Albert J. Fischer

Herbicide-Resistant Echinochloa oryzoides and E. phyllopogon in California Oryza sativa Fields

Abstract Echinochloa oryzoides and E. phyllopogon have become the most serious weeds in California Oryza sativa since continuous flooding was used to suppress E. crus-galli. Continuous use of a limited number of available graminicides and an increasing number of control failures led to the investigation of herbicide resistance in E. oryzoides and E. phyllopogon. Greenhouse dose-response studies with postemergence (POST) applications of molinate, thiobencarb, fenoxaprop-ethyl, and bispyribac-sodium estimating GR50 (herbicide dose to inhibit growth by 50%) values suggested resistance to all herbicides in two E. phyllopogon accessions and to molinate and thiobencarb in one E. oryzoides accession when compared with susceptible E. phyllopogon and E. oryzoides controls, respectively. No resistance was detected in dose-response studies with propanil. Minimum and maximum ratios (R/S) of the GR50 values of resistant to susceptible E. phyllopogon plants (in two experiments involving two resistant accessions) were 7.8 and >13.3 for thiobencarb, 2.2 and 4.3 for molinate, 16.5 and 428.7 for fenoxaprop-ethyl, and 2.0 and 12.0 for bispyribac-sodium. Minimum and maximum E. oryzoides R/S ratios (average of two experiments) were 21.9 and 4.6 for thiobencarb and molinate, respectively. A resistant E. phyllopogon (one accession tested) and the susceptible control were killed by POST applications of glyphosate, glufosinate, and clomazone, and by a preemergence application of pendimethalin. Thus, the repeated use of the few available grass herbicides in the predominantly monocultured O. sativa of California has selected for herbicide resistance in E. oryzoides and E. phyllopogon. The introduction of herbicides with new mechanisms of action will be useful to manage herbicide-resistant E. oryzoides and E. phyllopogon. However, cross- and multiple resistance emphasize the need to integrate herbicide use with nonchemical means of weed management. Nomenclature: Bispyribac-sodium, sodium 2,6-bis[(4,6-dimethoxypyrimidin-2-yl)oxy]benzoate; clomazone; fenoxaprop-ethyl; glyphosate; glufosinate; molinate; pendimethalin; propanil; thiobencarb; Echinochloa oryzoides (Ard.) Fritsch ECHOR, early watergrass; Echinochloa phyllopogon (Stapf) Koss ECHPH, late watergrass; Echinochloa crus-galli (L.) Beauv. var. crus-galli ECHCG, barnyardgrass; Oryza sativa L., rice.

Crop traits related to weed suppression in water-seeded rice (Oryza sativa L.)

Abstract Resistance to herbicides and the lack of viable control options have led to an interest in increasing the role of crop competition as a weed management tool in water-seeded rice production. Weed-suppressive rice cultivars have been suggested as a tool that could improve weed control and reduce the reliance of growers on herbicides. Field studies were conducted at Biggs, CA, in 1999 and 2000 with six to eight semidwarf rice cultivars to identify water-seeded rice traits related to the suppression of watergrass growth. Cultivars S-201 and M-302 were the most suppressive in both years. The dry weight (DW) of watergrass grown with the most suppressive cultivar was only 16% in 1999 and 57% in 2000 of the DW of watergrass grown with the least suppressive cultivar. Rice leaf area and root DW in weed-free plots were linearly related to watergrass DW in both years. Weed-suppressive traits were not inversely correlated with rice yields in monoculture; competitive cultivars also had high yields. This study suggests that an indirect selection program, based on traits that can be identified early in the season under weed-free conditions, has great potential for developing more competitive cultivars for water-seeded rice. Nomenclature: Early watergrass, Echinochloa oryzoides (Ard.) Fritsch ECHOR; late watergrass, Echinochloa phyllopogon (Stapf) Koss. ECHPH; rice, Oryza sativa L.

Cross-resistance to bispyribac-sodium and bensulfuron-methyl in Echinochloa phyllopogon and Cyperus difformis

Abstract Water-seeded and continuously flooded rice in California is mostly grown as a continuous crop and weeds are the most serious rice production problem. Echinochloa phyllopogon (Stapf) Koss and Cyperus difformis L. are adapted to this aquatic system and compete with rice, causing heavy economic losses. Flooding cannot fully suppress these weeds. Heavy reliance on few available herbicides resulted in the evolution of herbicide resistance in populations of both weeds in California rice fields. Resistance to bensulfuron-methyl (ALS inhibitor) is widespread among C. difformis populations. E. phyllopogon has evolved resistance to several herbicides, including bispyribac-sodium (ALS inhibitor), which has not yet been commercially used. A resistant (R) E. phyllopogon was also much more tolerant to bensulfuron-methyl than a susceptible (S) biotype. Understanding the patterns and mechanisms of cross-resistance in key weeds of rice to bensulfuron-methyl and bispyribac-sodium is relevant for the successful deployment of this new herbicide and for the management of herbicide resistance in California rice. Whole-plant bioassays were conducted to compare responses between E. phyllopogon and C. difformis to bispyribac-sodium and bensulfuron-methyl and to detect the involvement of cyt P-450 monooxygenases in E. phyllopogon resistance to bensulfuron-methyl using the cyt P-450 inhibitors piperonyl butoxide and malathion (previous studies had already shown cyt P-450-mediated resistance to bispyribac-sodium). ALS activity was assayed on leaf extracts from young R and S plants of both species for a range of bispyribac-sodium and bensulfuron-methyl concentrations. The dose–response studies confirmed cross-resistance in R E. phyllopogon ; minimum and maximum ratios (R/S) of the GR50 values of resistant to susceptible plants were 9 and >25.5 for bispyribac-sodium and bensulfuron-methyl, respectively. cyt P-450 contributed to bensulfuron-methyl resistance in R E. phyllopogon. C. difformis was also cross-resistant (R/S ratios: >10 for bispyribac-sodium and >26 for bensulfuron-methyl). ALS assays demonstrated that, unlike R E. phyllopogon, cross-resistance in R C. difformis was due to reduced ALS sensitivity. C. difformis ALS was more sensitive to bispyribac-sodium ( I 50 =138.87 nM ) than to bensulfuron-methyl ( I 50 =6724.56 nM ) . Also, ALS inhibition in R and S E. phyllopogon was higher with bispyribac-sodium. Thus, binding differences between both herbicides at the target site are suggested. This study reveals that cross-resistance between bensulfuron-methyl and bispyribac-sodium in both weeds involves degradation enhancement through monooxygenases and target-site alteration.

Relating rice traits to weed competitiveness and yield: a path analysis

Abstract Resistance to herbicides in the most important weeds threatens the sustainability of California rice. Weed-competitive rice cultivars could be a low-cost and safe nonchemical addition to an integrated weed management program. Trade-offs between competitiveness and productivity and inconsistent trait expression under weedy and weed-free conditions could complicate the breeding of competitive rice cultivars. A 2-year competition experiment was conducted in the greenhouse involving eight rice cultivars and two weed competition regimes (presence or absence of late watergrass) to examine the effects of rice weed-suppressive ability and tolerance to weed competition (weed tolerance) on rice yield. Competition reduced average rice yield from 32 to 48%, and watergrass biomass from 44 to 77%. Path analysis suggested that enhancing rice weed-suppressive ability and weed tolerance while minimizing possible productivity trade-offs should promote early (12 d after seeding) growth and light-capture traits followed by moderate growth rates before heading and a vigorous grain filling period. Crop growth rate (CGR) after heading was a relevant determinant of yield (direct path: 0.82, P < 0.01) and correlated (r = 0.30, P < 0.01) with weed tolerance. Late biomass accumulation was negatively correlated with harvest index and CGR during ripening (r = −0.46, P < 0.01); thus, late-season competitiveness can lower productivity. Rice traits conferring competitiveness were correlated across weed competition regimes (r = 0.36–0.81, P < 0.01). However, significant cultivar-by-competition and cultivar-by-year interactions suggest that selection efficiency would be greater when traits are identified under competition and in different environments. This study relates to the phenotypic expression of traits for competitiveness. Breeding competitive cultivars will require additional knowledge on trait heritability, genetic correlations with competitiveness, and on the effects of the environment upon gene expression. Nomenclature: Late watergrass, Echinochloa phyllopogon (Stapf) Koss., ECHPH; rice, Oryza sativa L.

Mechanism of resistance to penoxsulam in late watergrass [Echinochloa phyllopogon (Stapf) Koss.].

Late watergrass [ Echinochloa phyllopogon (Stapf.) Koss.] is a major weed of California rice that has evolved P450-mediated metabolic resistance to multiple herbicides. Resistant (R) populations are also poorly controlled by the recently introduced herbicide penoxsulam. Ratios (R/S) of the R to susceptible (S) GR(50) (herbicide rate for 50% growth reduction) ranged from 5 to 9. Although specific acetolactate synthase (ALS) activity was 1.7 higher in R than in S plants, the enzyme in R plants was about 6 times more susceptible to the herbicide. R plants exhibited faster (2.8 times) oxidative [(14)C]-penoxsulam metabolism than S plants 24 h after treatment. Addition of malathion (P450 inhibitor) enhanced herbicide phytotoxicity and reduced penoxsulam metabolism in R plants. Tank mixtures with thiobencarb (can induce P450) antagonized penoxsulam toxicity in R plants, suggesting penoxsulam may be broken down by a thiobencarb-inducible enzyme. These results suggest E. phyllopogon resistance to penoxsulam is mostly due to enhanced herbicide metabolism, possibly via P450 monooxidation.

Resistance to Glyphosate in Junglerice (Echinochloa colona) from California

Abstract A suspected glyphosate-resistant (R) junglerice population was collected from a glyphosate-R corn field near Durham in northern California where glyphosate had been applied at least twice a year for over 6 yr. Based on the amount of glyphosate required to reduce growth by 50% (ED50), the R population was 6.6 times more R than the susceptible (S) standard population. Based on the glyphosate concentration that inhibits EPSPS by 50% based on shikimate accumulation (I50) in leaf discs, R plants were four times more R than S plants. By 3 d after treatment with 0.42 kg ae ha−1 glyphosate, the S population had accumulated approximately five times more shikimate than the R population. No differences in [14C]-glyphosate uptake and translocation were detected between R and S plants. However, partial sequencing of the EPSPS gene revealed a mutation in R plants causing a proline to serine change at EPSPS position 106 (P106S). Our results reveal the first case of a P106S target site mutation associated with glyphosate resistance in junglerice. Nomenclature: Glyphosate; junglerice, Echinochloa colona (L.) Link.

Responses to clomazone and 5-ketoclomazone by Echinochloa phyllopogon resistant to multiple herbicides in Californian rice fields†

BACKGROUND Late watergrass [Echinochloa phyllopogon (Stapf.) Koss.] is a major weed of Californian rice that has evolved P450-mediated metabolic resistance to multiple herbicides. Resistant (R) populations are also poorly controlled by the recently introduced herbicide clomazone. The authors assessed whether this cross-resistance was also P450 mediated, and whether R plants also had reduced sensitivity to photooxidation. Understanding mechanism(s) of resistance facilitates the design of herbicide management strategies to delay resistance evolution. RESULTS Ratios (R/S) of R to susceptible (S) GR(50) were near 2.0. [(14)C]Clomazone uptake was similar in R and S plants. Clomazone and its metabolite 5-ketoclomazone reduced chlorophyll and carotenoids in S more than in R plants. The P450 inhibitors disulfoton and 1-aminobenzo-triazole (ABT) safened clomazone in R and S plants. Disulfoton safened 5-ketoclomazone only in S plants, while ABT synergized 5-ketoclomazone mostly against S plants. Paraquat was more toxic in S than in R plants. CONCLUSION Cross-resistance to clomazone explains failures to control R plants in rice fields, and safening by P450 inhibitors suggests that oxidative activation of clomazone is needed for toxicity to E. phyllopogon. Clomazone resistance requires mitigation of 5-ketoclomazone toxicity, but P450 detoxification may not significantly confer resistance, as P450 inhibitors poorly synergized 5-ketoclopmazone in R plants. Responses to paraquat suggest research on mechanisms to mitigate photooxidation in R and S plants is needed.

Concerted action of target‐site mutations and high EPSPS activity in glyphosate‐resistant junglerice (Echinochloa colona) from California

BACKGROUND Echinochloa colona is an annual weed affecting field crops and orchards in California. An E. colona population carrying a mutation in the EPSPS gene endowing resistance to glyphosate, the most widely used non-selective herbicide, was recently identified in the Northern Sacramento Valley of California. Plants from this population, from a suspected glyphosate-resistant (GR) population, and from one susceptible (S) population collected in the Northern Sacramento Valley of California, were used to generate three GR and one S selfed lines to study possible mechanisms involved in glyphosate resistance. RESULTS Based on the amount of glyphosate required to kill 50% of the plants (LD50 ), GR lines were 4-9-fold more resistant than S plants and accumulated less shikimate after glyphosate treatment. GR and S lines did not differ in glyphosate absorption, translocation or metabolism. A different target-site mutation was found in each of two of the GR lines corresponding to Pro106Thr and Pro106Ser substitutions; the mutations were found in different homoeologous EPSPS genes. No mutation was found in the third GR line, which exhibited 1.4-fold higher basal EPSPS activity and a fivefold greater LD50 than S plants. Quantitative RT-PCR revealed that GR lines had similar or lower EPSPS expression than S plants. CONCLUSION It is demonstrated that individuals with different glyphosate resistance mechanisms can coexist in the same population, individuals from different populations may carry different resistance mechanisms and different mechanisms can act in concert within single E. colona plants. However, other plant factors or resistance mechanisms appear to modulate plant expression of EPSPS sensitivity to glyphosate.

Cross-Resistance to Herbicides of Five ALS-Inhibiting Groups and Sequencing of the ALS Gene in Cyperus difformis L.

Resistance to ALS-inhibiting herbicides in Cyperus difformis has evolved rapidly in many rice areas worldwide. This study identified the mechanism of resistance, assessed cross-resistance patterns to all five chemical groups of ALS-inhibiting herbicides in four C. difformis biotypes, and attempted to sequence the ALS gene. Whole-plant and ALS enzyme activity dose-response assays indicated that the WA biotype was resistant to all ALS-inhibiting herbicides evaluated. The IR biotype was resistant to bensulfuron-methyl, orthosulfamuron, imazethapyr, and propoxycarbazone-sodium and less resistant to bispyribac-sodium and halosulfuron-methyl, and susceptible to penoxsulam. ALS enzyme activity assays indicated that resistance is due to an altered target site yet mutations previously found to endow target-site resistance in weeds were not detected in the sequences obtained. The inability to detect resistance mutations in C. difformis may result from the presence of additional ALS genes, which were not amplified by the primers used. This study reports the first ALS gene sequence from Cyperus difformis. Certain ALS-inhibiting herbicides can still be used to control some resistant C. difformis biotypes. However, because cross-resistance to all five classes of ALS-inhibitors was detected in other resistant biotypes, these herbicides should only be used within an integrated weed management program designed to delay the evolution of herbicide resistance.

Herbicide-resistant late watergrass (Echinochloa phyllopogon): similarity in morphological and amplified fragment length polymorphism traits

Abstract Late watergrass is a serious weed of California rice that has evolved resistance to molinate, thiobencarb, fenoxaprop-ethyl, and bispyribac-sodium. To obtain an insight into the origin and spread of resistant (R) late watergrass in California rice fields, we evaluated similarities in morphological traits and amplified fragment length polymorphism (AFLP) fingerprints among 15 R strains compared with susceptible (S) strains. All strains were derived by inbreeding from accessions collected in rice fields of the Sacramento Valley, CA. In the field, R plants were shorter than S plants; they also had narrower and shorter flag leaves and thinner culms. Spikelets also appeared smaller and more slender in R plants. There was greater morphological similarity among the 15 R strains than among the eight S strains. The mean coefficients of variation for morphological traits were much smaller among R strains, which in a cluster analysis (Wards method) were grouped morphologically apart at early clustering stages from the more variable S strains. AFLP electropherograms also showed greater similarity between R strains. R strains were grouped separately from the S strains in a cluster analysis based on calculated Nei and Li coefficients used in an unweighted pair group method using arithmetic means. However, small genetic differences also existed because the R strains were grouped into six clusters, suggesting that R strains were not samples from an identical strain. It was concluded that R strains originated from a preexisting and preadapted mutant late watergrass population in the Sacramento Valley. This study establishes that resistance moved by spikelet dispersal, not independent mutation events, most likely defined the geographical distribution of R late watergrass in California. Prevention and control of this dispersal combined with elimination of seed-producing survivors after herbicide treatment should be relevant components of the integrated management of herbicide-resistant late watergrass in California rice. Nomenclature: Bispyribac-sodium, sodium 2,6-bis[(4,6-dimethoxypyrimidin-2-yl)oxy]benzoate; fenoxaprop-ethyl; molinate; thiobencarb; early watergrass, Echinochloa oryzicola Vasing; late watergrass, Echinochloa phyllopogon (Stapf) Koss ECHPH; rice, Oryza sativa L.