Stoneville, MS, United States
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Molin W.T.,Crop Production Systems Research Unit | Wright A.A.,Crop Production Systems Research Unit | Lawton-Rauh A.,Clemson University | Saski C.A.,Clemson University
BMC Genomics | Year: 2017

Background: The expanding number and global distributions of herbicide resistant weedy species threaten food, fuel, fiber and bioproduct sustainability and agroecosystem longevity. Amongst the most competitive weeds, Amaranthus palmeri S. Wats has rapidly evolved resistance to glyphosate primarily through massive amplification and insertion of the 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene across the genome. Increased EPSPS gene copy numbers results in higher titers of the EPSPS enzyme, the target of glyphosate, and confers resistance to glyphosate treatment. To understand the genomic unit and mechanism of EPSPS gene copy number proliferation, we developed and used a bacterial artificial chromosome (BAC) library from a highly resistant biotype to sequence the local genomic landscape flanking the EPSPS gene. Results: By sequencing overlapping BACs, a 297 kb sequence was generated, hereafter referred to as the "EPSPS cassette." This region included several putative genes, dense clusters of tandem and inverted repeats, putative helitron and autonomous replication sequences, and regulatory elements. Whole genome shotgun sequencing (WGS) of two biotypes exhibiting high and no resistance to glyphosate was performed to compare genomic representation across the EPSPS cassette. Mapping of sequences for both biotypes to the reference EPSPS cassette revealed significant differences in upstream and downstream sequences relative to EPSPS with regard to both repetitive units and coding content between these biotypes. The differences in sequence may have resulted from a compounded-building mechanism such as repetitive transpositional events. The association of putative helitron sequences with the cassette suggests a possible amplification and distribution mechanism. Flow cytometry revealed that the EPSPS cassette added measurable genomic content. Conclusions: The adoption of glyphosate resistant cropping systems in major crops such as corn, soybean, cotton and canola coupled with excessive use of glyphosate herbicide has led to evolved glyphosate resistance in several important weeds. In Amaranthus palmeri, the amplification of the EPSPS cassette, characterized by a complex array of repetitive elements and putative helitron sequences, suggests an adaptive structural genomic mechanism that drives amplification and distribution around the genome. The added genomic content not found in glyphosate sensitive plants may be driving evolution through genome expansion. © 2016 The Author(s).


Gaines T.A.,University of Western Australia | Gaines T.A.,Bayer CropScience | Wright A.A.,Crop Production Systems Research Unit | Molin W.T.,Crop Production Systems Research Unit | And 6 more authors.
PLoS ONE | Year: 2013

Weed populations can have high genetic plasticity and rapid responses to environmental selection pressures. For example, 100-fold amplification of the 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene evolved in the weed species Amaranthus palmeri to confer resistance to glyphosate, the world's most important herbicide. However, the gene amplification mechanism is unknown. We sequenced the EPSPS gene and genomic regions flanking EPSPS loci in A. palmeri, and searched for mobile genetic elements or repetitive sequences. The EPSPS gene was 10,229 bp, containing 8 exons and 7 introns. The gene amplification likely proceeded through a DNA-mediated mechanism, as introns exist in the amplified gene copies and the entire amplified sequence is at least 30 kb in length. Our data support the presence of two EPSPS loci in susceptible (S) A. palmeri, and that only one of these was amplified in glyphosate-resistant (R) A. palmeri. The EPSPS gene amplification event likely occurred recently, as no sequence polymorphisms were found within introns of amplified EPSPS copies from R individuals. Sequences with homology to miniature inverted-repeat transposable elements (MITEs) were identified next to EPSPS gene copies only in R individuals. Additionally, a putative Activator (Ac) transposase and a repetitive sequence region were associated with amplified EPSPS genes. The mechanism controlling this DNA-mediated amplification remains unknown. Further investigation is necessary to determine if the gene amplification may have proceeded via DNA transposon-mediated replication, and/or unequal recombination between different genomic regions resulting in replication of the EPSPS gene.


Stetina S.R.,U.S. Department of Agriculture | Molin W.T.,Crop Production Systems Research Unit
Journal of Cotton Science | Year: 2014

Reniform nematode (Rotylenchulus reniformis) resistance is being transferred to Gossypium hirsutum from its distant relatives. Reports of fluometuron damage to LONREN lines with nematode resistance from G. longicalyx raised concerns about introducing herbicide sensitivity from other nematode resistance sources. The research objective was to evaluate 15 sources of reniform nematode resistance for their reaction to fluometuron three weeks after planting in a replicated greenhouse trial: two G. herbaceum accessions, four G. arboreum accessions, three G. barbadense accessions, three G. hirsutum accessions, and three G. hirsutum lines with resistance introgressed from G. barbadense (FR-05) or G. longicalyx (LONREN-1 and LONREN-2). The control genotype was G. hirsutum cultivar Deltapine 161 B2RF. Across all herbicide rates tested, mean herbicide injury ratings for G. arboreum accessions were greater than the control, whereas G. barbadense GB 713 and TX 110 were less. Regression analysis of herbicide rates indicated that injury increased linearly with increasing herbicide rate for all accessions, although G. arboreum A2-083 had more injury than the control. Regression analysis of herbicide rates indicated that biomass decreased linearly with increasing herbicide rate for all accessions, although G. barbadense GB 713 and Pima PHY 800 exhibited greater biomass reduction than the control. Across all herbicide rates tested, mean electron transport rates of all G. herbaceum and G. arboreum accessions and G. barbadense Pima PHY 800 were lower than the control. The relationship between herbicide rate and electron transport rate was curvilinear, with similar decreases in electron transport rate in response to increasing herbicide concentration for all lines. Increased sensitivity to fluometuron could be introduced into G. hirsutum through crosses with distantly related species, but with the exception of G. arboreum A2-083, the lines did not respond to the herbicide differently from the control. © The Cotton Foundation 2014.


Maroli A.S.,Clemson University | Nandula V.K.,Crop Production Systems Research Unit | Gerard P.,Crop Production Systems Research Unit | Tharayil N.,Clemson University
Journal of Agricultural and Food Chemistry | Year: 2015

Metabolomics and biochemical assays were employed to identify physiological perturbations induced by a commercial formulation of glyphosate in susceptible (S) and resistant (R) biotypes of Amaranthus palmeri. At 8 h after treatment (HAT), compared to the respective water-treated control, cellular metabolism of both biotypes were similarly perturbed by glyphosate, resulting in abundance of most metabolites including shikimic acid, amino acids, organic acids and sugars. However, by 80 HAT the metabolite pool of glyphosate-treated R-biotype was similar to that of the control S- and R-biotypes, indicating a potential physiological recovery. Furthermore, the glyphosate-treated R-biotype had lower reactive oxygen species (ROS) damage, higher ROS scavenging activity, and higher levels of potential antioxidant compounds derived from the phenylpropanoid pathway. Thus, metabolomics, in conjunction with biochemical assays, indicate that glyphosate-induced metabolic perturbations are not limited to the shikimate pathway, and the oxidant quenching efficiency could potentially complement the glyphosate resistance in this R-biotype. © 2015 American Chemical Society.


Weaver M.A.,Biological Control of Pests Research Unit | Boyette C.D.,Biological Control of Pests Research Unit | Hoagland R.E.,Crop Production Systems Research Unit
Biocontrol Science and Technology | Year: 2016

Among the most important and visible weeds in the Southeastern USA is the exotic invasive vine, kudzu (Pueraria montana var. lobata). Efforts to eradicate it typically involve many years of application of restricted-use pesticides. Recent availability of effective, non-restricted-use pesticides and developments with the application of the bioherbicide Myrothecium verrucaria has made possible new control programmes for kudzu management. Field trials at three sites over two years with aminocyclopyrachlor, aminopyralid, fluroxypyr, metsulfuron methyl and combinations of these herbicides achieved 99–100% reduction in aboveground kudzu biomass. Additionally, programmes were developed that eradicated kudzu while simultaneously establishing native vegetation. One of these successful programmes integrated bioherbicide application, mechanical removal of kudzu biomass and planting switchgrass (Panicum virgatum) in an entirely chemical herbicide-free system. These field tests demonstrate a variety of methods that can be used independently or in an integrated approach for rapid kudzu eradication. ©, The work was authored as part of the Authors' official duties as employees of the United States Government and is therefore a work of the United States Government. In accordance with 17 U.S.C. 105 no copyright protection is available for such works under U.S. law.


Ribeiro D.N.,Mississippi State University | Pan Z.,Natural Products Utilization Research Unit | Duke S.O.,Natural Products Utilization Research Unit | Nandula V.K.,Crop Production Systems Research Unit | And 3 more authors.
Planta | Year: 2014

The inheritance of glyphosate resistance in two Amaranthus palmeri populations (R1 and R2) was examined in reciprocal crosses (RC) and second reciprocal crosses (2RC) between glyphosate-resistant (R) and -susceptible (S) parents of this dioecious species. R populations and Female-R × Male-S crosses contain higher 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene copy numbers than the S population. EPSPS expression, EPSPS enzyme activity, EPSPS protein quantity, and level of resistance to glyphosate correlated positively with genomic EPSPS relative copy number. Transfer of resistance was more influenced by the female than the male parent in spite of the fact that the multiple copies of EPSPS are amplified in the nuclear genome. This led us to hypothesize that this perplexing pattern of inheritance may result from apomictic seed production in A. palmeri. We confirmed that reproductively isolated R and S female plants produced seeds, indicating that A. palmeri can produce seeds both sexually and apomictically (facultative apomixis). This apomictic trait accounts for the low copy number inheritance in the Female-S × Male-R offsprings. Apomixis may also enhance the stability of the glyphosate resistance trait in the R populations in the absence of reproductive partners. © 2013 Springer-Verlag Berlin Heidelberg.


Queiroz S.C.N.,EMBRAPA - Empresa Brasileira de Pesquisa Agropecuária | Cantrell C.L.,National United University | Duke S.O.,National United University | Wedge D.E.,National United University | And 3 more authors.
Journal of Agricultural and Food Chemistry | Year: 2012

Conyza canadensis (L.) Cronquist syn. (horseweed) is a problematic and invasive weed with reported allelopathic properties. To identify the phytotoxic constituents of the aerial parts, a systematic bioactivity-guided fractionation of the dichloromethane extract was performed. Three active enyne derivatives, (2Z,8Z)-matricaria acid methyl ester, (4Z,8Z)-matricaria lactone, and (4Z)-lachnophyllum lactone, were identified. The lactones inhibited growth of the monocot Agrostis stolonifera (bentgrass) and the dicot Lactuca sativa (lettuce) at 1 mg mL-1, while the (2Z,8Z)-matricaria acid methyl ester was less active. In a dose-response screening of the lactones for growth inhibitory activity against Lemna paucicostata, (4Z)-lachnophyllum lactone was the most active with an IC50 of 104 μM, while the (4Z,8Z)-matricaria lactone was less active (IC50 of 220 μM). In a fungal direct bioautography assay, the two lactones at 10 and 100 μg/spot inhibited growth of the plant pathogenic fungi Colletotrichum acutatum, Colletotrichum fragariae, and Colletotrichum gloeosporioides. In a dose-response screening of the lactones against six different plant pathogenic fungi, (4Z,8Z)-matricaria lactone was more active than the commercial fungicide azoxystrobin on Col. acutatum, Col. fragariae, and Col. gloeosporioides at 30 μM and about as active as the commercial fungicide captan against Col. gloeosporioides, while (4Z)-lachnophyllum lactone was less active. © 2012 American Chemical Society.


Reddy K.N.,Crop Production Systems Research Unit | Nandula V.K.,Crop Production Systems Research Unit
Indian Journal of Agronomy | Year: 2012

Advances in biotechnology have led to development and commercialization of several herbicide-resistant crops (HRCs) in the mid-1990s. HRCs survive herbicide treatment that previously would have killed the crop along with targetted weeds. Both transgenic (created through stable integration of a foreign gene) and non-transgenic (developed through traditional plant breeding) HRCs are commercially available to farmers. Although several HRCs are available, only transgenic HRCs, such as, glyphosate- and glufosinate-resistant crops appear to have greatest impact and dominate the market. HRCs are readily accepted in North and South America and are slowly making inroads into other parts of world. Farmers who have chosen HRCs must have seen some economic and weed control benefits; otherwise, the rapid increase in area planted to HRCs in recent years would not have occurred. There are benefits and risks associated with the use of HRCs as a weed management tool. The benefits of HRCs for weed management outweigh the risks based on current knowledge. HRCs should not be relied on solely to the exclusion of other weed control measures and should be used within integrated weed management programmes.


Bellaloui N.,Crop Genetics Research Unit | Bruns H.A.,Crop Production Systems Research Unit | Abbas H.K.,Biological Control of Pests Research Unit | Mengistu A.,Crop Genetics Research Unit | And 2 more authors.
Frontiers in Plant Science | Year: 2015

Information on the effects of management practices on soybean seed composition is scarce. Therefore, the objective of this research was to investigate the effects of planting date (PD) and seeding rate (SR) on seed composition (protein, oil, fatty acids, and sugars) and seed minerals (B, P, and Fe) in soybean grown in two row-types (RTs) on the Mississippi Delta region of the Midsouth USA. Two field experiments were conducted in 2009 and 2010 on Sharkey clay and Beulah fine sandy loam soil at Stoneville, MS, USA, under irrigated conditions. Soybean were grown in 102 cm single-rows and 25 cm twin-rows in 102 cm centers at SRs of 20, 30, 40, and 50 seeds m−2. The results showed that in May and June planting, protein, glucose, P, and B concentrations increased with increased SR, but at the highest SRs (40 and 50 seeds m−2), the concentrations remained constant or declined. Palmitic, stearic, and linoleic acid concentrations were the least responsive to SR increases. Early planting resulted in higher oil, oleic acid, sucrose, B, and P on both single and twin-rows. Late planting resulted in higher protein and linolenic acid, but lower oleic acid and oil concentrations. The changes in seed constituents could be due to changes in environmental factors (drought and temperature), and nutrient accumulation in seeds and leaves. The increase of stachyose sugar in 2010 may be due to a drier year and high temperature in 2010 compared to 2009; suggesting the possible role of stachyose as an environmental stress compound. Our research demonstrated that PD, SR, and RT altered some seed constituents, but the level of alteration in each year dependent on environmental factors such as drought and temperature. This information benefits growers and breeders for considering agronomic practices to select for soybean seed nutritional qualities under drought and high heat conditions. © 2015 by FUOC.


PubMed | Clemson University and Crop Production Systems Research Unit
Type: Journal Article | Journal: BMC genomics | Year: 2017

The expanding number and global distributions of herbicide resistant weedy species threaten food, fuel, fiber and bioproduct sustainability and agroecosystem longevity. Amongst the most competitive weeds, Amaranthus palmeri S. Wats has rapidly evolved resistance to glyphosate primarily through massive amplification and insertion of the 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene across the genome. Increased EPSPS gene copy numbers results in higher titers of the EPSPS enzyme, the target of glyphosate, and confers resistance to glyphosate treatment. To understand the genomic unit and mechanism of EPSPS gene copy number proliferation, we developed and used a bacterial artificial chromosome (BAC) library from a highly resistant biotype to sequence the local genomic landscape flanking the EPSPS gene.By sequencing overlapping BACs, a 297kb sequence was generated, hereafter referred to as the EPSPS cassette. This region included several putative genes, dense clusters of tandem and inverted repeats, putative helitron and autonomous replication sequences, and regulatory elements. Whole genome shotgun sequencing (WGS) of two biotypes exhibiting high and no resistance to glyphosate was performed to compare genomic representation across the EPSPS cassette. Mapping of sequences for both biotypes to the reference EPSPS cassette revealed significant differences in upstream and downstream sequences relative to EPSPS with regard to both repetitive units and coding content between these biotypes. The differences in sequence may have resulted from a compounded-building mechanism such as repetitive transpositional events. The association of putative helitron sequences with the cassette suggests a possible amplification and distribution mechanism. Flow cytometry revealed that the EPSPS cassette added measurable genomic content.The adoption of glyphosate resistant cropping systems in major crops such as corn, soybean, cotton and canola coupled with excessive use of glyphosate herbicide has led to evolved glyphosate resistance in several important weeds. In Amaranthus palmeri, the amplification of the EPSPS cassette, characterized by a complex array of repetitive elements and putative helitron sequences, suggests an adaptive structural genomic mechanism that drives amplification and distribution around the genome. The added genomic content not found in glyphosate sensitive plants may be driving evolution through genome expansion.

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