Bioprospection of the bacterial β-myrcene-biotransforming trait in the rhizosphere

The biocatalysis of β-myrcene into value-added compounds, with enhanced organoleptic/therapeutic properties, may be performed by resorting to specialized enzymatic machinery of β-myrcene-biotransforming bacteria. Few β-myrcene-biotransforming bacteria have been studied, limiting the diversity of genetic modules/catabolic pathways available for biotechnological research. In our model Pseudomonas sp. strain M1, the β-myrcene catabolic core-code was identified in a 28-kb genomic island (GI). The lack of close homologs of this β-myrcene-associated genetic code prompted a bioprospection of cork oak and eucalyptus rhizospheres, from 4 geographic locations in Portugal, to evaluate the environmental diversity and dissemination of the β-myrcene-biotransforming genetic trait (Myr+). Soil microbiomes were enriched in β-myrcene-supplemented cultures, from which β-myrcene-biotransforming bacteria were isolated, belonging to Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, and Sphingobacteriia classes. From a panel of representative Myr+ isolates that included 7 bacterial genera, the production of β-myrcene derivatives previously reported in strain M1 was detected in Pseudomonas spp., Cupriavidus sp., Sphingobacterium sp., and Variovorax sp. A comparative genomics analysis against the genome of strain M1 found the M1-GI code in 11 new Pseudomonas genomes. Full nucleotide conservation of the β-myrcene core-code was observed throughout a 76-kb locus in strain M1 and all 11 Pseudomonas spp., resembling the structure of an integrative and conjugative element (ICE), despite being isolated from different niches. Furthermore, the characterization of isolates not harboring the Myr+-related 76-kb locus suggested that they may biotransform β-myrcene via alternative catabolic loci, being thereby a novel source of enzymes and biomolecule catalogue for biotechnological exploitation. KEY POINTS: • The isolation of 150 Myr+ bacteria hints the ubiquity of such trait in the rhizosphere. • The Myr+ trait is spread across different bacterial taxonomic classes. • The core-code for the Myr+ trait was detected in a novel ICE, only found in Pseudomonas spp.

Open access funding provided by FCT|FCCN (b-on). This work was supported by the project VALEU (PTDC/EAM-AMB/30488/2017), by the strategic program UID/BIA/04050/2013 (POCI-01–0145-FEDER-007569), by the GenomePT project (POCI-01–0145-FEDER-022184), supported by COMPETE 2020—Opera tional Programme for Competitiveness and Internationalisation (POCI), Lisboa Portugal Regional Operational Programme (Lisboa2020), Algarve Portugal Regional Operational Programme (CRESC Algarve2020), under the Portugal 2020 Partnership Agreement, funded by national funds through the Fundação para a Ciência e a Tecnologia (FCT) I.P. and the European Regional Development Fund (ERDF), through the COMPETE2020-Programa Operacional Competitividade e Internacionalização (POCI). The work was supported as well through the Ph.D. grants to P.S-C (grant number SFRH/BD/76894/2011) and to F.S. (grant number PD/BD/146184/2019).