THE GLYCOPROTEINS OF PORCINE REPRODUCTIVE AND RESPIRATORY SYNDROME VIRUS AND THEIR ROLE IN INFECTION AND IMMUNITY

The porcine reproductive and respiratory syndrome virus (PRRSV) is an economically important pathogen of swine and is known to cause abortion and infertility in pregnant sows and respiratory distress in piglets. PRRSV contains a major glycoprotein (GP5) and three minor glycoproteins (GP2a, GP3, and GP4) on the virion envelope, all of which are required for infectious virus production. To study their interactions amongst each other and with a cellular receptor for PRRSV, CD163, I cloned each of the viral glycoproteins and CD163 in various expression vectors. My studies have shown that while the GP2a, GP3, and GP4 are co-translationally glycosylated, the GP5 is post-translationally glycosylated. By using co-immunoprecipitation (co-IP) assays, strong interaction was demonstrated between GP4 and GP5 proteins, although weak interactions among the other envelope glycoproteins were also detected. Further, GP4 was found to mediate interactions leading to formation of multiprotein glycoprotein complex. My results also show that GP2a and GP4 proteins are the only two GPs that specifically interact with the CD163 molecule and that glycosylation of these GPs is required for efficient interaction. Based on these studies, I have developed an interactome map of the viral GPs and CD163 and have proposed a model of the viral glycoprotein complex and its interaction with CD163. Studies reported here also show that glycan addition at residue 184 (N184) of GP2a, and residues N42, N50, and N131 of GP3 is essential for recovery of infectious virus. Although single site glycosylation mutants of GP4 had no effect on infectious virus production, introduction of double mutations was lethal. The loss of glycan moieties of GP2a, GP3, and GP4 proteins had no effect on host neutralizing antibody production. Overall, I conclude that the PRRSV glycoproteins are co-translationally and post-translationally glycosylated, the GP4 protein is central to mediating interglycoprotein interactions, and along with GP2a, serves as the viral attachment protein that is responsible for interactions with the viral receptor, CD163. Further, glycosylation of GP2a, GP3, and GP4 proteins is required for infectious virus production, efficient interaction with CD163, but does not play any role in neutralizing antibody response in infected animals.

Description of Paranoplocephala etholeni n. sp. (Cestoda: Anoplocephalidae) in the Meadow Vole Microtus pennsylvanicus, with a Synopsis of Paranoplocehala s. l. in Holarctic Rodents

Paranoplocephala etholeni n. sp, parasitizing the meadow vole Microtus pennsylvanicus in Alaska and Wisconsin, USA. is described Paranaplocephala etholeni is morphologically most closely related to the Nearctic Paranoplocephala ondatrae (Rausch, 1948). Available data suggest that P. etholeni is a host-specific, locally rare species that may have a wide but sporadic geographical distribution in North America. The finding of P. ondatrae-like cestodes in Microtus spp. suggests that this poorly known species may actually be a parasite of voles rather than muskrat (type host). A tabular synopsis of all the known species of Paranoplocephala s. I. in the Holarctic region with their main morphological features is presented.

Paranoplocephala sciuri (Cestoda: Anoplocephalidae), a Parasite of the Northern Flying Squirrel (Glaucomys sabrinus), with a Discussion of Its Systematic Status

This study redescribes Andrya sciuri Rausch, 1947 (Anoplocephalidae) from the northern flying squirrel, Glaucomys sabrinus (Shaw), in North America, to redefine the morphology and generic position of this poorly known cestode. Andrya sciuri is shown to belong unambiguously to the genus Paranoplocephala Lühe, 1910 sensu Haukisalmi and Wickström (2005). Paranoplocephala sciuri is compared with four species that resemble it morphologically, and features that can be used in its identification are presented. It is suggested that P. sciuri has speciated through a shift from arvicoline rodents (voles and lemmings) to G. sabrinus.

Nebraska Builder Award Presented to Dr. Mark Gustafson

Few Nebraskans are as devoted to the University of Nebraska as Mark Gustafson. Driven by his belief that a strong university is key to a strong Nebraska economy, Mark is an advocate for the university in the local, state, and national arenas. He is a Nebraska delegate to the Council for Agricultural, Research, Extension, and Teaching, a national advocacy organization for higher education. Since 1991, he's been a member of Agriculture Builders of Nebraska, Inc., which supports UNL's Institute of Agriculture and Natural Resources, as well as the entire University, and has served three terms as president. He has served on the advisory councils for the UNL chancellor and the NU president and served on UNL's Future Nebraska Taskforce. He holds baccalaureate and master's degrees from UNL and a Ph.D. from the University of California-Berkeley. When he's not volunteering his time, Mark operates the family farm near Mead. He and his wife, Dianne, are the parents of two children - Christopher, a UNL alumnus, and Anneke, a UNL junior.

Development of an Electrochemical Insulin Sensor Based on a High Affinity DNA Sequence Found in the Insulin-linked Polymorphic Region

The detection of pertinent biomarkers has the potential provide an early indication of disease progression before considerable damage has been incurred. A decrease in an individual’s sensitivity to insulin, which may be quantified as the ratio of insulin to glucose in the blood after a glucose pulse, has recently been reported as an early predictor of insulin-dependent diabetes mellitus. Routine measurement of insulin levels is therefore desirable in the care of diabetes-prone individuals. A rapid, simple, and reagentless method for insulin detection would allow for wide-spread screenings that provide earlier signs of diabetes onset. The aim of this thesis is to develop a folding-base electrochemical sensor for the detection of insulin. The sensor described herein consists of a DNA probe immobilized on a gold disc electrode via an alkanethiol linker and embedded in an alkanethiol self-assembled monolayer. The probe is labeled with a redox reporter, which readily transfers electrons to the gold electrode in the absence of insulin. In the presence of insulin, electron transfer is inhibited, presumably due to a binding-induced conformational or dynamic change in the DNA probe that significantly alters the electron-tunneling pathway. A 28-base segment of the insulin-linked polymorphic region that has been reported to bind insulin with high affinity serves as the capture element of the DNA probe. Three probe constructs that vary in their secondary structure and position of the redox label are evaluated for their utility as insulin-sensing elements on the electrochemical platform. The effects of probe modification on secondary structure are also evaluated using circular dichroism spectroscopy.

Genetic Mapping of Quantitative Trait Loci Associated with Bioenergy Traits, and The Assessment of Genetic Variability in Sweet Sorghum (Sorghum bicolor (L.). Moench)

Sweet sorghum, a botanical variety of sorghum is a potential source of bioenergy because high sugar levels accumulate in its stalks. The objectives of this study were to explore the global diversity of sweet sorghum germplasm, and map the genomic regions that are associated with bioenergy traits. In assessing diversity, 142 sweet sorghum accessions were evaluated with three marker types (SSR, SRAP, and morphological markers) to determine the degree of relatedness among the accessions. The traits measured (anthesis date [AD], plant height [PH], biomass yield [BY], and moisture content [MC]) were all significantly different (P<0.05) among accessions. Morphological marker clustered the accessions into five groups based on PH, MC and AD. The three traits accounted for 92.5% of the variation. There were four and five groups based on SRAP and SSR data respectively classifying accessions mainly on their origin or breeding history. The observed difference between SSR and SRAP based clusters could be attributed to the difference in marker type. SSRs amplify any region of the genome whereas SRAP amplify the open reading frames and promoter regions. Comparing the three marker-type clusters, the markers complimented each other in grouping accessions and would be valuable in assisting breeders to select appropriate lines for crossing. In evaluating QTLs that are associated with bioenergy traits, 165 recombinant inbred lines (RILs) were planted at four environments in Nebraska. A genetic linkage map constructed spanned a length of 1541.3 cM, and generated 18 linkage groups that aligned to the 10 sorghum chromosomes. Fourteen QTLs (6 for brix, 3 for BY, 2 each for AD and MC, and 1 for PH) were mapped. QTLs for the traits that were significantly correlated, colocalized in two clusters on linkage group Sbi01b. Both parents contributed beneficial alleles for most of traits measured, supporting the transgressive segregation in this population. Additional work is needed on exploiting the usefulness of chromosome 1 in breeding sorghum for bioenergy.

Student and Faculty Perceptions of Technology’s Usefulness in Community College General Education Courses

Educational institutions of all levels invest large amounts of time and resources into instructional technology, with the goal of enhancing the educational effectiveness of the learning environment. The decisions made by instructors and institutions regarding the implementation of technology are guided by perceptions of usefulness held by those who are in control. The primary objective of this mixed methods study was to examine the student and faculty perceptions of technology being used in general education courses at a community college. This study builds upon and challenges the assertions of writers such as Prensky (2001a, 2001b) and Tapscott (1998) who claim that a vast difference in technology perception exists between generational groups, resulting in a diminished usefulness of technology in instruction. In this study, data were gathered through student surveys and interviews, and through faculty surveys and interviews. Analysis of the data used Kendall’s Tau test for correlation between various student and faculty variables in various groupings, and also typological analysis of the transcribed interview data. The analysis of the quantitative data revealed no relationship between age and perception of technology’s usefulness. A positive relationship was found to exist between the perception of the frequency of technology use and the perception of technology’s effectiveness, suggesting that both faculty members and students believed that the more technology is used, the more useful it is in instruction. The analysis of the qualitative data revealed that both faculty and students perceive technology to be useful, and that the most significant barriers to technology’s usefulness include faulty hardware and software systems,lack of user support, and lack of training for faculty. The results of the study suggest that the differences in perception of technology between generations that are proposed by Prensky may not exist when comparing adults from the younger generation with adults from the older generation. Further, the study suggests that institutions continue to invest in instructional technology, with a focus on high levels of support and training for faculty, and more universal availability of specific technologies, including web access, in class video, and presentation software. Adviser: Ronald Joekel

Holocarboxylase Synthetase-dependent Biotinylation of Histone H4

Holocarboxylase synthetase (HCS) catalyzes the binding of biotin to lysine (K) residues in histones H3 and H4. Histone biotinylation marks play important roles in the repression of genes and retrotransposons. Preliminary studies suggested that K16 in histone H4 is a target for biotinylation by HCS. Here we demonstrated that H4K16bio is overrepresented in repeat regions {pericentromeric alpha satellite repeats; long terminal repeats (LTR)} compared with euchromatin promoters. H4K16bio was also enriched in the repressed interleukin-2 gene promoter. The enrichment at LTR22 and promoter 1 of the sodium-dependent multivitamin transporter (SMVT) depended on biotin supply; and was significantly lower in fibroblasts from an HCS-deficient patient compared with an HCS wild-type control. We conclude that H4K16bio is a real phenomenon and plays a role in the transcriptional repression of repeats and genes. HCS catalyzes the covalent binding of biotin to carboxylases, in addition to its role as a histone biotinyl ligase. HCS null individuals are not viable whereas HCS deficiency is linked to developmental delays and phenotypes such as short life span and low stress resistance. Here, we developed a 96-well plate assay for high-throughput analysis of HCS based on the detection of biotinylated p67 using IRDye-streptavidin and infrared spectroscopy. We demonstrated that the catalytic activity of rHCS depends on temperature and time, and proposed optimal substrate and enzyme concentrations to ensure ideal measurement of rHCS activity and its kinetics. Additionally, we demonstrated that this assay is sensitive enough to detect biotinylation of p67 by endogenous HCS from Jurkat lymphoid cells.

Nebraska Agribusiness Club 40th Annual Awards Banquet

Did you know if you look up the word "integrity" in the dictionary, the definition is Alan Moeller?

48th Annual Nebraska Agri-Business Association Meeting

Thank you so very much for this opportunity to speak today at this 48th Annual Nebraska Agri-Business Association meeting. Dr. Kyle Hoagland, Water Center Director in the Institute of Agriculture and Natural Resources at the University of Nebraska-Lincoln, and I are particularly pleased to be asked to talk with you today about water, a key concern in our state and in our world. New water regulations affecting Nebraska's agricultural producers, numerous legislative discussions, and stories of neighbors contesting water use by neighbors have been in the news and on our minds. We all know that will continue.

ABN Summer Meeting

It is such a pleasure to have you on campus with us today. While we couldn't do anything to turn the heat down outside, at least we've had some hot topics to go right along with it! I sincerely hope you've enjoyed the wide variety of information we've shared with you today. It is our pleasure to have this opportunity to host you here on campus, and I thank everyone who has been part of presenting and putting this day's program together. I also thank each of you, and every member of ABN, for all you do in support of Nebraska agriculture, the Institute, and the university.

Nebraska Pork Summit Remarks

I'd like to begin my brief remarks today with some thank you's. First, thank you, Governor Heineman, for sharing your views with us just now, and for your efforts on behalf of Nebraska agriculture as you work with others to grow Nebraska markets. We also thank you for your strong support of the University of Nebraska.

U.S. Meat Export Seminar Welcome

What a great pleasure it is to welcome you to our campus today. We are so very glad to have you here with us in beef country. I'm sure you are all very well aware beef is big business in our powerhouse agricultural state. And as a connoisseur of the product, let me tell you - it's very good beef!

LEAD Presentation

Thank you for inviting us to share with you some information about the Institute of Agriculture and Natural Resources, and what we are doing for Nebraska. As a stakeholder in Nebraska's land-grant university, you and every other Nebraskan have every right at any time to ask us what we've done for you and your neighbors lately. We welcome the question. There is so much excellent work being accomplished on this campus and throughout Nebraska that I am delighted each time I have the opportunity to talk about it. The biggest problem I find is stopping - there's always so much more to say - than there is time to say it in!

The Effects of High Pressure Processing on Peanut Sauce Inoculated with Salmonella

Recent Salmonella outbreaks have prompted the need for new processing options for peanut products. Traditional heating kill-steps have shown to be ineffective in lipid-rich matrices such as peanut products. High pressure processing is one such option for peanut sauce because it has a high water activity, which has proved to be a large contributing factor in microbial lethality due to high pressure processing. Four different formulations of peanut sauce were inoculated with a five strain Salmonella cocktail and high pressure processed. Results indicate that increasing pressure or increasing hold time increases log10 reductions. The Weibull model was fitted to each kill curve, with b and n values significantly optimized for each curve (p-value < 0.05). Most curves had an n parameter value less than 1, indicating that the population had a dramatic initial reduction, but tailed off as time increased, leaving a small resistant population. ANOVA analysis of the b and n parameters show that there are more significant differences between b parameters than n parameters, meaning that most treatments showed similar tailing effect, but differed on the shape of the curve. Comparisons between peanut sauce formulations at the same pressure treatments indicate that increasing amount of organic peanut butter within the sauce formulation decreases log10 reductions. This could be due to a protective effect from the lipids in the peanut butter, or it may be due to other factors such as nutrient availability or water activity. Sauces pressurized at lower temperatures had decreased log10 reductions, indicating that cooler temperatures offered some protective effect. Log10 reductions exceeded 5 logs, indicating that high pressure processing may be a suitable option as a kill-step for Salmonella in industrial processing of peanut sauces. Future research should include high pressure processing on other peanut products with high water activities such as sauces and syrups as well as research to determine the effects of water activity and lipid composition with a food matrix such as peanut sauces.