Progress 10/01/10 to 09/30/15
Outputs
Target Audience:Growers and custom applicators who attended the UIUC Agronomy Day in Urbana, as well asweed scientists from academia and the agrochemical industry who attended the North Central Weed Science Society annual meeting in Indianapolis inDecember of 2015. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?Students and scientists who have worked on this project include Rong Ma, PhD graduate student; Sarah O'Brien, MS graduate student; Dr. Anatoli Lygin, laboratory research associate; and Brittany Janney and Erin Lemley, undergraduate laboratory assistants. All people who have worked on this project have learned a great deal about plant physiology, biochemistry, molecular biology, and applied weed management in relation to understanding herbicide-resistance mechanisms in weeds. How have the results been disseminated to communities of interest?Results have been published in the high-impact journal Plant Physiology. Additionally, numerous presentations have been given to growers and custom applicators who attended the UIUC Agronomy Day and AgMaster conference in Urbana, as well as weed scientists from academia and the agrochemical industry who have attended the North Central Weed Science Society annual meetings and Weed Science Society of America annual meetings. What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
Waterhemp (Amaranthus tuberculatus) is a difficult-to-control weed in Illinois soybean and corn production systems. This is in part due to the evolution of multiple herbicide resistances in waterhemp, which is facilitated by its dioecious nature, outcrossing, prolific seed production, and high degree of genetic diversity. A population of waterhemp (designated MCR) from a seed corn field in McLean County, Illinois displays resistance to mesotrione and other 4-hydroxyphenylpyruvate dioxygenase (HPPD) inhibitors, as well as to atrazine and certain ALS-inhibiting herbicides. Our results showed this waterhemp biotype is resistant to mesotrione (plus all commercial HPPD-inhibiting herbicides used for weed control in corn) and atrazine, from both preemergence and postemergence applications, mainly due to rapid metabolism of mesotrione and atrazine (albeit by different detoxification mechanisms and enzymes). The fact that this population is resistant to both HPPD inhibitors and atrazine suggests the ability to achieve herbicide synergism for enhanced weed control may be attenuated under field conditions. These research findings are particularly significant and relevant to crop production and weed management with postemergence herbicides in Zea mays because several other waterhemp populations (and in a related weedy amaranth, A. palmeri) have recently been identified in seedcorn fields throughout the Midwest that possess this unique form of multiple, non-target site-based herbicide resistances. Additional achievements are the identification of a single, specific glutathione S-transferase (GST) gene that appears to confer metabolic-based atrazine resistance in the MCR population, and a cytochrome P450 gene that appears to be involved with mesotrione resistance in MCR. In addition to publications submitted or in preparation regarding gene identfication, these P450 and GST genes can be used as molecular markers to rapidly screen waterhemp populations for mesotrione or atrazine resistance, respectively.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2015
Citation:
Ma, R., Skelton, J.J. and Riechers, D.E. 2015. Measuring rates of herbicide metabolism in dicot weeds with an excised leaf assay. J Vis Exp, 103: e53236, doi:10.3791/53236.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2015
Citation:
Ma, R., Huffman, J., Hager, A.G., Tranel, P.J., Lambert, K.N. and Riechers, D.E. 2015. Evaluation of candidate cytochrome P450 expression in mesotrione-resistant, -sensitive and segregating F2 waterhemp (Amaranthus tuberculatus) populations. Proceedings of the North Central Weed Science Society 70:56.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2015
Citation:
O'Brien, S., Evans, A.F., Huffman, J., Tranel, P.J., Ma, R., Lambert, K.N. and Riechers, D.E. 2015. Correlating phenotype with GST expression in atrazine-resistant and -sensitive Amaranthus tuberculatus segregating F2 lines. Proceedings of the North Central Weed Science Society 70:55.
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Progress 10/01/13 to 09/30/14
Outputs
Target Audience: Growers and custom applicators who attended the UIUC Agronomy Day in Urbana, Illinois andweed scientists from academia and the agrochemical industry who attended the Weed Science Society of America annual meeting in Vancouver, BC, Canada in February of 2014 and the North Central Weed Science Society annual meeting in Minneapolis, December 2014. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided? Advanced undergraduate and graduate student training, as well as postdoctoral training, in methods for plant physiology and biochemistry has been provided to the following project participants: Rong Ma, PhD graduate student; Nick Hausman and Anton Evans, MS graduate students; Dr. Anatoli Lygin, laboratory research associate; and Erin Lemley, undergraduate laboratory assistant. Each person who has worked on this project has learned a great deal of detailed information regarding plant biology, biochemistry, agronomy, crop protection, and applied weed management in the field, growth chamber, greenhouse, and laboratory as part of working in my program. How have the results been disseminated to communities of interest? The basic and applied information generated by this research has been disseminated to weed scientists from academia and the agrochemical industry who attended the Weed Science Society of America annual meeting in Vancouver, BC, Canada in February 2014 and the North Central Weed Science Society meeting in Minneapolis, MN in December 2014, as well as growers and custom applicators who attended the UIUC Agronomy Day in Urbana, IL in August 2014. What do you plan to do during the next reporting period to accomplish the goals? Future work will quantify and compare waterhemp P450 gene expression levels in F2 segregating lines, derived from a cross of MCR and WCS, to further validate the correlation of P450 transcript abundance with phenotype (mesotrione-sensitive or mesotrione-resistant). In addition to further gene expression analyses, the entire P450 coding region from MCR, CHR, and WCS populations will be analyzed using rapid amplification of cDNA ends (RACE) with additional primers designed from the recently published genome and/or transcriptome of cultivated grain amaranth (Amaranthus hypochondriacus). Comparisons of coding regions among these waterhemp P450 proteins will allow for the detection of any alterations in the P450 enzyme’s active site in MCR or CHR compared with WCS. Expression of this P450 marker gene will be used for future molecular-genetic and functional genomics work to further investigate the association of its expression patterns with the resistant phenotype discovered in our prior research at the whole-plant level.
Impacts
What was accomplished under these goals?
A population of waterhemp (designated MCR) from a seed corn field in McLean County, Illinois displays resistance to mesotrione and other 4-hydroxyphenylpyruvate dioxygenase (HPPD) inhibitors, as well as to atrazine and certain acetolactate (ALS)-inhibiting herbicides. Our prior field and greenhouse results indicated this waterhemp biotype is resistant to mesotrione (plus all commercial HPPD-inhibiting herbicides used for weed control in maize) and atrazine, from both preemergence and postemergence applications, mainly due to rapid metabolism of mesotrione and atrazine (although by different detoxification mechanisms and enzymes).These research findings are particularly significant and relevant for crop production and weed management with postemergence (POST) herbicides in maize because several other waterhemp populations have recently been identified in seed corn fields throughout the Midwest that possess this unique form of multiple herbicide resistance, which is based on herbicide detoxification. Our previous research reported metabolic-based resistance to mesotrione in a waterhemp population designated MCR (for McLean County, Illinois HPPD-resistant). Elevated rates of oxidative metabolism, presumably catalyzed by cytochrome P450 monooxygenases (P450s), contributed significantly to mesotrione resistance within the MCR population. Experiments were conducted to test the hypothesis that higher expression levels of specific P450(s) correlate with mesotrione resistance in the MCR population. Another mesotrione-resistant population, CHR (for Champaign County, Illinois HPPD-resistant) and several other mesotrione-sensitive waterhemp populations were used for comparison with MCR. Total RNA extracted from meristem and new leaf tissue from individual waterhemp plants of the same ages and height (10 cm) were used for real-time quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) analysis to compare transcript levels of candidate P450 genes among populations. Primers were designed from conserved regions between several candidate maize and Helianthus P450 cDNAs and the most similar P450 contigs in the waterhemp transcriptome. qRT-PCR demonstrated that a P450 transcript most similar to a maize P450 gene is more highly expressed in meristem tissue of MCR and CHR seedlings (10 cm) compared with each mesotrione-sensitive waterhemp population. Significant differences in expression were not detected when comparing two additional candidate P450s among these waterhemp populations at a height of 10 cm. Expression of the maize P450 homolog in waterhemp, in meristem tissues from MCR seedlings harvested at 4, 6, 8, and 10 cm, was significantly higher than in WCS (for Wayne County, Illinois, herbicide-sensitive) seedlings, but not in 2 cm seedlings or in roots harvested from 10 cm plants. Therefore, only expression of this maize homolog in waterhemp correlated with POST mesotrione resistance, and growth-stage results in MCR and CHR suggested that P450 expression might be growth-stage dependent. Expression of this P450 marker gene can therefore be used for future molecular-genetic and functional genomics work to further investigate the association of its expression patterns with the resistant phenotype discovered in our research. Additionally, growth-stage results suggest that P450 expression may be growth-stage dependent, which negatively correlates with POST mesotrione control of waterhemp under field conditions. For example, significantly greater waterhemp control is obtained when mesotrione is applied POST to smaller waterhemp plants (2-5 cm tall) compared to plants at 5-10 or 10-15 cm tall. Current research is being conducted to quantify P450 expression in F2 segregating lines as well as obtain and compare the entire cDNA sequences of this gene in MCR, CHR, and WCS waterhemp populations.
Publications
- Type:
Journal Articles
Status:
Accepted
Year Published:
2014
Citation:
Hausman, N.E., Tranel, P.J., Riechers, D.E. and Hager, A.G. Responses of a waterhemp (Amaranthus tuberculatus) population resistant to HPPD-inhibiting herbicides to foliar-applied herbicides. Crop Protection.
- Type:
Conference Papers and Presentations
Status:
Accepted
Year Published:
2014
Citation:
Ma, R., Lambert, K.N. and Riechers, D.E. Evaluation of cytochrome P450 expression in mesotrione-resistant and -sensitive waterhemp (Amaranthus tuberculatus) populations. Proceedings North Central Weed Science Society. Vol. 69: pg 133.
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Progress 01/01/13 to 09/30/13
Outputs
Target Audience: Growers and custom applicators who attended the UIUC Agronomy Day in Urbana; weed scientists from academia and the agrochemical industry who attended the Weed Science Society of America annual meeting in Baltimore, Maryland in February 2013 and the North Central Weed Science Society meeting in Columbus, Ohio in December 2013. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided? Advanced undergraduate and graduate student training, as well as postdoctoral training, in methods for plant physiology and biochemistry has been provided to the following project participants: Rong Ma, PhD graduate student; Nick Hausman and Anton Evans, MS graduate students; Dr. Anatoli Lygin, laboratory research associate; Brittany Janney and Erin Lemley, undergraduate laboratory assistants. All people who have worked on this project have learned a great amount of information regarding plant biology, biochemistry, agronomy, crop protection, and applied weed management in the field, growth chamber, greenhouse, and laboratory in my program. How have the results been disseminated to communities of interest? The information generated by this research has been disseminated to weed scientists from academia and the agrochemical industry who attended the Weed Science Society of America annual meeting in Baltimore, Maryland in February 2013 as well as growers and custom applicators who attended the UIUC Agronomy Day in Urbana in August 2013 and the North Central Weed Science Society meeting in Columbus, Ohio in December 2013. What do you plan to do during the next reporting period to accomplish the goals? We will continue to investigate the role of cytochrome P450s in conferring resistance to HPPD-inhibiting herbicides by using chemical P450 inhibitors in field and greenhouse studies, and by conducting in vitro microsomal P450 enzyme activity assays with various herbicide substrates in the lab. Genetics and inheritance research in regards to the HPPD-resistance trait in McLean County waterhemp is ongoing in the greenhouse. In addition, growth chamber studies utilizing chlorophyll fluorescence techniques will be conducted to investigate the potential for achieving synergism between different HPPD inhibitors and photosystem II inhibitors when applied pre-emergence or post-emergence to resistant waterhemp seedlings, with or without P450 inhibitors.
Impacts
What was accomplished under these goals?
A population of waterhemp (designated MCR) from a seed corn field in McLean County, Illinois displays resistance to mesotrione and other 4-hydroxyphenylpyruvate dioxygenase (HPPD) inhibitors, as well as to atrazine and certain ALS-inhibiting herbicides. Our results indicate this waterhemp biotype is resistant to mesotrione (plus all commercial HPPD-inhibiting herbicides used for weed control in corn) and atrazine, from both preemergence and postemergence applications, mainly due to rapid metabolism of mesotrione and atrazine (by different detoxification mechanisms and enzymes). The fact that this population is resistant to both HPPD inhibitors and atrazine suggests the ability to achieve herbicide synergism for enhanced weed control may be attenuated under field conditions. These research findings are particularly significant and relevant to crop production and weed management with postemergence herbicides in maize because several other waterhemp populations have recently been identified in seedcorn fields throughout the Midwest that possess this unique form of multiple herbicide resistance. To address our experimental objectives, growth chamber, greenhouse, and laboratory experiments were conducted to determine if target-site or non-target-site mechanisms confer mesotrione resistance in MCR. In addition, the basis for atrazine resistance was investigated in MCR and an atrazine-resistant, but mesotrione-sensitive waterhemp population (ACR). A standard sensitive population (WCS) was used for comparison in all experiments. Experiments were designed to determine if differential herbicide uptake or metabolism are the basis for mesotrione and atrazine resistance in the MCR waterhemp population. Mesotrione-resistant (MCR) and mesotrione-sensitive (ACR and WCS) populations of waterhemp were treated with radiolabeled mesotrione for analyses of uptake and metabolism during time course experiments. Metabolism studies using whole plants and excised leaves revealed that the time for 50% of absorbed mesotrione to degrade (DT50) in MCR was significantly shorter than in ACR and WCS, which correlated with the phenotypic responses to mesotrione previously reported. The cytochrome P450 inhibitors malathion and tetcyclacis significantly reduced mesotrione metabolism in MCR and corn excised leaves, but not in ACR. Furthermore, malathion increased mesotrione postemergence activity (but not atrazine) in MCR seedlings treated in greenhouse tank-mix studies. Our results indicate that enhanced metabolism in MCR contributes significantly to mesotrione resistance, but further research is still needed to determine if additional non-target site mechanisms may also contribute to mesotrione resistance within the MCR population. In summary, elevated rates of metabolism via different detoxification mechanisms contribute to multiple herbicide resistance to mesotrione and atrazine within the MCR population. Future research with whole plants (utilizing chlorophyll fluorescence techniques) will examine herbicide synergism between mesotrione and atrazine in the greenhouse with the MCR population, and inheritance studies will be performed in the greenhouse to determine how many gene(s) confer resistance to HPPD inhibitors and atrazine in MCR or if mesotrione resistance is a dominant or recessive trait.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Ma, R., S.S. Kaundun, P.J. Tranel, C.W. Riggins, D.L. McGinness, A.G. Hager, T. Hawkes, E. McIndoe, and D.E. Riechers. Distinct detoxification mechanisms confer resistance to mesotrione and atrazine in a population of waterhemp. Plant Physiol. 163: 363-377.
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Hausman, N.E., P.J. Tranel, D.E. Riechers, D.J. Maxwell, L.C. Gonzini, and A.G. Hager. Responses of an HPPD inhibitor-resistant waterhemp (Amaranthus tuberculatus) population to soil-residual herbicides. Weed Technol. 27: 704-711.
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Progress 01/01/12 to 12/31/12
Outputs
OUTPUTS: Waterhemp (Amaranthus tuberculatus) is a difficult-to-control weed in Illinois soybean and corn production systems. This is in part due to the evolution of multiple herbicide resistances in waterhemp, which is facilitated by its dioecious nature, outcrossing, prolific seed production, and high degree of genetic diversity. A population of waterhemp (designated MCR) from a seed corn field in McLean County, Illinois displays resistance to mesotrione and other 4-hydroxyphenylpyruvate dioxygenase (HPPD) inhibitors, as well as to atrazine and certain ALS-inhibiting herbicides. Our results indicate this waterhemp biotype is resistant to mesotrione (plus all commercial HPPD-inhibiting herbicides used for weed control in corn) and atrazine, from both preemergence and postemergence applications, mainly due to rapid metabolism of mesotrione and atrazine (albeit by different detoxification mechanisms and enzymes). The fact that this population is resistant to both HPPD inhibitors and atrazine suggests the ability to achieve herbicide synergism for enhanced weed control may be attenuated under field conditions. These research findings are particularly significant and relevant to crop production and weed management with postemergence herbicides in Zea mays because several other waterhemp populations have recently been identified in seedcorn fields throughout the Midwest that possess this unique form of multiple herbicide resistance. These results have been disseminated at the UIUC Agronomy Day and AgMasters conference in Urbana, and at the Weed Science Society of America annual meeting at the Big Island of Hawaii. PARTICIPANTS: Rong Ma, PhD graduate student; Nick Hausman, MS graduate student; Dr. Anatoli Lygin, laboratory research associate; Jacqueline Janney and Wendy Zhang, undergraduate laboratory assistants. All people who have worked on this project have learned a great deal about plant physiology, biochemistry, and applied weed management. TARGET AUDIENCES: Growers and custom applicators who attend the UIUC Agronomy Day in Urbana, and weed scientists from academia and the agrochemical industry who attended the Weed Science Society of America annual meeting in Hawaii, February 2012. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
Growth chamber, greenhouse, and laboratory experiments were conducted to determine if target-site or non-target-site mechanisms confer mesotrione resistance in MCR. In addition, the basis for atrazine resistance was investigated in MCR and an atrazine-resistant, but mesotrione-sensitive waterhemp population (ACR). A standard sensitive population (WCS) was used for comparison in all experiments. Experiments were designed to determine if differential herbicide uptake or metabolism are the basis for mesotrione and atrazine resistance in the MCR waterhemp population. Atrazine-resistant (MCR and ACR) and atrazine-sensitive (WCS) populations of waterhemp were treated with radiolabeled atrazine to determine rates of metabolism. Mesotrione-resistant (MCR) and mesotrione-sensitive (ACR and WCS) populations of waterhemp were treated with radiolabeled mesotrione for analyses of uptake and metabolism during time course experiments. Mesotrione resistance in MCR is not due to an alteration in HPPD sequence, expression, or reduced herbicide absorption. Metabolism studies using whole plants and excised leaves revealed that the time for 50% of absorbed mesotrione to degrade (DT50) in MCR was significantly shorter than in ACR and WCS, which correlated with the phenotypic responses to mesotrione previously reported. The cytochrome P450 inhibitors malathion and tetcyclacis significantly reduced mesotrione metabolism in MCR and corn excised leaves, but not in ACR. Furthermore, malathion increased mesotrione postemergence activity (but not atrazine) in MCR seedlings treated in greenhouse tank-mix studies. Sequence analysis of atrazine-resistant (MCR and ACR) and atrazine-sensitive (WCS) populations of waterhemp showed no differences in the psbA target-site gene. The DT50s of atrazine in corn, MCR, and ACR leaves were shorter than in WCS, and a polar metabolite of atrazine was found in corn, MCR, and ACR that co-chromatographed with a synthetic atrazine-glutathione conjugate. Our results indicate that enhanced metabolism in MCR contributes significantly to mesotrione resistance, but further research is still needed to determine if additional non-target site mechanisms may also contribute to mesotrione resistance within the MCR population. In summary, elevated rates of metabolism via different detoxification mechanisms contribute to multiple herbicide resistance to mesotrione and atrazine within the MCR population. Additional research with whole plants (utilizing chlorophyll fluorescence techniques) will examine herbicide synergism between mesotrione and atrazine in the greenhouse with the MCR population, and inheritance studies will be performed in the greenhouse to determine how many gene(s) confer resistance to HPPD inhibitors and atrazine in MCR or if mesotrione resistance is a dominant or recessive trait.
Publications
- Ma, R., McGinness, D., Hager, A.G., Tranel, P.J., Kaundun, S., Hawkes, T., Vail, G. and Riechers, D.E. 2012. Investigation of resistance mechanisms to mesotrione in a waterhemp (Amaranthus tuberculatus) population from Illinois. Abstr. Weed Sci. Soc. Amer. 52:412.
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Progress 01/01/11 to 12/31/11
Outputs
OUTPUTS: Waterhemp (Amaranthus tuberculatus) is a difficult-to-control weed in Illinois soybean and corn production. This is in part due to the evolution of multiple herbicide resistances in waterhemp, which is facilitated by its dioecious biology and genetic diversity. A population of waterhemp (designated MCR) from a seed corn field in McLean County, Illinois displays resistance to mesotrione and other 4-hydroxyphenylpyruvate dioxygenase (HPPD) inhibitors, as well as to atrazine and certain ALS-inhibiting herbicides. Our results indicate that this waterhemp biotype is resistant to all commercial HPPD-inhibiting herbicides used for weed control in corn, from both preemergence and postemergence applications. The fact that this biotype is resistant to both HPPD inhibitors and atrazine suggests that the ability to achieve herbicide synergism for enhanced weed control with these herbicides may be attenuated. These results have been disseminated at the UIUC Agronomy Day and AgMasters conference in Urbana, and at the North Central Weed Science Society meeting in Milwaukee. PARTICIPANTS: Rong Ma, PhD graduate student; Nick Hausman, MS graduate student; Dan McGinness, laboratory technician; and Wendy Zhang, undergraduate laboratory assistant. TARGET AUDIENCES: Growers and custom applicators who attended the UIUC Agronomy Day in Urbana; weed scientists from academia and the agrochemical industry who attended the North Central Weed Science Society annual meeting. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
Growth chamber, greenhouse, and laboratory experiments were conducted to determine if differential herbicide uptake or metabolism are the basis for mesotrione and atrazine resistance in the MCR waterhemp population. Atrazine-resistant (MCR and ACR) and atrazine-sensitive (WCS) biotypes of waterhemp were treated with radiolabeled atrazine to determine rates of metabolism. The half-lives of atrazine in corn, MCR, and ACR were much shorter (less than 2 hours) than in WCS (greater than 12 hours). Thus, enhanced metabolism endows MCR and ACR with resistance to postemergence atrazine. Mesotrione-resistant (MCR) and mesotrione-sensitive (ACR and WCS) biotypes of waterhemp were treated with radiolabeled mesotrione for analyses of uptake and metabolism during time course experiments. Metabolism studies using whole plants and excised leaves revealed that the half-lives of mesotrione in MCR and corn were shorter than in ACR and WCS, which correlates with phenotypic responses to mesotrione applied postemergence in corn and in these waterhemp biotypes. The cytochrome P450 inhibitors malathion and tetcyclacis significantly inhibited mesotrione metabolism in excised leaves of MCR and corn at 6 and 24 HAT, but had no effect in ACR. Our results indicate that enhanced herbicide metabolism in MCR may contribute significantly to mesotrione resistance, but further research is needed to determine if additional non-target site mechanisms may also contribute to mesotrione resistance in MCR. In addition, whole-plant and chlorophyll fluorescence research examining herbicide synergism between mesotrione and atrazine will be conducted in the greenhouse with the MCR biotype, and inheritance studies will be performed to determine how many genes are involved in resistance to HPPD inhibitors and atrazine in MCR and if resistance is conferred by a dominant or recessive trait.
Publications
- Hausman, N.E., Singh, S., Tranel, P.J., Riechers, D.E., Kaundun, S.S., Polge, N.D., Thomas, D.A. and Hager, A.G. 2011. Resistance to HPPD-inhibiting herbicides in a population of waterhemp (Amaranthus tuberculatus) from Illinois, United States. Pest Manag. Sci. 67:258-261.
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