The Presence of the Hairy-Root-Disease-Inducing (Ri) Plasmid in Wheat Endophytic Rhizobia Explains a Pathogen Reservoir Function of Healthy Resistant Plants.

A large number of strains in the Rhizobium radiobacter species complex (biovar 1 Agrobacterium) have been known as causative pathogens for crown gall and hairy root diseases. Strains within this complex were also found as endophytes in many plant species with no symptoms. The aim of this study was to reveal the endophyte variation of this complex and how these endophytic strains differ from pathogenic strains. In this study, we devised a simple but effective screening method by exploiting the high resolution power of mass spectrometry. We screened endophyte isolates from young wheat and barley plants, which are resistant to the diseases, and identified seven isolates from wheat as members of the R. radiobacter species complex. Through further analyses, we assigned five strains to the genomovar (genomic group) G1 and two strains to G7 in R. radiobacter Notably, these two genomovar groups harbor many known pathogenic strains. In fact, the two G7 endophyte strains showed pathogenicity on tobacco, as well as the virulence prerequisites, including a 200-kbp Ri plasmid. All five G1 strains possessed a 500-kbp plasmid, which is present in well-known crown gall pathogens. These data strongly suggest that healthy wheat plants are reservoirs for pathogenic strains of R. radiobacterIMPORTANCE Crown gall and hairy root diseases exhibit very wide host-plant ranges that cover gymnosperm and dicot plants. The Rhizobium radiobacter species complex harbors causative agents of the two diseases. Recently, endophyte isolates from many plant species have been assigned to this species complex. We isolated seven endophyte strains belonging to the species complex from wheat plants and revealed their genomovar affiliations and plasmid profile. The significance of this study is the finding of the genomovar correlation between the endophytes and the known pathogens, the presence of a virulence ability in two of the seven endophyte strains, and the high ratio of the pathogenic strains in the endophyte strains. This study therefore provides convincing evidence that could unravel the mechanism that maintains pathogenic agents of this species and sporadically delivers them to susceptible plants.

Methylotenera oryzisoli sp. nov., a lanthanide-dependent methylotrophic bacteria isolated from rice field soil.

A new lanthanide (Ln3+)-dependent methanol-utilizing bacterial strain, La3113T, was isolated from rice field soil and its taxonomic position was investigated using polyphasic approaches. The strain was aerobic, Gram-stain-negative, strongly motile, catalase-positive and cytochrome oxidase-positive. It could neither catalyse the hydrolysis of urea nor reduce nitrate to nitrite. Growth was observed within a temperature range of 10-40 °C and a pH range of 6-8, with optimum growth at 28 °C and pH 7. Methylamine was utilized as the single source of energy, carbon and nitrogen, and it was oxidized by methylamine dehydrogenase. C16 : 1 ω7c, C16 : 1 ω6c and C16 : 0 were the dominant cellular fatty acids. Its draft genome (2.67 Mbp and 44.9 mol% G+C content) encodes genes including three Ln3+-dependent methanol dehydrogenase (XoxF-type MDH) genes, those for formaldehyde assimilation (ribulose monophosphate pathway), formate dehydrogenases and methylamine dehydrogenases, but not Ca2+-dependent MDH (MxaFI-MDH), which characterizes the species as a Ln3+-dependent methylotroph. The 16S rRNA gene sequence showed that strain La3113T belongs to the genus Methylotenera and is closely related to Methylotenera mobilis JLW8T (98.29 % identity). The digital DNA-DNA hybridization (dDDH) values (less than 30 %) and average nucleotide identity (ANI) values (less than 85 %) between genomes of strain La3113T and related type strains were lower than the thresholds for species delineation (70 % for dDDH and 95-96 % for ANI). On the basis of these polyphasic approaches, we propose a novel Methylotenera species, Methylotenera oryzisoli sp. nov. (type strain La3113T=NBRC 111954T=DSM 103219T).

Honeydew-associated microbes elicit defense responses against brown planthopper in rice.

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.

Bacteria with natural chemotaxis towards methanol revealed by chemotaxis fishing technique.

Motile bacteria often exhibit chemotaxis toward favorable compounds. However, the diversity of bacteria that are attracted to a given substance is largely unknown. This study aimed to reveal the diversity of bacteria with natural chemotaxis towards methanol. We tried to enrich environmental chemotactic bacteria using a glass capillary that is half-filled with methanol solidified with agarose as a trap ("chemotaxis fishing"). The pilot experiment using methanol-chemotactic Methylobacterium aquaticum strain 22A enriched the cells by 46-fold. The method was then applied to bacterial suspensions from paddy water and plants. Depending on the isolation sources and the methods of motility induction, methylotrophic bacteria were enriched 1.2-330-fold. The fished isolates belong to 32 species in 18 genera, mainly containing Acinetobacter, Methylobacterium and Pseudomonas species. Our chemotaxis fishing unveiled a part of diversity of the bacteria with natural chemotaxis towards methanol.

Isolation and genomic characterization of Novimethylophilus kurashikiensis gen. nov. sp. nov., a new lanthanide-dependent methylotrophic species of Methylophilaceae.

Recently, it has been found that two types of methanol dehydrogenases (MDHs) exist in Gram-negative bacterial methylotrophs, calcium-dependent MxaFI-MDH and lanthanide-dependent XoxF-MDH and the latter is more widespread in bacterial genomes. We aimed to isolate and characterize lanthanide-dependent methylotrophs. The growth of strain La2-4T on methanol, which was isolated from rice rhizosphere soil, was strictly lanthanide dependent. Its 16S rRNA gene sequence showed only 93.4% identity to that of Methylophilus luteus MimT , and the name Novimethylophilus kurashikiensis gen. nov. sp. nov. is proposed. Its draft genome (ca. 3.69 Mbp, G + C content 56.1 mol%) encodes 3579 putative CDSs and 84 tRNAs. The genome harbors five xoxFs but no mxaFI. XoxF4 was the major MDH in the cells grown on methanol and methylamine, evidenced by protein identification and quantitative PCR analysis. Methylamine dehydrogenase gene was absent in the La2-4T genome, while genes for the glutamate-mediated methylamine utilization pathway were detected. The genome also harbors those for the tetrahydromethanopterin and ribulose monophosphate pathways. Additionally, as known species, isolates of Burkholderia ambifaria, Cupriavidus necator and Dyadobacter endophyticus exhibited lanthanide dependent growth on methanol. Thus, lanthanide can be used as an essential growth factor for methylotrophic bacteria that do not harbor MxaFI-MDH.

Distribution of 1-aminocyclopropane-1-carboxylate deaminase and D-cysteine desulfhydrase genes among type species of the genus Methylobacterium.

The presence of 1-aminocyclopropane-1-carboxylate (ACC) deaminase determines the ability of bacteria to increase the resistance of plants to various types of stress. The genes of ACC deaminase (acdS) and the closely related enzyme D-cysteine desulfhydrase (dcyD) were searched in type strains of various representatives of the genus Methylobacterium. Using PCR screening and in silico searching in the available complete genome sequences of type strains, the genes were found in 28 of 48 species of the genus. Phylogenetic analysis of amino acid sequences of proteins revealed two large groups of sequences of the AcdS protein and one of the DcyD protein. The distribution of these groups correlates well with the phylogenetic tree based on the sequences of the 16S rRNA genes, which apparently indicates a different evolutionary adaptation to association with plants in the representatives of these groups. For the first time for aerobic methylotrophs it was demonstrated that the gene dcyD encodes D-cysteine desulfhydrase by cloning and recombinant protein characterization.

Oharaeibacter diazotrophicus gen. nov., sp. nov., a diazotrophic and facultatively methylotrophic bacterium, isolated from rice rhizosphere.

A novel facultatively methanol-utilizing bacterial strain, SM30T, was isolated from rice rhizosphere. Strain SM30T was Gram-stain-negative, aerobic, motile, short rods, and grew optimally at pH 7 and at 28 °C. It could tolerate 0 to 2 % (w/v) NaCl. Based on 16S rRNA gene sequence comparisons, strain SM30T was most closely related to Pleomorphomonas oryzae DSM 16300T, with a low similarity of 94.17 %. One of the lanthanide metals, lanthanum, could enhance its growth slightly on methanol. Phylogenetic trees, based on the mxaF, xoxF and cpn60 genes of SM30T showed its distinct phylogenetic position with respect to species with validly published names. Polymerase chain reaction (PCR) amplification of the nifH and growth on nitrogen-free medium indicated that strain SM30T is a diazotroph. The major cellular fatty acids were summed feature 8 (containing 18 : 1ω7c and 18 : 1ω6c) and cyclo 19 : 0ω8c. The major quinone was ubiquinone 10. The DNA G+C content was 74.6 mol%. Based on the genotypic and phenotypic characteristics, strain SM30T represents a novel genus and species, for which the name Oharaeibacter diazotrophicus gen. nov., sp. nov. is proposed with the type strain SM30T (=NBRC 111955T=DSM 102969T).

Widespread endogenization of genome sequences of non-retroviral RNA viruses into plant genomes.

Non-retroviral RNA virus sequences (NRVSs) have been found in the chromosomes of vertebrates and fungi, but not plants. Here we report similarly endogenized NRVSs derived from plus-, negative-, and double-stranded RNA viruses in plant chromosomes. These sequences were found by searching public genomic sequence databases, and, importantly, most NRVSs were subsequently detected by direct molecular analyses of plant DNAs. The most widespread NRVSs were related to the coat protein (CP) genes of the family Partitiviridae which have bisegmented dsRNA genomes, and included plant- and fungus-infecting members. The CP of a novel fungal virus (Rosellinia necatrix partitivirus 2, RnPV2) had the greatest sequence similarity to Arabidopsis thaliana ILR2, which is thought to regulate the activities of the phytohormone auxin, indole-3-acetic acid (IAA). Furthermore, partitivirus CP-like sequences much more closely related to plant partitiviruses than to RnPV2 were identified in a wide range of plant species. In addition, the nucleocapsid protein genes of cytorhabdoviruses and varicosaviruses were found in species of over 9 plant families, including Brassicaceae and Solanaceae. A replicase-like sequence of a betaflexivirus was identified in the cucumber genome. The pattern of occurrence of NRVSs and the phylogenetic analyses of NRVSs and related viruses indicate that multiple independent integrations into many plant lineages may have occurred. For example, one of the NRVSs was retained in Ar. thaliana but not in Ar. lyrata or other related Camelina species, whereas another NRVS displayed the reverse pattern. Our study has shown that single- and double-stranded RNA viral sequences are widespread in plant genomes, and shows the potential of genome integrated NRVSs to contribute to resolve unclear phylogenetic relationships of plant species.

Extensive reduction of cell viability and enhanced matrix production in Pseudomonas aeruginosa PAO1 flow biofilms treated with a D-amino acid mixture.

Treatment of Pseudomonas aeruginosa PAO1 flow biofilms with a D-amino acid mixture caused significant reductions in cell biomass by 75% and cell viability by 71%. No biofilm disassembly occurred, and matrix production increased by 30%, thereby providing a thick protective cover for remaining viable or persister cells.

Methylobacterium haplocladii sp. nov. and Methylobacterium brachythecii sp. nov., isolated from bryophytes.

Pink-pigmented, facultatively methylotrophic bacteria, strains 87e(T) and 99b(T), were isolated from the bryophytes Haplocladium microphyllum and Brachythecium plumosum, respectively. The cells of both strains were Gram-reaction-negative, motile, non-spore-forming rods. On the basis of 16S rRNA gene sequence similarity, strains 87e(T) and 99b(T) were found to be related to Methylobacterium organophilum ATCC 27886(T) (97.1% and 97.7%, respectively). Strains 87e(T) and 99b(T) showed highest 16S rRNA gene similarity to Methylobacterium gnaphalii 23e(T) (98.3 and 99.0%, respectively). The phylogenetic similarities to all other species of the genus Methylobacterium with validly published names were less than 97%. Major cellular fatty acids of both strains were C(18:1)ω7c and C(18:0). The results of DNA-DNA hybridization, phylogenetic analyses based on 16S rRNA and cpn60 gene sequences, fatty acid profiles, whole-cell matrix-assisted, laser-desorption/ionization time-of-flight mass spectrometry (MALDI-TOF/MS) analysis, and physiological and biochemical tests allowed genotypic and phenotypic differentiation of strains 87e(T) and 99b(T) from their phylogenetically closest relatives. We propose that strains 87e(T) and 99b(T) represent novel species within the genus Methylobacterium, for which the names Methylobacterium haplocladii sp. nov. (type strain 87e(T) =DSM 24195(T) =NBRC 107714(T)) and Methylobacterium brachythecii sp. nov. (type strain 99b(T) =DSM 24105(T) =NBRC 107710(T)) are proposed.

Dominant colonization and inheritance of Methylobacterium sp. strain OR01 on perilla plants.

Pink-pigmented facultative methylotrophs (PPFMs) are major inhabitants of the phyllosphere. In a preceding study, we found that perilla plants harbor a dominant population of PPFMs on their leaves and seeds, and that the closest relative of PPFMs (Methylobacterium sp. strain OR01 as representative strain) isolated from red perilla seeds was M. fujisawaense DSM5686(T). In the present study, the specific interaction between red perilla and Methylobacterium species was investigated. All the PPFMs isolated from red perilla seeds harvested in the Ohara area of Kyoto, Japan in 2009, 2010, and 2011 and the PPFMs isolated from red perilla leaves planted at four geographically different places in Japan had 16S rRNA sequences identical to that of strain OR01. Direct transmission of PPFMs from seeds to leaves and the competitiveness of strain OR01 were confirmed. This report is the first step toward understanding the species-level specificity of the interaction between perilla plants and Methylobacterium species.

Preparation and application of cinnamon-Litsea cubeba compound essential oil microcapsules for peanut kernel postharvest storage.

This research developed a novel, efficient and safe antimildew for peanut kernel postharvest storage. The antimildew, cinnamon-Litsea cubeba compound essential oil (CLCEO) microcapsule (CLCEOM), was synthesized with CLCEO as core materials and ß-cyclodextrin as wall materials. Fourier transform infrared spectroscopy and gas chromatography-mass spectrometry analyses indicated that major antifungal compounds of CLCEO were encapsulated in the cavity of ß-cyclodextrin. The inhibition zone experiment showed that CLCEOM retained antifungal effect on Aspergillus spp. strains even after storage for 2 months at 4 â„ƒ. Besides, CLCEOM reduced total number of fungal colonies, relative abundance of Aspergillus spp., and aflatoxin B1 content of peanut kernels, and had positive effect on slowing down the increase in acid value of peanut oil without causing any adverse effect on the viability and sensory properties during storage process. Overall, CLCEOM presented good preservative effects on peanut kernels, providing evidence for its potential use as antimildew for peanut storage.

Metabolism-linked methylotaxis sensors responsible for plant colonization in Methylobacterium aquaticum strain 22A.

Motile bacteria take a competitive advantage in colonization of plant surfaces to establish beneficial associations that eventually support plant health. Plant exudates serve not only as primary growth substrates for bacteria but also as bacterial chemotaxis attractants. A number of plant-derived compounds and corresponding chemotaxis sensors have been documented, however, the sensors for methanol, one of the major volatile compounds released by plants, have not been identified. Methylobacterium species are ubiquitous plant surface-symbiotic, methylotrophic bacteria. A plant-growth promoting bacterium, M. aquaticum strain 22A exhibits chemotaxis toward methanol (methylotaxis). Its genome encodes 52 methyl-accepting chemotaxis proteins (MCPs), among which we identified three MCPs (methylotaxis proteins, MtpA, MtpB, and MtpC) responsible for methylotaxis. The triple gene mutant of the MCPs exhibited no methylotaxis, slower gathering to plant tissues, and less efficient colonization on plants than the wild type, suggesting that the methylotaxis mediates initiation of plant-Methylobacterium symbiosis and engages in proliferation on plants. To examine how these MCPs are operating methylotaxis, we generated multiple gene knockouts of the MCPs, and Ca2+-dependent MxaFI and lanthanide (Ln3+)-dependent XoxF methanol dehydrogenases (MDHs), whose expression is regulated by the presence of Ln3+. MtpA was found to be a cytosolic sensor that conducts formaldehyde taxis (formtaxis), as well as methylotaxis when MDHs generate formaldehyde. MtpB contained a dCache domain and exhibited differential cellular localization in response to La3+. MtpB expression was induced by La3+, and its activity required XoxF1. MtpC exhibited typical cell pole localization, required MxaFI activity, and was regulated under MxbDM that is also required for MxaF expression. Strain 22A methylotaxis is realized by three independent MCPs, two of which monitor methanol oxidation by Ln3+-regulated MDHs, and one of which monitors the common methanol oxidation product, formaldehyde. We propose that methanol metabolism-linked chemotaxis is the key factor for the efficient colonization of Methylobacterium on plants.

Methylobacterium oxalidis sp. nov., isolated from leaves of Oxalis corniculata.

A pink-pigmented, facultatively methylotrophic bacterium, strain 35a(T), was isolated from the leaves of Oxalis corniculata. Cells of strain 35a(T) were Gram-reaction-negative, motile, non-spore-forming rods. The highest 16S rRNA gene pairwise sequence similarities for strain 35a(T) were found with the strains of Methylobacterium iners 5317S-33(T) (96.7%), 'Methylobacterium soli' YIM 48816 (96.6%) and Methylobacterium jeotgali S2R03-9(T) (96.3%). 16S rRNA gene sequence similarities with the type strains of all other recognized species of the genus Methylobacterium were below 96%. Major cellular fatty acids were C(18:1)ω7c, C(18:0) and C(16:0). The results of DNA-DNA hybridization experiments, analysis of cpn60 gene sequences, fatty acid profiles, whole-cell MALDI-TOF/MS spectral pattern analysis, and physiological and biochemical tests allowed genotypic and phenotypic differentiation of strain 35a(T) from its nearest phylogenetic neighbours. Strain 35a(T) is therefore considered to represent a novel species within the genus Methylobacterium, for which the name Methylobacterium oxalidis sp. nov. is proposed. The type strain is 35a(T) (=DSM 24028(T)=NBRC 107715(T)).

Methylobacterium gnaphalii sp. nov., isolated from leaves of Gnaphalium spicatum.

A pink-pigmented, facultatively methylotrophic bacterium, strain 23e(T), was isolated from the leaves of Gnaphalium spicatum (cudweed). The cells of strain 23e(T) were Gram-reaction negative, motile and non-spore-forming rods. On the basis of 16S rRNA gene sequence similarities, strain 23e(T) was related to Methylobacterium organophilum ATCC 27886(T) (97.1%) and Methylobacterium marchantiae JT1(T) (97%), and the phylogenetic similarities to all other Methylobacterium species with validly published names were less than 97%. Major cellular fatty acids were C(18:1)ω7c, C(16:00) and C(18:0). The results of DNA-DNA hybridization, phylogenetic analyses based on 16S rRNA and cpn60 gene sequences, fatty acid profiles, whole-cell matrix-assisted laser desorption/ionization time of flight/MS analysis, physiological and biochemical tests allowed genotypic and phenotypic differentiation of strain 23e(T) from the phylogenetically closest relatives. We propose that strain 23e(T) represents a novel species within the genus Methylobacterium, for which the name Methylobacterium gnaphalii sp. nov. is proposed. The type strain is 23e(T) (=DSM 24027(T)=NBRC 107716(T)).

Isolation and characterization of Kluyvera georgiana strain with the potential for acrylamide biodegradation.

Worldwide contamination by acrylamide, a neurotoxicant and carcinogen in animals, is becoming a significant problem. We isolated three novel acrylamide-degrading bacteria from domestic wastewater in Chonburi, Thailand. Using biochemical characteristics and 16S rRNA gene sequencing, the strains were identified as Klebsiella pneumoniae, Kluyvera georgiana and Enterococcus faecalis. K. georgiana strain No. 2 was selected for further characterization due to its degradation potential of high concentrations of acrylamide at the mesophilic temperatures. The strain grew well in the presence of acrylamide at concentrations to 0.5 % (w/v), pH 5.0 to 7.0 and 37°C. Degradation of acrylamide to acrylic acid began after 30 min of cultivation as a biomass-dependent manner. Mass balance analysis revealed 92.3 % conversion of acrylamide to acrylic acid and two lower polarity compounds. Strain No. 2 degraded many aliphatic amides but not iodoacetamide and thioacetamide. High degradation level (>80 %) was found with propionamide, cyanoacetamide and acetamide. Moderate degradation was obtained in the order of formamide > butyramide > lactamide > urea while sodium azide provided 34 % degradation. These findings render this novel bacterium attractive for biodegradation of acrylamide and other aliphatic amides in the environment.

A Periplasmic Lanthanide Mediator, Lanmodulin, in Methylobacterium aquaticum Strain 22A.

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 (Ln3+)-dependent MDH (XoxF), whose expression is regulated by the availability of Ln3+. 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 Ln3+. The transport of Ln3+ into the cytosol enhances XoxF expression. A unique protein named lanmodulin from Methylorubrum extorquens strain AM1 was identified as a specific Ln3+-binding protein, and its biological function was implicated to be an Ln3+ shuttle in the periplasm. In contrast, it remains unclear how Ln3+ levels in the cells are maintained, because Ln3+ 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 Ln3+). The expression level of mxcQE was altered to be negatively dependent on Ln3+ concentration in ∆lanM, whereas it was constant in the wild type. Furthermore, when exposed to La3+, ∆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.

The Antifungal Activity of Cinnamon-Litsea Combined Essential Oil against Dominant Fungal Strains of Moldy Peanut Kernels.

The antifungal activity of cinnamon (Cinnamomum cassia Presl), litsea [Litsea cubeba (Lour.) Pers.], clove (Syzygium aromaticum L.), thyme (Thymus mongolicus Ronn.) and citronella (Cymbopogon winterianus Jowitt) essential oils (EOs) against the dominant fungi isolated from moldy peanuts was investigated in this research. Firstly, strain YQM was isolated and identified by morphological characterization and 18S rRNA gene sequence analysis to be Aspergillus flavus (A. flavus). Next, antifungal effects of single or mixed EOs on strain YQM were evaluated by the inhibition zone test. The cinnamon-litsea combined essential oil (CLCEO, Vcinnamon oil:Vlitsea oil = 3:5) displayed the best antifungal effect on strain YQM. The chemical composition of CLCEO was identified and quantified by gas chromatograph-mass spectrometry (GC-MS), and results revealed that the major components of CLCEO were cinnamaldehyde and citral. Finally, the effect of EOs on the microstructure of strain YQM mycelia was observed under scanning electron microscope (SEM). The mycelia exposed to cinnamon essential oil (CEO) and litsea essential oil (LEO) were partly deformed and collapsed, while the mycelia treated with CLCEO were seriously damaged and the deformation phenomena such as shrinking, shriveling and sinking occurred. Therefore, CLCEO has great potential for using as anti-mildew agents during peanut storage.

Siderophore for Lanthanide and Iron Uptake for Methylotrophy and Plant Growth Promotion in Methylobacterium aquaticum Strain 22A.

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.

Pandoraea oxalativorans sp. nov., Pandoraea faecigallinarum sp. nov. and Pandoraea vervacti sp. nov., isolated from oxalate-enriched culture.

Five isolates, designated TA2, TA4, TA25(T), KOx(T) and NS15(T) were isolated in previous studies by enrichment in mineral medium with potassium oxalate as the sole carbon source and were characterized using a polyphasic approach. The isolates were Gram-reaction-negative, aerobic, non-spore-forming rods. Phylogenetic analyses based on 16S rRNA and DNA gyrase B subunit (gyrB) gene sequences confirmed that the isolates belonged to the genus Pandoraea and were most closely related to Pandoraea sputorum and Pandoraea pnomenusa (97.2-99.7 % 16S rRNA gene sequence similarity). The isolates could be differentiated from their closest relatives on the basis of several phenotypic characteristics. The major cellular fatty acid profiles of the isolates comprised C16:0, C18:1ω7c, C17:0 cyclo and summed feature 3 (C16:1ω7c and/or iso-C15:0 2-OH). On the basis of DNA-DNA hybridization studies and phylogenetic analyses, the isolates represent three novel species within the genus Pandoraea, for which the names Pandoraea oxalativorans sp. nov. (TA25(T)  = NBRC 106091(T)  = CCM 7677(T)  = DSM 23570(T)), Pandoraea faecigallinarum sp. nov. (KOx(T)  = NBRC 106092(T)  = CCM 2766(T)  = DSM 23572(T)) and Pandoraea vervacti sp. nov. (NS15(T)  = NBRC 106088(T)  = CCM 7667(T)  = DSM 23571(T)) are proposed.