Bacteria can maintain rRNA operons solely on plasmids for hundreds of millions of years.

It is generally assumed that all bacteria must have at least one rRNA operon (rrn operon) on the chromosome, but some strains of the genera Aureimonas and Oecophyllibacter carry their sole rrn operon on a plasmid. However, other related strains and species have chromosomal rrn loci, suggesting that the exclusive presence of rrn operons on a plasmid is rare and unlikely to be stably maintained over long evolutionary periods. Here, we report the results of a systematic search for additional bacteria without chromosomal rrn operons. We find that at least four bacterial clades in the phyla Bacteroidota, Spirochaetota, and Pseudomonadota (Proteobacteria) lost chromosomal rrn operons independently. Remarkably, Persicobacteraceae have apparently maintained this peculiar genome organization for hundreds of millions of years. In our study, all the rrn-carrying plasmids in bacteria lacking chromosomal rrn loci possess replication initiator genes of the Rep_3 family. Furthermore, the lack of chromosomal rrn operons is associated with differences in copy numbers of rrn operons, plasmids, and chromosomal tRNA genes. Thus, our findings indicate that the absence of rrn loci in bacterial chromosomes can be stably maintained over long evolutionary periods.

Chromosome-level genome assemblies of Cutaneotrichosporon spp. (Trichosporonales, Basidiomycota) reveal imbalanced evolution between nucleotide sequences and chromosome synteny.

BACKGROUND: Since DNA information was first used in taxonomy, barcode sequences such as the internal transcribed spacer (ITS) region have greatly aided fungal identification; however, a barcode sequence alone is often insufficient. Thus, multi-gene- or whole-genome-based methods were developed. We previously isolated Basidiomycota yeasts classified in the Trichosporonales. Some strains were described as Cutaneotrichosporon cavernicola and C. spelunceum, whereas strain HIS471 remained unidentified. We analysed the genomes of these strains to elucidate their taxonomic relationship and genetic diversity. RESULTS: The long-read-based assembly resulted in chromosome-level draft genomes consisting of seven chromosomes and one mitochondrial genome. The genome of strain HIS471 has more than ten chromosome inversions or translocations compared to the type strain of C. cavernicola despite sharing identical ITS barcode sequences and displaying an average nucleotide identity (ANI) above 93%. Also, the chromosome synteny between C. cavernicola and the related species, C. spelunceum, showed significant rearrangements, whereas the ITS sequence identity exceeds 98.6% and the ANI is approximately 82%. Our results indicate that the relative evolutionary rates of barcode sequences, whole-genome nucleotide sequences, and chromosome synteny in Cutaneotrichosporon significantly differ from those in the model yeast Saccharomyces. CONCLUSIONS: Our results revealed that the relative evolutionary rates of nucleotide sequences and chromosome synteny are different among fungal clades, likely because different clades have diverse mutation/repair rates and distinct selection pressures on their genomic sequences and syntenic structures. Because diverse syntenic structures can be a barrier to meiotic recombination and may lead to speciation, the non-linear relationships between nucleotide and synteny diversification indicate that sequence-level distances at the barcode or whole-genome level are not sufficient for delineating species boundaries.

Hyphal Growth in Trichosporon asahii Is Accelerated by the Addition of Magnesium.

Fungal dimorphism involves two morphologies: a unicellular yeast cell and a multicellular hyphal form. Invasion of hyphae into human cells causes severe opportunistic infections. The transition between yeast and hyphal forms is associated with the virulence of fungi; however, the mechanism is poorly understood. Therefore, we aimed to identify factors that induce hyphal growth of Trichosporon asahii, a dimorphic basidiomycete that causes trichosporonosis. T. asahii showed poor growth and formed small cells containing large lipid droplets and fragmented mitochondria when cultivated for 16 h in a nutrient-deficient liquid medium. However, these phenotypes were suppressed via the addition of yeast nitrogen base. When T. asahii cells were cultivated in the presence of different compounds present in the yeast nitrogen base, we found that magnesium sulfate was a key factor for inducing cell elongation, and its addition dramatically restored hyphal growth in T. asahii. In T. asahii hyphae, vacuoles were enlarged, the size of lipid droplets was decreased, and mitochondria were distributed throughout the cell cytoplasm and adjacent to the cell walls. Additionally, hyphal growth was disrupted due to treatment with an actin inhibitor. The actin inhibitor latrunculin A disrupted the mitochondrial distribution even in hyphal cells. Furthermore, magnesium sulfate treatment accelerated hyphal growth in T. asahii for 72 h when the cells were cultivated in a nutrient-deficient liquid medium. Collectively, our results suggest that an increase in magnesium levels triggers the transition from the yeast to hyphal form in T. asahii. These findings will support studies on the pathogenesis of fungi and aid in developing treatments. IMPORTANCE Understanding the mechanism underlying fungal dimorphism is crucial to discern its invasion into human cells. Invasion is caused by the hyphal form rather than the yeast form; therefore, it is important to understand the mechanism of transition from the yeast to hyphal form. To study the transition mechanism, we utilized Trichosporon asahii, a dimorphic basidiomycete that causes severe trichosporonosis since there are fewer studies on T. asahii than on ascomycetes. This study suggests that an increase in Mg2+, the most abundant mineral in living cells, triggers growth of filamentous hyphae and increases the distribution of mitochondria throughout the cell cytoplasm and adjacent to the cell walls in T. asahii. Understanding the mechanism of hyphal growth triggered by Mg2+ increase will provide a model system to explore fungal pathogenicity in the future.

Taxonomy of Pathogenic Yeasts Candida, Cryptococcus, Malassezia, and Trichosporon.

This review describes the changes in yeast species names in the previous decade. Several yeast species have been reclassified to accommodate the "One fungus=One name" (1F=1N) principle of the Code. As the names of medically important yeasts have also been reviewed and revised, details of the genera Candida, Cryptococcus, Malassezia, and Trichosporon are described in Section 3, along with the history of name changes. Since the phylogenetic positions of Candida species in several clades have not been clarified, revision of this species has not been completed. Among the species that remain unrevised despite their importance in the medical field, we propose the transfer of six Candida species to be reclassified in the Nakaseomyces clade, including Nakaseomyces glabratus and Nakaseomyces nivalensis.

[Malassezia Display a Hyphae-like "Spaghetti-and-Meatballs" Configuration in Keratotic Plugs].

Malassezia are lipophilic yeasts in the skin microbiome that abundantly colonize all parts of human skin except for the soles of the feet. Fungal microbiome analysis of keratotic plugs from the noses of 10 healthy individuals identified Malassezia restricta as the predominant species, followed by Malassezia globosa. Malassezia hyphae were observed in 5 of the 10 individuals. The hyphae were curved and thick-walled with spherical thick-walled and grouped blastoconidia, described as a "spaghetti-and-meatballs" configuration. In this study, we observed Malassezia hyphae in keratotic plugs of healthy subjects, although abundant Malassezia hyphae have previously only been observed in lesional sites of patients with pityriasis versicolor.

Nomenclatural issues concerning cultured yeasts and other fungi: why it is important to avoid unneeded name changes.

The unambiguous application of fungal names is important to communicate scientific findings. Names are critical for (clinical) diagnostics, legal compliance, and regulatory controls, such as biosafety, food security, quarantine regulations, and industrial applications. Consequently, the stability of the taxonomic system and the traceability of nomenclatural changes is crucial for a broad range of users and taxonomists. The unambiguous application of names is assured by the preservation of nomenclatural history and the physical organisms representing a name. Fungi are extremely diverse in terms of ecology, lifestyle, and methods of study. Predominantly unicellular fungi known as yeasts are usually investigated as living cultures. Methods to characterize yeasts include physiological (growth) tests and experiments to induce a sexual morph; both methods require viable cultures. Thus, the preservation and availability of viable reference cultures are important, and cultures representing reference material are cited in species descriptions. Historical surveys revealed drawbacks and inconsistencies between past practices and modern requirements as stated in the International Code of Nomenclature for Algae, Fungi, and Plants (ICNafp). Improper typification of yeasts is a common problem, resulting in a large number invalid yeast species names. With this opinion letter, we address the problem that culturable microorganisms, notably some fungi and algae, require specific provisions under the ICNafp. We use yeasts as a prominent example of fungi known from cultures. But viable type material is important not only for yeasts, but also for other cultivable Fungi that are characterized by particular morphological structures (a specific type of spores), growth properties, and secondary metabolites. We summarize potential proposals which, in our opinion, will improve the stability of fungal names, in particular by protecting those names for which the reference material can be traced back to the original isolate.

Draft Genome Sequence of Oleaginous Yeast Saitozyma sp. Strain JCM 24511, Isolated from Soil on Iriomote Island, Okinawa, Japan.

Here, we report draft genome sequence of oleaginous yeast strain Saitozyma sp. JCM 24511, which is phylogenetically closely related to Saitozyma podzolica These data will have implications not only for the study of the oleaginous activities of yeasts but also for the study of the plant-microorganism microbiome.

Description of four Apiotrichum and two Cutaneotrichosporon species isolated from guano samples from bat-inhabited caves in Japan.

Four new yeast species belonging to the genus Apiotrichum and two new yeast species belonging to Cutaneotrichosporon are described for strains isolated from guano samples from bat-inhabited caves in Japan. In 2005, we reported these isolates as Trichosporon species based on sequence analyses of the D1/D2 domain of large subunit (LSU) rRNA genes according to available basidiomycetous yeast classification criteria; however, to date, they have not been officially published as new species with descriptions. Their phylogenetic positions have been reanalysed based on comparison of internal transcribed spacer (ITS) region sequences (including the 5.8S rRNA gene) and the D1/D2 domain of the LSU rRNA gene with those of known species; we confirmed clear separation from previously described species. Physiological and biochemical properties of the isolates also suggest their distinctiveness. Therefore, we describe Apiotrichum akiyoshidainum (holotype JCM 12595T), Apiotrichum chiropterorum (JCM 12594T), Apiotrichum coprophilum (JCM 12596T), Apiotrichum otae (JCM 12593T), Cutaneotrichosporon cavernicola (JCM 12590T) and Cutaneotrichosporon middelhovenii (JCM 12592T) as new species. C. cavernicola showed particularly distinctive morphology including large inflated anomalous cells on the hyphae and germination from the cells, although clear clamp connections on the hyphae were not confirmed. Further study is needed to elucidate the morph of this species.

Takashi Nakase's last tweet: what is the current direction of microbial taxonomy research?

During the last few decades, type strains of most yeast species have been barcoded using the D1/D2 domain of their LSU rRNA gene and internal transcribed spacer (ITS) region. Species identification using DNA sequences regarding conspecificity in yeasts has also been studied. Most yeast species can be identified according to the sequence divergence of their ITS region or a combination of the D1/D2 and ITS regions. Studies that have examined intraspecific diversity have used multilocus sequence analyses, whereas the marker regions used in this analysis vary depending upon taxa. D1/D2 domain and ITS region sequences have been used as barcodes to develop primers suitable for the detection of the biological diversity of environmental DNA and the microbiome. Using these barcode sequences, it is possible to identify relative lineages and infer their gene products and function, and how they adapt to their environment. If barcode sequence was not variable enough to identify a described species, one could investigate the other biological traits of these yeasts, considering geological distance, environmental circumstances and isolation of reproduction. This article is dedicated to late Dr Takashi Nakase (1939-2018).

Heterocephalacria mucosa sp. nov., a new basidiomycetous yeast species isolated from a mangrove forest in Thailand.

Strains of yeast were isolated under a nitrogen-depleted culture condition from decaying tree bark (strain N-12.1) and from mangrove forest water (strain 1-7W.1) sampled at different locations within a mangrove forest site in Ranong province, Thailand. They were found to be genetically and phenotypically different from any currently recognised yeast species. Phylogenetic analysis of nucleotide sequence of three genes, the internal transcribed spacer region 1 and 2 plus 5.8S ribosomal RNA gene (ITS), D1/D2 domain of the large subunit ribosomal RNA gene (LSU D1/D2) and the small subunit of the ribosomal RNA gene (SSU), revealed that these two strains were related to but distinguished from Heterocephalacriaarrabidensis. Several distinct physiological characteristics of these two strains were detected, namely inability to assimilate glycerol, dl-lactic acid, succinic acid, citric acid, d-gluconic acid, and ability to grow well at 25 °C, which were different from those of H. arrabidensis. Accordingly, the name Heterocephalacria mucosa sp. nov., is proposed to accommodate this novel species. The MycoBank number is MB 828624.

Genome and transcriptome evolve separately in recently hybridized Trichosporon fungi.

Genome hybridization is an important evolutionary event that gives rise to species with novel capabilities. However, the merging of distinct genomes also brings together incompatible regulatory networks that must be resolved during the course of evolution. Understanding of the early stages of post-hybridization evolution is particularly important because changes in these stages have long-term evolutionary consequences. Here, via comparative transcriptomic analyses of two closely related, recently hybridized Trichosporon fungi, T. coremiiforme and T. ovoides, and three extant relatives, we show that early post-hybridization evolutionary processes occur separately at the gene sequence and gene expression levels but together contribute to the stabilization of hybrid genome and transcriptome. Our findings also highlight lineage-specific consequences of genome hybridization, revealing that the transcriptional regulatory dynamics in these hybrids responded completely differently to gene loss events: one involving both subgenomes and another that is strictly subgenome-specific.

Draft Genome Sequences of Basidiomycetous Epiphytic Phylloplane Yeast Type Strains Dioszegia crocea JCM 2961 and Dioszegia aurantiaca JCM 2956.

We report the draft genome sequences of type strains for Dioszegia crocea and its closely related species Dioszegia aurantiaca, which should improve our understanding of the epiphytic phylloplane yeasts. These data will also have implications for the plant microbiome, since Dioszegia is considered a microbial "hub" taxon.

Draft Genome Analysis of Trichosporonales Species That Contribute to the Taxonomy of the Genus Trichosporon and Related Taxa.

Many nomenclatural changes, including proposals of new taxa, have been carried out in fungi to adapt to the "One fungus = One name" (1F=1N) principle. In yeasts, while some changes have been made in response to 1F=1N, most have resulted from two other factors: i) an improved understanding of biological diversity due to an increase in number of known species, and ii) progress in the methods for analyzing and evaluating biological diversity. The method for constructing a backbone tree, which is a basal tree used to infer phylogeny, has also progressed from single-gene trees to multi-locus trees and further, to genome trees. This paper describes recent advances related to the contribution of genomic data to taxonomy, using the order Trichosporonales as an example.

Recognition and delineation of yeast genera based on genomic data: Lessons from Trichosporonales.

Delineation and characterization of genera in Trichosporonales (Agaricomycotina, Basidiomycota) was performed using 24 haploid and 3 naturally occurring hybrid genomes, with 3 Tremellales genomes used as outgroups. Orthologous group analysis of those genomes showed presence-absence patterns of orthologs that were consistent with the genus classifications. Many shared unique orthologs were identified in the well-supported lineages (genera Apiotrichum and Trichosporon), supporting the definitions of the genera Apiotrichum and Trichosporon from a genomic perspective. Specifically, we obtained 24 and 285 genus-specific genes from eight Apiotrichum and five Trichosporon species, respectively, and propose that these genus-specific genes can be used for delineation of those genera. On the other hand, the genus Cutaneotrichosporon shared only one genus-specific gene among eight genomes, indicating that this genus definition might require re-examination based on genomic data. In addition, taxonomic revisions are presented in this study, including the proposal of two genera, Pascua and Prillingera. Because genomic data can be systematically obtained and analyzed to compare species from a comprehensive viewpoint, they can be used not only to reconstruct reliable phylogenetic trees, but also to re-examine the definitions of taxonomic classifications. To our knowledge, this is the first report to discuss the 'natural system' of genus level classification in fungi based on genomic data.

Tempo and Mode of Genome Evolution in the Budding Yeast Subphylum.

Budding yeasts (subphylum Saccharomycotina) are found in every biome and are as genetically diverse as plants or animals. To understand budding yeast evolution, we analyzed the genomes of 332 yeast species, including 220 newly sequenced ones, which represent nearly one-third of all known budding yeast diversity. Here, we establish a robust genus-level phylogeny comprising 12 major clades, infer the timescale of diversification from the Devonian period to the present, quantify horizontal gene transfer (HGT), and reconstruct the evolution of 45 metabolic traits and the metabolic toolkit of the budding yeast common ancestor (BYCA). We infer that BYCA was metabolically complex and chronicle the tempo and mode of genomic and phenotypic evolution across the subphylum, which is characterized by very low HGT levels and widespread losses of traits and the genes that control them. More generally, our results argue that reductive evolution is a major mode of evolutionary diversification.

Lipid production via simultaneous conversion of glucose and xylose by a novel yeast, Cystobasidium iriomotense.

The yeast strains IPM32-16, ISM28-8sT, and IPM46-17, isolated from plant and soil samples from Iriomote Island, Japan, were explored in terms of lipid production during growth in a mixture of glucose and xylose. Phylogenetically, the strains were most closely related to Cystobasidium slooffiae, based on the sequences of the ITS regions and the D1/D2 domain of the LSU rRNA gene. The strains were oleaginous, accumulating lipids to levels > 20% dry cell weight. Moreover, kinetic analysis of the sugar-to-lipid conversion of a 1:1 glucose/xylose mixture showed that the strains consumed the two sugars simultaneously. IPM46-17 attained the highest lipid content (33%), mostly C16 and C18 fatty acids. Thus, the yeasts efficiently converted lignocellulosic sugars to lipids, aiding in biofuel production (which benefits the environment, promotes rural jobs, and strengthens fuel security). The strains constituted a novel species of Cystobasidium, for which we propose the name Cystobasidium iriomotense (type strain ISM28-8sT = JCM 24594T = CBS 15015T).

A Trichosporonales genome tree based on 27 haploid and three evolutionarily conserved 'natural' hybrid genomes.

To construct a backbone tree consisting of basidiomycetous yeasts, draft genome sequences from 25 species of Trichosporonales (Tremellomycetes, Basidiomycota) were generated. In addition to the hybrid genomes of Trichosporon coremiiforme and Trichosporon ovoides that we described previously, we identified an interspecies hybrid genome in Cutaneotrichosporon mucoides (formerly Trichosporon mucoides). This hybrid genome had a gene retention rate of ~55%, and its closest haploid relative was Cutaneotrichosporon dermatis. After constructing the C. mucoides subgenomes, we generated a phylogenetic tree using genome data from the 27 haploid species and the subgenome data from the three hybrid genome species. It was a high-quality tree with 100% bootstrap support for all of the branches. The genome-based tree provided superior resolution compared with previous multi-gene analyses. Although our backbone tree does not include all Trichosporonales genera (e.g. Cryptotrichosporon), it will be valuable for future analyses of genome data. Interest in interspecies hybrid fungal genomes has recently increased because they may provide a basis for new technologies. The three Trichosporonales hybrid genomes described in this study are different from well-characterized hybrid genomes (e.g. those of Saccharomyces pastorianus and Saccharomyces bayanus) because these hybridization events probably occurred in the distant evolutionary past. Hence, they will be useful for studying genome stability following hybridization and speciation events. Copyright © 2017 John Wiley & Sons, Ltd.

Current Status of Taxonomy of Pathogenic Yeasts.

Fungal taxonomy has been reconstructed on the basis of genome information, and new nomenclatural rules have been enacted from 2013. It has been proposed that Cryptococcus neoformans and Cryptococcus gattii be reclassified into two species (C. neoformans and Cryptococcus deneoformans) and five species (C. gattii, Cryptococcus bacillisporus, Cryptococcus deuterogattii, Cryptococcus tetragattii, and Cryptococcus decagattii), respectively. The genus Trichosporon has been reclassified into five genera. Trichosporon asahii, which is the causative agent of trichosporonosis, has been retained in the genus Trichosporon, while Trichosporon cutaneum has been transferred into a new genus, Cutaneotrichosporon.

Occultifur plantarum f.a., sp. nov., a novel cystobasidiomycetous yeast species.

Nine strains representing a single anamorphic novel yeast species in dispersed tropical and subtropical habitats were isolated from sugarcane leaf tissue (DMKU-SE24, DMKU-SE45T, DMKU-SE129 and DMKU-SE134) and corn leaf tissue (DMKU-CE36) in Thailand, phylloplane and rhizoplane of sugarcane in Brazil (IMUFRJ 52018 and IMUFRJ 52019), bromeliad leaf tissue in Brazil (IMUFRJ 51954) and plant leaf in Japan (IPM31-24). These strains showed identical or only 1 nt substitution in the sequences of the D1/D2 region of the LSU rRNA gene and 0-5 nt substitutions in the internal transcribed spacer (ITS) region. Phylogenetic analysis based on the combined sequences of the ITS and the D1/D2 regions showed that the eight of these strains represented a single species in the genus Occultifur that was distinct from related species. Occultifur kilbournensis was the most closely related species, but with 0.9-1.2 % nucleotide substitutions in the D1/D2 region of the LSU rRNA gene, and 2.4-2.6 % nucleotide substitutions in the ITS region. They are therefore considered to represent a novel species of the genus Occultifur although the formation of basidia was not observed. The name Occultifur plantarum f.a., sp. nov. is proposed. The type strain is DMKU-SE45T (=CBS 14554T=TBRC 6561T).