Undergraduate Research

Amber Kay

Analysis of the effects of phosphorylation on bovine herpesvirus-1 transcription

During productive infection Bovine Herpesvirus-1 (BHV-1) initiates transcription in a characteristic temporal pattern. Viral genes are classified as Immediate Early, Early, and Late, and their transcripts can be quantified by reverse-transcriptase polymerase chain reaction (RTPCR). We are interested in studying the effects of phosphorylation on viral transcription during productive infection. We designed primers to amplify three Immediate Early genes, bovine Infected Cell Protein 4 (bICP4), bICP-22, and bICP-0, two Early genes, Thymidine Kinase (TK) and Ribonucleotide Reductase (RR), two Late genes, Viral Protein 16 (VP 16) and Glycoprotein C (GC), and cellular control protein, bovine Growth Hormone (bGH) using the Primer3 Plus software. Optimization of primers for PCR was conducted by taking into account specific annealing temperature, MgCl2 concentration, DNA polymerase concentration, content of glycerol, and the total amount of DNA added. We have found optimum conditions for the amplification of all the selected genes. We infected cells in the presence or absence of phosphatase inhibitors at 2 and 4 hours post infection. Due to previous related experiments conducted in our lab, we expect to find that an over-phosphorylated environment will cause the virus to produce a decreased amount of mRNA transcripts therefore reducing the productivity of infection.

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Andrew House

Reproduction strategies for endangered frogs

He's always liked frogs and toads, but until he came to Mississippi State, Andrew House never thought he'd get a job working with them.

The junior from Kennedy, Alabama, works in an amphibian laboratory, a part of the Department of Biochemistry, Molecular Biology, Entomology, and Plant Pathology. House's work enhances his studies in biochemistry and natural resource conservation, he said.

"I'm really glad I got into the lab where students have the opportunity to do something different: to work with critically endangered frogs and try to help them reproduce," he said. "Frogs are environmental indicators that show the health of the ecosystem because they need water and land to reproduce and be able to thrive and survive."

The research laboratory houses critically endangered Mississippi gopher frogs, as well as Boreal toads. Researchers also work with Fowler's toads, which are not endangered but offer relevant information about the species' physiology. The team focuses on assistive reproductive technology to encourage species to reproduce, House explained.

After classes ended in May, he presented the researchers' work at the Amphibian Conservation Research Symposium 2014, held by the Zoological Society of London. House said getting to speak at the prestigious gathering was an exciting opportunity for him, and he hopes it will be the first of many.

Blaire Fleming

Effects of dietary melatonin supplementation on total serum nitrites and antioxidant capacity of late gestating Holstein heifers

The objective of my undergraduate research project was to determine total serum nitrites and antioxidant capacity of late gestating Holstein heifers following dietary melatonin supplementation. After obtaining serum samples and running the appropriate assays, it was found that total antioxidant capacity was increased by 40% in the heifers treated with melatonin compared to the heifers that were not. These heifers have better protection from oxidative stress experienced during pregnancy. The treatment had no effect on the total serum nitrites, but a main effect of gestational day was observed. This shows that nitric oxide concentrations increase as pregnancy progresses.

We decided to expand the study by determining serum components of the offspring from these heifers. I have obtained data regarding total antioxidant capacity and anti-Müllerian hormone concentrations and am currently working to obtain total serum nitrites for the offspring as well. Total antioxidant capacity for the offspring was not affected by the dietary melatonin supplementation during gestation. However, we did see a slight tendency for anti-Müllerian hormone concentrations to be higher in the calves whose dams received the supplementation. These calves could possibly have higher ovarian reserves which could increase their reproductive performance, but that is speculation at this point. Seeing as we did not see a treatment effect on serum nitrites in the heifers, I doubt that there will be an effect on the offspring serum. However, science can surprise us, so I will wait to interpret the data before drawing any definite conclusions. I hope to continue learning all that I can in the research field and am thankful for the opportunity I have been given.

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Caleb McKee

Development of a system to measure skinning incidence in sweetpotato

Sweetpotato is a high value crop which accounts for over $132 million in value for Mississippi's economy. Skinning during harvest and handling has a direct effect on storage performance and value. Skinning allows moisture to exit the root and allows fungal or bacterial rots to develop. Current methods of skinning incidence measurement focus on manually counting the number of scars and can include visual classification of the size of the scars.

The objectives of this study were to develop and validate a system to rapidly and accurately quantify skinning incidence. The image capture system was an off-the-shelf DSLR camera, a horizontal tripod, a computer running ImageJ software, and a fixture onto which the sweetpotato root is placed. A root on the fixture could be rotated at 15° intervals. System accuracy was assessed using targets with known ratios between colors representing sweetpotato skin and flesh.

The number of images used to estimate whole-root skinning percentage has a direct influence on information quality. Capturing an image at every 15° rotation results in 24 images which creates better data than two images captured at every 180°. Therefore differences in data quality was assessed using 2, 4, 8, 12, and 24 images captured at 180, 90, 45, 30, and 15 degrees of rotation, respectively. Three sweetpotato roots were analyzed at each increment and each analysis was repeated three times. At greater than 2% skinning, the mean system error was approximately 2%. Below 2% skinning, the error increased but in that range there would likely be little impact on storage quality.

There was no significant advantage to analyzing more than four images at 90° rotation as compared to 24 images at 15°. This system can open new research directions to analyze storage or cultivar performance based on the quantitative measure of sweetpotato skinning incidence.

Christina Cooper

Surface area and envelope volume of growing broilers from 35 d to 63 d of age

Poultry is Mississippi's most valuable agricultural commodity valued at $2.3 billion and produced on 1,478 farms in 2012. 3D models of broilers are needed to find ways to increase heat removal, thus be able to increase bird performance (better feed conversion ratios) or use fewer fans (less energy) or a combination of both. 3D models can be utilized in a broiler house computational fluid dynamics (CFD) model to improve our understanding of the bird-airflow interface. Therefore, the objective of this study was to characterize surface area and envelope volume of growing broiler birds from 35d to 63 d of age. Live broiler chickens were successfully scanned at 35, 49, 63 days of age with a commercially available 3D digitizer system. Adjustments were made to accommodate live broilers such as cameras angles and placement, light intensity, and bird movement. 3D surface models were successfully rendered however, issues arose with the calculation of surface area thus affecting volume and S/V ratio for both male and female birds. Models can currently be used to gather physical measurements but the study must be repeated to improve the model surface area measurements.

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Darby Dillard

Investigation of the seasonal variations of temperature and relative humidity on fertility potential of indoor boars: Preliminary data

The objective of my undergraduate research project was to evaluate the impacts of external temperature and relative humidity variations on semen production of boars maintained in a thermo-regulated barn. The study began November 2013, and will conclude October 2014. Currently, the winter results (November, December, January, and February) indicate that despite the thermoregulation of the barn, environmental temperature and relative humidity variations still affect semen production. Protein analysis and gene expression will be conducted with the expectations to find both qualitative and quantitative differences between samples collected on the warmest days and the coldest days. We expect to find that despite these boars being maintained in a temperature regulated barn, they are still suffering from seasonal variations.

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Emily West

Presence and absence of COX8 in reptile transcriptomes

I searched for genes encoding for COX8 (cytochrome c oxidase subunit 8) in Iguanids (composed of chameleons, iguanas, agamids). These genes are important because this subunit is needed to effectively make ATP in the electron transport chain, and they are present in all other eukaryotes. Other studies have shown that chameleons do not have COX8, and I am checking to see if I can specify further where the gene coding for it was lost as well as make sure it is gone for certain in iguanids.

I obtained the blood transcriptomes of twelve squamate reptiles (Texas blind snake, red eyed crocodile skink, tokay gecko, prairie rattlesnake, yellow racer, savannah monitor, blonde hognose snake, bearded dragon, boa constrictor, chameleon, Asian vine snake, gold tegu), and both COX8 isoforms from Ensembl. I used Blast to compare the snake transcriptomes (previous studies prove snakes contain COX8) to COX8. Then I extracted the contigs that coded for COX8 and compared them to the rest of the reptiles and recorded if COX8 was present. In order to validate my findings, I did a control search test using the nuclear encoding cytochrome c, and the mitochondrially encoding cytochrome b.

I queried for COX8 within my transcriptomes, and discovered that it was missing in iguanids and several other orders. While I believe that it completely missing from Iguanids, I cannot be certain unless I were to obtain more data, such as a complete genome.

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Ethan Norvell

A look at young cotton plant's roots in drought and low temperature situations

Better understanding of cotton's roots during drought and low temperatures is needed in order to grow it more effectively. An experiment was conducted by varying night and day temperatures (22/14, 30/22 degrees Celsius), and by varying water availability (well watered, drought stress). Nine cotton cultivars from nine different breeding programs in nine different states were grown in a pure sand soil under these conditions. Time of seed sprout, soil moisture, and evapotranspiration measurements were taken. Twenty days from the time of planting the plants will be pulled up in order to scan the roots. It is expected that the plants in the well watered, optimal temperature units will be healthier than the plants in the drought stressed or lower temperature units. In addition to this it is expected that the cultivars will perform based on their breeding origin; the cultivars from Arizona and New Mexico will do well in the drought stressed units. The relationship between roots, water availability, and temperature will be valuable in making decisions such as planting dates.

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Hazel Buka

An integrated watershed approach to water and sanitation hygiene priorities through a narrative review of Lake Chivero, Zimbabwe

Hazel Buka, originally from Harare, Zimbabwe, is an agricultural engineering technology and business major. She has a concentration in natural resources and management. Her projected graduation date is 2016. Buka attended her first two years of college at Chibero College of Agriculture in Zimbabwe before coming to MSU. Through a partnership established between these two educational institutions, she traveled to MSU to finish her bachelor's degree.

As an undergraduate research scholar, Buka's project is a review and assessment of studies on Lake Chivero, Zimbabwe, a main water source for the area. According to Buka's report, various factors have impacted the lake's water quality. After collecting and analyzing water quality data, Buka proposed two possible solutions: the reduction of the overflow of nutrients going into the lake and educational outreach on clean water practices. The work was presented at the Mississippi Water Resources Conference held in Jackson, Mississippi in April. The next step is to secure additional funding to implement solutions.

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Hillary Clements

The effects of phosphorylation during bovine herpesvirus-1 (BHV-1) infection

Phosphorylation is a major means of post-translational modification and has previously shown to be tightly regulated throughout the course of BHV-1 infection. Our goals included to study the effects of phosphorylation manipulation on viral replication and infection, to purify the BHV-1 virion, and to analyze the virion for its phosphoproteome. Wild-type virions and virions produced in an over-phosphorylated environment will be compared to identify any differences in constituents due to phosphorylation manipulation.

Currently, we have concluded there is very little difference in viral replication and infection in wild-type and over-phosphorylated BHV-1; although, a slight downward trend was seen in viral production when phosphatase inhibitors were added before infection. This lack of a significant difference in viral titer reinforces our belief that BHV-1 is regulating exactly what is incorporated into progeny virions. On the same note, the trend in decreased viral titer seen with phosphatase inhibitors added before infection, suggests a possible difference in the constituents of progeny virions produced normally and under over-phosphorylated conditions. We are purifying virus in preparation for enrichment of phosphoproteins and subsequent analysis using mass spectrometry. After identification of viral constituents via mass spectrometry, proteomics databases such as Proteome Discoverer will be used to deduce phosphorylation levels of predicted and identified phosphoproteins to determine any variance between the two types of virions.

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Jordan Holley

Locally grown: Examining attitudes and perceptions about organic cotton production and manufacturing between Mississippi cotton growers and consumers

The objective of my undergraduate research project is to spread the awareness of cotton production methods, particularly organic cotton production, and how cotton affects everyday consumer lifestyles. By examining the attitudes and perceptions about organic cotton of Mississippi cotton growers and producers in comparison to fashion conscious consumers, the goal is to understand how consumers and growers/producers feel about organic cotton production in relation to the initiatives the industry is taking to move toward sustainability.

My expected research outcomes are that undergraduate students are not eager about paying the higher price for organic fabric unless the fabric is associated with a popular brand or retailer. Farmers benefit from the higher profit margin and the elimination of harmful chemicals, but farmers will not have significant understand about the organic cotton demand. After analyzing data from 60 research participants, there is a disconnect of information between regional cotton growers/producers and consumers based on the advantages/disadvantages of growing and production processes, quality control, consumer preferences, and competitive price structures/profit margins. It is beneficial for the industry to understand disconnect of information exists in the supply chain because until efforts are made to increase the transparency of information, the organic cotton industry's ability to create a sustainable future will remain limited.

During the Spring 2014 semester, I have been able to make significant progress towards my research goal with the help of my mentor Dr. Charles Freeman, and I have an overall better understanding of how cotton growers/producers and fashion conscious consumers perceive organic cotton production in this region. For further study, I hope to expand my sample size to include other regions and a broader consumer base so that I can compare the attitudes and perceptions of growers/producers and undergraduate students in Mississippi to the regions where organic cotton production is more popular.

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Karley Parker

The effects of housing type and feeding frequency on growth and behavior in dairy calves

Karley Parker, originally from Ellisville, Mississippi, received her Bachelor of Science in Animal and Dairy Sciences in May 2014. She began veterinarian school in June 2014.

As an undergraduate research scholar, she studied the effects of housing type and feeding frequency on growth and behavior in dairy calves. Her research found paired calves had better health scores than the calves that were alone. In the dairy industry, it's often thought that housing calves together increases risk for infection, so the research findings could shape future practices.

The research was presented at the national meeting of the American Dairy Science Association in July 2014. The preliminary data was included in an animal research and Extension grant proposal as well.

Lee Peeples

The role of an absisic acid activated protein kinase in drought tolerance

Drought is the major environmental factor limiting crop productivity. There is therefore a great need for crop improvements that will increase drought tolerance and stabilize crop yield under drought conditions. When plants are subjected to drought, the absisic acid concentrations in leaves increases causing the stomata to close, which enables plants to reduce water loss under drought conditions. The aim of this project is to determine the role of a rice absisic acid activated protein kinase in plant response to drought stress. We were expecting that SAPK10 will be a positive regulator in drought tolerance because of the overexpression lines that it exhibits. And so far we have determined that the absisic acid activated protein kinase SAPK10 is a positive regulator of drought tolerance in rice. However, we are not yet finished. In further studies the target proteins of the SAPK10 kinase are going to be identified.

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Maryam Mohammadi Aragh

Rapid identification of pathogenic Vibrio parahaemolyticus using human red blood cell and urea

Vibrio parahaemolyticus is a pathogen found in fish and shellfish associated with acute gastroenteritis after consuming raw or improperly cooked seafood. Many people worldwide are affected by vibriosis each year and is a major food-borne pathogen plaguing the food industry. Interestingly, few strains of Vibrio parahaemolyticus are pathogenic and the cause of pathogenicity is still a mystery. Consequently, detecting contaminated food products is difficult and the development of rapid-detection methods is in high demand.

Thermostable-direct hemolysin (TDH) and TDH-related hemolysin (TRH) are proteins that are strongly associated with illness. When expressed, these pore-forming proteins rupture red blood cells (hemolysis). In this study, TDH and TRH were used as biomarkers to quickly identify pathogenic strains by observing hemolysis activity on Wagatsuma broth modified with fresh human blood. Traditional detection methods use urease tests to identify pathogenic Vibrio parahaemolyticus contamination; however, current research suggests urease is not a dependable biomarker.

The objective of this study was to investigate how TDH, TRH, and urease are related to hemolysis activity in modified-Wagatsuma broth. This data is valuable for developing an improved rapid-detection method that is fast, easy-to-use, and does not require expensive special equipment. Currently the media and TDH, TRH, urease relationship is complete and the project's new goal is to use Western blot to quantify TDH and TRH expression in modified-Wagatsuma media. This will reveal how modified-Wagatsuma broth affects the severity of TDH and TRH expression, and if altering the ingredients stimulates or inhibits hemolytic activity. Ultimately, this information will help strengthen the reliability of modified-Wagatsuma broth.

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Shelby Mathieu

Investigating the link between stress, temperature, and metabolic inertia in largemouth bass

Goal.—To relate temperature change and handling stress to metabolic inertia in largemouth bass.

Objectives.—To measure standard metabolic rate in largemouth bass acclimated to two temperatures, to measure metabolic rate and time required to return metabolic rate following an abrupt decrease of 4°C, and to measure metabolic rate and time required to stabilize metabolic rate after initiating standard handling stress to induce a stress response.

Current Progress.—In order to meet these objectives, a literature review has been done to determine what information is currently known linking stress, temperature, and metabolic rate in largemouth bass, the centrarchid family of fishes, and teleost fishes in general.

Expected Outcomes.—As seen in previous studies, metabolic rate will stabilize at the four hour mark after a stress response when fishes are acclimatized to 25C. Metabolic rate is expected to stabilize faster after the temperature drop as largemouth bass and teleost fishes in general recover best at lower temperatures.

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Taylor King

Effects Of genipin on decellularized porcine cartilage

Osteochondral articular transfer is the only procedure to repair articular cartilage and immediately restore functional hyaline cartilage to the joint surface. Drawbacks include donor site morbidity, limited tissue availability, risk of graft rejection and disease transmission.

This research aims to develop an alternative method using decellularized, crosslinked porcine osteochondral xenografts (OCXGs). Benefits of xenografts include low cost, abundant supply, and uniform prodcuts. Further benefits from decellularizing and crosslinking include decreased immunogenicity, reduced disease transmission and immediate "off-the-shelf" availability.

Crosslinking collagenous tissue results in greater mechanical strength, more resistance to enzymatic degradation, and reduced immunogenicity. Traditionally, glutaraldehyde has been used for crosslinking. However, it is cytotoxic and kills adjacent tissues. Instead, this study used genipin, a chemical substance from the Gardenia jasminoides Ellis fruit. Genipin is inexpensive and simple to use. Crosslinking density can be adjusted according to genipin concentration. In this study, porcine articular cartilage disks were decellularized via a previously published method for porcine nasal septal cartilage decellularization. The process extracts glycosaminoglycan to allow for greater infiltration of nutrients and host cells. The disks were then crosslinked with 0.01% and 0.1% aqueous genipin for 3 days at room temperature with agitation. Prior to decellularization, the cartilage disks' biphasic properties were determined by confined compression testing. The test was repeated after decellularization and after crosslinking. The aggregate modulus was notably lessened after decellularization but returned to, and in some instances, exceeded that of the fresh disks after crosslinking. This study demonstrates that genipin is a viable alternative for crosslinking OCXGs.

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