Antibiotic resistance is a major public health problem. For example, in people with the genetic disease cystic fibrosis, Pseudomonas aeruginosa causes chronic lung infections that resist treatment to all existing antibiotics. Gallium is a promising new antimicrobial agent that works through a novel mechanism of action. Bacterial uptake systems are unable to distinguish gallium from iron due to their nearly identical ionic radii. Once inside, bacterial gallium is thought to disrupt iron-dependent processes because Ga³⁺ cannot be reduced in physiological conditions, and iron's biological functions depend on its capacity for redox cycling. Here I sought to better understand gallium’s mechanism of action by identifying gene mutations that decrease P. aeruginosa gallium sensitivity. To accomplish this, I used transposon (Tn) mutagenesis, which transfers a genetic element to random locations on the bacterial chromosome thereby inactivating genes. Of the 295,000 mutants generated, I selected for mutants capable of growing in the presence of increased gallium compared to the parental strain. Using a procedure known as rescue cloning, a restriction digest was performed on the chromosomal DNA of these mutants to produce individual fragments. One of these DNA fragments will contain the Tn insert and chromosomal DNA flanking that insert. The Tn insert encodes resistance to gentamycin and holds the capacity to replicate as a plasmid in E. coli. I ligated the isolated fragments and transformed E. coli, selecting for gentamycin resistance. I isolated this plasmid DNA from two mutants and their Tn insertion sites (and the genes inactivated by the Tn) were identified by sequencing the DNA surrounding the Tn inserts. These genes were found to be involved in iron transport. By completing these experiments, I hope to suggest hypotheses for the roles these genes play in gallium sensitivity and to provide data useful toward guiding its development as an antibacterial agent.

Viral infection is associated with asthma exacerbations. Normally, viral presence is detected by Toll-like receptor 7 (TLR7)—an innate immune receptor in airway epithelium that responds to single-stranded RNA viruses (like influenza) and triggers an immune response to infection. TLR7 is also expressed on airway sensory nerves, but its specific role is unknown. Here, I tested the role of TLR7 on sensory nerves and identified which sensory nerve populations express TLR7 using nerves isolated from dorsal root ganglia from female Hartley guinea pigs (~400 g). Ganglia were isolated, plated onto matrigel, and treated with a TLR7 agonist R837 (0.1-100 microM) for 16 hours. Neurite number per cell was not changed by stimulating TLR7, but neurite length, and the number of branch points were significantly increased. Separate experiments measured which types of sensory nerves expressed TLR7. Vagal and dorsal root ganglia were isolated and fixed in zinc formalin. Nerves were immunolabeled with antibodies against TLR7 and either neurofilament-1 (NF-H) to identify A𝛿 fiber sensory nerves or transient receptor potential V1 (TRPV1) to identify C-fiber sensory nerves. I found that airway sensory nerves originating in the vagal and dorsal root ganglia expressed TLR7. However, TLR7 was expressed predominantly on small TRPV1-expressing C-fiber neurons, but not on large NF-H-positive A𝛿 neurons. In conclusion, TLR7 is highly expressed by airway sensory C-fibers nerves, and its activation stimulates neurite growth. These findings suggest TLR7 may increase airway C-fibers supplying the lungs which may enhance airway reactivity and be a mechanism for virus induced exacerbations of asthma.

Many genes have been found to be associated with microcephaly, a condition characterized by a smaller head size. Through exome-sequencing, we identified two de novo missense mutations in an actin polymerization gene, ARPC4, in four patients with microcephaly, mild developmental delay, and mild intellectual disability. To understand the molecular mechanisms that lead to the disease phenotype observed in the patients, I used a variety of genetic and biological assays. I began by using CADD, a measurement used to predict the deleteriousness of single nucleotide variants. The mutations identified in the patients were shown to be in highly conserved loci, indicating they might be pathogenic. We proceeded to study the functional effect of the pathogenic variants in actin polymerization. Actin is an essential component of a cell’s cytoskeleton which gives the cell its structure and aids in cell movement and cell division. I was able to establish fibroblast (skin) cell lines from two patients and perform an immunofluorescence assay staining for actin in patients and control fibroblasts. Additionally, I used western-blot, a technique used to detect a specific protein in a sample, to further quantify the amount of actin present in the patients’ cells. We were able to observe a greater abundance of actin filaments in the control sample compared to the patients’ cell. The results from this study elucidate the impact of ARPC4 in actin polymerization, establishing actin deficiency as a clinically recognizable cause of microcephaly.

Augmented reality is a promising field of study which aims to enhance our daily experiences by overlaying digital information into our view of the world, thus “augmenting” our perception of reality in useful ways. A key component of AR is computer vision which enables a computer to map and interpret the physical world. My proposed experiment will examine the effectiveness of several computer vision algorithms in balancing a pendulum attached to a movable cart. A camera, suspended by the pendulum, will collect observational data and relay this information to a microcontroller. Then I will utilize Hough Transform and Canny Edge Detection algorithms to recognize stable lines/edges in the environment, so a reference frame can be constructed in which to balance the pendulum. As the pendulum tips and the camera begins to fall, the system will compare the input frame to the reference frame and adjust the cart accordingly via servo motors. My research will compare several methods of video tracking and object recognition as well as SLAM (Simultaneous Localization and Mapping) as viable options to balance and orient systems with the real world. By designing a system that dynamically responds to its environment using visual cues, I aim to contribute to a critical component in the realization of emerging technologies such as autonomous vehicles, domestic robots and augmented reality systems.

Single electron devices (SEDs) are devices that can isolate individual electrons along a conducting path. SEDs have many proposed applications including their use in metrology, and quantum information science. However, there are many barriers that must be overcome before these applications are realized beyond the proof of concept experiments currently being performed in academic research labs. Our group’s work focuses on pathways to overcoming some of these barriers including the issues with material properties and device stability. My contribution focuses on data analysis of SED transport measurements. These analyses produce various metrics for both the SEDs under test as well as the measurement system probing them. They include determinations of charge offset drift, a metric for long term device stability, as well as electron temperature a limiting factor in magneto spectroscopy measurements.

Current studies have shown that changing climate is responsible for methane releases from the seafloor, making it important to understand how methane plumes can impact the ocean. Methane plumes impact the stratification and circulation within the water column which can influence primary productivity and seawater chemistry and potentially atmospheric greenhouse gas levels if the methane from the plume undergoes exchange with the atmosphere. Along the Washington margin there has been over 1772 individual bubble plumes located and identified in depths ranging from 40 meters to 1988 meters water depth. Compilation of archived CTD (conductivity, temperature, depth) data at active methane plume sites including Southern Hydrate Ridge, Grays Harbor, and Vancouver Island, can provide estimates of the plume fluids temperature and salinity. Using these estimates, the point where buoyancy reaches zero determines the possible depth of horizontal plume fluid intrusion into the water column and the entrainment coefficient to quantify the fluid that’s entrained by the plume can be calculated., I can determine the plumes’ ability to penetrate through sea water density interfaces such as the thermocline by calculating the Richardson number for the plumes. Using pre-established bubble models, I will also quantify sea water entrainment by the plumes. Through these methods, I will be able to determine how methane plumes impact water column stratification on the Washington Margin.

Our research seeks to more accurately define how the Pacific Northwest regional climate will look in fifty to one hundred years. Our results help quantify the uncertainty in our climate predictions, a major shortcoming in climate research. Our research is interesting because it utilizes a high resolution numerical weather prediction model to examine regional climate change here in the Pacific Northwest. We are creating a large set (an ensemble) of predictions of climate change in the Pacific Northwest. The tool used in our research is the Weather Research and Forecast (WRF) model. WRF requires General Circulation Model (GCM, also known as Global Climate Model) data to feed it information on the larger-scale climate. A big part of our project is collecting GCM data from a variety of research centers worldwide, and preparing it to be used as input data for WRF. We expect to complete our model simulations by the end of Spring Quarter 2018. These results could be of direct use to a number of local agencies and policy makers so that they can shape their decisions around the future changes that our local climate may experience. We also expect our data to be of great use for further research in quantifying climate prediction uncertainty.

How is star formation affected by its environment? How big is the effect of having differing metallicities during the star forming process? The goal of this research is to study star formation in low metallicity environments, which are well represented by dwarf galaxies. For this project, we used the Atacama Large Millimeter Array (ALMA) telescope, at an angular resolution of about 0.5 arcseconds (about 66 lys), to observe the 12CO (115 GHZ) distribution in the galaxy Henize 2-10 (He 2-10). He 2-10 is an irregular dwarf galaxy about 30 million lightyears (lys) away, about 1 kilolightyears (kly) across, and has a mass of about 10 billion solar masses. It has a high gas to dust ratio, is star forming, and has a low metallicity of about 12+log(O/H)~8.3. Additionally it contains a supermassive black hole candidate of about 1 billion solar masses, which could have significant dynamical interactions with the molecular gas. We created a zeroth moment map which we used to estimate several properties of the galaxy: including a molecular gas mass of about 38 million solar masses, an observable area of about 1850 square klys, and a molecular gas density of about 3.7x10^(-20) grams per cubic centimeter. We also created multiple channel maps which provide preliminary results that indicate the presence of at least 10 giant molecular clouds within He 2-10.  With what we have done so far, we will be able to compare our results to known giant molecular coulds in the Milky Way Galaxy, and begin to better understand the relationship, if any, between star formation and star forming environments.