege where obtaining the “American Dream” is easier for some and harder for others. So how can children from lower class families break out of the poverty cycle when society treats them unequally? This study examines social inequality in families, social institutions, and post-secondary education. This research focuses on racial/ethnic minorities in the United States within the age range of 21-25 years old and explores questions related to their family, childhood experiences, and future goals. This work aims to emphasize the great importance of breaking down the inequality barriers of classism.

Reactive oxygen species (ROS) are byproducts of photosynthesis that can damage plastid DNA (ptDNA), and 8-oxoguanine (8-oxoG) is the most common DNA lesion. Our lab previously showed that maize ptDNA damage, in general, increases during the development of non-green proplastids to green, photosynthetic chloroplasts. The objective here is to determine the amount of 8-oxoG in ptDNA during plastid development under light and dark growth conditions. DNA damage will be assessed using a semi-long qPCR procedure and the enzyme formamidopyrimidine DNA glycosylase (Fpg) which selectively removes 8-oxoG lesions. Testing of the semi-long qPCR method was performed with control DNA treated with damage-inducing UV light. After UV treatment, PCR amplification decreased, as expected for Taq DNA polymerase inhibition by bulky lesions. PCR amplification of ptDNA also decreased after UV light treatment of light-grown, dark-grown, and dark-to-light transferred seedlings. When ptDNA was extracted from plants grown in these three conditions, treatment with Fpg affected the PCR amplification, indicating removal of 8-oxoG lesions. 8-oxoG will now be assessed along the developmental gradient from proplastid to mature chloroplast. We may find more 8-oxoG lesions in mature chloroplast due to high ROS from photosynthesis than in the undeveloped proplastids, as well as more 8-oxoG in light than dark growth conditions.

Bone histology, the study of bone tissue, has become a critical tool in the field of paleobiology as a proxy for life-history information, such as growth rates, that is otherwise inaccessible from fossils. However, the bulk of bone histological research has focused on birds, crocodilians, and large-bodied mammals, leaving the extent to which bone microstructure acts as an indicator of growth rates among small mammals largely unknown. Therefore, we generated thin sections from the limb bones (e.g., tibiae, humeri, and femora) of a taxonomically diverse sample of 12 extant, small-bodied (<1 kg) mammals in an attempt to build a framework upon which we may make inferences about the evolution of mammalian growth dynamics. In general, vascularity is low or absent in our samples, and the degree of vascularity appears to decrease with decreasing body mass. All specimens exhibit two types of bone matrix: well-organized parallel-fibered bone and disorganized woven bone. Often, this parallel-fibered tissue lines the interior and exterior of the bone cortex, bordering an intermediate band of woven tissue. This middle band likely represents a period of rapid bone deposition, possibly corresponding with rapid growth in early ontogeny, while the parallel-fibered bone represents deposition after growth had slowed. We hypothesize that the structurally competent parallel-fibered bone bordering the cortex may act to buffer the intermediate, structurally weak, woven bone against mechanical strain during locomotion. To test these hypotheses, we will compile existing life-history data and quantify the histological features of our sample using NIS Elements photo documentation software, and apply multiple linear regression models to test for relationships of significance. By investigating the impact of life-history characteristics upon small mammal bone microstructure, we hope to make important inferences about the lives of extinct small mammals, and thus create a more comprehensive understanding of mammalian growth dynamics. 

Multiple factors interact to determine the lifespan of an organism. The Promislow lab uses the fruit fly Drosophila melanogaster to study the interaction between the metabolome (the profile of all small molecules within an organism), and the lifespan of a fly under stress. In a study of metabolome and lifespan data for many genotypes of Drosophila, our lab found that fly strains with relatively long lifespans when exposed to oxidative stress (peroxide food), had high levels of maltose, a disaccharide of glucose. We hypothesized that maltose was beneficial to flies on peroxide food, and tested this by supplementing the diet with maltose to see if this would extend lifespan. Flies fed supplemental maltose prior to exposure to peroxide food lived longer than flies fed unsupplemented food, supporting our hypothesis. Maltose could extend lifespan by providing energy to the fly via metabolism into glucose, or through another function as a disaccharide. To distinguish between these possibilities, we tested lactose, a disaccharide, to determine if any disaccharide could extend lifespan. Lactose did not extend lifespan, suggesting that disaccharides in general do not extend lifespan under stress. We found that glucose extended lifespan, supporting the hypothesis that maltose extends lifespan via conversion to glucose. Maltose can be stored as glycogen, a polysaccharide, and glucose is derived from glycogen by glycogen phosphorylase, encoded by the gene GlyP. To test the role of glycogen metabolism on lifespan under stress, we manipulated the expression of GlyP. Several transgenes were used to reduce the expression of GlyP by RNA interference (RNAi). RNAi of GlyP decreased lifespan, which supports our hypothesis that glucose derived from glycogen promotes survival. Our work suggests that glucose derived from glycogen or maltose is an important determinant of lifespan under stress, furthering our understanding of links between metabolism and complex phenotypes, like lifespan.

Magellanic penguins (Spheniscus magellanicus) have highly variable reproductive success, and while most eggs successfully hatch, about 50% of the chicks survive to fledging. What determines which of the two chicks in a brood fledges? Typically the first chick hatches two days before its sibling, and has an early size advantage in terms of weight. We predict that larger chicks will occupy the preferred nest position, switching from the warmer position early in development to the optimal feeding position. We also expect that chicks more often found in the “better” position at these different stages will be more likely to fledge. To assess optimal nest positioning we collected weight, nest position, and fledging data on over 1,200 chicks during the 2017-18 Magellanic penguin breeding season at Punta Tombo, Argentina. We extracted data from our SQL Database using a number of SQL queries, and we will use R Studio to analyze our results. We expect to find larger chicks preferentially in the back (warmer) position of the nest early in growth for better thermoregulation, and in the front of the nest later in development since they would be closer to the adult’s head during feeding. Developmental priorities should change as the chicks begin to independently thermoregulate at around 15 days old, so we expect a transition in chick positions at this time. Chicks consistently in the preferred position should have a higher likelihood of weight gain and fledging. These findings will further our understanding of the key factors and challenges in fledging and reproductive success at our study colony at Punta Tombo, Argentina.

Adaptive learning technologies are regarded as an innovative approach to personalize student learning in large college classrooms where interaction with instructors is limited. Such software allows students to learn at their own pace, and to concentrate their efforts on topics which need improvement. However, studies on adaptive learning systems across disciplines have yielded mixed results as to whether they have an impact on improving course mastery. Our research focused on testing how use of adaptive learning reading quizzes in the final quarter of an introductory biology series impacted student exam performance. Students (n=576) enrolled in this course were distributed across 25 lab sections. Students were randomly assigned by lab section to either static or adaptive learning pre-lecture reading quizzes. Both types of quizzes were provided to students through Macmillan’s LaunchPad portal, and points were awarded on a completion/ no completion basis. We used multiple linear regression with model selection to compare student performance between the two treatments, controlling for student characteristics (e.g., GPA, gender). When student performance on in-class reading quizzes and course exams was compared, no statistically significant differences were found between treatments. Similarly, through the administration of three surveys throughout the quarter, we were able to measure students’ attitudes toward different course components. Our findings show that there was no statistically significant difference as a result of adaptive learning reading quizzes on how prepared students felt for class, how much they enjoyed the course, how much they valued reading quizzes, or if they preferred the reading quiz format in this course to those in previous courses. Our findings suggest that given the additional financial cost of adaptive learning technologies to students, instructors should first verify that adaptive learning significantly impacts mastery of course material prior to fully integrating it into their course structure.

Cancer is one of the deadliest ailments in the United States, killing hundreds of thousands of people per year. One group of proteins that have been implicated in human cancers are the Phosphatase of Regenerating Livers, or PRLs. PRL-1 has been shown to have differing effects on cancer. Some research has found this protein to be an oncogene while others have found it to be a tumor suppressor. In our lab, past research has shown that for dPRL-1 to function as a tumor suppressor, it must localize to the adherens junction. The adherens junction helps glue epithelial cells together; malfunctioning adherens junctions, found in some cancers, no longer glue cells together thereby allowing them to metastasize. To better understand how dPRL-1 can function as a tumor suppressor, I performed a co-immunoprecipitation on the homolog, dPRL-1 in Drosophila melanogaster to discover the proteins that dPRL-1 interacts with in vivo at the adherens junction. To do this, I overexpressed dPRL-1 in the developing wing epithelium of the organisms and used an antibody to dPRL-1 for co-immunoprecipitation. This approach is an unbiased way to identify proteins that dPRL-1 interacts with while functioning as a tumor suppressor. My initial attempts at co-immunoprecipitation revealed one unknown protein of possible significance, but I have been unable to identify it yet. I am also testing my protocol to see if E-cadherin, a protein component of the adherens junction that contributes to cell adhesion, and dPRL-1 co-immunoprecipitate. We expect this interaction since our lab found that the two proteins colocalize via fluorescence microscopy. The proteins that dPRL-1 interacts with when functioning as a tumor suppressor could be mutated or missing in cancer cells thereby preventing dPRL-1 from functioning as a tumor suppressor. Hopefully, this knowledge will help clinicians make more effective chemotherapies to target cancers affected by PRLs.

Plants recycle resources from dying organs to feed the development of new ones. For example, chloroplast degradation in senescing leaves is a major source of nitrogen for seeds. Darkness or shade can induce leaf senescence and chloroplast degradation, but if light returns soon enough, new chloroplasts are made and the leaf remains photosynthetically active. Phytochrome B (PHYB) is a protein that serves a key role in resource allocation throughout the plant. phyB null mutants of the model crop species Brassica rapa (Br) exhibit low chloroplast count, poor seed production, and a reduced increased growth response to high CO2, as compared to wild type. We will investigate the underlying cause of these phenotypes by comparing the transcriptomic response of chloroplast-biogenesis and nitrogen-assimilation-related genes in wild-type and BrphyB null mutants before and during dark-induced senescence, as well as after recovery. Establishing a deeper understanding of the scope of PHYB's role in resource recycling will help us isolate appropriate targets for engineering more drought-resistant and nutrient efficient crops.

The trio of crops that are responsible for the bulk of human nutrition are pollinated by wind. Wind pollination has evolved repeatedly from insect pollination in angiosperms, however the mechanism for these shifts, such as the specific morphological traits and genes involved, has not been explored. Thalictrum (in the buttercup family Ranunculaceae) is a genus of herbaceous perennials with considerable floral diversity and multiple transitions to wind pollination in closely related species, making it an ideal system to study pollination syndrome shifts. Insect-pollinated species have larger, showier flowers with pigmented organs, while wind-pollinated species have green floral organs, exserted carpels and pendulous stamens. Here, I set out to characterize and quantify floral traits in wind and insect-pollinated species in Thalictrum to: 1) Test whether there is a suite of floral traits associated with wind pollination syndrome; 2) Prioritize the traits that contribute the most to pollination syndrome; and 3) Find the genetic basis of these traits. Flower scans from live plants, in addition to specimen from the University of Washington Herbarium, were used to measure floral traits using imageJ in 17 species across the Thalictrum phylogeny. Multivariate analysis of floral traits using Principal Component Analysis, showed that species cluster by pollination syndrome. Correcting for phylogenetic relatedness resulted in most of the variance being explained by the first principal component, clustering species by pollination syndrome even when they are distantly related. This evidence suggests that a specific combination of traits has been independently selected via convergent evolution in the transition from insect to wind pollination in this group. Next, I plan to conduct reconstruction of ancestral character states to track trait evolution and identify those that most closely correlate with shifts in pollination mode, followed by expression analysis of candidate genes.

Antibiotic resistance is an enormous public health problem and causes death for 23,000 Americans each year due to ineffective treatments. Many strategies to target resistance have emerged, the majority of which target resistance machinery directly. In Escherichia coli, the TEM-1 gene encodes for the β-lactamase enzyme which can degrade penicillins, a subclass of β-lactam antibiotics. Alleles of this gene can either improve or weaken the ability of β-lactamase to degrade other subclasses of β-lactam drugs. One strategy that can potentially slow the evolution of bacteria and select against antibiotic resistance is exploiting collateral sensitivity – mutations that confer resistance to one drug and confer sensitivity to another. Utilizing a technology called Deep Mutational Scanning, we are screening every amino acid variant of the TEM-1 gene, and examining growth patterns in twelve different β-lactam antibiotics. This will allow us to quantitatively measure the effect of every single protein variant on the fitness of the organism in hopes of identifying patterns of collateral sensitivity. This is done by first optimizing growth parameters and next sequencing the DNA of each amino acid variant before and after exposure to drug. The large data set produced allows us to uncover common evolutionary trends. This foundational knowledge will potentially allow us to predict drug pairings that exploit collateral sensitivity, leading to more successful treatments.

Carnivorous plants live in low nutrient environments and consume insects for heterotrophic nutrients. While previous research has typically focused on how pitcher plants capture prey, there is a lack of research concerning how pitcher plants attract pollinators. Scent is an important component of insect attraction, and this study explored whether pollinated and unpollinated plants produce different scents. This information will help us better understand the mechanism of pollination in carnivorous plants. We hypothesized that unpollinated plants will give off a different scent than the pollinated plants in order to attract more pollinators. The plants used for this experiment are the Sarracenia flava (yellow pitcher plant) which is endemic to the southeastern United States. We created two treatment groups consisting of pollinated and unpollinated plants inside a climate-controlled growth chamber at 25°C. Plants were watered from the base with unfertilized water. Scents were collected for 24 hours using Porapak Q adsorbent matrix and analyzed using a Gas Chromatograph Mass Spectrometer to identify the compounds. We anticipate that our results will further our understanding of how plants and insects interact. Future studies and analyses will focus on insect behavioral responses to plant scents, and on how insect antennae respond to each scent component.

Active student reasoning during peer discussion reinforces students’ conceptual understanding of lecture material and provides students with the opportunity to practice articulating the logical explanations required for exam questions. The quality of student reasoning during in-class discussion, and hence their usefulness in improving exam performance, may be affected by the questions instructors choose to ask. To determine which question properties prompt higher reasoning quality, we recorded and transcribed instructor questions asked and eight volunteer student group discussions to those questions over seven days in an active learning introductory biology classroom. We coded the in-class discussions for reasoning quality. We characterized questions in four ways: 1) Bloom’s taxonomy level (i.e., higher or lower order cognition questions), 2) question format (i.e., clicker question using peer instruction, think-pair-share, in-class Canvas questions where students must upload a picture of their work to Canvas), 3) number of dimensions of science learning using the 3-D Learning Assessment Protocol, and 4) if the question included a general model of physiology. Our results indicate that, in general, higher Bloom level questions encourage higher quality of reasoning. However, clicker questions using peer instruction prompt higher reasoning quality regardless of Bloom level. Additionally, the highest level of reasoning quality was seen during in-class Canvas questions. Both questions types that are incentivized by course credit (i.e., clicker and in-class Canvas) prompted higher reasoning quality than non-credited questions (i.e., Think-pair-share). We suggest that instructors should incorporate credited and higher Bloom level questions to increase the amount of high level reasoning during student discussions in their classrooms.

Active nests at the world’s largest Magellanic penguin (Spheniscus magellanicus) colony at Punta Tombo, Argentina have declined 40% since 1987. Climate change has increased rainfall and intense storms resulting in high chick mortality. Although these large-scale weather changes are known to factor into the decline, the effect of weather patterns on the scale of individual days is uncertain. To gain insight into penguins’ daily foraging patterns throughout the annual breeding season, automatic tread scales were placed on a penguin “highway” so when a penguin crossed we could document their time and direction of crossing, weight, and RFID tag. We sought to redesign the scales to improve energy efficiency by switching to solar power and increase accuracy and durability by changing the protective covers on the devices. Most importantly, the modified scales were required to be waterproof to protect internal components from heavy rain and flooding. The batteries used to power the scales had to last 7 days before needing to be replaced. Lastly, the materials used had to withstand temperature fluctuations and be animal-proof. The new design implements solar panels to charge batteries for the scales and cable glands to prevent water ingress through sides of the scales. The updated scales are made of wood and fiberglass then encased in heat-sealable fabric and sealant for waterproofing. Prototypes were firstly tested in the lab to see if they resisted flood-like conditions. Next, we tested whether the scales performed well after being exposed to large temperature changes using temperature controlled rooms. Additional tests were performed in the field to determine whether the design modifications decreased the amount of water ingress and increased battery life. Long-term tests in the field will be important to see how the materials weather over time when faced with temperature fluctuations, flooding, and animal usage. 

In mammals, daily rhythms of physiology and behavior are synchronized to a 24-hour light-dark (LD) cycle via input to the suprachiasmatic nucleus (SCN), the master circadian clock. Disruptions to the circadian system are associated with neurological and psychiatric disorders, including mood disorders like depression. These disorders are associated with symptoms such as sexual dysfunction and anhedonia, the inability to experience pleasure from activities that are typically enjoyable. Previous work has shown that rats exposed to a 22-hour LD cycle, which leads to desynchrony between neuronal oscillators in the SCN and mimics an internally desynchronized state, display behaviors indicative of these symptoms. Jet lag is a common challenge to the circadian system and, in rats, causes desynchrony between SCN neuronal oscillators. We hypothesized that desynchrony induced by jet lag could also lead to depressive symptoms. We measured sexual behavior before and after a 6-hour phase shift, simulating jet lag. Male rats were placed in an arena with a receptive female and sexual activity was quantified. We also performed a saccharin preference test where males had continuous access to water and saccharin solution. The volume drank from each bottle was measured daily; anhedonia is evident as a decrease in the preference for saccharin solution. Locomotor activity was recorded using infrared detectors, to associate behavior with the amount of time it took for males to re-entrain to the new LD cycle. Six-hour advances and delays of the LD cycle were not associated with behavioral manifestations of depression. This work, combined with previous work, provides greater insight into the consequences of circadian disruption and suggests that mental health consequences depend on the specific type of circadian misalignment. Whereas steady state misalignment induced by exposure to 22-hour LD cycles is associated with a depressive phenotype, the transient misalignment induced by jet lag is not.

Insect flight relies heavily on visual sensing. In many flight behaviors (e.g. navigating over long distances or through cluttered environments, finding food sources, or evading predators), insects must parse the visual scene to extract an estimate of their own motion and identify external objects or agents moving in their environment. Across numerous taxa and behaviors, there is a rich literature exploring behavioral responses to wide-field optic flow (visual stimuli arising from egomotion) and small-field target motion (cues corresponding to exogenous motion), primarily in the yaw dynamics involved in navigation. In contrast to yaw which is marginally stable, the equilibrium about pitch angle is inherently unstable, hence there are significant consequences to adjusting the flight attitude. To stabilize the visual scene under this constraint, insects can either reorient their body or move their head to redirect gaze. In this work, we investigate how the hawkmoth, Manduca sexta, modulates body pitch and gaze angle in response to wide- and small-field visual motion. Moths are tethered to a freely rotating armature at the center of a cylindrical arena and presented an image of a circular flower against a background grating. Figure and ground are oscillated both individually as well as simultaneously (both synchronously and incongruously). A multi-input--multi-output analysis reveals correlations that suggest moths employ parallel strategies for stabilizing posture and gaze dependent on the spatio-temporal content of the visual scene. The inherent instability in pitch dynamics necessitates these dual strategies. Distinguishing between and reacting to wide- and small-field visual stimuli are necessary parts of many animal behaviors; through this research we hope to better understand how biological systems assimilate motion and mechanical cues to coordinate effective movements.

Flowering plants exhibit striking diversity in their floral symmetry due to independent genetic changes throughout their evolutionary history. Diversity is also observed within the flowers of genus Rhododendron, which displays a variety of radially and bilaterally symmetric flowers across its approximately 1000 species. Floral symmetry is governed by an interaction between one TCP transcription factor, CYCLOIDEA (CYC), and two MYB-class genes, RADIALIS (RAD) and DIVARICATA (DIV). This interaction initiates when CYC is expressed in the floral meristem, leading to the activation of RAD in the dorsal domain. RAD then antagonizes DIV activity in the dorsal regions, leading to radial symmetry. In other flowering plants, duplications in CYC have been associated with changes in floral symmetry, but the role of RAD and DIV remain unstudied. A number of transitions to and reversals from radial symmetry have occurred throughout Rhododendron from a bilaterally symmetric ancestor. Focusing on RAD and DIV, the goal of this study is to better understand the evolutionary history of these floral symmetry genes in rhododendrons, and their correlation to floral symmetry changes. I sampled eight different species throughout Rhododendron in addition to various outgroups and reconstructed phylogenies of these genes. I obtained sequences from genomic and transcriptomic databases, and used these sequences to design primers for amplifying and sequencing these genes from laboratory samples from wild and cultivated specimens. I found two distinct copies resulting from one duplication, in both RAD and DIV. In future studies, we will expand our sampling of species to investigate the phylogenetic placement of the origin of these duplications in tandem with changes in flower symmetry, to determine whether duplications in RAD or DIV are associated with symmetry changes as has been shown for CYC.

Palms are a keystone species in tropical rainforests and are thought of as a model species due to their abundance and taxonomic diversity. Because over 90% of palm species diversity is concentrated in the tropical and subtropical biomes today, palm fossils are traditionally interpreted as indicators of warm and wet climate. However, recent work suggests that, in the past, palms may have played an important role also in other biomes. Palms produce great amounts of phytoliths (hardened silica bodies precipitated in and around plant cells), which, after the plant dies, are incorporated into the soil or sediment and preserved as fossils. Importantly, phytoliths are found in many environments where leaf impressions and other macrofossil are typically absent. Palm phytoliths are therefore a valuable potential source of information to reconstruct phylogenetic relationships and interpret the ecology of fossil palms and, by inference, paleoenvironments.Unfortunately, very little work has been done to comprehensively assess how much information about palm taxonomy and ecology can be inferred by palm phytolith morphology. To remedy this, we are conducting a morphometric study of palm phytoliths across the palm phylogenetic tree. We collected leaf samples from 80 palm taxa from herbaria and botanical gardens, then extracted phytoliths using strong oxidants and acids to decompose organic materials. Through measurement of their overall shape and size, as well as the density, size, and shape of phytolith ornamentation we hope to improve overall taxonomic resolution of the palm fossil record. In addition, for each of the taxa we recorded several habitat, climate, and ecology variables to test for biogeographical and ecological signal in palm phytolith morphology.

TOPLESS (TPL) is a transcriptional co-repressor protein that plays a central role in the regulation of plant growth and development. In the model plant Arabidopsis thaliana, TPL acts in essential hormone response pathways, including that of auxin, a small signaling molecule. TPL acts in the auxin response pathway by binding another co-repressor from the Aux/IAA family (henceforth referred to as IAAs). In the presence of auxin, IAAs are degraded, relieving TPL repression and allowing for transcription of auxin responsive genes. Despite its central role, the molecular mechanism by which TPL confers repression is not well understood. Recent structural analyses indicate that TPL has interfaces for both homodimerization and homotetramerization. Synthetic assays have shown that TPL truncations in which the tetramerization interface has been deleted have a significant decrease in repressive function. To determine the repressive mechanisms of TPL, we created a structure-function map. We have created full-length TPL variants with point mutations at the dimerization and tetramerization interfaces, as well as serial domain deletions. We tested the repressive function of these variants in a yeast synthetic assay in which TPL represses expression of a fluorescent protein. Repressive function was then quantified from fluorescent reporter output with stronger repression corresponding to decreased fluorescence. We then validated the results of our synthetic assays in planta by expressing TPL-IAA14 variants that negatively regulate lateral root development. The repressive strength of each TPL variant will be quantified by measuring the number of lateral roots, with fewer lateral roots corresponding to increased TPL repressive strength. TPL homologs exist in many species. Defects in TPL homologues have been implicated in the causes of many cancers and developmental diseases, therefore a better understanding of the functional mechanisms of TPL will have broad implications across organisms, including humans.

Auxin is a hormone that is crucial for plant growth and development. Auxin promotes the activity of auxin responsive transcription factors (ARFs) that induce transcription of target genes. Though, we know a lot about auxin response in the model plant Arabidopsis, very little is known about ARF behavior in maize. It has been previously shown that auxin and ARFs mediate many diverse developmental responses in Arabidopsis and maize. We hypothesize this diversity of response may be caused by different ARFs binding to different promoters, allowing different genes to be activated. The goal of the project is to examine the activity of maize ARFs on different promoter structures. We tested if maize ARFs show similar rules of ARF-promoter interactions as Arabidopsis ARFs. By comparing Arabidopsis and maize ARF activity, we are be able to understand how the two species have evolved over time, and if ARF function and auxin response are conserved in the two species. Working on maize is also important because it such an essential food crop for people and animals and can help us resolve issues such as food shortage. To investigate how ARFs regulate the activation of genes, we used a synthetic yeast system to quantify transcriptional activation. We engineered yeast to express both a maize ARF and a synthetic auxin-responsive promoter regulating transcription of a fluorescent reporter. We quantified fluorescence, a readout of ARF activation of the promoter, by flow cytometry. We systematically tested how activation is impacted by the sequence, orientation, and number of cis-elements within a promoter sequence, and how these effects differ among different ARFs.

A seed’s successful transition to a seedling depends on the suitability of the surrounding environment. All plants need water, sunlight, favorable temperatures, and soil nutrients to survive. On top of these essentials, interactions with other organisms through competition, herbivory, or disease may also have an impact. The goal of this study is to assess which factors influence tree germination in Washington’s temperate forests. Are there essentials without which seeds fail to complete the pivotal transition between embryo and tree? This question was explored using data collected since 2014 in a 25.6 hectare plot located in the Gifford Pinchot National Forest (WA). The dominant trees here, including Western hemlock and Douglas fir, were the focus species of this study. Forty seed traps distributed throughout the plot collected falling tree seeds so that they could later be identified to species and counted in the lab. Germinants and seedlings were recorded within a square meter adjacent to each seed trap, along with environmental factors such as soil moisture and sun exposure. Regression analyses were used to assess whether germination rates (the proportion of seeds that successfully germinated) were influenced by environmental factors, other organisms, seed densities, or year. While analyses and data collection are ongoing, results thus far indicate a negligible role for soil moisture and sunlight in germination. On the other hand, the presence of other vegetation decreased germination, while the presence of nurse logs increased it. Germination is an important and delicate life stage for every tree, and through their influence on it environmental factors may shape the species composition of these communities. Identifying relevant factors may also provide some ability to anticipate how these trees will respond to the warmer and drier conditions expected to define climate change in the Pacific Northwest.

Manila clams, Ruditapes philippinarum, are a widely used species in Washington shellfish aquaculture. Juveniles in particular have high mortality rates due in part to predation, but the extent of this predation is unknown. This study investigates the prey size range and preference shown by the crabs Hemigrapsus nudus, Metacarcinus gracilis, Cancer productus, and Romaleon antennarium when preying on R. philippinarum. To test if crabs prefer to prey on larger clams due to their higher food content, we placed crabs and clams together in microcosms and measured mortality rates of the clams. It was found that all crabs could eat all tested clam sizes. Cancrid crabs preferred to eat larger clams; R. antennarium ate the most clams. In contrast, H. nudus had no preference for a particular clam size. Based on these results, countermeasures against juvenile Manila predation should have a special focus on cancrid crabs, and further research on the reasons behind the trend of cancrid preference for larger juveniles should be considered. 

Non-vascular epiphytes have been determined to grow in a non-random distribution in the canopies of Acer macrophyllum in the Hoh Rainforest, Washington. Nitrogen (N) source has been suggested as a potention driver of the distribution, and stable isotopic analyses were used to consider the influence of this factor. Three epiphyte species, the lycophyte Selaginella oregana, and bryophytes Rhytidiadelphus loreus, and Neckera douglasii, and four trunk dwelling species, Selaginella oregana, and the bryophytes Hypnum subimponens, Leucolepis acanthoneuron, and Metaneckera menziesii, were selected for analyses on the basis of their distribution on five host trees. Samples of plant tissues, as well as canopy and trunk soil were analyzed using isotope ratio mass spectrometry to identify the N source of each species. N source varied significantly by species and growing location. δ¹⁵N values of all bryophyte species confirmed an atmospheric N source. Results for the lycophyte S. oregana varied significantly in response to its relationship with R. loreus, which grows above it in mats in the inner canopy. Selaginella oregana appears to use canopy soil N in the presence of R. loreus, and atmospheric N in its absence on the trunk. These findings suggest the role of niche partitioning in the coexistence of S. oregana and R. loreus within the canopy, and illustrate the likely ability of S. oregana to vary its N source based on competition for the resource. No study of this kind has been conducted in the Hoh Rainforest to date, and this work highlights the intersection of abiotic and biotic factors in determining epiphyte species distributional patterns in the temperate forest ecosystem. 

Active learning has often been described as the key to increase retention rates among undergraduate STEM majors because students are engaged in meaningful learning. Common activities associated with active learning include completing worksheets in groups, clicker questions, or discussion questions followed by reporting out by random call during class. Random call differs from cold call as the instructor allows discussion time where students can work together to formulate an answer prior to random call. However, previous research has demonstrated that implementing active learning principles may increase student anxiety. Specifically, answering questions in class, either by volunteering or random call, may cause the highest level of anxiety for students. We have conducted a study investigating how student demographics impact the likelihood of students responding to random call. Our study was conducted in an Introductory Biology course with 576 students, in a lecture-hall style classroom. The instructor utilized random call, then recorded which students either gave a response or passed the question when called upon. A binary logistic regression was performed to create a best fit model in which student demographics could predict which students were more likely to answer or pass. Students who are the first in their family to go to college (1st generation students) were 57% less likely to respond to random call questions compared to other students, even when controlling for incoming GPA. This result indicates that students with diverse demographics participate differently in some active learning activities and further investigations are necessary to determine if this has an impact on their learning. To bridge this “responding” gap between 1st generation and Non-1st generation students, instructors should implement additional techniques such as group random call as opposed to individual random call to increase student comfort during random call, thereby increasing the likelihood of students responding regardless of student demographics.

The hindbrain, including vital sensory and motor neurons, develops into the pons and medulla of the brainstem. During development, waves of spontaneous activity (electrical activity present without stimulation; SA) influence neuronal circuitry and change spatiotemporally in a characteristic pattern. The upregulation of a resting potassium channel, KCNK9, mediates the gradual decrease in SA in the hindbrain, beginning at embryonic stage (E)11.5. Cannabinoids derive from marijuana and act on CB1 receptors in the central nervous system to affect motor control, mood and memory through altered neurotransmitter release. Previous reports in the literature have demonstrated that AM251, a CB1 antagonist, binds to KCNK9, opening the channel. The goal of my research project has been to use intracellular calcium imaging to test the hypothesis that cannabinoids act on the KCNK9 channel to alter or regulate spontaneous activity. This involves determining whether cannabinoids affect hindbrain SA by utilizing the CB1 receptor or by binding directly to the KCNK9 channel. To test this, I used both AM251 and a CB1 agonist, ACEA (arachindonyl-2’-chloroethylamide). I predicted that AM251 would open or augment current through KCNK9 channels, either by binding directly to the channel or via CB1 receptor binding, and would slow SA by hyperpolarizing the cells; in contrast, I predicted that ACEA would decrease KCNK9 current, increasing SA. Increasing concentrations of AM251 (ranging from 200 nM to 2 µM) do indeed decrease SA; the effect of ACEA is less clear and requires further study. Should a clear effect on hindbrain SA be found, this will demonstrate that embryonic spontaneous activity can be modulated by use of cannabinoids, perhaps altering the neuronal circuitry during development.

Computer science and neurobiology have an undeniable entanglement with one another in the 21st century with the advent of advanced artificial intelligence and attempts to understand the black-box model of a brain. However, most neurobiology research has revolved around understanding existing behaviors in large animals, with commonly researched organisms including zebrafish and mice. Introducing novel behaviors to animals in a controlled environment is undeniably attractive in modern neurobiology. With the use of mass data-collection in the online Aquarium lab environment, this study shows the potential to model and produce novel behaviors in Hydra vulgaris (freshwater anemones with simple nervous systems). We aim to create a high-throughput, data-oriented, model for gene-based behavior analysis and editing in neurobiology research. By creating a single controlled environment to perform all research jobs on mass-produced H. vulgaris strains with clockwork husbandry, extraneous factors affecting the organisms are significantly reduced. By taking data on the success of every modification of these Hydra, we can model the effects of specific factors on these animals and progress toward creating a model organism for neurobiology research akin to knockout mice in medicinal genetics. With these methods we have been able to produce a significantly more efficient process of gene editing in Hydra and with several different kinds of DNA integrants. Over the long term, the wide availability of the online Aquarium lab can yield the discovery of unknown trends in genetic knockouts and the discovery of novel behaviors through analysis of all data collected in this controlled environment. Here we highlight successes thus far and future developments necessary to make this system a viable model for gene-based neural engineering over this century.

In a warming world, where growing human population continually intensifies demand for agricultural yield, studying genetic and molecular mechanisms underlying plant development and morphology becomes increasingly critical. As in other multicellular organisms, development, growth, reproduction and stress-response in plants are regulated through a complex network of signaling pathways, which, though spatially tiny, have a mammoth impact on organismal-level fitness. In the model angiosperm Arabidopsis thaliana, a symphony of such signals coordinates during early development to dictate the patterning of stomata, small epidermal guard cell–pore complexes that facilitate gas exchange vital for driving photosynthesis and thus biomass production. Several tiers of signal cascades, each executed by a distinct group of molecular players, regulate this process to prohibit two adjacent protodermal cells from becoming stomata, creating a spacing pattern critical for plant fitness and thus our sustenance. In one such tier, perception of several characterized members of the Epidermal Patterning Factor (EPF) peptide family by transmembrane receptor like kinases tunes downstream signaling that regulates stomatal cell fate. EPF1 and EPF2 act in this manner to inhibit a cell from advancing towards a stomatal fate, while EPFL9 acts to promote it, but actions of several other EPF members remain unknown. One such “mystery,” EPFL7, shares closest sequence homology with EPF2, and curiously is expressed in vascular tissue during early plant development. As the epidermis above vascular tissue in Arabidopsis and most other vascular plants is regularly devoid of stomata, it is possible that EPFL7’s expression confers an arresting signal similar to that of EPF2, preventing differentiation of protodermal cells above vasculature into stomata lineage cells. My project evaluates this and two other possibilities: EPFL7 functions in the stomata development pathway, either (1) inhibiting or (2) promoting it, or (3), EPFL7 may be involved in other processes, such as vascular development.

Incorporating research into the undergraduate experience provides students with opportunities to develop transferable skills by engaging in the process of science. Historically, most students who have participated in research did so through apprenticeships in faculty research labs. However, the apprenticeship model limits the numbers of students who have access to authentic research experiences early in their academic careers. In contrast, Course-based Undergraduate Research Experiences (CUREs) engage larger numbers of students in firsthand research investigating questions that are relevant to the scientific community. CUREs provide access to undergraduates who might not otherwise have the awareness, confidence or social capital needed to pursue scientific research opportunities through traditional pathways. The University of Washington (UW) Introductory Biology CURE provides students with authentic research experiences studying the evolution of antibiotic resistance in E. coli through credit-bearing lab sections. We investigated the CURE’s impacts on students’ personal development as scientists along with their mastery of fundamental biological concepts and principles of experimental design. At the beginning, middle, and end of the first two quarters of the Introductory Biology series, CURE and non-CURE students completed concept surveys to detect changes in their: (1) attitudes about science and their own abilities, (2) competency in experimental design, and (3) conceptual understanding of key evolutionary concepts, such as the relationship between genotype, phenotype, and fitness. Although all students in this study attended the same lectures, we anticipate that students who participated in the CURE lab modules will attain greater gains relative to their peers enrolled in the non-CURE labs. Specifically, we predict that CURE students will demonstrate greater understanding of the mechanisms of evolution via natural selection, and improved ability to design controlled experiments and reason scientifically. In addition, we expect that alumni of the CURE will gain an increased sense of belonging and enhanced retention in science-related fields.

Flowering is crucial to a plant's lifecycle because it determines reproductive success for the plant. Scientific understanding of the molecular mechanisms that direct this process are constantly being revised and expanded as more is discovered about how plants sense their environment and adjust their flowering time in order to optimize reproductive success. In this study we explore the molecular mechanisms involved in the FKF1 photoreceptor signaling pathway. FKF1 is a blue light sensitive photoreceptor that influences FLOWERING LOCUS T (FT) gene expression, which induces flowering in plants. The molecular mechanism of activation of FT expression under high light has not yet been fully characterized. By comparing relative gene expression in cat2 mutants we found six transcription factors that have the potential to bind to the FT promoter and show significantly higher expression levels under high light conditions. Our research focuses on these six transcription factors and their potential impact on flowering time in the plant. To study these transcription factors, we are developing transgenic lines to analyze the effects of knocking down or overexpressing these genes. We will look at the resulting phenotypes of our transgenic plants in order to develop models that explain the suppressive or activating effects of our transcription factors. This will further our understanding of how plants regulate flowering under high light conditions. By achieving a deeper understanding of plant development, we can devise ways to maximize agricultural output in order to feed an ever growing global population.

Pooideae is the largest grass subfamily, comprising over one third of all grass species. Pooid grasses are dominant in C3 grasslands which were once more widespread, but today are restricted to temperate or high elevation areas. During the last 8 million years, many C3 Pooideae were replaced by C4 grasses as regional climates became hotter and drier at low- to mid latitudes, or by forests when local conditions became wetter. Understanding when and where C3 grasslands first formed and when they were replaced by tropical C4 grasslands or forests is important for reconstructing global ecosystems and tracking climate change through time. Since grass macrofossils are rare, microfossils, particularly phytoliths (solid silica particles formed in plant tissue), which can persist in soils for millions of years, are the best paleontological evidence for the presence and abundance of different grasses through time. Grass phytoliths form highly regular shapes which may be specific to clades at low taxonomic levels; however, phytolith shape variation in Pooideae has not been analysed using quantitative methods. To better understand differences in shape within the subfamily, we extracted phytoliths from 36 representative Pooideae species. The phytoliths were imaged using confocal microscopy, and processed into 3D surface meshes for geometric morphometric analysis. We used the meshes as a reference collection to compare similarly processed fossil phytoliths from paleontological sites. Our goal was to use quantitative methods to confirm that the fossil phytoliths were correctly assigned to Pooideae and then to classify the phytoliths to lower taxonomic levels when possible. We used these results to infer a new time sequence for Pooideae evolution. In the future, this reference collection could be used to study domestication and spread of agriculturally important species in the subfamily (e.g., wheat, rye and oats).

Understanding the principles that underlie physiological problems can help students succeed in the mastery of a wide range of physiology topics. Understanding the principle of flux, which governs pressure gradients, can greatly improve a student’s ability to solve problems related to bulk flow and pressure in a variety of physiological systems. In general terms, the principle of flux describes the rate of movement of a substance from one compartment to another. It is defined as the magnitude of the driving force (gradient) divided by resistance. We examined how students in an introductory physiology class think about flux in the context of a pressure gradient within the cardiovascular system. We began by incrementally refining a variety of gradient-related questions through successive rounds of think-aloud interviews with individual student volunteers. Using these responses, we generated a cardiovascular pressure gradient question that elicited a wide variation of student answers. We collected responses from 200 introductory physiology students both before and after relevant course content to be sure our question captured both novice and advanced reasoning. We examined the responses and constructed a rubric to categorize student responses into a hierarchy of response sophistication and completeness. Generally, level 1 represents the most novice responses, while level 5 represents the most advanced. In our rubric, level 1 answers relied on vague reasoning relating to homeostasis, equilibrium, or intuition. Responses categorized at levels 2 and 3 showed an understanding of the components that contribute to pressure without successfully identifying how they connect to changing pressures. Lastly, answers at levels 4 and 5 showed an understanding of these components as well as the ability to successfully relate them to changes in pressure. Identifying patterns in student responses to a pressure gradient problem can give instructors a starting point from which they can target common misconceptions to ensure conceptual mastery in their classrooms.

Plasma, the fourth state of matter, is created by introducing a gas - in this case, helium - to an electric field. This field ionizes the helium atoms, which in turn ionize other molecules, creating a wide variety of highly reactive species. Previous work at Edmonds Community College demonstrated the efficacy of an atmospheric pressure plasma jet (APPJ) in killing endospores of Bacillus atrophaeus for the purpose of spacecraft sterilization for NASA (Bernard et al., 2017). With increasing rates of antibiotic resistance, there is interest in using an APPJ in healthcare settings to eliminate harmful pathogens and therefore promote wound healing. However, given its destruction of endospores, it is unclear if the APPJ would damage living tissue. The purpose of this research was to analyze the effect of an APPJ on multicellular eukaryotic organisms. The impact of different exposure times of the APPJ upon both germinated and ungerminated Zinnia elegans seeds, and the effects on plant growth, was studied. The optimal exposure time for germinated seeds was between one and two minutes, while treatments greater than 2 minutes may increase germination rate.

In plants, the hormone auxin plays a crucial role in the regulation of many different genes during growth, embryogenesis, differentiation, and many more developmental processes. The promoters of auxin responsive genes are bound by a transcription factor called an Auxin Response Factor (ARF), which is repressed by a protein complex consisting of a linker protein called an IAA and a corepressor known as TOPLESS (TPL). In the presence of auxin, the IAA is degraded resulting in the dissociation of TPL and subsequent activation of transcription of these auxin responsive genes. IAA20, IAA30 and IAA31 are members of a closely related sub-clade of IAAs that are known to be auxin insensitive. These genes are transcribed at low levels in nearly every cell type, yet their role in the auxin signaling network is completely unknown. I hypothesize that the auxin insensitive IAAs create an auxin concentration threshold to reduce the noise of auxin signaling output. In this model, auxin insensitive IAAs interact with all ARFs to generally increase the concentration of auxin required to elicit a transcriptional response. To test my hypothesis, I examined phenotypic changes in single, double and triple-mutants for the auxin insensitive IAAs, which I have generated via CRISPR/Cas9 mutagenesis combined with existing mutations. I am currently testing whether these mutants have quantifiable auxin-related defects in the highly stereotyped root development and arrangement of organs along the stem, or phyllotaxy. If I observe that mutants in the auxin insensitive IAAs have defects with these processes, it will suggest that these IAAs buffer noise present within the auxin signaling network. This work will enhance our understanding of how auxin coordinates transcriptional repression and how signaling noise is reduced in a multi-component molecular system.

The acrosome is a sperm-specific organelle required for the fertilization of the ovum in most animals. However, the molecules required for its formation during development are largely unknown. Toward identifying the function of candidate proteins, we are testing a system known as deGradFP in Drosophila melanogaster which targets fluorescently-tagged fusion proteins for degradation. This system depends on the ubiquitin-tagged proteasomal degradation pathway. Currently, we are testing whether this system works at different stages of spermatogenesis by targeting Yellow Fluorescent Protein (YFP) tagged Rabs, a small GTPase required for vesicular trafficking. Our preliminary results suggest successful targeting of the YFP-tagged Rab6. We are currently testing additional YFP-tagged Rabs, as well as sperm-specific proteins. The results should reveal whether the proteasomal pathway is working in all stages of spermatogenesis and aid in the understanding of proteins required in the acrosome biosynthesis pathway. Overall, our findings will expand our knowledge of sperm formation and function.

Keeping exotic pets carries considerable risk for both the animals and their owners. Unlike with domesticated pets, acessibility to appropriate veterinary services and adequate care information for exotic animals are often minimal and difficult to obtain. Exotic animal laws vary considerably state-by-state, and even county-by-county; leaving generous loopholes for irresponsible and impulsive animal ownership. In this literature review, I've surveyed the physical and psychological impact that captivity has on exotic pets, as well as the health and safety risks posed on humans by the prevalence of zoonotic disease. I have also investigated the plausible options for decreasing the popularity of exotic pet ownership. High mortality rates, inadequate nutrition and living enclosures, and poor socialization have demostrated that exotic animals are at significant risk of not receiving the care they require. Literature on various exotic animals laws from across the United States show that enforceable, well-communicated laws, as well as easily accessible information on exotic animals concerning their associated diseases can be effective in discouraging people from owning exotic animals. Cohesive interstate laws are needed for easier enforcement, and legally requiring pets wholesalers to explicitly publicize the health and safety risks of owning exotic animals would go a long way in decreasing the popularity of owning exotic pets. 

Global climate change and population growth pose a significant risk to global food security. Estimates suggest that animal products alone, such as animal feed, must increase by 60-70% by 2050 to keep up with population growth. Crop species are needed to feed people directly and to feed livestock. How crop species respond to predicted climate change compounds the complexity of this challenge. Phytochrome B (phyB) is a photoreceptor that affects the plant’s ability to respond to changing resource availability. Plants with a reduced phyB function make less seeds, whether measured in the model plant, Arabidopsis thaliana, or in the crop, Brassica rapa. The reason for this seed yield phenotype is unknown. Characterizing reproductive structures in Brassica rapa from before fertilization through the production of viable seeds to identify the role phyB plays in seed production will give insight in this process. Doing this may ultimately lead to the identification of potential targets for engineering higher yielding oil seed varieties.

What drives change in populations of endangered species such as the northern spotted owl (Strix occidentalis caurina)? Despite northern spotted owl populations in Washington declining yearly, variability in year to year population performance suggests that understanding what facilitates a “good” year for this endangered species may improve management practices. It is possible that resource pulses, distinct increases in ephemeral resources occurring at infrequent time intervals, influence this variation. “Masting” is a type of resource pulse in terrestrial ecosystems characterized by large increases in seed or fruit production, which may raise abundance of small mammals that consume seed. For example, northern flying squirrels (Glaucomys sabrinus), have been found to respond positively to mast events from the previous autumn. Because northern flying squirrels are a primary prey item of S. occidentalis, constituting up to 81% of its diet by biomass, spotted owl numbers and seed masting events may be correlated. To elucidate this relationship, I used seed production data collected in Mt. Rainier National Park (MORA) since 2008 by the Hille Ris Lambers Lab, in conjunction with northern spotted owl demographic data from the National Park Service. An estimated 80,000 acres of suitable spotted owl habitat exist in MORA extending to altitudes of 4,800 feet, dominated by Psuedotsuga menziesii, three Abies species, Thuja plicata, and Tsuga heterophylla — trees which mast and may be consumed by small mammals. We hypothesize that a mast event will produce a resource pulse of seeds which may heighten flying squirrel abundance, resulting in increased northern spotted owl population two years after the seed mast event. This hypothesis predicts a positive correlation between seed production and adult northern spotted owl population two years post mast event. This study will improve knowledge of spotted owl population dynamics and potentially improve management protocol of Strix occidentalis caurina in MORA.

The behaviors of female mosquitoes related to disease transmissions such as host-seeking and egg laying are heavily studied. It is also known that both male and female mosquitoes acquire sugar from plants. However, mosquito-plant interactions are relatively understudied even though both male and female mosquitoes rely on plants for sugar. This mosquito-plant interaction allows for mosquitoes to act as potential pollinators. Some researchers have argued that mosquitoes are generally nectar robbers, while other studies have shown mosquitoes can indeed contribute to pollination. However, their significance as pollinators has not been explored. Here, we discuss a species of snowmelt mosquito, Ochlerotatus communis, a major pollinator of Platanthera obtusata, the blunt-leaved rein orchid. Orchids utilize pollinia, or pollen sacs, which attach to stereotyped locations on their pollinators. This consistency ensures successful pollination on subsequent visits. In order to maintain this consistency, the morphology of mosquitoes should match that of the flower for successful pollination leading to higher reproductive success for the plant. Through multivariate linear measurements of mosquito head characteristics, we found that the proboscis length was a distinguishing character that accounts for the morphological variation among different species of mosquitoes that are sympatric with the P. obtusata. Our finding indicates that there is a possible morphological match and suggests that there is a possible match with the nectary length of the orchid and morphological characters of the mosquito. Our study demonstrates the first-ever analysis of mosquito morphology within the context of pollination. To further test this, additional studies can be done on verifying the significance of the lengths of the proboscis and nectary for successful pollinia removal.

There are more than 3,500 species of mosquitoes and within this group, there can be a very diverse range of host preferences. However, very little is known about the decision-making process in choosing a host. The main question behind our research is: do different species of mosquitoes learn odors associated with their preferred host? It has been previously shown that Aedes Aegypti have the ability to learn some odors and not others, but it was unknown whether the ability to learn odors varies across mosquito species. To test this, females of each species were trained using classical conditioning utilizing an aversive procedure. Learning was then tested in a Y- Maze two-choice test, using 3 different odors; octenol, hexanoic acid, and linalool. We also tested whether a dopamine agonist abolishes this learning. Our results showed that learning ability is related to known host preference; mosquitoes learned the odors associated with their preferred host. Understanding more about mosquito host preference will be beneficial to understanding patterns of mosquito transmitted diseases and impact of mosquitoes on global health.

In many plant species, temperature and herbivory have been shown to affect photosynthetic rates, and thus, capacity for growth. However, few studies have looked at  the combined effects of temperature and herbivory on plant performance. This is important, as increases in temperature and rates of herbivory are predicted to coincide with climate change. To address this issue, Lupinus latifolius, a perennial plant native to the western coast of the United States, was studied. L. latifolius was chosen because it is a nitrogen-fixer, and it can facilitate the growth of other plant species in disturbed habitat. As such, observing how L. latifolius responds to future conditions we expect with climate change can provide insight about how plant communities will respond in the future. In this experiment, two growth chambers were used, and L. latifolius plants were placed in one of four treatment groups. The treatment groups were organized so the combined effects of temperature and herbivory could be compared to the standalone effects of temperature and herbivory. For the herbivory treatments, grasshoppers were used. MultispeQ, a portable plant-phenotyping instrument, was used to capture photosynthetic parameters and relative chlorophyll. Certain morphological traits, such as plant height and leaf mass per area (LMA), were also measured. I expect that higher temperatures will result in higher photosynthetic rates, but that herbivory will have a larger negative impact on photosynthesis. As a result, lower photosynthetic rates are likely to be observed under increased temperature and herbivory. T tests and ANOVA were conducted to determine whether differences between treatments are statistically significant. By conducting this research, we can gain a better understanding of how changes in temperature and herbivory rates associated with climate change can interact to affect plant performance and health.

Symbiotic relationships are vital to the survival of a vast array of organisms. A well-documented set of symbiotic relationships in the marine environment are between bivalve mollusks and the variety of epibionts that encrust their shells. Marine sponges are some of the most frequently observed epibionts of bivalves, and can confer both positive and negative effects on their hosts. The Puget Sound sponge species Myxilla incrustans lives almost exclusively on the valves of the scallops Chlamys rubida and Chlamys hastata. Studies have found that this symbiosis is mutualistic, with both the scallop and the sponge acting as predator deterrent for the other, among other interactions. Additional research has shown that presence of sponge does not significantly affect the swimming distance of scallops, however little has been done to look at the effect of different predators on scallop swims, as different scallop species may invest different amounts into protecting their sponge symbionts. This study aimed to evaluate the responses of two species of scallops to two species of predatory sea star to determine if they react differently and if one species of scallop is a better host to the sponge. Scallops were placed in tanks with either the sponge predator Pteraster tesselatus or the scallop predator Pycnopodia helianthoides and the scallop-sea star interactions were recorded. Time and distance of swims were measured for each scallop swim and were categorized by what initiated the swim. Distance and duration of swims were used to evaluate the response of the scallops to different predators to determine if there was a difference between the two species.

Changing developmental patterns is an essential strategy for plants to survive environmental stressors, and the hormone auxin coordinates much of plant development. In the auxin signaling pathway, Aux/IAAs are co-repressors that block expression of auxin target genes. When auxin is present, Aux/IAAs are ubiquinated by an E3 ligase and then degraded via the proteasome. This results in an increase of auxin response gene expression. Past research in the Nemhauser lab found that a dominant mutant in the Aux/IAA known as IAA28 alters the placement of lateral organs around the stem (a process called phyllotaxy). IAA28 loss-of-function mutants show no phenotype, so we hypothesized that another gene or genes could be acting in a redundant manner. IAA28 is closely related to two other Aux/IAAs called IAA18 and IAA26. Using CRISPR/Cas9 technology, I am making triple mutants that lack function of IAA18, IAA26, and IAA28. Once I have these mutants, I will measure phyllotaxy and other developmental phenotypes to look for differences when compared to wild-type plants.

Fecal hormone analysis has been extremely useful in evaluating the physiological condition of wildlife populations. One of the most important steps in the testing process is sifting fecal samples to remove undigested materials. This allows the sample to be more easily homogenized so that the subsample to be extracted has a uniform hormone distribution. Sifting can be quite time-consuming for predators whose scats contain large amounts of hair and bone. It could take over an hour to process each of these scats by hand. Currently, we have thousands of samples to analyze and were able to increase the process by tenfold with a new sifting tool. Using radioimmunoassay, which measures the concentration of hormone metabolites, we validate the high-throughput sifting tool by comparing hormone concentrations obtained from different subsamples. Results suggest that the new tool is reliable and has tremendous potential to reduce the time needed to process large numbers of wolf and coyote scats.

Scientists agree that recent climate changes result in decreased plant growth and agricultural efficiency. Because the timing of flowering affects plant development and biomass, it is critical to understand how environmental factors influence the flowering regulation. Photoperiod (length of a day) and temperature are the most important factors that regulate flowering timing in a model plant, Arabidopsis thaliana. In order to understand how these factors affect seasonal flowering in nature, the Imaizumi Lab grew Arabidopsis outside in Seattle, Washington during the summer solstice (16 hours light, 8 hours darkness, highest average temperature 21°C, 1:1 Red:Far-red ratio), where day-length and temperature were similar to long-day (LD) lab conditions (16 hours light, 8 hours darkness, constant 22°C, 2:1 Red:Far-red ratio). They discovered a unique expression pattern of the florigen gene, FLOWERING LOCUS T (FT), which promotes flowering. The lab also recreated the FT expression pattern by changing temperature and light settings of the LD lab conditions and found that CONSTANS (CO) and EARLY FLOWERING 3 (ELF3) act as positive and negative regulators of the FT expression. Then, we established a double mutant co101 elf3-1 to investigate the genetic relation between CO and ELF3. Since there is a known linear correlation between the number of leaves a plant produces and its developmental state of age, we used that as a proxy to estimate the plants’ flowering timing phenotype. The FT expression under the recreated natural condition was also analyzed. Taken together, we found that co101 elf3-1 flowered slightly earlier than co101. This suggests that ELF3 regulates the flowering timing through both CO-dependent and -independent pathways. To further understand how ELF3 regulates the flowering timing, we are currently analyzing flowering phenotype of a double mutant ft101 elf3-1. Our results will progress to a better understanding of the molecular mechanism of seasonal flowering in nature.