Oxford University Press (OUP) Acta Medica Okayama 2730-6151 5 1 2025 Methanol chemoreceptor MtpA- and flagellin protein FliC-dependent methylotaxis contributes to the spatial colonization of PPFM in the phyllosphere ycaf092 EN Shiori Katayama Graduate School of Agriculture, Kyoto University Kosuke Shiraishi Graduate School of Agriculture, Kyoto University Kanae Kaji Graduate School of Agriculture, Kyoto University Kazuya Kawabata Graduate School of Agriculture, Kyoto University Naoki Tamura Department of Anatomy and Histology, School of Medicine, Fukushima Medical University Akio Tani Institute of Plant Science and Resources, Okayama University Hiroya Yurimoto Graduate School of Agriculture, Kyoto University Yasuyoshi Sakai Graduate School of Agriculture, Kyoto University Pink-pigmented facultative methylotrophs (PPFMs) capable of growth on methanol are dominant and versatile phyllosphere bacteria that provide positive effects on plant growth through symbiosis. However, the spatial behavior of PPFMs on plant surfaces and its molecular basis are unknown. Here, we show that Methylobacterium sp. strain OR01 inoculated onto red perilla seeds colonized across the entire plant surface in the phyllosphere concomitant with the plant growth. During its transmission, strain OR01 was found to be present on the entire leaf surface with a preference to sites around the periphery, vein, trichome, and stomata. We found that methanol-sensing chemoreceptor MtpA-dependent chemotaxis (methylotaxis; chemotaxis toward methanol) and flagellin protein FliC-dependent motility facilitated the bacterial entry into the stomatal cavity and their colonization in the phyllosphere. No potential conflict of interest relevant to this article was reported.

Springer Science and Business Media LLC Acta Medica Okayama 0003-6072 118 10 2025 Duganella hordei sp. nov., Duganella caerulea sp. nov., and Duganella rhizosphaerae sp. nov., isolated from barley rhizosphere 146 EN Katsumoto Kishiro Institute of Plant Science and Resources, Okayama University Nurettin Sahin Egitim Fakultesi, Mugla Sitki Kocman University Daisuke Saisho Institute of Plant Science and Resources, Okayama University Naoki Yamaji Institute of Plant Science and Resources, Okayama University Jun Yamashita Institute of Plant Science and Resources, Okayama University Yuki Monden Graduate School of Environmental, Life, Natural Science and Technology, Okayama University Tomoyuki Nakagawa Faculty of Applied Biological Sciences, Gifu University Keiichi Mochida RIKEN Center for Sustainable Resource Science Akio Tani Institute of Plant Science and Resources, Okayama University Duganella sp. strains R1T, R57T, and R64T, isolated from barley roots in Japan, are Gram-stain-negative, motile, rod-shaped bacteria. Duganella species abundantly colonized barley roots. Strains R1T, R57T, and R64T were capable of growth at 4 °C, suggesting adaptation to colonize winter barley roots. Strains R57T and R64T formed purple colonies, indicating violacein production, while strain R1T did not. Based on 16S rRNA gene sequence similarities, strains R1T, R57T, and R64T were most closely related to D. violaceipulchra HSC-15S17T (99.10%), D. vulcania FT81WT (99.45%), and D. violaceipulchra HSC-15S17T (99.86%), respectively. Their genome sizes ranged from 7.05 to 7.38 Mbp, and their genomic G+C contents were 64.2–64.7%. The average nucleotide identity and digital DNA–DNA hybridization values between R1T and D. violaceipulchra HSC-15S17T, R57T and D. vulcania FT81WT, R64T and D. violaceipulchra HSC-15S17T were 86.0% and 33.2%, 95.7% and 67.9%, and 92.7% and 52.6%, respectively. Their fatty acids were predominantly composed of C16:0, C17:0 cyclo, and summed feature 3 (C16:1 ω7c and/or C16:1 ω6c). Based on their distinct genetic and phenotypic characteristics, and supported by chemotaxonomic analyses, we propose that strains R1T, R57T, and R64T represent novel species within the Duganella genus, for which the names Duganella hordei (type strain R1T = NBRC 115982 T = DSM 115069 T), Duganella caerulea (type strain R57T = NBRC 115983 T = DSM 115070 T), and Duganella rhizosphaerae (type strain R64T = NBRC 115984 T = DSM 115071 T) are proposed. No potential conflict of interest relevant to this article was reported.

Frontiers Media SA Acta Medica Okayama 1664-302X 13 2022 Siderophore for Lanthanide and Iron Uptake for Methylotrophy and Plant Growth Promotion in Methylobacterium aquaticum Strain 22A 921635 EN Patrick Otieno Juma Institute of Plant Science and Resources, Okayama University Yoshiko Fujitani Institute of Plant Science and Resources, Okayama University Ola Alessa Institute of Plant Science and Resources, Okayama University Tokitaka Oyama Graduate School of Science, Kyoto University Hiroya Yurimoto Graduate School of Agriculture, Kyoto University Yasuyoshi Sakai Graduate School of Agriculture, Kyoto University Akio Tani Institute of Plant Science and Resources, Okayama University Methylobacterium and Methylorubrum species are facultative methylotrophic bacteria that are abundant in the plant phyllosphere. They have two methanol dehydrogenases, MxaF and XoxF, which are dependent on either calcium or lanthanides (Lns), respectively. Lns exist as insoluble minerals in nature, and their solubilization and uptake require a siderophore-like substance (lanthanophore). Methylobacterium species have also been identified as plant growth-promoting bacteria although the actual mechanism has not been well-investigated. This study aimed to reveal the roles of siderophore in Methylobacterium aquaticum strain 22A in Ln uptake, bacterial physiology, and plant growth promotion. The strain 22A genome contains an eight-gene cluster encoding the staphyloferrin B-like (sbn) siderophore. We demonstrate that the sbn siderophore gene cluster is necessary for growth under low iron conditions and was complemented by supplementation with citrate or spent medium of the wild type or other strains of the genera. The siderophore exhibited adaptive features, including tolerance to oxidative and nitrosative stress, biofilm formation, and heavy metal sequestration. The contribution of the siderophore to plant growth was shown by the repressive growth of duckweed treated with siderophore mutant under iron-limited conditions; however, the siderophore was dispensable for strain 22A to colonize the phyllosphere. Importantly, the siderophore mutant could not grow on methanol, but the siderophore could solubilize insoluble Ln oxide, suggesting its critical role in methylotrophy. We also identified TonB-dependent receptors (TBDRs) for the siderophore-iron complex, iron citrate, and Ln, among 12 TBDRs in strain 22A. Analysis of the siderophore synthesis gene clusters and TBDR genes in Methylobacterium genomes revealed the existence of diverse types of siderophores and TBDRs. Methylorubrum species have an exclusive TBDR for Ln uptake that has been identified as LutH. Collectively, the results of this study provide insight into the importance of the sbn siderophore in Ln chelation, bacterial physiology, and the diversity of siderophore and TBDRs in Methylobacterium species. No potential conflict of interest relevant to this article was reported.

Frontiers Media SA Acta Medica Okayama 1664-302X 13 2022 A Periplasmic Lanthanide Mediator, Lanmodulin, in Methylobacterium aquaticum Strain 22A 921636 EN Yoshiko Fujitani Institute of Plant Science and Resources, Okayama University Takeshi Shibata K.K. AB SCIEX Akio Tani Institute of Plant Science and Resources, Okayama University Methylobacterium and Methylorubrum species oxidize methanol via pyrroloquinoline quinone-methanol dehydrogenases (MDHs). MDHs can be classified into two major groups, Ca2+-dependent MDH (MxaF) and lanthanide (Ln(3+))-dependent MDH (XoxF), whose expression is regulated by the availability of Ln(3+). A set of a siderophore, TonB-dependent receptor, and an ABC transporter that resembles the machinery for iron uptake is involved in the solubilization and transport of Ln(3+). The transport of Ln(3+) into the cytosol enhances XoxF expression. A unique protein named lanmodulin from Methylorubrum extorquens strain AM1 was identified as a specific Ln(3+)-binding protein, and its biological function was implicated to be an Ln(3+) shuttle in the periplasm. In contrast, it remains unclear how Ln(3+) levels in the cells are maintained, because Ln(3+) is potentially deleterious to cellular systems due to its strong affinity to phosphate ions. In this study, we investigated the function of a lanmodulin homolog in Methylobacterium aquaticum strain 22A. The expression of a gene encoding lanmodulin (lanM) was induced in response to the presence of La3+. A recombinant LanM underwent conformational change upon La3+ binding. Phenotypic analyses on lanM deletion mutant and overexpressing strains showed that LanM is not necessary for the wild-type and XoxF-dependent mutant's methylotrophic growth. We found that lanM expression was regulated by MxcQE (a two-component regulator for MxaF) and TonB_Ln (a TonB-dependent receptor for Ln(3+)). The expression level of mxcQE was altered to be negatively dependent on Ln(3+) concentration in increment lanM, whereas it was constant in the wild type. Furthermore, when exposed to La3+, increment lanM showed an aggregating phenotype, cell membrane impairment, La deposition in the periplasm evidenced by electron microscopy, differential expression of proteins involved in membrane integrity and phosphate starvation, and possibly lower La content in the membrane vesicle (MV) fractions. Taken together, we concluded that lanmodulin is involved in the complex regulation mechanism of MDHs and homeostasis of cellular Ln levels by facilitating transport and MV-mediated excretion. No potential conflict of interest relevant to this article was reported.

Frontiers Media Acta Medica Okayama 1664-302X 12 2021 Comprehensive Comparative Genomics and Phenotyping of Methylobacterium Species 740610 EN Ola Alessa Institute of Plant Science and Resources, Okayama University Yoshitoshi Ogura Division of Microbiology, Department of Infectious Medicine, Kurume University School of Medicine Yoshiko Fujitani Institute of Plant Science and Resources, Okayama University Hideto Takami Atmosphere and Ocean Research Institute, The University of Tokyo Tetsuya Hayashi Department of Bacteriology, Graduate School of Medical Sciences, Kyushu University Nurettin Sahin Egitim Fakultesi, Mugla Sitki Kocman University Akio Tani Institute of Plant Science and Resources, Okayama University The pink-pigmented facultative methylotrophs (PPFMs), a major bacterial group found in the plant phyllosphere, comprise two genera: Methylobacterium and Methylorubrum. They have been separated into three major clades: A, B (Methylorubrum), and C. Within these genera, however, some species lack either pigmentation or methylotrophy, which raises the question of what actually defines the PPFMs. The present study employed a comprehensive comparative genomics approach to reveal the phylogenetic relationship among the PPFMs and to explain the genotypic differences that confer their different phenotypes. We newly sequenced the genomes of 29 relevant-type strains to complete a dataset for almost all validly published species in the genera. Through comparative analysis, we revealed that methylotrophy, nitrate utilization, and anoxygenic photosynthesis are hallmarks differentiating the PPFMs from the other Methylobacteriaceae. The Methylobacterium species in clade A, including the type species Methylobacterium organophilum, were phylogenetically classified into six subclades, each possessing relatively high genomic homology and shared phenotypic characteristics. One of these subclades is phylogenetically close to Methylorubrum species; this finding led us to reunite the two genera into a single genus Methylobacterium. Clade C, meanwhile, is composed of phylogenetically distinct species that share relatively higher percent G+C content and larger genome sizes, including larger numbers of secondary metabolite clusters. Most species of clade C and some of clade A have the glutathione-dependent pathway for formaldehyde oxidation in addition to the H4MPT pathway. Some species cannot utilize methanol due to their lack of MxaF-type methanol dehydrogenase (MDH), but most harbor an XoxF-type MDH that enables growth on methanol in the presence of lanthanum. The genomes of PPFMs encode between two and seven (average 3.7) genes for pyrroloquinoline quinone-dependent alcohol dehydrogenases, and their phylogeny is distinctly correlated with their genomic phylogeny. All PPFMs were capable of synthesizing auxin and did not induce any immune response in rice cells. Other phenotypes including sugar utilization, antibiotic resistance, and antifungal activity correlated with their phylogenetic relationship. This study provides the first inclusive genotypic insight into the phylogeny and phenotypes of PPFMs. No potential conflict of interest relevant to this article was reported.

Nature Research Acta Medica Okayama 2058-5276 1 1 2016 A capsidless ssRNA virus hosted by an unrelated dsRNA virus 15001 EN Rui Zhang Agrivirology Laboratory, Institute of Plant Science and Resources, Okayama University Sakae Hisano Agrivirology Laboratory, Institute of Plant Science and Resources, Okayama University Akio Tani Agrivirology Laboratory, Institute of Plant Science and Resources, Okayama University Hideki Kondo Agrivirology Laboratory, Institute of Plant Science and Resources, Okayama University Satoko Kanematsu NARO Institute of Fruit Tree Science Nobuhiro Suzuki Agrivirology Laboratory, Institute of Plant Science and Resources, Okayama University Viruses typically encode the capsid that encases their genome, while satellite viruses do not encode a replicase and depend on a helper virus for their replication1. Here, we report interplay between two RNA viruses, yado-nushi virus 1 (YnV1) and yado-kari virus 1 (YkV1), in a phytopathogenic fungus, Rosellinia necatrix2. YkV1 has a close phylogenetic affinity to positive-sense, single-stranded (+)ssRNA viruses such as animal caliciviruses3, while YnV1 has an undivided double-stranded (ds) RNA genome with a resemblance to fungal totiviruses4. Virion transfection and infectious full-length cDNA transformation has shown that YkV1 depends on YnV1 for viability, although it probably encodes functional RNA-dependent RNA polymerase (RdRp). Immunological and molecular analyses have revealed trans-encapsidation of not only YkV1 RNA but also RdRp by the capsid protein of the other virus (YnV1), and enhancement of YnV1 accumulation by YkV1. This study demonstrates interplay in which the capsidless (+)ssRNA virus (YkV1), hijacks the capsid protein of the dsRNA virus (YnV1), and replicates as if it were a dsRNA virus. No potential conflict of interest relevant to this article was reported.

MDPI Acta Medica Okayama 2076-2607 8 6 2020 Lanthanide-Dependent Methanol and Formaldehyde Oxidation inMethylobacterium aquaticumStrain 22A 822 EN Patcha Yanpirat Institute of Plant Science and Resources, Okayama University Yukari Nakatsuji Institute of Plant Science and Resources, Okayama University Shota Hiraga Institute of Plant Science and Resources, Okayama University Yoshiko Fujitani Institute of Plant Science and Resources, Okayama University Terumi Izumi Institute of Plant Science and Resources, Okayama University Sachiko Masuda Institute of Plant Science and Resources, Okayama University Ryoji Mitsui Department of Biochemistry, Faculty of Science, Okayama University of Science Tomoyuki Nakagawa The United Graduate School of Agricultural Science, Gifu University Akio Tani Institute of Plant Science and Resources, Okayama University Lanthanides (Ln) are an essential cofactor for XoxF-type methanol dehydrogenases (MDHs) in Gram-negative methylotrophs. The Ln(3+)dependency of XoxF has expanded knowledge and raised new questions in methylotrophy, including the differences in characteristics of XoxF-type MDHs, their regulation, and the methylotrophic metabolism including formaldehyde oxidation. In this study, we genetically identified one set of Ln(3+)- and Ca2+-dependent MDHs (XoxF1 and MxaFI), that are involved in methylotrophy, and an ExaF-type Ln(3+)-dependent ethanol dehydrogenase, among six MDH-like genes inMethylobacterium aquaticumstrain 22A. We also identified the causative mutations in MxbD, a sensor kinase necessary formxaFexpression andxoxF1repression, for suppressive phenotypes inxoxF1mutants defective in methanol growth even in the absence of Ln(3+). Furthermore, we examined the phenotypes of a series of formaldehyde oxidation-pathway mutants (fae1,fae2,mchin the tetrahydromethanopterin (H4MPT) pathway andhgdin the glutathione-dependent formaldehyde dehydrogenase (GSH) pathway). We found that MxaF produces formaldehyde to a toxic level in the absence of the formaldehyde oxidation pathways and that either XoxF1 or ExaF can oxidize formaldehyde to alleviate formaldehyde toxicity in vivo. Furthermore, the GSH pathway has a supportive role for the net formaldehyde oxidation in addition to the H4MPT pathway that has primary importance. Studies on methylotrophy inMethylobacteriumspecies have a long history, and this study provides further insights into genetic and physiological diversity and the differences in methylotrophy within the plant-colonizing methylotrophs. No potential conflict of interest relevant to this article was reported.

Oxford University Press Acta Medica Okayama 0022-0957 70 5 2019 Honeydew-associated microbes elicit defense responses against brown planthopper in rice 1683 1696 EN David Wari Institute of Plant Science and Resources, Okayama University Md Alamgir Kabir Institute of Plant Science and Resources, Okayama University Kadis Mujiono Institute of Plant Science and Resources, Okayama University Yuko Hojo Institute of Plant Science and Resources, Okayama University Tomonori Shinya Institute of Plant Science and Resources, Okayama University Akio Tani Institute of Plant Science and Resources, Okayama University Hiroko Nakatani Institute of Plant Science and Resources, Okayama University Ivan Galis Institute of Plant Science and Resources, Okayama University Feeding of sucking insects, such as the rice brown planthopper (Nilaparvata lugens; BPH), causes only limited mechanical damage on plants that is otherwise essential for injury-triggered defense responses against herbivores. In pursuit of complementary BPH elicitors perceived by plants, we examined the potential effects of BPH honeydew secretions on the BPH monocot host, rice (Oryza sativa). We found that BPH honeydew strongly elicits direct and putative indirect defenses in rice, namely accumulation of phytoalexins in the leaves, and release of volatile organic compounds from the leaves that serve to attract natural enemies of herbivores, respectively. We then examined the elicitor active components in the honeydew and found that bacteria in the secretions are responsible for the activation of plant defense. Corroborating the importance of honeydew-associated microbiota for induced plant resistance, BPHs partially devoid of their microbiota via prolonged antibiotics ingestion induced significantly less defense in rice relative to antibiotic-free insects applied to similar groups of plants. Our data suggest that rice plants may additionally perceive herbivores via their honeydew-associated microbes, allowing them to discriminate between incompatible herbivores―that do not produce honeydew―and those that are compatible and therefore dangerous. No potential conflict of interest relevant to this article was reported.

PUBLIC LIBRARY SCIENCE Acta Medica Okayama 1932-6203 7 3 2012 Practical Application of Methanol-Mediated Mutualistic Symbiosis between Methylobacterium Species and a Roof Greening Moss, Racomitrium japonicum e33800 EN Akio Tani Yuichiro Takai Ikko Suzukawa Motomu Akita Haruhiko Murase Kazuhide Kimbara Bryophytes, or mosses, are considered the most maintenance-free materials for roof greening. Racomitrium species are most often used due to their high tolerance to desiccation. Because they grow slowly, a technology for forcing their growth is desired. We succeeded in the efficient production of R. japonicum in liquid culture. The structure of the microbial community is crucial to stabilize the culture. A culture-independent technique revealed that the cultures contain methylotrophic bacteria. Using yeast cells that fluoresce in the presence of methanol, methanol emission from the moss was confirmed, suggesting that it is an important carbon and energy source for the bacteria. We isolated Methylobacterium species from the liquid culture and studied their characteristics. The isolates were able to strongly promote the growth of some mosses including R. japonicum and seed plants, but the plant-microbe combination was important, since growth promotion was not uniform across species. One of the isolates, strain 22A, was cultivated with R. japonicum in liquid culture and in a field experiment, resulting in strong growth promotion. Mutualistic symbiosis can thus be utilized for industrial moss production. No potential conflict of interest relevant to this article was reported.

PUBLIC LIBRARY SCIENCE Acta Medica Okayama 1932-6203 7 7 2012 High-Throughput Identification and Screening of Novel Methylobacterium Species Using Whole-Cell MALDI-TOF/MS Analysis e40784 EN Akio Tani Sahin Nurettin Yumiko Matsuyama Takashi Enomoto Naoki Nishimura Akira Yokota Kazuhide Kimbara Methylobacterium species are ubiquitous α-proteobacteria that reside in the phyllosphere and are fed by methanol that is emitted from plants. In this study, we applied whole-cell matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis (WC-MS) to evaluate the diversity of Methylobacterium species collected from a variety of plants. The WC-MS spectrum was reproducible through two weeks of cultivation on different media. WC-MS spectrum peaks of M. extorquens strain AM1 cells were attributed to ribosomal proteins, but those were not were also found. We developed a simple method for rapid identification based on spectra similarity. Using all available type strains of Methylobacterium species, the method provided a certain threshold similarity value for species-level discrimination, although the genus contains some type strains that could not be easily discriminated solely by 16S rRNA gene sequence similarity. Next, we evaluated the WC-MS data of approximately 200 methylotrophs isolated from various plants with MALDI Biotyper software (Bruker Daltonics). Isolates representing each cluster were further identified by 16S rRNA gene sequencing. In most cases, the identification by WC-MS matched that by sequencing, and isolates with unique spectra represented possible novel species. The strains belonging to M. extorquens, M. adhaesivum, M. marchantiae, M. komagatae, M. brachiatum, M. radiotolerans, and novel lineages close to M. adhaesivum, many of which were isolated from bryophytes, were found to be the most frequent phyllospheric colonizers. The WC-MS technique provides emerging high-throughputness in the identification of known/novel species of bacteria, enabling the selection of novel species in a library and identification without 16S rRNA gene sequencing. No potential conflict of interest relevant to this article was reported.

PUBLIC LIBRARY SCIENCE Acta Medica Okayama 1932-6203 7 11 2013 A Catalytic Role of XoxF1 as La3+-Dependent Methanol Dehydrogenase in Methylobacterium extorquens Strain AM1 e50480 EN Tomoyuki Nakagawa Ryoji Mitsui Akio Tani Kentaro Sasa Shinya Tashiro Tomonori Iwama Takashi Hayakawa Keiichi Kawai In the methylotrophic bacterium Methylobacterium extorquens strain AM1, MxaF, a Ca2+-dependent methanol dehydrogenase (MDH), is the main enzyme catalyzing methanol oxidation during growth on methanol. The genome of strain AM1 contains another MDH gene homologue, xoxF1, whose function in methanol metabolism has remained unclear. In this work, we show that XoxF1 also functions as an MDH and is La3+-dependent. Despite the absence of Ca2+ in the medium strain AM1 was able to grow on methanol in the presence of La3+. Addition of La3+ increased MDH activity but the addition had no effect on mxaF or xoxF1 expression level. We purified MDH from strain AM1 grown on methanol in the presence of La3+, and its N-terminal amino acid sequence corresponded to that of XoxF1. The enzyme contained La3+ as a cofactor. The ΔmxaF mutant strain could not grow on methanol in the presence of Ca2+, but was able to grow after supplementation with La3+. Taken together, these results show that XoxF1 participates in methanol metabolism as a La3+-dependent MDH in strain AM1. No potential conflict of interest relevant to this article was reported.

Springer International Publishing Acta Medica Okayama 2193-1801 2 2013 Evaluation of rhizosphere, rhizoplane and phyllosphere bacteria and fungi isolated from rice in Kenya for plant growth promoters 606 EN Mwashasha Rashid Mwajita Hunja Murage Akio Tani Esther M Kahangi Rice (Oryza sativa L.) is the most important staple food crop in many developing countries, and is ranked third in Kenya after maize and wheat. Continuous cropping without replenishing soil nutrients is a major problem in Kenya resulting to declining soil fertility. The use of chemical fertilizers to avert the problem of low soil fertility is currently limited due to rising costs and environmental concerns. Many soil micro-organisms are able to solubilize the unavailable phosphorus, increase uptake of nitrogen and also synthesize growth promoting hormones including auxin. The aim of this study was to isolate and characterize phyllosphere, rhizoplane and rhizosphere micro-organisms from Kenyan rice with growth promoting habits. In this study whole plant rice samples were collected from different rice growing regions of Kenya. 76.2%, over 80% and 38.5% of the bacterial isolates were positive for phosphate solubilization, nitrogenase activity and IAA production whereas 17.5% and 5% of the fungal isolates were positive for phosphate solubilization and IAA production respectively. Hence these micro-organisms have potential for utilization as bio-fertilizers in rice production. No potential conflict of interest relevant to this article was reported.

岡山大学環境管理センター Acta Medica Okayama 0917-1533 31 2009 岡山大学資源生物科学研究所における屋上緑化による建物冷却効果 21 25 EN Maki Katsuhara Shigemi Tanakamaru Izumi C. Mori Akio Tani Shigeko Utsugi Takashi Enomoto Toshihiko Maitani Roof greening is known to be environmentally friendly technology. Recently developed new roof greening systems, such as the thin-layer/Excel soil© system and the wetland type greening system, were tested at the roof top of buildings of Research Institute for Bioresources, Okayama University. After a multi-year test, these new systems have been established during high-temperature and less-rainfall summer seasons in the south Okayama region. Data indicated that roof greening effectively reduced the temperature of the concrete surface (more than 10°C). The room temperature under the green roof was also reduced both in a stock room (up to 6°C) and in an office room (about 2°C). We also provided the estimation indicating that this roof greening is useful for the decrease in CO(2) emission through the reduction of the electric power for air-conditioning in the summer. No potential conflict of interest relevant to this article was reported.