A new species of Presbistus Kirby, 1896, Presbistus vitivorus sp. nov., is described from Cambodia based on both sexes, nymphs, and eggs. Male genitalia and vomer are described and figured. Illustrations of adults, nymphs, specimens in situ, host plants, a distribution map and records on biology and breeding in captivity are provided. The host plants of the species belong to the family Vitaceae. The genus Presbistus and the family Aschiphasmatidae are recorded from Cambodia for the first time. The species diversity and the distribution of the genus are discussed, and it is shown that the genus is restricted to Sundaland. A nomenclature for the morphology of the dissected vomer is proposed and tries to homologize the previously used terms.

Aedeagus, Aschiphasmatini, Phasmatodea, stick insect, Vitaceae, vomer

The genus Presbistus Kirby, 1896 belongs to the Aschiphasmatini in the subfamily Aschiphasmatinae and currently contains nine species. Phylogenetic studies have shown Aschiphasmatinae to be monophyletic and the sister group to all remaining Euphasmatodea (Simon et al. 2015).

Presbistus is currently recorded from India, Sri Lanka, Peninsular Malaysia, Sumatra, Java, and Borneo (Brock et al. 2022). Within the Aschiphasmatinae, the genus is characterized by triangular or spine-like tegmina, uniform translucent brown anal region of hind wings, and cylindrical or conical cerci in both sexes. The male has a well-developed vomer and has the end of the abdomen distinctively swollen and club-shaped. The eggs are slightly longer than high and lack setae (Bragg 2001, Seow-Choen 2016).

The vomer, a sclerotized derivative of sternum X, was first introduced by Pantel (1890) as “vomer sous-anal”. The vomer is used during mating and clasps the posterior margin of sternum VII of the female, which has evolved into a praeopercular organ (Bradler 2003). Pantel (1915) discussed the structure and variability of the vomer in different subgroups of Phasmatodea and detected a vomer in three genera of “Ascepasmini” (= Aschiphasmatini) including Presbistus. Bradler (1999, 2009) argued that the vomer was already present in the phasmatodean ground plan and that it represents a plesiomorphic character for all Euphasmatodea. Most authors have only described the external part of the vomer, and a standardized nomenclature for the different parts of this organ is still needed.

During the study of stick insect specimens collected in the framework of the Global Taxonomy Initiative project “A step further in the entomodiversity of Cambodia,” a species belonging to the genus Presbistus could not be attributed to any known species.

The phasmid fauna of Cambodia remains poorly studied (Bresseel and Constant 2018b), and the present paper describes the new species as Presbistus vitivorus sp. nov., provides useful characters for differentiation, and gives information on its biology and distribution. A nomenclature for the morphological characterization of the vomer in stick insects is proposed.

Due to their nocturnal behavior (as in most Phasmida), the specimens of P. vitivorus sp. nov. were collected at night. A lightweight, water-proof Petzl MYO RXP head torch was used during collecting. The females were kept alive in a mesh pop-up cage (exo terra explorarium) for producing eggs. The wild caught specimens were euthanized by injection with ethanol. The specimens were then stored in airtight plastic “zip” bags containing wood chips commonly used in rodent cages and sprinkled with etylacetate (EtOAc) to prevent rotting, mould, and to keep the specimens flexible. The bags were frozen on arrival, and the specimens were mounted at a later date.

A number of pictures of each specimen were taken using a Canon 700D camera (Canon Inc., Ota City, Tokyo, Japan) equipped with a Sigma 50 mm macro lens (Sigma Corporation, Kawasaki, Japan) for adults or with a Leica EZ4W stereomicroscope (Leica Microsystems Ltd., Wetzlar, Germany) with integrated camera for eggs and male genitalia. The images were stacked using CombineZ software (https://combinezp.software.informer.com) and optimized with Adobe Photoshop CS3. The distribution map was produced using SimpleMappr (Shorthouse 2010). Observations were done with a Leica EZ4W stereomicroscope, and measurements were taken with an electronic calliper. The dissection of the vomer and aedeagus was done on a fresh specimen using a needle blade. The vomer was extracted after cutting the membrane around the connective ring that separates the internal apodemes from the externally visible body; the aedeagus was extracted after spreading the poculum and cutting the inner membrane around the integument that separates the internal basal apodeme from the external remaining parts of the aedeagus. After extraction, the vomer and aedeagus were boiled for a few minutes in 10% KOH solution to remove the remaining soft parts then rinsed and examined in 70% ethanol. The vomer and aedeagus were preserved in glycerine in a polyethylene genitalia tube and stored under the label of the corresponding specimen.

The nomenclature of the morphological characters follows Bragg (2001) and that of the egg morphology follows Clark-Sellick (1997, 1998). The description of the coloration is based on live specimens.

Acronyms used for the collections.—

RBINS Royal Belgian Institute of Natural Sciences, Brussels, Belgium.

RUPP Royal University of Phnom Penh, Cambodian Entomology Initiative, Phnom Penh, Cambodia.

VNMN Vietnam National Museum of Nature, Hanoi, Vietnam

Abbreviations.—

HT holotype

PT paratype

Male terminalia.—The morphology of the external terminalia in Aschiphasmatidae was recently studied (Valotto et al. 2016a, b), but the internal male genitalia have remained almost completely undocumented. Several studies provide basic information on phasmid genitalia (Chopard 1920, Walker 1922, Snodgrass 1937), but Helm et al. (2011) provided the first detailed study based on the Oriental species Oxyartes lamellatus Kirby, 1904 (Lonchodidae, Necrosciinae). Genitalia in Neotropical taxa were recently studied by Heleodoro and Rafael (2019), Chiquetto-Machado and Cancello (2021), and Ghirotto (2021), highlighting the importance of the phallic organ characters for phasmid systematics. At least the large sclerotized and specialized dorsal portion in Presibistus vitivorus sp. nov. seems to be homologous to the dorsal sclerite found in all species examined in the previously mentioned studies and coincides with the hypothesis by Helm et al. (2011) that the dorsal sclerite may be present in all or nearly all species of stick insects.

In Presbistus, the aedeagus is moderately sclerotized and shows considerable differences when compared to other studied taxa. The aedeagus provides several interesting characters for species differentiation and future phylogenetic considerations, but since our study represents the first documentation of a dissected aedeagus for the subfamily, no conclusions regarding these subjects can be drawn.

The characters of the male vomer are frequently used when describing or differentiating stick insect taxa (Bradler 2009, Bresseel and Constant 2018a, Cumming et al. 2021). The nomenclature proposed here for the dissected vomer can be used in nearly all species, and the structure of the vomer provides useful differential characters when studied from different angles. The suggested nomenclature (Fig. 4) tries to homologize with previously used terms. The vomer of a number of genera has been dissected, such as Timema Scudder, 1895 (Timematoidea), Cryptophyllium Cumming et al., 2021 (Phyllioidea), Dajaca Brunner von Wattenwyl, 1893 (Vallotto et al. 2016a), and Orthomeria Kirby, 1904 (Aschiphasmatoidea) (Bradler 1999, Vallotto et al. 2016a, b, Cumming et al. 2021), and these studies show the presence of two basal apodemes (Fig. 4: lba, rba) embedded in the body. The structure and direction of these apodemes can differ considerably between genera.

The body of the vomer (Fig. 4: b) refers to the external part of the vomer including the distal ending and is connected to the tegument by the connective ring (Fig. 4: cr). The body of the vomer shows considerable variation in shape but most often narrows towards the posterior. This organ can be functionally replaced or can be strongly reduced in genera with a modified tergum X or specialized cerci for clasping the female (Pantel 1915, Bradler 2003, 2009, Hennemann and Conle 2008).

The apical portion of the body of the vomer is often upcurved and armed with one to multiple spines. In many species the vomer ends in a single apical spine (Fig. 4, as), referred to as hook(s) or prong(s) by several authors (Bradler 1999, Bradler et al. 2014, Cumming et al. 2021). The apex can sometimes be more complex, bearing two (Neooxyartes Ho, 2018, Cryptophyllium) or more (Spinohirasea Zompro, 2002, Paramenexenus Redtenbacher, 1908) sometimes blunt, spines (Hennemann 2007, Bresseel and Constant 2018a, Bradler et al. 2014, Cumming et al. 2021). The body of the vomer can be symmetrical (Neooxyartes) to asymmetrical (Cryptophyllium) depending on the size and direction of the spine(s) (Bresseel and Constant 2018a, Cumming et al. 2021).

Species diversity and distribution.—With the description of this new taxon there are now ten known Presbistus Kirby, 1896 species. The family Aschiphasmatidae is for the first time recorded from Cambodia, with Presbistus vitivorus sp. nov being only the fifth species of stick insect described from the country (Brock et al. 2022).

Presbistus peleus (Gray, 1835), the type species, is the most widespread species in the genus and is recorded from India, Peninsular Malaysia, Singapore, Sumatra, and Borneo (Brock et al. 2022). The holotype originates from “Ora Malabariensi” (Gray 1835), considered the Malabar coast (India) by subsequent authors (Brock 1999, Bragg 2001). However, both authors expressed doubts about the accuracy of the original locality, as all subsequent records originate from Sundaland and the genus has never been confirmed in India. Other species described by Gray (1835) bearing the same collecting data and all “ex collection D. Children” have since been recorded from Peninsular Malaysia (Malacca) (Brock 1999, Bragg 2001). Therefore, the Indian record of P. peleus is regarded as erroneous. Furthermore, from photographs of the holotype female available from Brock et al. 2022, it appears that P. crudelis (Westwood, 1859) from Sri Lanka is not congeneric with the type species of Presbistus, P. peleus (Gray, 1835), and belongs in a separate, yet undescribed genus that differs from Presbistus by the almost straight profemora, the comparatively shorter mesonotum, and the spoon-shaped subgenital plate. Hence, the genus Presbistus is removed from the list of stick insects from India and Sri Lanka.

With P. vitivorus sp. nov extending the distribution of Presbistus to Cambodia, the genus is shown to be distributed over most of Sundaland, comprising Indochina, the Malay Peninsula, Sumatra, Java, and Borneo.

We thank Sophany Phauk (RUPP) and his students as well as Dr. Hong Thai Pham (Vietnam National Museum of Nature, Hanoi, Vietnam) and Xavier Vermeersch (RBINS collaborator) for their help, enthusiasm, and friendship during the collecting trips. Myriam Bourguignon and the staff and students of KTA Horteco (Vilvoorde, Belgium) are thanked for cultivating the plants needed for rearing the species in captivity. We also thank Tim Bollens (Herselt, Belgium) for his help in captive rearing of the species and providing additional study material and Cédric Cauquil (Soual, France) for additional information on incubation. Dr. Anna Trias-Blasi (Royal Botanic Gardens, Kew, UK) is thanked for her help in identifying the food plants from our photographs. Thank you to the authorities of Phnom Samkos Wildlife Sanctuary for their support of this study, to Sharyn and Benjamin J. Davis, the managers of Be Treed and the Cambodian Ministry of Environment for allowing and facilitating our research at BeTreed Adventures. Dr. Patrick Grootaert, Dr. Yves Samyn, and Dr. Marie-Lucie Susini (RBINS) are thanked for their permanent support of the project in Cambodia, Mado Berthet (RBINS) for improving the illustrations, and Gerard Chartier (Tatai, Cambodia) for his help in the field and hospitality. This paper is a result of the project “A step further in the entomodiversity of Cambodia” (2016–2017) supported through grants issued by the capacity building program of the Belgian Global Taxonomy Initiative National Focal Point that runs under the CEBioS program with financial support from the Belgian Directorate-General for Development Cooperation.