Agrobacterium tumefaciens-Mediated Transformation of the Aquatic Fungus Phialemonium inflatum FBCC-F1546.

Phialemonium inflatum is a useful fungus known for its ability to mineralise lignin during primary metabolism and decompose polycyclic aromatic hydrocarbons (PAHs). However, no functional genetic analysis techniques have been developed yet for this fungus, specifically in terms of transformation. In this study, we applied an Agrobacterium tumefaciens-mediated transformation (ATMT) system to P. inflatum for a functional gene analysis. We generated 3689 transformants using the binary vector pSK1044, which carried either the hygromycin B phosphotransferase (hph) gene or the enhanced green fluorescent protein (eGFP) gene to label the transformants. A Southern blot analysis showed that the probability of a single copy of T-DNA insertion was approximately 50% when the co-cultivation of fungal spores and Agrobacterium tumefaciens cells was performed at 24-36 h, whereas at 48 h, it was approximately 35.5%. Therefore, when performing gene knockout using the ATMT system, the co-cultivation time was reduced to ≤36 h. The resulting transformants were mitotically stable, and a PCR analysis confirmed the genes' integration into the transformant genome. Additionally, hph and eGFP gene expressions were confirmed via PCR amplification and fluorescence microscopy. This optimised transformation system will enable functional gene analyses to study genes of interest in P. inflatum.

Arthonia dokdoensis and Rufoplaca toktoana - Two New Taxa from Dokdo Islands (South Korea).

Arthonia dokdoensis sp. nov., a lichenicolous fungus from the subcosmopolitan Arthonia molendoi complex growing on crustose thalli of species of the genus Orientophila (subfamily Xanthorioideae, Teloschistaceae), as well as the lichen species Rufoplaca toktoana sp. nov. (subfamily Caloplacoideae, Teloschistaceae) similar to Rufoplaca kaernefeltiana, both from Dokdo Islands, Republic of Korea, are described, illustrated, and compared with closely related taxa. In the phylogenetic tree of the Arthoniaceae based on 12S mtSSU and RPB2 gene sequences, the phylogenetic position of the A. dokdoensis and the relationship with the A. molendoi group are illustrated, while the position of the newly described R. toktoana is confirmed by phylogenetic tree based on ITS nrDNA data.

Arthothelium punctatum (Arthoniaceae, Arthoniales), A New Lichen Species from South Korea.

A total of 121 species of lichens belonging to the genus Arthothelium have been described to date, most of which have been found in tropical regions. Here, we describe the discovery of a novel Arthothelium species for the first time in South Korea. Until now, Arthothelium ruanum was the only Arthothelium species reported in South Korea. Among the 113 specimens collected in this study, we identified A. ruanum and a putative new species, Arthothelium punctatum (J. S. Park & J.-S. Hur, sp. nov.). The diagnostic characters of A. punctatum are as follows: apothecia punctate, shortly elongate to branched, small, 0.1-0.2 mm wide, hypothecium hyaline to pale brown and obovate to broadly ellipsoid, muriform ascospores, 29.5-44.6 × 12.2-18.2 µm. The new species was found in Mt. Seokbyeong at an altitude of 790 m on smooth bark. Upon phylogenic analysis, the putative new species, A. punctatum, was separated from other Arthothelium species although the specimens analyzed were clustered with Arthoniaceae in phylogenetic trees based on both the mitochondrial small subunit (mtSSU) sequence and combined mtSSU and nuclear ribosomal large subunit sequences. Our data clearly indicate that this species is a new species belonging to the family Arthoniaceae. To elucidate the taxonomic characteristics of the new species, we provide morphological descriptions and a distribution map.

Three New Monotypic Genera of the Caloplacoid Lichens (Teloschistaceae, Lichen-Forming Ascomycetes).

Three monophyletic branches are strongly supported in a phylogenetic analysis of the Teloschistaceae based on combined data sets of internal transcribed spacer and large subunit nrDNA and 12S small subunit mtDNA sequences. These are described as new monotypic genera: Jasonhuria S. Y. Kondr., L. Lokös et S. -O. Oh, Loekoesia S. Y. Kondr., S. -O. Oh et J. -S. Hur and Olegblumia S. Y. Kondr., L. Lokös et J. -S. Hur. Three new combinations for the type species of these genera are proposed.

Notes on the lichen genus leptogium (collemataceae, ascomycota) in South Korea.

Leptogium (Ach.) Gray is distributed throughout South Korea; however, for nearly two decades no detailed taxonomic or revisionary research on this lichen genus has been conducted. This study examined the specimens deposited in the lichen herbarium at the Korean Lichen Research Institute, and samples were identified using descriptions recently published in the scientific literature. In this revisionary study, a total of fourteen species of Leptogium were documented, including new records of Leptogium delavayi Hue, Leptogium denticulatum Nyl., and Leptogium trichophoroides P. M. Jørg. & A. K. Wallace. Detailed descriptions of each species are given, including their morphological, anatomical, and chemical characteristics. A key to all Leptogium species known to occur in South Korea is also presented.

An easy, rapid, and cost-effective method for DNA extraction from various lichen taxa and specimens suitable for analysis of fungal and algal strains.

Lichen studies, including biodiversity, phylogenetic relationships, and conservation concerns require definitive species identification, however many lichens can be challenging to identify at the species level. Molecular techniques have shown efficacy in discriminating among lichen taxa, however, obtaining genomic DNA from herbarium and fresh lichen thalli by conventional methods has been difficult, because lichens contain high proteins, polysaccharides, and other complex compounds in their cell walls. Here we report a rapid, easy, and inexpensive protocol for extracting PCR-quality DNA from various lichen species. This method involves the following two steps: first, cell breakage using a beadbeater; and second, extraction, isolation, and precipitation of genomic DNA. The procedure requires approximately 10 mg of lichen thalli and can be completed within 20 min. The obtained DNAs were of sufficient quality and quantity to amplify the internal transcribed spacer region from the fungal and algal lichen components, as well as to sequence the amplified products. In addition, 26 different lichen taxa were tested, resulting in successful PCR products. The results of this study validated the experimental protocols, and clearly demonstrated the efficacy and value of our KCl extraction method applied in the fungal and algal samples.