ItemCardiac performance as a function of temperature in the larvae of the American lobster (Homarus americanus)Karlsson, Julie; Jane, Aubrey; Frederich, Markus; Rasher, Douglas; Waller, Jesica; Annis, Eric; Annis, Eric; Hood College Biology; Hood College Departmental HonorsAmerican lobster larvae are subject to rapidly warming waters in the Gulf of Maine due to climate change, yet gaps remain in our understanding of their physiological response to temperature stress. Cardiac performance and respiration are metabolic metrics for assessing response to thermal stress. We measured heart rate, stroke volume, cardiac output and gill bailer beats in lab-reared stage I-IV larvae, and wild-caught stage IV larvae at 4, 8, 18, 26 and 30°C. Heart rate increased with temperature in all stages but there were no significant differences between stages of laboratory reared larvae, nor between lab reared and wild-caught stage IV larvae. Stroke volume and cardiac output decreased at high and low temperature extremes. Gill bailer beats increased with temperature but decreased at 30°C. Our results suggest that heart rate alone is an insufficient metric for thermal stress in lobster larvae and misses non-linear relationships between cardiac function and temperature. Reduced cardiac output at high and low temperatures is consistent with the hypothesis that thermal tolerances are defined by limitations in the ability to deliver oxygen to the tissues. ItemUsing Angler Surveys to Record Recreational Fishing Interactions with Turtles in the Chesapeake Bay(2018-04-23) Bulgarelli, Kyle R.; Ferrier, Drew; Hood College Biology; Hood College Departmental HonorsThe conservation of sea turtles is of concern on a global level. Because of human impacts like global warming, pollution, and commercial fishing, sea turtle populations are being threatened. Conservation groups are working to counteract commercial fishing to protect sea turtles from becoming bycatch. This protects turtles in major bodies of water and from large commercial vessels, but the sea turtles are still threatened by recreational fishing, which is generally undocumented. Large bays and other coastal bodies of water are areas where sea turtles may interact with fishermen. ItemDirected Evolution of Pectin Methylesterase to Higher Activity(2018-04) Decker, Kelsey; Laufer, Craig; Hood College Biology; Hood College Departmental HonorsBiofuels have become an increasing interest over the last decade as the cost of fuel has continued to fluctuate and the sources of fossil fuels have been slowly depleting (Faaij 2006). In general, biofuels, also known as alternative fuel, are renewable energy sources that are produced from raw biological resources referred to as biomass (Demirbas 2009). Several advantages to biofuels include the high availability of biomass, the reduction in greenhouse gas emissions, and the elimination of dependence on fossil fuels found in other countries. Unlike fossil fuels, biofuels are renewable energy sources meaning they can be produced or replenished within a short period of time (Ragauskas 2006). Fossil fuels, such as crude oil that is formed from the buried remains of plants and animals that lived millions of years ago, will soon run out at the rate at which the fossil fuels are currently being consumed. On the other hand, biofuels are produced at a much faster rate and the supply of biomass feedstocks is not fixed. These feedstocks can include plant and algal material such as food and agricultural waste as well as municipal solid waste and yard clippings. These wastes, that would normally be unused, can now be used to generate energy, thus recycling waste that would be otherwise unusable. In addition, energy biomass crops such as switchgrass, energy beets, or poplars are being considered as other potential feedstocks (Demirbas 2009). ItemCloning and Characterization of the Pectin Methylesterase Gene of Pectobacterium wasabiae(2016-05) Spall, Ammarah; Laufer, Craig; Hood College Biology; Hood College Departmental HonorsAs worldwide fossil fuel consumption continues to occur at an increasing rate, the need to look for alternative energy sources becomes apparent. Fossil fuels, such as oil, natural gas, and coal, constitute a finite source of energy that powers factories, furnaces, and homes all over the world. The Energy Information Administration (EIA) reported in its Annual Energy Outlook 2015 that fossil fuels contribute to 67.2% of the United States electric energy consumption. The EIA also sees fossils fuels maintaining their status as America’s leading source of energy consumption until 2040 (1). As global climate change continues to occur due to the burning of fossil fuels, this fact should cause for alarm. ItemCharacterization and Identification of a Novel Pectinolytic Bacteria(2016-05) Bullard-Sisken, Jonathan; Laufer, Craig; Hood College Biology; Hood College Departmental HonorsOne of the main contributing factors to global climate change is the emission of greenhouse gases produced from combustion of fossil fuels, which trap heat by preventing infrared radiation from leaving the atmosphere. Carbon dioxide is the most abundant greenhouse gas and accounted for 80.9% of all greenhouse gas emissions in the United States in 2014, of which 26% were related to transportation. Due to the depletion of non-renewable energy sources and concern for the environment, focus is beginning to shift toward developing renewable energy sources. In the past couple of decades, production of electricity from solar and wind energy has seen much growth. However, electric cars are few and far between, so biomass is currently the best option for producing clean transportation fuels. ItemDirected Evolution of Pectin Methylesterases to Optimize Their Use in a Biofuels Application(2017-04) Mankowitz, Rachel; Laufer, Craig; Hood College Biology; Hood College Departmental HonorsOne of the biggest challenges facing our society today is finding more effective ways to produce cleaner energy. With the uncertainty of crude oil prices, depletion of non-renewable energy resources, rising levels of greenhouse gas emissions, as well as political instability in producing countries, biofuels are a promising alternative (2). Biofuel is defined as fuel that is predominantly produced from biomass, which is simply organic matter (5). The key players in the production of biofuels are the enzymes produced by microbes that are capable of breaking down plant biomass into individual sugar monomers that can be fermented into biofuels such as ethanol. ItemAnalyzing Egg Laying Behaviors in C. elegans Based on Bacterial Food Sources(2016-04) Jones, Georgette; Hood College Biology; Hood College Departmental HonorsCaenorhabditis elegans is a non-parasitic nematode that feeds on soil bacteria. It has been shown that these organisms prefer to consume strains of E. coli bacteria, which are Gram negative, over strains of Bacillus bacteria, which are Gram positive. It was hypothesized that the food preference of the nematodes relates to the survival of the fittest concept, so that worms who consume Gram negative bacteria have greater reproductive success than worms who consume Gram positive bacteria. To test this hypothesis, the nematodes were fed two types of Gram negative bacteria and one type of Gram positive bacteria, and reproductive success was measured as the number of eggs the worms could lay after a specific time period. The results showed that the number of eggs laid by the worms fed Gram positive bacteria was significantly less than the number of eggs laid by worms fed Gram negative bacteria. In addition, a gene expression study using real-time PCR to amplify two egg-laying genes and two dauer-formation genes was conducted. Three of the four genes investigated are part of the TGFβ superfamily of genes that are known to be involved in many stages of reproduction and embryogenesis. The results showed that worms fed Gram negative bacteria had a higher expression of the genes tested than the worms fed Gram positive bacteria, concluding that reproductive abilities of nematodes can be affected by a food source. The mechanism by which bacterial food source affects reproduction needs further investigation, but these data indicate there may be a link between the immune system, diet, and reproduction. ItemAnalysis of the Stability of Microbial Consortia Grown on Pectin(2017-04) Sellers, Ian; Laufer, Criag; Hood College Biology; Hood College Departmental HonorsBacteria are social organisms that interact within and between other species while simultaneously responding to external stimuli from their surrounding environment (11). Bacterial communities are among the most diverse of ecological communities in that they vary drastically in species composition, niches occupied, and influence on different environments. Understanding the development and implications of these bacterial communities is of great importance, because they not only influence our personal health, but also our surrounding environment. Determining exactly how these communities form is a complex issue that is not completely understood. ItemIdentification of HIV Mutations in Donors Experiencing Virologic Failure on Raltegravir-Containing Regimens(2022-04-25) Barthlow, Cailyn; Smith, Oney; Hood College Department of Biology; Departmental HonorsHIV still remains a public global health concern with 38 million people living with HIV. There is no cure to HIV, however combination antiretroviral therapy (ART) is a highly effective treatment that suppresses the virus. Despite the effectiveness of ART, drug resistance mutations can arise, typically found in the target gene. However, previous in vitro studies have identified drug resistance mutations to integrase inhibitors in non-target genes. In this study, we investigated whether mutations in non-target genes, gag Nucleocapsid (NC) and/or nef (3’PPT) might also contribute to drug resistance and virologic failure in five individuals with HIV failing Raltegravir (an integrase inhibitor). Single- Genome Sequencing (SGS) was performed on plasma-derived viruses from pre-ART and early and/or late ART failure samples from these participants, then the gag and nef sequences were analyzed for mutations. We specifically identified a mutation, V24I, in the Nucleocapsid region of gag in vivo that is similar to previously identified resistance mutations in vitro. This analysis suggests that non-target genes may contribute to the emergence of drug resistance in vivo. ItemStudies on the Cellulase Inhibition by Phenols from Hydrothermal Pretreated Soybean WastesJackson, Armoni; Hood College Department of Biology; Hood College Departmental HonorsBiofuels can be generated through the processing of lignocellulosic plant material. Chemical and biological processes known as pretreatment are utilized to process plant material. Pretreatment contributes to the breakdown of the crystalline structure of lignocellulosic materials. A method of pretreatment is hydrothermal pretreatment with various acids and bases. This study addresses the effect of hydrothermal pretreatment with sodium hydroxide (NaOH) or hydrogen peroxide (H2O2) on enzymatic hydrolysis of soybean straw. Hydrothermal pretreatment of soybean straws (10% w/v) was carried out with either sodium hydroxide (1% v/v, NaOH) or hydrogen peroxide (1% v/v, H2O2) at 121°C for 60 min to evaluate the effect of water-soluble inhibitors (released during the pretreatment) on cellulolytic enzymes. The cellulose in pretreated solids (1% w/v glucan) was enzymatically hydrolyzed for 72 h with 25 mg enzyme protein/g glucan in the presence of either buffer or pretreated liquor (rich in phenols and lignin-derived molecules). The hydrolysis of NaOH treated solids in buffer gave a 57% cellulose conversion to glucose versus a 39% glucose yield from the H2O2 treated solids. When pretreated liquor was applied, NaOH and H2O2 treated solids had a 20% and 30% glucose yield, respectively, indicating the suppression of enzyme activity by non-productive bindings between enzyme proteins and inhibitors. ItemDetermining the Thermal Tolerance of the Stage V Larvae of Homarus americanus(2021-04) Sever, ReidAnn; Annis, Dr. Eric; Hood College Biology; Hood College Departmental HonorsIn response to warming water temperatures in the Gulf of Maine, this experiment attempts to define the thermal tolerance of Stage IV and Stage V Homarus americanus in order to test for an ontogenetic shift between the tolerances. I measured oxygen consumption in active and resting trials of Stage IV and Stage V Homarus americanus larvae to determine the average aerobic scope of each stage from 4℃ to 32℃. The Stage V larvae had twice the aerobic scope between 6℃ and 16℃ compared to that of the Stage IV’s. This supports the hypothesis that the larvae of H. americanus are more cold tolerant in Stage V. ItemEngineering melting temperatures of carbohydrate binding modules through site-directed mutagenesis(2019-05-01) Jablunovsky, Anastazia; Laufer, Craig; Chemistry and Biology; Hood College Departmental HonorsBiofuels are a promising alternative to environmentally harmful fossil fuels. However, in order to displace fossil fuels, biofuels must be competitive in cost. Currently, the production process is the most significant issue regarding their cost, despite feedstock material being fairly inexpensive. One of the largest contributors to this cost is the use of enzymes to breakdown the biomass into simple sugars for subsequent use. If the enzymes used in this process could be easily recycled, the total cost of production would be significantly reduced. This project targets carbohydrate binding modules (CBMs) in an attempt to lower their melting temperature (Tm) through mutagenesis. These modules allow the holoenzyme (CBM with the catalytic domain) to bind to the substrate. Engineering a Tm of the CBM that was below the Tm of the catalytic domain would allow binding to be turned “off” for enzyme recovery, and later turned back “on” for future reactions. Tm values were established for three CBMs as well as refolding capabilities after heat treatment. Assays were developed for CBMs fused with green fluorescent protein by evaluating fluorescence signal on cellulose substrates in order to assess binding activity. Potential amino acid substitutions were identified in CBMs 11 and 44 from Ruminoclostridium thermocellum that were predicted to lower the Tm. Two of these mutations in CBM 44 that specifically interfere with Ca+ stabilization were created and sequenced. These mutants will be tested for changes in Tm and binding capabilities. ItemEvaluation of Biological and cDNA Clone-Derived Stocks of Venezuelan Equine Encephalitis Virus TC-83 Vaccine Strain(2019-04) Ball, Matthew; Boyd, Ann; Smith, Oney; Glass, Pamela; Department of Biology; Departmental HonorsThe objective of this research study was to evaluate the potential antibody escape mutations identified. The specific aims of this study were as follows: Aim 1: Generate mutant virus stocks containing the identified mutations, singly or in combination; Aim 2: Characterize the mutant viruses, along with the biological or clone-derived wild-type viruses, for fitness and ability to bind antibody 1A3B-7. The results of these studies demonstrated that the biological and clone-derived VEEV TC-83 viruses behaved similarly with regards to replication in cell culture and antibody neutralization. Though there appeared to be some differences in virulence in the C3H/HeN mouse model, the difference in survival was not statistically different. The results of these studies were important to provide the necessary foundation for characterization of the potential antibody escape mutations as the mutants viruses must be clone-derived from cDNA. Future research will observe the two escape mutant viruses with nucleotide changes in the E2 glycoprotein at nucleotide 9177 and 9189. Evaluation of the escape mutants may suggest the most effective way to develop a therapeutic against VEEV would be through the administration of a broadly neutralizing antibody or a cocktail of monoclonal antibodies which recognize multiple epitopes on the surface of the virus particle.