Comparative genomics of plant growth promoting phosphobacteria isolated from acidic soils.

Despite being one of the most abundant elements in soil, phosphorus (P) often becomes a limiting macronutrient for plants due to its low bioavailability, primarily locked away in insoluble organic and inorganic forms. Phosphate solubilizing and mineralizing bacteria, also called phosphobacteria, isolated from P-deficient soils have emerged as a promising biofertilizer alternative, capable of converting these recalcitrant P forms into plant-available phosphates. Three such phosphobacteria strains-Serratia sp. RJAL6, Klebsiella sp. RCJ4, and Enterobacter sp. 198-previously demonstrated their particular strength as plant growth promoters for wheat, ryegrass, or avocado under abiotic stresses and P deficiency. Comparative genomic analysis of their draft genomes revealed several genes encoding key functionalities, including alkaline phosphatases, isonitrile secondary metabolites, enterobactin biosynthesis and genes associated to the production of indole-3-acetic acid (IAA) and gluconic acid. Moreover, overall genome relatedness indexes (OGRIs) revealed substantial divergence between Serratia sp. RJAL6 and its closest phylogenetic neighbours, Serratia nematodiphila and Serratia bockelmanii. This compelling evidence suggests that RJAL6 merits classification as a novel species. This in silico genomic analysis provides vital insights into the plant growth-promoting capabilities and provenance of these promising PSRB strains. Notably, it paves the way for further characterization and potential application of the newly identified Serratia species as a powerful bioinoculant in future agricultural settings.

Downsizing Class II Lasso Peptides: Genome Mining-Guided Isolation of Huascopeptin Containing the First Gly1-Asp7 Macrocycle.

A new lasso peptide, huascopeptin, was isolated following genome-mined discovery of a new biosynthetic gene cluster in extremotolerant Streptomyces huasconensis HST28T from Salar de Huasco, Atacama Desert, Chile. Compound 1 is a 13-residue class II lasso peptide containing a novel Gly1-Asp7 macrolactam ring, a three-residue loop, and a three-residue tail, making it the smallest lasso peptide isolated to date. The lasso structure was confirmed using NOE restraint-based molecular dynamics simulations.

Pseudomonas khazarica sp. nov., a polycyclic aromatic hydrocarbon-degrading bacterium isolated from Khazar Sea sediments.

A novel Gram-negative, aerobic, motile and rod-shaped bacterium with the potential to biodegrade polycyclic aromatic hydrocarbons, was isolated from Khazar (Caspian) Sea. Strain TBZ2T grows in the absence of NaCl and tolerates up to 8.5% NaCl. Growth occurred at pH 3.0-10.0 (optimum, pH 6.0-7.0) and 10-45 °C (optimum, 30 °C). The major fatty acids are C18:1ω7C, C16:1ω7C/ C15:0 iso 2-OH, C16:0, C12:0, C10:0 3-OH, C12:0 3-OH. The major polar lipids include diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, phosphatidylcholine and the predominant respiratory quinone is ubiquinone Q-9. The 16S rRNA gene sequence analysis showed that strain TBZ2T is a member of the genus Pseudomonas with the highest similarity to P. oleovorans subsp. oleovorans DSM 1045T (98.83%), P. mendocina NBRC 14162T (98.63%), P. oleovorans subsp. lubricantis RS1T (98.61%) and P. alcaliphila JCM 10630T (98.49%) based on EzBioCloud server. Phylogenetic analyses using housekeeping genes (16S rRNA, rpoD, gyrB and rpoB) and genome sequences demonstrated that the strain TBZ2T formed a distinct branch closely related to the type strains of P. mendocina and P. guguanensis. Digital DNA-DNA hybridisation and average nucleotide identity values between strain TBZ2T and its closest relatives, P. mendocina NBRC 14162T (25.3%, 81.5%) and P. guguanensis JCM 18146T (26.8%, 79.0%), rate well below the designed threshold for assigning prokaryotic strains to the same species. On the basis of phenotypic, chemotaxonomic, genomic and phylogenetic results, it is recommended that strain TBZ2T is a novel species of the genus Pseudomonas, for which the name Pseudomonas khazarica sp. nov., is proposed. The type strain is TBZ2T (= LMG 29674T = KCTC 52410T).

Streptomyces huasconensis sp. nov., an haloalkalitolerant actinobacterium isolated from a high altitude saline wetland at the Chilean Altiplano.

Streptomyces strain HST28T isolated from the Salar de Huasco, an athalassohaline and poly-extreme high altitude saline wetland located in northern Chile, was the subject of a polyphasic taxonomic study. Strain HST28T showed morphological and chemotaxonomic features in line with its classification in the genus Streptomyces. Optimal growth of strain HST28T was obtained at 28 °C, pH 8-9 and up to 10 % (w/v) NaCl. Single (16S rRNA) and multi-locus gene sequence analyses showed that strain HST28T had a distinct phylogenetic position from its closest relatives, the type strains of Steptomyces aureus and Streptomyces kanamyceticus. Digital DNA-DNA hybridization (23.3 and 31.0 %) and average nucleotide identity (79.3 and 85.6 %) values between strain HST28T and its corresponding relatives mentioned above were below the threshold of 70 and 96 %, respectively, defined for assigning a prokaryotic strains to the same species. Strain HST28T was characterised by the presence of ll-diaminopimelic acid in its peptidoglycan layer; galactose, glucose, ribose and traces of arabinose and mannose as whole-cell sugars; phosphatidylmethylethanolamine, phosphatidylinositol, aminolipid, glycophospholipid and an unidentified lipid as polar lipids; and the predominating menaquinones MK-9(H6), MK-9(H8) and MK-9(H4) (>20 %) as well as anteiso-C15 : 0 and anteiso-C17 : 0 as major fatty acids (>15 %). Based on the phenotypic and genetic results, strain HST28T (DSM 107268T=CECT 9648T) merits recognition as a new species named Streptomyces huasconensis sp. nov.

Mycolicibacterium stellerae sp. nov., a rapidly growing scotochromogenic strain isolated from Stellera chamaejasme.

A polyphasic study was undertaken to establish the taxonomic provenance of a rapidly growing Mycolicibacterium strain, CECT 8783T, recovered from the plant Stellera chamaejasme L. in Yunnan Province, China. Phylogenetic analyses based upon 16S rRNA and whole-genome sequences showed that the strain formed a distinct branch within the evolutionary radiation of the genus Mycolicibacterium. The strain was most closely related to Mycolicibacterium moriokaense DSM 44221T with 98.4 % 16S rRNA gene sequence similarity, but was distinguished readily from this taxon by a combination of chemotaxonomic and phenotypic features and by low average nucleotide identity and digital DNA-DNA hybridization values of 79.5 and 21.1 %, respectively. Consequently, the strain is considered, to represent a novel species of Mycolicibacterium for which the name Mycolicibacterium stellerae sp. nov is proposed; the type strain is I10A-01893T (=CECT 8783T=KCTC 19843T=DSM 45590T).

Streptomyces altiplanensis sp. nov., an alkalitolerant species isolated from Chilean Altiplano soil, and emended description of Streptomyces chryseus (Krasil'nikov et al. 1965) Pridham 1970.

A polyphasic approach was used for evaluating the taxonomic status of strain HST21T isolated from Salar de Huasco in the Atacama Desert. The results of 16S rRNA gene and multilocus sequence phylogenetic analyses assigned strain HST21T to the genus Streptomyceswith Streptomyces albidochromogenes DSM 41800Tand Streptomyces flavidovirens DSM 40150T as its nearest neighbours. Digital DNA-DNA hydridization (dDDH) and average nucleotide identity (ANI) values between the genome sequences of strain HST21T and S. albidochromogenes DSM 41800T (35.6 and 88.2 %) and S. flavidovirens DSM 40105T (47.2 and 88.8 %) were below the thresholds of 70  and 95-96 % for prokaryotic conspecific assignation. Phenotypic, chemotaxonomic and genetic results distinguished strain HST21T from its closest neighbours. Strain HST21T is characterized by the presence of ll-diaminopimelic acid in its peptidoglycan layer; glucose and ribose as whole cell sugars; diphosphatidylglycerol, phosphatidylmethylethanolamine, phosphatidylethanolamine, phosphatidylinositol, glycophospholipids, unknown lipids and phospholipids as polar lipids; and anteiso-C15 : 0 (21.6 %) and anteiso-C17 : 0 (20.5 %) as major fatty acids (>15 %). Based on these results, strain HST21T merits recognition as a novel species, for which the name Streptomyces altiplanensis sp. nov. is proposed. The type strain is HST21T=DSM 107267T=CECT 9647T. While analysing the phylogenies of strain HST21T, Streptomyces chryseus DSM 40420T and Streptomyces helvaticus DSM 40431T were found to have 100 % 16S rRNA gene sequence similarity with digital DNA-DNA hydridization (dDDH) and average nucleotide identity (ANI) values of 95.3 and 99.4 %, respectively. Therefore, S. helvaticus is considered as a later heterotypic synonym of S. chryseus and, consequently, we emend the description of S. chryseus.

Comparative Genomic Study of Vinyl Chloride Cluster and Description of Novel Species, Mycolicibacterium vinylchloridicum sp. nov.

Advanced physicochemical and chemical absorption methods for chlorinated ethenes are feasible but incur high costs and leave traces of pollutants on the site. Biodegradation of such pollutants by anaerobic or aerobic bacteria is emerging as a potential alternative. Several mycobacteria including Mycolicibacterium aurum L1, Mycolicibacterium chubuense NBB4, Mycolicibacterium rhodesiae JS60, Mycolicibacterium rhodesiae NBB3 and Mycolicibacterium smegmatis JS623 have previously been described as assimilators of vinyl chloride (VC). In this study, we compared nucleotide sequence of VC cluster and performed a taxogenomic evaluation of these mycobacterial species. The results showed that the complete VC cluster was acquired by horizontal gene transfer and not intrinsic to the genus Mycobacterium sensu lato. These results also revealed the presence of an additional xcbF1 gene that seems to be involved in Coenzyme M biosynthesis, which is ultimately used in the VC degradation pathway. Furthermore, we suggest for the first time that S/N-Oxide reductase encoding gene was involved in the dissociation of the SsuABC transporters from the organosulfur, which play a crucial role in the Coenzyme M biosynthesis. Based on genomic data, M. aurum L1, M. chubuense NBB4, M. rhodesiae JS60, M. rhodesiae NBB3 and M. smegmatis JS623 were misclassified and form a novel species within the genus Mycobacterium sensu lato. Mycolicibacterium aurum L1T (CECT 8761T = DSM 6695T) was the subject of polyphasic taxonomic studies and showed ANI and dDDH values of 84.7 and 28.5% with its close phylogenetic neighbour, M. sphagni ATCC 33027T. Phenotypic, chemotaxonomic and genomic data considering strain L1T (CECT 8761T = DSM 6695T) as a type strain of novel species with the proposed name, Mycolicibacterium vinylchloridicum sp. nov.

Genomic Virulence Features of Two Novel Species Nocardia barduliensis sp. nov. and Nocardia gipuzkoensis sp. nov., Isolated from Patients with Chronic Pulmonary Diseases.

Strains 335427T and 234509T, isolated from two 76-year-old patients with chronic pulmonary diseases, were the subject of polyphasic taxonomic studies and comparative genomic analyses for virulence factors. The 16 rRNA gene sequence similarity between strains 335427T and 234509T and their closest phylogenetic neighbors Nocardia asiatica NBRC 100129T and Nocardia abscessus NBRC 100374T were 99.5% and 100%, respectively. Digital DNA-DNA hybridization values between the aforementioned studied strains were well below the 70% threshold for assigning prokaryotic strains to a novel species. Strains 335427T and 234509T have genome sizes of 8.49 Mpb and 8.07 Mpb, respectively, with G + C content of 68.5%. Isolate 335427T has C16:0, C18:1 ω9c, C18:0 and C18:0 10 methyl as major fatty acids (>15%) and mycolic acids formed of 52-54 carbon atoms. However, only C18:1 ω9c was detected for isolate 234509T, which had mycolic acids with 44-56 carbon. Based on phenotypic and genetic data, strains 335427T (DSM 109819T = CECT 9924T) and 234509T (DSM 111366T = CECT 30129T) merit recognition as novel species, which are named Nocardia barduliensis sp. nov. and Nocardia gipuzkoensis sp. nov., respectively. All the strains studied had homologous VF-associated genes to those described in M. tuberculosis, including experimentally verified virulence genes in humans related to tuberculosis. The narGHIJ (nitrate reduction pathway) and gvpAFGOJLMK (gas vesicles) genetic maps of strains 335427T, 234509T, NBRC 100129T and NBRC 100374T showed the same syntenic block and raise the question of whether their functions are interlinked during the infection of the human host. However, further research is required to decipher the role of the gas vesicle in the pathogenicity mechanism of Nocardia spp.

Cutting the Gordian knot: early and complete amino acid sequence confirmation of class II lasso peptides by HCD fragmentation.

Lasso peptides are a diverse class of ribosomally synthesized and post-translationally modified peptides (RiPPs). Their proteolytic and thermal stability alongside their growing potential as therapeutics has increased attention to these antimicrobial peptides. With the advent of genome mining, the discovery of RiPPs allows for the accurate prediction of putatively encoded structures, however, MSn experiments only provide partial sequence confirmation, therefore 2D NMR experiments are necessary for characterisation. Multiple MS/MS techniques were applied to two structurally characterized lasso peptides, huascopeptin and leepeptin, and one uncharacterized lasso peptide, citrulassin C, which was not isolable in sufficient quantity for NMR analysis. We have shown that MS2 can be used to elucidate the full amino acid sequences previously predicted with genome mining for this compound class. HCD was able to open the macrocycles and fragment the newly opened linear peptides, confirming the complete amino acid sequences of the characterised lasso peptides. In addition, to determine if this technique could be applied at the earliest stages of the isolation process, we targeted a lasso peptide found by genome mining, citrulassin C, and were able to fully elucidate the amino acid sequence using only MS2 from a semi-crude extract of Streptomyces huasconensis HST28T.

Genomic Insights Into Plant-Growth-Promoting Potentialities of the Genus Frankia.

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.