Water Lily Type Plant Phylogenetic Studies

Water lilies, or the order Nymphaeales sensu lato, are an important group in primitive angiosperms, which plays a key role in study of origin and evolution of angiosperms. This group includes 10 genera, i.e., Barclaya, Brasenia, Cabomba, Ceratophyllum, Euryale, Nelumbo, Nuphar, Nymphaea, Ondinea, and Victoria, and about 110 species. The phylogenetic relationships among and within these genera have long been controversial. Twenty-four morphological, anatomical, and palynological characters in water lilies were used in phylogenetic analyses. The orders Magnoliale, Laurales, Piperales, Illiciales, and Ranunculales were selected as a complex outgroup. The evolutionary polarity of each character was determined using the outgroup comparison method. On the basis of the character-coding matrix, the phylogenetic trees were constructed with Camin-Sokal method and Farris-Wagner method, implemented in PHYLIP package. The results of phylogenetic analyses showed that there are major differences between the two trees and existing classifications of water lilies, with none of the classifications was well supported by the trees. New evidence from extensive studies, especially molecular study, for solved the intergeneric relationships seems to be needed.The DNA sequences of chloroplast gene large subunit of the enzyme ribulose-1,5-bisphosphate carboxylase (rbcL) and nuclear ribosomal 18S RNA gene obtained from GenBank were analyzed using maximum parsimony (MP), neighbor-joining (NJ) and maximum likelihood (ML) methods, with Amborella as an outgroup according to the Angiosperm Phylogeny Group (APG) system. The intergeneric phylogenetic trees of water lilies showed that Nelumbo and Ceratophyllum form a sister group, but with relatively high sequence divergence, supporting that place the two genera into two distinct families; and Brasenia and Cabomba form a sister group, with relatively low sequence divergence, supporting that the two genera were placed the same family Cabombaceae. In addition, the sister-group relationship between Nymphaea and Ondinea was supported by some trees. The sister-group relationship between Euryale and Victoria did not support thatEuryale can be promoted to an independent family. The distinctness of Nuphar suggesting the Nupharaceae should also be reconsidered.The sequences of internal transcribed spacer (ITS) regions of nuclear ribosomal DNA (nrDNA) of 9 samples of Nymphaea and related water lilies as outgroups were sequenced and assembled with the sequences in GenBank to analyze the specific and infraspecific relationships. The results indicate that ITS sequences can be applied to detect the infraspecific variations, and the maximum likelihood method is more effective than the maximum parsimony. These provide a new approach to germplasm identification and phylogenetic analysis of the genus Nymphaea.In addition, fanwort (Cabomba caroliniana), an invasive species of water lilies in China, was investigated. This provides not only a new material for the phylogenetic studies of water lilies but also its distribution in eastern China and potential ecological consequences. Since fanwort has a different ecological niche than other aquatic weeds such as alligator alternanthera (Alternanthera philoxeroides) and common water hyacinth (Eichhornia crassipes), which could imply "joint" impact on native species diversity, further biological and ecological investigations of fanwort are urgently needed.In conclusion, a comprehensive analysis of water lilies based on morphological and molecular data was performed, showing the phylogenetic relationships among the 10 genera and providing new evidence for classification at family level. The specific and infraspecific relationships within the genus Nymphaea were revealed using the ITS sequences, providing a new approach to germplasm identification and phylogenetic analysis of water lilies. The investigation of invasive fanwort in China may also provide a sound basis of its ecological studies.