Molecular ecological studies on the naturalization of weedy type velvetleaf (Abutilon theophrasti) in Japan

In order to elucidate the source of the recent velvetleaf (Abutilon theophrasti) invasion and to understand the process of naturalization, I examined firstly intra-specific variation in morphological characters and growth habits, using accessions obtained from the USDA collection as well as from imported grains. The best index of morphological variation was capsule color, either ivory or ebony. Variation in capsule color was found to be governed by nuclear gene(s), with ebony being dominant. Cluster analysis based on the ISSR polymorphism showed two major clusters that corresponded well with variation in capsule color. All six accessions obtained from imported grains united into one major cluster, while all of the old Japanese accessions occurred in another major cluster. A sequencing analysis of approx. 6% of the chloroplast genome detected two chloroplast DNA (cpDNA) haplotypes. By combining the cpDNA haplotypes with capsule color variation, I was able to classify all the accessions into three genotypes (Types I, II and III). Type II was recognized as derived from the crossing of Type I (maternal) and Type III (paternal). All Japanese cultivars were identified as Type I, while USDA accessions included Types I, II and III. All the accessions obtained from imported grains and from the Corn Belt of the United States were Type III, and most of the accessions presently distributed in Japan were Type III. A molecular genotyping method using historical herbarium specimens of velvetleaf was developed as a potential tool to understand the naturalization process of the species in Japan. PCR-based markers were shown to be applicable for genotyping the historical herbarium specimens. The earliest record of haplotypes B was in 1893 in Tokyo. Most of the accessions obtained from fields of Japanese forage crops were classified as Type III, which had the cpDNA haplotype of the accessions obtained from seeds mixed with imported grains. The result appears to strongly support the hypothesis that the recent weedy velvetleaf occurring in Japan was derived from seeds mixed in with imported grains. However, it was found that Type II, which had a different cpDNA haplotype from Type III, also occurred sporadically in Japan. In addition, Type III was also found in herbarium specimens collected 110 years ago. These facts suggest that seeds contaminating imported grains might not be the only source of recent weedy velvetleaf in Japanese forage crop fields.