Hydrogen peroxide (H2O2) inhibits microbial growth at a specific concentration. However, we previously isolated two environmental bacterial strains that exhibited sensitivity to a lower H2O2 concentration in agar plates. Putative catalase genes, which degrade H2O2, were detected in their genomes. We herein elucidated the characteristics of these putative genes and their products using a self-cloning technique. The products of the cloned genes were identified as functional catalases. The up-regulation of their expression increased the colony-forming ability of host cells under H2O2 pressure. The present results demonstrated high sensitivity to H2O2 even in microbes possessing functional catalase genes.

Streptococcus mutans is a major caries-causing bacterium that forms firmly attached biofilms on tooth surfaces. Biofilm formation by S. mutans consists of polysaccharide-dependent and polysaccharide-independent processes. Among polysaccharide-independent processes, extracellular DNA (eDNA) mediates the initial attachment of cells to surfaces. We previously reported that the secreted peptide signal, competence-stimulating peptide (CSP) induced cell death in a subpopulation of cells, leading to autolysis-mediated eDNA release. The autolysin gene lytF, the expression of which is stimulated by CSP, has been shown to mediate CSP-dependent cell death, while cell death was not entirely abolished in the lytF deletion mutant, indicating the involvement of other factors. To identify novel genes involved in CSP-dependent cell death, we herein compared transcriptomes between live and dead cells derived from an isogenic population. The results obtained revealed the accumulation of several mRNAs in dead cells. The deletion of SMU_1553c, a putative bacteriocin gene, resulted in significant reductions in CSP-induced cell death and eDNA production levels from those in the parental strain. Moreover, in the double mutant strain of lytF and SMU_1553c, cell death and eDNA production in response to synthetic CSP were completely abolished under both planktonic and biofilm conditions. These results indicate that SMU_1553c is a novel cell death-related factor that contributes to CSP-dependent cell death and eDNA production.

Pseudomonas putida is a major species belonging to the genus Pseudomonas. Although several hundred strains of P. putida have been deposited in culture collections, they potentially differ from the genetically defined "true Pseudomonas putida" because many were classified as P. putida based on their phenotypic and metabolic characteristics. A phylogenetic ana-lysis based on the concatenated sequences of the 16S rRNA and rpoD genes revealed that 46 strains of P. putida deposited in Japanese culture collections were classified into nine operational taxonomic units (OTUs) and eleven singletons. The OTU7 strain produces N-acylhomoserine lactone as a quorum-sensing signal. One of the OTU7 strains, JCM 20066, exhibited a ppuI-rsaL-ppuR quorum-sensing system that controls biofilm formation and motility. The P. putida type strain JCM 13063T and six other strains were classified as OTU4. Classification based on the calculation of whole-genome similarity revealed that three OTU4 strains, JCM 20005, 21368, and 13061, were regarded as the same species as JCM 13063T and defined as true P. putida. When orthologous genes in the whole-genome sequences of true P. putida strains were screened, PP4_28660 from P. putida NBRC 14164T (=JCM 13063T) was present in all true P. putida genome sequences. The internal region of PP4_28660 was successfully amplified from all true P. putida strains using the specific primers designed in this study.

When viruses infect microbial cells, their phenotypes depend on the host's genotype and on the environmental conditions. Here we describe such an effect in laboratory strains of the chlorovirus PBCV-1 and its algal host Chlorella variabilis. We studied the growth of six virus isolates, and found that the mean lysis time was 1.34±0.05 times longer at multiplicity of particles (MOP) 10 than at MOP 1. We could not detect any associated changes in burst size. This is a novel plastic trait for chloroviruses, and we hypothesize that it is caused by our specific laboratory algae.

Anammox bacteria produce N2 gas by oxidizing NH4+ with NO2-, and hydroxylamine (NH2OH) is a potential intermediate of the anammox process. N2 gas production occurs when anammox bacteria are incubated with NH2OH only, indicating their capacity for NH2OH disproportionation with NH2OH serving as both the electron donor and acceptor. Limited information is currently available on NH2OH disproportionation by anammox bacteria; therefore, the stoichiometry of anammox bacterial NH2OH disproportionation was examined in the present study using 15N-tracing techniques. The anammox bacteria, Brocadia sinica, Jettenia caeni, and Scalindua sp. were incubated with the addition of 15NH2OH, and the production of 15N-labeled nitrogenous compounds was assessed. The anammox bacteria tested performed NH2OH disproportionation and produced 15-15N2 gas and NH4+ as reaction products. The addition of acetylene, an inhibitor of the anammox process, reduced the activity of NH2OH disproportionation, but not completely. The growth of B. sinica by NH2OH disproportionation (-240.3 kJ mol NH2OH-1 under standard conditions) was also tested in 3 up-flow column anammox reactors fed with 1) 0.7 mM NH2OH only, 2) 0.7 mM NH2OH and 0.5 mM NH4+, and 3) 0.7 mM NH2OH and 0.5 mM NO2-. NH2OH consumption activities were markedly reduced after 7 d of operation, indicating that B. sinica was unable to maintain its activity or biomass by NH2OH disproportionation.

Phosphorus (P) is an essential macronutrient for plant growth and is mainly present in agricultural soil in unavailable forms. Phosphate-solubilizing microorganisms (PSMs) increase soil P availability. The objective of the present study was to assess the population and type of PSMs and their relationships with soil characteristics in the agricultural soil of Manokwari. Twenty-one composite soil samples (0-20 cm) were collected at the rhizospheres of plants in the Prafi and Masni Districts. A dilution technique and plate count method on Pikovskayas agar medium were used to examine the PSM population, phosphate-solubilizing index (PSI), and various soil properties. The results obtained showed that the total population of phosphate-solubilizing bacteria ranged between 25×103 and 550×103 CFU g-1 of soil, while that of phosphate-solubilizing fungi was between 2.0×103 and 5.0×103 CFU g-1 of soil at all locations. The PSI of the isolates ranged between 1.1 to 3.6 mm, with the most efficient and highest PSI being obtained for Bacillus sp. (strain 8) and the lowest for Pseudomonas sp. (strain 15). Six isolates found at all locations were identified at the genus level: Chromobacterium sp., Pseudomonas sp., Bacillus sp., Micrococcus sp., Caulobacter sp., and Aspergillus sp. A correlation was observed between the number of PSMs and the level of soil P availability and moisture content, indicating an increase in soil P availability with a greater abundance of PSMs in soil.

Integrative and conjugative elements (ICEs) play a role in the horizontal transfer of antibiotic resistance genes (ARGs). We herein report an ICE from Shewanella halifaxensis isolated from fish intestine with a similar structure to both a clinical bacterial ICE and marine bacterial plasmid. The ICE was designated ICEShaJpn1, a member of the SXT/R391 family of ICEs (SRIs). ICEShaJpn1 has a common core structure with SRIs of clinical and fish origins and an ARG cassette with the pAQU1 plasmid of Photobacterium damselae subsp. damselae, suggesting that the common core of SRIs is widely distributed and ARG cassettes are collected from regional bacteria.

The rhizosphere microbiome of the native Antarctic hairgrass Deschampsia antarctica from the central maritime Antarctic was investigated using 16S RNA metagenomics and compared to those of the second native Antarctic plant Colobanthus quitensis and closely related temperate D. cespitosa. The rhizosphere microbial communities of D. antarctica and D. cespitosa had high taxon richness, while that of C. quitensis had markedly lower diversity. The majority of bacteria in the rhizosphere communities of the hairgrass were affiliated to Proteobacteria, Bacteroidetes, and Actinobacteria. The rhizosphere of C. quitensis was dominated by Actinobacteria. All microbial communities included high proportions of unique amplicon sequence variants (ASVs) and there was high heterogeneity between samples at the ASV level. The soil parameters examined did not explain this heterogeneity. Bacteria belonging to Actinobacteria, Bacteroidetes, and Proteobacteria were sensitive to fluctuations in the soil surface temperature. The values of the United Soil Surface Temperature Influence Index (UTII, Iti) showed that variations in most microbial communities from Galindez Island were associated with microscale variations in temperature. Metabolic predictions in silico using PICRUSt 2.0, based on the taxonomically affiliated part of the microbiomes, showed similarities with the rhizosphere community of D. antarctica in terms of the predicted functional repertoire. The results obtained indicate that these communities are involved in the primary processes of soil development (particularly the degradation of lignin and lignin-derived compounds) in the central maritime Antarctic and may be beneficial for the growth of Antarctic vascular plants. However, due to the limitations associated with interpreting PICRUSt 2.0 outputs, these predictions need to be verified experimentally.

Agricultural soil is the primary N2O sink limiting the emission of N2O gas into the atmosphere. Although Gemmatimonadetes bacteria are abundant in agricultural soils, limited information is currently available on N2O reduction by Gemmatimonadetes bacteria. Therefore, the effects of pH and temperature on N2O reduction activities and affinity constants for N2O reduction were examined by performing batch experiments using an isolate of Gemmatimonadetes bacteria, Gemmatimonas aurantiaca (NBRC100505T). G. aurantiaca reduced N2O at pH 5-9 and 4-50°C, with the highest activity being observed at pH 7 and 30°C. The affinity constant of G. aurantiaca cells for N2O was 4.4 μM. The abundance and diversity of the Gemmatimonadetes 16S rRNA gene and nosZ encoding nitrous oxide reductase in agricultural soil samples were also investigated by quantitative PCR (qPCR) and amplicon sequencing ana-lyses. Four N2O-reducing agricultural soil samples were assessed, and the copy numbers of the Gemmatimonadetes 16S rRNA gene (clades G1 and G3), nosZ DNA, and nosZ mRNA were 8.62-9.65×108, 5.35-7.15×108, and 2.23-4.31×109 copies (g dry soil)-1, respectively. The abundance of the nosZ mRNA of Gemmatimonadetes bacteria and OTU91, OUT332, and OTU122 correlated with the N2O reduction rates of the soil samples tested, suggesting N2O reduction by Gemmatimonadetes bacteria. Gemmatimonadetes 16S rRNA gene reads affiliated with OTU4572 and OTU3759 were predominant among the soil samples examined, and these Gemmatimonadetes OTUs have been identified in various types of soil samples.

Recent studies suggested the presence of magma chambers from the Tatun volcano group under northern Taiwan's surface, the result of episodic volcanism for 0.2-2.8 million years. However, the microbial community in volcanic soil has not yet been characterized. Therefore, the present study investigated the spatial distribution of microbial communities and their relationships with environmental variables, including heavy metals. Next-generation sequencing was used to analyze the microbial community structures in three areas with different land uses: Lengshuikeng (recreational area), Zhuzihu (agricultural area), and Huangzuishan (conservation area). High contents of environmental factors, such as nitrogen (0.46-1.14%) and phosphorus (2.01-13.88 ppm), were detected. Large concentrations of heavy metals, such as copper (55.90-127.60 ppm) and zinc (36.13-147.73 ppm), were found among the three sites, whereas those of lead (83.13 ppm) and chromium (48.33 ppm) were higher in the Zhuzihu area. The most prevalent phylum across all sites was Proteobacteria, followed by Actinobacteria, Acidobacteria, and Chloroflexi, while the most abundant bacterial species was Koribacteraceae: NA_01, followed by Cyanobacteria: NA. A network ana-lysis showed that Koribacteracea: NA_01 positively correlated with bacterial groups, including Flavisolibacter sp., Oxalobacteraceae: NA, and Actinomycetales: NA_01. Based on Shannon and Simpson's diversity indices, the diversity of bacteria was significantly less in the Huangzuishan area than in the Lengshuikeng and Zhuzihu areas. Bacterial assemblages also significantly differed (P<0.05) among the three sites. The present results provide clear evidence to show that environmental variables, including heavy metals, are key factors affecting the bacterial community structure in volcanic soil.

Propionate oxidation in Pelotomaculum thermopropionicum is performed under a thermodynamic limit. The most energetically unfavorable reaction in the propionate oxidation pathway is succinate oxidation. Based on previous genomic and transcriptomic ana-lyses, succinate oxidation in P. thermopropionicum under propionate-oxidizing conditions is conducted by the membrane-bound forms of two succinate dehydrogenases (SDHs). We herein examined the activity of SDH, the mechanisms underlying the succinate oxidation reaction in P. thermopropionicum, and the importance of the protein sequences of related genes. SDH activity was highly localized to the membrane fraction. An ana-lysis of the soluble fraction revealed that fumarate reductase received electrons from NADH, suggesting the involvement of membrane-bound SDH in propionate oxidation. We utilized an uncoupler and inhibitors of adenosine triphosphate (ATP) synthase and membrane-bound SDH to investigate whether the membrane potential of P. thermopropionicum supports propionate oxidation alongside hydrogen production. These chemicals inhibited hydrogen production, indicating that membrane-bound SDH requires a membrane potential for succinate oxidation, and this membrane potential is maintained by ATP synthase. In addition, the phylogenetic distribution of the flavin adenine dinucleotide-binding subunit and conserved amino acid sequences of the cytochrome b subunit of SDHs in propionate-oxidizing bacteria suggests that membrane-bound SDHs possess specific conserved amino acid residues that are strongly associated with efficient succinate oxidation in syntrophic propionate-oxidizing bacteria.

We herein propose a fast and easy DNA and RNA co-extraction method for environmental microbial samples. It combines bead beating and phenol-chloroform phase separation followed by the separation and purification of DNA and RNA using the Qiagen AllPrep DNA/RNA mini kit. With a handling time of ~3 h, our method simultaneously extracted high-quality DNA (peak size >10-15‍ ‍kb) and RNA (RNA integrity number >6) from lake bacterioplankton filtered samples. The method is also applicable to low-biomass samples (expected DNA or RNA yield <50‍ ‍ng) and eukaryotic microbial samples, providing an easy option for more versatile eco-genomic applications.

The umbilicus accumulates more dirt than other body surfaces and is difficult to clean. Hygiene in this area is vital, particularly for surgery, because of its proximity to the laparotomy site. Although microorganisms in the umbilicus have been extensively examined, those in umbilical dirt have not due to the lack of an efficient method of collection. We previously established a technique to extract umbilical dirt using the anchor effect of polymers, which are injected into the umbilicus. In the present study, we applied this technique for the first time to investigate umbilical dirt. The results obtained revealed an abundance of Corynebacterium among various bacteria, whereas Cutibacterium and Staphylococcus, which are abundant at other skin sites, were rare. The relationships between the microbiota and issues related to the umbilicus were investigated and some covariates, including the odor score and several bacteria, were identified. A detailed ana-lysis of the genera associated with odor revealed no correlation with Corynebacterium; however, some minor anaerobic bacteria, such as Mobiluncus, Arcanobacterium, and Peptoniphilus, were more abundant in the high odor score group. Therefore, this technique to collect umbilical dirt provided insights into the microbiota in umbilical dirt and suggested functions for minor anaerobes. Furthermore, since various pathogenic microorganisms were detected, their control may contribute to the prevention of both odor production and infectious diseases caused by these microorganisms.

Post-mega-earthquake geochemical and microbiological properties in subseafloor sediments of the Japan Trench accretionary wedge were investigated using core samples from Hole C0019E, which was drilled down to 851‍ ‍m below seafloor (mbsf) at a water depth of 6,890 m. Methane was abundant throughout accretionary prism sediments; however, its concentration decreased close to the plate boundary decollement. Methane isotope systematics indicated a biogenic origin. The content of mole-cular hydrogen (H2) was low throughout core samples, but markedly increased at specific depths that were close to potential faults predicted by logging-while-drilling ana-lyses. Based on isotopic systematics, H2 appeared to have been abundantly produced via a low-temperature interaction between pore water and the fresh surface of crushed rock induced by earthquakes. Subseafloor microbial cell density remained constant at approximately 105‍ ‍cells‍ ‍mL-1. Amplicon sequences revealed that predominant members at the phylum level were common throughout the units tested, which also included members frequently found in anoxic subseafloor sediments. Metabolic potential assays using radioactive isotopes as tracers revealed homoacetogenic activity in H2-enriched core samples collected near the fault. Furthermore, homoacetogenic bacteria, including Acetobacterium carbinolicum, were isolated from similar samples. Therefore, post-earthquake subseafloor microbial communities in the Japan Trench accretionary prism appear to be episodically dominated by homoacetogenic populations and potentially function due to the earthquake-induced low-temperature generation of H2. These post-earthquake microbial communities may eventually return to the steady-state communities dominated by oligotrophic heterotrophs and hydrogenotrophic and methylotrophic methanogens that are dependent on refractory organic matter in the sediment.

Roseateles depolymerans is an obligately aerobic bacterium that produces a photosynthetic apparatus only under the scarcity of carbon substrates. We herein examined changes in the transcriptomes of R. depolymerans cells to clarify the expression of photosynthesis genes and their upstream regulatory factors under carbon starvation. Transcriptomes 0, 1, and 6‍ ‍h after the depletion of a carbon substrate indicated that transcripts showing the greatest variations (a 500-fold increase [6 h/0 h]) were light-harvesting proteins (PufA and PufB). Moreover, loci with more than 50-fold increases (6 h/0‍ ‍h) were fully related to the photosynthetic gene cluster. Among 13 sigma factor genes, the transcripts of a sigma 70 family sigma factor related to RpoH (SP70) increased along photosynthesis genes under starvation; therefore, a knockout experiment of SP70 was performed. ΔSP70 mutants were found to lack photosynthetic pigments (carotenoids and bacteriochlo-rophyll a) regardless of carbon starvation. We also examined the effects of heat stress on ΔSP70 mutants, and found that SP70 was also related to heat stress tolerance, similar to other RpoH sigma factors (while heat stress did not trigger photosystem production). The deficient accumulation of photosynthetic pigments and the heat stress tolerance of ΔSP70 mutants were both complemented by the introduction of an intact SP70 gene. Furthermore, the transcription of photosynthetic gene operons (puf, puh, and bch) was markedly reduced in the ΔSP70 mutant. The RpoH homologue SP70 was concluded to be a sigma factor that is essential for the transcription of photosynthetic gene operons in R. depolymerans.

Current information on the diversity and evolution of eukaryotic RNA viruses is biased towards host lineages, such as animals, plants, and fungi. Although protists represent the majority of eukaryotic diversity, our understanding of the protist RNA virosphere is still limited. To reveal untapped RNA viral diversity, we screened RNA viruses from 30 marine protist isolates and identified a novel RNA virus named Haloplacidia narnavirus 1 (HpNV1). A phylogenetic ana-lysis revealed that HpNV1 is a new member of the family Narnaviridae. The present study filled a gap in the distribution of narnaviruses and implies their wide distribution in Stramenopiles.

Filamentous fungi grow by the elongation of tubular cells called hyphae and form mycelia through repeated hyphal tip growth and branching. Since hyphal growth is closely related to the ability to secrete large amounts of enzymes or invade host cells, a more detailed understanding and the control of its growth are important in fungal biotechnology, ecology, and pathogenesis. Previous studies using fluorescence imaging revealed many of the molecular mechanisms involved in hyphal growth. Raman microspectroscopy and imaging methods are now attracting increasing attention as powerful alternatives due to their high chemical specificity and label-free, non-destructive properties. Spatially resolved information on the relative abundance, structure, and chemical state of multiple intracellular components may be simultaneously obtained. Although Raman studies on filamentous fungi are still limited, this review introduces recent findings from Raman studies on filamentous fungi and discusses their potential use in the future.

Patescibacteria are widely distributed in various environments and often detected in activated sludge. However, limited information is currently available on their phylogeny, morphology, and ecophysiological role in activated sludge or interactions with other microorganisms. In the present study, we identified microorganisms that interacted with Patescibacteria in activated sludge via a correlation ana-lysis using the 16S rRNA gene, and predicted the metabolic potential of Patescibacteria using a metagenomic ana-lysis. The metagenome-assembled genomes of Patescibacteria consisted of three Saccharimonadia, three Parcubacteria, and one Gracilibacteria, and showed a strong positive correlation of relative abundance with Chitinophagales. Metabolic predictions from ten recovered patescibacterial and five Chitinophagales metagenome-assembled genomes supported mutualistic interactions between a member of Saccharimonadia and Chitinophagales via N-acetylglucosamine, between a member of Parcubacteria and Chitinophagales via nitrogen compounds related to denitrification, and between Gracilibacteria and Chitinophagales via phospholipids in activated sludge. The present results indicate that various interactions between Patescibacteria and Chitinophagales are important for the survival of Patescibacteria in activated sludge ecosystems.

The effects of different types of additional fertilizations (a compound fertilizer and Chiyoda-kasei) on the root-associated microbes of napa cabbage grown in an Andosol field were investigated by molecular community ana-lyses. Most of the closest known species of the bacterial sequences whose relative abundance significantly differed among fertilizers were sensitive to nitrogen fertilization and/or related to the geochemical cycles of nitrogen. The fungal community on the roots of napa cabbage was dominated by two genera, Bipolaris and Olpidium. The relative abundance of these two genera was affected by the types of fertilizers to some extent and showed a strong negative correlation.

The presence of Listeria monocytogenes in piggery effluents intended for irrigation crops may be a source of bacterial dissemination in agriculture. The occurrence and diversity of L. monocytogenes in the farm environment were examined in two pig manure treatment systems (S1 and S2). Samples collected over the course of one year consisted of manure, the liquid fraction of treated manure (lagoon effluent), and soil surrounding the lagoon. L. monocytogenes was enumerated using the Most Probable Number (MPN) method, serotyped by PCR, genotyped by pulsed-field gel electrophoresis (PFGE), and sequenced for multilocus sequence typing (MLST). L. monocytogenes was detected in 92% of manure samples and in approximately 50% of lagoon effluent and soil samples. Concentrations ranged between 5 and 103 MPN 100‍ ‍mL-1. Serogroups IIa, IIb, and IVb were identified. Diversity was high with 44 PFGE profiles (252 isolates) and 17 clonal complexes (CCs) (96 isolates) with higher diversity in manure at site S1 supplied by four farms. Some PFGE profiles and CCs identified in manure or in pig feces from a previous study were also detected in lagoons and/or soil, reflecting pig L. monocytogenes circulation throughout the manure treatment and in the vicinity of the sampling sites. However, some PFGE profiles and CCs were only found in the lagoon and/or in soil, suggesting an origin other than pigs. The present study highlights the limited ability of biological treatments to eliminate L. monocytogenes from pig manure. The persistence of some PFGE profiles and CCs throughout the year in the lagoon and soil shows the ability of L. monocytogenes to survive in this type of environment.