Naoto Tanaka

Salt-free lacto-fermentation is gaining attention owing to the global demand for sodium reduction. Sunki, a traditional unsalted Japanese turnip pickle, provides a unique ecological model. In this study, we investigated the microbial balance and systemic metabolic pathways that promote organic acid accumulation using shotgun metagenomics. We revealed that a high-quality sunki cluster exhibiting an abundant accumulation of organic acids (lactic acid 8.58 ± 2.42 g/L, acetic acid 2.70 ± 0.76 g/L, succinic acid 1.50 ± 0.52 g/L) and low pH (3.91 ± 0.15) was characterized by the co-dominance of specific lactic acid bacteria along with the foundational species Limosilactobacillus fermentum. This co-dominance enables the synergistic operation of both the homofermentative and heterofermentative pathways, whose functional potentials positively correlated with lactic acid (r = 0.50) and acetic acid (r = 0.48) concentrations, respectively, thereby achieving both effective preservation and flavor complexity. Furthermore, we propose a dual-pathway model for succinic acid production involving the reductive tricarboxylic acid cycle from malate and the γ-aminobutyric acid (GABA) shunt. Succinic acid positively correlated with the functional potential of the GABA shunt enzyme (puuE, r = 0.57) and negatively correlated with that of glutamate dehydrogenase (gdhA, r = −0.51), suggesting a metabolic switch redirecting glutamate flux toward the GABA shunt. This mechanism highlights an advanced ecological adaptation for pH homeostasis, emphasizing that a balanced co-dominance structure determines the quality of salt-free fermented foods.

The taxonomic status of two gram-negative, whitish-pigmented motile bacteria KMM 9576^T and KMM 9553 isolated from a sandy sediment sample from the Sea of Japan seashore was defined. Phylogenetic analysis revealed that strains KMM 9576^T and KMM 9553 represent a distinct lineage within the family Rhizobiaceae, sharing 100% 16S rRNA sequence similarity and 99.5% average nucleotide identity (ANI) to each other. The strains showed the highest 16S rRNA sequence similarities of 97.4% to Sinorhizobium garamanticum LMG 24692^T, 96.9% to Ensifer adhaerens NBRC 100388^T, and 96.8% to Pararhizobium giardinii NBRC 107135^T. The ANI values between strain KMM 9576^T and Ensifer adhaerens NBRC 100388^T, Sinorhizobium fredii USDA 205^T, Pararhizobium giardinii NBRC 107135^T, and Rhizobium leguminosarum NBRC 14778^T were 79.9%, 79.6%, 79.4%, and 79.2%, respectively. The highest core-proteome average amino acid identity (cpAAI) values of 82.1% and 83.1% were estimated between strain KMM 9576^T and Rhizobium leguminosarum NBRC 14778^T and ‘Rhizobium album’ NS-104, respectively. The DNA GC contents were calculated from a genome sequence to be 61.5% (KMM 9576^T) and 61.4% (KMM 9553). Both strains contained the major ubiquinone Q-10 and C_18:1ω7c as the dominant fatty acid followed by 11-methyl C_18:1ω7c and C_19:0 cyclo, and polar lipids consisted of phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, an unidentified aminophospholipid, and two unidentified phospholipids. Based on phylogenetic and phylogenomic analyses, and phenotypic characterization, strains KMM 9576^T and KMM 9553 are concluded to represent a novel genus and species, for which the name Fererhizobium litorale gen. nov., sp. nov. is proposed. The type strain of the type species is strain KMM 9576^T (=NRIC 0957^T).

This study was designed to assess the suppressive effects of various anaerobically digested slurries (ADSs), and the microorganisms inhabiting them, against Fusarium wilt in spinach. We used five different ADSs from a range of source materials (dairy cow manure, sewage sludge, food garbage, pig manure, night soil sludge), combined in different proportions. All five raw ADSs suppressed the growth of Fusarium oxysporum f. sp. spinaciae (Fos) on agar plates using a co-culture test. In contrast, filtrate ADSs did not suppress the growth of Fos. In total, 32 bacterial strains were isolated from five ADSs, and eight isolates showed antagonistic activities against Fos. Based on 16S rDNA sequences, the strain AD-3 isolated from ADS from dairy cow manure belonged to Bacillus velezensis. Genome analysis revealed that AD-3 had two kinds of genes related to the production of the non-ribosomal lipopeptides, fengycin/plipastatin (pps genes), and surfactin (srf genes). In pot assays, inoculation of AD-3 (1.0 × 10⁶ CFU·g⁻¹ dry soil) into Fos-infected soil (1.0 × 10⁵ bud-cells·g⁻¹ dry soil) significantly reduced the severity of Fusarium wilt disease at 28 d after seedling. The percentage reductions in disease severity in two replicates were 64.3% and 44.3%, respectively. Thus, bacterial strain AD-3 could be applied to reduce Fusarium wilt in spinach.

This study was designed to assess the suppressive effects of various anaerobically digested slurries (ADSs), and the microorganisms inhabiting them, against Fusarium wilt in spinach. We used five different ADSs from a range of source materials (dairy cow manure, sewage sludge, food garbage, pig manure, night soil sludge), combined in different proportions. All five raw ADSs suppressed the growth of Fusarium oxysporum f. sp. spinaciae (Fos) on agar plates using a co-culture test. In contrast, filtrate ADSs did not suppress the growth of Fos. In total, 32 bacterial strains were isolated from five ADSs, and 8 isolates showed antagonistic activities against Fos. Based on 16S rDNA sequences, the strain AD-3 isolated from ADS from dairy cow manure belonged to Bacillus velezensis. Genome analysis revealed that AD-3 had two kinds of genes related to the production of the non-ribosomal lipopeptides, fengycin/plipastatin (pps genes), and surfactin (srf genes). In pot assays, inoculation of AD-3 (1.0 × 106 CFU·g−1 dry soil) into Fos-infected soil (1.0 × 105 bud-cells·g−1 dry soil) significantly reduced the severity of Fusarium wilt disease at 28 d after seedling. The percentage reductions in disease severity in two replicates were 64.3% and 44.3%, respectively. Thus, bacterial strain AD-3 could be applied to reduce Fusarium wilt in spinach.

Siderophores are produced by several bacteria that utilise iron in various environments. Elucidating the structure of a specific siderophore may have valuable applications in drug development. Stenotrophomonas maltophilia , a Gram-negative bacterium that inhabits a wide range of environments and can cause pneumonia, produces siderophores. However, the structure was unknown, and therefore, in this study, we aimed to elucidate it. We purified siderophores from cultures of S. maltophilia K279a using preparative reversed-phase HPLC. The structure was analysed through LC-MS and ¹H and ¹³C NMR. The results demonstrated that S. maltophilia K279a produces 2,3-dihydroxybenzoylserine (DHBS), a monomer unit of enterobactin. We suggested the uptake of Iron(III) by the DHBS complex. DHBS production by S. maltophilia K279a could be attributed to an incomplete enterobactin pathway. Drugs targeting DHBS synthesis could prevent S. maltophilia infection.

This study investigated endophytic nitrogen-fixing bacteria isolated from two species of yam (water yam, Dioscorea alata L.; lesser yam, Dioscorea esculenta L.) grown in nutrient-poor alkaline soil conditions on Miyako Island, Okinawa, Japan. Two bacterial strains of the genus Rhizobium , S-93^T and S-62, were isolated. The phylogenetic tree, based on the almost-complete 16S rRNA gene sequences (1476 bp for each strain), placed them in a distinct clade, with Rhizobium miluonense CCBAU 41251^T, Rhizobium hainanense I66^T, Rhizobium multihospitium HAMBI 2975^T, Rhizobium freirei PRF 81^T and Rhizobium tropici CIAT 899^T being their closest species. Their bacterial fatty acid profile, with major components of C_19 : 0 cyclo ω8c and summed feature 8, as well as other phenotypic characteristics and DNA G+C content (59.65 mol%) indicated that the novel strains belong to the genus Rhizobium . Pairwise average nucleotide identity analyses separated the novel strains from their most closely related species with similarity values of 90.5, 88.9, 88.5, 84.5 and 84.4 % for R. multihospitium HAMBI 2975^T, R. tropici CIAT 899^T, R. hainanense CCBAU 57015^T, R. miluonense HAMBI 2971^T and R. freirei PRF 81^T, respectively; digital DNA–DNA hybridization values were in the range of 26–42 %. Considering the phenotypic characteristics as well as the genomic data, it is suggested that strains S-93^T and S-62 represent a new species, for which the name Rhizobium dioscoreae is proposed. The type strain is S-93^T (=NRIC 0988^T=NBRC 114257^T=DSM 110498^T).

A deep-rooting upland rice variety (Kinandang Patong) was evaluated for its growth response to bio-fertilization at early stages. Five non-native plant growth-promoting bacteria previously isolated from yams (Dioscorea spp.) were inoculated to upland rice under growth chamber and greenhouse conditions. Effects of the inoculation varied depending on bacterial strains and growing conditions. Growth of 14-day rice seedlings was improved by all tested bacterial strains. Under growth chamber, the strain S-333 increased plant length, shoot dry weight and nitrogen content as compared to the control, but total dry weight, nitrogen uptake, leaf chlorophyll content and number of tillers were higher with N fertilizer application. Under greenhouse conditions, most rice growth parameters were improved by inoculation with the strain S-7. The correlations between the bacterial plant-growth-promoting traits and rice growth parameters under growth chamber conditions were all negative for phosphate solubilization indexes. Our results suggest that bacterial inoculation can replace half (S-343 and S-611) of or the full (S-7) rate of chemical N fertilizer required, depending on bacterial strains and growing environments, although δ¹⁵N value in control plants was lower than in inoculated plants under growth chamber conditions, suggesting that the bacteria improve rice growth through mechanisms other than biological nitrogen fixation.

乳酸菌は多岐にわたってヒトの生活に貢献する有用な微生物であり、資源として今後もさらに開拓されていく主要微生物であることは間違いない。長年に渡る乳酸菌の研究により、乳酸菌の性質をうまく活用した効率的な分離技術が確立されているため、微生物を扱う基本的な技術があれば、初めてであっても容易に分離できる。ここでは特にこれから乳酸菌の分離を始める方を対象に、乳酸菌という生き物についての説明も交えながらその乳酸菌の分離方法と乳酸菌を分離する時に知っておくべきことおよび考えるべきことについて述べる。

Fructobacillus spp. in fructose-rich niches belong to the family Leuconostocaceae. They were originally classified as Leuconostoc spp., but were later grouped into a novel genus, Fructobacillus, based on their phylogenetic position, morphology and specific biochemical characteristics. The unique characters, so called fructophilic characteristics, had not been reported in the group of lactic acid bacteria, suggesting unique evolution at the genome level. Here we studied four draft genome sequences of Fructobacillus spp. and compared their metabolic properties against those of Leuconostoc spp. Fructobacillus species possess significantly less protein coding sequences in their small genomes. The number of genes was significantly smaller in carbohydrate transport and metabolism. Several other metabolic pathways, including TCA cycle, ubiquinone and other terpenoid-quinone biosynthesis and phosphotransferase systems, were characterized as discriminative pathways between the two genera. The adhE gene for bifunctional acetaldehyde/alcohol dehydrogenase, and genes for subunits of the pyruvate dehydrogenase complex were absent in Fructobacillus spp. The two genera also show different levels of GC contents, which are mainly due to the different GC contents at the third codon position. The present genome characteristics in Fructobacillus spp. suggest reductive evolution that took place to adapt to specific niches.

An aerobic, Gram-stain-negative, agarolytic rod-shaped bacterium, designated KMM 9504^T, was isolated from a sediment sample collected from the seashore of the Sea of Japan. Comparative 16S rRNA gene sequence analysis showed that strain KMM 9504^Tbelonged to the genus Simiduia as it was most closely related to Simiduia areninigrae KCTC 23293^T(97.3 % sequence similarity). Strain KMM 9504^Twas characterized by the major ubiquinone Q-8, and by the predominance of C_16 : 1ω7c, C_17 : 1ω8c, followed by C_16 : 0, C_15 : 0, C_17 : 0and C_12 : 1in its fatty acid profile. Polar lipids consisted of phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, an unknown aminophospholipid, an unknown aminolipid, unknown phospholipids, and unknown lipids. Based on the distinctive phenotypic characteristics, phylogenetic analysis and DNA–DNA hybridization results, it is concluded that strain KMM 9504^Trepresents a novel species of the genus Simiduia , for which the nameSimiduialitoreasp. nov. is proposed. The type strain of the species is strain KMM 9504^T( = NRIC 0917^T = JCM 19759^T).