Genomic characterisation and gene editing of Marinibacterium sp. CCB-SX1 as a new marine chassis for polyhydroxyalkanoate production

The development of robust microbial platform with customised genetic traits is crucial for advancing polyhydroxyalkanoate (PHA) production as a biodegradable plastic alternative. This study genomically characterised a new marine isolate, Marinibacterium sp. CCB-SX1, for its potential as a PHA-producing chassis. The complete genome comprises a 6.14 Mb chromosome and nine plasmids. Phylogenomic analysis placed CCB-SX1 within Marinibacterium, with genomic metrics (average nucleotide identity and digital DNA-DNA hybridisation) suggesting it represents a new species. Comparative genomics of the family Paracoccaceae revealed an open pangenome with a small core and a large accessory genome, abundant in functions for energy production and conversion, replication, recombination and repair, and transcription. Mobile genetic elements were dominated by integration/excision and transfer-associated genes, reflecting extensive horizontal gene transfer and genomic plasticity. PHA-related genes (phaC, phaB, phaP, phaR, phaZ) were conserved in the soft-core genome, mostly organised as phaR-phaP-phaC-phaZ. Genome annotation of CCB-SX1 revealed a complete pathway for 3-hydroxybutyrate synthesis and methylmalonyl-CoA enzymes enabling 3-hydroxyvalerate formation. CCB-SX1 synthesised PHA from multiple carbon sources, with acetate yielding the highest PHA content (27.3 wt%) and producing the copolymer poly(3-hydroxybutyrate-co-3-hydroxyvalerate) with 1.2 mol% 3HV, while other carbon sources formed poly(3-hydroxybutyrate). A CRISPR-Cas9-nickase system was established to generate single and double knockouts of intracellular depolymerases (phaZ1, phaZ2). Disruption of phaZ1 significantly increased PHA accumulation, while phaZ2 deletion had negligible effect. These findings establish Marinibacterium sp. CCB-SX1 as a genetically tractable marine chassis with potential for metabolic engineering and biopolymer production.