BIODIVERSITAS ISSN: 1412-033X Volume 23, Number 8, August 2022 E-ISSN: 2085-4722 Pages: 4159-4168 DOI: 10.13057/biodiv/d230838 Comparative leaf epidermis study in Habenaria spp. (Orchidaceae) from Thailand NOODUAN MUANGSAN 1, , PIYAPORN SAENSOUK 2 , THOTSAPORN CHANOKKHUN 1 , SANTI WATTHANA 1 1 School of Biology, Institute of Science, Suranaree University of Technology. Nakhon Ratchasima 30000, Thailand. Tel./Fax.: +66-44-223-000,  email: nooduan@g.sut.ac.th 2 Diversity of Family Zingiberaceae and Vascular Plants for Its Applications Research Unit, Mahasarakham University. Mahasarakham 44150, Thailand Manuscript received: 30 May 2022. Revision accepted: 28 July 2022. Abstract. Muangsan N, Saensouk P, Chanokkhun T, Watthana S. 2022. Comparative leaf epidermis study in Habenaria spp. (Orchidaceae) from Thailand. Biodiversitas 23: 4159-4168. The variation of epidermal and stomatal characteristics could be used for plant classification, including orchids. This study aimed to investigate leaf epidermis characters of Habenaria from Thailand and focused on the H. rhodocheila group and related species. Fourteen accessions belonging to 10 species were studied and compared. Epidermal peels were taken from the upper and lower surfaces of the leaves and were observed under a light microscope. The stomata of all accessions studied were distributed only in the lower epidermis, where they were hypostomatic and anomocytic. The epidermal cell density showed significant differences between the two surface sides. The stomatal size is significantly different among all samples. The highest stomatal index was presented in H. rostrata and the lowest on H. rhodocheila (red). There is no gap variation of epidermis characteristics in H. rhodocheila and its closely related species. It is indicated that the leaf epidermal characters are the shared characters of the genus Habenaria from Thailand. H. rostrata has tentatively separated from other species based on epidermal anatomical characteristics. Keywords: Anatomy, epidermis, Habenaria, Orchidaceae, stomata INTRODUCTION Habenaria Willd. (Orchidaceae) is a large terrestrial orchid genus, belonging to the subfamily Orchidoideae. The genus consists of 928 species, distributed throughout the tropical, subtropical, and temperate regions (Batista et al. 2013; Govaerts et al. 2022). Some species are well known for ornamental purposes and their therapeutic properties (Teoh 2016). The phylogenetic relationship between Habenaria and its alliance revealed that it is not monophyletic (Jin et al. 2014, 2017). Kränzlin (1893) proposed 32 sections for intrageneric classification, then Pridgeon et al. (2001) modified Kränzlin’s system and proposed 37 sections. Habenaria is reported 46 species in Thailand (Kurzweil 2011; Kurzweil et al. 2017). Kurzweil (2009) grouped Thai Habenaria based on morphological characteristics into 9 groups. The H. rhodocheila group consists of H. rhodocheila and H. carnea (Kurzweil 2009). Judging from morphological characteristic a recently new species, H. janellehayneana should be placed in this group. The leaf shape of the genus Habenaria is very diverse, varying from linear to broadly ovate, lanceolate to elliptic. The leaves arise in the rainy season and bloom in the mid- to-late rainy season. Their flowers are visible and recognizable only for one or two months of the year (Kurzweil 2009). Flowers of Habenarias are mainly white, green, or yellow, except in H. rhodocheila, H. carnea and H. janellehayneana with white, pink, red, orange, or yellow (Kurzweil 2009; Sinumporn et al. 2020). They are usually deciduous plants and have underground tubers. Foliar epidermal characters, such as epidermal cells, stomata, trichomes, cuticles, and wax have been used as taxonomic and phylogenetic characters across the Orchidaceae (Angela et al. 2015; Rindyastuti et al. 2018; Saensouk and Saensouk 2019). Some studies on the foliar epidermis features at the generic level in orchids have been investigated. Singh et al. (2020) reported that stomatal traits, glandular trichome type, distribution, and cuticular thickening pattern were potentially important for the characterization and differentiation of Bulbophyllum species, whereas only cuticular wax characteristics distinguished Holcoglossum from related genera (Fan et al. 2014). Darmawati et al. (2018) demonstrated that using a squared Euclidean distance of 34.9% similarity, two Dendrobium species were distinguished from other groups based on leaf anatomical features. Pradhan and Bajracharya (2018) found the distribution of stomata on the leaf surface, stomata type, and epidermal cell shape in Dendrobium were taxonomically significant. Later, Pradhan and Bajracharya (2020) used cluster analysis with anatomical and micromorphological characters to classify 26 Dendrobium species of Nepal into 6 sections. Several reports on leaf epidermal characters of Habenaria were demonstrated (Stern 1997; Dangat and Gurav 2016; Verma et al. 2018), but no defined characters among the genera have been reached. More members of this genus need to be done to complement the existing taxonomic information.