Bioscience Methods 2025, Vol.16, No.3, 137-153 http://bioscipublisher.com/index.php/bm 149 reminds us to be cautious in dealing with the mixing of the two varieties to prevent genetic contamination and economic losses. From a conservation perspective, while the large-scale cultivation of H. discus hannai should be carried out, the genetic resources of local abalone should be retained, such as building isolated original breeding farms or protected areas to prevent it from being replaced in competition with foreign species. 6.3 Phylogenetic and taxonomic assessment of Japanese, Australian, and East Asian abalones There have been some disputes in the taxonomy of abalone, focusing on species with close geographical distribution or similar morphology. For example, Japanese abalone includes two main types, H. discus hannai and H. discus black, Australian abalone includes green abalone and black-lipped abalone, and there are also H. discus and H. variegata in the coastal areas of East Asia. This section discusses the relationship and taxonomic treatment of abalone in these regions based on mitochondrial pedigree information. First, Japanese H. discus hannai and Japanese black abalone (H. discus discus) have long been regarded as different subspecies of the same species, but there are different opinions on taxonomy. Molecular phylogenetic results show that although the two abalones form a sister group relationship, they are still clearly distinguished in branches, and the genetic distance reaches the typical species level (Hsu and Gwo, 2017). In particular, they each form a monophyletic branch without haplotype confusion, which supports the view that they are independent species. Chinese scholars have also found in aquaculture practice that the survival rate of hybrid offspring of the two is extremely low, which further confirms the taxonomic distinction from the perspective of reproductive isolation. Therefore, in recent years, most researchers tend to formally recognize the Japanese black abalone (also known as the true abalone) as an independent species H. discus, while retaining the original scientific name of the wrinkled disc abalone H. hannai. In contrast, the controversy between the Taiwan nine-hole abalone and the mainland variegated abalone is just the opposite - they were once considered different species, but are now mostly merged into the same species. The Taiwan nine-hole abalone (traditionally called H. diversicolor supertexta) is slightly different from the mainland variegated abalone in shell shape and growth rate, which has led some scholars to advocate that it is a subspecies or even species-level distinction. However, molecular evidence has repeatedly shown that the mitochondrial sequences of the two are almost indistinguishable and intertwined in the phylogenetic tree. For example, a comparison of 16S and COI sequences of Taiwan and mainland abalone revealed that the interspecific genetic distance was only about 1%, far lower than the typical species difference. Classical genetic studies such as isozymes and electrophoretic markers have also failed to clearly separate the two. Therefore, the current mainstream view tends to believe that Taiwan abalone is only a geographical population, not an independent species, and its species name supertexta can be reduced to a synonym or infraspecific taxonomic unit of H. diversicolor (Yang et al., 2023). The taxonomic controversy of Australian abalone mainly involves the interspecific relationship between green abalone (H. laevigata) and black-lipped abalone (H. rubra). They are distributed adjacently and have natural hybridization zones. At one time, some people suggested that they may belong to a highly variable population. However, mitochondrial DNA shows that the sequence similarity between the two is about 92%, and the interspecific differences are significant. Although hybridization occurs, the frequency is low and mostly one generation, without fusion of the two gene pools (Cook, 2025). The phylogenetic tree divides green abalone and black-lipped abalone into different branches, supporting that each is a species. It is also worth noting that there are several local species in Australia, such as H. conicopora, which are closely related to the above two species. The latest whole genome study found that they had a second contact after the ice age, and some genes showed selective introgression. This reminds us that in addition to distinguishing by morphology and mitochondria in classification, we should also monitor the exchange of nuclear genes to fully understand the species boundaries. By comparing the mitochondrial lineages of Japanese, Australian and East Asian abalone, many taxonomic disputes can be clarified. Molecular evidence generally supports the following viewpoints: Japanese wrinkled abalone and black abalone should be different species; Taiwan's nine-hole abalone and mainland variegated abalone should be considered the same species; Australian green abalone and black-lipped abalone are independent species, although there is limited hybridization in nature. For some other controversial taxa, such as the blue abalone and red abalone in Mexico, and the population division of rare abalone in South Africa, the same method can also be used for evaluation.
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