Strategy of Echinochloa oryzicola Vasing. for survival in flooded rice

Echinochloa oryzicola Vasing. (= Echinochloa phyllopogon Stapf ex Kessenko) is an obligate weed with an elaborated survival strategy in the flooded rice of Japan. In this review various adaptive characters of the weed, which comprise the survival strategy, are discussed through the life cycle. The weed is distributed only in flooded rice. Seeds (spikelets) buried in the soil exhibit annual cycles between dormant and non‐dormant state, and non‐dormant seeds recurrently appear in spring when rice growers start to prepare seedling beds and fields for rice transplanting. The non‐dormant seeds have unique characters metabolically adapted to submerged conditions to germinate and grow by the anaerobic respiration through alcohol fermentation. The weed has seemingly perfect mimicry of the rice plants throughout its development from seedling to heading, by which the weed escapes from manual weeding. In a rice paddy, the weed starts heading coincidentally with the rice plants at the period when the growers are reluctant to walk in the rice paddy to weed. Irrespective of plant height of the rice cultivar, the weed develops a few upper leaves above the rice canopy during the heading period of rice. This phenotypic plasticity of E. oryzicola in plant height is one of the characters conferring its competitive aggressiveness in flooded rice. When weeding is begun again after heading, the dormant weed seeds escape weeding by shattering and join the soil seedbank. The dormant seeds express the gene of an enzyme catalyzing ATP synthesis through the mitochondrial oxidative phosphorylation more abundantly, and have larger oxygen absorption and enzyme activity of the aerobic respiration than the non‐dormant seeds, suggesting that the dormant seeds maintain viability by the conventional aerobic respiration in the paddy soil drained from rice harvesting in fall to the next early spring. The various adaptive characters comprising the survival strategy of E. oryzicola in flooded rice consist of those inherited from the wild progenitor and those selected by the crop cultivation pressure. It is suggested that both the mimicry of the weed and the heading coincident with the rice plants have been acquired by the large selection pressure of frequent weeding, which has been done over the past hundred years. However, today, the manual weeding is substituted with herbicides, which cannot detect the mimicry and heading photoperiodic sensitivity. As a result, the dominant species of Echinochloa weeds in flooded rice is changing from E. oryzicola to Echinochloa crus‐galli var. crus‐galli that has neither mimicry nor photoperiodic sensitivity synchronizing to that of rice, but is more competitive against rice.     

Microsatellite variability of sulfonylurea‐resistant and susceptible populations of Schoenoplectus juncoides (Cyperaceae) in Kinki,Japan

Schoenoplectus juncoides is one of the most harmful weeds found in East Asian paddy fields. Recent emergence of biotypes that are resistant to the herbicide sulfonylurea (SU) has made weed control difficult. To examine the effect of the evolution of this herbicide resistance on genetic diversity within local populations, we investigated microsatellite variability within and among paddy field populations of S. juncoides in Kinki, Japan. In vivo assay of acetolactate synthase activity and root elongation assay in the presence of SU revealed that of 21 populations, five were sulfonylurea‐susceptible (SU‐S) and eight were completely sulfonylurea‐resistant (SU‐R). The remaining eight populations were a mixture of SU‐S and SU‐R individuals. The average gene diversity for SU‐R populations (H_S = 0.168) was lower than those for SU‐S (H_S = 0.256) and mixed (H_S = 0.209) populations, but the difference was not significant. This indicates that positive selection for SU‐R phenotype did not cause a genome‐wide reduction in genetic diversity. Genetic differentiation among S. juncoides populations was higher than that observed for most weed species studied previously. Although populations in neighbouring paddy fields showed a high level of differentiation, Bayesian clustering analyses suggested that some level of gene flow occurs among them and that the genetic exchange or colonisation between neighbouring populations could contribute to the geographical expansion of the resistant allele.     

Simple sequence repeat analysis of genetic diversity among Acetyl‐CoA carboxylase inhibitor‐resistant and ‐susceptible Echinochloa crus‐galli and E. oryzicola populations in Korea

Echinochloa species are amongst the most problematic weeds in rice fields of Korea. The steady reliance on the Acetyl‐CoA carboxylase (ACCase) and acetolactate synthase inhibiting herbicides for control of these weeds has led to resistance to these herbicides. The objective of this study was to assess the genetic diversity among populations of ACCase inhibitor‐resistant and ‐susceptible Echinochloa crus‐galli and E. oryzicola in Korea, to better understand their population structure and possible origins of resistance. Seven simple sequence repeat markers were applied to assess the genetic diversity between resistant and susceptible E. crus‐galli and E. oryzicola from 12 populations in Korea. Genetic diversity was slightly higher in the resistant group. The Unweighted Pair Group Method using Arithmetic algorithm (UPGMA) dendrogram generated two distinct clades. One clade consisted of Echinochloa spp. from three populations, i.e. Anmyeondo, Gimje 4 and Gongju, which are resistant and differentiated from the susceptible populations, and the other clade contained the rest of the populations. Structure modelling supported two clades of UPGMA clustering. Based on these data, we can infer that some resistant populations are greatly differentiated, whereas other resistant biotypes are still building up resistance in rice fields in Korea. Resistance traits will be fixed and continue to spread over time without proper control measures.     

Misidentifications of Echinochloa crus‐galli var. formosensis as Echinochloa oryzicola as a result of similar cpDNA sequences

Echinochloa crus‐galli (L.) Beauv. var. formosensis Ohwi (2n = 6x = 54, AABBCC genomes) and Echinochloa oryzicola (Vasinger) Vasinger (2n = 4x = 36, AABB) are major paddy weeds in East and Southeast Asia. E. oryzicola has been generally considered to be a paternal genome donor of E. crus‐galli s. l., which includes E. crus‐galli var. formosensis based on cpDNA sequences. Thus, molecular characterization using polymerase chain reaction‐restriction fragment length polymorphism analysis of cpDNA has been proposed as a reliable method for discriminating between the two species. In this study, we report that four accessions of E. crus‐galli var. formosensis from Okinawa, Nagasaki, Shizuoka and Tokyo had similar cpDNA sequences to E. oryzicola and had been misidentified as E. oryzicola using molecular methods. In addition, our results demonstrated that these accessions likely inherited their chloroplast genomes from E. oryzicola and not from an anonymous diploid species during polyploidization. Our findings provide new insights into the evolution of E. crus‐galli s. l. and suggest that identification using the cpDNA molecular method alone is not an appropriate approach to differentiate E. crus‐galli var. formosensis and E. oryzicola.     

Performance of Bangladesh indigenous rice in a weed infested field and separation of allelopathy from resource competition

This study aimed to identify the potential allelopathic indigenous rice (Oryza sativa L. ssp. indica) varieties from Bangladesh using a performance study in a weed‐infested field and to assess the extent of allelopathic interference relative to resource competition in a glasshouse experiment. Six varieties – namely, “Boterswar,” “Goria,” “Biron” and “Kartiksail” as the most allelopathic, “Hashikolmi” as weakly allelopathic and “Holoi” as nonallelopathic – were raised following a nonweed control method. The infestation levels of weed species were calculated using Simpson's Diversity Index (SDI), which ranged from 0.2 to 0.56. However, a significant correlation coefficient (0.87, P < 0.001) was obtained from these field data compared with the root inhibition percentage from the laboratory bioassay, and the “Boterswar” variety was the most allelopathic. The interactions between the allelopathic variety “Boterswar,” weakly allelopathic variety “Hashikolmi” and Echinochloa oryzicola via a target (rice)‐adjacent (E. oryzicola) cogrowth culture were determined in a hydroponic arrangement. The relative competitive intensity (RCI) and the relative neighbor effect (RNE) values showed that the crop–weed interaction was facilitation for “Boterswar” and competition for “Hashikolmi” and E. oryzicola in rice/E. oryzicola cogrowth cultures. The allelopathic effects of “Boterswar” were much higher than the resource competition in rice/E. oryzicola cogrowth cultures. The converse was observed for “Hashikolmi.” Moreover, the mineral content of E. oryzicola was severely affected by “Boterswar”/E. oryzicola cogrowth cultures’ exudate solution. Therefore, the allelopathic potential of “Boterswar” variety might be useful for developing the weed‐suppressing capacity of rice, which will likely have a significant influence on paddy weed control.     

Glyphosate‐resistant Italian ryegrass (Lolium multiflorum) on rice paddy levees in Japan

The rapid range expansion of naturalized Italian ryegrass (Lolium multiflorum Lam.) in farmland is a serious problem in Fukuroi city in Shizuoka Prefecture, Japan. Glyphosate has been used to control Italian ryegrass in the levees of rice paddy fields and wheat fields for ～20 years, but this weed in Fukuroi city is poorly controlled by glyphosate. In order to elucidate the level of resistance to glyphosate in Italian ryegrass populations, seed bioassays and a foliar application experiment, using seeds collected from 16 wild populations in and around Fukuroi city and from three susceptible cultivars, were conducted. For the susceptible cultivars and one population from a site where glyphosate had not been applied for >10 years, the shoot length in the seed bioassays was greatly suppressed at a glyphosate concentration of 10 mg ai L^?1 and no seedling survived after the foliar application of glyphosate at a rate of 2.3 kg ai ha^?1. Nine wild populations from levees in the southern part of Fukuroi city showed vigorous shoot growth at a glyphosate concentration of 10 mg ai L^?1 and had at least a 78% survival rate after the application of glyphosate at 2.3 kg ai ha^?1. Four wild populations from levees in the northern part of Fukuroi city showed a slight suppression of the shoot growth as a result of the glyphosate treatment and their survival rates ranged from 20 to 64%. The results suggested that resistance to glyphosate has evolved in the wild populations of Italian ryegrass that are growing on the levees. This is the first report of a glyphosate‐resistant weed in Japan.     

Effects of ferrous iron (Fe) on the germination and root elongation of paddy rice and weeds

Laboratory experiments were conducted to analyze the iron (Fe) tolerance of paddy weeds and rice varieties (Oryza sativa) for germination and root elongation. Under a waterlogged soil condition, the Fe(II) content in a soil solution increased with an increase in the ratio of rice straw to the soil. In the presence of 0.9% (w/w) straw to soil, which corresponds approximately to 8 t of straw applied to an area of 1 ha × 10 cm depth in the field, ～80 mg L^?1 of Fe(II) was produced in the soil solution. Based on this result, the seeds of rice and the weeds were incubated in a solution with <100 mg L^?1 of Fe(II). The presence of 100 mg L^?1 of Fe(II) suppressed the germination of Echinochloa crus‐galli var. crus‐galli, Cyperus serotinus, Cyperus difformis, and Monochoria korsakowii. However, it had no effect on the germination of Echinochloa oryzicola, Schoenoplectus juncoides (= Scirpus juncoides var. ohwianus), and Monochoria vaginalis. This level of Fe tolerance was the same as that of rice. These findings suggest that E. oryzicola, S. juncoides, and M. vaginalis can grow under more severe conditions than E. crus‐galli, C. serotinus, C. difformis, and M. korsakowii. In relation to seminal root elongation, the order of tolerance of Fe toxicity was O. sativa cv. Dunghan Shali > O. sativa cv. Hoshinoyume > E. oryzicola > M. vaginalis > S. juncoides. Thus, the results show that the tolerance of rice is greater than that of E. oryzicola, which had a comparatively strong tolerance among the weeds examined, and also that there are differences in tolerance among the rice varieties. These findings suggest that the difference in Fe tolerance is involved in weed control systems when organic materials are applied. If this difference is an important factor in the weed control system, Fe‐tolerant rice varieties, like cv. D. Shali, could facilitate weed control systems due to their higher Fe tolerance ability.     

Genetic relationship between Echinochloa crus‐galli and Echinochloa oryzicola accessions inferred from internal transcribed spacer and chloroplast DNA sequences

Barnyardgrass, hexaploid Echinochloa crus‐galli, is considered to arise from the hybridization between tetraploid Echinochloa oryzicola and an unknown diploid species. The genetic relationship between E. crus‐galli and E. oryzicola was examined to investigate the position of E. oryzicola in the evolutionary process of E. crus‐galli, based on the nuclear DNA internal transcribed spacer (ITS) and the chloroplast cpDNA trnT‐L, trnL intron, and trnL‐F regions. New World E. crus‐galli was clearly separated from Eurasian E. crus‐galli and showed a close relationship to the American taxa, Echinochloa crus‐pavonis and Echinochloa walteri, in both the ITS and chloroplast DNA. The nrDNA ITS sequences indicated no differentiation between the Eurasian E. crus‐galli and E. oryzicola, in contrast to their clear divergence in the cpDNA sequence. The present results suggest that E. oryzicola is the male donor of E. crus‐galli.     

Frequency and distribution of herbicide resistance in Raphanus raphanistrum populations randomly collected across the Western Australian wheatbelt

In 2003, a random survey was conducted across the Western Australian wheatbelt to establish the extent and frequency of herbicide resistance in Raphanus raphanistrum populations infesting crop fields. Five hundred cropping fields were visited, with 90 R. raphanistrum populations collected, representative of populations present in crop fields throughout the Western Australian wheatbelt. Collected populations were screened with four herbicides of various modes of action that are commonly used for the control of this weed. The majority of Western Australian R. raphanistrum populations were found to contain plants resistant to the acetolactate synthase (ALS)‐inhibiting herbicide chlorsulfuron (54%) and auxin analogue herbicide, 2,4‐D amine (60%). This survey also determined that over half (58%) of these populations were multiple resistant across at least two of the four herbicide modes of action used in the screening. Only 17% of R. raphanistrum populations have retained their initial status of susceptibility to all four herbicides. The distribution patterns of the herbicide‐resistant populations identified that there were higher frequencies of resistant and developing resistance populations occurring in the intensively cropped northern regions of the wheatbelt. These results clearly indicate that the reliance on herbicidal weed control in cropping systems based on reduced tillage and stubble retention will lead to higher frequencies of herbicide‐resistant weed populations. Therefore, within intensive crop production systems, there is a need to diversify weed management strategies and not rely entirely on too few herbicide control options.     

Effect of naringenin on the growth and lignin biosynthesis of gramineous plants

Lignin biosynthesis is essential for plant growth. 4‐Coumarate CoA ligase (4‐CL, EC6.2.1.12) is involved in the monolignol synthesis and occupies a key role in regulating carbon flow into the phenylpropanoid metabolism pathway. Naringenin, one of the metabolites in this pathway, is known as a potent in vitro inhibitor of 4‐CL. The growth of rice (Oryza sativa L. cv. Koshihikari), maize (Zea mays L. cv. Yellow corn) and Echinochloa oryzicola Vasing seedlings at the 2nd leaf stage was inhibited after continuous root application with 0.1 mmol L^?1 naringenin for 1 week, although naringenin did not kill these gramineous plants. The highest inhibition of fresh weight increase was observed in maize, followed by rice and E. oryzicola. The symptoms in these plants were root browning, delay of leaf/root development and shoot dwarfing. Naringenin treatment increased the contents of 4‐CL substrates, cinnamic acid, 4‐coumaric acid, caffeic acid and ferulic acid from 1.2 to 7.2 times and from 1.2 to 3.5 times in shoots and roots, respectively, except for ferulic acid in E. oryzicola roots. It also caused a slight decrease of the lignin content and alteration of lignin constitutions in rice plants. These results suggested that the monolignol pathways after 4‐CL towards lignin has the possibility to be the novel action sites of plant growth retardants, although further investigations are needed to clarify the mode of action.     

Control of <Emphasis Type="Italic">Verticillium dahliae</Emphasis> at a strawberry nursery by paddy-upland rotation

Effects of crop rotation between rice paddy fields and strawberry nurseries on the control of Verticillium wilt of strawberry were studied. For detecting Verticillium dahliae, the causal agent of Verticillium wilt, in soil, eggplant was used as an indicator plant. We were thus able to detect as low as 1 microsclerotium/g dry soil. In field surveys of Chiba and Hokkaido from 2000 to 2003, V. dahliae was detected in 9 of 10 upland fields but in none of 21 paddy-upland fields. In Hokkaido during 2000–2007, strawberry mother plants were planted, and plantlets were produced in upland and paddy-upland fields to assess V. dahliae infestation. Verticillium wilt of strawberry had never occurred in 72 tested paddy-upland fields, compared to 13.2–73.9% of plantlets infected with V. dahliae in upland fields. In a pot experiment in a greenhouse, two flooding treatments or two paddy rice cultivations suppressed Verticillium wilt symptoms on eggplant. In field experiments, one paddy rice cultivation in Chiba and two in Hokkaido prevented development of Verticillium wilt symptoms on eggplant. Verticillium wilt of strawberry was controlled completely with one paddy rice cultivation in infested fields in Chiba. In these field experiments, the number of microsclerotia of V. dahliae decreased under the flooding conditions for paddy rice cultivation. Based on the reduction in microsclerotia, a crop rotation system with paddy rice for 3 years (three times), green manure for 1 year, and strawberry nursery for 1 year was designed for Hokkaido.     

Cross‐resistance of Echinochloa species to acetolactate synthase inhibitor herbicides

This study was conducted to evaluate the cross‐resistance of acetolactate synthase (ALS) inhibitors with different chemistries, specifically azimsulfuron (sulfonylurea), penoxsulam (triazolopyrimidine sulfonanilide) and bispyribac‐sodium (pyrimidinyl thio benzoate), in Echinochloa oryzicola and Echinochloa crus‐galli that had been collected in South Korea and to investigate their herbicide resistance mechanism. Both Echinochloa spp. showed cross‐resistance to the ALS inhibitors belonging to the above three different chemistries. In a whole plant assay with herbicides alone, the resistant/susceptible ratios for azimsulfuron, penoxsulam and bispyribac‐sodium were 12.6, 28.1 and 1.9 in E. oryzicola and 21.1, 13.7 and 1.8 in E. crus‐galli, respectively. An in vitro ALS enzyme assay with herbicides showed that the I ₅₀‐values of the resistant accessions were approximately two‐to‐three times higher than the susceptible accessions, with no statistical difference, suggesting that the difference in ALS sensitivity cannot explain ALS inhibitor resistance in Echinochloa spp. for azimsulfuron, penoxsulam and bispyribac‐sodium. A whole plant assay with fenitrothion showed that the GR ₅₀‐values significantly decreased in both the resistant E. oryzicola and E. crus‐galli accessions when azimsulfuron, penoxsulam and bispyribac‐sodium were applied with the P450 inhibitor, while no significant decrease was observed in the susceptible accessions when the P450 inhibitor was used. Thus, these results suggest that ALS inhibitor cross‐resistance for azimsulfuron, penoxsulam and bispyribac‐sodium is related to enhanced herbicide metabolism.     

Occurrence of sulfonylurea resistance in Sagittaria trifolia,a basal monocot species,based on target‐site and non‐target‐site resistance

Sagittaria trifolia L. is one of the most serious weeds in paddy fields in Japan. Since the late 1990s, severe infestations of S. trifolia have occurred following applications of sulfonylurea herbicides in Akita prefecture. In this study, two accessions of S. trifolia, R1 and R2, were collected from paddy fields with severe infestations and their resistance profiles were determined in comparison to a susceptible accession, S1. R1 and R2 were highly resistant to bensulfuron‐methyl. R1 was also highly resistant to pyrazosulfuron‐ethyl, but R2 was susceptible. Relative to S1, R1 had an amino acid substitution at the Pro₁₉₇ residue of acetolactate synthase (ALS), a well‐known mutation that confers sulfonylurea resistance, suggesting that R1 has a target‐site‐based resistance (TSR) mechanism. The sequence of the ALS gene in R2 was identical to that in S1. A Southern blot analysis indicated that there was only one copy of the ALS gene in S1 and R2. These results suggest that R2 has a non‐target‐site‐based resistance (NTSR) mechanism. R2 was moderately resistant to imazosulfuron but susceptible to thifensulfuron‐methyl. R2 and S1 were susceptible to pretilachlor, benfuresate, MCPA‐ethyl and bentazon. The results reveal the occurrence of two sulfonylurea‐resistant biotypes of S. trifolia that show different mechanisms of cross‐resistance to sulfonylureas related to TSR in R1 and NTSR in R2.     

Micro‐elevations in paddy fields affect the efficacy of mechanical weeding: Evaluation of weeding machines to control Monochoria vaginalis in herbicide‐free farming

Differences in local topography (micro‐elevation) within a paddy field that constitute a source of variability in agronomical indicators have not been considered thoroughly as a block factor in weed studies. This study investigated and evaluated the performance of weeding machines (weeders) in two herbicide‐free paddy fields that contained micro‐elevations. The plant density of Monochoria vaginalis, a typical and harmful paddy weed unless controlled with herbicides, was used as the indicator of the efficacy of the weeders. Among the three weeders that were tested, one suppressed M. vaginalis dramatically at low elevations and the others were less sensitive to micro‐elevation. For comparison across the fields, micro‐elevations at weed sampling locations were converted to the initial depth of water by using the records of hydrographs that had been set in each field. The relationship between the initial depth of water and the plant density of M. vaginalis was very clear with the use of the elevation‐sensitive weeder. Moreover, this relationship was valid, even with the less‐sensitive weeders. The finding that the greater the depth of water, the less the plant density was significant, even for M. vaginalis, a difficult aquatic paddy weed that was controlled with any of the weeders tested. Thus, micro‐elevation within a paddy field needs to be treated as a crucial block factor in weed‐sampling studies. A coarse survey of the level of a field and the installation of a hydrograph are recommended for a clear analysis of the background of weed control practices.     

Influence of weed management measures on glyphosate resistance and endophyte infection in naturalized Italian ryegrass (Lolium multiflorum)

Recently, glyphosate‐resistant Italian ryegrass (Lolium multiflorum Lam.) was found on rice paddy levees in a western region of Shizuoka Prefecture, Japan. Naturalized populations of Italian ryegrass are frequently infected with fungal Epichloë endophytes. Endophytes often confer benefits to their host grasses. This study investigated the influence of five weed management treatments on glyphosate resistance and endophyte infection in Italian ryegrass that was growing on paddy levees where glyphosate‐resistant individuals were dominant. The weed management treatments were: (i) mowing once before the grass flowered; (ii) mowing once during flowering; (iii) mowing twice during flowering; (iv) glyphosate application before flowering; and (v) no treatment. The seeds were collected from the treatment plots in 2013 and 2014. The seeds were examined for endophyte infection and the seedlings that had been grown from the seeds were tested for the frequency of glyphosate resistance. The seedlings that had been derived from the glyphosate treatment showed higher frequencies of glyphosate resistance than those seedlings that had been derived from all the other treatments. Endophytes were found in all populations of the seeds from the paddy levees, with higher infection rates in the seeds that had been derived from the glyphosate treatment and the twice‐mowed treatment. There was a significant relationship between the endophyte infection frequency in the seeds and glyphosate resistance in the seedlings that had been grown from the same populations. The results indicate that where glyphosate herbicides are frequently used, selection for glyphosate‐resistant Italian ryegrass occurs, and along with this, the frequency of endophyte infection also increases.     

Scanning electron microscopy of the establishment of compatible and incompatible Xanthomonas campestris pathovars on the leaf surface of Italian ryegrass and maize1

Scanning electron-microscopy observations made on the leaves of Italian ryegrass and maize, inoculated with Xanthomonas campestris pvs graminis, oryzae and oryzicola clearly showed a difference in the distribution pattern among the different pathovars tested. Pvs oryzae and oryzicola could be detected on the leaf trichomes of ryegrass, while pv. graminis was not. On maize leaves, the attachment of pvs oryzae and oryzicola to trichomes was much more pronounced than on ryegrass. In addition, pv. oryzae spread further into leaves of maize than did pvs graminis and oryzicola and could be detected in masses until at least the sixth day after inoculation. These observations suggest that non-host plants such as maize could function as alternate inoculum sources of pv. oryzae for nearby rice plants.     

Improved physical control of glyphosate‐resistant Italian ryegrass (Lolium multiflorum) on rice paddy levees in Japan

Italian ryegrass ( Lolium multiflorum Lam.) is a non‐native annual winter grass that has seriously infested rice paddy levees and wheat fields in Japan. Recently, glyphosate‐resistant Italian ryegrass was found on paddy levees in central Japan, thereby making control of the grass by using glyphosate less effective. In this study, physical control methods were tested that combined the timing and frequency of mowing in order to more effectively control glyphosate‐resistant Italian ryegrass on rice paddy levees. A 3 year field experiment was conducted from 2012 to 2014 in a western region of Shizuoka Prefecture, where glyphosate‐resistant Italian ryegrass has become dominant. Five treatments were tested: (i) mowing once before the flowering of the grass (i.e. conventional mowing measure); (ii) mowing once during flowering; (iii) mowing twice during flowering; (iv) glyphosate application before flowering (i.e. one of the conventional mowing measures); and (v) no treatment. The above‐ground biomass, seed production, soil seed bank and seedling occurrence of Italian ryegrass were measured to determine the effectiveness of these treatments. Mowing during the flowering period resulted in reduced above‐ground biomass, seed production and soil seed bank when compared with the other treatments. Additionally, mowing twice during the flowering period resulted in a lower seedling density than mowing once. The results suggest that, in this region, physical control by mowing during the flowering period would be more effective than conventional measures for controlling glyphosate‐resistant Italian ryegrass.     

Varied occurrence of diverse sulfonylurea‐resistant biotypes of Schoenoplectus juncoides [Roxb.] Palla in Japan,as classified by an acetolactate synthase gene mutation

Suspected sulfonylurea (SU)‐resistant Schoenoplectus juncoides plants were collected from rice paddy fields at 24 sites in Japan in order to discover the occurrence pattern of target‐site substitutions on a nationwide scale and at a local field scale. A genetic analysis of the two acetolactate synthase (ALS) genes, ALS1 and ALS2, of the collected plants confirmed that a single‐nucleotide mutation at the Pro₁₉₇, Asp₃₇₆ or Trp₅₇₄ site of either ALS1 or ALS2 existed in each suspected SU‐resistant plant. On a nationwide scale, it was shown that the ALS1 mutations and the ALS2 mutations occurred at a similar frequency, that the P197S and the P197L substitutions were found most frequently among all the substitutions, and that the W574L substitutions (known as global resistance to any ALS‐inhibiting herbicide) were found at a relatively low frequency but in a geographically wide range. In the local field‐scale survey, which was conducted at two sites in Hyogo Prefecture, it was shown that the substitutions were less diverse, compared to on a nationwide scale, probably because the investigation involved a limited number of local fields, and that several substitutions and a susceptible biotype were found in single fields suggesting that a number of collections is required in order to understand the local SU‐resistant status of S. juncoides. In addition, this study reported new findings, that of the P197R, P197T and D376E substitutions in S. juncoides. This set of diverse substitutions in a weed species can be used for further research purposes.     

Current situation with invasive Erigeron annuus (l.) Pers. (daisy fleabane) in the Republic of Macedonia

Detailed surveys of populations of Erigeron annuus (L.) Pers., an invasive plant species native to Eastern North America, were made in three locations in north‐western, northern and eastern parts of the Republic of Macedonia. The population densities were not quantified, but several stands of different sizes were found. A rapid ecological risk assessment, mainly based on knowledge of invasion histories in South‐Eastern and Central European countries, showed that this species is a serious threat to Macedonian biodiversity, particularly in the north‐western part, where monospecific stands of E. annuus were recorded. Biological invasions of E. annuus affect biodiversity worldwide through its rapid growth and high seed production, phenotypic plasticity in the native range with regard to the availability of soil nutrients and release of compounds to the soil over the period of plant growth. Consequently, invaded ecosystems suffer from significant loss in economic and cultural value.     

PCR-SSCP analysis of <Emphasis Type="Italic">Fusarium</Emphasis> diversity in asparagus decline in Japan

The diversity of Fusarium populations in asparagus (Asparagus officinalis L.) decline fields in Japan was estimated by PCR-SSCP (single-stranded conformational polymorphism) analysis of the ITS2 regions of the nuclear rRNA genes. This method was used to rapidly and objectively identify pathogens associated with roots of plants showing symptoms of asparagus decline collected from fields in five regions across Japan. Over 651 fusarial isolates were obtained, and were easily differentiated into three principal species. Fusarium oxysporum f. sp. asparagi was most frequently isolated from the domestic five regions (68%), whereas Fusarium proliferatum (28.6%) was less frequent. Fusarium solani was found much rarely (2.5%). The frequency of isolation of Fusarium proliferatum increased gradually from the north to the south of Japan, though considerable differences were found between fields in each region, as well as regional differences among the Fusarium populations. Most of the fusarial isolates were highly pathogenic in vitro. These results reveal that Fusarium oxysporum f. sp. asparagi and Fusarium proliferatum are important biotic factors which lead to asparagus decline in Japan.