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Estimating the role of specific processes in the spread of alien species necessitates the determination of introduction pathways and source populations of invaded areas. Alien grasses in the genus Lolium that have extensively invaded Japan provide a unique opportunity to estimate the expansion process through direct comparison between source and naturalised populations because the introduction pathways, contaminants in grain commodities and commercial cultivars for fodder crops or revegetation materials are well‐known. Therefore, by directly comparing source and naturalised populations, we estimated the introduction pathways and whether adaptative evolution occurred in Lolium species on sandy coasts in Japan.Lolium individuals sampled from naturalised populations in croplands, seaports, and sandy coasts were compared with those from two introduction sources for morphological and genetic variations based on a genome‐wide single nucleotide polymorphism analysis and a common garden experiment. Furthermore, we conducted a reciprocal transplant experiment between cropland and sandy coast.Populations naturalised in croplands were closely related to the cultivars, whereas those naturalised in seaports and sandy coasts were associated with contaminants. These results indicate that the cropland and sandy coast populations are derived from cultivars and contaminants, respectively. In addition, asymmetric gene flow from cropland populations to sandy coast populations was observed. The reciprocal transplant experiment clearly demonstrated the home site advantage; populations derived from croplands yielded higher floret numbers than those derived from other habitats at the cropland site; sandy coast populations had higher survival rates than those from croplands at the coastal site. Port populations exhibited a similar tendency as sandy coast populations, indicating that contaminants may be originally adapted to salty and dry environments, such as that in sandy coasts. The flowering phenology in the sandy coast populations evolved in the late flowering; therefore, late flowering alleles may have been transferred from cropland populations to sandy coast populations.Synthesis. We demonstrated that two congeneric species with different ecological characteristics were introduced through multiple introduction pathways and spread across different habitats. A direct comparison between source and naturalised populations can considerably elucidate the patterns and processes of biological invasions. [ABSTRACT FROM AUTHOR]