Neeravi, A. | Search bioRxiv

bioRxiv2026

Three Plasmid Strategies, One Intermediate Convergence State: Lineage-Specific Resistance, Virulence Architecture in Dominant Indian Carbapenem-Resistant Klebsiella pneumoniae Clones

Carbapenem-resistant Klebsiella pneumoniae (CRKp) is a critical global healthcare threat driven by high-risk multidrug-resistant (MDR) clones that acquire hypervirulence genes. Although resistance-virulence co-occurrence is extensively documented, the plasmid-level mechanisms facilitating this convergence remain unclear. In this study, we utilized hybrid short- and long-read whole-genome sequencing of 376 clinical CRKp strains to define the evolutionary trajectories and structural plasmid dynamics of three predominant high-risk clones: ST147 (n=157), ST231 (n=108), and ST2096 (n=111). Carbapenemase genes were present in 90% of isolates, predominantly blaOXA-48-like and blaNDM-5 co-harbored with blaCTX-M-15. Virulence profiling indicated high aerobactin (iuc) prevalence (62.7%), while salmochelin and colibactin were undetected. Hypermucoviscosity occurred infrequently (6.6%) and was independent of rmpA/rmpA2, confirming a clear genotype-phenotype discordance. Comparative plasmid mapping revealed three distinct, lineage-specific plasmid configurations underlying this intermediate convergent pathotype: ST147 exhibited dynamic, mosaic hybrid IncFIB-IncHI1B plasmids; ST2096 showed structurally stabilized hybrids; and ST231 retained virulence and resistance determinants on separate, segregated plasmids. These findings show that convergence is regulated by multiple, clone-specific evolutionary routes rather than a single path, highlighting the critical need for more in-depth genomic surveillance capable of identifying convergent plasmids along with high-risk lineages

microbiology↗

bioRxiv2025

Sorafenib, a clinically approved kinase inhibitor attenuates Streptococcus pneumoniae pathogenesis in vivo by targeting serine/threonine kinase StkP

Streptococcus pneumoniae is a respiratory commensal bacterium responsible for over one million annual fatalities globally, particularly among children under five years of age. The rapid emergence of macrolide-resistant strains led the WHO in 2024 to designate S. pneumoniae as a priority pathogen, underscoring the need for alternative strategies such as anti-virulence therapy. Here, we repurposed the FDA-approved cancer drug, sorafenib identified by in silico screening of compounds targeting the bacterial Serine/Threonine kinase protein StkP, an essential regulator of cell division and peptidoglycan synthesis conserved across many bacterial pathogens. Sorafenib interacts with the StkP-kinase domain and exhibited broad-spectrum activity against diverse pneumococcal serotypes including multi-drug-resistant clinical isolates. Ectopic expression of StkP in both wild-type and isogenic mutant strains conferred partial resistance to sorafenib, confirming on-target activity. Scanning electron microscopy revealed aberrant cell-wall morphology, and differential viability staining demonstrated increased membrane permeability. Consistently, sorafenib-treated bacteria showed significantly higher complement C3 deposition and consequent killing in human serum. In human lung epithelial cells, sorafenib reduced bacterial adherence and invasion without detectable host cytotoxicity. Serial passaging at sub-microbicidal concentrations indicated a low propensity for resistance development in vitro. In vivo, sorafenib administration reduced mortality and lung bacterial load in a mouse model of pneumonia. Taken together, our data identify StkP as target of sorafenib in S. pneumoniae and justify its continued preclinical development for therapeutic intervention.

microbiology↗