ABSTRACT
Nearly 50 years after the ground-breaking isolation of the primary Comptonia peregrina microsymbiont under axenic conditions, efforts to isolate a substantial number of Protofrankia and Frankia strains continue with enduring challenges and complexities. This study aimed to streamline genomic insights through comparative and predictive tools to extract traits crucial for isolating specific Frankia in axenic conditions. Pangenome analysis unveiled significant genetic diversity, suggesting untapped potential for cultivation strategies. Shared metabolic strategies in cellular components, central metabolic pathways, and resource acquisition traits offered promising avenues for cultivation. Ecological trait extraction indicated that most uncultured strains exhibit no apparent barriers to axenic growth. Despite ongoing challenges, potential caveats, and errors that could bias predictive analyses, this study provides a nuanced perspective. It highlights potential breakthroughs and guides refined cultivation strategies for these yet-uncultured strains. We advocate for tailored media formulations enriched with simple carbon sources in aerobic environments, with atmospheric nitrogen optionally sufficient to minimize contamination risks. Temperature adjustments should align with strain preferences-28-29°C for Frankia and 32-35°C for Protofrankia-while maintaining an alkaline pH. Given potential extended incubation periods (predicted doubling times ranging from 3.26 to 9.60 days, possibly up to 21.98 days), patience and rigorous contamination monitoring are crucial for optimizing cultivation conditions.
ABSTRACT
A comprehensive polyphasic taxonomic investigation integrating taxongenomic criteria was conducted on strain IRAMC:0171T isolated from the root nodules of Retama raetam in Tunisia. This Gram-stain-negative and aerobic bacterium thrived within a temperature range of 5-45 °C, optimal at 28 °C, and tolerated salt concentrations from 0-6â% NaCl, with an optimal range of 0-3â%. It displayed pH tolerance from pH 4 to 10, thriving best at pH 6.8-7.5. Chemotaxonomically, strain IRAMC:0171T was characterized by diphosphatidylglycerol, phosphatidylglycerol, phosphatidylcholine, and phosphatidylethanolamine as polar lipids. Its predominant fatty acid composition was C18â:â1 ω7c (61.2â%), and the primary ubiquinone was Q10 (97â%). Analysis of the 16S rRNA gene of strain IRAMC:0171T showed 99.08â% similarity to Mesorhizobium waimense ICMP 19557T, Mesorhizobium amorphae ACCC 19665T, and Mesorhizobium huakuii IAM 14158. However, digital DNA-DNA hybridization and average nucleotide identity analyses revealed values ranging from 21.1 to 25.2â% and 77.05 to 82.24â%, respectively, signifying significant deviation from established species demarcation thresholds. Phylogenetic studies, encompassing 16S rRNA, whole-genome-based tree reconstruction, and core protein analysis, positioned strain IRAMC:0171T closest to Mesorhizobium terrae KCTC 72278T and 'Mesorhizobium hungaricum' UASWS1009T, forming together a distinct branch within the genus Mesorhizobium. In consideration of this comprehensive data, we propose strain IRAMC:0171T (=DSM 112841T=CECT 30767T) as the type strain of a new species named Mesorhizobium retamae sp. nov.
Subject(s)
Fatty Acids , Mesorhizobium , Phylogeny , Root Nodules, Plant , Bacterial Typing Techniques , Base Composition , DNA, Bacterial/genetics , Fatty Acids/analysis , Fatty Acids/chemistry , Mesorhizobium/genetics , Mesorhizobium/isolation & purification , Mesorhizobium/classification , Nucleic Acid Hybridization , RNA, Ribosomal, 16S/genetics , Root Nodules, Plant/microbiology , Sequence Analysis, DNA , Tunisia , UbiquinoneABSTRACT
Root exudates from host plant species are known to play a critical role in the establishment and maintenance of symbiotic relationships with soil bacteria. In this study, we investigated the impact of root exudates from compatible host plant species; Elaeagnus angustifolia on the exoproteome of Parafrankia soli strain NRRL B-16219. A total of 565 proteins were evidenced as differentially abundant, with 32 upregulated and 533 downregulated in presence of the plant exudates. Analysis of the function of these proteins suggests that the bacterial strain is undergoing a complex metabolic reprogramming towards a new developmental phase elicited in presence of host plant root exudates. The upregulation of Type II/IV secretion system proteins among the differentially expressed proteins indicates their possible role in infecting the host plant, as shown for some rhizobia. Additionally, EF-Tu, proteins upregulated in this study, may function as an effector for the T4SSs and trigger plant defense responses. These findings suggest that Parafrankia soli may use EF-Tu to infect the actinorhizal host plant and pave the way for further investigations of the molecular mechanisms underlying the establishment of symbiotic relationships.
ABSTRACT
The taxonomic position of strain BMG 8361T, isolated from sandstone collected in the Sahara Desert of Southern Tunisia, was refined through a polyphasic taxonomic investigation. Colonies of BMG 8361T were pale-orange coloured, irregular with a dry surface and produced a diffusible pink or brown pigment depending on media. The Gram-positive cells were catalase-positive and oxidase-negative. The strain exhibited growth at 10-40â°C and pH values ranging from 5.5 to 9.0, with optima at 28-35â°C and pH 6.5-8.0. Additionally, BMG 8361T demonstrated the ability to grow in the presence of up to 1â% NaCl (w/v) concentration. The peptidoglycan of the cell wall contained meso-diaminopimelic acid, glucose, galactose, xylose, ribose, and rhamnose. The predominant menaquinones consisted of MK-9(H4) and MK-9. The main polar lipids were phosphatidylcholine, phosphatidylinositol, glycophosphatidylinositol, diphosphatidylglycerol, phosphatidylethanolamine, and two unidentified lipids. Major cellular fatty acids were iso-C16â:â0, iso-C16â:â1 h, and C17â:â1 ω8c. Phylogenetic analyses based on both the 16S rRNA gene and whole-genome sequences assigned strain BMG 8361T within the genus Blastococcus. The highest pairwise sequence similarity observed in the 16S rRNA gene was 99.5â% with Blastococcus haudaquaticus AT 7-14T. However, when considering digital DNA-DNA hybridization and average nucleotide identity, the highest values, 48.4 and 86.58â%, respectively, were obtained with Blastococcus colisei BMG 822T. These values significantly undershoot the recommended thresholds for establishing new species, corroborating the robust support for the distinctive taxonomic status of strain BMG 8361T within the genus Blastococcus. In conjunction with the phenotyping results, this compelling evidence leads to the proposal of a novel species we named Blastococcus brunescens sp. nov. with BMG 8361T (=DSM 46845T=CECT 8880T) as the type strain.
Subject(s)
Actinomycetales , Fatty Acids , Tunisia , Phylogeny , RNA, Ribosomal, 16S/genetics , Fatty Acids/chemistry , Sequence Analysis, DNA , DNA, Bacterial/genetics , Bacterial Typing Techniques , Base CompositionABSTRACT
In many frankia, the ability to nodulate host plants (Nod+) and fix nitrogen (Fix+) is a common strategy. However, some frankia within the Pseudofrankia genus lack one or two of these traits. This phenomenon has been consistently observed across various actinorhizal nodule isolates, displaying Nod- and/or Fix- phenotypes. Yet, the mechanisms supporting the colonization and persistence of these inefficient frankia within nodules, both with and without symbiotic strains (Nod+/Fix+), remain unclear. It is also uncertain whether these associations burden or benefit host plants. This study delves into the ecological interactions between Parafrankia EUN1f and Pseudofrankia inefficax EuI1c, isolated from Elaeagnus umbellata nodules. EUN1f (Nod+/Fix+) and EuI1c (Nod+/Fix-) display contrasting symbiotic traits. While the prediction suggests a competitive scenario, the absence of direct interaction evidence implies that the competitive advantage of EUN1f and EuI1c is likely contingent on contextual factors such as substrate availability and the specific nature of stressors in their respective habitats. In co-culture, EUN1f outperforms EuI1c, especially under specific conditions, driven by its nitrogenase activity. Iron-depleted conditions favor EUN1f, emphasizing iron's role in microbial competition. Both strains benefit from host root exudates in pure culture, but EUN1f dominates in co-culture, enhancing its competitive traits. Nodulation experiments show that host plant preferences align with inoculum strain abundance under nitrogen-depleted conditions, while consistently favoring EUN1f in nitrogen-supplied media. This study unveils competitive dynamics and niche exclusion between EUN1f and EuI1c, suggesting that host plant may penalize less effective strains and even all strains. These findings highlight the complex interplay between strain competition and host selective pressure, warranting further research into the underlying mechanisms shaping plant-microbe-microbe interactions in diverse ecosystems. IMPORTANCE: While Pseudofrankia strains typically lack the common traits of ability to nodulate the host plant (Nod-) and/or fix nitrogen (Fix-), they are still recovered from actinorhizal nodules. The enigmatic question of how and why these unconventional strains establish themselves within nodule tissue, thriving either alongside symbiotic strains (Nod+/Fix+) or independently, while considering potential metabolic costs to the host plant, remains a perplexing puzzle. This study endeavors to unravel the competitive dynamics between Pseudofrankia inefficax strain EuI1c (Nod+/Fix-) and Parafrankia strain EU1Nf (Nod+/Fix+) through a comprehensive exploration of genomic data and empirical modeling, conducted both in controlled laboratory settings and within the host plant environment.
Subject(s)
Elaeagnaceae , Frankia , Nitrogen Fixation , Root Nodules, Plant , Symbiosis , Frankia/genetics , Frankia/physiology , Frankia/metabolism , Elaeagnaceae/microbiology , Root Nodules, Plant/microbiology , Coculture Techniques , Genome, BacterialABSTRACT
A comprehensive polyphasic investigation was conducted to elucidate the taxonomic position of an actinobacterium, designated BMG 814T, which was isolated from the historic ruins of Carthage city in Tunisia. It grew as pink-orange pigmented colonies and displayed versatile growth capabilities, thriving within a temperature range of 20-40â°C, across a pH spectrum ranging from pH 5.5 to 10 and in the presence of up to 4â% NaCl. Chemotaxonomic investigations unveiled specific cell components, including diphosphatidylglycerol, phosphatidylcholine, phosphatidylethanolamine, phosphatidylinositol, glycophosphatidylinositol, an unidentified aminoglycophospholipid, six unidentified aminolipids, two unidentified phospholipids and one unidentified lipid in its polar lipid profile. Furthermore, galactose, glucose and ribose were identified as the primary cell-wall sugars. Major menaquinones identified were MK-9(H4), MK-9(H2) and MK-9, while major fatty acids comprised iso-C15â:â0, iso-C16â:â0, C17â:â1 ω8c and C18â:â1 ω9c. Through phylogenetic analysis based on the 16S rRNA gene sequence, the strain was positioned within the genus Blastococcus, with Blastococcus capsiensis BMG 804T showing the closest relationship (99.1â%). In light of this, draft genomes for both strains, BMG 814T and BMG 804T, were sequenced in this study, and comparative analysis revealed that strain BMG 814T exhibited digital DNA-DNA hybridization and average nucleotide identity values below the recommended thresholds for demarcating new species with all available genomes of type strains of validly names species. Based on the polyphasic taxonomy assessment, strain BMG 814T (=DSM 46848T=CECT 8878T) was proposed as the type strain of a novel species named Blastococcus carthaginiensis sp. nov.
Subject(s)
Actinomycetales , Fatty Acids , Tunisia , Phylogeny , RNA, Ribosomal, 16S/genetics , Fatty Acids/chemistry , Sequence Analysis, DNA , DNA, Bacterial/genetics , Bacterial Typing Techniques , Base CompositionABSTRACT
A nitrogen-fixing actinobacterium strain (Cc1.17T) isolated from a root nodule of Colletia cruciata was subjected to polyphasic taxonomic studies. The strain was characterized by the presence of meso-diaminopimelic acid in its peptidoglycan, galactose, glucose, mannose, rhamnose, ribose and xylose as cell-wall sugars, phosphatidylinositol, diphosphatidylglycerol, glycophospholipids, phosphatidylglycerol, glycophospholipid and uncharacterized lipids as its polar lipids, and C16â:â0, iso-C16â:â0, C17â: 1 ω9 and C18â:â1 ω9 as major fatty acids (>10â%). Strain Cc1.17T showed 16S rRNA gene sequence similarities of 97.4-99.8â% to validly named Frankia species. Phylogenetic trees based on 16S rRNA gene and genome sequences placed strain Cc1.17T in a new lineage within the genus Frankia. Digital DNA-DNA hybridization and average nucleotide identity values between strain Cc1.17T and its closest phylogenomic neighbours were well below the thresholds recommended for prokaryotic species delineation. Therefore, strain Cc1.17T (=DSM 43829T=CECT 9313T) merits recognition as the type strain of a new species for which the name Frankia colletiae sp. nov. is proposed.
Subject(s)
Frankia , Rubiaceae , Fatty Acids/chemistry , Phospholipids , Phylogeny , RNA, Ribosomal, 16S/genetics , Sequence Analysis, DNA , DNA, Bacterial/genetics , Bacterial Typing Techniques , Base Composition , Rubiaceae/geneticsABSTRACT
The status of four Frankia strains isolated from a root nodule of Alnus glutinosa was established in a polyphasic study. Taxogenomics and phenotypic features show that the isolates belong to the genus Frankia. All four strains form extensively branched substrate mycelia, multilocular sporangia, vesicles, lack aerial hyphae, but contain meso-diaminopimelic acid as the diamino acid of the peptidoglycan, galactose, glucose, mannose, ribose, xylose and traces of rhamnose as cell wall sugars, iso-C16:0 as the predominant fatty acid, diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol as the major polar lipids, have comparable genome sizes to other cluster 1, Alnus-infective strains with structural and accessory genes associated with nitrogen fixation. The genome sizes of the isolates range from 7.0 to 7.7 Mbp and the digital DNA G + C contents from 71.3 to 71.5 %. The four sequenced genomes are rich in biosynthetic gene clusters predicted to express for novel specialized metabolites, notably antibiotics. 16S rRNA gene and whole genome sequence analyses show that the isolates fall into two lineages that are closely related to the type strains of Frankia alni and Frankia torreyi. All of these taxa are separated by combinations of phenotypic properties and by digital DNA:DNA hybridization scores which indicate that they belong to different genomic species. Based on these results, it is proposed that isolates Agncl-4T and Agncl-10, and Agncl-8T and Agncl-18, be recognised as Frankia gtarii sp. nov. and Frankia tisai sp. nov. respectively, with isolates Agncl-4T (=DSM 107976T = CECT 9711T) and Agncl-8T (=DSM 107980T = CECT 9715T) as the respective type strains.
Subject(s)
Alnus , Frankia , Phospholipids/chemistry , RNA, Ribosomal, 16S/genetics , Sequence Analysis, DNA , DNA, Bacterial/genetics , Phylogeny , Fatty Acids/chemistry , Bacterial Typing TechniquesABSTRACT
The actinorhizal plant, Coriaria myrtifolia, is a neurotoxic plant species endemic to the western Mediterranean area, which forms a nitrogen-fixing symbiosis with members of Frankia cluster 2. Contrarily to other Frankia clusters, the occurrence and mode of dispersal for infective cluster 2 units outside of the host plant rhizosphere remains controversial. The present study was designed to investigate the structure of the microbiomes of C. myrtifolia phytosphere, rhizosphere, and soil samples extending outward linearly up to 1 km. Results showed that the epiphyte and endophyte communities were not significantly different from each other for most of the plant tissues. The communities associated with the below-ground tissues (nodule and root) were significantly different from those found on the above-ground tissues (fruit, leaves, and stems) and had a higher community richness. Coriaria myrtifolia phytomicrobiomes were dominated by Cyanobacteria for leaf, stem, and fruit while Actinobacteria and Proteobacteria were dominant in the root and nodule organelles. The nodule, a special niche for nitrogen fixation, was mainly inhabited by Frankia but contained several non-Frankia bacteria. Beside Frankia cluster 2, the presence of clusters 1, 4, and large numbers of cluster 3 strains have been detected in nodules, roots, and rhizospheres of C. myrtifolia. Despite Frankia being found in all plots using plant trapping bioassays with C. myrtifolia seedlings, Frankia cluster 2 was not detected in soil metagenomes showing the limits of detection by this approach. This result also suggests that in the absence of appropriate host plant species, Frankia cluster 2 has a reduced number of infective units present in the soil outward from the rhizosphere.
ABSTRACT
Coriaria myrtifolia occurs as natural flora of warm temperate climates of northern Algeria which commonly found in hedges, forest and ravine edges. This actinorhizal species was known to establish a mutualistic symbiosis with members of phylogenetic cluster 2 (including strains associated to Coriaria spp., Ceanothus, Datiscaceae, and Dryadoideae) within the genus Frankia. Attempts to isolate C. myrtifolia microsymbionts from native plants growing in 4 locations in Algeria permitted to only recover asymbiotic Frankia strains (unable to reestablish nodulation and to fix nitrogen) from phylogenetic cluster 4 and several non-Frankia actinobacteria including members of Micrococcus, Micromonospora, Nocardia, Plantactinospora, and Streptomyces genera. The biodiversity of Frankia microsymbionts of C. myrtifolia root nodules was assessed using PCR-amplification followed by partial nucleotide sequencing of glnA1 (glutamine synthetase type 1) gene. On the 12 different glnA1 gene sequences obtained in this study, 9 were detected for the first time, and were mainly closelyrelated to Mediterranean genotypes previously described in the Grand Maghreb countries (Morocco and Tunisia) and in Europe (France) but without clear separations from other cluster 2 genotypes.
ABSTRACT
A new actinobacterium strain, designated BMG 823T, was isolated from a limestone sample collected in Tunisia. Its taxonomic position was scrutinized using a polyphasic approach. Colonies of strain BMG 823T were pink orange-coloured, regular and had a moist surface. Cells are Gram-stain-positive, catalase-negative and oxidase-negative. The strain grew at pH 5.5-9, 10-40 °C and in presence of up to 4â% NaCl (w/v). Chemotaxonomically, strain BMG 823T was characterized by cell-wall type III containing meso-diaminopimelic acid as diamino acid, glucose, ribose and rhamnose as whole-cell sugars, MK-9(H4) as predominant menaquinone, and phosphatidylcholine, diphosphadidylglycerol, phosphatidethanolamine, phosphatidylcholine, phosphatidylinositol, unidentified glycolipid, unidentified aminophospholipids and unidentified glycophospholipid as major polar lipids. The fatty acid profile consisted of iso-C16â:â0 and iso-C17â:â1 ω9. Phylogenetic trees based on 16S rRNA gene and genome sequences placed strain BMG 823T within the genus Blastococcus and separated it from all type strains of validly published species. Comparison of 16S rRNA gene sequence similarity, digital DNA-DNA hybridization and average nucleotide identity indicated that strain BMG 823T was most closely related to Blastococcus litoris DSM 106127T and Blastococcus colisei BMG 822T with pairwise values well below the species differentiation thresholds. The distinct phenotypic and genotypic features of strain BMG 823T (=DSM 46838T=CECT 8881T) within the genus Blastococcus warrant its recognition as the type strain for the new species for which we propose the name Blastococcus tunisiensis sp. nov.
Subject(s)
Actinomycetales , Calcium Carbonate , Bacterial Typing Techniques , Base Composition , DNA, Bacterial/genetics , Fatty Acids/chemistry , Phosphatidylcholines , Phospholipids/chemistry , Phylogeny , RNA, Ribosomal, 16S/genetics , Sequence Analysis, DNA , TunisiaABSTRACT
Frankia and actinorhizal plants exchange signals in the rhizosphere leading to specific mutual recognition of partners and nitrogen-fixing nodule organogenesis. Frankia soli strain NRRL B-16219, from the Elaeagnus specificity group, colonizes the root tissues of its actinorhizal host through direct intercellular penetration of root epidermis cells and cortex. Here, we studied the early proteogenomic response of strain NRRL B-16219 to treatment with root exudates from compatible Elaeagnus angustifolia, and incompatible Ceanothus thyrsiflorus and Coriaria myrtifolia, host plants grown in nitrogen depleted hydroponic medium. Next-generation proteomics was used to identify the main Frankia proteins differentially expressed in response to the root exudates. No products of the nod genes present in B-16219 were detected. Proteins specifically upregulated in presence of E. angustifolia root exudates include those connected to nitrogen fixation and assimilation (glutamate synthetase, hydrogenase and squalene synthesis), respiration (oxidative phosphorylation and citric acid cycle pathways), oxidative stress (catalase, superoxide dismutase, and peroxidase), proteolysis (proteasome, protease, and peptidase) and plant cell wall degrading proteins involved in the depolymerization of celluloses (endoglucanase, glycosyltransferase, beta-mannanases, glycoside hydrolase and glycosyl hydrolase). Proteomic data obtained in this study will help link signaling molecules/factors to their biosynthetic pathways once those factors have been fully characterized.
Subject(s)
Elaeagnaceae/microbiology , Frankia , Plant Exudates , Plant Roots/microbiology , Proteome , Frankia/genetics , Proteome/metabolism , Proteomics , SymbiosisABSTRACT
Actinorhizal plants host mutualistic symbionts of the nitrogen-fixing actinobacterial genus Frankia within nodule structures formed on their roots. Several plant-growth-promoting bacteria have also been isolated from actinorhizal root nodules, but little is known about them. We were interested investigating the in planta microbial community composition of actinorhizal root nodules using culture-independent techniques. To address this knowledge gap, 16S rRNA gene amplicon and shotgun metagenomic sequencing was performed on DNA from the nodules of Casuarina glauca. DNA was extracted from C. glauca nodules collected in three different sampling sites in Tunisia, along a gradient of aridity ranging from humid to arid. Sequencing libraries were prepared using Illumina NextEra technology and the Illumina HiSeq 2500 platform. Genome bins extracted from the metagenome were taxonomically and functionally profiled. Community structure based off preliminary 16S rRNA gene amplicon data was analyzed via the QIIME pipeline. Reconstructed genomes were comprised of members of Frankia, Micromonospora, Bacillus, Paenibacillus, Phyllobacterium, and Afipia. Frankia dominated the nodule community at the humid sampling site, while the absolute and relative prevalence of Frankia decreased at the semi-arid and arid sampling locations. Actinorhizal plants harbor similar non-Frankia plant-growth-promoting-bacteria as legumes and other plants. The data suggests that the prevalence of Frankia in the nodule community is influenced by environmental factors, with being less abundant under more arid environments.
ABSTRACT
Frankia sp. strain BMG5.11, which was isolated from Elaeagnus angustifolia nodules, is able to infect other actinorhizal plants, including Elaeagnaceae, Rhamnaceae, Colletieae, Gymnostoma, and Myricaceae Here, we report the 11.3-Mbp draft genome sequence of Frankia sp. strain BMG5.11, with a G+C content of 69.9% and 9,926 candidate protein-encoding genes.
ABSTRACT
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 C17â:â1ω8c, iso-C16â:â0, C15:0, C16â:â0 and C17â:â0. 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 (99.78â%) while their digital DNA-DNA hybridization value was 41.1â%. 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.
Subject(s)
Ceanothus/microbiology , Frankia/classification , Phylogeny , Soil Microbiology , Bacterial Typing Techniques , Base Composition , DNA, Bacterial/genetics , Diaminopimelic Acid/chemistry , Fatty Acids/chemistry , Frankia/isolation & purification , Multilocus Sequence Typing , Nucleic Acid Hybridization , Phospholipids/chemistry , RNA, Ribosomal, 16S/genetics , Sequence Analysis, DNA , United States , Vitamin K 2/analogs & derivatives , Vitamin K 2/chemistryABSTRACT
Stone surfaces are extreme environments that support microbial life. This microbial growth occurs despite unfavourable conditions associated with stone including limited sources of nutrients and water, high pH and exposure to extreme variations in temperature, humidity and irradiation. These stone-dwelling microbes are often resistant to extreme environments including exposure to desiccation, heavy metals, UV and Gamma irradiation. Here, we report on the effects of climate and stone geochemistry on microbiomes of Roman stone ruins in North Africa. Stone microbiomes were dominated by Actinobacteria, Cyanobacteria and Proteobacteria but were heavily impacted by climate variables that influenced water availability. Stone geochemistry also influenced community diversity, particularly through biologically available P, Mn and Zn. Functions associated with photosynthesis and UV protection were enriched in the metagenomes, indicating the significance of these functions for community survival on stones. Core members of the stone microbial communities were also identified and included Geodermatophilaceae, Rubrobacter, Sphingomonas and others. Our research has helped to expand the understanding of stone microbial community structure and functional capacity within the context of varying climates, geochemical properties and stone conditions.
Subject(s)
Extreme Environments , Microbiota , Africa, Northern , Bacteria/genetics , Bacteria/isolation & purification , Metagenome , Microbiota/genetics , Photosynthesis , Ultraviolet RaysABSTRACT
This study was designed to determine the plant growth promoting (PGP) potential of members of the genus Frankia. To this end, the genomes of 21 representative strains were examined for genes associated directly or indirectly with plant growth. All of the Frankia genomes contained genes that encoded for products associated with the biosynthesis of auxins [indole-3-glycerol phosphate synthases, anthranilate phosphoribosyltransferases (trpD), anthranilate synthases, and aminases (trpA and B)], cytokinins (11 well-conserved genes within the predicted biosynthetic gene cluster), siderophores, and nitrogenases (nif operon except for atypical Frankia) as well as genes that modulate the effects of biotic and abiotic environmental stress (e.g., alkyl hydroperoxide reductases, aquaporin Z, heat shock proteins). In contrast, other genes were associated with strains assigned to one or more of four host-specific clusters. The genes encoding for phosphate solubilization (e.g., low-affinity inorganic phosphate transporters) and lytic enzymes (e.g., cellulases) were found in Frankia cluster 1 genomes, while other genes were found only in cluster 3 genomes (e.g., alkaline phosphatases, extracellular endoglucanases, pectate lyases) or cluster 4 and subcluster 1c genomes (e.g., NAD(P) transhydrogenase genes). Genes encoding for chitinases were found only in the genomes of the type strains of Frankia casuarinae, F. inefficax, F. irregularis, and F. saprophytica. In short, these in silico genome analyses provide an insight into the PGP abilities of Frankia strains of known taxonomic provenance. This is the first study designed to establish the underlying genetic basis of cytokinin production in Frankia strains. Also, the discovery of additional genes in the biosynthetic gene cluster involved in cytokinin production opens up the prospect that Frankia may have novel molecular mechanisms for cytokinin biosynthesis.
ABSTRACT
Strain CpI1T was, in 1978, the first isolate of the genus Frankia to be obtained from Comptonia peregrina root nodules. In this study, a polyphasic approach was performed to identify the taxonomic position of strain CpI1T among the members of the genus Frankia. The strain contains meso-diaminopimelic acid as the diagnostic diamino acid and galactose, glucose, mannose, rhamnose, ribose and xylose as cell wall sugars. The polar lipids were found to consist of phosphatidylinositol, diphosphatidylglycerol, glycophospholipids, phosphatidylglycerol, an aminophospholipid and unidentified phospholipids and lipids. The predominant menaquinone was identified as MK-9 (H8), while the major fatty acid are iso-C16:0 and C17:1ω 8c. The 16S rRNA gene sequence identity varies from 97.4 to 99.6% with the type strains of currently described Frankia species. Phylogenetic analyses based on 16S rRNA gene sequences and multi-locus sequence analysis (MLSA) using atp1, ftsZ, dnaK, gyrA and secA gene sequences showed that strain CpI1T is closely related to Frankia alni ACN14aT. The genome size of strain CpI1T is 7.6 Mb with a digital DNA G+C content of 72.4%. Digital DNA:DNA hybridization (values between strain CpI1T and its close phylogenetic relative F. alni ACN14aT was 44.1%, well below the threshold of 70% for distinguishing between bacterial genomic species. Based on the phenotypic, phylogenetic and genomic data, strain CpI1T (= DSM44263T = CECT9035T) warrants classification as the type strain of a novel species, for which the name Frankia torreyi sp. nov. is proposed.
Subject(s)
Frankia/isolation & purification , Root Nodules, Plant/microbiology , Axenic Culture , Base Composition , DNA, Bacterial/genetics , Fatty Acids/chemistry , Fatty Acids/metabolism , Frankia/classification , Frankia/genetics , Frankia/metabolism , Myricaceae/microbiology , Phylogeny , RNA, Ribosomal, 16S/geneticsABSTRACT
Actinorhizal plants form a symbiotic association with the nitrogen-fixing actinobacteria Frankia. These plants have important economic and ecological benefits including land reclamation, soil stabilization, and reforestation. Recently, many non-Frankia actinobacteria have been isolated from actinorhizal root nodules suggesting that they might contribute to nodulation. Two Nocardia strains, BMG51109 and BMG111209, were isolated from Casuarina glauca nodules, and they induced root nodule-like structures in original host plant promoting seedling growth. The formed root nodule-like structures lacked a nodular root at the apex, were not capable of reducing nitrogen and had their cortical cells occupied with rod-shaped Nocardiae cells. Both Nocardia strains induced root hair deformation on the host plant. BMG111209 strain induced the expression of the ProCgNin:Gus gene, a plant gene involved in the early steps of the infection process and nodulation development. Nocardia strain BMG51109 produced three types of auxins (Indole-3-acetic acid [IAA], Indole-3-Byturic Acid [IBA] and Phenyl Acetic Acid [PAA]), while Nocardia BMG111209 only produced IAA. Analysis of the Nocardia genomes identified several important predicted biosynthetic gene clusters for plant phytohormones, secondary metabolites, and novel natural products. Co-infection studies showed that Nocardia strain BMG51109 plays a role as a "helper bacteria" promoting an earlier onset of nodulation. This study raises many questions on the ecological significance and functionality of Nocardia bacteria in actinorhizal symbioses.
Subject(s)
Fagales/growth & development , Nocardia/physiology , Root Nodules, Plant/microbiology , Fagales/microbiology , Indoleacetic Acids/metabolism , Nocardia/genetics , Nocardia/isolation & purification , Plant Growth Regulators/metabolism , Root Nodules, Plant/growth & development , SymbiosisABSTRACT
It has been 40 years since the first meeting dedicated to Frankia and actinorhizal plants, which was held at Petersham, Massachusetts (reported in Torrey and Tjepkema, 1979). Since then biennial meetings have been organised and held in different venues around the globe (Table 1). The most recent meeting, the "19th International Meeting on Frankia and Actinorhizal Plants", organised in Hammamet, Tunisia from 17th to 19th of March, 2018, gathered scientists from Algeria, Argentina, Belgium, China, Egypt, France, India, Portugal, Senegal, Sweden, UK, USA and Tunisia. The event was a stimulating opportunity for active researchers to share many advances since the previous meeting held in Montpellier, France (Franche et al. 2016) and to discuss new perspectives in this research field.