Genus Rhodococcus Research Articles

Degradation of long-chain n-alkanes by a novel thermal-tolerant Rhodococcus strain.

A novel bacterial strain, CH91, was isolated from a high-temperature oil reservoir. Morphological characterization, phylogenetic analyses of 16S rRNA gene sequence and genome relatedness indicated that the strain is a potential new species in the genus Rhodococcus. Strain CH91 could grow in the temperature range of 25-50°C (optimally at 37°C) and utilize a broad range of long-chain n-alkanes from hexadecane to hexatriacontane. The utilization of the n-alkanes mixture of strain CH91 revealed that the degradation rate was correlated to the length of the carbon chain. Two novel alkB genes encoding alkane 1-monooxygenase were found in the genome of this strain. The protein sequences of both alkane 1-monooxygenases showed a remarkable phylogenetic distance to other reported AlkB protein sequences. These results would help broaden our knowledge about alkane degradation by Rhodocuccus and its potential ecological role. The ability of the strain in the long-chain alkane degradation and thermal tolerance could also be further exploited for bioremediation of oil contaminations and microbial enhanced oil recovery.

Transcriptomic Analysis of the Dual Response of Rhodococcus aetherivorans BCP1 to Inorganic Arsenic Oxyanions.

Bacterial strains belonging to the genus Rhodococcus are able to degrade various toxic organic compounds and tolerate high concentrations of metal(loid)s. We have previously shown that Rhodococcus aetherivorans BCP1 is resistant to various levels of the two arsenic inorganic species, arsenite [As(III)] and arsenate [As(V)]. However, while arsenite showed toxic effects at concentrations as low as 5 mM, arsenate at 30 mM boosted the growth rate of BCP1 cells and was toxic only at concentrations of >100 mM. Since such behavior could be linked to peculiar aspects of its metabolism, the transcriptomic analysis of BCP1 cells exposed to 5 mM As(III) and 30 mM As(V) was performed in this work. The aim was to clarify the mechanisms underlying the arsenic stress response of the two growth phenotypes in the presence of the two different oxyanions. The results revealed that As(III) induced higher activity of reactive oxygen species (ROS)-scavenging enzymes than As(V) in relation to the expression of enzymes involved in cellular damage recovery and redox buffers/cofactors (ergothioneine, mycofactocin, and mycothiol). Further, As(III) downregulated pathways related to cell division, while both oxyanions downregulated genes involved in glycolysis. Notably, As(V) induced the expression of enzymes participating in the synthesis of metallophores and rearranged the central and energetic metabolism, also inducing alternative pathways for ATP synthesis and glucose consumption. This study, in providing transcriptomic data on R. aetherivorans exposed to arsenic oxyanions, sheds some light on the plasticity of the rhodococcal response to arsenic stress, which may be important for the improvement of biotechnological applications. IMPORTANCE Members of the genus Rhodococcus show high metabolic versatility and the ability to tolerate/resist numerous stress conditions, including toxic metals. R. aetherivorans BCP1 is able to tolerate high concentrations of the two inorganic arsenic oxyanions, arsenite [As(III)] and arsenate [As(V)]. Despite the fact that BCP1 intracellularly converts As(V) into As(III), this strain responds very differently to the presence of these two oxyanions in terms of cell growth and toxic effects. Indeed, while As(III) is highly toxic, exposure to specific concentrations of As(V) seems to boost cell growth. In this work, we investigated the transcriptomic response, ATP synthesis, glucose consumption, and H2O2 degradation in BCP1 cells exposed to As(III) and As(V), inducing two different growth phenotypes. Our results give an overview of the transcriptional rearrangements associated with the dual response of BCP1 to the two oxyanions and provide novel insights into the energetic metabolism of Rhodococcus under arsenic stress.

Chemical and microbiological changes on the surface of microplastic after long term exposition to different concentrations of ammonium in the environment

The increasing production of plastic in the world has resulted in the widespread pollution of the environment with microplastics (MP). MP enter facilities such as wastewater treatment plants or landfills characterized by various ammonium concentrations. The aim of this study was to determine the structure of the microbial community on MP surfaces at various concentrations of ammonium nitrogen, and in particular, to identify microorganisms capable of polyethylene terephthalate (PET) degradation. Moreover, changes in the chemical characteristics of the MP surface resulting from microbial activity were also investigated, and the potential of MP to serve as a vector for pollutants was determined. The tests were carried out in a reactor filled with PET for a period of 260 days. The experiment was carried out in 3 phases: in I and III phase, the concentration of N-NH4 was about 70 mg/L, while in II phase, it was about 430 mg/L. On the MP surface, biofilm-forming microorganisms from the genera Rhodococcus, Pseudomonas and Xantomonas were identified at the lower ammonium concentration. At this concentration, MP-degraders belonging to genera Acidovorax, Gordonia, Pseudomonas, Sphingomonas, and Sphingopyxis were identified in the biofilm. At the higher N-NH4 concentration, the biomass was enriched with bacteria from genera Nitrosospira, Nitrosomonas and Terrimonas, and the number of microorganisms with the potential to degrade MP decreased. Analysis of the MP surface during the experiment has showed the loss of carbonyl groups and formation of carboxyl and hydroxyl groups, which indicated the degradation of MP. Independent of the ammonium concentration in the environment, MP was a carrier of pathogenic microorganisms from the genera Mycobacterium, Enterobacter and Brevundimonas.

Effects of Nickel Nanoparticles on Rhodococcus Cell Surface Morphology and Nanomechanical Properties

Nickel nanoparticles (NPs) are used for soil remediation and wastewater treatment due to their high adsorption capacity against complex organic pollutants. However, despite the growing use of nickel NPs, their toxicological towards environmental bacteria have not been sufficiently studied. Actinobacteria of the genus Rhodococcus are valuable bioremediation agents degrading a range of harmful and recalcitrant chemicals. Both positive and negative effects of metal ions and NPs on the biodegradation of organic pollutants by Rhodococcus were revealed, however, the mechanisms of such interactions, in addition to direct toxic effects, remain unclear. In the present work, the influence of nickel NPs on the viability, surface topology and nanomechanical properties of Rhodococcus cells have been studied. Bacterial adaptations to high (up to 1.0 g/L) concentrations of nickel NPs during prolonged (24 and 48 h) exposure were detected using combined confocal laser scanning and atomic force microscopy. Incubation with nickel NPs resulted in a 1.25–1.5-fold increase in the relative surface area and roughness, changes in cellular charge and adhesion characteristics, as well as a 2–8-fold decrease in the Young’s modulus of Rhodococcus ruber IEGM 231 cells. Presumably, the treatment of rhodococcal cells with sublethal concentrations (0.01–0.1 g/L) of nickel NPs facilitates the colonization of surfaces, which is important in the production of immobilized biocatalysts based on whole bacterial cells adsorbed on solid carriers. Based on the data obtained, cell surface functionalizing with NPs is possible to enhance adhesive and catalytic properties of bacteria suitable for environmental applications.

Identification of a Novel Biosurfactant with Antimicrobial Activity Produced by Rhodococcus opacus R7.

Rhodococcus members excrete secondary metabolites, especially compounds which act as biosurfactants. In this work, we demonstrated the ability of Rhodococcus opacus R7 to produce a novel bioactive compound belonging to the class of biosurfactants with antimicrobial properties during the growth on naphthalene. Chemical and biochemical analyses of the isolated compound demonstrated that the biosurfactant could be classified as a hydrophobic peptide. The ESI-full mass spectrometry revealed that the isolated biosurfactant showed a molecular weight of 1292 Da and NMR spectra evidenced the composition of the following amino acid residues: Ala, Thr, Asp, Gly, Ser. Surfactant activity of the R. opacus R7 compound was quantified by the critical micelle dilution (CMD) method and the critical micelle concentration (CMC) was estimated around 20 mg L−1 with a corresponding surface tension of 48 mN m−1. Moreover, biological assays demonstrated that R. opacus R7 biosurfactant peptide exhibited antimicrobial activity against Escherichia coli ATCC 29522 and Staphylococcus aureus ATCC 6538 with the minimum inhibition growth concentration (MIC) values of 2.6 mg mL−1 and 1.7 mg mL−1, respectively. In this study for the first time, a hydrophobic peptide with both biosurfactant and antimicrobial activity was isolated from a bacterium belonging to Rhodococcus genus.

Antrihabitans stalagmiti sp. nov., isolated from a larva cave and a proposal to transfer Rhodococcus cavernicola Lee et al. 2020 to a new genus Spelaeibacter as Spelaeibacter cavernicola gen. nov. comb. nov.

An actinobacterial strain, designated YC3-6T, was isolated from a larva cave in Jeju, Republic of Korea. The novel isolate was found to grow at 10-30°C, pH 5.0-10.0 and 0-4% (w/v) NaCl. The 16S rRNA gene phylogeny showed that the novel isolate formed a distinct subline within the family Nocardiaceae. Levels of 16S rRNA gene similarity indicated that the close relatives are Rhodococcus cavernicola (98.4% sequence similarity) and "Rhodococcus psychrotolerans" (98.2%) followed by Antrihabitans stalactiti (96.8%). However, the core gene-based phylogeny revealed that the novel isolate formed a tight cluster with A. stalactiti and was separated from R. cavernicola and other members of the family Nocardiaceae. The morphological and chemotaxonomic characteristics of strain YC3-6T are in line with those of the genus Antrihabitans. Strain YC3-6T showed an average nucleotide identity of 75.5% and a digital DDH of 20.3% with A. stalactiti. In addition, the core gene analysis showed that R. cavernicola formed a distinct subline between an Antrihabitans cluster and Aldersonia kunmingensis, and well separated from members of the genus Rhodococcus. The average amino acid identity values of R. cavernicola to closely related neighbours were 69.3-69.4% with members of the genus Antrihabitans and 67.3% with Ald. kunmingensis, while the POCP values ranged from 56.9 to 63.6%. On the basis of results obtained here, strain YC3-6T is concluded to represent a novel species of the genus Antrihabitans, for which the name Antrihabitans stalagmiti sp. nov. (type strain, YC3-6T = KACC 19963T = DSM 107561T) is proposed. Based on overall genome relatedness and chemotaxonomic differences, it is also proposed that R. cavernicola Lee et al. 2020 be transferred to a new genus Spelaeibacter as Spelaeibacter cavernicola gen. nov., comb. nov.

POS-722 PERITONEAL DIALYSIS-ASSOCIATED PERITONITIS CAUSED BY GORDONIA AICHIENSIS: A CASE REPORT AND LITERATURE REVIEW

Gordonia species are aerobic and slow-growing gram-positive pathogens that rarely cause human infections. However, previous published case reports indicate that Gordonia species can cause unusual human infections in patients who are immunocompromised or have indwelling catheters. Furthermore, they are uncommon bacilli in patients with peritoneal dialysis-associated peritonitis (PDAP). Gordonia aichiensis originally belonged to the genus Rhodococcus and was classified in 1994 as Gordona on the basis of wall characteristics.

Complete Genome Sequence and Benzophenone-3 Mineralisation Potential of Rhodococcus sp. USK10, A Bacterium Isolated from Riverbank Sediment

Benzophenone-3 (BP3) is an organic UV filter whose presence in the aquatic environment has been linked to detrimental developmental impacts in aquatic organisms such as coral and fish. The genus Rhodococcus has been extensively studied and is known for possessing large genomes housing genes for biodegradation of a wide range of compounds, including aromatic carbons. Here, we present the genome sequence of Rhodococcus sp. USK10, which was isolated from Chinese riverbank sediment and is capable of utilising BP3 as the sole carbon source, resulting in full BP3 mineralisation. The genome consisted of 9,870,030 bp in 3 replicons, a G+C content of 67.2%, and 9722 coding DNA sequences (CDSs). Annotation of the genome revealed that 179 of these CDSs are involved in the metabolism of aromatic carbons. The complete genome of Rhodococcus sp. USK10 is the first complete, annotated genome sequence of a Benzophenone-3-degrading bacterium. Through radiolabelling, it is also the first bacterium proven to mineralise Benzophenone-3. Due to the widespread environmental prevalence of Benzophenone-3, coupled with its adverse impact on aquatic organisms, this characterisation provides an integral first step in better understanding the environmentally relevant degradation pathway of the commonly used UV filter. Given USK10′s ability to completely mineralise Benzophenone-3, it could prove to be a suitable candidate for bioremediation application.

Features of the Bioconversion of Pentacyclic Triterpenoid Oleanolic Acid Using Rhodococcus Actinobacteria

The ability of actinobacteria of the genus Rhodococcus to transform oleanolic acid (OA), a plant pentacyclic triterpenoid, was shown for the first time using bioresources of the Regional Specialized Collection of AlkanotrophicMicroorganisms (IEGM; WDCM #768;www.iegmcol.ru). The most promising strains (R.opacus IEGM 488 and R.rhodochrousIEGM 285) were selected, and these catalyzed80% bioconversion of OA (0.5 g/L) in the presence of n-hexadecane (0.1% v/v) for seven days. The process of OA bioconversion was accompanied by a gradual decrease in the culture medium pH. Adaptive responses of bacterial cells to the OA effects included the formation of compact cellular aggregates, a marked change in the surface-to-volume ratio of cells, and a significant increase in the Zeta potential values. The results demonstrated that the process of OA bioconversion was catalyzed by membrane-bound enzyme complexes. Participation of cytochrome P450-dependent monooxygenases in the oxidation of the OA moleculewas confirmedusing specific inhibitors. The obtained data expand our knowledge on the catalytic activity of actinobacteria of the genus Rhodococcus and their possible use as biocatalysts for the bioconversion of complex hydrophobic compounds. The results can also be used inthe searchfor promising OA derivatives to be used in the synthesis of biologically active agents.
 Keywords: bioconversion, oleanolic acid, Rhodococcus, biologically active compounds

Влияние одностенных углеродных нанотрубок на биопленкообразование актинобактерий рода Rhodococcus и протеобактерий активного ила

The effect of single-walled carbon nanotubes (SWCNTs) on the formation and eradication of biofilms of activated sludge proteobacteria and actinobacteria of the genus Rhodococcus was studied. It has been shown that SWCNTs do not inhibit the formation of biofilms of these bacteria and do not cause their destruction, while in the presence of SWCNTs, much more massive biofilms of Alcaligenes faecalis 2 and Acinetobacter guillouiae 11h are formed. It was shown that the level of dehydrogenase activity, estimated by the reduction of tetrazolium salt to formazan, in biofilm cells formed in the presence of SWCNTs, exceeded that of biofilms in the control.

Characterization Of A Bacterium – Producer Of Amidase

We analyzed 11 samples of wastewater, sludge and soil contaminated with acrylic acid nitrile, acrylamide and acrylic acid from the territory of Navoiyazot JSC (Navoi, Uzbekistan). As a result, 19 strains belonging to the bacteria of the genus Rhodococcus were isolated into a pure culture. In the process of selection, 2 strains of bacteria were selected that exhibit the highest amidase activity for acrylic acid (0.712 U / mg and 0.868 U / mg). Strains of Rhodococcus sp. - 8/4/1 and Rhodococcus sp. - 3/4/3 are potential amidase producers.

Microbial community and metabolic characteristics evaluation in start-up stage of electro-enhanced SBR for aniline wastewater treatment

An electro-enhanced sequencing batch reactor (E-SBR) was constructed to treat 400 mg/L aniline wastewater and realized system rapid start-up. The aniline degradation efficiency was almost 100% and the total nitrogen removal rate was 80% in the E-SBR. High-throughput sequencing showed that the microbial community in E-SBR changed greatly. At genus level, Anaerolineae, Rhodococcus, Luteococcus, Deniratisoma, and SM1A02 enriched in activated sludge. Thauera, Ellin6067, Arenimonas, Hydrogenophaga enriched in the electrode biofilm. These functional microorganisms accounted for a large proportion and played an important role in the removal of pollutants. Meanwhile, the microorganism metabolism in the E-SBR system was significantly different from that in the traditional SBR system, the E-SBR system had higher nitrogen metabolic and carbohydrate metabolism abundance to ensure the effective removal of aniline and nitrogen.

Impact of pipe material and chlorination on the biofilm structure and microbial communities

Pipe material and residual chlorine are key factors for the drinking water distribution system, and understanding the biofilm ecosystem is vital for water quality safeguard. The aim of our study was to determine the influence of pipe materials (ductile iron, steel, polyethylene) and chlorination on the biofilm structure and microbial community, as shown by the physicochemical properties, extracellular polymeric substances (EPS) structural characteristics, bacterial community composition, and functional traits. EPS spatial properties were studied based on a semi-quantitative confocal laser scanning microscope (CLSM) description. Regarding the impact of chlorination, residule chlorine (1.0 ± 0.3 mg L−1 free chlorine) could inhibit the bacteria colonization, and initiate a potential response to external disinfectants revealed by the EPS spatial distribution changes and communities variation compared to unchlorinated system. Regarding the impact of pipe material, polyethylene (PE) biofilms displayed lower biomass, loose zoogloea structure, lower proteins and polysaccharides content, and poor microbial diversity in contrast to ductile iron and steel biofilms. Pipe material was the more possible driving factor of the biofilm community composition compared to the chlorination based on principal coordinates analysis (PCoA) and permutational multivariate analysis of variance (PERMANOVA). Actinobacteria was dominant in the PE biofilms (45.57%–83.32%), while Alphaproteobacteria (34.30%–73.22%) and Gammaproteobacteria (6.46%–36.82%) were the major classes in the steel and ductile iron biofilms. The genus Rhodococcus was predominant in the PE biofilms. Rhodococcus, Pseudomonas, and Sphingomonas seemed to have a better growth advantage in the chlorinated system and display a stronger disinfectant resistance. Functional sketch prediction indicated the potential impact of pipe material and chlorination on functional pathway abundnce, possible functional pathways associated with infectious disease included. This study provides insights into the impact of pipe material and chlorination on biofilm structure and microbial community and might help to develop monitoring or maintenance strategies to protect the biosafety of the drinking water.

Rausuquinone, a non-glycosylated pluramycin-class antibiotic from Rhodococcus.

A new pluramycin-class polyketide, rausuquinone (1), and its known congener hydramycin (2) were isolated from the culture extract of the deep-sea water-derived Rhodococcus sp. RD015140. Compound 1 possesses a γ-pyrone-fused anthraquinone core with a 3-butene-1,2-diol side chain. Structures of 1 was determined by extensive analysis of 1D and 2D NMR spectroscopic data. Compound 1 showed antimicrobial activity against Gram-positive bacteria. This is the first discovery of aromatic polyketides from the genus Rhodococcus.

Pangenomic and functional investigations for dormancy and biodegradation features of an organic pollutant-degrading bacterium Rhodococcus biphenylivorans TG9

Environmental bacteria contain a wealth of untapped potential in the form of biodegradative genes. Leveraging this potential can often be confounded by a lack of understanding of fundamental survival strategies, like dormancy, for environmental stress. Investigating bacterial dormancy-to-degradation relationships enables improvement of bioremediation. Here, we couple genomic and functional assessment to provide context for key attributes of the organic pollutant-degrading strain Rhodococcus biphenylivorans TG9. Whole genome sequencing, pangenome analysis and functional characterization were performed to elucidate important genes and gene products, including antimicrobial resistance, dormancy, and degradation. Rhodococcus as a genus has strong potential for degradation and dormancy, which we demonstrate using R. biphenylivorans TG9 as a model. We identified four Resuscitation-promoting factor (Rpf) encoding genes in TG9 involved in dormancy and resuscitation. We demonstrate that R. biphenylivorans TG9 grows on fourteen typical organic pollutants, and exhibits a robust ability to degrade biphenyl and several congeners of polychlorinated biphenyls. We further induced TG9 into a dormant state and demonstrated pronounced differences in morphology and activity. Together, these results expand our understanding of the genus Rhodococcus and the relationship between dormancy and biodegradation in the presence of environmental stressors.

Alterations of gut microbiota in gestational diabetes patients during the second trimester of pregnancy in the Shanghai Han population

BackgroundThe causes of gestational diabetes mellitus (GDM) are still unclear. Recent studies have found that the imbalance of the gut microbiome could lead to disorders of human metabolism and immune system, resulting in GDM. This study aims to reveal the different gut compositions between GDM and normoglycemic pregnant women and find the relationship between gut microbiota and GDM.MethodsFecal microbiota profiles from women with GDM (n = 21) and normoglycemic women (n = 32) were assessed by 16S rRNA gene sequencing. Fasting metabolic hormone concentrations were measured using multiplex ELISA.ResultsMetabolic hormone levels, microbiome profiles, and inferred functional characteristics differed between women with GDM and healthy women. Additionally, four phyla and seven genera levels have different correlations with plasma glucose and insulin levels. Corynebacteriales (order), Nocardiaceae (family), Desulfovibrionaceae (family), Rhodococcus (genus), and Bacteroidetes (phylum) may be the taxonomic biomarkers of GDM. Microbial gene functions related to amino sugar and nucleotide sugar metabolism were found to be enriched in patients with GDM.ConclusionOur study indicated that dysbiosis of the gut microbiome exists in patients with GDM in the second trimester of pregnancy, and gut microbiota might be a potential diagnostic biomarker for the diagnosis, prevention, and treatment of GDM.

Genome-Based Characterization of Plant-Associated Rhodococcus qingshengii RL1 Reveals Stress Tolerance and Plant-Microbe Interaction Traits.

Stress tolerant, plant-associated bacteria can play an important role in maintaining a functional plant microbiome and protecting plants against various (a)biotic stresses. Members of the stress tolerant genus Rhodococcus are frequently found in the plant microbiome. Rhodococcus qingshengii RL1 was isolated from Eruca sativa and the complete genome was sequenced, annotated and analyzed using different bioinformatic tools. A special focus was laid on functional analyses of stress tolerance and interactions with plants. The genome annotation of RL1 indicated that it contains a repertoire of genes which could enable it to survive under different abiotic stress conditions for e.g., elevated mercury concentrations, to interact with plants via root colonization, to produce phytohormones and siderophores, to fix nitrogen and to interact with bacterial signaling via a LuxR-solo and quorum quenching. Based on the identified genes, functional analyses were performed in vitro with RL1 under different growth conditions. The R. qingshengii type strain djl6 and a closely related Rhodococcus erythropolis BG43 were included in the experiments to find common and distinct traits between the strains. Genome based phylogenetic analysis of 15 available and complete R. erythropolis and R. qingshengii genome sequences revealed a separation of the R. erythropolis clade in two subgroups. First one harbors only R. erythropolis strains including the R. erythropolis type strain. The second group consisted of the R. qingshengii type strain and a mix of R. qingshengii and R. erythropolis strains indicating that some strains of the second group should be considered for taxonomic re-assignment. However, BG43 was clearly identified as R. erythropolis and RL1 clearly as R. qingshengii and the strains had most tested traits in common, indicating a close functional overlap of traits between the two species.

Rhodococcus as Biofactories for Microbial Oil Production.

Bacteria belonging to the Rhodococcus genus are frequent components of microbial communities in diverse natural environments. Some rhodococcal species exhibit the outstanding ability to produce significant amounts of triacylglycerols (TAG) (>20% of cellular dry weight) in the presence of an excess of the carbon source and limitation of the nitrogen source. For this reason, they can be considered as oleaginous microorganisms. As occurs as well in eukaryotic single-cell oil (SCO) producers, these bacteria possess specific physiological properties and molecular mechanisms that differentiate them from other microorganisms unable to synthesize TAG. In this review, we summarized several of the well-characterized molecular mechanisms that enable oleaginous rhodococci to produce significant amounts of SCO. Furthermore, we highlighted the ability of these microorganisms to degrade a wide range of carbon sources coupled to lipogenesis. The qualitative and quantitative oil production by rhodococci from diverse industrial wastes has also been included. Finally, we summarized the genetic and metabolic approaches applied to oleaginous rhodococci to improve SCO production. This review provides a comprehensive and integrating vision on the potential of oleaginous rhodococci to be considered as microbial biofactories for microbial oil production.

Responses to Ecopollutants and Pathogenization Risks of Saprotrophic Rhodococcus Species.

Under conditions of increasing environmental pollution, true saprophytes are capable of changing their survival strategies and demonstrating certain pathogenicity factors. Actinobacteria of the genus Rhodococcus, typical soil and aquatic biotope inhabitants, are characterized by high ecological plasticity and a wide range of oxidized organic substrates, including hydrocarbons and their derivatives. Their cell adaptations, such as the ability of adhering and colonizing surfaces, a complex life cycle, formation of resting cells and capsule-like structures, diauxotrophy, and a rigid cell wall, developed against the negative effects of anthropogenic pollutants are discussed and the risks of possible pathogenization of free-living saprotrophic Rhodococcus species are proposed. Due to universal adaptation features, Rhodococcus species are among the candidates, if further anthropogenic pressure increases, to move into the group of potentially pathogenic organisms with “unprofessional” parasitism, and to join an expanding list of infectious agents as facultative or occasional parasites.

Rhodococcus comparative genomics reveals a phylogenomic-dependent non-ribosomal peptide synthetase distribution: insights into biosynthetic gene cluster connection to an orphan metabolite.

Natural products (NPs) are synthesized by biosynthetic gene clusters (BGCs), whose genes are involved in producing one or a family of chemically related metabolites. Advances in comparative genomics have been favourable for exploiting huge amounts of data and discovering previously unknown BGCs. Nonetheless, studying distribution patterns of novel BGCs and elucidating the biosynthesis of orphan metabolites remains a challenge. To fill this knowledge gap, our study developed a pipeline for high-quality comparative genomics for the actinomycete genus Rhodococcus, which is metabolically versatile, yet understudied in terms of NPs, leading to a total of 110 genomes, 1891 BGCs and 717 non-ribosomal peptide synthetases (NRPSs). Phylogenomic inferences showed four major clades retrieved from strains of several ecological habitats. BiG-SCAPE sequence similarity BGC networking revealed 44 unidentified gene cluster families (GCFs) for NRPS, which presented a phylogenomic-dependent evolution pattern, supporting the hypothesis of vertical gene transfer. As a proof of concept, we analysed in-depth one of our marine strains, Rhodococcus sp. H-CA8f, which revealed a unique BGC distribution within its phylogenomic clade, involved in producing a chloramphenicol-related compound. While this BGC is part of the most abundant and widely distributed NRPS GCF, corason analysis unveiled major differences regarding its genetic context, co-occurrence patterns and modularity. This BGC is composed of three sections, two well-conserved right/left arms flanking a very variable middle section, composed of nrps genes. The presence of two non-canonical domains in H-CA8f’s BGC may contribute to adding chemical diversity to this family of NPs. Liquid chromatography-high resolution MS and dereplication efforts retrieved a set of related orphan metabolites, the corynecins, which to our knowledge are reported here for the first time in Rhodococcus. Overall, our data provide insights to connect BGC uniqueness with orphan metabolites, by revealing key comparative genomic features supported by models of BGC distribution along phylogeny.