Cooperative State Research Research Articles

IPM: Where to Next?

American Entomologist • Summer 2010 In this paper, we present a brief overview of IPM’s history, its present status, and current and future trends we see developing, along with our suggestions for actions that we believe will help IPM implementation grow and prosper as a unifying concept. Our careers as colleagues and collaborators have spanned over 30 years, beginning in 1978 in the Department of Entomology at North Dakota State University (Kopp with an extension appointment and Meyer with a research/teaching appointment). For the past 12 years, we have had program leadership roles in the USDA Cooperative State Research, Education, and Extension Service (CSREES). In Washington, working with colleagues and stakeholders, we have had the opportunity to conceptualize, prioritize, direct, manage and implement IPMrelated programs at the national level. At least in some small ways, we have shaped and focused directions for federally funded USDA IPM programs. We have served in leadership roles for the following USDA programs: National Agricultural Pesticide Impact Assessment Program (NAPIAP), Regional IPM Centers, Pest Management Alternatives Program (PMAP), Crops at Risk (CAR), Risk Avoidance and Management Program (RAMP), Methyl Bromide Transitions (MBT), Minor Crops Pest Management (IR-4) Program, National Plant Diagnostic Network (NPDN), and Critical Issues: Emerging and New Plant and Animal Pests and Diseases Program. The ideas presented here have evolved over the course of our shared state and federal careers: these perceptions, ideas and suggestions represent our own views and are not those of the USDA, CSREES or that of the federal government. CSREES has now been reorganized, consistent with requirements of the 2008 Farm Bill legislation, and became the National Institute of Dennis D. Kopp and H.J. (Rick) Meyer

Linking Information to Real-Life Problems: An Interdisciplinary Collaboration of Librarians, Departments, and Food Businesses

Through a United States Department of Agriculture Cooperative State Research, Education, and Extension Service (USDA CSREES) Higher Education Challenge Grant, a team of teaching faculty and librarians at Rutgers University developed an interdisciplinary course, “Food and Nutrition Business Information and Communication.” The course employed an active learning approach that involved a unique collaboration between various academic disciplines; the Rutgers University Libraries; the School of Communication, Information, and Library Studies (SCILS); and the Rutgers Food Manufacturing Technology Facility (FMT). In the course, teams of students from the departments involved worked collaboratively to help solve an actual research problem from industry partners working at FMT. Three examples of student problem solving are presented and assessed using the rubric developed for the course. The course actively involves librarians and demonstrates the importance of information literacy skills in today's information economy.

The New USDA: Cultivating Change

In proclaiming the week of August 23–29 National Community Gardening Week, Agriculture Secretary Tom Vilsack noted, “Community gardens provide numerous benefits including opportunities for local food production, resource conservation, and neighborhood beautification. But they also promote family and community interaction and enhance opportunities to eat healthy, nutritious foods. Each of these benefits is something we can and should strive for.” Vilsack’s statement was the latest in a string of signals in the past 7 months that suggest significant changes are afoot at the U.S. Department of Agriculture (USDA). On 29 July 2009 Agriculture Deputy Secretary Kathleen Merrigan inaugurated a new rooftop garden at the offices of the USDA Economic Research Service. In early spring, Vilsack “broke pavement” for a vegetable garden known as the People’s Garden, which was planted in front of USDA headquarters across from the National Mall. And a March 2009 planning meeting to discuss the People’s Garden and other sustainability initiatives, chaired by Vilsack, included people not typically seen at USDA meetings in the past: representatives from community garden associations, local food policy councils, botanical gardens, and the Rodale Institute, a nonprofit organization in Pennsylvania dedicated to organic farming research. Vilsack “talked about sustainability, linking agriculture, food, and human health in a way that you haven’t heard [from USDA],” recalls Rose Hayden-Smith, a fellow at the Minneapolis-based nonprofit Institute for Agriculture and Trade Policy and master gardener for the California Cooperative Extension service.

Washington update

Jesse and Doris Lyons are the recipients of the 2008 Poultry Extension Special Recognition Award for their exceptional contributions to state, regional, and national extension programs. The National Poultry Extension Workshop continues to be possible only through the dedicated efforts of volunteers. In a welcome break from tradition, the workshop was moved from Sunday to Monday morning, which reduced personal conflicts related to Saturday travel and early Sunday meetings. The Future Trends in Animal Agriculture 2008 Symposium was “Complementary Relationships in Animal Agriculture.” The 2009 Quadrennial Poultry Extension Workshop will be held in Raleigh, North Carolina. The National Poultry Waste Management Symposium coordinator for 2008 is Casey Ritz, University of Georgia. The USDA grants, deadlines, and procedures can be found on the Cooperative State Research, Education, and Extension Service Internet site (http://www.csrees.usda.gov). The Cooperative State Research, Education, and Extension Service, Plant and Animal Systems has yearly internal reviews of its various portfolios that summarize the accomplishments and impacts of system personnel. Program accountability requires documentation by everyone. Now and in the future, cooperation, networking, teamwork, and participation in multistate research committees are essential to our ability to make meaningful contributions to the poultry system. Several universities now provide animal welfare education programs, including participation in assessment competitions and distance education. The Animal Welfare Assessment Contest will be coordinated by Michigan State University for the next few years. Tyson Foods sponsors the Poultry Science Association Support Personnel Award, and Felicity Dennis of the University of Alberta was the 2008 recipient.

Antagonism of the Sphingosine 1-Phosphate (S1P) Signaling Pathway in Pregnant Ewes Alters Placentome Architecture, Amino Acid Transport, and Fetal Growth.

Angiogenesis is a critical factor in the process of placentation. In ruminants, angiogenesis occurs primarily in placentomes, formed by interdigitation and branching morphogenesis of placental cotyledons and maternal endometrial caruncles, which supply hematotrophic nutrition to the developing conceptus. We hypothesized that the lysosphingolipid sphingosine-1-phosphate (S1P) stimulates angiogenesis within developing sheep placentomes through activation of the S1P receptors (S1P1-S1P5) and that their antagonism would alter placentomal angiogenesis and subsequent uteroplacental nutrient exchange during early to mid pregnancy. In Study 1, passage 4-5 primary ovine uterine artery endothelial cells (oUAEC) from 3 late pregnant sheep were seeded as monolayers onto three dimensional (3D) collagen matrices containing angiogenic growth factors and S1P. S1P dose-dependently (1–1000 nM) enhanced endothelial cell invasion compared to VEGF and FGF2 alone. Using RT-PCR to detect S1P1-S1P5, only S1P1 and S1P3 receptors were found to be expressed in oUAEC. Antagonism of these receptors with FTY720 and VPC 23019 blocked S1P-induced endothelial cell invasion in vitro. In Study 2, we examined potential physiological mechanisms for actions of endogenous S1P in the control of ovine placental angiogensis. Ewes with a single fetus were given daily intravenous injections of FTY720 (n=6; 1.5 mg/day) or vehicle (n=5) from Days 30 to 60 prior to collection of tissues on Day 60. FTY720 treatment altered the histoarchitecture of the placentome as evidenced by decreased stromal area of caruncular crypts and diminished thickness of connective tissue forming the placentomal capsule. Concomitant with these effects on interdigitation of uterine and placental tissues, the total volume of cotyledonary tissue per placentome was greater in FTY720-treated ewes. Immunofluorescence localization of the intermediate filament proteins, cytokeratin and vimentin indicated no overt change in synctial epithelia, whereas stromal tissue thickness and density were decreased with FTY720 treatment. Immunofluorescence localization of alpha-smooth muscle actin and the endothelial marker, von Willebrand Factor, in placentomes revealed that FTY720 treatment increased diameter of the lumen, thickness of the tunica muscularis of vessels, but decreased the total number of vessels in caruncles as compared to controls. FTY720 treatment also affected transport of amino acids across the placentae and fetal development as evidenced by decreases in methionine levels in allantoic fluid (P<0.05), aspartate and glutamate levels in amniotic fluid (P<0.005), and crown-rump length of fetuses (P<0.05), however, total fetal weight was not affected (P>0.05). Results of this study indicates that antagonism of the S1P signaling pathway using FTY720 interfered with placentomal development, including angiogenesis, transport of micronutrients from the uterus to the placenta and fetal growth. We conclude that S1P is a key regulatory molecule responsible for mediating development of the angiogenic network in placentomes of sheep that provide hematatrophic support for the developing conceptus. (Research supported by National Research Initiative Competitive Grants Program, Cooperative State Research, Education, and Extension Service, USDA award 2008-35203-18830 to KAD and AHA Scientist Development Grant #053020N to KJB) (poster)

Disruption of Cytokeratin 18-Containing Intermediate Filaments in Bovine Luteal Cells: Effects of Acrylamide on Progesterone Secretion, Fas Expression, and FasL-Induced Apoptosis.

One proposed mechanism of cell death during regression of the bovine corpus luteum (CL) is Fas-Fas Ligand (FasL)-induced apoptosis. Fas is a protein receptor found on the surface of luteal cells that binds with FasL and induces cell death through a series of signaling pathways involving caspase enzymes and cytokeratin-containing intermediate filaments. Others have shown in non-ovarian cell types that cytokeratin 18 (CK18)-containing intermediate filaments impair FasL-induced apoptosis, yet these filaments are also directly cleaved by caspases during apoptotic signaling. The purpose of the present study was to determine whether disruption of CK18-containing intermediate filaments in bovine luteal cells increases the cell surface expression of Fas and enhances FasL-induced apoptosis. Early and late stage CL (Days 5 and 16-18 postovulation, respectively) were collected and prepared for primary cell culture following collagenase dissociation. The cultures were exposed initially to culture medium (control) or acrylamide (5mM) for 4 hours to disrupt CK18-containing intermediate filaments. Subsequently, the culture medium in both groups was exchanged and the cells were then exposed to vehicle (control) or a cytokine cocktail containing FasL (50ng/mL) for 24 hours to induce cell death. Immunocytochemical staining revealed that acrylamide effectively disrupted CK18-containing intermediate filaments, resulting in perinuclear aggregation. In contrast, acrylamide had no effect on microtubule organization, progesterone secretion (177 versus 197 ng/5x105cells in control and acrylamide cultures, respectively), or cell viability (19% versus 14% cell death in control and acrylamide treated cultures, respectively) regardless of the stage of CL. Flow cytometric analysis revealed that Fas expression on the surface of cultured luteal cells from early and late stage CL was relatively high (an average of 84% and 61% of luteal cells from early and late stages, respectively). Pretreatment of the cultures with acrylamide, however, did not substantially enhance Fas expression (86% and 63% of luteal cells from early and late stage CL, respectively). Similarly, acrylamide did not augment FasL-induced cell death (an average of 42% versus 40% cell death for control and acrylamide-treated cultures, respectively), regardless of stage of CL. Consistent with the relatively high expression of Fas on cells from early stage CL, the incidence of FasL-induced death was greater in cultures from early stage CL than late stage CL (52% versus 29% cell death, respectively). The results indicate that while acrylamide effectively disrupts CK18-containing intermediate filaments in bovine luteal cells, its transient actions fail to increase Fas expression on the cell surface or to promote FasL-induced apoptosis. This project was supported by National Research Initiative Competitive Grant no. 2007-35203-18074 from the USDA Cooperative State Research, Education, and Extension Service. (poster)

Genetic Over-Expression of Cytokeratin 18 in Bovine Luteal Cells: Effects on Intermediate Filament Formation and FasL-Induced Death.

The Fas-Fas ligand (FasL) system is an integral component of luteal regression, yet the intrinsic mechanism(s) sensitizing luteal cells to FasL-induced cell death are unknown. Susceptibility to FasL in some cell types is influenced by cytokeratin-containing intermediate filaments, and within these cells the filaments impart resistance to cell death. In previous work we have observed that steroidogenic cells of the bovine corpus luteum (CL) contain cytokeratin 18 (CK18) filaments and thus might be resistant to FasL-induced cell death. Here, we tested this possibility by infecting primary cultures of bovine luteal cells with a full-length cDNA construct for hCK18 in a GFP-containing adenovirus vector with cytomegalovirus promoter. The cells were obtained from bovine CL in the early (Day 5) and late stages (Days 16-18) post-ovulation and prepared for primary cell culture. Cells were cultured overnight and exposed to culture medium (Control), GFP-containing adenovirus vector (Ad.GFP transduced), or CK18-adenovirus vector, also containing GFP (Ad.CK18 transduced) at an optimized MOI for 1 hr. Subsequently, the cells were incubated an additional 16-24 hr in virus-free media, and then exposed to either vehicle or a cytokine cocktail containing FasL (50ng/mL FasL) for 24 hr to induce cell death. The cells were fixed and prepared for detection of GFP, immunocytochemical detection of CK18, and an assessment of cell death. Overall infection efficiency averaged 75% in cultures from early stage CL; whereas only 40% efficiency was achieved for cultures from late stage CL. Approximately 45% of the cells from early stage CL expressed CK18 filaments prior to transduction; whereas >75% of the cells expressed CK18 following treatment with Ad.CK18. In cells from late stage CL, endogenous expression of CK18 filaments was relatively low (~16% of all cells), but increased to ~40% of the cells following treatment with Ad.CK18. Intriguingly, virtually all transduced cells from early and late stage CL contained abundant CK18 aggregates rather than filamentous structures. These aggregates were not observed in Control or GFP-transduced cells nor did they affect the viability of the cells in CK18-transduced cultures. FasL-induced cell death was similar among Control, Ad.GFP-, and Ad.CK18-transduced cultures (14%, 18%, and 16% death, respectively) regardless of CL stage, indicating aggregation of CK18 in transduced cultures did not prevent FasL-induced death. Aggregation of CK18 also did not influence progesterone secretion by cells of early CL before or after FasL treatment (62 versus 60 ng/ml, respectively), but potentially enhanced progesterone secretion by cells of late stage CL (82 versus 181 ng/ml for control and FasL-treated cultures, respectively). Knowing that intermediate filaments consist of heterodimeric proteins (e.g., CK18 paired with CK8), the aggregation of CK18 following transduction might have resulted from a lack of other available cytokeratins. In conclusion, genetic over-expression of CK18 in bovine luteal cells results in CK18 aggregation rather than filament formation, which fails to impair FasL-induced cell death but may influence progesterone secretion. This project was supported by National Research Initiative Competitive Grant no. 2007-35203-18074 from the USDA Cooperative State Research, Education, and Extension Service. (poster)

In Vivo Temporal Expression of Protein Levels of Tissue Inhibitor of Metalloproteinase-3 (TIMP-3) During Prostaglandin F2alpha-Induced Luteolysis in Sheep.

Multiple pulses of uterine prostaglandin F2alpha (PGF2α) are required to cause the complete functional decline and structural regression of the sheep corpus luteum (CL). We have used a physiological model in sheep that mimics the effects of PGF2α pulses to study the family of proteins known to play key roles in remodeling the extracellular matrix (ECM), the matrix metalloproteinases (MMPs) and TIMPs. Following the first, second and third pulse of PGF2α our findings to date indicate that the activities of the gelatinases, MMP-2 and MMP-9, exhibit minor fluctuations. However, TIMP-1 and TIMP-2 protein expression decreases following the first and second PGF2α pulses, while they rebound to control levels following the third pulse. Because other members of this family may also have a role in luteolysis, the objective of the present study was to determine the temporal protein expression of TIMP-3 in the sheep CL following one, two and three infusions of PGF2α. Among the four known TIMPs, TIMP-3 is the only member that is bound to the ECM and it is associated with apoptosis. During the mid-luteal phase (day 10/11), sheep were given three 1 hour-long systemic infusions of PGF2α (~0.22μg/kg/min/hr) at intervals matching endogenous pulses of PGF2α during luteolysis. Luteectomies (n=4 per time point) were performed at 0 hour (pretreatment control) and at -1, 1, 8, 16, or 24 hours after each PGF2α infusion. Immunoblotting was used to determine TIMP-3 protein expression. Densitometry of immunoblots was performed before the Mixed Procedure of SAS was used to determine differences in protein expression between time points. Two forms of TIMP-3 migrating at relative molecular masses of 48 and 24 kDa were present in sheep luteal extracts. The expression of the 48 kDa form of TIMP-3, compared to the 0 hour pretreatment control, was unchanged (p>0.05) after the first infusion of PGF2α until the 16 hour time point, when expression increased (p<0.05) by 3-fold. Thereafter, following the second and third infusions of PGF2α, TIMP-3 expression overall was increased (p<0.05) by 1.5-fold above the 0 hour pretreatment control and throughout the remaining sampling period. In contrast, the expression of the 24 kDa form of TIMP-3, compared to the 0 hour pretreatment control, decreased (p<0.05) at the 1 hour time point and remained depressed (p<0.05) throughout the remaining period and following both the two and three infusions of PGF2α. In summary, the concomitant increase and decrease in the expression of the 48 and 24 kDa forms, respectively, of TIMP-3 in the sheep CL following multiple infusions of PGF2α may be related to their roles in remodeling of the ECM and in causing apoptosis during luteolysis. Future studies are warranted to determine the functional role(s) of TIMP-3 on luteal function in the sheep. (Supported by National Research Initiative Competitive Grant # 2004-35203-14176 from the USDA Cooperative State Research Education and Extension Service). (poster)

Identification of Ovarian Genes Regulated by Follicle-Stimulating Hormone During Early Secondary Oocyte Growth in Salmon.

It is well established in mammals that follicle-stimulating hormone (FSH) plays a major role in regulating oocyte growth and development by modulating the expression of follicle cell and oocyte factors. In fishes, however, little is known about the function(s) of FSH and regulation of early oocyte growth, a period of development that may affect later egg quality and fecundity. In species that exhibit synchronous gonad development (e.g., salmonids, catfish, striped bass), plasma FSH and/or pituitary gonadotropin subunit expression increases significantly during early secondary oocyte growth and continues to rise during vitellogenesis, suggesting FSH may play a role in regulating follicle development during this period. The goal of this study was to gain a fundamental understanding of how FSH acts at the level of the fish ovary by identifying FSH regulated ovarian genes. To examine time course effects of FSH, coho salmon (Oncorhynchus kisutch) previtellogenic ovarian follicles were cultured with or without 500 ng salmon FSH for 12-72 h. Culture media was collected for estradiol-17beta (E2) measurement, ovarian RNA was isolated, and abundance of candidate oocyte and follicle cell mRNAs was measured by quantitative PCR. Medium E2 levels were significantly elevated in response to FSH by 12 h and plateaued after 48 h. Messenger RNAs encoding steroidogenic enzymes, such as steroid acute regulatory protein (star) and 3beta-hydroxysteroid (hsd3b) were upregulated by 12 h, while P450 aromatase (cyp19a1a) showed no change in response to FSH. Other putative follicle cell transcripts such as FSH receptor (fshr) and lipoprotein lipase (lpl) were depressed initially by FSH treatment and later recovered to control levels. Putative oocyte transcripts such as zona pellucida glycoproteins, alveolin, and a C-type egg lectin showed no change over the 72-h experiment. Additional FSH regulated ovarian genes were subsequently identified through construction of four suppression subtractive hybridization (SSH) libraries with RNA isolated from FSH-treated and untreated whole ovarian follicles, as well as follicle/interstitial cell enriched ovarian tissues. Many of the genes identified by SSH are involved in signal transduction, cell cycle regulation, cell growth and proliferation, apoptosis, cholesterol metabolism and steroidogenesis. Experiments utilizing steroidogenesis inhibitors are underway to distinguish ovarian genes directly regulated by FSH versus those regulated by sex steroids produced in response to FSH. As evident by the rapid and significant increase in E2 release and changes in steroidogenic enzyme gene expression within the ovary, we can conclude that FSH strongly influences steroidogenesis of the salmon ovary during early secondary oocyte growth and may be a critical signal for puberty onset. This is the first comprehensive assessment of the biological actions of FSH in the fish ovary. Discoveries will aid in developing methods to reduce common reproductive dysfunctions in aquaculture, such as delayed puberty and poor egg quality. This project was supported in part by National Research Initiative Competitive Grant no. 2007-35203-18082 from the USDA Cooperative State Research, Education, and Extension Service to JAL. (platform)

Downregulation of Reproductive Homeobox Gene 6 (Rhox6) Interferes with Male Germ Cell Differentiation.

Homeodomain proteins contain a 60 amino acid DNA binding motif called homeobox that regulates a variety of developmental events such as determination of body segment identity, limb development and organogenesis. Recently, a novel cluster of homeobox genes has been discovered on the X chromosome of the mouse. Because of their selective expression in reproductive organs, they are named Reproductive homeobox genes on the X chromosome (Rhox). Rhox6, a member of the Rhox6 family, is expressed in primordial germ cells (PGCs) in the developing gonad and the placenta, although the biological function of Rhox6 in determination of PGCs and placental cells remains unknown. To investigate the Rhox6 function, we generated male chimeric mice by injection of the gene-trapped embryonic stem cells (ESCs) harboring a hypomorphic mutation in Rhox6 into wild-type blastocysts. Almost all of the male chimeric mice exhibited infertility associated with smaller testes, reduced sperm number and motility and abnormal morphology of the epididymal sperm. Next, to obtain a better understanding of the Rhox6 function in germ cell differentiation, we took advantage of in vitro ESC culture. ESCs can differentiate into PGCs when ESCs are cultured as aggregates called embryoid bodies (EBs) in the medium containing Ascorbic acid, 1-Thioglycerol and Transferrin without the leukemia inhibitory factor. By using a mouse ESC line that expresses the enhanced green fluorescent protein (EGFP) under the promoter of Oct3/4, a central regulator of cellular pluripotency, the presence of PGCs within EBs was monitored in real time by EGFP expression. While this ESC line was induced to differentiate into PGCs, expression of Rhox6 was downregulated by stable expression of the short-hairpin (sh) RNA against the Rhox6 transcripts. The level of Rhox6 knockdown was validated by RT-PCR. As a negative control, a scrambled nucleotide sequence that does not show any match with the mouse genome was used to express shRNA molecules. EBs were generated from multiple independent clones that express either one of the shRNAs, which were isolated by electroporation of the ESC line with each vector, followed by drug selection. We found that 0% of the EBs having the shRNA against the Rhox6 transcripts expressed (n=29; the number of clones used to generate EBs is 17) had EGFP-positive cells, whereas 43% of the EBs having the control shRNA expressed (n=35; the number of clones used to generate EBs is 25) had EGFP-positive cells. To validate differentiation of PGCs, these EBs were pooled according to EGFP expression and used to examine expression of PGC markers by semi-quantitative RT-PCR. Because expression of Rex1, a marker for undifferentiated ESCs, was significantly downregulated in these EBs than in undifferentiated ESCs, EGFP expression detected in the EBs indicates the presence of PGCs, but not the presence of undifferentiated ESCs. Rhox6 as well as other PGC markers including Oct3/4, Rnh2, Piwil2, and Fgls were downregulated in the EBs expressing the shRNA against the Rhox6 transcripts, whereas the EBs expressing the control shRNA with EGFP-positive cells expressed Rhox6 and these PGC markers. Collectively, these in vivo and in vitro studies strongly suggest that downregulation of Rhox6 impairs differentiation of PGCs. This study is supported by the Departmental Startup fund including the USDA Cooperative State Research, Education and Extension Service, Hatch project (ILLU-538-323). (poster)

A Health Care Cooperative Extension Service

RIMARY CARE IS THE ESSENTIAL FOUNDATION FOR AN effective, efficient, and equitable health care system.Callstorebuildthecrumblingprimarycareinfrastructure in the United States are reaching receptive ears, with public and private advisory groups includingtheMedicarePaymentAdvisoryCommissionand the National Business Group on Health recommending increased payments for primary care. 1 The American RecoveryandReinvestmentAct(ARRA) 2 of2009appropriated$19 billion for the purchase of health information technology (HIT), with primary care physicians’ offices slated to be among the beneficiaries. Policy makers expect that new investments will transform primary care by creating more effective and efficient patient-centered medical homes. The primary care physician community acknowledges the need for new practice models that provide accessible, comprehensive, integrated care based on healing relationships over time. 3 New investment in primary care is necessary but not sufficienttorevitalizeprimarycareunlesscombinedwithastrategy for disseminating and implementing innovations and best practices. Acquiring an electronic health record (EHR) will not create a highly functioning medical home unless it can be used to create functional patient registries. Receivingenhancedpaymentsforcarecoordinationwithoutaworkable plan for hiring and training health coaches for patient self-management leaves a gap between expectations and reality. Large, organized delivery systems such as Geisenger, Kaiser Permanente, and the Veterans Administration have theinstitutionalwherewithalandeconomiesofscaletoimplement practice redesign in a systematic and successful manner. However, two-thirds of office-based physicians work in practices of 4 or fewer physicians. 4 These clinicians oftenhavelittleornotechnicalassistancetodeployandmaintain new practice improvements like EHRs. To successfully redesign practices requires knowledge transfer, performance feedback, facilitation, and HIT support provided by individuals with whom practices have established relationships over time. The farming community learned these principles a century ago. Primary care practices are like small farms of that era, which were geographically dispersed, poorly resourced for change, and inefficient in adopting new techniques or technology but vital tothenation’swell-being.Practicingphysiciansneedsomething akin to the agricultural extension agent who was so transformativeforfarming. 5,6 AnationwidePrimaryCareCooperative Extension Service, modeled after the US Department of Agriculture’s Cooperative State Research, Education,andExtensionService(CooperativeExtension),which so successfully accelerated farm transformation, should be created.County-basedhealthextensionorganizationswould supportprimarycarecliniciansinthesamemannerthatthe agricultural model assists family farmers, providing infrastructureforlocallearningcommunitiesandpracticetransformation. ARRA establishes a Health Information Technology Extension Program 2 to “assist health care providers toadopt,implement,andeffectivelyusecertifiedEHRtechnology,” which could serve as the nidus for a broader program to revitalize primary care and community health.

Cooking schools improve food intake patterns and diabetes knowledge of people with diabetes

Implementation of nutrition recommendations is often difficult for people with diabetes. The objective was to determine if cooking schools delivered by the Cooperative Extension Service in collaboration with registered dietitians and certified diabetes educators improved dietary patterns and knowledge of people with diabetes. The research design was quasi‐experimental with a pretest, posttest design. Each subject served as his or her own control. Kitchen Creations cooking schools intervention consisted of four, three‐hour long lessons. The cooking schools featured current nutrition recommendations for people with diabetes and hands‐on food preparation and cooking experiences. Participants completed three‐day food records and demographics and knowledge questionnaires prior to attending Kitchen Creations and one month after completing the Kitchen Creations series. Following participation in Kitchen Creations, participants showed significant (p&lt;0.05) decreases in energy, fat energy, fat grams, percent of calories from fat, fat exchanges, saturated fat energy, saturated fat grams, carbohydrate grams, cholesterol (mg) and sodium (mg). Bread exchanges and "other carbohydrate" exchanges decreased (p&lt;0.05.) Vegetable exhanges increased in women (p&lt;0.05) but not in men. Knowledge of nutrition recommendations for people with diabetes significantly (p&lt;0.05) improved. Results indicate that a cooking school approach can successfully change food intake patterns. Funding provided by the USDA Cooperative State Research, Education &amp; Extension Service, New Mexico State University and the New Mexico Department of Health, Diabetes Prevention and Control Program.

Using genomics to contemplate microbial physiology

A major objective of a microbial physiology curriculum is to cement a comprehensive understanding of central carbon metabolism (glycolysis/gluconeogenesis, the pentose phosphate pathway, and the citric acid cycle). One way to meet this goal is to have students consider the catabolic and anabolic roles of these pathways in organisms with diverse physiologies (e.g., respiratory heterotrophs; nonrespiratory, fermentative heterotrophs; autotrophs). In MCB4404 Microbial Physiology Lab, we query microbial genome sequences to elucidate the presence and absence of genes encoding enzymes catalyzing steps of central carbon metabolism in a variety of microorganisms. This genome‐level approach exposes students to bioinformatics tools and allows them to test hypotheses for gene presence/absence based on their understanding of microbial physiology. This project is generously supported by the USDA Cooperative State Research and Extension Service: US Department of Agriculture Higher Education Challenge Grants Program.

The growth performance, carcass traits, meat pH and color, and CO2 production in pigs supplementated arginine with conjugated linoleic acid

This experiment tested the hypothesis that the combination of dietary conjugated linoleic acid (CLA) plus arginine will increase the growth performance, carcass traits, meat quality, and CO2 production. Eight pigs (80kg) were assigned to treatments in a 2 x 2 factorial design with two pigs per treatment (control: 2.05% alanine + 1% canola oil; CLA: 2.05% alanine + 1% CLA (mixed isomers); arginine: 1% arginine + 1% canola oil; arginine + CLA: 1% arginine + 1% CLA). Preliminary tests showed up to 2% arginine plus 1% CLA supplementation can be acceptable without interfering lysine absorption. There was no significant difference in growth performance including average daily feed, average daily gain over the five week treatment period. Dietary arginine and CLA did not affect either carcass or meat quality traits including loin area, back fat thickness, pH45min, pH24hr, meat color (L*, a*, and b*), and drip loss. Rates of glucose and palmitate conversion to CO2 in vitro in liver, longissimus dorsi muscle, abdominal and subcutaneous fat, and enterocytes were not affected by CLA. However, arginine increased CO2 production in all tissue except muscle. Also, CO2 production in both adipose tissue deposits were increased by CLA and arginine (P &lt; 0.05), (This project was supported by National Research Initiative Competitive Grant no. 2008‐35206‐18762 from the USDA Cooperative State Research, Education, and Extension Service).

100 ASSESSMENT OF LOCUS-SPECIFIC DNA METHYLATION: OPTIMIZATION FOR BOVINE EMBRYOS

Assessment of chromatin remodeling in early embryos is a major focus of studies today, and evaluation of DNA methylation at specific loci is one approach to study these epigenetic modifications. Our objective was to optimize the bisulfite sequencing methodology for use with very small cell numbers originating from pre-implantation embryos, making the process more time- and cost-efficient. The optimized steps include bisulfite conversion of small samples, bisulfite primer design, high-throughput plasmid DNA amplification, and preparation for sequencing. Methylation at 2 loci, Satellite I and Oct4, was investigated in bovine in vitro-produced (IVP) embryos collected at the 2-cell, 8-cell, and blastocyst stages. Bovine skin fibroblasts were first used to optimize the particular steps of the process. All reactions were run in duplicate and no-template negative and somatic cell-positive controls were treated alongside samples. Incorporating the use of Methyl Primer Express (Applied Biosystems, Foster City, CA), MacVector (Oxford Molecular Ltd., Campbell, CA), and Mfold software (Mathews DH et al. 1999 J. Mol. Biol. 288, 911–940; Zuker M 2003 Nucleic Acids Res. 31, 3406–3415) improved the specificity of bisulfite primers by exclusion of secondary or tertiary structures. The DNA from bisulfite treatment for 15 to 16 h was of better quality than DNA treated for 18 h. After initial PCR optimization, different cell concentrations were used to establish that detectable PCR products and subsequent methylation data could be obtained from DNA isolated from as few as 8 cells. Treating single blastocysts and pools of ten 8-cell and forty 2-cell embryos was sufficient for the entire scope of the experiment, allowing use of the same samples across all loci. After molecular cloning, plasmid DNA was amplified by 3 different methods and evaluated for efficiency: miniprep, TempliPhi (GE Healthcare, Piscataway, NJ), or 96-well glycerol stocks and automated TempliPhi format. Although TempliPhi alone was better than miniprep for small-scale experiments, it was the 96-well format that saved weeks of time and was most cost-effective. Sequencing was performed on a minimum of 8 clones/sample using ABI Prism sequencers (Applied Biosystems), and results were analyzed using Chromas Pro software (Technelysium Pty. Ltd., Helensvale, Australia). Percentage methylation of bovine IVP 2-cell, 8-cell, and blastocyst stage embryos for Satellite I was 25, 10, and 22%, respectively, and for Oct4 was 88, 88, and 79%, respectively. However, somatic cell methylation was 74% for Satellite I and 88% for Oct4, implying that Satellite I is demethylated during early embryo development, whereas Oct4 remains hypermethylated. In conclusion, these improved methods will benefit further studies of chromatin remodeling in early bovine pre-implantation embryos. This project was supported by National Research Initiative Competitive Grant no. 2006-35203-16620 from the USDA Cooperative State Research, Education, and Extension Service.

193 DIFFERENTIAL GENE EXPRESSION IN CUMULUS CELLS OF DEVELOPMENTALLY COMPETENT V. CHALLENGED BOVINE OOCYTES

It is well established that exposure of cumulus–oocyte complexes (COC) to heat stress during the first 12 h of maturation reduces blastocyst development by 42 to 65%. Previous research supports the notion that some of the effects of heat stress on oocyte competence may be cumulus-mediated. To determine the extent to which this may occur, COC were matured at 38.5°C for 24 h (control) or 41°C for the first 12 h of maturation followed by 38.5°C for remaining 12 h (heat stress). A subset of COC underwent IVF with Percoll-prepared sperm and then was cultured in KSOM containing 0.5% BSA to assess developmental competence. Remaining oocytes were denuded. Cumulus cells, kept separate by treatment, were stored in lysis buffer at –80°C until RNA extraction. Total RNA from cumulus was amplified prior to hybridization to bovine Affymetrix GeneChips (Affymetrix Inc., Santa Clara, CA, USA; n = 8 pools per treatment collected on 8 different occasions; n = 16 chips). Following pre-processing using the MAS5.0 algorithm, microarray data were subjected to linear modeling and empirical Bayes analyses (Bioconductor, Limma package). False discovery rate was controlled using the Benjamini and Hochberg method, and differentially expressed genes were selected by an adjusted P-value (P &lt; 0.05). Functional annotation of selected genes was performed using NetAffx (Affymetrix Inc.) and Database for Annotation, Visualization and Integrated Discovery (DAVID; NIAID, NIH, Bethesda, MD, USA). Heat stress of COC reduced blastocyst development (27.2 v. 16.1% for control v. heat stress, respectively; SEM = 1.6; P &lt; 0.002). Approximately 66 and 65% of 24 000 possible genes were called present (i.e. expressed) in RNA from cumulus of competent (control) v. challenged (heat-stressed) oocytes, respectively. In cumulus from developmentally challenged COC, increased abundance of 42 genes (36 currently annotated) was noted. Use of DAVID demonstrated enrichment of genes important for electron transport and energy generation (NOS2A, MAOB, CYP11A1, HSD11B1L, LTB4DH). Further examination of gene ontology identified genes associated with mitochondrial function (SLC25A10, MAOB, CYP11A1), cell signaling (similar to JAK-3, FSHR, CYP11A1, WNT2B), cytoskeleton (ACTA1), antioxidant activity (GSTA1), and extracellular region (FMOD). In contrast, cumulus from developmentally competent COC had increased expression of 22 genes (20 currently annotated), of which 15% were related to protein binding (CAV1, MMP9, TGFB2) according to DAVID. Further analysis using gene ontology revealed genes associated with extracellular matrix formation (MMP9, MMP19, PCOLCE2) and neural tissue (METRNL). In summary, alterations in cumulus gene expression were associated with differences in developmental competence of oocytes. Additional research is necessary to examine the extent to which identified genes account for functional differences in oocyte competence. This research was supported in part by National Research Initiative Competitive Grant no. 2004-35203-14772 from the USDA Cooperative State Research, Education, and Extension Service.

The Impact of Networks and the Context of Reception on Asset Accumulation Strategies of Latino Newcomers in New Settlement Communities of the Midwest

American Journal of Agricultural EconomicsVolume 90, Issue 5 p. 1319-1325 Principal Paper Session The Impact of Networks and the Context of Reception on Asset Accumulation Strategies of Latino Newcomers in New Settlement Communities of the Midwest Corinne Valdivia, Corinne Valdivia Associate professor Department of Agricultural Economics, University of Missouri (MU)Search for more papers by this authorPedro Dozi, Pedro Dozi Graduate research assistant Department of Agricultural Economics, MUSearch for more papers by this authorStephen Jeanetta, Stephen Jeanetta Assistant extension professor Department of Rural Sociology, MUSearch for more papers by this authorLisa Y. Flores, Lisa Y. Flores Associate Professor of Counseling Psychology College of Education, MUSearch for more papers by this authorDomingo Martínez, Domingo Martínez Director of Cambio Center for Research and Outreach on Latinos and Changing Communities MUSearch for more papers by this authorAnne Dannerbeck, Anne Dannerbeck Research manager of Missouri State Courts Administration MUSearch for more papers by this author Corinne Valdivia, Corinne Valdivia Associate professor Department of Agricultural Economics, University of Missouri (MU)Search for more papers by this authorPedro Dozi, Pedro Dozi Graduate research assistant Department of Agricultural Economics, MUSearch for more papers by this authorStephen Jeanetta, Stephen Jeanetta Assistant extension professor Department of Rural Sociology, MUSearch for more papers by this authorLisa Y. Flores, Lisa Y. Flores Associate Professor of Counseling Psychology College of Education, MUSearch for more papers by this authorDomingo Martínez, Domingo Martínez Director of Cambio Center for Research and Outreach on Latinos and Changing Communities MUSearch for more papers by this authorAnne Dannerbeck, Anne Dannerbeck Research manager of Missouri State Courts Administration MUSearch for more papers by this author First published: 01 December 2008 https://doi.org/10.1111/j.1467-8276.2008.01224.xCitations: 9 The authors are grateful for the support of many community organizations in the development of their research, and the Office of Socio Economic Data Analysis OSEDA, MU. This project was supported by National Research Initiative Grant No. 2006-35401-17429 from the USDA Cooperative State Research, Education, and Extension Service Rural Development Program. This article was presented in a principal paper session at the 2008 AAEA annual meeting Orlando, FL. The articles in these sessions are not subjected to the journal's standard refereeing process. All are fellows at Cambio Center. Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat Citing Literature Volume90, Issue5December 2008Pages 1319-1325 RelatedInformation

The Growing Role of Local Food Markets

American Journal of Agricultural EconomicsVolume 90, Issue 5 p. 1289-1295 Principal Paper Session The Growing Role of Local Food Markets Shermain D. Hardesty, Shermain D. Hardesty Extension specialist Department of Agricultural and Resource Economics, University of California, DavisSearch for more papers by this author Shermain D. Hardesty, Shermain D. Hardesty Extension specialist Department of Agricultural and Resource Economics, University of California, DavisSearch for more papers by this author First published: 01 December 2008 https://doi.org/10.1111/j.1467-8276.2008.01219.xCitations: 32 This research was supported by the National Research Initiative of the USDA Cooperative State Research, Education and Extension Service, grant number 2006-55618-17015. This article was presented in a principal paper session at the 2008 AAEA annual meeting in Orlando, FL. The articles in these sessions are not subjected to the journal's standard refereeing process. Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat Citing Literature Volume90, Issue5December 2008Pages 1289-1295 RelatedInformation

The first five years of the Conservation Effects Assessment Project

T he Conservation Effects Assessment Project (CEAP) was initiated by the USDA Natural Resources Conservation Service (NRCS), Agricultural Research Service (ARS), and Cooperative State Research, Education, and Extension Service (CSREES) in response to a general call for better accountability of how society would benefit from the 2002 farm bill's substantial increase in conservation program funding (Mausbach and Dedrick 2004). The original goals of CEAP were to establish the scientific understanding of the effects of conservation practices at the watershed scale and to estimate conservation impacts and benefits for reporting at the national and regional levels. Other federal agencies and nongovernmental organizations with conservation and natural resource interests are currently partners in various CEAP activities, often through jointly funded research projects. CEAP activities are organized into three interconnected efforts: 1. Bibliographies, literature reviews, and a scientific workshop to establish what is known about the environmental effects of conservation practices at field and watershed scales, and what kinds of research and data collection are needed to assess conservation practice benefits. 2. Watershed assessment studies to provide in-depth quantification of water quality and soil quality impacts of conservation practices at the local level and to provide insight on what practices are needed…

US Governmental Interagency Programs, Opportunities, and Collaboration

Improving the quality, value, and safety of national food supplies is the common mission of several Federal agencies. Under different authorities, these agencies conduct basic and applied research, develop curriculum at higher educational institutions, and disseminate new scientific knowledge about food processing, formulation, and preservation to a broad range of stakeholders. Emerging food processing technologies, including various thermal and nonthermal processes, as well as chemical processes, have received increased attention and investment in recent years. Some agencies dealing with Emerging Technologies include the Cooperative State Research, Education, and Extension Service (CSREES), Agricultural Research Service (ARS), and Food Safety Inspection Service (FSIS), and all of the United States Department of Agriculture (USDA). The U.S. Army Natick Research Center of Department of Defense (DOD), National Aeronautics and Space Administration (NASA), and Food and Drug Administration (FDA) of Department of Human Health Service (DHHS) also are involved in this research area. These agencies have their vision, mission, strategic goals, and current programs and activities related to emerging food processing technologies. The synergy of effective collaboration and cooperation among all stakeholders is the key to generating an impact greater than the simple sum of all.