Springsnails are a broad group of highly diverse North American gastropods, with the most speciose genus being Pyrgulopsis (139 species). These tiny aquatic gastropods are often found in thermally-influenced freshwater springs, where endemism is common. Restricted occurrence and narrow habitat requirements of many species has generated conservation concern because of anthropogenic threats to spring systems and associated aquafers. The Blue Point Pyrg is endemic to Blue Point Spring in Lake Mead National Recreation Area within the eastern Mojave Desert. Two other springsnail species also occupy this unique spring. Little is known, however, about the distribution, abundance, and habitat associations of these springsnails. As a first step in a larger study investigating these parameters, we addressed methodological questions associated with the use of artificial tile samplers for long-term monitoring of springsnail abundance. We determined that the area of our tile samplers was adequate for sampling these springsnails and that our multiple, replicate tiles did not produce significantly different measures of abundance within our one meter sampling areas. These results indicate that monitoring could be accomplished with fewer tile replicates. We identified all three springsnail species within our samples and determined that identification of adult springsnails by shell morphology under magnification appears possible; we are currently developing genetic markers to confirm the morphological assessment. The relative abundance of springsnails peaked at approximately four meters below the springhead, and then dropped precipitously further downstream. Limited numbers of springsnails downstream may be related to the presence of nonnative, predatory fishes. We are currently investigating this hypothesis, and the findings of our methodological assessment have allowed us to use half of our sampler replicates in a fish exclusion experiment to assess the impact of predation.

The Lake Washington, and Sammamish basin contains a complex mix of life history strategies of Oncorhynchus nerka. These life history strategies include 1) Anadromous sockeye that remain in the lake for one or two years before migrating to the ocean and returning to fresh water as mature adults; 2) “Residual” sockeye salmon that breed with and are genetically a part of the anadromous population but do not migrate to the ocean, and 3) “kokanee” salmon that are genetically distinct from anadromous and residual sockeye. Kokanee salmon are native to the basin and are thought to have evolved when glaciers or other barriers restricted access to the ocean. Although once extremely abundant throughout the basin, native kokanee are now thought to be found only in Lake Sammamish. Large numbers of kokanee-like fish continue to occasionally migrate from Lake Washington into the Sammamish river and its tributaries. Intriguingly, these “mystery nerka” migrate and spawn later than the sockeye/residual population and may represent a fourth distinct O. nerka population (e.g. a remnant of native Lake Washington kokanee, or a newly evolved kokanee population derived from sockeye ancestors). Sockeye, residuals, and kokanee use gill rakers, bony extensions in the throat, to capture prey. In other cases where kokanee have evolved from sockeye ancestors, the number and size of gill rakers differs – a reflection of the of the different types environments they mature in and the different types of prey available in freshwater vs. saltwater. In this research we document the variation in gill raker number and length in the Lake Washington/Sammamish populations of O. nerka in an attempt to 1) investigate trophic adaptations within the basin associated with life history strategy and location of maturation, and 2) to assess the relationship of “mystery nerka” to the other populations known to occur in the basin.

The natural history and morphology of blue mud shrimp, Upogebia pugettensis, burrows indicate that they farm their burrow linings. We tested the farming hypothesis by comparing carbon and nitrogen isotope, Carbon-13 and Nitrogen-15 ratios in tissues of U. pugettensis, their commensal clams, Naearomya rugifera and Cryptomya californica, and their isopod parasite, Orthione griffenis with other species and trophic groups co-occurring in the same mudflat, including the burrowing ghost shrimp Neotrypaea californiensis and Neotrypaea gigas, benthic clams and polychaetes. Ratios of these isotopes can indicate the relative trophic positions of organisms. We found similar isotope ratios among U. pugettensis and their commensal clams and that were different from isotope ratios in other clams and trophic groups occurring outside of the burrows in the same mudflat. The common isotope ratios among U. pugettensis and its commensal clams relative to these isotope ratios in other suspension and deposit feeding species in the same community are consistent with farming of the burrow lining.

The life history traits of parasites are dependent on the host environment and host immune response. Heligmosomoides bakeri, is an intestinal nematode parasite that infects laboratory mice (Mus musculus). This investigation used rapamycin, an immunosuppressant drug, in the diet of the host. Rapamycin is currently being considered as an over the counter drug to extend life expectancy. The purpose of this experiment was to determine if inhibition of the host immune system by rapamycin had a significant effect on parasite life history traits during a secondary infection. Mice were fed either a rapamycin diet or control diet for two months. Mice were then infected with H. bakeri, drug cleared of the primary infection, then re-infected with a secondary H. bakeri infection. Starting eight days after the secondary infection fecal egg counts were done daily to determine worm reproduction in vivo. Eighteen days after the secondary infection the mice were euthanized and worms were removed, sexed and counted. Length of ten female and ten male worms from each mouse was then measured, and 10 females were cultured to measure ex vivo reproduction. After 24 hours of incubation in culture media, egg output and worm motility (an index of viability) were measured. For this experiment I was involved in recording the worm length and was responsible for performing the ex vivo egg count and motility measurements. Results will provide important information about potential over-the-counter use of rapamycin in humans to slow biological aging and how changes in immune function may affect susceptibility to parasite infection.

Age related immune decline may play a role in susceptibility to parasitic helminth infections among the octogenarian population. Few studies have examined very old age on susceptibility to helminth infections in people. Effects of age on host susceptibility using mouse models also has not been thoroughly studied for older ages of laboratory mice (Mus musculus), such as beyond about one year. In this study, the intestinal nematode (Heligmosomoides bakeri), a model for hookworm infection in humans, was used to infect 1-year-old and 2-year-old Swiss Webster mice. At 40 days post-infection, worms were removed and worm survival, sex ratio, ex vivo egg output, and worm length (as an index of growth) were measured and analyzed. I expect to find an increase in worm survival, ex-vivo egg output, and worm length in older mice. These results will be valuable because they will model the helminths capacity to proliferate in older hosts in comparison to younger hosts during a human hookworm infection.

Stem cells are known for their ability to self-renew and differentiate into specialized cells. A key factor that drives self-renewal and differentiation is the specific localization of cell-fate determinants inside the cell cytoplasm. In this work, we focused on a protein called Numb, commonly known to inhibit the Notch signaling pathway responsible for the regulation cell-fate. We sought to understand the correlation between cell spreading, morphology and Numb localization. We studied human adipose-derived stem cells (HADSCs) in two conditions: 1) Cells fully spread after 24 hours and 2) Cells that were actively spreading or retracting. In each condition we analyzed the distribution of Numb fluorescence expression inside the cell cytoplasm. We found that in cells that were allowed to fully spread, Numb was highly localized only on cells with pseudopodia (“arm-like” cytoplasmic protrusions). Numb was evenly distributed when cells were fully spread, had a large area and circular morphology. In the second condition, cells that were actively spreading or retracting were small and circular in morphology. Interestingly, these cells had a high fluorescence expression of Numb specifically localized at the cell edges. These results seem to indicate that Numb localization was found in areas of membrane activity, such as pseudopodia and spreading/retracting cells. This could be due to the fact that Numb has been found to participate in integrin turn-over (cell adhesion). Based on these results our next step will be to modify membrane activity and cell adhesion and observe the effects on Numb localization and stem cell differentiation. Regulating stem cell fate via protein localization will allow us to enhance the production of differentiated cells for regenerative medicine applications.

Glutaredoxins (GRXs) are small oxidoreductase enzymes that can reduce disulfide bonds in target proteins. The genome of the model plant Arabidopsis thaliana has more than 30 GRX genes, but the biological function of most of these GRXs is unknown. We previously found that a small group of Arabidopsis GRX genes is specifically activated by nitrate, a common source of nitrogen in the soil. In order to better characterize the function of one of these nitrate-regulated GRXs, AtGRX660, we generated transgenic Arabidopsis plants that continuously overexpress the AtGRX660 gene. We isolated RNA from 12 independent transgenic lines and quantified AtGRX660 mRNA levels via real-time reverse transcriptase PCR. Three elite lines displaying >100-fold increase in basal AtGRX660 transcript levels were selected for further analysis. AtGRX660-overexpression lines and wild-type plants were grown on soil in a controlled environment growth chamber for characterization of shoot phenotypes, and on vertically-orientated plates of plant growth media for characterization of root phenotypes. All AtGRX660-overexpression lines displayed a dwarf shoot phenotype, with significant reductions in shoot biomass, total leaf area, and silique length compared to wild-type plants. In addition, root system architecture was highly altered. While primary root growth was normal in the transgenic plant lines, lateral roots were almost completely absent. Phase contrast microscopy demonstrated that lateral root primordia develop in the transgenic lines, but these primordia do not elongate and emerge from the primary root. Overall, these results suggest that AtGRX660 acts a negative regulator of shoot organ development and inhibits lateral root elongation. Our findings could have agricultural relevance in plant drought tolerance, since AtGRX660 differentially affects primary root system growth (root system depth) and lateral root system growth (root system breadth).

Growing reliance on antibiotics in livestock production, commercial agriculture, and human healthcare has created evolutionary pressures on bacteria. These pressures have given rise to a new threat to public health, drug-resistant microbes. In addition, antibiotics eliminate all microbes, including the beneficial ones, and are not advisable for long-term use. Recent advances in the understanding of bacterial virulence via quorum-sensing (QS) has presented the scientific community with a promising new approach for alleviating the ongoing overuse of broad-spectrum antibiotics. Quorum-sensing is a form of coordinated gene expression mediated through detection of specific population density. Bacteria use QS as a way to regulate behaviors including biofilm formation, virulence, and motility. Quorum-sensing inhibition (QSI), is the ability to prevent QS and is used by organisms across kingdoms. This prevalence suggests that disrupting prokaryotic communication is either a common defense tactic against infection or acts as a competitive advantage in resource acquisition. Fungi, a kingdom in direct competition with prokaryotes are prime candidates for broadening our understanding of the mechanisms behind quorum-sensing. With its abundance of endemic fungi, the Puget Sound region provides a unique opportunity to screen species for QSI compounds. In this study, fungi representing diverse ecological niches were collected from local forests around the Puget Sound and isolated on potato dextrose agar plates. To screen fungal isolates we used C. violaceum, a bacterium that forms purple colonies when able to quorum-sense and changes to white when a QSI is present. Our results suggest fungi are an underexploited and unexplored source of novel bioactive molecules that could provide a method to inhibit virulent effects of bacteria without damaging an organism’s microbiome.

The sensory mechanisms by which organisms orient towards potential food prior to initiating movement are well-researched. However, behavioral mechanisms in stimulus-poor environments which determine foraging through seemingly stochastic movement variability or random-walk models remain poorly understood. Existing literature describe analyses of a variety of single-organism emergent movement patterns but do not appear to offer comparative analysis between single- and multi-individual environments nor within degrees of resource availability. We introduce a new analysis of observed fluctuations in spontaneous movement by Caenorhabditis elegans exposed to diverse conditions of competition and resource abundance. We describe a sequence of experiments which quantify the movement patterns of C. elegans through video imaging pattern recognition codified by run-length-time and turn-angle-time in comparison to recursively updating algorithmic position estimation. We expect to observe movement patterned on optimized explore-exploit strategies—such as simple random walk or Lévy flight—with frequency of implementation influenced by both population and resource density.

The Zea mays (maize) crop is essential to global agriculture and food security, and is also vital to the US economy. Like many crop plants, maize is often subjected to extreme environmental stresses, such as drought, extreme cold and high salinity, that negatively impact crop yield. Therefore, it is important to characterize the molecular (epigenetic) changes that occur to a gene under environmental stress conditions in order to manipulate these responses to improve crop yield under stress. We are interested in characterizing the epigenetic regulation of stress-responsive transcription factors under normal (control) and abiotic stress (drought) conditions in maize seedlings. Using bisulfite conversion of genomic DNA, we previously identified that the Zea mays basic transcription factor 3 (ZmBTF3) gene displays some variations in methylation patterns in parts of the gene promoter between the two treatments. To confirm these results, we used an alternative method: a methylation sensitive restriction digest technique, which uses methylation sensitive restriction enzymes (MSREs) that cut DNA molecules at precise locations and are sensitive to DNA methylation. Our results show some treatment (control vs. drought) and restriction enzyme (sequence/context) specific methylation patterns. Using these two independent techniques, we identify stress-responsive epigenetic variations (DNA methylation) in the promoter region of the ZmBTF3 gene, previously uncharacterized in maize. Ongoing research includes (i) characterizing the sequence contexts of these methylation patterns between the two treatments, (ii) determining if the observed DNA methylation changes correlate with transcription of the ZmBTF3 gene, and (iii) characterizing the drought response phenotype in maize plants with a mutation in the ZmBTF3 gene. This study will help us gain a better understanding of how the BTF3 gene in maize is regulated through epigenetic modifications.

The level of plastid DNA (ptDNA) declines as plastids develop from colorless proplastids to green chloroplasts. The decline in ptDNA is associated with an increase in DNA damage resulting from oxidative stress and ultraviolet (UV) radiation. 8-oxoguanine (8-oxoG) is a lesion in ptDNA that results from the oxidation of guanine. Our lab previously found a reduced amount of ptDNA in light-grown plants compared to dark-grown plants, likely due to increased oxidative stress that increases ptDNA damage. Here, our objective is to quantify ptDNA damage by assessing 8-oxoG lesions during the greening of maize leaves. We hypothesize that as plastids mature, 8-oxoG lesions increase. Our experimental outline involves the quantification of 8-oxoG by the enzyme-linked immunosorbent assay (ELISA), as well as immunofluorescence microscopy using antibodies that target 8-oxoG. We are examining plastids isolated from light-grown and dark-grown stalk and leaf tissues. As 8-oxoG lesions are one of the markers of oxidative DNA damage, our results will be used to assess DNA damage during development of maize plants. This research will provide a better understanding of role of oxidative stress in plant development

 RNA-directed DNA methylation (RdDM) is an epigenetic pathway that induces transcriptional gene silencing through DNA methylation of target genes. The Zea mays (maize) RMR1 protein has an important role in the RdDM pathway as it functions to assist the process of making double stranded RNAs, required for progression of the pathway. Abscisic acid (ABA) is a plant hormone that accumulates under abiotic stress, and can inhibit seed germination. We determine if the RMR1 protein is involved in epigenetic regulation under abiotic stress by measuring maize phenotype and identifying transcriptional changes in ABA treated (vs. control) rmr1-1 mutants. To determine the effect of the rmr1-1 mutation on maize phenotype, we are measuring shoot length, primary root length and additional roots in maize germinating seeds from a segregating family. The genotype of the individual seeds is determined by amplifying the region of the rmr1 gene that includes the point mutation (C to T) that is on the 4th exon of the gene, followed by Sanger sequencing. The genotype is then associated with the measured phenotype. We are also interested in determining if ABA-induced transcriptional factors are differently expressed in rmr1 mutant compared to wildtype under ABA, and this will be measured by quantitative reverse transcription and PRC(qRT-PCR). This research is in progress and results will be presented. Maize and other crop plants are often negatively affected by abiotic stress, therefore, understanding how epigenetic pathways are involved in these responses is important to agricultural productivity.

Zea mays (maize, corn) is an essential crop plant; important to global agriculture and the U.S. economy. However, maize productivity and yield can be drastically affected by abiotic environmental stress. Therefore, a priority for many plant breeding programs is to select for crops displaying phenotypic traits of enhanced tolerance to abiotic stress. A subset of abiotic stresses induce the plant hormone, abscisic acid (ABA). The mediator of paramutation1 (mop1) gene encodes an RNA-dependent RNA polymerase that functions in the RNA-directed DNA methylation (RdDM) pathway. The mop1-1 mutation results in the loss of DNA methylation which in turn causes a variety of genes to be expressed abnormally. We determined how a mutation in a mop1-1 affects RNA expression under abiotic stress by conducting a computational analysis of multiple RNA-seq datasets of stress-treated maize seedlings. We compared RNA-seq data from mop1-1 and WT seedlings treated with exogenous ABA control (no ABA treatment) with a publicly available dataset of WT maize plants treated with heat, cold, drought, salinity, and control (no stress treatment). Genes commonly down-regulated in the four stresses and in MOP1 WT ABA, but up-regulated in mop1-1 ABA represent genes potentially silenced under stress that require MOP1 for gene silencing. The presence of these genes in the given stress treatment allows us to identify the abiotic stress responsive genes that require ABA and MOP1 mediated regulation.

In the US more than 100 million people are living with diabetes or pre-diabetes. The economic burden caused by these conditions, including medical costs, is approximately $327 billion annually. Conventionally, transgenic Escherichia coli has been the primary source of commercial insulin production, a process that requires extensive purification to ensure shelf stability and complete removal of contaminants. This study seeks to establish an alternative mode of insulin production using polyethelyne glycol (PEG) and agrobacterium to transform the oyster mushroom, Pleurotus ostreatus, with the human insulin gene. P. ostreatus is a valuable target for genetic transformation due to its lack of endotoxins, rapid growth, and fully sequenced genome. P. ostreatus was transformed using PEG and agrobacterium with a plasmid containing the human insulin gene and a carboxin resistance gene. Transformed cells were selected using carboxin, extracted, and regenerated on plates composed of yeast extract, malt extract, and glucose (YMG). Integration of the human insulin gene in to the mushroom genome was confirmed through PCR analysis of the transformants. Successful transformation of P.ostreatus offers a new avenue for insulin production, potentially diversifying the market and treatment options for diabetics.

In Zea mays (maize), MEDIATOR OF PARAMUTATION 1 (MOP1) is involved in paramutation and epigenetic regulation of gene expression. MOP1 in an RNA-dependent RNA polymerase that functions as part of the RNA-directed DNA methylation (RdDM) pathway, by producing double-stranded RNA molecules that are further processed into small interfering RNAs (siRNAs). These siRNAs are able to trigger DNA methylation which can decrease or silence gene expression. The MOP1-FLAG transgene was created by overexpressing the mop1 gene with the FLAG epitope tag. As a consequence of using agrobacterium-mediated transformation, the chromosomal location of the MOP1-FLAG transgene is unknown. To identify the sequences flanking the transgene, we used an adapter ligation-mediated PCR and cloning technique, as described by O’Malley. To do so, we first digested the MOP1-FLAG DNA with restriction enzymes designed to cut the DNA in specific locations along the genome. We then ligated adapters onto the DNA fragments. Through PCR using gene-specific and adapter-specific primers, we were then able to amplify the fragments that contain MOP1-FLAG and the surrounding genomic sequence, clone and then sequence them. We used the recovered flanking sequence to BLAST search the maize genome in order to identify the maize chromosome that the MOP1-FLAG transgene is on. Chromosomal mapping of the MOP1-FLAG transgene is useful to distinguish between the endogenous mop1 gene in ongoing complementation and chromatin immunoprecipitation (ChIP) experiments.

Dynamic coordination of animal motion depends on the interaction between the neuromuscular system and body mechanics. Changes in muscle length resulting from neuromuscular activation drive the control of locomotion. This project seeks to predict change in muscle length over time based on electrical activity in the complex physiological environment of a living organism. During energetically costly insect flight, the antagonistic shortening of the dorsal longitudinal muscles (DLMs) and dorsal ventral muscles (DVMs) deform the thorax to indirectly power wing flapping. Using electromyography (EMG), we recorded both DLM and DVM activity in vivo during tethered flight of the insect model Manduca sexta. Simultaneously, we captured high-speed video of thorax deformation to measure the change in length of the DLM muscles, which attach directly to the exoskeleton. Unlike other insects, in M. sexta there is a one-to-one relationship between muscle electrical activation and length-wise contraction, allowing direct comparison between these two temporal data sets. Using machine learning, we created a model to predict the amplitude and time course of changes in muscle length 𝓁(𝑡) based on characteristics of the EMG recording. Predicting downstream insect flight mechanics based on EMG data is an exciting application of machine learning which enables us both to better understand the factors influencing muscle length in vivo, and to connect multi-scale muscle structural data through the common link of EMG data. For example, we will predict organism-level changes in muscle length that would have occurred in existing data sets of sub-cellular muscle kinematics based on EMG data collected simultaneously.

In eukaryotes, conserved mechanisms ensure that cell growth is coordinated with nutrient availability. Overactive growth during nutrient limitation (“nutrient-growth dysregulation”) can lead to rapid cell death. Here, we find that cells can adapt to nutrient-growth dysregulation by evolving major metabolic defects. Specifically, when yeast lysine auxotrophic mutant lys- encountered lysine limitation, an evolutionarily-novel stress, they suffered nutrient-growth dysregulation, and a sub-population repeatedly evolved to lose the ability to synthesize organosulfurs (lys-orgS-). The purpose of this study is to identify the types of yeast cells that recovered by being lys- and orgS- and their frequencies through generations. Using 96-well plates, we inoculated single colonies into three separate conditions (control, +lys, +lys+orgS) to test for auxotrophs. Surprisingly, we found drastically different frequencies of lys-orgS- when lys- evolved as a monoculture in lysine-limited chemostat versus in a coculture with a lysine-releasing strain. We also found mutants defective in synthesizing glutamine and arginine. Our work suggests that under stressful conditions, multiple metabolites are released, which can facilitate the evolution of new interactions and mechanisms. 

The Puget Sound is an economically and culturally important marine ecosystem, and insects are a little understood part of this ecosystem. Research suggests that insects are an important food source for near shore juvenile salmon which has wider ecological and conservation implications, including at higher trophic levels; while some work has been done in coastal British Columbia, little work has been done on the insects themselves within the environment of the Puget Sound. To close this biodiversity knowledge gap, field and museum collection surveys were and are being conducted to gain a better understanding of the insects of Puget Sound. Various hand sampling techniques were utilized including net, aspirator and insect vacuum. So far, 15 individual species of 6 families of 2 different orders of insects were confirmed present in Puget Sound in this study. Additionally, 10 species of 2 families of 2 different orders of insects were reported in similar conditions in the literature, but were not observed in this survey or found in museum collections. The most abundant order was Diptera. In all, 14 beaches on the Puget Sound have assessed and some contained unique species. Further studies, in seasonality (and food availability), habitat substrate preference, insect behavior, insect population dynamics and a deeper look at predation of marine insects by juvenile salmon could be investigated based off the work done in this study.

How can we improve healthcare in under-served South East Asian Countries? One possibility is via mobile-health technology. Mobile-health or m-health is the practice of medical and public health supported by mobile and wireless devices. This literature review looks at over 20 years of the emergence of m-health and the prediction of future possibilities. The goal is to look at areas of improvement such as cost efficiency, adapting to the specific country’s infrastructure and integration of m-health in healthcare. In countries like Nepal, m-health has helped to eradicate diseases like malaria, improve neonatal health care and empower female health volunteers. It has also helped gather information such as statistics on usage and distribution of drugs, healthcare workers' interactions with patients and disease surveillance. With the rise of mobile users, healthcare facilities can create a structured international framework to regulate and increase the efficiency of m-health. This research will help facilitate conversations about the potential of m-health and where it needs to be improved so that it can play a pivotal role in healthcare of South East Asian countries.

Coevolution is a process where two or more species reciprocally affect each other's evolution over time. The traits of one species evolves in response to the other. This process can lead to the diversification of organisms with unique adaptive traits. Previous experiments tested whether coevolution occurred during the 2000 generations of evolution between the bacteria Desulfovibrio vulgaris and archaea Methanococcus maripaludis by pairing populations from 1000 generations with mutualist partners from their evolutionary past or future. Results suggested patterns of coevolution, but hypothesizing that those patterns occurred by accident, we tested whether or not patterns can occur that look like coevolution with microbes that could not possibly coevolve. We conducted a timeshifts control experiment using freezer stocks of D. vulgaris and M. maripaludis that evolved alone for 2000 generations and created pairings of the mutualists with five different generational pairings. They then were paired with an ancestral control group and a modern test group from the 1000th generation. Growth rates were calculated for each coculture. Graphs of the effects the 1000th generation of M. maripaludis and D. vulgaris had on coculture growth rate showed that M1000 had similar growth rates irrespective of its partners evolution while D1000 results were variable. An ANOVA test showed the mean log ratios statistically indistinguishable across all timepoints, indicating that the null hypothesis should be accepted. This result suggests that both partners have the same effects on fitness no matter how long they evolved. However, due to variations in results between D. vulgaris and M. maripaludis it is still unclear whether these populations have the ability to create patterns of coevolution due to their adaptation to their abiotic environment.

Control of the cell-cycle is a topic of fundamental importance in cell biology. We are using the unicellular eukaryotic green alga Chlamydomonas reinhardtii to better understand how DNA damage affects transition of these cells in and out of the mitotic cell cycle. We find that UV treatment of cultures of C. reinhardtii gametes (in Go) delays re-entry of these cells into the mitotic cell cycle upon addition of nitrogen to cultures of gametes, a standard treatment which releases these cells to re-enter mitosis. Using a C. reinhardtii mutant which is unable to carry out photorepair of UV-induced DNA damage, we are characterizing the degree to which the kinetics of re-entry into mitosis depends on this very important repair system; we are thus beginning the 'parsing out' of the relative importance of the various DNA repair systems in controlling the transition from Go, a non-dividing stage, to G1,  the start of DNA replication and cell division. This will potentially provide insight into other downstream effects of UV exposure, such as mutagenesis. Our work is of a very basic nature, but with application to understanding e.g. dynamics of natural populations of microalgae in a changing global environment.

Campylobacter jejuni is a Gram-negative spirochete bacteria that colonizes the guts of many bird species. They are pathogenic to humans and typically consumed with undercooked poultry or contaminated water. In this study, two species of crow, separated by 7,000 miles, have been examined for their ability to carry C. jejuni. Crow fecal samples were collected from Bothell, Washington (WA) and Kolkata, India (KK). All of the crow Campylobacter isolates were confirmed by qPCR and PCR methods to be C. jejuni. No other species are isolated. We compared these isolates by method of MLST (Multilocus Sequence Typing) and searched for known allelic profiles on the PubMLST database.Thirteen  isolates were analyzed for seven housekeeping genes : aspA, glnA, glyA, gltA, pgm, tkt, uncA. Using primers specified in the PubMLST database portions of the genes were amplified by PCR, verified by gel-electrophoresis and sent to Eurofin Genomics for sequencing. Once the sequence files were returned, they were aligned and assembled using Mega7 software. An allele number was obtained for each sequence for each sequence from PubMLST database. The allelic profile of the 7 genes for each isolate was used to obtain the Sequence type (ST) of the isolate. MLST of 15 crow isolates that represented different phylogenetic clusters based on fla-SVR sequencing, showed a majority of the WA isolates to belong to sequence types ST2678, ST9271, ST3322, ST5472. The Kolkata isolates were ST8288 and ST5472. All types were shown to occur in wild birds. One crow’s ST3174 from Kolkata has been shown to occur in human stool, while 4 isolates including Kolkata and Washington were of unknown ST’s. Although Kolkata and WA isolates belonged to different ST’s, majority were from wild birds that have rarely been isolated from humans.

Beside its academia, University of Washington Bothell (UWB) is also known for its phenomenal nature where more than 15,000 crows roost every evening, especially during the autumn and winter months, in the wetlands. Since these crows have infested these wetlands, and because free living birds can be significant contributors of antibiotic resistant (AR) bacteria to the environment they were chosen for this study. E. coli, which lives as a harmless commensal in the gut of all animal and birds, has proved to be not only an indicator of fecal coliform but also of antibiotic resistance present in the environment. This study seeks to find out the different sequence types(STs) of E. coli found in UWB wetlands using the Multi Locus Sequence Typing (MLST) technique, to find out about the lineage, and potentially how these isolates have ended up in the UWB wetlands. Antibiotic resistance possessed by the isolates was performed against 13 different antibiotics by other students in the laboratory. MLST was performed according to methods specified in the EntroBase database; Specifically 7 housekeeping genes, were amplified and sequenced. Extended spectrum beta lactamase (ESBL) and non-ESBL containing isolates were found. Two non-ESBL isolates, one from water and one from the fecal samples collected on the same date, with the same antibiotic resistant pattern, turned out to be from the human isolated clone ST 58, suggesting a link between crow and water. ESBL E.coli isolate ST 131, a highly virulent, and multi drug resistant isolate was found in 6 fecal and 1 water isolate. Therefore it is concluded that crows are potential vectors of spreading multiple drug resistant strains in wetlands, which poses health risk since these strains may be carried further during the winter months.

Reactive oxygen species (ROS) are partially reduced oxygen molecules produced during cellular metabolism in all aerobic organisms including plants. ROS derivatives such as superoxide (O2–) and hydrogen peroxide (H2O2) cause several types of cell damage in plants. Chloroplasts and mitochondria are major sources of ROS because of photosynthesis and aerobic respiration in these organelles, respectively. Previously, our lab showed that light-grown maize plants have more damage in plastid DNA (ptDNA) and mitochondrial DNA (mtDNA) than dark-grown plants and that ptDNA and mtDNA levels decline during leaf development. Here, we hypothesize that increased damage to ptDNA and mtDNA in light-grown leaves is linked to increased ROS generation compared to dark-grown and germline stalk tissues. We used absorbance- and/or fluorescence-based assays to quantify levels of ROS in chloroplasts and mitochondria isolated from leaf and stalk tissues during seedling development. Our data suggest that light-grown leaf has more ROS than light-grown stalk, dark-grown leaf, and dark-grown stalk. Our findings indicate that highly damaged DNA is a consequence of ROS generation in light-grown leaves of maize. Overall, this research will help us further understand the oxidative damage caused by various reactive oxygen species during the development of maize plants.