Target-Site Mutations and Expression of ALS Gene Copies Vary According to Echinochloa Species

The sustainability of rice cropping systems is jeopardized by the large number and variety of populations of polyploid Echinochloa spp. resistant to ALS inhibitors. Better knowledge of the Echinochloa species present in Italian rice fields and the study of ALS genes involved in target-site resistance could significantly contribute to a better understanding of resistance evolution and management. Using a CAPS-rbcL molecular marker, two species, E. crus-galli (L.) P. Beauv. and E. oryzicola (Vasinger) Vasing., were identified as the most common species in rice in Italy. Mutations involved in ALS inhibitor resistance in the different species were identified and associated with the ALS homoeologs. The relative expression of the ALS gene copies was evaluated. Molecular characterization led to the identification of three ALS genes in E. crus-galli and two in E. oryzicola. The two species also carried different point mutations conferring resistance: Ala122Asn in E. crus-galli and Trp574Leu in E. oryzicola. Mutations were carried in the same gene copy (ALS1), which was significantly more expressed than the other copies (ALS2 and ALS3) in both species. These results explain the high resistance level of these populations and why mutations in the other ALS copies are not involved in herbicide resistance.

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Confirmation of Glyphosate-Resistant Kochia (Kochia scoparia) from Sugar Beet Fields in Idaho and Oregon

Weed Technology ◽

10.1017/wet.2017.80 ◽

2017 ◽

Vol 32 (1) ◽

pp. 27-33 ◽

Cited By ~ 3

Author(s):

Vipan Kumar ◽

Joel Felix ◽

Don Morishita ◽

Prashant Jha

Keyword(s):

Sugar Beet ◽

Copy Number ◽

Cropping Systems ◽

Gene Copy Number ◽

Glyphosate Resistance ◽

Gene Copy ◽

Resistance Level ◽

Control Programs ◽

Gene Copies

AbstractGlyphosate-resistant (GR) kochia is an increasing management concern in major cropping systems of the northwestern US. In 2014, we investigated four putative GR kochia accessions (designated as ALA, VAL, WIL, DB) collected from sugar beet fields in eastern Oregon and southwestern Idaho to characterize the level of evolved glyphosate resistance and determine the relationship between the 5-enol-pyruvylshikimate-3-phospate synthase (EPSPS) gene copy number and level of glyphosate resistance. TheEPSPSgene copy number was used as a molecular marker to detect GR kochia in subsequent surveys in 2015 and 2016. Based on LD50values from a whole-plant dose-response study, the four putative GR kochia populations were 2.0- to 9.6-fold more resistant to glyphosate than the glyphosate-susceptible (GS) accession. In anin vivoleaf-disk shikimate assay, leaf disks of GS kochia plants treated with 100-μM glyphosate accumulated 2.4- to 4.0-fold higher amounts of shikimate than the GR plants. The four GR accessions had 2.7 to 9.1 relativeEPSPSgene copies compared with the GS accession (<1EPSPSgene copies), and there was a linear relationship betweenEPSPSgene copy number and glyphosate resistance level (LD50values). The 2015 and 2016 GR kochia survey results indicated that about half of the collected populations from sugar beet fields in eastern Oregon had developed resistance to glyphosate whereas only one population from the Idaho collection was confirmed glyphosate resistant. This is the first confirmation of GR kochia in sugar beet fields in eastern Oregon and southwestern Idaho. Diversified weed control programs will be required to prevent further development and spread of GR kochia in sugar beet-based rotations in this region.

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Multiple herbicide resistance in California Italian ryegrass (Lolium perenne ssp. multiflorum): characterization of ALS-inhibiting herbicide resistance

Weed Science ◽

10.1017/wsc.2019.1 ◽

2019 ◽

Vol 67 (3) ◽

pp. 273-280 ◽

Cited By ~ 4

Author(s):

Parsa Tehranchian ◽

Vijay K. Nandula ◽

Maor Matzrafi ◽

Marie Jasieniuk

Keyword(s):

Lolium Perenne ◽

Herbicide Resistance ◽

Resistance Mechanism ◽

Acetolactate Synthase ◽

Target Site ◽

Italian Ryegrass ◽

Resistance Level ◽

Als Inhibitors ◽

Inhibitor Resistance ◽

Als Inhibitor

AbstractMultiple resistance to glyphosate, sethoxydim, and paraquat was previously confirmed in two Italian ryegrass [Lolium perenne L. ssp. multiflorum (Lam.) Husnot] populations, MR1 and MR2, in northern California. Preliminary greenhouse studies revealed that both populations were also resistant to imazamox and mesosulfuron, both of which are acetolactate synthase (ALS)-inhibiting herbicides. In this study, three subpopulations, MR1-A (from seed of MR1 plants that survived a 16X rate of sethoxydim), MR1-P (from seed of MR1 plants that survived a 2X rate of paraquat), and MR2 (from seed of MR2 plants that survived a 16X rate of sethoxydim), were investigated to determine the resistance level to imazamox and mesosulfuron, evaluate other herbicide options for the control of these multiple resistant L. perenne ssp. multiflorum, and characterize the underlying ALS-inhibitor resistance mechanism(s). Based on LD50 values, the MR1-A, MR1-P, and MR2 subpopulations were 38-, 29-, 8-fold and 36-, 64-, and 3-fold less sensitive to imazamox and mesosulfuron, respectively, relative to the susceptible (Sus) population. Only MR1-P and MR2 plants were cross-resistant to rimsulfuron, whereas both MR1 subpopulations were cross-resistant to imazethapyr. Pinoxaden (ACCase inhibitor [phenylpyrazoline 'DEN']) only controlled MR2 and Sus plants at the labeled field rate. However, all plants were effectively controlled (>99%) with the labeled field rate of glufosinate. Based on I50 values, MR1-A, MR-P, and MR2 plants were 712-, 1,104-, and 3-fold and 10-, 18-, and 5-fold less responsive to mesosulfuron and imazamox, respectively, than the Sus plants. Sequence alignment of the ALS gene of resistant plants revealed a missense single-nucleotide polymorphism resulting in a Trp-574-Leu substitution in MR1-A and MR1-P plants, heterozygous in both, but not in the MR2 plants. An additional homozygous substitution, Asp-376-Glu, was identified in the MR1-A plants. Addition of malathion or piperonyl butoxide did not alter the efficacy of mesosulfuron on MR2 plants. In addition, the presence of 2,4-D had no effect on the response of mesosulfuron on the MR2 and Sus. These results suggest an altered target site is the mechanism of resistance to ALS inhibitors in MR1-A and MR1-P plants, whereas a non–target site based resistance apparatus is present in the MR2 plants.

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Identification of Japanese Foxtail (Alopecurus Japonicus) Resistant to Haloxyfop Using Three Different Assay Techniques

Weed Science ◽

10.1614/ws-07-016.1 ◽

2007 ◽

Vol 55 (6) ◽

pp. 537-540 ◽

Cited By ~ 31

Author(s):

Caihong Yang ◽

Liyao Dong ◽

Jun Li ◽

Stephen R. Moss

Keyword(s):

Oilseed Rape ◽

Coenzyme A ◽

Resistance Index ◽

Target Site ◽

The Other ◽

Resistance Level ◽

Acetyl Coenzyme A ◽

Whole Plant ◽

Low Degrees ◽

Acetyl Coenzyme A Carboxylase

The objective of this study was to investigate the resistance level of Japanese foxtail to haloxyfop, an acetyl coenzyme A carboxylase (ACCase; EC 6.4.1.2)–inhibiting herbicide. Eleven biotypes were collected from oilseed rape fields in different areas in Jiangsu and Anhui provinces where haloxyfop had been continuously applied for various periods. Biotypes were assessed by two different methods, a seed bioassay and whole-plant assay, to identify the most resistant and susceptible biotypes for further studies on the activity of the target enzyme ACCase. A good correlation was obtained between the two different bioassay methods. The Jurong and Chuzhou biotypes were the most resistant and susceptible biotypes, respectively, whereas the other nine biotypes showed variable and relatively low degrees of haloxyfop resistance. Furthermore, target-site enzyme sensitivity results confirmed that the Jurong biotype was resistant to haloxyfop with a concentration of herbicide causing 50% inhibition of ACCase activity (IC50) of 9.19 µM, whereas the IC50 of the susceptible biotype (Chuzhou) was 0.76 µM, giving a resistance index of 12.

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Different Mutations Providing Target Site Resistance to ALS- and ACCase-Inhibiting Herbicides in Echinochloa spp. from Rice Fields

Agriculture ◽

10.3390/agriculture11050382 ◽

2021 ◽

Vol 11 (5) ◽

pp. 382

Author(s):

Ignacio Amaro-Blanco ◽

Yolanda Romano ◽

Jose Antonio Palmerin ◽

Raquel Gordo ◽

Candelario Palma-Bautista ◽

...

Keyword(s):

Integrated Control ◽

Point Mutations ◽

Acetolactate Synthase ◽

Rice Fields ◽

Target Site ◽

Cross Resistance ◽

Resistance Evolution ◽

Resistance Patterns ◽

Frequent Mutations ◽

Western Spain

Echinochloa spp. is one of the most invasive weeds in rice fields worldwide. Acetolactate synthase (ALS) and acetyl-CoA carboxylase (ACCase) inhibiting herbicides are two of the most widely used rice herbicides. However, overuse has led to the resistance evolution of Echinochloa spp. to penoxsulam (ALS-inhibitor) and cyhalofop-methyl (ACCase-inhibitor). In this work, 137 different Echinochloa spp. populations were collected in different rice fields in Extremadura (western Spain) where lack of control was detected. Target-site based resistance (by sequencing ALS and ACCase gene) and characterization of Echinochloa species at the molecular level (based on PCR-RFLP analyses) were carried out in those populations. Most of the populations studied (111 of 137) belong to the E. oryzicola/E. oryzoides group. Three-point mutations were identified in ALS genes: Pro197Ser, Pro197Thr, and Ser653Asn, the first being the most frequent substitution in resistant plants. In the ACCase gene, the Ile1781Leu substitution was found. In both ALS and ACCase sequencing, evidence of heterozygosity was also observed. To assess whether cross-resistance patterns differed between mutations, two populations belonging to the E. oryzicola/E. oryzoides group had its most frequent mutations (Pro197Ser, population ech3-14 and Ile1781Leu, population ech114-10) chosen to be carried out in a dose-response assay. It was confirmed that Pro197Ser conferred resistance to triazolopyrimidine, imidazolinone, sulfonylurea, and pyrimidinyl benzoate families. On the other hand, the Ile1781Leu change gave resistance to aryloxyphenoxypropionate and cyclohexanedione families. Of the authorized herbicides in rice in Spain, more that 80% belong to these families. It is therefore important that farmers carry out an integrated control system that combines both chemical and non-chemical tools.

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Target-site Mutation and Fitness Cost of Acetolactate Synthase Inhibitor-resistant Annual Bluegrass

HortScience ◽

10.21273/hortsci13512-18 ◽

2019 ◽

Vol 54 (4) ◽

pp. 701-705

Author(s):

Te-Ming Tseng ◽

Swati Shrestha ◽

James D. McCurdy ◽

Erin Wilson ◽

Gourav Sharma

Keyword(s):

Dna Sequencing ◽

Acetolactate Synthase ◽

Target Site ◽

Alternative Methods ◽

Resistance Level ◽

Annual Bluegrass ◽

Mechanism Of Resistance ◽

Als Inhibitors ◽

Inhibitor Resistance ◽

Als Inhibitor

Annual bluegrass (Poa annua L.) is an annual weed that is particularly troublesome in managed turfgrass. It has been controlled conventionally with herbicides, including acetolactate synthase (ALS) inhibitors. However, resistance to ALS inhibitors has been documented throughout the southeastern United States since 2012. A rate–response trial was conducted to confirm and determine the resistance level of suspected resistant P. annua biotypes from Mississippi (Reunion), followed by DNA sequencing to determine whether the mechanism of resistance is a target-site mutatio n. In addition, a fitness assay was conducted together with a susceptible biotype to determine whether resistance to ALS inhibitors is associated with decreased fitness. Reunion was at least 45 times more resistant to foramsulfuron than the standard susceptible biotype based on I50 estimates [I50 is the rate of herbicide giving a 50% response (50% visual necrosis)], requiring a predicted 331 g a.i./ha foramsulfuron for 50% control. DNA sequencing results identified a Trp574-to-Leu mutation in the ALS gene of the Reunion biotype, which has been shown by other studies to confer resistance to ALS inhibitors. Measurement of fitness parameters among the Reunion and susceptible biotypes demonstrated reduced seed yield, tillering, and flowering time in the resistant Reunion biotype, suggesting that ALS inhibitor resistance is possibly correlated to decreased fitness in P. annua. Alternative methods to control P. annua need to be considered as a result of the evolution of herbicide-resistant biotypes. An integrated management strategy to control P. annua weeds will help prevent further evolution of resistance. Because this study evaluated only the target-site mechanism of resistance, it is also necessary to determine whether the resistant biotype has reduced uptake, translocation, or enhanced metabolism as additional mechanisms of resistance. Consequently, a fitness study encompassing a more comprehensive list of plant parameters will provide conclusions of the fitness costs associated with ALS inhibitor resistance in P. annua. Chemical names: Foramsulfuron {1-(4,6-dimethoxypyrimidin-2-yl)-3-[2-(dimethylcarbamoyl)-5-formamidophenylsulfonyl] urea}.

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Expression of rRNA Genes and Nucleolus Formation at Ectopic Chromosomal Sites in the Yeast Saccharomyces cerevisiae

Molecular and Cellular Biology ◽

10.1128/mcb.02324-05 ◽

2006 ◽

Vol 26 (16) ◽

pp. 6223-6238 ◽

Cited By ~ 18

Author(s):

Melanie L. Oakes ◽

Katsuki Johzuka ◽

Loan Vu ◽

Kristilyn Eliason ◽

Masayasu Nomura

Keyword(s):

The Other ◽

Rrna Genes ◽

Gene Copy ◽

Rrna Gene ◽

Yeast Saccharomyces Cerevisiae ◽

Pol I ◽

Gene Copies ◽

Tandem Repetition ◽

Polymerase I ◽

Single Versus

ABSTRACT We constructed yeast strains in which rRNA gene repeats are integrated at ectopic sites in the presence or absence of the native nucleolus. At all three ectopic sites analyzed, near centromere CEN5, near the telomere of chromosome VI-R, and in middle of chromosome V-R (mid-V-R), a functional nucleolus was formed, and no difference in the expression of rRNA genes was observed. When two ribosomal DNA (rDNA) arrays are present, one native and the other ectopic, there is codominance in polymerase I (Pol I) transcription. We also examined the expression of a single rDNA repeat integrated into ectopic loci in strains with or without the native RDN1 locus. In a strain with reduced rRNA gene copies at RDN1 (∼40 copies), the expression of a single rRNA gene copy near the telomere was significantly reduced relative to the other ectopic sites, suggesting a less-efficient recruitment of the Pol I machinery from the RDN1 locus. In addition, we found a single rRNA gene at mid-V-R was as active as that within the 40-copy RDN1. Combined with the results of activity analysis of a single versus two tandem copies at CEN5, we conclude that tandem repetition is not required for efficient rRNA gene transcription.

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Target Site–Based and Non–Target Site Based Resistance to ALS Inhibitors in Palmer Amaranth (Amaranthus palmeri)

Weed Science ◽

10.1017/wsc.2017.43 ◽

2017 ◽

Vol 65 (6) ◽

pp. 681-689 ◽

Cited By ~ 16

Author(s):

Sridevi Nakka ◽

Curtis R. Thompson ◽

Dallas E. Peterson ◽

Mithila Jugulam

Keyword(s):

Cropping Systems ◽

Acetolactate Synthase ◽

The United States ◽

Palmer Amaranth ◽

Target Site ◽

Nucleotide Sequence Analysis ◽

Response Analysis ◽

Common Mutation ◽

Weed Species ◽

Als Inhibitors

Resistance to acetolactate synthase (ALS)-inhibitor herbicides due to continuous and repeated selection is widespread in many troublesome weed species, including Palmer amaranth, throughout the United States. The objective of this research was to investigate the physiological and molecular basis of resistance to ALS inhibitors in a chlorsulfuron-resistant Palmer amaranth population (KSR). Our results indicate that the KSR population exhibits a high level of resistance to chlorsulfuron compared with two known susceptible populations, MSS and KSS from Mississippi and Kansas, respectively. MSS is highly susceptible to chlorsulfuron, whereas KSS is moderately sensitive. Dose–response analysis revealed that KSR was more than 275-fold more resistant compared with KSS. Nucleotide sequence analysis of theALSgene from the plants that survived chlorsulfuron treatment revealed the possibility of evolution of both target site–based and non–target site based resistance to ALS inhibitors in the KSR population. The most common mutation (Pro-197-Ser) in theALSgene associated with resistance to the sulfonylureas in many weed species was found only in 30% of the KSR population. A preliminary malathion study showed that the remaining 70% of resistant plants might have cytochrome P450–mediated non–target site resistance. This is the first report elucidating the mechanism of resistance to ALS inhibitors in Palmer amaranth from Kansas. Presence of both target site– and non–target site based mechanisms of resistance limits the herbicide options to manage Palmer amaranth in cropping systems.

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EPSPSGene Amplification Primarily Confers Glyphosate Resistance among Arkansas Palmer amaranth (Amaranthus palmeri) Populations

Weed Science ◽

10.1017/wsc.2017.83 ◽

2018 ◽

Vol 66 (3) ◽

pp. 293-300 ◽

Cited By ~ 5

Author(s):

Shilpa Singh ◽

Vijay Singh ◽

Amy Lawton-Rauh ◽

Muthukumar V. Bagavathiannan ◽

Nilda Roma-Burgos

Keyword(s):

Copy Number ◽

Gene Copy Number ◽

Glyphosate Resistance ◽

Resistance Mechanisms ◽

Palmer Amaranth ◽

Gene Copy ◽

Amaranthus Palmeri ◽

Resistance Level ◽

Relative Copy Number ◽

Gene Copies

AbstractResearch was conducted to determine whether resistance to glyphosate among Palmer amaranth (Amaranthus palmeriS. Watson) populations within the U.S. state of Arkansas was due solely to increasedEPSPSgene copy number and whether gene copy number is correlated with resistance level to glyphosate. One hundred and fifteenA. palmeriaccessions were treated with 840 g ae ha−1glyphosate. Twenty of these accessions, selected to represent a broad range of responses to glyphosate, underwent further testing. Seven of the accessions were controlled with this dose; the rest were resistant. The effective dose to cause 50% injury (ED50) for susceptible accessions ranged from 28 to 207 g ha−1. The glyphosate-resistant (GR) accessions had ED50values ranging from 494 to 1,355 g ha−1, a 3- to 48-fold resistance level compared with the susceptible standard (SS). The 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene relative copy number was determined for 20 accessions, 4 plants accession−1. Resistant plants from five GR accessions (38% of resistant plants tested) did not have increasedEPSPSgene copies. Resistant plants from the remaining eight GR accessions (62% of resistant plants tested) had 19 to 224 moreEPSPSgene copies than the SS. Among the accessions tested, injury declined 4% with every additionalEPSPScopy. ED50values were directly correlated withEPSPScopy number. The highly resistant accession MIS11-B had an ED50of 1,355 g ha−1and 150 gene copies. Partial sequences ofEPSPSfrom GR accessions withoutEPSPSamplification did not contain any of the known resistance-conferring mutations. Nearly 40% of GR accessions putatively harbor non–target site resistance mechanisms. Therefore, elevatedEPSPSgene copy number is associated with glyphosate resistance amongA. palmerifrom Arkansas.

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The use of random amplified polymorphic DNA to evaluate the genetic variability of Ponkan mandarin (Citrus reticulata Blanco) accessions

Genetics and Molecular Biology ◽

10.1590/s1415-47572000000100031 ◽

2000 ◽

Vol 23 (1) ◽

pp. 169-172 ◽

Cited By ~ 8

Author(s):

Helvécio Della Coletta Filho ◽

Marcos Antonio Machado ◽

M. Luiza P.N. Targon ◽

Jorgino Pompeu Jr.

Keyword(s):

Genetic Variability ◽

Rapd Analysis ◽

Random Amplified Polymorphic Dna ◽

Point Mutations ◽

The Other ◽

Citrus Reticulata ◽

Random Primers ◽

Citrus Reticulata Blanco ◽

Ponkan Mandarin

RAPD analysis of 19 Ponkan mandarin accessions was performed using 25 random primers. Of 112 amplification products selected, only 32 were polymorphic across five accessions. The absence of genetic variability among the other 14 accessions suggested that they were either clonal propagations with different local names, or that they had undetectable genetic variability, such as point mutations which cannot be detected by RAPD.

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Natural infection of phlebotomines (Diptera: Psychodidae) in a visceral-leishmaniasis focus in Mato Grosso do Sul, Brazil

Revista do Instituto de Medicina Tropical de São Paulo ◽

10.1590/s0036-46652007000200011 ◽

2007 ◽

Vol 49 (2) ◽

pp. 119-122 ◽

Cited By ~ 21

Author(s):

João Cezar do Nascimento ◽

Byanca Regina de Paiva ◽

Rosely dos Santos Malafronte ◽

Wedson Desidério Fernandes ◽

Eunice Aparecida Bianchi Galati

Keyword(s):

Visceral Leishmaniasis ◽

Natural Infection ◽

Leishmania Species ◽

Common Species ◽

The Other ◽

Lutzomyia Longipalpis ◽

Leishmania Chagasi ◽

Mato Grosso ◽

Negative Results ◽

Mato Grosso Do Sul

The main purpose of this study was to investigate natural infection by Leishmania in phlebotomine females in a visceral-leishmaniasis focus in Antonio João county in Mato Grosso do Sul State, Brazil. Between June and October 2003, the digestive tracts of 81 females captured in Aldeia Campestre, Aldeia Marangatu and Povoado Campestre were dissected. The females were separated by species, location, area and date of capture into 13 groups and kept in ethanol 70%. To identify the Leishmania species using the PCR technique, amplifications of the ribosomal-DNA (rDNA) and mini-exon genes were analyzed. Of the 81 specimens, 77 (95%) were Lutzomyia longipalpis, making this the most common species; only one specimen of each of the species Brumptomyia avellari, Evandromyia cortelezzii, Evandromyia lenti and Nyssomyia whitmani was found. Trypanosomatids were identified in eight of the nine groups of Lutzomyia longipalpis (10.39%) one group from Aldeia Campestre, one from Aldeia Marangatu and six from Povoado Campestre; of the eight groups, one from Aldeia Marangatu and another, with promastigotes forms also confirmed by dissection (1.23%) from Povoado Campestre, were identified by PCR as Leishmania chagasi (2.6%). The other groups gave negative results. These findings indicate that there is a high risk of leishmaniasis transmission in this area.

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