Research Article |
Corresponding author: Feng-Li Hui ( fenglihui@yeah.net ) Academic editor: Thorsten Lumbsch
© 2021 Wan-Li Gao, Ying Li, Chun-Yue Chai, Zhen-Li Yan, Feng-Li Hui.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Gao W-L, Li Y, Chai C-Y, Yan Z-L, Hui F-L (2021) New species of Yamadazyma from rotting wood in China. MycoKeys 83: 69-84. https://doi.org/10.3897/mycokeys.83.71156
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Yamadazyma is one of the largest genera in the family Debaryomycetaceae (Saccharomycetales, Saccharomycetes) with species mainly found in rotting wood, insects and their resulting frass, but also recovered from flowers, leaves, fruits, tree bark, mushrooms, sea water, minerals, and the atmosphere. In the present study, several strains obtained from rotting wood in Henan and Yunnan Provinces of China were isolated. Based on morphology and a molecular phylogeny of the rDNA internal transcribed spacer region (ITS) and the D1/D2 domain of the large subunit (LSU) rDNA, these strains were identified as three new species: Yamadazyma luoyangensis, Y. ovata and Y. paraaseri; and three previously described species, Y. insectorum, Y. akitaensis, and Y. olivae. The three new species are illustrated and their morphology and phylogenetic relationships with other Yamadazyma species are discussed. Our results indicate a high undiscovered diversity of Yamadazyma spp. inhabiting rotting wood in China.
Debaryomycetaceae, phylogeny, rotting wood-inhabiting yeast, taxonomy, Yamadazyma
The genus Yamadazyma
Yamadazyma species can be originally found in tropical, subtropical, and temperate regions of different continents, but most known species appear to exist in Asia and South America (
Samples of rotting wood were collected in the Xishuangbanna Primeval Forest Park (Yunnan Province, China) and the Tianchi Mountain National Forest Park (Henan Province, China). The Xishuangbanna Primeval Forest Park (21°98'N, 100°88'E) is 500 m above sea level (MASL), with a hot and humid climate. The average annual temperature is between 16 °C and 28 °C, and the average annual rainfall is above 1,100 mm. The Tianchi Mountain National Forest Park (34°33'N, 112°28'E) is at 850 MASL, with a continental monsoon climate, average annual temperature of 14–16 °C, and average annual rainfall between 800 mm and 900 mm. Fifty decayed wood samples were collected during July and August in 2018–2020. The samples were stored in sterile plastic bags and transported under refrigeration to the laboratory over a period of no more than 24 h. The yeast strains were isolated from rotting wood samples in accordance with the methods described by
Morphological and physiological properties were determined according to those used by
Genomic DNA was extracted from the yeast using an Ezup Column Yeast Genomic DNA Purification Kit, according to the manufacturer’s instructions (Sangon Biotech, Shanghai, China). The internal transcribed spacer (ITS) and the D1/D2 domain of the large subunit (LSU) rDNA were respectively amplified using ITS5/ITS4 (
The 25 µL total volume of PCR mixture contained 9.5 µL of ddH2O, 12.5 µL of 2X PCR Master Mix (TIANGEN Co., China), 1 µL of DNA template, and 1 µL of forward and reverse primers (10 µM each) in each reaction. PCR amplified products were checked on 1% agarose electrophoresis gels stained with GoldView I nuclear staining dye (1 µL/10 mL of agarose). Purification and sequencing of PCR products were done by Sangon Biotech (Shanghai) Co., Ltd., Shanghai, China. A consensus sequence for each gene region was assembled in SeqMan (DNAStar, Inc., Madison, WI, USA). The newly-generated sequences were deposited in GenBank (https://www.ncbi.nlm.nih.gov/genbank/ (accessed on 30 May 2021); Table
Abbreviations:
NYNU Microbiology Lab, Nanyang Normal University, Henan, China;
T type strain.
The sequences obtained in this study and the reference sequences downloaded from GenBank (Table
Sequences used in molecular phylogenetic analysis. Entries in bold are newly generated in this study.
Species | Strain no. | Locality | Sample | GenBank accession numbers | |
---|---|---|---|---|---|
ITS | D1/D2 | ||||
Candida aaseri | CBS 1913T | Norway | Sputum | AY821838 | U45802 |
C. amphixiae | CBS 9877T | Panama | Beetle | EU491501 | AY520327 |
C. andamanensis | CBS 10859T | Thailand | Estuarine water | AB525239 | AB334210 |
C. atlantica | CBS 5263T | Portugal | Shrimp egg | AJ539368 | U45799 |
C. atmosphaerica | CBS 4547T | Spain | Atmosphere | AJ539369 | U45779 |
C. blattariae | CBS 9876T | Panama | Cockroach | FJ715435 | AY640213 |
C. buinensis | CBS 6796T | Papua New Guinea | Gelatinous exudate | HQ283376 | U45778 |
C. cerambycidarum | CBS 9879T | Panama | Beetle | AY964669 | AY520299 |
C. conglobata | CBS 2018T | – | Tubercular lung | AJ539370 | U45789 |
C. dendronema | CBS 6270T | South Africa | Frass | HQ283365 | U45751 |
C. diddensiae | CBS 2214T | USA | Shrimp | AY580315 | U45750 |
C. diospyri | CBS 9769T | China | Kaki fruit | AY450919 | AY450918 |
C. endomychidarum | CBS 9881T | Panama | Beetle | AY964672 | AY520330 |
C. friedrichii | CBS 4114T | Germany | D-glucitol solution | HQ283377 | U45781 |
C. germanica | CBS 4105T | Germany | Atmosphere | HQ283366 | AF245401 |
C. gorgasii | CBS 9880T | Panama | Beetle | AY964670 | AY520300 |
C. insectorum | CBS 6213T | South Africa | Frass | HQ283372 | U45791 |
C. insectorum | NYNU 1672 | China | Rotten wood | MZ314279 | MZ314278 |
C. jaroonii | CBS 10790T | Thailand | Frass | AB360437 | DQ404493 |
C. kanchanaburiensis | CBS 11266T | Thailand | Mushroom | NR_137581 | KY106534 |
C. keroseneae | CECT 13058T | UK | Aircraft fuel | FJ235128 | FJ357698 |
C. khao-thaluensis | CBS 8535T | Thailand | Leaf | HQ283374 | HQ283383 |
C. koratica | CBS 10789T | Thailand | Frass | AB360443 | AB354232 |
C. lessepsii | CBS 9941T | Panama | Beetle | AY964671 | AY640214 |
C. membranifaciens | CBS 1952T | India | Urine | AJ606465 | U45792 |
C. michaelii | CBS 9878T | Panama | Beetle | AY964673 | AY520329 |
C. naeodendra | CBS 6032T | South Africa | Frass | AY580316 | U45759 |
C. oceani | CBS 11857T | Atlantic Ocean | Deep-sea coral | NR_156008 | GU002284 |
C. pseudoaaseri | CBS 11170T | Germany | Blood culture | JN241686 | JN241689 |
C. sinolaborantium | CBS 9940T | Panama | Beetle | NR_111343 | NG_055206 |
C. songkhlaensis | CBS 10791T | Thailand | Frass | AB360438 | DQ404499 |
C. spencermartinsiae | CBS 10894T | Seawater | Florida | FJ008050 | FJ008044 |
C. tallmaniae | CBS 8575T | French Guiana | Flower | HQ283378 | HQ283385 |
C. tammaniensis | CBS 8504T | USA | Frass | HQ283375 | AF017243 |
C. taylori | CBS 8508T | Belize | Sea water | FJ008051 | FJ008045 |
C. temnochilae | CBS 9938T | Panama | Beetle | AY964678 | AY242344 |
C. trypodendroni | CBS 8505T | Canada | Beetle | FJ153212 | AF017240 |
C. vaughaniae | CBS 8583T | French Guiana | Flower | HQ283364 | HQ283381 |
C. vrieseae | CBS 10829T | Brazil | Bromeliad | FJ755905 | EU200785 |
Yamadazyma akitaensis | CBS 6701T | Japan | Exudate | DQ409164 | U45766 |
Y. akitaensis | NYNU 16719 | China | Rotten wood | MZ314281 | MZ314280 |
Y. barbieri | CBS 14301T | Brazil | Sea water | LT547714 | LT547716 |
Y. cocois |
|
Vietnam | Fruits of the coconut palm | MN764369 | MN764369 |
Y. dushanensis | CBS 13914T | China | Rotten wood | KM272249 | KM272248 |
Y. endophytica | CBS 14163T | Thailand | Corn leaf | KT307981 | KT307981 |
Y. epiphylla | CBS 13384T | Thailand | Rice leaf | LC006082 | LC006026 |
Y. insecticola | CBS 13382T | Thailand | Frass | LC006081 | DQ400379 |
Y. kitorensis | CBS 14158T | Japan | Red viscous gel | LC060995 | LC060995 |
Y. laniorum | CBS 14780T | USA | Bark | KY588337 | KY588136 |
Y. luoyangensis | NYNU 201023T | China | Rotting wood | MW365549 | MW365545 |
Y. luoyangensis | NYNU 201035 | China | Rotting wood | MZ318445 | MZ318422 |
Y. mexicana | CBS 7066T | Mexico | Agria cactus | AB054110 | U45797 |
Y. nakazawae | CBS 6700T | Japan | Exudate | EU343867 | U45748 |
Y. olivae | CBS 11171T | Greece | Fermenting olive | FJ715432 | FJ715430 |
Y. olivae | NYNU 167106 | China | Rotting wood | MZ314288 | MZ314282 |
Y. olivae | NYNU 209116 | China | Rotting wood | MZ318443 | MZ318444 |
Y. ovata | NYUN 191125T | China | Rotting wood | MT990560 | MT990559 |
Y. ovata | NYUN 19130 | China | Rotting wood | MZ318424 | MZ318425 |
Y. ovata | NYUN 19116 | China | Rotting wood | MZ318442 | MZ318423 |
Y. paraaseri | NYNU 1811114T | China | Rotting wood | MK682794 | MK682805 |
Y. paraaseri | NYNU 181033 | China | Rotting wood | MZ318421 | MZ318460 |
Y. paraphyllophila | CBS 9928T | China, Taiwan | Pencil wood leaf | AY559447 | AY562397 |
Y. philogaea | CBS 6696T | South Africa | Soil | AB054107 | U45765 |
Y. phyllophila | CBS 12572T | Thailand | Corn leaf | AB734050 | AB734047 |
Y. riverae | CBS 14121T | Brazil | Rotting wood | KP900044 | KP900043 |
Y. scolyti | CBS 4802T | USA | Frass | EU343807 | U45788 |
Y. siamensis | CBS 12573T | Thailand | Sugarcane leaf | AB734049 | AB734046 |
Y. takamatsuzukensis | CBS 10916T | Japan | Air | AB365470 | AB365470 |
Y. tenuis | CBS 615T | Russia | Beetle | HQ283371 | U45774 |
Y. terventina | CBS 12510T | Italy | Olive oil | JQ247717 | JQ247717 |
Y. triangularis | CBS 4094T | Japan | Tamari soya | EU343869 | U45796 |
Y. tumulicola | CBS 10917T | Japan | Stone chamber | AB365463 | AB365463 |
Y. ubonensis | CBS 12859T | Thailand | Tree bark | NR_155998 | AB759913 |
Scheffersomyces coipomoensis | CBS 8178T | – | – | NR_111424 | U45747 |
Babjeviella inositovora | CBS 8006T | – | – | NR_111018 | U45848 |
MP analysis was run using a heuristic search option of 1,000 search replicates with random-addition of sequences and tree bisection and reconnection (TBR) as the branch-swapping algorithm. Gaps were treated as missing data. Bootstrapping with 1,000 replicates was performed to determine branch support (
The alignment based on the combined nuclear dataset (ITS and D1/D2 LSU) included 65 taxa and two outgroup taxa (Scheffersomyces coipomoensis and Babjeviella inositovora), and was comprised of 1,103 characters including gaps (576 for ITS and 527 for D1/D2 LSU) in the aligned matrix. Of these characters, 351 were constant, 455 variable characters were parsimony-uninformative, and 297 characters were parsimony-informative. The heuristic search using MP analysis generated the most parsimonious tree (TL = 979, CI = 0.297, RI = 0.653, RC = 0.248). The best model applied in the ML analysis was GTR+I+G. The ML analysis yielded a best scoring tree with a final optimization likelihood value of –11,006.61. Both methods for phylogenetic tree inference resulted in a similar topology. Therefore, only the best scoring PhyML tree is shown with BS and BT values simultaneously in Figure
Maximum likelihood phylogenetic tree of Yamadazyma inferred from the combined ITS and D1/D2 LSU dataset and rooted with Scheffersomyces coipomoensis (CBS 8178) and Babjeviella inositovora (CBS 8006). ML and MP bootstrap support values above 50% are respectively shown at the first and second positions. Newly sequenced collections are in blue boldface.
According to the phylogenetic tree (Figure
Physiological characteristics of the new Yamadazyma species and their closely related taxa.
Characteristics | Y. luoyangensis | Y. mexicana | Y. ovata | C. trypodendroni | Y. paraaseri | C. aaseri |
---|---|---|---|---|---|---|
Fermentation of | ||||||
d-Glucose | + | + | + | + | - | v |
Assimilation of | ||||||
l-Sorbose | – | – | + | – | + | s |
d-Glucosamine | + | + | + | – | + | – |
l-Rhamnose | + | + | – | + | – | – |
Melibiose | – | v | + | – | – | – |
Lactose | – | + | – | – | + | v |
Raffinose | – | + | – | – | – | – |
Inulin | + | – | – | – | + | – |
Xylitol | + | + | – | – | + | |
Galactitol | + | v | + | – | – | – |
2-Keto-d-Gluconate | – | s | + | + | – | – |
Cadaverine | – | n | – | + | – | + |
Growth tests | ||||||
10%Nacl/5%glucose | + | + | + | v | – | + |
Growth at 37 °C | – | + | + | – | + | + |
China, Henan Province, Luoyang City, Song County, in rotting wood from a forest park, September 2020, J.Z. Li & Z.T Zhang (holotype NYNU 201023T, culture ex-type CBS 16666, CICC 33509).
The species name luoyangensis refers to the geographical origin of the type strain of this species.
The cells are ovoid to ellipsoid (2–4 × 3.5–7 μm) and occur singly or in pairs after being placed in YM broth for three days at 25 °C (Figure
China, Henan Province, Luoyang City, Song County, in rotting wood from a forest park, September 2020, J.Z. Li & Z.T Zhang, NYNU 201035.
Two isolates representing Y. luoyangensis were resolved in a well-supported clade and are most closely related to Y. mexicana (Figure
China, Henan Province, Luoyang City, Song County, in rotting wood from a forest park, September 2019, J.Z. Li & Z.T Zhang (holotype NYNU 191125T, culture ex-type CBS 16655, CICC 33500).
The species name ovata refers to the ovoid cell morphology of the type strain.
The cells are ovoid to ellipsoid (2–3 × 3–6.5 μm) and occur singly or in pairs after growth in a YM broth for three days at 25 °C (Figure
China, Henan Province, Luoyang City, Song County, in rotting wood from a forest park, September 2019, J.Z. Li & Z.T Zhang, NYNU 19116, NYNU 19130.
We generated sequences for three isolates of Y. ovata, NYNU 191125, NYNU 19116, and NYNU 19130. This new species is phylogenetically most closely related to C. trypodendroni (Figure
China, Yunnan Province, Jinghong City, Mengyang Town, in rotting wood from a tropical rainforest, July 2018, K.F. Liu & Z.W. Xi (holotype NYNU 1811114T, culture ex-type CBS 16010, CICC 33365).
The species name paraaseri refers to its phylogenetic similarity to C. aaseri.
The cells are ovoid to elongate (2–2.5 × 3–8.5 μm) and occur singly or in pairs after being placed in YM broth for three days at 25 °C (Figure
China, Yunnan Province, Jinghong City, Mengyang Town, in rotting wood from a tropical rainforest, July 2018, K.F. Liu & Z.W. Xi, NYNU 181033.
Two strains representing Y. paraaseri were clustered in a well-supported clade and were phylogenetically related to C. aaseri [7]. Yamadazyma paraaseri can be distinguished from C. aaseri based on ITS and D1/D2 LSU loci (8/573 in ITS and 8/531 in D1/D2 LSU). Physiologically, the ability to assimilate d-glucosamine and inulin and the inability to assimilate xylitol and d-glucono-1, 5-lactone are the primary differences between Y. paraaseri and its closest relative, C. aaseri. Additionally, C. aaseri can grow in 10% NaCl with 5% glucose, while Y. paraaseri cannot (Table
In this work, six Yamadazyma species were identified based on morphology and molecular phylogeny. All species were isolated from rotting wood collected in Henan and Yunnan Provinces, China. Yamadazyma luoyangensis, Y. ovata, and Y. paraaseri are proposed as new species in Yamadazyma due to their well-supported phylogenic positions and distinctive physiological traits. Also, three known species of Yamadazyma, Y. insectorum, Y. akitaensis, and Y. olivae, were clearly identified by both morphological and molecular approaches.
In the past, methods of species identification of Yamadazyma were based only on morphology and physiological characters such as the shape of ascospores and reactions in standard growth and fermentation tests (
Yamadazyma species have a worldwide distribution and are isolated from diverse substrates. They can be found in flowers, leaves, fruits, tree bark, mushrooms, sea water, mineral and atmosphere, but most known species appear to exist in rotting wood, insects and their resulting frass (
We sincerely thank Dr. Jing-Zhao Li, Dr. Zheng-Tian Zhang, Dr. Kai-Fang Liu, and Dr. Zhi-Wen Xi for their help with collecting specimens. This project was supported by Grant No. 31570021 from the National Natural Science Foundation of China, China, No. 2018001 from the State Key Laboratory of Motor Vehicle Biofuel Technology, Henan Tianguan Enterprise Group Co., Ltd., China.