Genomic and functional characterization of the rare thiosulfonate thiolutin dioxide as a low toxicity antibacterial candidate from a novel Streptomyces lineage

Abstract The rapid emergence of antibiotic-resistant bacteria drives an urgent need for novel antimicrobial agents. Member of the genus Streptomyces are widely recognized as a rich source of potent bioactive natural products. This study focused on the isolation and characterization of antimicrobial compounds from the soil-derived Streptomyces sp. PKA51. Based on 16S rRNA gene sequencing and whole genome analysis, strain PKA51 represents a potentially novel species within the same clade as Streptomyces luteosporeus . Chromatographic purification and structural characterization revealed two bioactive dithiolopyrrolone metabolites, thiolutin (C 8 H 8 N 2 O 2 S 2 ) and its rare oxidized derivative, thiolutin dioxide (C 8 H 8 N 2 O 4 S 2 ). Both compounds demonstrated broad-spectrum antibacterial activity against high-priority pathogens, including methicillin-resistant Staphylococcus aureus (MRSA) DMST 20654, methicillin-resistant Staphylococcus epidermidis (MRSE) SUTH, and multidrug-resistant clinical isolates ( Escherichia coli 2026 and Klebsiella pneumoniae 1617). Thiolutin and thiolutin dioxide exhibited minimum inhibitory concentration values ranging from 2 to 128 µg/mL and 4 to 64 µg/mL, respectively. Thiolutin showed the greatest inhibitory effect against MRSA (2 µg/mL), whereas thiolutin dioxide was most effective against MRSE (4 µg/mL). Notably, in vitro cytotoxicity assays on Vero cells demonstrated that thiolutin dioxide possesses significantly lower host toxicity (IC 50 = 39.73 µg/mL) compared to parent thiolutin (IC 50 = 1.52 µg/mL), yielding a superior Selectivity Index and a safer profile for therapeutic development. Whole-genome analysis of strain PKA51 revealed a putative strain-specific biosynthetic gene cluster (BGC) variant involved in directing the synthesis of both dithiolopyrrolones. To the best of our knowledge, thiolutin dioxide remains a rarely reported thiosulfonate derivative whose natural occurrence is poorly characterized. This study broadens current knowledge of thiolutin dioxide by establishing its production in Streptomyces sp. PKA51, detailing its antibacterial spectrum, identifying its putative strain-specific BGC variant, and confirming its low mammalian cytotoxicity. These findings highlight the potential of this bioactive natural product in combating antibiotic-resistant bacteria and its relevance for pharmaceutical drug development.