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A series of new metallophores, referred to as frankobactins, were extracted from cultures of the symbiotic and nitrogen-fixing actinobacterium Frankia sp. CH37. Structure elucidation revealed a 2-hydroxyphenyl-substituted oxazoline core and a chain composed of five proteinogenic and nonproteinogenic amino acids, suggesting nonribosomal peptide synthesis as the biosynthetic origin. By whole-genome sequencing, bioinformatic analysis, and comparison with other Frankia strains, the genetic locus responsible for the biosynthesis was detected. Spectrophotometric titration of frankobactin with Fe(III) and Cu(II) and mass spectrometry established the 1:1 (metal:frankobactin) coordination. Uptake experiments suggested that frankobactin A1 (1) did not serve to recruit iron, but to detoxify Cu(II). As frankobactin A1 prevents the cellular entry of Cu(II), it could play a crucial role in the symbiosis of Frankia sp. and its host in the reclamation of copper-contaminated soil.

The phoD-harbouring bacterial community is responsible for much of the hydrolysis of organic phosphorus (Po) in soils and is therefore significant for the improvement of soil phosphorus (P) availability. However, the distribution of phoD-harbouring bacterial community structure and how it regulates the soil P fractions in steppe soils remain largely unknown. It is necessary to assess these relationships to maintain sustainable development on the steppe. We sampled soils from three steppe types across Inner Mongolia, China. Illumina high-throughput sequencing was used to quantify the bacterial phoD gene. The dominant phoD-harbouring genera were Amycolatopsis (5–11%), Bacillus (6–13%), Bradyrhizobium (3–8%) and Pseudomonas (3–5%) across all steppe soils. The relative abundances of phoD-harbouring Amycolatopsis and Bacillus increased significantly as available P (AP) decreased, while the relative abundances of Bradyrhizobium, Pseudomonas and Methylobacterium were significantly positively correlated with AP content. Redundancy analysis showed that the soil stoichiometric ratio of carbon (C), nitrogen (N) and P had a strong effect on the phoD-harbouring bacterial community structure. Correlation analyses further indicated that only phoD-harbouring Dietzia and Sphingomonas had a significant correlation with alkaline phosphatase activity and that they increased P availability by mineralizing Po. phoD-harbouring Frankia and Methylobacterium were positively correlated with labile-Po and negatively correlated with non-labile inorganic P. Moreover, phoD-harbouring Bacillus and Bradyrhizobium promoted the conversion of the non-labile Po pool into the labile Po pool, which can be attributed to microbial immobilization. Not all bacteria carrying the phoD gene promote soil P availability through mineralization or are induced not only in a P-repressible manner. Members of the phoD-harbouring bacterial community employ flexible P use strategies and can be strongly activated by their nutrient preferences and environmental conditions.

Casuarina species are evergreen angiosperm trees native to Australia, South East Asia and the Pacific archipelagos. The genus comprises 14 confirmed species that are able to grow on a wide range of soils. Casuarina spp. are able to enter into an actinorhizal root nodule symbiosis with Frankia casuarinae, nitrogen-fixing soil actinobacteria; the symbiosis renders them independent of soil nitrogen sources. Casuarina spp. can also form symbiotic associations with ectomycorrhizal fungi (EMF) and/or arbuscular mycorrhizal fungi (AMF) for better phosphate supply. Under severe deficiency of phosphate and iron, several Casuarina spp. can form cluster roots. These trees are highly tolerant to salinity, drought, flooding and heavy metal pollution and are extensively used for the rehabilitation of degraded sites. Casuarina spp. plantations also play a significant role in prevention of soil erosion, coastal reclamation, dune stabilization, and as windbreaks in agricultural lands. Due to their multipurpose applications in agriculture and forestry, they have been widely introduced outside their native habitat especially in tropical, arid and semiarid countries across the globe. Therefore, the global distribution of Casuarina spp. has been significantly affected by anthropogenic influences. While in most of these countries, Casuarina spp. have become well-integrated into the regional farming systems, they have become invasive in other regions.

The symbiosis between plants and nitrogen‐fixing bacteria is widespread among legumes and actinorhizal plants within the nitrogen‐fixing root nodule (NFN) clade. However, there are major differences, as well as similarities, in the symbioses between actinorhizal plants and Frankia and those of legumes and their associated rhizobia. This review provides an overview of NFN symbioses. We outline the evolution and biogeography of actinorhizal plants and legumes and compare and contrast their microsymbionts and symbiotic processes. Within the NFN clade, a far greater number of nodulated legumes exists, compared with actinorhizal plants, and legumes have a much wider biogeographical distribution. There are genetic and physiological differences between free‐living diazotrophic Frankia and the phylogenetically diverse rhizobia, most strains of which are unable to fix N2 ex planta. Actinorhizal nodules are modified lateral roots with a central vascular system, whereas legume nodules are stem‐like organs with peripheral vascular systems. Most legumes contain their microsymbionts within symbiosomes, rather than the infection threads found in actinorhizal nodule cells. Legumes have greater control of their microsymbionts, and those within the Inverted Repeat Lacking Clade impose terminal differentiation on their bacteroids. Legumes also have effective processes for autoregulation of nodulation and downregulation of N2 fixation in response to high levels of soil N. These features of the legume‐rhizobia symbiosis have led to increased efficiencies in N2 fixation. Synthesis. We suggest that these characteristic features of the legume‐rhizobia symbiosis, specifically legumes' greater flexibility in the choice of microsymbiont partner and the evolution of increased efficiencies in N2 fixation, are factors that can explain why the majority of species within the Leguminosae have retained the ability to nodulate and how this has contributed to their evolutionary success.

Root and rust rot are common fungal diseases that decrease both the yield and quality of ginseng, resulting in severe economic losses for the ginseng industry. In this study, we explored the bacter...

In this study, we describe the genomes of two novel candidate species of non-nitrogen fixing Frankia that were isolated from the root nodules of Coriaria nepalensis and Alnus glutinosa, genospecies CN and Ag, respectively. Comparative genomic analyses revealed that both genospecies lack genes essential for nitrogen-fixation and possess genes involved in the degradation of plant cell walls. Additionally, we found distinct biosynthetic gene clusters in each genospecies. The availability of these genomes will contribute to the study of the taxonomy and evolution of actinorhizal symbioses.

Nitrogen occurs as inert and inaccessible dinitrogen gaseous form (N

Frankia is a mycelium-forming actinobacterial genus that is found in actinorhizal plants as a nitrogen-fixing facultative endo-symbiont. Frankia has been a commonly uncultured endophytic actinobacteria with very slow growth germination. In the present study, the root nodules from Casuarina spp. were collected at hyper-saline and terrestrial habitats in Tamil Nadu, South India. The root nodules were surface-sterilized, dried, homogenized, and inoculated on the DPM medium with sodium propionate as a carbon source and biotin as a vitamin source. A total of 11 Frankia isolates were obtained from the specified habitats in a solid and liquid medium. The larger number of isolates was recorded in the hyper-saline habitats compared to terrestrial habitats. Finally, 2 predominant growth occurrences of isolates were selected; from each habitat, unique isolates were molecularly characterized and identified as Frankia sp. DDNSF-01 and Frankia casuarinae DDNSF-02. The physico-chemical property analysis of Casuarina rhizosphere soil of hyper-saline habitat showed higher pH, salinity and micro-nutrients like Fe−, Mn−, Cu−, and Zn− than terrestrial habitats, which could be the major factors responsible for the distribution dynamics of Frankia in hyper-saline habitats. Similarly, the biologically active metabolites of Frankia spp. were extracted by the ethyl acetate extraction method and tested by MIC assay, which was performed against emerging multi-host pathogens like Pseudomonas sp. and Candida sp. Likewise, the antibiofilm activity assay was done against the above pathogens. The biofilm inhibition of live/dead cells was confirmed by CLSM microscopic analysis. The F. casuarinae was inhibited the high level of biofilm formation of both pathogens when compared with Frankia sp. The present findings conclude the F. casuarinae is efficient in the control of emerging multi-host pathogens in actinorhizal plants.

Background Alpine alder vegetation acts upon the nearby grass and dwarf shrub vegetation by the nitrogen supply from the symbiotic bacteria Frankia alni of Alnus viridis. This has been studied in two transects concerning plant distribution, plant diversity, nitrate concentration in soil and photosynthetic performance of specific marker plants. Results Away from the alder stand, a band of some meters was dominated by Calamagrostis varia which then was followed by alpine dwarf shrub vegetation. Nitrate in the soil showed a concentration decrease away from the alder stand leading to values near the detection limit in the dwarf shrub zone. Within these three zones, plant species were distributed according to their N-index, given in the ecological literature. Three dominant species, Calamagrostis varia, Rhododendron ferrugineum and Vaccinium myrtillus were examined at sites of different N-availability in the horizontal transect for their photosynthetic performance, by measuring the prompt fluorescence, the OJIP named polyphasic rise of chlorophyll-a fluorescence. All three plant species showed signs of stress in the fluorescence rise kinetics at decreased nitrate availability. These are similar to other known stress effects such as faster reduction of the primary acceptor or an electron supply limitation on the donor site of photosystem II. Conclusion Prompt chlorophyll-a fluorescence data of the examined leaves in a natural vegetation system showed the effects of a decrease in the essential nutrient nitrogen and in a manner parallel to changes in plant diversity. The selected marker plants behaved differently towards decreasing nitrogen concentrations in soil.

The physicochemical profiling of e-waste was done by AAS, XRD, and SEM with EDX that reveals the composition of valuable metals like Au−, Ag−, Zn−, and Cu−. Thus, the present findings provide the important resource of precious metal in the e-waste (PCBs) as pollutant in the environment and human impact. The native isolates Frankia sp. DDNSF-01 and Frankia casuarinae DDNSF-02 were subjected for bioleaching of gold and precious elements bearing e-waste under one and two-step leaching methods. The two-step bioleaching protocol was found effectual in expression with high level of metal leaching within the culture medium of Frankia metabolites and organic acids which were obtained by the leaching of metals. The presence of organic acids functional groups was determined by FT-IR spectrum and by low pH of growth medium. The AAS analysis quantified the initial concentration of metals and their significant increase brought by the leaching. The initial concentration of precious metal elements was found in the e-waste like Au−, Ag−, Cu−, and Zn− (0.04, 0.05, 0.35, 0.1 mg/g) respectively. Frankia sp. leached out precious metals like Au−, Ag−, Cu−, and Zn− (0.2, 0.14, 0.68, 0.2 mg/g) respectively. Frankia casuarinae leached out precious metals like Au−, Ag−, Cu−, and Zn− (0.25, 0.15, 0.55, and 0.2 mg/g) respectively. The precious metal gold was highly leached out by F. casuarinae than Frankia sp.. The valuable metal copper was significantly increased by Frankia sp. than F. casuarinae. Higher gold recovery, at 75%, was obtained for F. casuarinae and copper 94% recovery by Frankia sp. bioleaching of the biooxidised e-waste at 0.2% low pulp densities. The results were confirmed by XRD and SEM with EDX analysis.

The use of wastewater for irrigation in agroforestry is cost-effective for water management. It is well established that rhizospheric microorganisms such as N2-fixing bacteria are able to modulate rhizobioaugmention and to boost phyoremediation process. To date, no study has been conducted to evaluate biological effects of rhizobioaugmentation in Casuarina glauca trees induced by their symbiont N-fixing actinobacteria of the genus Frankia. The objective of the present study was to evaluate the main effects of rhizobioaugmentation on the biological activity in the C. glauca’s rhizosphere and on C. glauca growth in soils irrigated with industrial wastewater. Two Frankia strains (BMG5.22 and BMG5.23) were used in a single or dual inoculations of C. glauca seedlings irrigated with industrial wastewater. Soil enzymes activity related to carbon, phosphorus, sulfur and nitrogen cycling were measured. Results revealed that the BMG5.22 Frankia strain increases significantly the size (dry weight) of C. glauca shoots and roots while dual inoculation increased significantly the root length. Surprisingly, β-glucosidase (BG), cellobiohydrolase (CBH), β-N-acetylglucosaminidase (NAGase), aryl sulfatase (AS), acid phosphatase (AP), alkaline phosphatase (AlP), glycine aminopeptidase (GAP), leucine aminopeptidase (LAP), and peroxidase (PER) activity in the rhizosphere decreased significantly in soils treated with the two strains of symbionts. This suggests no positive correlations between enzymatic activity and C. glauca growth.

Plant interactions with single symbionts of nitrogen fixing bacteria or mycorrhizal fungi are well studied. However, less is known about how plants interact with multiple symbiont species. The aim of this study was to assess whether multiple symbionts had a synergistic effect on host plant performance compared with a single symbiont and whether increasing the number of non-specific fungal species would increase plant performance due to niche complementarity. Green alder (Alnus viridis ssp. crispa) were inoculated with non-specific ectomycorrhizal fungi (Lactarius torminosus, Lactarius theiogalus, Hebeloma crustuliniforme) alone or in combination, with and without Frankia, and grown in nutrient-poor soil. Frankia significantly increased plant growth by 48.5%, doubled the proportion of plants forming ectomycorrhizae, as well as increased root extracellular phosphatase activity by 8.8% compared to non-Frankia treatments. However, increasing number of fungal species decreased nodulated plant biomass. Plants that formed a Hartig net had reduced nodule number and nodule biomass allocation, as well as total nodule activity compared with plants with no Hartig net. Our results indicated that Frankia inoculation provided benefit for both host plants and ectomycorrhizal fungi in the nutrient-poor soil, while ectomycorrhizal fungi did not. These negative effects of mycorrhizal fungi are likely dependent on soil nutrient P availability.

Actinobacterial strain CjT was directly isolated from soil beneath Ceanothus jepsonii growing in the USA. The strain formed cell structures typical of the genus Frankia including extensive hyphae, vesicles and sporangia, and it effectively nodulated members of the actinorhizal Colletieae, Elaeagnaceae and Myricaceae. The whole-cell hydrolysate of strain CjT was rich in meso-diaminopimelic acid and galactose, glucose, mannose, xylose, ribose and a trace of rhamnose. Tbe polar lipid profile contained phosphatidylinositol, phosphatidylglycerol, diphosphatidylglycerol and glycophospholipid. The menaquinone was predominantly MK-9(H4). The fatty acid profile predominantly consisted of C17 : 1ω8c, iso-C16 : 0, C15:0, C16 : 0 and C17 : 0. A multilocus sequence analysis phylogeny based on atp1, ftsZ, dnaK, gyrA and secA gene sequences positioned the strain within Elaeagnaceae- and Colletieae -nodulating species together with Frankia elaeagni DSM 46783T, Frankia discariae DSM 46785T and Frankia irregularis DSM 45899T. Pairwise 16S rRNA gene sequence similarities showed that strain CjT was most closely related to F. discariae DSM 46785T (99.78 %) while their digital DNA–DNA hybridization value was 41.1 %. Based on the overall analyses, strain CjT (=DSM 100623T=CECT 9041T) warrants classification as the type strain of a novel species, for which the name Frankia soli sp. nov. is proposed.

Frankia sp. strain B2 was isolated from Casuarina cunninghamiana nodules. Here, we report the 5.3-Mbp draft genome sequence of Frankia sp. strain B2 with a G+C content of 70.1 % and 4,663 candidate protein-encoding genes. Analysis of the genome revealed the presence of high numbers of secondary metabolic biosynthetic gene clusters.

Rhizobium nitrogen-fixing nodule symbiosis occurs in two taxonomic lineages: legumes (Fabaceae) and the genus Parasponia (Cannabaceae). Both symbioses are initiated upon the perception of rhizobium-secreted lipochitooligosaccharides (LCOs), called Nod factors. Studies in the model legumes Lotus japonicus and Medicago truncatula showed that rhizobium LCOs are perceived by a heteromeric receptor complex of distinct Lys motif (LysM)-type transmembrane receptors named NOD FACTOR RECEPTOR1 (LjNFR1) and LjNFR5 (L. japonicus) and LYSM DOMAIN CONTAINING RECEPTOR KINASE3 (MtLYK3)-NOD FACTOR PERCEPTION (MtNFP; M. truncatula). Recent phylogenomic comparative analyses indicated that the nodulation traits of legumes, Parasponia spp., as well as so-called actinorhizal plants that establish a symbiosis with diazotrophic Frankia spp. bacteria share an evolutionary origin about 110 million years ago. However, the evolutionary trajectory of LysM-type LCO receptors remains elusive. By conducting phylogenetic analysis, transcomplementation studies, and CRISPR-Cas9 mutagenesis in Parasponia andersonii, we obtained insight into the origin of LCO receptors essential for nodulation. We identified four LysM-type receptors controlling nodulation in P. andersonii: PanLYK1, PanLYK3, PanNFP1, and PanNFP2. These genes evolved from ancient duplication events predating and coinciding with the origin of nodulation. Phylogenetic and functional analyses associated the occurrence of a functional NFP2-orthologous receptor to LCO-driven nodulation. Legumes and Parasponia spp. use orthologous LysM-type receptors to perceive rhizobium LCOs, suggesting a shared evolutionary origin of LCO-driven nodulation. Furthermore, we found that both PanLYK1 and PanLYK3 are essential for intracellular arbuscule formation of mutualistic endomycorrhizal fungi. PanLYK3 also acts as a chitin oligomer receptor essential for innate immune signaling, demonstrating functional analogy to CHITIN ELECITOR RECEPTOR KINASE-type receptors.

International audience; Since their introduction at the beginning of the 19th century, Casuarina species have become important components of the Algerian landscape, yet the numbers of species present in Algeria and their identity, distribution, and symbiotic status are still not well understood. A survey conducted on 2,304 trees in 96 Casuarina stands across Algeria indicates that they are represented exclusively by five species: Casuarina cristata Miq., C. cunninghamiana Miq., C. equisetifolia L., C. glauca Sieber ex Spreng. and C. junghuhniana Miq. Two species, C. cunninghamiana and C. glauca, were the most common – the former occurring more often in the relatively humid and temperate areas of the country while the latter is most frequent in the drier and warmer areas. Production of seedlings of C. cunninghamiana and C. glauca in 12 nurseries resulted in low rates of spontaneous mycorrhizal infection frequency and nodulation, suggesting that inoculation of seedlings with selected mycorrhizal fungi and/or Frankia strains in the nursery would be necessary to improve their establishment in the field. This study is the first survey of the geographical distribution of Casuarina species in Algeria and should lead to more efficient utilization of this widely used tree genus in Algeria.; Malgré l’importance prise par les Casua-rina en Algérie depuis leur introductionau début du 19 e siècle, le nombre d’es-pèces aujourd’hui présentes dans lepays, leur identité, leur distribution ainsique leur statut symbiotique sont encorelargement méconnus. Une étude menéesur 2 304 arbres dans 96 peuplementsrépartis dans toute l’Algérie indiquequ’ils sont représentés exclusivementpar cinq espèces du genre Casuarina :Casuarina cristata Miq., C. cunningha-miana Miq., C. equisetifolia L., C. glaucaSieber ex Spreng. et C. junghuhnianaMiq. Deux espèces, C. cunninghamiana etC. glauca, sont largement prédominanteset différemment réparties, la premièreétant plus fréquente dans les zonesrelativement humides et tempérées dupays tandis que la seconde est présentedans les régions les plus arides et lesplus chaudes. La production de plantsde C. cunninghamiana et C. glauca dans12 pépinières situées dans le nord et lesud du pays indique qu’ils présententde faibles taux de mycorhization et denodulation spontanée. Cela suggèrequ’il est nécessaire d’inoculer les jeunesplants avec des souches sélectionnéesde champignons mycorhiziens et/oude Frankia au sein des pépinières afind’améliorer leur installation durable surle terrain. Cette étude constitue le pre-mier inventaire de la répartition géogra-phique des Casuarina en Algérie et peutconduire à une meilleure exploitationdes arbres de ce genre largement utiliséen Algérie

Background Recent studies highlighted the biosynthetic potential of nocardiae to produce diverse novel natural products comparable to that of Streptomyces, thereby making them an attractive source of new drug leads. Many of the 119 Nocardia validly named species were isolated from natural habitats but little is known about the diversity and the potential of the endophytic nocardiae of root nodule of actinorhizal plants. Results The taxonomic status of an actinobacterium strain, designated ncl2T, was established in a genome-based polyphasic study. The strain was Gram-stain-positive, produced substrate and aerial hyphae that fragmented into coccoid and rod-like elements and showed chemotaxonomic properties that were also typical of the genus Nocardia. It formed a distinct branch in the Nocardia 16S rRNA gene tree and was most closely related to the type strains of Nocardia nova (98.6%), Nocardia jiangxiensis (98.4%), Nocardia miyuensis (97.8%) and Nocardia vaccinii (97.7%). A comparison of the draft genome sequence generated for the isolate with the whole genome sequences of its closest phylogenetic neighbours showed that it was most closely related to the N. jiangxiensis, N. miyuensis and N. vaccinii strains, a result underpinned by average nucleotide identity and digital DNA-DNA hybridization data. Corresponding taxogenomic data, including those from a pan-genome sequence analysis showed that strain ncl2T was most closely related to N. vaccinii DSM 43285T. A combination of genomic, genotypic and phenotypic data distinguished these strains from one another. Consequently, it is proposed that strain ncl2T (= DSM 110931T = CECT 30122T) represents a new species within the genus Nocardia, namely Nocardia alni sp. nov. The genomes of the N. alni and N. vaccinii strains contained 36 and 29 natural product-biosynthetic gene clusters, respectively, many of which were predicted to encode for a broad range of novel specialised products, notably antibiotics. Genome mining of the N. alni strain and the type strains of its closest phylogenetic neighbours revealed the presence of genes associated with direct and indirect mechanisms that promote plant growth. The core genomes of these strains mainly consisted of genes involved in amino acid transport and metabolism, energy production and conversion and transcription. Conclusions Our genome-based taxonomic study showed that isolate ncl2T formed a new centre of evolutionary variation within the genus Nocardia. This novel endophytic strain contained natural product biosynthetic gene clusters predicted to synthesize novel specialised products, notably antibiotics and genes associated with the expression of plant growth promoting compounds.