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Symposium Proceedings

 

Note: Presentations are grouped by the student’s area of research (based on the faculty mentor’s academic department), not the student’s academic major.

 

Poster Session A: 

8:30 - 9:30 am

Animal and Food Science; Entomology and Plant Pathology; Horticulture and Landscape Architecture; Integrative Biology; and Microbiology and Molecular Genetics (35 posters)

 

Presentations:

 

A-01       Hunter McConnell

Research Collaborators:  Abigail R. Rathert-Williams, Andrew P. Foote

Research Presentation Title:  Effect of increased ruminal propionate on the expression of hepatic gluconeogenic genes in cattle on a finishing ration

Faculty Research Mentor:  Andrew Foote, Animal and Food Science

 

The objective of this experiment was to ascertain if supplementing calcium propionate (CaP) in varying amounts would result in the increased expression of genes related to glucose metabolism in the liver. The study utilized cannulated Holstein steers (n = 6) in a 3 ´ 6 Latin rectangle with three 15-d periods. The treatments were as follows: Control (no CaP), low propionate (100 g/d CaP), and high propionate (300 g/d CaP). The treatments were administered in halves twice a day through rumen cannulas. The steers were provided with ad libitum finishing ration, using Insentec feeders to record feed intake, and unrestricted access to water. Liver biopsies were taken on d15 of each period, a day after a glucose tolerance test, and flash frozen. RNA was extracted from the liver tissue, reverse transcribed for cDNA, and analyzed through quantitative real-time PCR. Five target genes involved in gluconeogenesis were analyzed and included solute carrier family 16 member 1 (SLC16A1), phosphoenolpyruvate carboxykinase 1 (PCK1), phosphoenolpyruvate carboxykinase 2 (PCK2), glucose-6-phosphatase (G6PC), and solute carrier family 2 member 2 (SLC2A2). Data were analyzed using a mixed model with treatment, period, and their interaction included as fixed effects and steer as a random effect. There was no treatment effect on hepatic gene expression (P ≥ 0.57). SLC16A1 showed a negative, correlation with d7 plasma lactate concentration (r = -0.84, P < 0.001) and a negative relationship with fasting plasma lactate concentration (r = -0.55, P = 0.028). SLC2A2 tended to show a positive correlation with fasting plasma glucose (r = 0.44, P = 0.09), fasting plasma lactate concentration (r = 0.43, P = 0.09), and glucose area under the curve (r = 0.46, P = 0.07). These data indicate that increased propionate may not have an impact on hepatic gene expression. Keywords: propionate, cattle, gene expression, liver

 

A-02       Morgan Patterson

Research Collaborators:  Lily Hernandez, Cassidy Reddout

Research Presentation Title:  Chronic Stress and Welfare Assessment in Short Dock Versus Long Dock Tailed Lambs

Faculty Research Mentor:  Janeen Salak-Johnson, Animal and Food Science

 

Tailing docking in lambs is a common practice that can improve lamb welfare by reducing fecal soiling and ultimately reducing the incidence of fly strike. Fly strike is a painful condition that results when maggots burrow into the underside of the tail, which irritates the flesh and attracts more flies and releases ammonia which can poison sheep (AVMA, 2014). Sheep can die within 3-6 days after the onset of fly strike (Morris, 2000). Tail-docked sheep often have greater reproductive capacity, improved feed efficiency, and body weight gain. These sheep are also more tolerant to heat stress, improving sheep welfare (Alkass et al., 2014). Despite these positive benefits, there is much controversy centered around the tail length of the docked tail in terms of lamb health and welfare. Currently, data are sparse as to what is too short based on welfare outcomes since the length defined as "short" is arbitrary and often associated with an increased incidence of rectal prolapse. Therefore, this study was designed to assess the effect of tail dock length of 0.5 to 0.7” (short) versus 1.5” (long) on lamb welfare using multiple measures of behavior, physiology, and productivity. Methods include collecting behavioral data via video footage of lambs interacting in social group housing partially separated from the ewes; the area of observation is only accessible by the lambs, but the test subjects are still able to be with their mothers at any time. Data collected for this study will also include blood samples run to test cortisol levels and assess the severity of each treatment's stress.  

 

A-03       Kaylee Rumbaugh

Research Collaborators:  Divya Jaroni, Jordan Drake

Research Presentation Title:  Selection of Lactic Acid Bacteria for a Direct-Fed-Microbial Against Shigatoxigenic Escherichia coli in Food Animals

Faculty Research Mentor:  Divya Jaroni, Animal and Food Science

 

Introduction: Shigatoxigenic Escherichia coli (STEC) have caused numerous foodborne outbreaks and recalls, with meat products and fresh produce as implicated vehicles. Food animals, especially ruminants, are well-known reservoirs. Appropriate measures to reduce this group of pathogens at the farm-level are warranted. Strains of lactic acid bacteria (LAB) may be utilized as direct-fed-microbials (DFM) in food animals, however, they must be carefully selected to maximize their inhibitory activity. Purpose: To evaluate LAB strains for acid and bile tolerance and inhibition towards STEC for potential DFM. Methods: Viability of several (n=205) strains of lactic acid bacteria (Genus: Lactobacillus, Lactococcus, Leuconostoc, or Streptococcus) were evaluated for inhibition against STEC, using agar-spot-test. Cocktails (5x107 CFU/ml) of 4 STEC strains per serotype (O157, O111, O103, O26, O121, O45, O145) were used. LAB that inhibited STEC significantly (inhibition zones >10.0 mm) were further tested for acid and bile tolerance. Each strain (1x108 CFU/ml) was added to MRS broth adjusted to pH (2, 4, 5) and bile (0.1, 0.3, 0.5%), along with controls, and incubated for 0, 1, 3, and 6 h at 37°C. Bacterial growth was determined by measuring absorbance at 620 nm and 660 nm for acid and bile tolerance, respectively. Results: Fifty of the revived strains were further tested for inhibition against STEC serotypes. Among these, 15% showed excellent (>15 mm), 32% very-good (>10 mm), and 29% good (>5 mm) inhibition against STEC. Isolates showing >10 mm inhibition were further screened for acid and bile tolerance. Compared to 0 h (A620=0.222-0.736), all isolates showed stable growth up to 6 h (A620=0.243-0.986) at all pH values. For bile tolerance, 80% isolates at 0.1%, 40% at 0.3% and 30% at 0.5% showed increased growth over 6 h, indicating that they were better able to tolerate 0.1% compared to the 0.3 and 0.5% bile concentrations. Significance: Lactic acid bacteria showed potential as DFM due to good survival capabilities for the GI-tract environment and significant inhibition against STEC.

 

A-04       Camilla Austin

Research Collaborators:  Rafaela Gomes Ruschel

Research Presentation Title:  First report of pokeweed mosaic virus (PkMV) in Oklahoma and Michigan based on molecular detection methods

Faculty Research Mentor:  Francisco Ochoa Corona, Entomology and Plant Pathology

 

The Pokeweed mosaic virus (PkMV) is a single-stranded RNA virus that belongs to the Potyviridae family and Potyvirus genus. This virus is known to infect Phytolacca decandra, or American Pokeweed, that can be found throughout the United States. The virus has been reported in New Jersey, Maryland, Pennsylvania, and other eastern states, each reported state having a different strain of this virus. American Pokeweed found in Michigan and Oklahoma present symptoms of irregular leaf mottling, stunted growth, and curled leaves. To our knowledge, there are no reports of PkMV being present in these two states prior to this finding. According to previous reports, mutated virus is present in every state PkMV is reported in. Thus, we hypothesize that PkMV from Michigan and Oklahoma could represent new strains of the virus. In order to confirm that this Potyvirus species was present in the Phytolacca decandra samples from Michigan and Oklahoma, RNA extraction was performed using RNeasy Plant Mini Kit (Qiagen), and general potyvirus primers were tested. Moreover, High-throughput Sequencing (HTS) with MinION was also applied for diagnostics. For confirmation of virus presence Klenov et al. 2018 primers were used. Additionally, eleven sets of primers were designed based on alignment of all PkMV sequences deposited in NCBI GeneBank. Oligonucleotide primers were designed using Geneoius v. 2021.1.1. The obtained RT-PCR products were Sanger sequenced. General Poyvirus primers and HTS sequencing confirmed the presence of the virus. According to the ICTV criteria new Potyvirus strains are 82% identical in amino acid and <76% identical in nucleotide sequence, in their complete ORF (polyprotein) sequences. We propose that the PkMV found in Oklahoma and Michigan compose new strains of pokeweed mosaic virus spp.

 

A-05       Makenzie Driever

Research Collaborators:  Makenzie Driever, Rafaela Ruschel,  Francisco Ochoa-Corona

Research Presentation Title:  Exploring the Virome of Chrysanthemums in Oklahoma

Faculty Research Mentor:  Francisco Ochoa Corona, Entomology and Plant Pathology

 

A farm in Tulsa, Oklahoma, distributes chrysanthemums nationally. A large number of varieties showed signs of disease creating a risk for virus spreading nationwide and economic losses. A pool of 165 varieties of chrysanthemum were sampled. The 165 samples were subjected to serological testing (ELISA) and 15 viruses were assayed. The results showed that 157 samples were infected with at least one virus. Several varieties showed multiple virus infections, one variety was infected by seven viruses simultaneously. The presence of viroids was also tested in the 165 varieties using hybridization assay. Two viroids were tested, CchMVd and CSVD. The results showed that only four samples tested negative for either viroid. Recombinase polymerase amplification (RPA) assays are currently under development. Two sets of primers, one for each viroid, were designed using Primer3 adapted for use in Replicase Polymerase Amplification (RPA). Reverse transcriptase polymerase chain reaction (RT- PCR) was also performed for confirmation. This project and its results are beneficial to the future of research in agriculture and the surrounding viral ecosystems. The ability to detect the presence of viruses and viroids efficiently in plants can help to prevent the spread of viruses and viroids throughout. Further research will allow for successful and more efficient detection practices in the near future.

 

A-06       Isabella Hinojosa

Research Collaborators:  Rafaela G Ruschel

Research Presentation Title:  Preservation at Room Temperature of rose rosette emaravirus Infected Tissue

Faculty Research Mentor:  Francisco Ochoa Corona, Entomology and Plant Pathology

 

The storage of biological evidence in microbial forensics is important because investigations may take years. Biological material such as RNA and DNA require storage at freezing temperatures. Cold storage for extended periods of time requires equipment, capital expenditures, expensive maintenance, energy, and substantial space. The ability to store biological material such as RNA at room temperature would curb these issues. This study analyzes 12 samples of plant tissue infected with rose rosette emaravirus stored at room temperature for different time-lapses. RNA extraction will be conducted using a RNeasy Kit and a method named plastic trapping method. Extracted RNA will be subject to testing methods that include cDNA synthesis, end-point RT-PCR, and gel electrophoresis. Results will determine if RNA can be stored at room temperature for a duration exceeding a year. This study explores whether viral RNA in lyophilized samples stored at room temperature remains preserved for further biological research usage.

 

A-07       Ethan Shaw

Research Collaborators:  Greg Middleton

Research Presentation Title:  Determining Activity Patterns of Oklahoma Dung Beetles and Their Response to Prescribed Fire

Faculty Research Mentor:  Wyatt Hoback, Entomology and Plant Pathology

 

Producers of livestock benefit from the activities of dung beetles (Coleoptera: Scarabaeidae) in a variety of ways. Burial of dung can prevent pest fly species from reproducing, and adds nutrients and aeration to soils, encouraging future plant growth in pastures. The estimated ecosystem value of dung beetles exceeds $380 million with Oklahoma ranking second in U.S. cattle production. Despite their recognized importance and the characterization of behavioral groups and timing of reproduction, many dung beetles have limited natural history information about daily activity patterns.  Because dung is an ephemeral resource, competition is predicted to lead to niche partitioning in activity patterns.  We observed daily activity patterns of Phanaeus vindex, Copris fricator, Canthon vigilians, and Onthophagus hecate to determine if they are diurnal or nocturnal.  We discovered that P. vindex, C. vigilans, and O. hecate were primarily diurnal while C. fricator appeared to be exclusively nocturnal. We also observed how some of these species interacted with the resource of dung. C. vigilans rolled dung balls along the surface before burying it, and C. fricator also buried their dung but showed no signs of any surface dung rolling. O. hecate appears to have taken pieces off of the dung they were provided, and buried said pieces. These results can potentially benefit producers by allowing treatments to occur when beneficial insects are inactive.

 

A-08       Abigail Hobbs

Research Collaborators:  Niels Maness, Shehbaz Singh

Research Presentation Title:  Carbohydrate content of rhizomes as potential indicator of sod tensile strength in Cynodon spp.

Faculty Research Mentor:  Charles Fontanier, Horticulture and Landscape Architecture

 

Bermudagrass (Cynodon spp.) is an important turfgrass for golf, sports fields, and amenity lawns in Oklahoma. The predominant means for establishing bermudagrass is from sod, and new cultivars of bermudagrass should be evaluated to ensure sufficient sod tensile strength (STS) for commercial production. The measurement of STS in turfgrass research traditionally requires the establishment and subsequent destruction of large field plots. It is hypothesized that the concentration of structural and non-structural carbohydrates in rhizomes corresponds to STS. As part of a previous project, twenty-five bermudagrasses were planted in the field at the Oklahoma State University Turf Research Center in summer 2019, and subsequently evaluated for STS using standard field-based methods. The present study measured the carbohydrate concentration in rhizomes of these twenty-five bermudagrasses varying in STS and developed a model predicting STS from rhizome carbohydrate concentration. In spring 2020, rhizomes were collected from each one for determination of physical properties and structural and non-structural carbohydrate concentration. Starch content was measured via spectrophotometer using the Nelson-Somogyi colorimetric procedure at 520 nm; soluble sugars (glucose, fructose, and sucrose) were measured via gas chromatograph injections using a DB 5 column; and structural carbohydrates were quantified as a function of acid detergent fiber, neutral detergent fiber, and acid detergent lignin. Data were analyzed using multiple linear regression with the backward elimination option to identify variables most predictive of STS. Lignin, glucose, and rhizome dry weight were considered the best predictors of STS. Predicted STS was compared against measured STS, resulting in a moderately good fit to the data (R = 0.67). Variation in overall rhizome density within a sod pad likely influenced model accuracy. Combining these physiological traits with morphological data may provide an alternative to traditional STS field measurements.

 

A-09       Grace Rembold

Research Collaborators:  Geano Dong

Research Presentation Title:  Effective Irrigation Scheduling Using Daily Time-Steps

Faculty Research Mentor:  Charles Fontanier, Horticulture and Landscape Architecture

 

Efficient irrigation scheduling is critical to the function, health, and aesthetics of turfgrasses and conservation of water resources in Oklahoma. Current guidance for irrigation scheduling of turfgrasses have typically been developed using monthly average rainfall and reference evapotranspiration.  Using these recommendations, it is commonly recommended to irrigate warm season turfgrasses at “one inch per week”.  Observations under replicated experiments and real-world case studies suggest this recommendation overestimates actual turfgrass water requirements. This study aims to build a model to improve recommendations for irrigation scheduling in Oklahoma.  The model uses daily meteorological data from the Oklahoma Mesonet and functions from the Oklahoma State Soil Physics Toolbox. The MATLAB model creates a daily soil water balance using meteorological data and prescribed irrigation schedules.  After the compilation of the initial code into one congruent function, the model was calibrated using 2016 data from two Mesonet sites (Copan and Perkins).  The model was then expanded to predict a daily water balance and irrigation water requirements for ten separate locations over ten years. Model outputs included monthly irrigation depth, annual irrigation depth, annual irrigation event count, and number of days at various soil moisture thresholds (no stress, moderate stress, severe stress).  Input parameters (crop coefficient, allowed depletion) were varied, and outputs compared to traditional recommendations (“one inch per week”). In a state like Oklahoma, which has a highly variable climate, a daily time-step model could enhance turfgrass management practices and create opportunities for further automation.

 

A-10       Amanda Ayon

Research Collaborators:  Angela Riley

Research Presentation Title:  The Effects of Paternal Deprivation on Beak Coloration

Faculty Research Mentor:  Jennifer Grindstaff, Integrative Biology

 

For ornamented species, beak coloration is a key factor that is considered by the female. In zebra finches (Taeniopygia guttata), females prefer males that exhibit redder beaks in comparison to those that exhibit more orange beaks. This carotenoid coloration signals bird health, with many factors influencing adult beak color development, including stress and immune challenges during early life. Like many bird species, zebra finches are biparental. However, few studies have tested how paternal removal affects offspring development. We tested how paternal removal affected beak coloration of sons. We set up two treatment groups: one group that had the father removed from the nest at hatching (early removal) and another that had the father removed at fledging (late removal), as well as a control group with no paternal removal. Once adults, we measured the brightness, hue, and saturation of the upper mandible of male offspring using a spectrophotometer. Birds in the late removal group had breaks with significantly lower red reflectance in comparison to the control group. These results suggest that paternal removal during fledging disrupts the proper development of beak coloration.

 

A-11       Madisen Brown

Research Collaborators:  Michael Reichert, Rachel Atherton, Clarissa Gottfried

Research Presentation Title:  Feather Color as an Indicator of Bird Health and Fitness

Faculty Research Mentor:  Michael Reichert, Integrative Biology

 

Bird feathers come in many beautiful colors, and in addition to being pleasing to the eye, they could potentially help scientists understand certain aspects of that bird’s health and fitness. For example, variations in brightness could be correlated with other measurements of bird health. Additionally, something very unique about birds is that they have a fourth type of cone in their eyes that allow them to see in the ultraviolet light range and as a result, their feathers can be brightly colored in not just the visible light spectrum, but also the ultraviolet light spectrum. By measuring the spectroscopy of the bright yellow breast feathers collected from over 1,200 individuals of the Great Tit, a European songbird, and comparing these measurements with other data from the same individuals, we can determine if these bright colors in the visible light range and in the ultraviolet range can be correlated with other indicators of bird health, such as fat score, weight, and age. In addition, since many of these feathers were collected repeatedly from the same individuals at different times of the year, we can also observe how their feather color changes across seasons and with age. With this information, we can determine if feather color could be used as a reliable indicator of overall bird fitness.

 

A-12       Hailey Freeman

Research Collaborators:  Jennifer Grindstaff

Research Presentation Title:  The Effects of Early Life Adversity and Multiple Stressors on Zebra Finch Stress Resilience

Faculty Research Mentor:  Jennifer Grindstaff, Integrative Biology

 

Chronic stress has been shown to have adverse physiological effects and may impact specific biomarkers (hemoglobin, blood glucose, plasma protein, and triglycerides). However, these biomarkers vary among individuals and while some individuals may experience an increase in vulnerability due to chronic stress, others may increase in resilience. There is much known about the effects of single-point stressors, but less has been done to explore the effects of stressors across developmental stages. The objective of this study was to test the effects of multiple stressors on Zebra Finches (Taeniopygia guttata) and quantify biomarkers as an indicator of stress resilience. In addition, we are testing alternative ways to quantify physiological dysregulation through a measure of multivariate deviation from normality called Mahalanobis distances. Zebra finches are biparental, social birds, and reductions in parental care and social isolation are perceived as stressors. We set up Zebra Finch families to either be exposed to different combinations of stressors or in a control group exposed to no stressors. In early life, paternal removal occurred at either the beginning of the nestling period or at the beginning of the fledgling period. For the adolescent stressor, independent offspring were separated for 2 hours per day from their social cage mate for 12 days using a screen. After 140 days post-hatch, we tested adult sensitivity to the environment through social disruption and measured physiological biomarkers. All individuals were exposed to social disruption by moving to new cages with a novel, same-sex cage mate. If early life adversity canalizes physiological reactivity to the environment, then we would expect that individuals from the paternal removal and adolescent separation groups will demonstrate less of a change in biomarkers after social disruption in adulthood than control individuals. This study will help us further understand environmental sensitivity and physiological reactivity.

 

A-13       Brooke Jacob and Aimee Zimmerman

Research Collaborators:  Paige Stevens, Jason Bruck

Research Presentation Title:  Cognition in Dolphins: The effects of anthropogenic noise and the COVID-19 anthropause on attention in Tursiops truncatus

Faculty Research Mentor:  Matt Lovern, Integrative Biology

 

Human-generated noise pollution is an increasing threat to marine mammals, yet the extent of anthropogenic sound pollution’s effects on cognition are poorly understood. Bottlenose dolphins, Tursiops truncatus, use cognitive skills to hunt, maintain social groups, and communicate. Thus, they are at an increased risk of poor welfare in the presence of anthropogenic sounds that may function as cognitive distractors. To test the effects sound has on dolphin cognition, I will examine three different aspects of cognition (attention, learning, and memory) in the presence of anthropogenic sound. To test how these sounds affect attention, I will travel to multiple dolphinarium facilities, both inland (where the animals experience no boat or sonar sounds) and coastal (where the animals experience regular anthropogenic noise from cruise ships, sonar, and personal water crafts). Each subject will be presented with four different playback sessions, and the results will be compared between facility types as well as across individual, sex, and noise type. Preliminary data using look duration of animals from Dolphin Quest Bermuda and Georgia Aquarium suggest that cruise ship fits a habituation response, while Navy Low Frequency sonar fits a sensitization pattern. Jet ski response does not fit a habituation or a sensitization profile. To study learning, I will train ten dolphins two novel behaviors. Each dolphin will learn one of the novel tasks in the presence of added anthropogenic noise and the other task without the added noise. To assess long-term memory effects, I will return to retest the same individuals after one year. Testing the cognitive response to anthropogenic noise will allow for discussion of broader implications and expectations related to habituation and sensitization in cetaceans.

 

A-14       Mason Miller

Research Presentation Title:  Exploring the Winner-Loser Effect on Emotion in Crickets

Faculty Research Mentor:  Michael Reichert, Integrative Biology

 

Individual variance in behavioral traits, commonly referred to as “personality,” has been exhibited in numerous animal species, including invertebrates. The well-studied personality trait of exploration tendency may be affected due to positive/optimistic or negative/pessimistic internal emotional states. Animal contests can induce affective profiles, as winning against one’s opponent increases an individual’s “optimism”, while losing causes “pessimism”; these states could then go on to affect behavior in other contexts. This winner-loser effect is prominent in aggressive contests of house crickets (Acheta domesticus), thus the experiment investigates if a cricket’s exploratory behavior is influenced by the outcome of its contest. Exploration tendency is first measured as an individual’s latency time in exiting from a shelter, tested twice for repeatability. Each cricket then participates in an aggressive contest, with opponents chosen to be either smaller or larger, to predetermine the winner and loser. The individual’s latency is again measured, both directly after the contest, as well as 48 hours afterwards to examine if the winner-loser effects persist over longer periods of time. Data collection is still in progress, with over 100 crickets already tested in the span of two months. If the findings support the hypothesis that crickets exhibit varying emotional conditions, the implications on human mental health and medicine could be extensive; their simplistic neural architecture could facilitate studies of the mechanisms behind mental states and result in advanced therapeutic techniques and drugs.

 

A-15       Jordan Palmer and William Lewis

Research Collaborators:  Rachel Hamrock, Jason Bruck

Research Presentation Title:  Kin Recognition in Bottlenose Dolphins

Faculty Research Mentor:  Matthew Lovern, Integrative Biology

 

Kin recognition has been known to aid fitness in mammals by preventing inbreeding and allowing individuals to provide support based on relatedness. Previous research has shown that Bottlenose Dolphins (Tursiops truncatus), use signature whistles to transmit social identity and maintain group cohesion when they are used as contact calls.  The purpose of this project was to discover if dolphins were capable of recognizing kin they had never met, through the utilization of signature whistles. To test this, we performed playbacks of known kin and unknown kin signature whistles to dolphins at two facilities in Hawaii and measured their behavioral responses. We discuss implications for kin recognition mechanisms mediated by learned innovated acoustic signals. Differing levels of response to familiar and unfamiliar kin would further support the hypothesis that there is a potential aspect of signature whistles that conveys relatedness. The implication of these findings indicates possible nuances within signature whistles that may convey kinship. 

 

A-16       Casey Sergott, Emmy Fowler and Shayla Nguyen

Research Collaborators:  Jesse Hurd

Research Presentation Title:  The Effects of Pair Bonding and Sperm Competition on Epididymal Sperm Characteristics

Faculty Research Mentor:  Elizabeth McCullagh, Integrative Biology

 

Prairie Voles are a social, monogamous species which have gained traction in the fields of neurobiology, neuroendocrinology, and psychology as a potential model species for studying human social behavior and associated biology, due to similarities in brain chemistry and social tendencies as well as the similarities between rodent and human physiology. Reproductive work in prairie voles is somewhat limited, with more emphasis placed on behavior than physiology. The female prairie vole estrus cycle is unique in that the females do not begin estrus cycling until exposed to a non-relative male. It is unknown if/how female exposure and pair bonding affect male sperm characteristics and/or production. Here we utilize male prairie voles as a study model for if/how pair bonding, potential sperm competition, and monogamy affect sperm production and characteristics. Prior to comparing sperm characteristics between these groups, we established an effective methodology for epididymal sperm extraction utilizing established techniques in other rodents, modern theriogenology concepts, and trial and error. This methodology displays statistical significance and consistency based on established reproductive parameters in mammals and is thus appropriate to utilize for study group comparisons moving forward. We now turn our attention to the differences between unmated males, mated monogamous males, and males exposed to a variety of sperm competition. Preliminary data indicates a statistical difference in sperm characteristics between unmated males and mated monogamous males, but more data is needed to reach an objective conclusion. This study will contribute to a better understanding of the relationship between monogamy, sperm competition, and pair bonding and their effects on reproductive physiology.

 

A-17       Elise Ballinger

Research Collaborators:  Jeffrey Hadwiger

Research Presentation Title:  Role of Dictyostelium Transcription Factor GtaC in Chemotaxis

Faculty Research Mentor:  Jeffrey Hadwiger, Microbiology and Molecular Genetics

 

Dictyostelium discoideum, a social amoeba, is a model system used to examine and understand chemotaxis and development. These bacterivores forage for bacteria by sensing and chemotaxing to folate that is released by bacteria. Dictyostelium discoideum can also find neighboring amoebas and form multicellular aggregates via chemotaxis of cAMP. The atypical MAPK (Erk2) is required for the chemotaxis of Dictyostelium discoideum in both folate and cAMP. Additionally, Erk2 function is required for the translocation of the GATA transcription factor (GtaC) in response to folate and cAMP signaling. In order to determine the role of GtaC in chemotaxis, we assayed the movements of gtaC- mutants and wild type cells in response to folate and cAMP. The gtaC- cells chemotaxis to cAMP was similar to wild type cells, suggesting that GtaC is not required for this chemotaxis in early development. Moreover, the gtaC- chemotaxis to folate was greater than that displayed by wild type cells, indicating that GtaC function may inhibit this response. The gtaC- cells increased chemotaxis in response to folate is consistent with GtaC repression of the far1 gene that encodes the receptor required for folate chemotaxis.

 

A-18       Catalina Bradley

Research Collaborators:  Wouter Hoff, Rosalie Dohmen, Natali Edwards, Mary Erdmann, Saylor Hampton, Brenden Heise, Gunnar Hoogerwerf, Clarice Huffman, Scout Powell, Sarah Marie Teeman

Research Presentation Title:  Photoactive Yellow Protein Bioinformatics Analysis

Faculty Research Mentor:  Wouter Hoff, Microbiology and Molecular Genetics

 

Photoactive Yellow Protein (PYP) is a highly researched protein receptor that prompts several responses in different bacteria. Many bacteria have biofilm formation, photo protective pigments, phototaxis, chemotaxis, and other various responses. What remains as a question is what is in the signal transduction chain that brings the signal from the PYP to the actual response and if those proteins are functionally related to the PYP. We have utilized bioinformatics to analyze the multi-domain proteins that showed a PYP domain. The function of the other domains in the proteins can lead us to find similarities in the PYP domain. With this information we can analyze the flanking genes in the operon structures of the multi-domain PYP’s. In analyzing the flanking genes in the operon structures, we have found multiple possible protein functions such as Histidine Kinase, MCP, TAL, GGDEF, and more. Based on our findings, we can further our research to fully understand the regulations and indications of the gene expressions that lead to bacterial responses.

 

A-19       Emily Chandler

Research Collaborators:  Carrie Pratt

Research Presentation Title:  Culturing Novel Anaerobic Gut Fungi from Marsupials

Faculty Research Mentor:  Mostafa Elshahed, Microbiology and Molecular Genetics

 

Anaerobic gut fungi (AGF) from the phylum Neocallimastigomycota are an essential part of the microbiome in herbivores. These fungi aid in digestion in ruminants, pseudoruminants, and nonruminants alike, where they adhere to plant biomass and produce enzymes and bioactive molecules that break down complex sugars. This includes the degradation of cellulose, which is essential in the development of biofuels and other biotechnologies converting plant biomass to products of commercial value. Marsupials, non-placental mammals who split from placental mammals ~125 Mya, have shown strong microscopic and sequence-based evidence for harboring AGF. Marsupial herbivores are promising hosts for uncovering novel AGF because some species are foregut fermenters (kangaroos and wallabies) and others are hindgut fermenters (koalas and wombats), both digestive methods that are associated with AGF presence in other herbivores. Due to their laborious maintenance procedures, strict anaerobic nature, and technical difficulties in genomic sequencing, these microorganisms are under-researched and poorly sampled. We hypothesized that marsupials represent a yet-untapped reservoir for discovering novel AGF taxa. To test this hypothesis, we collected fresh fecal samples from a wide range of marsupials and are currently attempting to enrich and isolate novel AGF taxa from these samples. Using complex media amended with plant substrates, strict anaerobic techniques, and multiple antibiotics for suppressing growth, we were able to enrich for AGF, as evident by visual production of hyphal filaments in enrichment tubes. We are currently undertaking purification procedures to isolate axenic cultures of AGF for subsequent identification and characterization. Culturing novel AGF from marsupials will allow for deeper perspectives on the diversity and biology of anaerobic gut fungi and will enable their applications in veterinary medicine, biofuels, and biomedical engineering.

 

A-20       Kaitlyn Cotton

Research Collaborators:  Benjamin N. Nelson, Nikolay Gerasimchuk, Karen L. Wozniak

Research Presentation Title:  Effects of Organoantimony Compounds on Fungal Pathogens Cryptococcus neoformans and Candida albicans

Faculty Research Mentor:  Karen Wozniak, Microbiology and Molecular Genetics

 

Cryptococcus neoformans is an opportunistic pathogen that causes pulmonary cryptococcosis, or an acute or chronic infection in the lungs, and cryptococcal meningitis, an infection of the brain and spinal column, in immune-compromised individuals. Similarly, Candida albicans can be an opportunistic lung pathogen (often in cystic fibrosis patients), causing an infection called pulmonary candidiasis in addition to genitourinary tract infections, candidemia (a bloodstream infection), and oral candidiasis. Overall, fungal infections are responsible for approximately 1.7 million deaths each year. In contrast to antibacterial drugs, the quantity of antifungal drugs capable of combating fungal infections remains low. With the high toxicity and increased resistance to antifungals in recent years, the importance of finding new options for antifungal therapy is even more crucial. We hypothesized that a series of organoantimony compounds would be effective at restricting fungal growth. For this, we tested approximately 20 compounds against C. neoformans and C. albicans in minimum inhibitory concentration (MIC) assays. Of those tested, compounds A, B, E, I, F, and G were most effective against C. neoformans with MIC concentrations of 10.94 μg/ml, 19.79 μg/ml, 18.75 μg/ml, 12.5 μg/ml, 20.83 μg/ml, and 2.60 μg/ml respectively, and compounds E and G were most effective against C. albicans with MIC concentrations of 15.625 μg/ml and 25 μg/ml respectively. In addition, compounds I and G were also fungicidal against C. neoformans at concentrations 50 μg/ml and 25 μg/ml respectively and compound G was fungicidal against C. albicans at 50 μg/ml. Cytotoxicity assays with mammalian cells were performed on Compounds A, B, E, I, F, and G, and all six compounds were found to have low to negative cytotoxicity. RNA sequencing studies have identified several C. neoformans genes involved with the compounds’ inhibitory effects. Future studies will continue to examine the mechanism(s) of antifungal activity using mutant libraries of these fungal organisms and through scanning electron microscopy (SEM) and transmission electron microscopy (TEM) studies.

 

A-21       John Dorlon IV

Research Collaborators:  Ross Waslker, Robert L. Burnap

Research Presentation Title:  Design and implementation of a CRISPR knockout system in Synechococcus elongatus sp PCC 7942

Faculty Research Mentor:  Robert Burnap, Microbiology and Molecular Genetics

 

Synechococcus elongatus PCC 7942 is a model cyanobacteria used widely for research on the mechanics of photosynthesis. Our lab studies the cyanobacterial NADPH dehydrogenase-like (NDH) complex. The NDH complex, in addition to being an important part of all respiratory chains, is part of the cyanobacterial carbon concentrating mechanism (CCM), which serves to concentrate inorganic carbon near RuBisCO [1]. The CCM greatly increases the concentration of inorganic carbon (Cᵢ) around RuBisCO. Although the cyanobacterial NDH complex has several known functions, including proton transport across the thylakoid membrane and carbonic anhydrase-like activity, the mechanics of the NDH complex constituent proteins have not yet been determined. The project goal is to develop a CRISPR system which can create markerless deletions and potential point mutations in the NDH complex’s constituent proteins.  In previous work, vector pCpf1b was used in the creation of mutant gene libraries through use of the CRISPR system Cas12a [2,3]. This project aims to modify the vector pCpf1b with enhanced selection markers in order to more easily create mutations in the NDH complex of cyanobacteria, thereby allowing for further research into the mechanisms by which this vital complex functions.

 

A-22       Mary Erdmann

Research Collaborators:  Mary J. Erdmann, Somalisa Pan, Matthew Cabeen

Research Presentation Title:  A Novel Lipase Plays a Role in PstN-Mediated Biofilm Suppression in Pseudomonas aeruginosa

Faculty Research Mentor:  Matthew Cabeen, Microbiology and Molecular Genetics

 

Pseudomonas aeruginosa strain PA14 is an opportunistic pathogen that causes nosocomial infections and whose antibiotic resistance poses an ongoing medical threat. One mechanism of drug therapy resistance is the production of biofilm - a self-secreted substance made of polysaccharides, DNA, and proteins that binds cells together, shielding bacteria from their environment. Unraveling the mechanisms behind P. aeruginosa biofilm regulation would provide better an understanding for how this species manages virulence. In PA14, PtsN, part of a phosphorelay called the Nitro-PTS, was identified as a negative regulator of biofilm formation when unphosphorylated. The Nitro-PTS is known to regulate various cellular functions; identifying potential biofilm-suppressive targets of unphosphorylated PstN would contribute to understanding how unphosphorylated PstN decreases biofilm. Using transposon mutagenesis, an unnamed gene (04030) was observed to have a biofilm-suppressive function in an unphosphorylated PtsN background. Predicted protein structures suggested that 04030 has a folding pattern like lipase enzymes. Furthermore, our studies suggest that 04030 has a catalytic serine that is important for 04030 function and appears in a signature catalytic motif typical of lipase enzymes. When unphosphorylated, PstN is potentially interacting with a downstream gene encoding a putative lipase, 04030, to achieve biofilm suppression in PA14.

 

A-23       Kale Goodwin

Research Collaborators:  Mostafa Elshahed, Carrie Pratt

Research Presentation Title:  Culturing anaerobic gut fungi from an avian herbivore

Faculty Research Mentor:  Noha Youssef, Microbiology and Molecular Genetics

 

Anaerobic gut fungi (AGF, phylum Neocallimastigomycota) are an understudied group of microorganisms that reside in the digestive tracts of herbivorous mammals and aid in plant biomass degradation. This degradation is facilitated by adhesion to the biomass and production of enzymes that break down complex sugars including cellulose. The presence of AGF in non-mammalian herbivores is not yet well understood, however, digestive tracts that promote fermentation are most likely to host AGF. One of few truly herbivorous birds, the ostrich, is a hindgut fermenter, with a very long colon and high retention time. These traits are strong indications that ostriches likely harbor AGF within their digestive tracts. We hypothesized that ostriches represent not only a new host for AGF but likely also house novel AGF taxa. To test this hypothesis, we obtained fecal samples from four ostriches and are attempting to enrich and isolate known and novel AGF taxa. Using strict anaerobic techniques, a complex media amended with plant substrates, and antibiotics for bacterial growth suppression, we were able to enrich for AGF, which was visualized as hyphal filaments. We are currently undertaking procedures to isolate pure cultures for identification and characterization. Whether isolates are known or novel, isolation of any AGF from an avian host represents new knowledge gained about these understudied fungi. Culturing AGF from ostriches will allow for further understanding of how AGF evolved in non-mammalian hosts and expand their known evolutionary lineage.

 

A-24       Ashley Harmon

Research Collaborators:  Bekkah Friske, Rolf Prade

Research Presentation Title:  A measure of extracellular vesicle actively during growthon Aspergillus nidulans mutant A773

Faculty Research Mentor:  Bekkah Friske, Microbiology and Molecular Genetics

 

There are still many mysteries about fungal growth and behavior. Research shows that extracellular vesicles (EV) have a system of production that assisting with digestion of media substrate that may let the fungi metabolize and allow for faster growth. Learning more about this form of extra cellular activity in fungi could lead to more understanding over how fungi change their environment. This research could help lead to ability to target and adjust growth in fungi at early stages of development, opening the doors to methods that could increase industrial production give new tools for treating fungal pathogens. In this research we follow the growth curve of the Aspergillus nidulans mutant A773 over the course of its 37OC incubation of multiple 1% Carbon inoculated liquid media. Media on an inoculated six well plate had their absorbance measures taken on a 96 well reader to have as base growth data. The samples we centrifuged down to pellet cell debris and the supernatant was then filtered through an Ultra Filtrate 300 Kilo Dalton Filter. A Bradford protein assay was done on a sample of each step of filtration in order measure the protein concentrations of the raw media, the flow through media (unfiltered supernatant) and the isolated EV. The rest of the isolated vesicles were stained by Cell Brite 488 Membrane Stain and measured in accordance with the time-lapse using Flow Cytometry. The Bradford showed that the EV sample for CMC had the highest concentration of proteins, glucose had the highest concentrations in both other samples, and glycerol had the lowest across the board. As for the time-lapse the flow reading showed that glucose and glycerol held a notable spike at the 48-hour mark, where CMC had a spike at 24 hours with an overall smaller count. Where glucose continued to trend down however, CMC and glycerol were shown to bounce back a bit after the hour mark. The trend that spike in EV counts spiked during early in the growth cycle can indicate that this is way that newly growing fungi may influence the environment to optimize their growth.

 

A-25       Anna Jackson

Research Collaborators:  Amy G. Hurst

Research Presentation Title:  Using DNA barcoding to molecularly identify Apis mellifera sspp.

Faculty Research Mentor:  Amy G. Hurst, Microbiology and Molecular Genetics

 

Apis mellifera, the Western Honeybee, is a primary pollinator that improves biodiversity and produces food for both society and animals.  Morphological identification methods of Apis species and subspecies have long been used.  However, newer molecular identification methods, such as DNA barcoding, provide a genetic approach to identification that may better preserve breed purity and bee health.  DNA barcoding protocol was carried out by collecting an A. mellifera sample, extracting and amplifying a DNA fragment, verifying the amplified DNA by gel electrophoresis, sequencing the DNA fragment at GENEWIZ, and comparing the sequence to known identities in NCBI’s GenBank database.  Gel electrophoresis verified the bee’s DNA fragment by its size.  The sequence returned from GENEWIZ and resulted in 99% matches to three A. mellifera subsecies in GenBank, including A. m. ligustica, A. m. causica, and A. m. carnica.  The sample’s morphological characteristics and geographical location contributed to the conclusion of its identity as A. m. ligustica.

 

A-26       Elizabeth Kent

Research Presentation Title:  How Do Novel Nanospring Patterns Impact Bacterial Growth?

Faculty Research Mentor:  Matthew Cabeen, Microbiology and Molecular Genetics

 

Previous research studying the antibacterial effects of nanopatterned surfaces has found evidence showing that these patterns can reduce or increase bacterial colonization and pathogen growth. The topographical surface patterns may be applied to future medical technology and have the potential to reduce infections and bacterial growth without the use of antibacterial products. Although commercial antibiotics are commonly used to kill pathogens, the overuse of these products has resulted in bacterial resistance, which can pose a serious health threat (F. Arisoy et al). Antifouling and bactericidal capabilities occur naturally on the surfaces of various plants, insect wings, and the skin of reptiles and sharks (M. Mischalska et al) and may provide an effective solution for killing bacteria or preventing their growth without the need for antibacterial products (M. Mischalska et al). Synthetic nanopatterned surfaces may similarly retard or encourage bacterial attachment and growth. We have collaborated with Dave McIlroy’s lab in the Oklahoma State University Department of Physics to test the impact of novel silicon nanosprings (L. Wang et al) on bacterial colonization and growth. These nanosprings are on the same scale as biologically patterned structures but are distinct and have never been tested for their effect on microbial growth. Thus, testing nanospring-covered surfaces for bacterial growth was an ideal opportunity to test the ability and efficiency of novel nanospring patterns to retard or encourage bacterial colonization and growth. To test bacterial colonization on nanosprings, we inoculate sterile control and nanospring-covered substrates with Pseudomonas aeruginosa, an opportunistic human pathogen. We then incubate the substrate to allow bacterial growth. We assess attachment and growth directly, via microscopy, and indirectly by dislodging the attached bacteria, serially diluting, and growing them on agar plates. Data collection and measurements for this experiment are currently ongoing, preventing any strong conclusions at present. Alternative means of reducing bacterial growth and biofilm formation is increasingly important as many strains of bacteria resist traditional antibiotics. Assessing untested nano-patterned surfaces is one promising avenue for discovering new ways to prevent unwanted bacterial growth.

 

A-27       Creed Killgore and Trenton Skinner

Research Collaborators:  Matthew Cabeen

Research Presentation Title:  Genetic regulation of bacterial killing complexes in Pseudomonas aeruginosa

Faculty Research Mentor:  Matthew Cabeen, Microbiology and Molecular Genetics

 

Pseudomonas aeruginosa is an opportunistic pathogenic bacterium that has the unique ability to produce intraspecific killing complexes called pyocins, typically in response to DNA damage. The pyocins kill any nearby susceptible bacteria. However, to permit the relatively large pyocin complexes to escape the P. aeruginosa cells that produce these molecules, producer cells die by programmed cell lysis. Our laboratory recently found that deletion of the xerC gene, encoding a tyrosine recombinase, initiates an alternative pathway for strong pyocin production (and the accompanying cell lysis) that is independent of DNA damage. Deletion of xerC also increases the effectiveness of fluoroquinolone antibiotics. Most of the molecular mechanisms behind the newly discovered alternative pathway are not yet understood. We aim to uncover the details of this pathway and leverage it for therapeutic advantage. To do so, we have mutated, manipulated, and deleted specific genes that are believed to be critical in the pyocin production and release pathway. We find that not only holin and lysin genes encoded in the pyocin gene cluster, but also another holin called AlpB, are involved in cell lysis and death. Even with lysis blocked, pyocin-producing cells stop growing. We also find that the pyocin gene activator PrtN is necessary but not sufficient for pyocin production, suggesting the involvement of other factors. By making mutations in a repressor of prtN expression, PrtR, we gained evidence that PrtR has additional targets in pyocin expression and release. Our ultimate goal is to discover information that will lead to the discovery of drugs that can mimic xerC deletion, eliciting increased antibiotic susceptibility and pyocin production in clinical P. aeruginosa infections.

 

A-28       Jacob Lieberman

Research Collaborators:  Tim Hubin

Research Presentation Title:  Antifungal Activity of Novel Macrocycle Derivatives

Faculty Research Mentor:  Karen Wozniak, Microbiology and Molecular Genetics

 

Introduction. Cryptococcus neoformans is a fungal pathogen that causes meningitis in approximately 225,000 individuals with AIDS each year, resulting in over 181,000 annual deaths. C. neoformans is inhaled and then escapes the lungs and spreads to the central nervous system where life-threatening meningitis occurs. There are currently only four classes of antifungal drugs available to treat this infection. These medications are not very effective, they are highly toxic to humans, and the organism is developing resistance against these drugs. Therefore, new antifungal drugs are desperately needed. Our collaborator has developed macrocycle derivatives that have activity against other fungal strains, parasites, and bacterial pathogens, and we were interested in testing their activity against C. neoformans strain H99. Based on their activity against other pathogens, we hypothesized that these compounds would have antifungal activity against C. neoformans. Methods. For these studies, 12 novel macrocycle compounds provided by our collaborator were tested against the C. neoformans strain H99. Antifungal activity was determined using minimum inhibitory concentration (MIC) assays in two media - RPMI-MOPS and YNB-glucose. Results. Thus far, our results show that 3 of the tested compounds have MIC values below 20ug/ml in RPMI-MOPS and 4 of the tested compounds have MIC values below 10ug/ml in YNB-glucose. Conclusion. We conclude that several of the compounds show antifungal activity against the C. neoformans strain, H99. Future studies will test the cytotoxicity of the antifungal compounds against mammalian macrophage cells, RNA collection and analysis, and the efficacy of the compounds against a C. neoformans mutant library. Relevance of the study. These results and future studies will determine whether these compounds could be developed as a therapy for deadly cryptococcosis.

 

A-29       Rebecca Lindley

Research Collaborators:  Ramee Aranda and Jeffrey Hadwiger

Research Presentation Title:  Examination of Protein Kinase Specificity Relating to the C-terminal Motif in Atypical MAPKs

Faculty Research Mentor:  Jeffrey Hadwiger, Microbiology and Molecular Genetics

 

The soil-dwelling amoeba Dictyostelium discoideum is often used as a model organism for cellular signaling due to its similarity to mammalian cells, specifically leukocytes. Mitogen-activated protein kinases (MAPKs) are important in eukaryotic signaling pathways, cell growth/division, cell differentiation, and cell movement and therefore might play a role in cancer or other diseases. Little is known about atypical MAPKs that are found in mammals and amoebae. Dictyostelium discoideum has two MAPKs, one typical (Erk1) and one atypical (Erk2). Erk2 is required for chemotactic movement in response to cAMP or folate. Erk1 is not required for chemotaxis but is activated as a secondary response to chemoattractants. It is also known that loss of Erk2 has differing phenotypes than the loss of Erk1. To define regions of these MAPKs that are unique for typical or atypical MAPKs, chimeric MAPKs were created at an area of interest called the Carboxyl Terminal Motif (CTM). The CTM region is unique to atypical MAPKs which may mean it plays an important role in their function. In one chimera the atypical MAPK Erk2 CTM region was replaced with the typical MAPK Erk1 region. In the other chimera the typical Erk1 region was replaced with an atypical Erk2 CTM region. The chimeras were expressed in cells lacking either Erk1 or Erk2 and tested for MAPK function. This included developmental and aggregation assays, shuttling of the GtaC transcription factor, and phosphorylation. It was found that replacing the CTM of Erk2 with Erk1 greatly reduces the function of Erk2 and adding the CTM of Erk2 to Erk1 does not grant Erk2 function to Erk1. Our data also suggests that the amino terminal half of MAPKs seem to be important for their specificity.

 

A-30       Brannon Maravich

Research Collaborators:  Matthew Cabeen, Rabindra Khadka

Research Presentation Title:  The impact of specific amino acid mutations in the Bacillus subtilis stressosome on environmental stress response patterns and fitness

Faculty Research Mentor:  Matthew Cabeen, Microbiology and Molecular Genetics

 

The natural ability of bacteria to survive a range of conditions depends heavily on their ability to sense and respond to stressors in the environment. However, the ability to sense external stimuli and convert them to internal cues is incompletely understood. It has been demonstrated that Bacillus subtilis, a model organism for the stress response, mounts a response to various external stressors through a cytoplasmic multi-protein complex called the stressosome, which consists of RsbR, RsbS, and RsbT proteins. In the presence of environmental stress, this complex releases RsbT, which activates a cascade of events leading to the release of the transcription factor σB, which promotes the expression of stress-response genes. The RsbR protein, and specifically its N-terminal region, is the proposed sensor for the stressosome. In this study, we created a stain containing only a single RsbR paralog, RsbRA (wild-type cells encode 5 different RsbR paralogs). We then used alanine scanning to replace specific amino acid residues within this N-terminal region of RsbRA and measure their impact on σB-directed expression in the presence of ethanol by measuring β-galactosidase activity. In comparison to the wild type, the mutant strains displayed altered response patterns that we categorized as reduced, delayed, or delayed but increasing, suggesting that these amino acids impact cells’ ability to respond to stress. To further test the effects of these mutations, we will perform competition experiments between different mutant strains to gauge the impact of each allele on fitness. We will tag strains with specific fluorescent tags, co-culture them, and then count colonies or use fluorescence microscopy to quantify their relative proportions (fitness). This analysis will contribute to our understanding of the importance of individual amino acid residues of the RsbRA protein, helping us connect stress response patterns to fitness in B. subtilis.

 

A-31       Vanessa Moore

Research Collaborators:  Sudhir Doranga

Research Presentation Title:  WT and ΔompA E.coli compete in the same niche for colonization in the mouse intestine

Faculty Research Mentor:  Tyrrell Conway, Microbiology and Molecular Genetics

 

Escherichia Coli is known to contain several outer membrane porin proteins that potentially play a role in its ability to colonize its host. One of the most abundant is the Beta barrel outer membrane protein A, or ompA. To study its role in colonization, an ompA-deletion mutant of commensal E.coli MG1655 was constructed. The ΔompA strain was administered to mice in a series of experiments to determine how the absence of ompA would affect the colonization of E.coli in the mouse intestine. The ΔompA mutant alone colonized in near equivalence to the wild type, supporting that ompA is not vital to colonization. However, when WT and ΔompA E.coli strains were administered simultaneously, WT outcompeted the ΔompA by almost 4 log fold within 15 days of competition. To determine if ompA mutant competes in the same niche as the WT, mice were associated with WT then challenged with the ompA mutant. A second experiment was also done in reverse order. When WT was administered first followed by ompA, ompA was unable to colonize. Likewise, when ompA was administered first followed by WT, WT was unable to colonize. This determined that both strains are competing in the same niche and once the niche is fully occupied by one strain, another cannot colonize.

 

A-32       Kayli Nail

Research Collaborators:  Erika Lutter, Christian Holcomb

Research Presentation Title:  Bacterial two hybrid analysis of Chlamydia trachomatis inclusion membrane protein, CT226

Faculty Research Mentor:  Erika Lutter, Microbiology and Molecular Genetics

 

Chlamydia trachomatis is an obligate intracellular bacterial pathogen which poses severe health problems throughout the world. There are over 90 million new cases annually, making it the most common sexually transmitted disease in the world. Chlamydia can pose significant problems during and after infection. It is imperative to understand how Chlamydia manipulates the host cell and potentially develop future treatments. The mechanisms by which C. trachomatis alters immune response is not well understood, but recent work has identified an interaction between the chlamydial inclusion membrane protein, CT226, and the potential interacting host proteins, Flightless homologue II (FLII), and Leucine Rich-Repeat Flightless-Interacting Proteins 1&2 (LRRFIP1 and LRRFIP2). FLII, LRRFIP1 and LRRFIP2 are known to interact as a complex and are upstream regulators of the inflammasome. Currently, it is unknown if CT226 interacts with one or all of the interacting partners and needs the actual interaction needs investigation. My hypothesis is that CT226 will directly interact with one of the 3 potential interacting partners (LRRFIP1, LRRFIP2, and FLII) and that we will be able to detect this interaction in the bacterial two hybrid system. Current efforts have focused on cloning CT226 into PUT18 and the three host proteins (FLII, LRRFIP1 and LRRFIP2) individually into PKNT25. PUT18 and PKNT25 are the bait and prey plasmids that have been adapted for use in the bacterial two hybrid system. Once each pair of plasmids are properly transformed (each E. coli strain will have two plasmids, one carrying CT226 and one carrying the potential interacting partner), they will be screened using traditional Beta-galactosidase assays or cAMP assays to determine the specific interactions.

 

A-33       Brenda Rodriguez

Research Collaborators:  Ross Walker, Robert L. Burnap

Research Presentation Title:  Measuring Accumulation of a Unique Carbonic Anhydrase-like CO2 Uptake Protein in Synechococcus elongatus sp. PCC 7942

Faculty Research Mentor:  Robert Burnap, Microbiology and Molecular Genetics

 

Synechococcus elongatus sp. PCC 7942 is a cyanobacteria, a group of single-celled bacteria capable of performing photosynthesis. This organism is used widely for research on the molecular mechanisms of photosynthesis, as it is genetically tractable. An important component of photosynthesis is RuBisCO, which requires highly concentrated CO2 to function efficiently. PCC 7942 possesses the ability to uptake inorganic carbon in the form of CO2 from its environment and concentrate it within the cell for use in metabolic processes in a process called the carbon concentrating mechanism (CCM). CO2 uptake in cyanobacteria is mediated by specialized type 1 NAD(P)H Dehydrogenase (NDH-1) complexes in the thylakoid membrane [1]. These unique NDH-1 complexes house the CO2 uptake proteins A and B (CupA and CupB), which are known to be involved in the CCM due to their carbonic anhydrase-like activity of converting CO2 into HCO3- [2]. However, the mechanism by which the Cup proteins are able to accomplish this reaction within the cyanobacterium remains unknown. To gain a better understanding of the workings of CupA, this project aims to design and create a tool for measuring accumulation of CupA in strains of PCC 7942 with mutations in CupA. Using a variety of genetic and proteomic tools, CupA will be overexpressed, isolated, and injected into a host animal to raise antibody against CupA.

 

A-34       Andrew Thomas

Research Collaborators:  Adam Soriano, Peyton Thompson

Research Presentation Title:  Economic Biofuel Production from Recombinant Trichoderma reesei Transformation

Faculty Research Mentor:  Rolf Prade, Microbiology and Molecular Genetics

 

Energy plays a crucial role in the development of society and its future. The energy obtained from biomass is the fourth largest source of energy in the world, following only oil, natural gas, and coal (Wu et al. 2012). However, biofuel is, comparatively, a much newer industry, whose true potential is still waiting to be tapped into. Lignocellulosic biomass (LCB) used in the production of these biofuels contain cellulose (C6-sugars), hemicellulose (C5-sugars), and lignin (Ballmann et al. 2019). Trichoderma reesei – the leading industrial fungus used for biomass degradation –  is remarkably effective at producing enzymes to break down C5-sugars and C6-sugars, excluding lignin; therefore, lignin is removed through various biomass pretreatments allowing T. reesei to efficiently hydrolyze cellulose (Reese 1976, Gupta et al. 2016). LCB pretreatment methods – including both physical and chemical processes – allow for the catabolism of cellulose but also form a C5-sugar-rich liquor, pentosan-containing pretreated biomass liquors (PPTBs), through the partial breakdown of hemicellulose; the pentose contained in PPTBs can serve as an inducer, allowing us to cheaply manufacture enzymes (Ballmann et al. 2019). However, in the presence of these PPTBs, T. reesei is incapable of producing the vast amount of cellulases it normally does (Kiesenhofer et al. 2018). The barrier, established by the PPTBs blocking cellulase production, prevents the onsite production of cellulases and, consequently, biofuels, drastically increasing production expenses. The aim of this project is to modify the genetic makeup of T. reesei, enabling the use of the otherwise obstructing and often discarded PPTBs’ pentose to produce cellulases, converting cellulose to glucose and allowing for the complete onsite production of biofuels. 

 

A-35       Pau Von

Research Collaborators:  Clark Jett

Research Presentation Title:  Growth Characteristic of Various Fast Growing Synechococcus Elongatus Strains

Faculty Research Mentor:  Robert Burnap, Microbiology and Molecular Genetics

 

Cyanobacteria are unique photosynthetic microorganisms that have the ability to fix carbon at a rapid rate using CO2 concentrating mechanism (CCM) that functions to ‘turbocharge’ the process. The cyanobacterial CCM utilizes a modified NADH dehydrogenase oxidoreductase aka respiratory complexes I, that drives the hydration of CO2 à bicarbonate reaction to accumulate relatively large amounts of inorganic carbon (Ci) in their cytoplasm. This allows optimal enzyme kinetics for the CO2 fixing enzyme rubisco. Understanding this unique and highly effective carbon concentrating mechanism could provide solutions to mitigate the current rising levels of CO2 in the atmosphere. Until recently it has been very hard to study cyanobacteria because of their exceptionally slow growth rate. However, recently several strains have been isolated and/or genetically modified and are shown to have much faster growth rates.  Recently isolated in Port Aransas, Texas, Synechococcus elongatus 2973 (S2973) has been shown to have a much faster growth rate than widely studied Synechococcus elongatus 7942 and Synechocystis 6803. Shortly after, S2973 was genetically modified as to allow for natural competence (2973T). This revelation allows for structure/function studies of the photosynthetic mechanism to be performed faster than usual. Unfortunately, the transformable, fast-growing strain S2973T has recently been discovered to contain a contaminant. That being said, it is possible that the 2973T strain may be in a commensalistic or symbiotic relationship with its contaminant, partially contributing to its fast-growing phenotype. The purpose of this study is to decontaminate the valuable 2973T strain and probe its ability to grow on its own, compared to its contaminated and non-genetically modified counterparts. In addition. This will allow for the quantification of the growth phenotypes of all the current lab strains.

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