Boletoid fungi (Boletaceae, Basidiomycota) of protected areas of Kon Tum Plateau (Central Highlands of Vietnam)

The article continues the series of publications devoted to the boletoid fungi of Vietnam. It summarizes the results of the observation of their diversity in three nature protected areas of the Central Highlands (Tây Nguyên) – Kon Ka Kinh National Park, Kon Chu Rang (Kon Chư Răng) Nature Reserve and Kon Plong Protected Forest, where middle-mountain evergreen broad-leaved and coniferous-deciduous forests prevail at altitudes of 900–1500 m. An annotated list of 22 revealed species is presented. Of these, two species (Fistulinella aurantioflava and Tylopilus subotsuensis) have been recently described from this territory as new to science. Hortiboletus rupicapreus is reported “ad interim”. Information on eight species (Aureoboletus sinobadius, Hourangia nigropunctata, Ionosporus longipes, Neoboletus multipunctatus, Phylloporus luxiensis, Pulveroboletus subrufus, Tylopilus atripurpureus, T. atroviolaceobrunneus) is published for the first time for Vietnam. Strobilomyces aff. echinocephalus, S. aff. glabriceps, and Tylopilus aff. balloui (two different lines) are close to known species but genetically not identical. Six more specimens are identified only to the genus, and they probably represent species new to science. The color photographs of new to Vietnam and noteworthy species are presented. The nucleotide sequences obtained during the study were deposited in NCBI GenBank.


Introduction
This article continues the series of publications devoted to the diversity of the boletoid fungi in Vietnam (Pham et al., 2018;Pham, Morozova, 2020). The results of the work in three nature protected areas of the Central Highlands (Tây Nguyên) -Kon Chu Rang (Kon Chư Răng) Nature Reserve, Kon Ka Kinh National Park and Kon Plong Protected Forest -are summarized here. Before the beginning of our research, there was no information on the mycobiota of this territory. Then, the data on the agaricoid genera Entoloma, Volvariella, Pluteus were published (Morozova et al., 2018;Malysheva et al., 2019Malysheva et al., , 2020. An annotated list of 22 species of Boletaceae is presented here. Of these, two species (Fistulinella aurantioflava and Tylopilus subotsuensis) have been recently described by us from this territory as new to science (Crous et al., 2020(Crous et al., , 2021. Hortiboletus rupicapreus is reported here "ad interim", it will be published soon. Eight species (Aureoboletus sinobadius, Hourangia nigropunctata, Ionosporus longipes, Neoboletus multipunctatus, Phylloporus luxiensis, Pulveroboletus subrufus, Tylopilus atripurpureus, and T. atroviolaceobrunneus) are published for the first time for Vietnam. But most of them (by the exclusion of Aureoboletus sinobadius and Tylopilus atripurpureus) were reported previously in the manuscript of the dissertation work of the first author (Pham, 2020).

Materials and methods
Collections were made by route method in different types of tropical forests on Kon Tum plateau in 2015-2017 in course of the investigation of mycobiota of the Central Highlands (Tây Nguyên).
The information on the nature conditions of the territories studied is provided below.

Kon Chu Rang Nature Reserve
The KBTTN Kon Chu Răng is located in the northeastern part of Gia Lai Province (K'Bang District, Son Lang Commune), between 14.5°N-14.58°N and 108.5°E-108.65°E, and is bordered by the provinces of Binh Dinh, Quang Ngai, and Kon Tum. The area of the reserve is 159 km 2 , of which 156.1 km 2 (99 %) are primary and intact forests. The relief of the reserve is mostly hilly, mountainous in the northern part, with heights from 800 to 1452 m a. s. l. (Kon Chư Răng). The average annual temperature is about 21 °C (maximum in May +28 °C, minimum in January +12 °C). The average annual precipitation is about 1900-2000 mm, the peak of precipitation falls in September (340 mm). The dry season is relatively short, with up to 60 mm of precipitation per month in January -April (New et al., 2002;Sourcebook of Existing ..., 2004). Numerous rivers flowing through the reserve belong to basin of the Kon River (Sông Kôn), which has a number of waterfalls, the most famous and tallest of which is 50 m high.
The main forest type is middle-mountain evergreen broad-leaved and mixed forest distributed at elevations between 900 and 1500 m in the northwest of the nature reserve. Canopy cover of this forest type is 70 to 80 %, and the tree flora is dominated by species from the Fagaceae (Lithocarpus, Quercus, Castanopsis), Lauraceae, Fabaceae, Clusiaceae, Myrtaceae, Ericaceae, Burseraceae, and Magnoliaceae, mixed with gymnosperms (Dacrycarpus imbricatus, Dacrydium elatum). Lowland evergreen forest occurs at elevations below 900 m. Only 2 % of the nature reserve is covered by secondary vegetation, mainly scrub with scattered trees.

Kon Plong Protected Forest
The Kon Plong Protected Forest is located in the northeast of Kon Tum Province (Kon Plong County), within the state forestry enterprises of Mang Canh II, Tan Lap, and the Ngok tem (Ngọk tem) Water Protection Forest. The massif has an area of 650.8 km 2 and is located on the highly dissected mountain basalt plateau Kon Ha Nung (Kon Hà Nừng), at an altitude of 1100 to 1757 m a. s. l. The studied territory is located in the basin of the Lo (Đắk Lo), Te (Nước Che) rivers and the lower reaches of the Khe (Đắk Khê) river within 14.7125°N-14.7589°N and 108.3°E-108.3233°E. The climate is characterized by moderate temperatures (average annual temperature is about +18 °C, and the coldest month +16 °C) and heavy rainfall (up to 2800 mm per year).
The vegetation cover above 1000 m a. s. l. is dominated by middle-mountain evergreen mixed forests, the upper tier of which is dominated by

Morphological study
Macromorphological features were studied on the basis of fresh and dried material, as well as through the analysis of photographs and descriptions taken in the field.
Micromorphological structures were studied using herbarium material. The collected specimens were examined using light and electron microscopy, as well as by analyzing DNA sequences (ITS and tef1a sites). Microscopical characters were studied with a light Zeiss Axioscope A1 microscope with AxioCam1Cc 3 camera and program tools AxioVisionRel.4.6. (Carl Zeiss, Germany). Basidiospores, basidia, and hymenial cystidia were observed in squash preparations of small parts of the tubes in 5 % KOH. The pileipellis was examined on a radial section of the pileus, the stipitipellis -on longitudinal slice of the stipe in 5 % KOH. Basidiospore dimensions are based on 20 measurements, whereas cystidia and basidia dimensions are based on observing at least 10 structures per collection. Boletoid fungi of the Kon Tum Plateau (Vietnam) SEM photos were also used for the identification of material. They have been made by L. A. Kartseva on a scanning analytical electron microscope JEOL JSM-6390LA of Core Facility Centre 'Cell and Molecular Technologies in Plant Science' of Komarov Botanical Institute and by A. V. Alexandrova (Lomonosov State University) on scanning analytical electron microscopes JSM-6380LA and Camscan-S2 (Cambridge Instruments, 1990).

Molecular-genetic study
The molecular study is based on a phylogenetic species recognition (Taylor et al., 2000). To assess the taxonomic status of the identified phylogenetic lineages and put forward species hypotheses, we relied on the accepted boundaries of the possible variability of the ITS1-5.8S-ITS2 and tef1α nDNA regions within the species (Petersen et al., 2008;Hughes et al., 2009).
Collections studied are kept in the Mycological Herbarium of the Komarov Botanical Institute (LE).

Results and discussion
Twenty-two species revealed in the Kon Tum plateau are listed below including ten species published for the first time for Vietnam (marked with "!").    Totally, the presence of 28 species of Boletaceae in the studied area were confirmed by the molecular data (Table). Strobilomyces aff. echinocephalus, S. aff. glabriceps, and Tylopilus aff. balloui (two different lines) morphologically are close to known species but genetically not identical with the sequences kept in the NCBI GenBank. In addition, 6 more genotypes were revealed on the base of the molecular study. They could not be attributed to any known species either by molecular or morphological data; there are: Boletus sp. 1, Boletus sp. 2, Chalciporus sp. 1, Chalciporus sp. 2, Phylloporus sp. 1, Phylloporus sp. 2.
The species are distributed between 17 genera belonging to 5 subfamilies and the Pulveroboletus group. Among the territories studied, the highest diversity of boletoid species was found in the Kon Ka Kinh National Park due to the higher diversity of the communities represented there and more favorable weather conditions during the collection period. The mountain evergreen broad-leaved and coniferous-deciduous forests of this park are richest in the diversity of the Boletaceae species.