A newly developed Escherichia coli isolate panel from a cross section of U.S. animal production systems reveals geographic and commodity-based differences in antibiotic resistance gene carriage.

There are limited numbers of Escherichia coli isolate panels that represent United States food animal production. The majority of existing Escherichia coli isolate panels are typically designed: (i) to optimize genetic and/or phenotypic diversity; or (ii) focus on human isolates. To address this shortfall in agriculturally-related resources, we have assembled a publicly-available isolate panel (AgEc) from the four major animal production commodities in the United States, including beef, dairy, poultry, and swine, as well as isolates from agriculturally-impacted environments, and other commodity groups. Diversity analyses by phylotyping and Pulsed-field Gel Electrophoresis revealed a highly diverse composition, with the 300 isolates clustered into 71 PFGE sub-types based upon an 80% similarity cutoff. To demonstrate the panel's utility, tetracycline and sulfonamide resistance genes were assayed, which identified 131 isolates harboring genes involved in tetracycline resistance, and 41 isolates containing sulfonamide resistance genes. There was strong overlap in the two pools of isolates, 38 of the 41 isolates harboring sulfonamide resistance genes also contained tetracycline resistance genes. Analysis of antimicrobial resistance gene patterns revealed significant differences along commodity and geographical lines. This panel therefore provides the research community an E. coli isolate panel for study of issues pertinent to U.S. food animal production.

Evaluation of soybean genotypes for reaction to natural field infection by Cercospora species causing purple seed stain.

Purple seed stain (PSS) of soybean (Glycine max (L.) Merr.) is a prevalent seed disease. It results in poor seed quality and reduced seed lot market grade, and thus undermines value of soybean worldwide. The objectives of this research were to evaluate the reaction of selected soybean genotypes collected from 15 countries representing maturity groups (MGs) III, IV, and V to PSS, and to identify new sources of resistance to PSS based on three years of evaluation of natural field infection by Cercospora spp. in the Mississippi Delta of the U. S. In this study, 42 soybean genotypes were evaluated in 2010, 2011, and 2012. Seventeen lines including six MG III (PI 88490, PI 504488, PI 417361, PI 548298, PI 437482, and PI 578486), seven MG IV (PI 404173, PI 346308, PI 355070, PI 416779, PI 80479, PI 346307, and PI 264555), and four MG V (PI 417567, PI 417420, PI 381659, and PI 407749) genotypes had significantly lower percent seed infection by Cercospora spp. than the susceptible checks and other genotypes evaluated (P ≤ 0.05). These genotypes of soybean can be used in developing soybean cultivars or germplasm lines with resistance to PSS and for genetic mapping of PSS resistance genes. In addition, among these 17 lines with different levels of resistance to PSS, nine soybean genotypes (PI 417361, PI 504488, PI 88490, PI 346308, PI 416779, PI 417567, PI 381659, PI 417567, and PI 407749) were previously reported as resistant to Phomopsis seed decay. Therefore, they could be useful in breeding programs to develop soybean cultivars with improved resistance to both seed diseases.

Validation of Yield Component Traits Identified by Genome-Wide Association Mapping in a tropical japonica × tropical japonica Rice Biparental Mapping Population.

The Rice Diversity Panel 1 (RDP1) was developed for genome-wide association (GWA) studies to explore five rice ( L.) subpopulations (, , , , and ). The RDP1 was evaluated for over 30 traits, including agronomic, panicle architecture, seed, and disease traits and genotyped with 700,000 single nucleotide polymorphisms (SNPs). Most rice grown in the southern United States is and thus the diversity in this subpopulation is interesting to U.S. breeders. Among the RDP1 accessions, 'Estrela' and 'NSFTV199' are both phenotypically and genotypically diverse, thus making them excellent parents for a biparental mapping population. The objectives were to (i) ascertain the GWA QTLs from the RDP1 GWA studies that overlapped with the QTLs uncovered in an Estrela × NSFTV199 recombinant inbred line (RIL) population evaluated for 15 yield traits, and (ii) identify known or novel genes potentially controlling specific yield component traits. The 256 RILs were genotyped with 132 simple sequence repeat markers and 70 QTLs were found. Perl scripts were developed for automatic identification of the underlying candidate genes in the GWA QTL regions. Approximately 100 GWA QTLs overlapped with 41 Estrela × NSFTV199 QTL (RIL QTL) regions and 47 known genes were identified. Two seed trait RIL QTLs with overlapping GWA QTLs were not associated with a known gene. Segregating SNPs in the overlapping GWA QTLs for RIL QTLs with high values will be evaluated as potential DNA markers useful to breeding programs for the associated yield trait.

Control of Pecan Weevil With Microbial Biopesticides.

The pecan weevil, Curculio caryae (Horn) (Coleoptera: Curculionidae), is a key pest of pecans Carya illinoinensis ([Wangenh.] K. Koch) (Fagales: Juglandaceae). Control recommendations rely on broad spectrum chemical insecticides. Due to regulatory and environmental concerns, effective alternatives for C. caryae control must be sought for pecan production in conventional and organic systems. We explored the use of microbial biopesticides for control of C. caryae in Georgia pecan orchards. Three experiments were conducted. The first investigated an integrated microbial control approach in an organic system at two locations. Three microbial agents, Grandevo (based on byproducts of the bacterium Chromobacterium subtsugae Martin, Gundersen-Rindal, Blackburn & Buyer), the entomopathogenic nematode Steinernema carpocapsae (Weiser), and entomopathogenic fungus Beauveria bassiana (Balsamo) Vuillemin, were applied to each treatment plot (0.6 ha) at different times during the season. A second experiment compared the effects of S. carpocapsae and B. bassiana applied as single treatments relative to application of both agents (at different times); survival of C. caryae was assessed approximately 11 mo after larvae were added to pots sunk in an organic pecan orchard. In a conventional orchard (with 1.0 ha plots), the third experiment compared Grandevo applications to a commonly used regime of chemical insecticides (carbaryl alternated with a pyrethroid). All experiments were repeated in consecutive years. The combined pest management tactic (experiment 1) reduced C. caryae infestation relative to non-treated control plots in both locations in 2014 and one of the two locations in 2015 (the other location had less than 1% infestation). In experiment 2, no differences among combined microbial treatments, single-applied microbial treatments or different numbers of application were observed, yet all microbial treatments reduced C. caryae survival relative to the control. In the third experiment, both Grandevo and standard chemical insecticide applications resulted in lower weevil infestation than the control (both years) and there was no difference between the insecticide treatments in 2014 although the chemical insecticide regime had slightly lower infestation in 2015. These results provide evidence that microbial biopesticides can substantially reduce pecan weevil infestations in organic and nonorganic systems.

Assessment of Five Chilling Tolerance Traits and GWAS Mapping in Rice Using the USDA Mini-Core Collection.

Rice (Oryza sativa L.) is often exposed to cool temperatures during spring planting in temperate climates. A better understanding of genetic pathways regulating chilling tolerance will enable breeders to develop varieties with improved tolerance during germination and young seedling stages. To dissect chilling tolerance, five assays were developed; one assay for the germination stage, one assay for the germination and seedling stage, and three for the seedling stage. Based on these assays, five chilling tolerance indices were calculated and assessed using 202 O. sativa accessions from the Rice Mini-Core (RMC) collection. Significant differences between RMC accessions made the five indices suitable for genome-wide association study (GWAS) based quantitative trait loci (QTL) mapping. For young seedling stage indices, japonica and indica subspecies clustered into chilling tolerant and chilling sensitive accessions, respectively, while both subspecies had similar low temperature germinability distributions. Indica subspecies were shown to have chilling acclimation potential. GWAS mapping uncovered 48 QTL at 39 chromosome regions distributed across all 12 rice chromosomes. Interestingly, there was no overlap between the germination and seedling stage QTL. Also, 18 QTL and 32 QTL were in regions discovered in previously reported bi-parental and GWAS based QTL mapping studies, respectively. Two novel low temperature seedling survivability (LTSS)-QTL, qLTSS3-4 and qLTSS4-1, were not in a previously reported QTL region. QTL with strong effect alleles identified in this study will be useful for marker assisted breeding efforts to improve chilling tolerance in rice cultivars and enhance gene discovery for chilling tolerance.

Evaluation of Commercial Soybean Cultivars for Reaction to Phomopsis Seed Decay.

Phomopsis seed decay (PSD), caused by Phomopsis longicolla (syn. Diaporthe longicolla), is an economically important soybean disease causing poor seed quality. Planting resistant cultivars is one of the most effective means to control PSD. In this study, 16 commercially available maturity groups IV and V soybean cultivars, including two previously identified PSD-resistant and two PSD-susceptible checks, were evaluated for seed infection by P. longicolla in inoculated and noninoculated plots, and harvested promptly or with a 2-week delay in harvest. The test was conducted at Stoneville, Mississippi, in 2012 and 2013. Seed infection by P. longicolla ranged from 0.5 to 76%, and seed germination ranged from 18 to 97%. One MG IV cultivar (Morsoy R2 491) and five MG V cultivars (Progeny 5650, Progeny 5706, Asgrow 5606, Asgrow 5831, and Dyna-Gro33C59) had significantly (P ≤ 0.05) lower percent seed infected by P. longicolla than their respective susceptible checks and other cultivars in the same tests. Information obtained from this study will be useful for soybean growers and breeders for selection of cultivars for planting or breeding and future genetic studies in the development of cultivars with improved resistance to PSD.

Estimating Broad Sense Heritability and Investigating the Mechanism of Genetic Transmission of Cold Tolerance Using Mannitol as a Measure of Post-freeze Juice Degradation in Sugarcane and Energycane (Saccharum spp.).

In approximately 25% of the sugarcane-producing countries worldwide, conventional sugarcane (Saccharum spp. hybrids) is exposed to damaging freezes. A study was conducted during the 2009 and 2010 harvest seasons to compare late-season freeze tolerance among three groups: commercial Louisiana sugarcane genotypes, early generation genotypes selected for cold tolerance in the U.S. Department of Agriculture sugarcane breeding programs at Houma, LA, and Canal Point, FL, and potential energycane genotypes selected for high total biomass per acre. Mannitol concentrations in cane juice following freezing temperatures were determined to evaluate levels of cold tolerance. Genotypes selected for cold tolerance in Houma, LA, had significantly more late-season freeze tolerance than commercial sugarcane genotypes and genotypes selected in Canal Point, FL. Genotypes showing the most cold tolerance were Ho02-146 and Ho02-152, and those that were most highly susceptible were US87-1006 and US87-1003 (early-generation breeding genotypes) and L99-233 (commercial genotype). Broad-sense heritability for late-season cold tolerance in the two-year study was estimated at g(2) = 0.78. The enzymatic mannitol analysis successfully differentiated high-fiber energycane genotypes from those from other sources.

Scab Susceptibility of a Provenance Collection of Pecan in Three Different Seasons in the Southeastern United States.

Pecan scab (caused by Fusicladium effusum) is the most economically destructive disease of pecan in the Southeast United States. Wet, humid conditions typical of the Southeast are known to provide conditions conducive to epidemics. A provenance collection of pecan from 19 locations representing the native range of the tree is located in Byron, Georgia, and was assessed for pecan scab severity in 1998, 2013, and 2014. There were significant differences among the 19 provenances (F = 5.6 to 62.5, P < 0.0001). Provenances from wetter locations (generally north of Texas) had the greatest proportion of scab resistant trees, while provenances from the drier southern areas (Texas and Mexico) tended to be the most susceptible to scab. The association with rainfall was borne out by correlation analysis (r = -0.625 to -0.823 [P < 0.0001 to 0.004]). Other factors consistently associated with scab severity included leaflet tilt and droop angle (r = -0.533 to -0.883 [P < 0.0001 to 0.02]). Multiple regression analysis demonstrated that leaflet droop angle was a particularly good predictor of provenance susceptibility. Leaflet characteristics vary with provenance location, and whether there is a direct relationship between scab severity and leaflet characteristics is not established. Estimates of heritability were not entirely consistent among years, but different methods were used to assess scab severity in 1998 (a 1 to 5 category scale) compared with 2013 and 2014 (the percent ratio scale). Despite using different methods, there was generally good agreement among years in regard to severity of disease on individual trees. In conclusion, trees from more northern populations (in areas with greater annual rainfall) are most likely to provide valuable and diverse sources of resistance to scab. The provenance collection contains a range of scab-resistant genotypes from diverse locations that can contribute to genetic improvement regarding scab resistance.

Genome-Wide Association Study of Ureide Concentration in Diverse Maturity Group IV Soybean [Glycine max (L.) Merr.] Accessions.

Ureides are the N-rich products of N-fixation that are transported from soybean nodules to the shoot. Ureides are known to accumulate in leaves in response to water-deficit stress, and this has been used to identify genotypes with reduced N-fixation sensitivity to drought. Our objectives in this research were to determine shoot ureide concentrations in 374 Maturity Group IV soybean accessions and to identify genomic regions associated with shoot ureide concentration. The accessions were grown at two locations (Columbia, MO, and Stuttgart, AR) in 2 yr (2009 and 2010) and characterized for ureide concentration at beginning flowering to full bloom. Average shoot ureide concentrations across all four environments (two locations and two years) and 374 accessions ranged from 12.4 to 33.1 µmol g(-1) and were comparable to previously reported values. SNP-ureide associations within and across the four environments were assessed using 33,957 SNPs with a MAF ≥0.03. In total, 53 putative loci on 18 chromosomes were identified as associated with ureide concentration. Two of the putative loci were located near previously reported QTL associated with ureide concentration and 30 loci were located near genes associated with ureide metabolism. The remaining putative loci were not near chromosomal regions previously associated with shoot ureide concentration and may mark new genes involved in ureide metabolism. Ultimately, confirmation of these putative loci will provide new sources of variation for use in soybean breeding programs.

Evaluation of Diverse Soybean Germplasm for Resistance to Phomopsis Seed Decay.

Phomopsis seed decay (PSD), caused primarily by the fungal pathogen Phomopsis longicolla, is one of the most important diseases reducing seed quality and yield of soybean. Few cultivars have been identified as resistant. To identify new sources of resistance to PSD, 135 soybean germplasm accessions, originating from 28 countries, were field screened in Arkansas, Mississippi, and Missouri in 2009. Based on seed assays of natural field infection by P. longicolla in 2009, 42 lines, including the most resistant and susceptible lines, were reevaluated in the field in 2010, 2011, and 2012 with P. longicolla-inoculated and noninoculated treatments. Six maturity group (MG) III (PI 189891, PI 398697, PI 417361, PI 504481, PI 504488, and PI 88490), four MG IV (PI 158765, PI 235335, PI 346308, and PI 416779), and five MG V (PI 381659, PI 381668, PI 407749, PI 417567, and PI 476920) lines had significantly lower percent seed infection by P. longicolla than the susceptible checks and other lines in the same test (P ≤ 0.05). They appeared to have some levels of resistance to PSD. These new sources of PSD resistance can be used in developing soybean breeding lines or cultivars with resistance to PSD, and for genetic mapping of PSD resistance genes.

Influence of post-treatment strategies on the properties of activated chars from broiler manure.

There are a myriad of carbonaceous precursors that can be used advantageously to produce activated carbons or chars, due to their low cost, availability and intrinsic properties. Because of the nature of the raw material, production of granular activated chars from broiler manure results in a significant ash fraction. This study was conducted to determine the influence of several pre- and post-treatment strategies in various physicochemical and adsorptive properties of the resulting activated chars. Pelletized samples of broiler litter and cake were pyrolyzed at 700 °C for 1h followed by a 45 min steam activation at 800 °C at different water flow rates from 1 to 5 mL min(-1). For each activation strategy, samples were either water-rinsed or acid-washed and rinsed or used as is (no acid wash/rinse). Activated char's physicochemical and adsorptive properties towards copper ions were selectively affected by both pre- and post-treatments. Percent ash reduction after either rinsing or acid washing ranged from 1.1 to 15.1% but washed activated chars were still alkaline with pH ranging from 8.4 to 9.1. Acid washing or water rinsing had no significant effect in the ability of the activated char to adsorb copper ions, however it significantly affected surface area, pH, ash content and carbon content. Instead, manure type (litter versus cake) and the activation water flow rate were determining factors in copper ion adsorption which ranged from 38 mg g(-1) to 104 mg g(-1) of activated char. Moreover, strong positive correlations were found between copper uptake and concentration of certain elements in the activated char such as phosphorous, sulfur, calcium and sodium. Rinsing could suffice as a post treatment strategy for ash reduction since no significant differences in the carbon properties were observed between rinsed and acid wash treatments.

Distribution patterns of imported fire ants (Hymenoptera: Formicidae) on a sheep and goat farm in Oklahoma.

Imported fire ant colonies were quantified in 1,000-m(2) circular subplots spaced approximately 125 m apart on a sheep and goat farm in Oklahoma. Social form (percent polygyny), mound density, cumulative above-ground mound volume, and average mound volume were subjected to multiple regression analyses to examine trends related to landscape metrics and habitat characteristics. Monogyne populations were spatially autocorrelated, and polygyne mounds tended to be smaller and more numerous. A model incorporating the effects of percent polygyny, canopy cover, and 1-d cumulative incident solar radiation explained 34% of the variation in mound density. Percent polygyny was not significantly related to cumulative mound volume, which provides a better estimate of overall ant biomass. A model incorporating the effects of 1-d cumulative incident solar radiation on the summer solstice, elevation, canopy cover, distance from cisterns, distance from water, and distance from trees explained 42% of the variation in cumulative mound volume. Monogyne mounds in areas that were flat and close to water in low-lying areas were largest. Results indicate that remotely sensed data in combination with publicly available U.S. Geological Survey data may be useful in predicting areas of high and low fire ant abundance at a field scale.

Monitoring Bacillus thuringiensis-susceptibility in insect pests that occur in large geographies: how to get the best information when two countries are involved.

The adoption of cotton producing insecticidal proteins of Bacillus thuringiensis, commonly referred to as Bt cotton, around the world has proven to be beneficial for growers and the environment. The effectiveness of this important genetically-modified crop can be jeopardized by the development of resistance to Bt cotton by pests it is meant to control, with the possibility that this phenomenon could develop in one country and spread to another by means of insect migration. To preserve the effectiveness of this agricultural biotechnology, regulatory agencies have developed plans to mitigate the development of resistance, and research institutions constantly monitor for shifts in Bt-susceptibility in important pests. If Bt-resistance is detected, this finding needs to be corroborated by an independent laboratory according to current regulatory requirements; a process that presents numerous challenges. We investigated the biological activity of Bt-incorporated diet on Helicoverpa virescens L. after it was stored for several days at different temperatures. Diet stored up to nine days at different temperatures (-14 to 27 degrees C) produced the same biological effect on H. virescens as freshly-prepared diet. Elevating the temperature of Bt stock solution to 76 degrees C as compared to 26 degrees C yielded significantly higher reading of apparent Cry1Ac concentration from MVP II, but not enough to elicit a significant biological response when these stock solutions were incorporated into insect artificial diet. These findings are important particularly when the confirmation of resistance is done at a distant location, such as Mexico, or when diet is shared between laboratories, and must be stored for later use, as in the case of international collaboration.

Environmental influences on flight activity of USDA-ARS Russian and Italian stocks of honey bees (Hymenoptera: Apidae) during almond pollination.

Differences in flight activity and in the percentages of pollen foragers between commercially managed honey bees, Apis mellifera L. (Hymenoptera: Apidae), of two stocks (USDA-ARS Russian, n = 41 colonies; and Italian, n = 43 colonies) were evaluated in an almond, Prunus dulcis (Miller) D. A. Webb, orchard in Kern Co., CA, during February and March 2002. Flight activity was measured by taking 1-min counts of bees exiting colonies on each of 9 d. Flight activity was best predicted with a model containing the effects of colony size (populations of adult bees and sealed brood), temperature, time of day, the interaction of adult bee population with temperature, and the interaction of adult bee population with time of day. Flight increased linearly with adult bee and brood population, had a quadratic relationship with temperature (increasing, but less so at higher temperatures), and had a quadratic relationship with time of day (decreasing, but less so at later times). Larger colonies had more response to changing temperatures and less response to different times of day than small colonies. Bee type had no direct influence on flight activity at any given colony size, temperature, or time of observation or when evaluated using a reduced data set retaining 34 Italian colonies and 32 Russian colonies whose mean sizes were equal. Overall, however, Russian colonies were less populous by about one-fourth and so fielded on average 71% of the foragers that Italian colonies did. Pollen collection was measured by capturing returning foragers on 4 d. The percentages of foragers with pollen were not different for the bee types.