Phylogenetic and functional analyses of N6-methyladenosine RNA methylation factors in the wheat scab fungus Fusarium graminearum.

In eukaryotes, N6-methyladenosine (m6A) RNA modification plays a crucial role in governing the fate of RNA molecules and has been linked to various developmental processes. However, the phyletic distribution and functions of genetic factors responsible for m6A modification remain largely unexplored in fungi. To get insights into the evolution of m6A machineries, we reconstructed global phylogenies of potential m6A writers, readers, and erasers in fungi. Substantial copy number variations were observed, ranging from up to five m6A writers in early-diverging fungi to a single copy in the subphylum Pezizomycotina, which primarily comprises filamentous fungi. To characterize m6A factors in a phytopathogenic fungus Fusarium graminearum, we generated knockout mutants lacking potential m6A factors including the sole m6A writer MTA1. However, the resulting knockouts did not exhibit any noticeable phenotypic changes during vegetative and sexual growth stages. As obtaining a homozygous knockout lacking MTA1 was likely hindered by its essential role, we generated MTA1-overexpressing strains (MTA1-OE). The MTA1-OE5 strain showed delayed conidial germination and reduced hyphal branching, suggesting its involvement during vegetative growth. Consistent with these findings, the expression levels of MTA1 and a potential m6A reader YTH1 were dramatically induced in germinating conidia, followed by the expression of potential m6A erasers at later vegetative stages. Several genes including transcription factors, transporters, and various enzymes were found to be significantly upregulated and downregulated in the MTA1-OE5 strain. Overall, our study highlights the functional importance of the m6A methylation during conidial germination in F. graminearum and provides a foundation for future investigations into m6A modification sites in filamentous fungi.IMPORTANCEN6-methyladenosine (m6A) RNA methylation is a reversible posttranscriptional modification that regulates RNA function and plays a crucial role in diverse developmental processes. This study addresses the knowledge gap regarding phyletic distribution and functions of m6A factors in fungi. The identification of copy number variations among fungal groups enriches our knowledge regarding the evolution of m6A machinery in fungi. Functional characterization of m6A factors in a phytopathogenic filamentous fungus Fusarium graminearum provides insights into the essential role of the m6A writer MTA1 in conidial germination and hyphal branching. The observed effects of overexpressing MTA1 on fungal growth and gene expression patterns of m6A factors throughout the life cycle of F. graminearum further underscore the importance of m6A modification in conidial germination. Overall, this study significantly advances our understanding of m6A modification in fungi, paving the way for future research into its roles in filamentous growth and potential applications in disease control.

Transcriptional Divergence Underpinning Sexual Development in the Fungal Class Sordariomycetes.

Gene expression divergence through evolutionary processes is thought to be important for achieving programmed development in multicellular organisms. To test this premise in filamentous fungi, we investigated transcriptional profiles of 3,942 single-copy orthologous genes (SCOGs) in five related sordariomycete species that have morphologically diverged in the formation of their flask-shaped perithecia. We compared expression of the SCOGs to inferred gene expression levels of the most recent common ancestor of the five species, ranking genes from their largest increases to smallest increases in expression during perithecial development in each of the five species. We found that a large proportion of the genes that exhibited evolved increases in gene expression were important for normal perithecial development in Fusarium graminearum. Many of these genes were previously uncharacterized, encoding hypothetical proteins without any known functional protein domains. Interestingly, the developmental stages during which aberrant knockout phenotypes appeared largely coincided with the elevated expression of the deleted genes. In addition, we identified novel genes that affected normal perithecial development in Magnaporthe oryzae and Neurospora crassa, which were functionally and transcriptionally diverged from the orthologous counterparts in F. graminearum. Furthermore, comparative analysis of developmental transcriptomes and phylostratigraphic analysis suggested that genes encoding hypothetical proteins are generally young and transcriptionally divergent between related species. This study provides tangible evidence of shifts in gene expression that led to acquisition of novel function of orthologous genes in each lineage and demonstrates that several genes with hypothetical function are crucial for shaping multicellular fruiting bodies. IMPORTANCE The fungal class Sordariomycetes includes numerous important plant and animal pathogens. It also provides model systems for studying fungal fruiting body development, as its members develop fruiting bodies with a few well-characterized tissue types on common growth media and have rich genomic resources that enable comparative and functional analyses. To understand transcriptional divergence of key developmental genes between five related sordariomycete fungi, we performed targeted knockouts of genes inferred to have evolved significant upward shifts in expression. We found that many previously uncharacterized genes play indispensable roles at different stages of fruiting body development, which have undergone transcriptional activation in specific lineages. These novel genes are predicted to be phylogenetically young and tend to be involved in lineage- or species-specific function. Transcriptional activation of genes with unknown function seems to be more frequent than ever thought, which may be crucial for rapid adaption to changing environments for successful sexual reproduction.

Effect of Isolation Conditions on Diversity of Endolichenic Fungal Communities from a Foliose Lichen, Parmotrema tinctorum.

Endolichenic fungi (ELF) are emerging novel bioresources because their diverse secondary metabolites have a wide range of biological activities. Metagenomic analysis of lichen thalli demonstrated that the conventional isolation method of ELF covers a very limited range of ELF, and the development of an advanced isolation method is needed. The influence of four variables were investigated in this study to determine the suitable conditions for the isolation of more diverse ELF from a radially growing foliose lichen, Parmotrema tinctorum. Four variables were tested: age of the thallus, severity of surface-sterilization of the thallus, size of a thallus fragment for the inoculation, and nutrient requirement. In total, 104 species (1885 strains) of ELF were isolated from the five individual thalli of P. tinctorum collected at five different places. Most of the ELF isolates belong to Sordariomycetes. Because each part of lichen thallus (of different age) has unique ELF species, the whole thallus of the foliose lichen is needed to isolate diverse ELF. Moderate sterilization is appropriate for the isolation of diverse ELF. Inoculation of small fragment (1 mm2) of lichen thallus resulted in the isolation of highest diversity of ELF species compared to larger fragments (100 and 25 mm2). Moreover, ELF species isolated from the small thallus fragments covered all ELF taxa detected from the medium and the large fragments in this study. The use of two media-Bold's basal medium (nutrient poor) and potato dextrose agar (nutrient rich)-supported the isolation of diverse ELF. Among the tested variables, size of thallus fragment more significantly influenced the isolation of diverse ELF than other three factors. Species composition and richness of ELF communities from different lichen thalli differed from each other in this study.

Identification of Headspace Volatile Compounds of Blended Coffee and Application to Principal Component Analysis.

Coffee can be blended to create a variety of products to meet consumer's needs. In order to uncover the blending effect of coffee beans, we performed an experiment using principal component analysis (PCA). Twelve varieties of green beans were tested in 11 experimental groups, and the volatile compounds of the beans were analyzed. A total of 41 volatile compounds were identified. PCA was performed on 13 compounds that had a low odor threshold value or a high concentration among the identified compounds. PCA of total volatile compounds showed that principal component (PC) 1 and PC2 were extracted within 80% cumulative dispersion level. In PC1 and PC2, furfuryl alcohol and formic acid ethyl ester showed the greatest positive correlation coefficients among all the volatile compounds. The largest negative correlation coefficients in PC1 and PC2 were 4-hydroxy-2-butanone and 3-(ethylthio)propanal, respectively. Using PCA of the major volatile compounds in coffee, propanal and 1-methylpyrrole were found to have the largest positive correlation coefficients in PC1 and PC2, respectively. In the score plot of the major volatile components, 4 kinds of blended coffee were closely grouped, therefore showing similar aroma qualities. However, 5 kinds of other blended coffees showed a positive correlation with PC2. This is probably due to 3-(ethylthio)propanal acting as a specific value. The application of statistical methods to blended coffee allows for logical and systematic data analysis of data and may be used as a basis for quality evaluation.

Short- and Long-term Effects of Case Management on Suicide Prevention among Individuals with Previous Suicide Attempts: a Survival Analysis.

BACKGROUND: This study examined the long-term preventive effects of a case management service on suicide reattempts, and clarified the factors related to suicide reattempts. METHODS: We reviewed the medical records of suicide attempters who visited the emergency department of Ulsan University Hospital from August 28, 2013 to July 31, 2017. A 4-week case management service was provided to consenting participants, either face-to-face or by telephone. Using survival analysis, we analyzed differences in the time to the next emergency department visit for a suicide attempt according to whether participants completed the case management service. We also assessed which characteristics of participants were associated with suicide reattempts. RESULTS: We found no overall difference in time to suicide reattempt between case-managed participants and controls over the entire observation period (median period: 19 months). However, in the first 24 weeks after the initial suicide attempt, the case-managed group showed a longer time to reattempt than did the control group (log-rank test = 4.243; P = 0.039). A higher risk of reattempt was found among participants with the medical benefit type of health insurance compared to those with national health insurance (hazard ratio [HR], 5.134; P < 0.001) and among participants aged 20-39 compared to those aged ≥ 60 (HR, 3.502; P = 0.05). CONCLUSION: Case management had only short-term benefits (within 24 weeks of initial suicide attempt). Risk factors for suicide reattempts were having a medical benefit health insurance and being aged 20-39 years.