Leaf anatomy and cytogenetics analyses of selected acanthaceae species in Peninsular Malaysia

Abstract

Acanthaceae is one of the potential medicinal plant families that occurred in Peninsular Malaysia. A great contribution of the Acanthaceae was observed by the documentation of several Acanthaceae species in Malay Medical Manuscripts. However, the treatment of Acanthaceae subfamilies previously seems to have many changes including the placement of the genus Avicennia L. into the Acanthaceae. The placement of this genus somehow is made by molecular study without a strong justification from leaf anatomy, micromorphology and cytogenetics points of view. A systematic study thereby was conducted on ten selected species from nine genera of Acanthaceae from Peninsular Malaysia. This research aims to investigate the significance of leaf anatomy and micromorphology characteristics that can be used in the identification of Acanthaceae as well as determine the genome size data to support the systematic study of Acanthaceae. The leaf anatomical study involved several methods such as cross-section using a sliding microtome, leaf epidermal peeling, leaf clearing and observation under a light microscope. The leaf micromorphology method involved the observation of adaxial and abaxial surfaces under a Scanning Electron Microscope (Zeiss Model EVO 50). Cytogenetics study method involved the determination of genome size by using Flow Cytometry Method. The dichotomous key of species was constructed based upon leaf anatomical and micromorphological features gathered in this study. Phylogeny analysis was conducted based on leaf anatomical and micromorphological features by using Multivariate Statistical Package. The analysis of One-Way ANOVA based on genome size was done to investigate the interspecific variations among Acanthaceae species. Results revealed three common characteristics, fourteen variation characteristics and forty diagnostic characteristics of leaf anatomy and micromorphology among the Acanthaceae species. Results also revealed the separation of the genus Avicennia from other Acanthaceae species based on the constructed phylogeny tree. For the cytogenetics part, the genome size of Acanthaceae species was measured by using Glycine max cv. Polanka (2C=2.5pg) as external reference standard. Results revealed the genome size of Acanthaceae species ranged from 2.05 to 4.09 pg. The determination of genome size however was only obtained for six species studied, whilst the other four species were unable to represent a good peak of histogram. This might be due to the insufficient of nuclei quantities and interference of DNA staining during the analysis. The One-Way ANOVA of genome size showed a statistical difference, thus indicating the presence of interspecific variation among the Acanthaceae species. In conclusion, the leaf anatomy, micromorphology and genomes size data possessed taxonomic values and proven to be additional data for the identification and classification of Acanthaceae.