THE FIRST RECORDS OF TWO RARE POLYPOROID FUNGI PICIPES SUBMELANOPUS AND PICIPES ULLEUNGENSIS IN RUSSIA Текст научной статьи по специальности «Биологические науки»

ISSN 1560-7259 (print edition)

TURCZANINOWIA

ISSN 1560-7267 (online edition)

УДК 582.284+581.95(571.14+571.150)

The first records of two rare polyporoid fungi Picipes submelanopus and Picipes ulleungensis in Russia

V. A. Vlasenko1' 3*, S. V. Volobuev2 4, A. V. Vlasenko1 5

1 Central Siberian Botanical Garden SB RAS, Zolotodolinskaya St., 101, Novosibirsk, 630090, Russian Federation 2 Komarov Botanical Institute RAS, Professora Popova St., 2, St. Petersburg, 197022, Russian Federation 3E-mail: vlasenkomyces@mail.ru; ORCID iD: https://orcid.org/0000-0001-5928-0041 4E-mail sergvolobuev@binran.ru; ORCID iD: https://orcid.org/0000-0003-1217-5548 5E-mail: anastasiamix81@mail.ru; ORCID iD: https://orcid.org/0000-0002-4342-4482

* Corresponding author

Keywords: DNA barcoding, new records, polyporoid fungi, rare species, Western Siberia.

Summary. Molecular genetic studies of the morphological genus Polyporus s. l. recently made it possible to detect the presence of several phylogenetic lineages among its representatives, to clarify the boundaries and scope of the taxon. Species with a black cuticle covering the stipe of the basidiomata have been separated in the genus Picipes. Most representatives of this genus are known from China. In Russia, out of 29 species of the genus, there are three widespread species of Picipes - P. badius, P. melanopus, P. tubaeformis, and one rare species - P. rhizophilus. Phylogenetic analysis of the nucleotide sequences of selected nrDNA loci obtained from the herbarium materials collected by us, originally assigned to Picipes melanopus, revealed the presence of two rare, morphologically similar species, P. submelanopus and P. ulleungensis, which turned out to be new to Russia. Picipes submelanopus is characterized by a terrestrial fruiting bodies and differentiated from P. melanopus, that grows on dead wood. All specimens of fruiting bodies of Picipes submelanopus collected by us in Western Siberia were found on the roots of woody plants, as in the typе locality in China. Picipes ulleungensis differs from P. melanopus by larger fruiting bodies, slightly larger pores. In order to differentiate Picipes submelanopus and P. ulleungensis from morphologically similar species, in addition to morphological characters and ecological preferences, it is desirable to use DNA barcoding methods.

Первые находки двух редких полипороидных грибов Picipes submelanopus и Picipes ulleungensis в России

В. А. Власенко1, С. В. Волобуев2, А. В. Власенко1

1 Центральный сибирский ботанический сад СО РАН, ул. Золотодолинская, д. 101, г. Новосибирск, 630090, Россия

2 Ботанический институт им. В. Л. Комарова РАН, ул. Профессора Попова, д. 2, г. Санкт-Петербург, 197022, Россия

Ключевые слова: ДНК-штрихкодирование, Западная Сибирь, новые находки, полипороидные грибы, редкие виды.

Аннотация. Молекулярно-генетические исследования морфологического рода Polyporus s. l. за последнее время позволили обнаружить среди его представителей наличие нескольких филогенетических линий, уточнить границы и объем таксона. Виды с черной кутикулой, покрывающей ножку плодового тела, были выделены в род Picipes. Большая часть представителей этого рода известна из Китая. В России из 29 ви-

Поступило в редакцию 21.12.2022 Принято к публикации 11.07.2023

Submitted 21.12.2022 Accepted 11.07.2023

дов рода встречается три широко распространенных вида Ртрея - Р. Ъа&ш, Р. шг1апорш, Р. tubaeformis и один редкий вид - Р. гЫхорИИш. Филогенетический анализ нуклеотидных последовательностей избранных локусов ярДНК, полученных из собранных нами гербарных материалов, первоначально отнесённых к Picipes ше1апорш\ позволил выявить наличие двух редких, морфологически сходных видов - Р. submelanopus и Р. ulleungensis, оказавшихся новыми для России. Picipes ^'иЬтеапорш' характеризуется наземными плодовыми телами и отличается от Р. melanopus, растущего на мертвой древесине. Все образцы плодовых тел Picipes submelanopus, собранных нами в Западной Сибири, были обнаружены на корнях древесных растений, как и в типовом местонахождении в Китае. Picipes ulleungensis отличается от Р. melanopus более крупными плодовыми телами, несколько более крупными порами. Для дифференциации Picipes submelanopus и Р. ulleungensis от морфологически сходных видов, помимо морфологических признаков и экологических предпочтений, желательно использовать методы ДНК-штрихкодирования.

Introduction

The genus Picipes was described by Ivan Zmitro-vich and Aleksandr Kovalenko (Zmitrovich, Kovalenko, 2016) with the type species Picipes badius (Pers.) Zmitr. et Kovalenko. Three species were assigned to the new taxon, including also Picipes mela-nopus (Pers.) Zmitr. et Kovalenko and P. tubaeformis (P. Karst.) Zmitr. et Kovalenko, segregated from the morphological genus Polyporus P. Micheli ex Adans., belonging to a separate phylogenetic lineage. Earlier, these species were combined into the "Melanopus" morphological group based on the presence of a black cuticle covering the entire or most of the stipe of the basidiomata (Nunez, Ryvarden, 1995). Later, polyporoid fungi of this group were actively studied by mycologists from China, where their high species diversity is observed. Molecular genetic studies made it possible to clarify the taxonomic status and transfer another 10 species to the genus Picipes, and another 18 species were described as new to science in the genus, two of which were transferred to the genus Polyporus s. str. (Dai et al., 2014; Zhou et al., 2016; Cui et al., 2019; Bhunjun et al., 2022; Ji et al., 2022). Thus, based on the data below, the genus Pici-pes currently includes 29 taxa.

Material and methods

We collected specimens in the Western Siberia, the Novosibirsk Region and the Altai Territory. We took photographs of fresh fungal fruiting bodies, as well as dry basidiomata from the herbarium collection. We used a Zeiss Axio Imager A1 light microscope (Carl Zeiss Microscopy, Germany) for the detailed morphological examination.

DNA extraction and sequencing

To extract DNA, we crushed fragments of dried fungal fruiting bodies (0.01 g) using aluminium oxide (Al2O3). Then, we homogenized them in the

Phyto-Sorb kit lysis buffer (Synthol, Moscow). We amplified the ITS1-5.8S-ITS2 nrDNA regions with ITS1F and ITS4B primers (Gardes, Bruns, 1993). We used HS Taq DNA Polymerase (Evrogen, Moscow). We used C1000 Thermal Cycler (Bio-Rad, USA) and visualized amplicons in Gel Doc XR+ Imager (BioRad, USA) to perform the PCR. We executed Sanger DNA sequencing at SB RAS Genomics Core Facilities (ICBFM SB RAS, Novosibirsk, Russia).

Sequence alignments and phylogenetic analyses

We analyzed the Picipes species' complete ITS1-5.8S-ITS2 sequenograms in Chromas, version 2.6.6 (http://technelysium.com.au/wp/chromas/). We compared these sequences with the existing data in GenBank using the Nucleotide BLAST tool (Altschul et al., 1990). Based on the BLAST search, we incorporated additional ITS sequences of other Polyporus s. l. species into the analyses.

Sequences and alignment

The total ITS dataset consists of 23 sequences including an outgroup. For this study we produced newly generated ITS sequences. Table provides an overview of the taxa we used, information on the herbarium specimens, GenBank accession numbers, and references. We aligned the sequences online in MAFFT (Katoh et al., 2002), version 7 (http://mafft. cbrc.jp/alignment/server/) using the E-INS-I strategy (Katoh, Toh, 2008). We chose Trametes conchi-fer (Schwein.) Pilât as an outgroup (Justo, Hibbett, 2011).

Phylogenetic analysis

We used MEGA X software (Kumar et al., 2018) for the statistical analyses of our nucleotide datasets. We reconstructed phylogenic relations using the Maximum Likelihood (ML) method (Felsenstein, 2004). We built phylogenetic trees in the online version (Trifinopoulos et al., 2016, http://iqtree.

cibiv.univie.ac.at/) of IQ-TREE software (Nguyen as statistical support in this analysis. We visualized et al., 2015) with the standard parameters. We used the phylogenetic trees in Fig Tree version 1.4.4 nonparametric bootstrapping with 1000 replicates (Rambaut, 2018).

Table

Sequences we used for the alignments with their corresponding GenBank access numbers and references

Species Herbarium voucher/ isolate (ITS) Genbank accession numbers (ITS) Reference

Lentinus (Polyporus) brumalis Dai 14946 KX851609 Zhou et Cui (GenBank)

Lentinus tigrinus - KY565250 Akbarikiarood et Rahnama (GenBank)

Lentinus substrictus (Polyporus ciliatus) H6012970 LN610426 Miettinen (GenBank)

Neofavolus alveolaris CBS 207.48 MH856311 Vu et al., 2019

Picipes badius UBC:F19745 HQ604799 Berbee et al., (GenBank)

Picipes rhizophilus Dai 16082 KX851634 Ji et al., 2022

Picipes submelanopus HMAS 290127 MK966675 Wei (GenBank)

Picipes submelanopus NSK1014847 OP458518 in this study

Picipes ulleungensis Cui12410 KX900022 Ji et al., 2022

Picipes ulleungensis SFC 20120814-41 KY038467 Tibpromma et al., 2017

Picipes ulleungensis NSK1014990 OP964916 in this study

Picipes ulleungensis KUC20130809-19 KJ668475 Jang et Kim (GenBank)

Podofomes mollis CLZhao 4429 MK343523 He et Zhao, 2022

Polyporus chozeniae LE301310 KJ595567 Zmitrovich et al., 2014

Polyporus fraxineus Dai 2494 KC572023 Zhou et al., 2016

Polyporus melanopus H 6003449 JQ964422 Xue et Zhou, 2012

Polyporus pseudobetulinus TFM:F-27567 AB587644 Zhou et al., 2011

Polyporus squamosus SP-16 MK386841 Farooq et al., (GenBank)

Polyporus tuberaster LE-BIN 3865 MG735335 Senik et al., (GenBank)

Polyporus tubaeformis CLZhao 4429 MK894074 He et Zhao, 2022

Polyporus umbellatus - AY322495 Xing et Guo, 2005 (GenBank)

Polyporus varius MHHNU 30391 MK182303 Chen et al., (GenBank)

Trametes conchifer FP106793sp JN164924 Justo et Hibbett, 2011

Results and discussion

Picipes submelanopus (H. J. Xue et L. W. Zhou) J. L. Zhou et B. K. Cui (= Polyporus submelanopus H. J. Xue et L. W. Zhou) (Fig. 1A, B, E).

Specimens examined. RUSSIAN FEDERATION: "Altai Territory, Troitsky district, Borovlyanka village, 52.6170°N, 84.4379°E, 229 m a. s. l., birch-pine forest, on the roots of Betula pendula in soil. 11 VIII 2009. V. A. Vlasenko" (NSK 1014992, GenBank OP964915 (ITS)); "Novosibirsk Region, Novosibirsk City, Akademgorodok, Botanical Garden, 54.8148°N, 83.1019°E, 166 m a. s. l., plantations of willows, on the roots of Salix sp. in soil. 21 VII 2009. V. A. Vlasenko" (NSK 1014993); ibid, "Akademgorodok, st. Tereshkovoy 9, 54.8431°N, 83.1058°E, 175 m a. s. l., planting birches along the road, on the roots of Betula pendula in soil. 03 VII

2021. V. A. Vlasenko" (NSK 1014847, GenBank OP458518 (ITS), OP965388 (LSU)).

Distribution. North America - Canada, Europe - Estonia, Latvia; Asia - Russia (in this study), China, Pakistan (Xue, Zhou, 2012; Runnel et al., 2021; Picipes submelanopus, 2023).

Comments. The ML analysis of the ITS1-5.8S-ITS2 nrDNA region confirmed that the studied specimen belongs to the Picipes submelanopus (Fig. 2). The species as an independent taxon was introduced only in 2012, based on molecular genetic studies. On a global scale, the species is known from four localities, with a type locality in China. Picipes submelanopus is morphologically similar to P. mela-nopus.

Picipes submelanopus is characterized by a terrestrial habit, centrally or laterally black-stipitate basid-iomata, straw-yellow to honey-yellow pore surface,

circular to angular pores, two types of generative hyphae bearing either simple septa or clamp connections, and cylindrical basidiospores. However, Picipes melanopus has smaller pores and clamped generative hyphae only (Xue, Zhou, 2012). All specimens of the species collected by us in Western Siberia were found on the roots of woody plants - on a birch and willow. In the type locality, in China, Picipes submelanopus has also been found on the soil on tree roots (Xue, Zhou, 2012).

According to this ecological feature, the species is differentiated from Picipes melanopus, which develops on dead wood. Records from China, Pakistan, and Russia have been confirmed by molecular genetic studies. In Latvia, the species is known from metagenomic soil studies data (Picipes submelano-pus, 2023).

Fig. 1. Picipes submelanopus: A, B - fresh basidiomata; E - hymenial surface; Picipes ulleungensis: C, D - dry basidi-omata; F - hymenial surface. Bars: A-D - 1 cm; E-F - 1 mm.

Picipes ulleungensis (H. Lee, N. K. Kim et Y. W.

Lim) B. K. Cui, Xing Ji et J. L. Zhou (= Polyporus ulleungensis H. Lee, N. K. Kim et Y. W. Lim) (Fig. 1C, D, F).

Specimens examined. RUSSIAN FEDERATION: "Altai Territory, Talmensky district, near Vy-polzovo village, 53.8296°N, 83.7521°E, 162 m a. s. l., birch forest, on a fallen trunk of Betula pendula. 09

VII 2008. V. A. Vlasenko" (NSK 1014990, GenBank OP964916 (ITS)).

Distribution. Europe - Estonia; Asia - Russia (in this study), China, South Korea (Tibpromma et al., 2017; Runnel et al., 2021; Ji et al., 2022).

Comments. The ML analysis of the ITS1-5.8S-ITS2 nrDNA region confirmed that the studied specimen nests within the Picipes ulleungensis (Fig. 2). The species as an independent taxon was de-

scribed in 2017, based on molecular genetic studies. On a global scale, the species is known from four localities, with a type locality in South Korea. Picipes ulleungensis is morphologically similar to P. melanopus. Specimens of the species, collected by us in Western Siberia, were found on dead wood - on a birch. In the type locality in Korea, Picipes ulleungensis has also been found on a woody plant

belonging to the genus Betula - B. platyphylla var. japonica (Tibpromma et al., 2017). Picipes ulleungensis, according to the original description, differs from P. melanopus larger basidiomata and slightly larger pores (5-6 on mm vs. 6-8 on mm). Records from South Korea, China, and Russia have been confirmed by molecular genetic studies.

Fig. 2. The maximum likelihood tree based on ITS1-5.8S-ITS2 nrDNA sequences shows the phylogenetic relationships between Picipes submelanopus, P. ulleungensis, and other closely related Polyporus s. l. species. Numbers on the tree demonstrate genetic distances (branches) and bootstrap values (nodes). Each sequence name contains its GenBank access number.

Acknowledgements

The work of V. A. Vlasenko and A. V. Vlasenko was carried out within the framework of the State Task for the Central Siberian Botanical Garden, the

Siberian Branch of the Russian Academy of Sciences, project AAAA-A21-121011290024-5. The work of S. V. Volobuev was carried out within the framework of the State Task for the Komarov Botanical Insti-

tute (project AAAA-A18-118022090078-2). DNA isolation and sequencing were financially supported by the Ministry of Education and Science of Russia under Agreement No. 075-15-2021-1056 of September 28, 2021 between the BIN RAS and the Ministry of Science and Higher Education of

the Russian Federation, also under Agreement No. En/29-10-21-4 of October 29, 2021 between BIN RAN and CSBG SB RAS. Fungal specimens are deposited in M. G. Popov Herbarium, USU 440537, Novosibirsk.

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