Mesoterricola silvestris gen. nov., sp. nov., Mesoterricola sediminis sp. nov., Geothrix oryzae sp. nov., Geothrix edaphica sp. nov., Geothrix rubra sp. nov., and Geothrix limicola sp. nov., six novel members of Acidobacteriota isolated from soils.

Forty-eight Acidobacteriota strains were isolated from soils and sediments in Japan. Among them, six representative strains, designated W79T, W786T, Red222T, Red802T, Red803T, and Red804T, were subjected to the taxonomic classification. These six strains are Gram-stain-negative, non-spore-forming, rod-shaped, and facultative anaerobic bacterium that can reduce ferric iron. Phylogenetic and phylogenomic trees based on 16S rRNA genes and multiple single-copy gene sequences showed that strains Red222T, Red802T, Red803T, and Red804T formed a cluster with the type strains of Geothrix species, but strains W79T and W786T created an independent cluster from any other type strains. The former four strains shared 97.95-99.08% similarities of 16S rRNA gene sequence with the type strains of the genus Geothrix, whereas the latter two strains 94.86-95.49% similarities. The average amino acid identity of strains W79T and W786T were <63 % to any other type strains, which were below the genus delineation thresholds. Moreover, colonies of these two strains were white, while those of the other four isolated strains were reddish-yellow as well as the type strain Geothrix fermentans H-5T. Although the known type strains of Geothrix species have been reported to be non-motile, five strains (W79T, W786T, Red222T, Red803T, and Red804T) except for strain Red802T displayed motility. Furthermore, multiple genomic, phylogenetic, and phenotypic features supported the discrimination between these isolated strains. Based on the study evidence, we propose these six isolates as novel members within the Acidobacteriota/Holophagae/Holophagales/Holophagaceae, comprising two novel species of a novel genus, Mesoterricola silvestris gen. nov., sp. nov., and Mesoterricola sediminis sp. nov., and four novel species of the genus Geothrix: Geothrix oryzae sp. nov., Geothrix edaphica sp. nov., Geothrix rubra sp. nov., and Geothrix limicola sp. nov.

Biological nitrogen fixation in the long-term nitrogen-fertilized and unfertilized paddy fields, with special reference to diazotrophic iron-reducing bacteria.

Biological nitrogen fixation (BNF) is important to sustain nitrogen fertility of paddy soil and rice yield, while could be affected by nitrogen fertilization. Iron-reducing bacteria, Anaeromyxobacter and Geobacter, are newly found diazotrophic bacteria predominant in paddy soil. Experimental field of this study is a long-term (35 years) nitrogen fertilized (6.0 g N/m2/year) and unfertilized paddy field, where ca. 70% of rice yield was obtained yearly in nitrogen unfertilized plot (443 ± 37 g/m2) compared to fertilized plot (642 ± 64 g/m2). Effects of long-term nitrogen fertilization/unfertilization on soil properties related to BNF were investigated with special reference to diazotrophic iron-reducing bacteria. Soil chemical/biochemical properties, soil nitrogen-fixing activity, and community composition of diazotrophic bacteria were similar between nitrogen fertilized and unfertilized plot soils. In both plot soils, Anaeromyxobacter and Geobacter were the most predominant diazotrophs. Their nifD transcripts were detected at similar level, while those of other general diazotrophs were under detection limit. It was concluded that long-term use/unuse of nitrogen fertilizer in this field did not affect the predominance and nitrogen-fixing activity of diazotrophic iron-reducing bacteria, composition of other general diazotrophs, and the resulting soil nitrogen-fixing activity. BNF, primarily driven by diazotrophic iron-reducing bacteria, might significantly contribute to sustain soil nitrogen fertility and rice yield in both plot soils. Appropriate soil management to maintain BNF, including diazotrophic iron-reducing bacteria, will be important for sustainable soil nitrogen fertility and rice production.

Anaeromyxobacter oryzae sp. nov., Anaeromyxobacter diazotrophicus sp. nov. and Anaeromyxobacter paludicola sp. nov., isolated from paddy soils.

Three bacterial strains (Red232T, Red267T and Red630T) were isolated from paddy soils sampled in Japan. Cells of these strains were Gram-stain-negative, facultative anaerobic, long rod-shaped with monotrichous flagella or pilus-like structures for motility, and formed red colonies on agar plates. Phylogenetic trees based on 16S rRNA gene and multiple single-copy gene sequences showed that the three strains formed a cluster with the type strains of Anaeromyxobacter species, independent from any other strain genera. Similarity values of the 16S rRNA gene sequences and genomes among the three isolated strains and the type strain of Anaeromyxobacter, Anaeromyxobacter dehalogenans 2CP-1T, were 95.4-97.4% for 16S rRNA gene sequence, 75.3-79.5% for average nucleotide identity, 19.6-21.7% for digital DNA-DNA hybridization and 64.1-72.6% for average amino acid identity, all of which are below the species delineation thresholds. Nitrogenase genes were observed in the genomes of the three novel strains, but not in A. dehalogenans 2CP-1T. Moreover, multiple genomic, physiological and chemotaxonomic features supported the discrimination between these three strains. Based on the evidence in this study, the three isolates represent three novel independent species for which the following names are proposed: Anaeromyxobacter oryzae sp. nov., Anaeromyxobacter diazotrophicus sp. nov. and Anaeromyxobacter paludicola sp. nov. The type strains are Red232T (=NBRC 114074T=MCCC 1K03954T), Red267T (=NBRC 114075T=MCCC 1K04211T), and Red630T (=NBRC 114076T=MCCC 1K03957T), respectively.

Investigation of Rice Yields and Critical N Losses from Paddy Soil under Different N Fertilization Rates with Iron Application.

The application of iron powder stimulated the growth of iron-reducing bacteria as a respiratory substrate and enhanced their nitrogen (N)-fixing activity in flooded paddy soils. High N fertilization (urea) in the flooded paddy soils has caused adverse environmental impacts such as ammonia (NH3) volatilization, nitrous oxide (N2O) emissions, and nitrate (NO3-) leaching. This study aims to investigate the effects of N fertilization rates in combination with an iron amendment on rice yields and N losses from flooded paddy fields. We performed a 2-year field plot experiment with traditional rice-wheat rotation in China's Yangtze River Delta. The investigation consisted of seven treatments, including 100%, 80%, 60%, and 0% of the conventional N (urea and commercial organic manure) fertilization rate, and 80%, 60%, and 0% of the conventional N with the iron powder (≥99% purity) amendment. The rice yields decreased with a reduction in the conventional N fertilization rate, whereas they were comparable after the iron application under the 80% and 60% conventional N rate. The critical N losses, including NH3 volatilization, N2O emissions, and NO3- and NH4+ leaching, generally decreased with a reduction in the conventional N fertilization rate. These N losses were significantly greater after the iron amendment compared with the non-amended treatments under the 80% and 60% conventional N fertilization rate in the first rice-growing season. However, it was comparable between the iron-amended and the non-amended treatments in the second season. Furthermore, NO3- leaching was the most significant N loss throughout the two rice seasons, followed by NH3 volatilization. The iron amendment significantly increased soil Fe2+ content compared with the non-amended treatments irrespective of N fertilization, suggesting the reduction of amended iron by iron-reducing bacteria and their simultaneous N fixation. A combination of the iron application with 60-80% of the conventional N fertilization rate could maintain rice yields similar to the conventional N fertilization rate while reducing the critical N losses in the flooded paddy field tested in this study. Our study leads to the establishment of novel and practical rice cultivation, which is a step towards the development of green agriculture.

Undervalued Pseudo-nifH Sequences in Public Databases Distort Metagenomic Insights into Biological Nitrogen Fixers.

Nitrogen fixation, a distinct process incorporating the inactive atmospheric nitrogen into the active biological processes, has been a major topic in biological and geochemical studies. Currently, insights into diversity and distribution of nitrogen-fixing microbes are dependent upon homology-based analyses of nitrogenase genes, especially the nifH gene, which are broadly conserved in nitrogen-fixing microbes. Here, we report the pitfall of using nifH as a marker of microbial nitrogen fixation. We exhaustively analyzed genomes in RefSeq (231,908 genomes) and KEGG (6,509 genomes) and cooccurrence and gene order patterns of nitrogenase genes (including nifH) therein. Up to 20% of nifH-harboring genomes lacked nifD and nifK, which encode essential subunits of nitrogenase, within 10 coding sequences upstream or downstream of nifH or on the same genome. According to a phenotypic database of prokaryotes, no species and strains harboring only nifH possess nitrogen-fixing activities, which shows that these nifH genes are "pseudo"-nifH genes. Pseudo-nifH sequences mainly belong to anaerobic microbes, including members of the class Clostridia and methanogens. We also detected many pseudo-nifH reads from metagenomic sequences of anaerobic environments such as animal guts, wastewater, paddy soils, and sediments. In some samples, pseudo-nifH overwhelmed the number of "true" nifH reads by 50% or 10 times. Because of the high sequence similarity between pseudo- and true-nifH, pronounced amounts of nifH-like reads were not confidently classified. Overall, our results encourage reconsideration of the conventional use of nifH for detecting nitrogen-fixing microbes, while suggesting that nifD or nifK would be a more reliable marker. IMPORTANCE Nitrogen-fixing microbes affect biogeochemical cycling, agricultural productivity, and microbial ecosystems, and their distributions have been investigated intensively using genomic and metagenomic sequencing. Currently, insights into nitrogen fixers in the environment have been acquired by homology searches against nitrogenase genes, particularly the nifH gene, in public databases. Here, we report that public databases include a significant amount of incorrectly annotated nifH sequences (pseudo-nifH). We exhaustively investigated the genomic structures of nifH-harboring genomes and found hundreds of pseudo-nifH sequences in RefSeq and KEGG. Over half of these pseudo-nifH sequences belonged to members of the class Clostridia, which is supposed to be a prominent nitrogen-fixing clade. We also found that the abundance of nitrogen fixers in metagenomes could be overestimated by 1.5 to >10 times due to pseudo-nifH recorded in public databases. Our results encourage reconsideration of the prevalent use of nifH as a marker of nitrogen-fixing microbes.

Volatile Components of the Kuromoji Essential Oil (Lindera umbellata Thunb.) and the Utilization for Touch Care Treatment.

The volatile components of kuromoji oil (Lindera umbellata Thunb.) obtained in Shizuoka Pref. were analyzed by GC/MS. Linalool, α-pinene, limonene, camphene, cis- and trans-dihydrocarvone, 1,8-cineol, 4-terpinenol, α-terpineol, piperitone, geranyl acetate, geraniol, and trans-nerolidol were identified as major components. Using enantio-MDGC-MS, the enantiomeric ratio ((R)-(-) vs (S)-(+)) of linalool in this oil was determined to be 67.8/32.2. Touch care treatment while sniffing this oil was done on cancer patients. We found that the relaxation effect persisted longer after the treatment compared to treatment without aroma.

Genome-Based Taxonomic Rearrangement of the Order Geobacterales Including the Description of Geomonas azotofigens sp. nov. and Geomonas diazotrophica sp. nov.

Geobacterales is a recently proposed order comprising members who originally belonged to the well-known family Geobacteraceae, which is a key group in terrestrial ecosystems involved in biogeochemical cycles and has been widely investigated in bioelectrochemistry and bioenergy fields. Previous studies have illustrated the taxonomic structure of most members in this group based on genomic phylogeny; however, several members are still in a pendent or chaotic taxonomic status owing to the lack of genome sequences. To address this issue, we performed this taxonomic reassignment using currently available genome sequences, along with the description of two novel paddy soil-isolated strains, designated Red51T and Red69T, which are phylogenetically located within this order. Phylogenomic analysis based on 120 ubiquitous single-copy proteins robustly separated the species Geobacter luticola from other known genera and placed the genus Oryzomonas (fam. Geobacteraceae) into the family 'Pseudopelobacteraceae'; thus, a novel genus Geomobilimonas is proposed, and the family 'Pseudopelobacteraceae' was emended. Moreover, genomic comparisons with similarity indexes, including average amino acid identity (AAI), percentage of conserved protein (POCP), and average nucleotide identity (ANI), showed proper thresholds as genera boundaries in this order with values of 70%, 65%, and 74% for AAI, POCP, and ANI, respectively. Based on this, the three species Geobacter argillaceus, Geobacter pelophilus, and Geobacter chapellei should be three novel genera, for which the names Geomobilibacter, Geoanaerobacter, and Pelotalea are proposed, respectively. In addition, the two novel isolated strains phylogenetically belonged to the genus Geomonas, family Geobacteraceae, and shared genomic similarity values higher than those of genera boundaries, but lower than those of species boundaries with each other and their neighbors. Taken together with phenotypic and chemotaxonomic characteristics similar to other Geomonas species, these two strains, Red51T and Red69T, represent two novel species in the genus Geomonas, for which the names Geomonas azotofigens sp. nov. and Geomonas diazotrophica sp. nov. are proposed, respectively.

A case of vastus lateralis muscle metastasis of tongue carcinoma.

Distant metastasis is the most important prognostic factor for head and neck cancer. This report presents the case of a 50-year-old man with distant metastasis of tongue carcinoma to the vastus lateralis muscle which presented to Nihon University Itabashi Hospital, Tokyo, Japan. Tumourectomy was performed with a diagnosis of tongue carcinoma (cT2N0M0, Stage II). Seven months later, radical neck dissection was performed for lymph node metastasis to a left supraclavicular lymph node. In addition, metastasis was then detected outside the neck dissection region. Tumourectomy and radiotherapy (50 Gy) were, therefore, added to the treatment regimen. However, left-sided vastus lateralis muscle metastasis was then observed. To the best of our knowledge, this is the first report of distant metastasis of oral squamous cell carcinoma to the vastus lateralis muscle.

Geomesophilobacter sediminis gen. nov., sp. nov., Geomonas propionica sp. nov. and Geomonas anaerohicana sp. nov., three novel members in the family Geobacterecace isolated from river sediment and paddy soil.

Bacteria in the family Geobacteraceae have been proven to fill important niches in a diversity of anaerobic environments and global biogeochemical processes. Here, three bacterial strains in this family, designated Red875T, Red259T, and Red421T were isolated from river sediment and paddy soils in Japan. All of them are Gram-staining-negative, strictly anaerobic, motile, flagellum-harboring cells that form red colonies on agar plates and are capable of utilizing Fe(III)-NTA, Fe(III) citrate, ferrihydrite, MnO2, fumarate, and nitrate as electron acceptors with acetate, propionate, pyruvate, and glucose as electron donors. Phylogenetic analysis based on the 16S rRNA gene and 92 concatenated core proteins sequences revealed that strains Red259T and Red421T clustered with the type strains of Geomonas species, whereas strain Red875T formed an independent lineage within the family Geobacteraceae. Genome comparison based on  average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values clearly distinguished these three strains from other Geobacteraceae members, with lower values than the thresholds for species delineation. Moreover, strain Red875T also shared low average amino acid identity (AAI) and percentage of conserved proteins (POCP) values with the type species of the family Geobacteraceae. Based on these physiological, chemotaxonomic, and phylogenetic distinctions, we propose that strain Red875T (=NBRC 114290T = MCCC 1K04407T) represents a novel genus in the family Geobacteraceae, namely, Geomesophilobacter sediminis gen. nov., sp. nov., and strains Red259T (=NBRC 114288T = MCCC 1K05016T) and Red421T (=NBRC 114289T = MCCC 1K06216T) represent two novel independent species in the genus Geomonas, namely, Geomonas propionica sp. nov. and Geomonas anaerohicana sp. nov., respectively.

Mitigating N2O emissions from agricultural soils with fungivorous mites.

Nitrous oxide (N2O) is an important greenhouse gas and an ozone-depleting substance. Due to the long persistence of N2O in the atmosphere, the mitigation of anthropogenic N2O emissions, which are mainly derived from microbial N2O-producing processes, including nitrification and denitrification by bacteria, archaea, and fungi, in agricultural soils, is urgently necessary. Members of mesofauna affect microbial processes by consuming microbial biomass in soil. However, how microbial consumption affects N2O emissions is largely unknown. Here, we report the significant role of fungivorous mites, the major mesofaunal group in agricultural soils, in regulating N2O production by fungi, and the results can be applied to the mitigation of N2O emissions. We found that the application of coconut husks, which is the low-value part of coconut and is commonly employed as a soil conditioner in agriculture, to soil can supply a favorable habitat for fungivorous mites due to its porous structure and thereby increase the mite abundance in agricultural fields. Because mites rapidly consume fungal N2O producers in soil, the increase in mite abundance substantially decreases the N2O emissions from soil. Our findings might provide new insight into the mechanisms of soil N2O emissions and broaden the options for the mitigation of N2O emissions.

3D-CT stress test for the assessment of CFL insufficiency.

BACKGROUND: Following ankle sprains, some patients complain of their ankles "giving way," characterized by functional instability with no positive findings in traditional stress tests. The calcaneofibular ligament (CFL) may contribute to the stabilization of the subtalar and talocrural joints, and some functional instability may be due to CFL insufficiency. We aimed to clarify and quantitatively assess CFL insufficiency with three-dimensional stress computer tomography (CT) using the Pronation-External Rotation Stress Test (PERST). METHODS: Ten patients who tested positive under PERST and underwent an isolated CFL reconstruction were included. Using a custom-made loading jig, we used the Supination-Internal Rotation Stress Test (SIRST) and PERST to assess the function of anterior talofibular ligament (ATFL) and CFL, respectively. 3D-CT in neutral position was used as a baseline, and we quantified the distance between the origin and insertion of the CFL and ATFL at 2 years pre- and postoperatively. RESULTS: Postoperative scores improved in all patients with no giving way symptoms. The preoperative length of the CFL increased by 14.0% from baseline under PERST, while the postoperative length only increased by 2.0% and was significantly restricted (P < .01). The pre- and postoperative length of ATFL was increased by 7.5% and 9.0% from baseline under SIRST, respectively, with no significant difference (P = .41). The clinical function improved with significantly less change in distance between the origin and insertion under PERST and showed no difference under SIRST. CONCLUSION: The 3D-CT stress test may be useful for quantifying pre- and postoperative CFL function. CFL insufficiency is one of the main causes of subtalar joint instability; therefore, measuring the distance between the origin and insertion of the CFL could provide the means to quantify the instability of the subtalar joint.

Geomonas silvestris sp. nov., Geomonas paludis sp. nov. and Geomonas limicola sp. nov., isolated from terrestrial environments, and emended description of the genus Geomonas.

Three bacterial strains, designated Red330T, Red736T and Red745T, were isolated from forest and paddy soils in Japan. Strains Red330T, Red736T and Red745T are flagella-harbouring and strictly anaerobic bacteria forming red colonies. A 16S rRNA gene sequence-based phylogenetic tree showed that all three strains were located in a cluster, including the type strains of Geomonas species, which were recently separated from the genus Geobacter within the family Geobacteraceae. Similarities of the 16S rRNA gene sequences among the three strains and Geomonas oryzae S43T, the type species of the genus Geomonas, were 96.3-98.5 %. The genome-related indexes, average nucleotide identity, digital DNA-DNA hybridization, and average amino acid identity, among the three strains and G. oryzae S43T were 74.7-86.8 %, 21.2-33.3 % and 70.4-89.8 %, respectively, which were lower than the species delineation thresholds. Regarding the phylogenetic relationships based on genome sequences, the three strains clustered with the type strains of Geomonas species, which were independent from the type strains of Geobacter species. The distinguishableness of the three isolated strains was supported by physiological and chemotaxonomic properties, with the profile of availability of electron donors and cellular fatty acids composition being particularly different among them. Based on genetic, phylogenetic and phenotypic properties, the three isolates represent three novel independent species in the genus Geomonas, for which the names Geomonas silvestris sp. nov., Geomonas paludis sp. nov. and Geomonas limicola sp. nov. are proposed. The type strains are Red330T (=NBRC 114028T=MCCC 1K03949T), Red736T (=NBRC 114029T=MCCC 1K03950T) and Red745T (=NBRC 114030T=MCCC 1K03951T), respectively.

Diazotrophic Anaeromyxobacter Isolates from Soils.

Biological nitrogen fixation is an essential reaction in a major pathway for supplying nitrogen to terrestrial environments. Previous culture-independent analyses based on soil DNA/RNA/protein sequencing could globally detect the nitrogenase genes/proteins of Anaeromyxobacter (in the class Deltaproteobacteria), commonly distributed in soil environments and predominant in paddy soils; this suggests the importance of Anaeromyxobacter in nitrogen fixation in soil environments. However, direct experimental evidence is lacking; there has been no research on the genetic background and ability of Anaeromyxobacter to fix nitrogen. Therefore, we verified the diazotrophy of Anaeromyxobacter based on both genomic and culture-dependent analyses using Anaeromyxobacter sp. strains PSR-1 and Red267 isolated from soils. Based on the comparison of nif gene clusters, strains PSR-1 and Red267 as well as strains Fw109-5, K, and diazotrophic Geobacter and Pelobacter in the class Deltaproteobacteria contain the minimum set of genes for nitrogenase (nifBHDKEN). These results imply that Anaeromyxobacter species have the ability to fix nitrogen. In fact, Anaeromyxobacter PSR-1 and Red267 exhibited N2-dependent growth and acetylene reduction activity (ARA) in vitro Transcriptional activity of the nif gene was also detected when both strains were cultured with N2 gas as a sole nitrogen source, indicating that Anaeromyxobacter can fix and assimilate N2 gas by nitrogenase. In addition, PSR-1- or Red267-inoculated soil showed ARA activity and the growth of the inoculated strains on the basis of RNA-based analysis, demonstrating that Anaeromyxobacter can fix nitrogen in the paddy soil environment. Our study provides novel insights into the pivotal environmental function, i.e., nitrogen fixation, of Anaeromyxobacter, which is a common soil bacterium.IMPORTANCEAnaeromyxobacter is globally distributed in soil environments, especially predominant in paddy soils. Current studies based on environmental DNA/RNA analyses frequently detect gene fragments encoding nitrogenase of Anaeromyxobacter from various soil environments. Although the importance of Anaeromyxobacter as a diazotroph in nature has been suggested by culture-independent studies, there has been no solid evidence and validation from genomic and culture-based analyses that Anaeromyxobacter fixes nitrogen. This study demonstrates that Anaeromyxobacter harboring nitrogenase genes exhibits diazotrophic ability; moreover, N2-dependent growth was demonstrated in vitro and in the soil environment. Our findings indicate that nitrogen fixation is important for Anaeromyxobacter to survive under nitrogen-deficient environments and provide a novel insight into the environmental function of Anaeromyxobacter, which is a common bacterium in soils.

Description of Three Novel Members in the Family Geobacteraceae, Oryzomonas japonicum gen. nov., sp. nov., Oryzomonas sagensis sp. nov., and Oryzomonas ruber sp. nov.

Bacteria of the family Geobacteraceae are particularly common and deeply involved in many biogeochemical processes in terrestrial and freshwater environments. As part of a study to understand biogeochemical cycling in freshwater sediments, three iron-reducing isolates, designated as Red96T, Red100T, and Red88T, were isolated from the soils of two paddy fields and pond sediment located in Japan. The cells were Gram-negative, strictly anaerobic, rod-shaped, motile, and red-pigmented on agar plates. Growth of these three strains was coupled to the reduction of Fe(III)-NTA, Fe(III) citrate, and ferrihydrite with malate, methanol, pyruvate, and various organic acids and sugars serving as alternate electron donors. Phylogenetic analysis based on the housekeeping genes (16S rRNA gene, gyrB, rpoB, nifD, fusA, and recA) and 92 concatenated core genes indicated that all the isolates constituted a coherent cluster within the family Geobacteraceae. Genomic analyses, including average nucleotide identity and DNA-DNA hybridization, clearly differentiated the strains Red96T, Red100T, and Red88T from other species in the family Geobacteraceae, with values below the thresholds for species delineation. Along with the genomic comparison, the chemotaxonomic features further helped distinguish the three isolates from each other. In addition, the lower values of average amino acid identity and percentage of conserved protein, as well as biochemical differences with their relatives, indicated that the three strains represented a novel genus in the family Geobacteraceae. Hence, we concluded that strains Red96T, Red100T, and Red88T represented three novel species of a novel genus in the family Geobacteraceae, for which the names Oryzomonas japonicum gen. nov., sp. nov., Oryzomonas sagensis sp. nov., and Oryzomonas ruber sp. nov. are proposed, with type strains Red96T (= NBRC 114286T = MCCC 1K04376T), Red100T (= NBRC 114287T = MCCC 1K04377T), and Red88T (= MCCC 1K03694T = JCM 33033T), respectively.

Trichoblastic infundibular cyst should be renamed infundibular cyst with unique papillary projections.

Trichoblastic infundibular cyst (TBIC) was previously reported as a unique keratinous cystic lesion, which was characterized by the papillary projections of follicular germinative-like cells emanating from the cyst wall. Here, we report three additional cases of this cyst and discuss the pathogenesis of this unique entity. In all cases, a unilocular cyst contained keratin, and the cyst wall was composed of squamous epithelium. A number of cords and papillary projections emanated from the basal layer of the cyst wall. They were composed of cells with large nuclei and scant cytoplasm arranged in a peripheral palisade. Immunohistochemically, anti-cytokeratin 15, anti-cytokeratin 20, and anti-epithelial cell adhesion molecule antibodies were negative. Thus, these cells resembled follicular germinative cells or sebaceous mantle morphologically, but we failed to prove the differentiation immunohistochemically. The cyst was surrounded by fibrotic stroma and inflammatory cells, suggesting previous rupture of the cyst. We speculate that the cells of the projections possibly differentiate into the mantle rather than follicular germinative cells, even though we could not provide sufficient immunohistochemical evidence. We also suggest that they may be induced by special reaction to fibrohistiocytic stroma surrounding the infundibular cyst. Therefore, TBIC should be renamed infundibular cyst with unique papillary projections.

Geomonas oryzae gen. nov., sp. nov., Geomonas edaphica sp. nov., Geomonas ferrireducens sp. nov., Geomonas terrae sp. nov., Four Ferric-Reducing Bacteria Isolated From Paddy Soil, and Reclassification of Three Species of the Genus Geobacter as Members of the Genus Geomonas gen. nov.

In paddy soil, bacteria from the family Geobacteraceae have been shown to strongly contribute to the biogeochemical cycle. However, no Geobacteraceae species with validly published names have been isolated from paddy soil. In this study, we isolated and characterized four novel ferric reducing bacteria in the family Geobacteraceae from the paddy soils of three different fields in Japan. The four strains, S43T, Red53T, S62T, and Red111T, were Gram-stain negative, strictly anaerobic, chemoheterotrophic, and motile with peritrichous flagella. Phylogenetic studies based on 16S rRNA gene sequences, five concatenated housekeeping genes (fusA, rpoB, recA, nifD, and gyrB) and 92 concatenated core genes revealed that the four strains belong to the family Geobacteraceae and are most closely related to Geobacter bemidjiensis BemT (97.4-98.2%, 16S rRNA gene sequence similarities) and Geobacter bremensis Dfr1T (97.1-98.0%). Genomic analysis with average nucleotide identity (ANI) and digital DNA-DNA hybridization (GGDC) calculations clearly distinguished the four isolated strains from other species of the family Geobacteraceae and indicated that strains S43T, Red53T, S62T, and Red111T represent independent species, with values below the thresholds for species delineation. Chemotaxonomic characteristics, including major fatty acid and whole cell protein profiles, showed differences among the isolates and their closest relatives, which were consistent with the results of DNA fingerprints and physiological characterization. Additionally, each of the four isolates shared a low 16S rRNA gene sequence similarity (92.4%) and average amino acid identity (AAI) with the type strain of the type species Geobacter metallireducens. Overall, strains S43T, Red53T, S62T, and Red111T represent four novel species, which we propose to classify in a novel genus of the family Geobacteraceae, and the names Geomonas oryzae gen. nov., sp. nov. (type strain S43T), Geomonas edaphica sp. nov. (type strain Red53T), Geomonas ferrireducens sp. nov. (type strain S62T), and Geomonas terrae sp. nov. (type strain Red111T) are proposed. Based on phylogenetic and genomic analyses, we also propose the reclassification of Geobacter bremensis as Geomonas bremensis comb. nov., Geobacter pelophilus as Geomonas pelophila comb. nov., and Geobacter bemidjiensis as Geomonas bemidjiensis comb. nov.

A Fragrant Environment Containing α-Pinene Suppresses Tumor Growth in Mice by Modulating the Hypothalamus/Sympathetic Nerve/Leptin Axis and Immune System.

The environment is thought to affect outcomes in patients with cancer; however, this relationship has not been proven directly. Recently, an enriched environment, as a model of a positive environment, has been shown to suppress tumor growth by lowering leptin production through a pathway involving the hypothalamus/sympathetic nerve/leptin axis. We previously reported that a fragrant environment (FE) containing α-pinene suppressed tumor growth in mice; however, the underlying mechanism has not been elucidated. Accordingly, in this study, we investigated changes in the neuroendocrine and immune systems following exposure to an FE. Mice were exposed to α-pinene (5 h/day) for 4 weeks prior to tumor implantation with murine melanoma cells and 3 weeks after transplantation. In addition to the evaluation of tumor growth, the blood, spleen, and hypothalamus were collected 3 weeks after transplantation, and neuroendocrinological and immunological parameters were measured. Tumor size was ~40% smaller in mice exposed to FE. Moreover, plasma noradrenaline concentrations, which reflected sympathetic nervous activity, tended to increase, and leptin levels were significantly decreased in FE-exposed mice. Levels of stress hormones, such as plasma corticosterone and adrenaline, did not change in the 2 groups. In the hypothalamus, brain-derived neurotrophic factor protein levels and glucose-1-phosphate concentrations were decreased in the FE group. Additionally, numbers of B cells, CD4+ T cells, CD8+ T cells, and natural killer cells increased in the FE-exposed mice. These neurohormonal and immunological changes in the FE-exposed mice suggested that the FE may activate the hypothalamus/sympathetic nerve/leptin axis and immune system, thereby retarding tumor growth.

Predominant but Previously-overlooked Prokaryotic Drivers of Reductive Nitrogen Transformation in Paddy Soils, Revealed by Metatranscriptomics.

Waterlogged paddy soils possess anoxic zones in which microbes actively induce reductive nitrogen transformation (RNT). In the present study, a shotgun RNA sequencing analysis (metatranscriptomics) of paddy soil samples revealed that most RNT gene transcripts in paddy soils were derived from Deltaproteobacteria, particularly the genera Anaeromyxobacter and Geobacter. Despite the frequent detection of the rRNA of these microbes in paddy soils, their RNT-associated genes have rarely been identified in previous PCR-based studies. This metatranscriptomic analysis provides novel insights into the diversity of RNT microbes present in paddy soils and the ecological function of Deltaproteobacteria predominating in these soils.

Whole exome sequencing detects variants of genes that mediate response to anticancer drugs.

Certain interindividual differences affecting the efficacy of drug treatment and adverse drug reactions are caused by genetic variants, and their phenotypic effects differ among ethnic groups. In this study, we used whole exome sequencing (WES) systematically to identify germline mutations that influence the activities of drug-metabolizing enzymes, as well as that of a transporter. We analyzed DNA isolated from blood samples from 2,042 Japanese patients with diverse cancers. We identified sequence variants of CYP2B6 (rs3745274), CYP2C9 (rs1057910), CYP2C19 (rs4986893), CYP2C19 (rs4244285), TPMT (rs1142345), NAT2 (rs1799930), NAT2 (rs1799931), UGT1A1 (rs4148323), COMT (rs4680), ABCB1 (rs1045642), and CDA (rs60369023). Wider application of WES will help to determine the effects of mutations on the activities of proteins encoded by drug response genes, and the information gained will accelerate the development of personalized therapies for patients with cancer. Moreover, this knowledge may provide clues for preventing cancer before the onset of symptoms.