5th Annual Texas A&M University System Pathways to the Doctorate Student Research Symposium
November 2-3, 2007
Hosted by Tarleton State University

Abstracts of Student Presentations: Life Sciences

JEANETTE ANDERSON, VERONICA DUQUE GIBSON, and DAVID NGUYEN. "Development of a qPCR Assay to Detect Zygomycetes"
Faculty Mentors: Dr. Sally Lewis, Dr. Sally Hoger, Dr. Sara Taylor

Zygomycetes infections are rare and potentially life-threatening.  Two of the most common members of this family include Rhizopus and Mucor.  Traditional microscopic identification techniques are cumbersome, tedious, and imprecise.  A sensitive and specific molecular assay will provide prompt diagnosis and treatment.  Thus, we are developing a  quantitative real-time (RT) PCR assay with specificity for Rhizopus and Mucor species.  Utilizing the National Center for Biotechnology Information (NCBI) database, primers were designed for a unique partial gene of Rhizopus stolonifer.  Primers were validated by amplification of the target with RT-PCR technology employing a fluorescein (FAM) labeled hydrolysis probe.  Although RT amplification successfully detected a known strain of Rhizopus stolonifer (ATCC isolate 6227b), further validation using clinical specimens is required.


AMANDA BARMORE. "Habitat Selection in Blacktail Shiners."
Faculty Mentor: Dr. Christopher Higgins

Streams of the southwestern United States are subject to a wide array of environmental factors ranging from severe droughts to flash flooding.  In addition, many of these streams are faced with environmental perturbations due to anthropogenic modifications, such as the construction of reservoirs and current practices in the agriculture industry.  Stream fish are capable of responding to environmental perturbations by voluntarily selecting a different habitat within the stream where the conditions may be more suited for their survival and reproduction. The objective of this study was to examine habitat selection in blacktail shiner (Cyprinella venusta), which is an abundant species in many Texas streams.  We used backpack electrofishing to collect fish at one site in each of the Pedernales, San Saba, and South Llano Rivers.  In addition, we quantified habitat availability and habitat use at each site, which involved assessing water temperature, current velocity, water depth, substrate, and geomorphology.  All three sites were sampled once each season.  Blacktail shiners selected cobble substrates regardless of season and selected waters with velocities between 1 and 3 meters per second. During the spring and summer seasons, blacktail shiners preferred riffles where depths were less than 20 cm. However, blacktail shiners selected deeper waters (21-40 cm) during the fall and winter seasons. Blacktail shiners consistently selected cooler waters regardless of season.  The results of this study indicate that blacktail shiners do in fact select certain habitats and that they could be used to assess changes in environmental conditions.


EMILY BRUCKS. "Mathematical Models of the Barred Tiger Salamander."
Faculty Mentors: Dr. Keith Emmert, Dr. Peter White

Amphibians are a great indicator for the overall health of the environment.  Both continuous and discrete models are analyzed for disease free barred tiger salamander, Ambystoma Tigrinum Mavoritium.  Conditions are derived for the stability of equilibria of both the discrete and continuous models.  Simulations are used to compare and contrast as well as explore different aspects of these two types of models.


ARMANDO DE LA CERDA and AMANDA FRANKLIN. "Development of a qPCR Assay to Detect Campylobacter jejuni."   *First place award - Life Sciences undergraduate student*
Faculty Mentors: Dr. Sally Lewis, Dr. Sally Hoger, Dr. Sara Taylor

Campylobacter jejuni and C. coli are the most commonly reported bacterial cause of foodborne infection in the United States. Traditionally, Campylobacter species detection requires a special incubation environment as well as specific growth media for an extended growth period.  The development of a rapid and reliable diagnostic tool for the detection of Campylobacter species would be a valuable aid to the medical diagnostic decision process, especially to rule out Campylobacter infection during the  enteric pre-surgical time period.  Improved patient outcomes might result if this rapid assay could reduce the number of enteric surgeries. In this study, our purpose is to develop a real-time PCR assay with specificity for Camplylobacter jejuni and C. coli.  We designed and validated  specific primers targeting a unique sequence of 80 base pairs within the cadF gene specific for Camplylobacter jejuni and C. coli..  Our assay utilized  fluorescein (FAM) labeled hydrolysis probes for the detection of the amplification of the cadF gene.  Initial results were able to amplify and detect ATCC isolate 33291 of Campylobacter jejuni with appropriate negative controls. This experimental assay is still in its developmental process and further research will prospectively validate our assay’s ability to detect Campylobacter jejuni and C. coli isolates in clinical specimens.


SARA HARSLEY. "A Flora of the Vascular Plants of Erath County, Texas"   *Second place award - Life Sciences Masters Degree student*
Faculty Mentor: Dr. Allan Nelson

Knowledge from native floras and introduced plants in a region is essential to understanding and conserving plant biodiversity.  This type of data is unavailable to conservation managers in the Western Cross Timbers of Texas. A checklist of the vascular plant species from 34 sites and herbarium collections over the past 100 years are being tabulated for Erath County, Texas. Erath County, approximately 1,086 square miles, is located in the Western Cross-Timbers region of Texas. The plant species richness of two large sites, Hunewell Ranch and Tarleton State University’s Agricultural Center will be compared, using species area-relationships, to protected natural areas occurring in north central Texas. The number of endemic, native, and invasive species of the county flora will also be compared to a previous flora of plants occurring in Erath County, compiled by Lula C. Gough in 1921.


SAM KIESCHNICK. "Genetic Diversity of Baird's Pocket Gopher, Geomys breviceps"   *First place award - Life Sciences Masters Degree student* and *First place award - ALL Masters Degree students*
Faculty Mentors: Dr. Phillip Sudman, Dr. Russell Pfau

Baird´s pocket gopher (Geomys breviceps) is found in central and southeastern Texas eastward into western Louisiana and north into southwestern Arkansas and eastern Oklahoma.  A recent study has indicated species-level divergence within this taxon based on analyses of the mitochondrial cytochrome b gene.  However, these conclusions were tentative due to both limited sample size and limited geographic sampling. Our goal is to examine the nuclear genomic DNA of pocket gophers sampled from throughout the range of G. breviceps using the molecular technique of AFLP (amplified fragment length polymorphisms).  We hypothesize that this study will help to clarify patterns of genetic divergence across the distribution of G. breviceps and either support or refute the existence of a cryptic species as previously postulated.


MARK NELSON and BETH WATSON. "Using AFLP to Examine Genetic Diversity in the Texas Kangaroo Rat."
Faculty Mentors: Dr. Allan Nelson, Jim Goetze

The Texas Kangaroo Rat’s (Dipodomys elator)  range once included 10 counties in North Central Texas and Southern Oklahoma.   Its range has been in a state of  decline and today it may only occur in five counties in Texas:  Archer, Childress, Hardeman, Motley, and Wichita.  Previously it was known from state-owned land including Copper Breaks State Park in Hardeman County.  Based on preliminary trapping surveys, it does not occur in the park anymore because it has been managed as a mid-grass prairie without disturbances such as fire and grazing.  However, we have collected individuals  near the park in more suitable habitat.  The decline in populations throughout the range of the Texas kangaroo rat is likely due to loss of short-grass prairie habitat and fragmentation caused by agriculture.  Isolation of populations by farm fields has likely created genetic bottlenecks that have lowered genetic diversity.  To examine genetic diversity, we propose the use of Amplified Fragment Length Polymorphism (AFLP) markers to examine populations occurring in Wichita and Hardeman counties of Texas.These two counties represent the eastern and western margins of the known range of the species.   This data could provide critical information needed  to update the conservation status of D. elator and possibly to use as a baseline for making decisions about restoring populations on state-owned lands.   The objective of this study is to use AFLP to:  (1) ascertain the genetic diversity within populations of D. elator and  (2) examine patterns of population structuring that could be important in managing this state-threatened species.


MARY NGUYEN and SAMSHER RAWAL. "Development of a qPCR Assay to Detect Penicillium marneffei."
Faculty Mentors: Dr. Sally Lewis, Dr. Sally Hoger, Dr. Sara Taylor

Penicillium marneffei is a very pathogenic, dimorphic fungus/yeast that is endemic to Southeast Asia. P. marneffei is an opportunistic pathogen, establishing itself in immunocompromised individuals.  In the United States, infection with P. marneffei should be of concern considering today’s increased globalization and travel by the world population.  Identification of yeasts and molds involves time consuming culture, resulting in delayed diagnosis and treatment.  The development of a rapid and reliable diagnostic tool for the identification of Campylobacter species is warranted in order to improve patient outcomes.  In this study, our purpose is to develop a real-time PCR assay with specificity for Penicillium marneffei.  We designed and validated  specific primers targeting a unique sequence of 80 base pairs within the MP-1 gene specific for P. marneffei.  Our assay utilized the fluorescein (FAM) labeled hydrolysis probes for the detection of the amplification of the specific gene.  Initial results were able to amplify and detect ATCC isolate 24100 of P. marneffei with appropriate negative controls. This experimental assay is still in its developmental process and further research utilizing clinical specimens is necessary.


BETHANY STEVENS and ALICIA SANDERS. "Development of Microsatellite Markers for the Cotton Rat (Sigmodon Hispidus)
Faculty Mentor: Dr. Russell Pfau

This study was performed to develop microsatellite markers for the hispid cotton rat (Sigmodon hispidus).  S. hispidus occurs throughout the southern United States and, because of their abundance, has been the subject of much ecological research.  There are two genetic lineages that are morphological indistinguishable, one occurring in the southeastern U. S., and the other in the southwestern U. S.  These two lineages are known to hybridize in eastern Texas.  Microsatellite markers are a type of repetitive DNA consisting of short repeat units and have a high mutation rate resulting in much genetic variation that can be used to address population genetic questions. We isolated microsatellites using the FIASCO (Fast Isolation by AFLP of Sequences Containing Repeats) technique. FIASCO works by cutting the genome into fragments and attaching known DNA fragments onto the ends.  Fragments are then amplified by polymerase chain reaction and biotinylated microsatellite probes are allowed to hybridize to microsatellite-containing DNA fragments.  Streptavidin-coated magnetic beads are added and bind to the biotinylated probes.  A magnet is then used to separate fragments containing microsatellites from those that do not.  Fragments are then inserted into plasmids and cloned into Escherichia coli.  Plasmids are then isolated and sequenced to verify the presence of a microsatellite and to determine the flanking DNA sequences.  We designed primers for 2 microsatellite loci (named C30 and D21) using DNA from a single individual of the western lineage.  Both loci were used to assess genetic variation in two populations, one western and one eastern, and both loci were found to be polymorphic.  For locus C30, genotypes were obtained for nineteen individuals of the western population and three individuals of the eastern population. Nine alleles were identified in the western population and 84.2% of the individuals were heterozygous.  The eastern population had four alleles and a heterozygosity of 67.7%.  For locus D21, genotypes were obtained for seventeen individuals in the western population and six individuals in the eastern population.  Twelve alleles were observed in the western population and heterozygosity was 82.4%. The eastern population had 4 alleles and a heterozygosity of 100%.  Additional primer pairs have been designed but have not been tested.  Once additional primers have been designed and tested, these microsatellite markers can be used in a variety of studies including analysis of the hybrid zone, assessment of multiple paternity, and effects of landscape on population structure.


ELIZABETH WATSON, MARK NELSON and JIM GOETZE. "Comparison of Ecological Preferences for the Texas Kangaroo Rat."
Faculty Mentor: Dr. Allan Nelson

Assessing the habitat preferences of the Texas Kangaroo Rat (Dipodomys elator) is critical to ensuring the continuation of this state-threatened species.  Knowledge of habitat preference can assist researchers in locating new populations and play a role in prairie restorations that include this endemic species.  To this end, investigations of burrow habitats were conducted in a heavily-grazed pasture site in north-central Texas.  Ten total burrows were examined in late May of 2005.   The Texas kangaroo rat appears to favor elevated, well-drained, clay loam soils.  Burrows were also associated with high percentages of bare ground, and short vegetation.  Vegetation was indicative of disturbance caused by heavy grazing.  Little barley (Hordeum pusillum) was always the dominant grass and Virginia pepperweed (Lepidium virginicum) the dominant forb. Co-dominant woody vegetation included mesquite (Prosopis glandulosa) and lotebush (Zizyphus obtusifolia).  A second population of kangaroo rats were examined in May 2007 in a moderately grazed, well-drained, clay loam pastureland in Wichita County.   The objective of this study is to compare burrow ecology at this site with that of the heavily grazed site in relation to coverage, vegetation height, richness, and dominant vegetation heights.  This type of data is critical in understanding habitat preferences and for locating suitable areas for prairie restoration.