Tohru Tominaga

Inheritance of Seed Shattering in Lolium temulentum and L. persicum Hybrids

Lolium temulentum and L. persicum are non-crop species found in wheat and barley fields. L. temulentum has non-shattering seeds like the associated grain crops, whereas L. persicum seeds shatter after maturity. We analyzed the inheritance mode of shattering tendency by comparing the F2 of L. temulentum and L. persicum hybrids. The selfed progeny of L. temulentum and L. persicum exhibited typical non-shattering (1.6% shattering) and shattering phenotypes (70.8%), respectively. F1 hybrids of L. temulentum×L. persicum and its reciprocals were of the shattering phenotype (71.4% and 63.8%, respectively), indicating that shattering is dominant to non-shattering. When the phenotype ratio was assumed to be 15 shattering: 1 non-shattering, the χ2 value for F2 segregation was not significant at the 5% level, and the reciprocal effect was not detected. This indicates that the non-shattering tendency is controlled by two recessive genes. The two-gene inheritance model of shattering tendency suggests that harvest is the selector for seed shattering in cultivated fields, thus the alternative tendency for non-shattering seeds of L. temulentum or shattering of L. persicum would be better adapted to cultivated fields.

Copy Number Variation in Acetolactate Synthase Genes of Thifensulfuron-Methyl Resistant Alopecurus aequalis (Shortawn Foxtail) Accessions in Japan

Severe infestations of Alopecurus aequalis (shortawn foxtail), a noxious weed in wheat and barley cropping systems in Japan, can occur even after application of thifensulfuron-methyl, a sulfonylurea (SU) herbicide. In the present study, nine accessions of A. aequalis growing in a single wheat field were tested for sensitivity to thifensulfuron-methyl. Seven of the nine accessions survived application of standard field rates of thifensulfuron-methyl, indicating that severe infestations likely result from herbicide resistance. Acetolactate synthase (ALS) is the target enzyme of SU herbicides. Full-length genes encoding ALS were therefore isolated to determine the mechanism of SU resistance. As a result, differences in ALS gene copy numbers among accessions were revealed. Two copies, ALS1 and ALS2, were conserved in all accessions, while some carried two additional copies, ALS3 and ALS4. A single-base deletion in ALS3 and ALS4 further indicated that they represent pseudogenes. No differences in ploidy level were observed between accessions with two or four copies of the ALS gene, suggesting that copy number varies. Resistant plants were found to carry a mutation in either the ALS1 or ALS2 gene, with all mutations causing an amino acid substitution at the Pro197 residue, which is known to confer SU resistance. Transcription of each ALS gene copy was confirmed by reverse transcription PCR, supporting involvement of these mutations in SU resistance. The information on the copy number and full-length sequences of ALS genes in A. aequalis will aid future analysis of the mechanism of resistance.

Genetic Diversity of Darnel (Lolium Temulentum L.) in Malo, Ethiopia Depends on Traditional Farming Systems

Genetic Diversity of Darnel (Lolium temulentum L.) in Malo, Ethiopia Depends on Traditional Farming Systems. Economic Botany 58(4):568–577, 2004. Darnel (Lolium temulentum L.) is a mimic weed associated with wheat and barley cultivation. Mimic weeds have evolved along with cultivated crops. Human impact on the genetic diversity of agricultural weeds was elucidated using darnel as a model. Three strains in each field in Gaitza village, Malo region, Ethiopia were randomly sampled, and the genetic composition of 120 samples was analyzed using RAPD analysis. Genetic diversity and distances were calculated using Nei’s analysis, and cluster analysis was performed. These results indicated the occurrence of seed migration among the fields and a high genetic diversity of darnel in this region. Crop seed exchange and contamination of crop seeds with darnel seeds lead to the unintended artificial gene flow of darnel. Because darnel seeds and seedlings resemble their associated crop seeds and seedlings, traditional methods of crop cultivation have the unintended consequence of conserving the genetic diversity of darnel.

Evaluation of Genetic Variation in High Molecular Weight Glutenin Subunits of Seed Storage Protein Using Landraces of Common Wheat from Pakistan

SDS-PAGE was used to determine the variation in high molecular weight glutenin subunits (HMW-GS) alleles present in 170 landraces of common wheat from Sindh, Balochistan and NWFP in Pakistan. Three alleles for Glu-A1, six alleles for Glu-B1 and four alleles for Glu-D1 were detected. These included three new alleles, namely Glu-B1e, Glu-B1h and Glu-D1n, which had not previously been found in landraces of Pakistani wheat. Furthermore, Glu-D1m and Glu-D1n, which are considered to be endemic alleles in Pakistan and Afghanistan, were both detected from Balochistan. Twenty genotypes were identified based on combinations of alleles at the three Glu-1 loci. The present study showed that Pakistani landraces of common wheat have maintained a broad diversity of HMW-GS alleles, and that they may serve as genetic resources to improve wheat varieties adapted to Pakistani climatic and edaphic conditions.

Crop-Associated Weeds

Some weeds are always associated with a particular crop and inhabit only the cropland, rarely expanding into other habitats. They have adapted to the habitats which man disturbs for the crop cultivation. Such weeds are the product of evolution under the specific selection pressures imposed by the crop cultivation over many years. Over time, they developed strategies adapted to the cultural practices or environments encountered throughout their life cycles that enable them to survive and even dominate there for generations. These cultural practices include field operations such as hand-weeding, but also crop harvest, threshing, and winnowing.

Isolation and characterization of polymorphic microsatellite loci for Imperata cylindrica, an invasive perennial grass

Cogongrass (Imperata cylindrica) is an invasive perennial grass widely distributed in the world, and two ecotypes (C-type and E-type) are found in Japan. We isolated and characterized eight microsatellite loci from the two ecotypes. The number of alleles ranged from 4 to 12 in C-type, 1 to 7 in E-type and 4 to 12 in the hybrid between them. The range of expected heterozygosities was 0.561–0.910 in C-type, 0.000–0.823 in E-type and 0.541–0.918 in the hybrid. These markers will be useful for investigating expansion process, natural hybridization between two ecotypes and reproductive strategy of this weed.

Preliminary Observations of Insect Visitation to Flowers of Vincetoxicum pycnostelma (Apocynaceae: Asclepiadoideae), an Endangered Species in Japan

Abstract Vincetoxicum pycnostelma Kitag. (Apocynaceae) is a perennial herb that grows in grassland habitats in East Asia and the western part of Amur. Urbanization, land development, abandonment of traditional management of grasslands, or any combination of these factors are rapidly reducing grassland habitats, and V. pycnostelma is now ranked as a “Near Threatened” species on the Red List of Japan. Identification and characterization of pollinators of this endangered species are important in its conservation. We observed and collected insect visitors to the flowers at night and in the morning in the populations of V. pycnostelma on the banks of the Kidzu River, Kyoto, Japan from June to August in 2011. Pollinators were characterized as those insects on which flower pollinaria were attached. Twenty-seven species (9 families; 3 orders) were observed visiting the flowers; individuals of 14 lepidopteran species (6 families) had pollinaria attached to their bodies and most visited the flowers at night. Adult lepidopterans with a proboscis length of 0.9 - 4.5 mm were the primary carriers of pollinaria. Examination of stigmatic chambers of randomly selected flowers showed that the percentage of flowers in which pollinaria was inserted was much higher than the percentage of fruit set observed. Reasons for the observed low fruit set may be self-incompatibility or resource limitation. These preliminary observations were conducted at only 1 site and were restricted to a short period of time; yet, these data could be useful for elucidation of effective pollinators of this species and, thus, its conservation.

Non-target-site mechanism of glyphosate resistance in Italian ryegrass (Lolium multiflorum): NTSR to glyphosate in ryegrass

In Shizuoka Prefecture, Japan, glyphosate-resistant Lolium multiflorum is a serious problem on the levees of rice paddies and in wheat fields. The mechanism of resistance of this biotype was analyzed. Based on LD50, the resistant population was 2.8–5.0 times more resistant to glyphosate than the susceptible population. The 5-enolpyruvyl-shikimate-3-phosphate synthase (EPSPS) gene sequence of the resistant biotype did not show a non-synonymous substitution at Pro106, and amplification of the gene was not observed in the resistant biotype. The metabolism and translocation of glyphosate were examined 4 days after application through the direct detection of glyphosate and its metabolite aminomethylphosphonic acid (AMPA) using liquid chromatograph-tandem mass spectrometer (LC-MS/MS). AMPA was not detected in either biotype in glyphosate-treated leaves or the other plant parts. The respective absorption rates of the susceptible and resistant biotypes were 37.90 3.63% and 41.09 3.36%, respectively, which were not significantly different. The resistant biotype retained more glyphosate in a glyphosate-treated leaf (91.36 1.56% of absorbed glyphosate) and less in the untreated parts of shoots (5.90 1.17%) and roots (2.76 0.44%) compared with the susceptible biotype, 79.58 3.73%, 15.77 3.06% and 4.65 0.89%, respectively. The results indicate that the resistance mechanism is neither the acquisition of a metabolic system nor limiting the absorption of glyphosate but limited translocation of the herbicide in the resistant biotype of L. multiflorum in Shizuoka Prefecture.

The Expansion Route of Ryegrasses (Lolium spp.) into Sandy Coasts in Japan

Although an increasing number of investigations have been made into the evolution of alien species once introduced, few studies have identified the invasion routes of these introduced species. Because multiple introductions are common in invasive species, failing to take into account the introduced lineages can be misleading when studying evolutionary change in alien species after they begin to extend their ranges. In Japan, diverse lineages of ryegrasses (Lolium spp.) were introduced as forage crops and contaminants in trading grain and have expanded to sandy coasts. We studied the expansion route of populations established along the coasts of three geographic regions within Japan by comparing variations in morphology and nuclear microsatellite and chloroplast DNA in the two habitats where ryegrasses were first introduced: croplands and international seaports. Chloroplast DNA haplotypes did not differ significantly among habitats and regions, but the coastal and seaport populations displayed similar microsatellite genetic compositions and morphological characteristics. Our results revealed that coastal populations originated from seaport populations derived from contaminants. Selective forces from the past, including domestication and naturalization, may have assisted the introduced lineages in colonizing new habitats. Nomenclature: Italian ryegrass, Lolium multiflorum L.; rigid ryegrass, Lolium rigidum Gaudin.

Variation in seed germination of cultivated and weedy perilla (Perilla frutescens var. frutescens)

Seed germination was investigated in four populations of Perilla (Perilla frutescens var. frutescens) collected in Nagano, Japan. One of the four grew as a weed in a plum orchard, and the other three were cultivated by different practices. Seeds obtained from the four populations were sown on May 21, 1993, and mature seeds were harvested from raised plants from October 1 to November 1 of that year. The seeds of each population were divided into two groups. One was stored at a constant 5°C, and the other was stored at an alternate temperature. The germination of one hundred seeds in each population of both groups kept at 25°C was recorded at intervals of one month with three replications. The germination of freshly harvested seeds was investigated with three replications.The germination percentage of each population increased with longer storage and was higher in the seeds stored under an alternate temperature than those stored under a constant 5°C. Freshly harvested seeds and those stored for less than two months in weedy perilla (No. 25 population) did not germinate, whereas 14% of the freshly harvested seeds germinated in no. 11 population, which was assumed to be the most domesticated type. After two months of storage under an alternate temperature, 80% of the seeds of no. 11 population germinated. The other two populations showed an intermediate pattern of germination between the no. 11 and 25 populations. The differences in the seed germination pattern seemed to be due to the differences in the degree of domestication.