Temperatures above 37°C increase virulence of a convergent Klebsiella pneumoniae sequence type 307 strain

Hypermucoviscosity in Klebsiella pneumoniae is often related to the overexpression of capsular polysaccharides, regulated by complex biosynthetic mechanisms in response to external cues. However, little is known about the processes involved in hypermucoviscosity in convergent K. pneumoniae, which combine extensive drug resistance with high bacterial virulence, under pathophysiological conditions. This study aimed to fill this gap by investigating the temperature dependence of hypermucoviscosity and overall virulence in a convergent K. pneumoniae strain isolated during a clonal outbreak belonging to the high-risk sequence type (ST)307. Hypermucoviscosity, biofilm formation, and mortality rates in Galleria mellonella larvae were examined at different temperatures (room temperature, 28{degrees}C, 37{degrees}C, 40{degrees}C and 42{degrees}C) and with various phenotypic experiments including electron microscopy. The underlying mechanisms of the phenotypic changes were explored via qPCR analysis to evaluate plasmid copy numbers, and transcriptomics. Our results indicate a temperature-dependent "switch" above 37{degrees}C to a hypermucoviscous phenotype, correlating with increased biofilm formation capacity and in vivo mortality, which might be due to a bacterial response to pathophysiological conditions, i.e., fever. In addition, we detected upregulation of a hybrid plasmid encoding both carbapenemase and the mucoid regulator rmpA genes. Surprisingly, rmpA did not exhibit temperature-dependent differential gene expression, suggesting other drivers. Apparent co-regulation of hypermucoviscosity and fimbrial expression was also identified. This study not only revealed the impact that increased temperatures above 37{degrees}C have on hypermucoviscosity and virulence in a convergent K. pneumoniae strain but contributes to the understanding of previously unrecognized dimension of K. pneumoniaes behavior, emphasizing its adaptability to changing environments. Abstract importanceUnderstanding the temperature-dependent dynamics of hypermucoviscosity in Klebsiella pneumoniae is crucial for unraveling the intricacies of its hypervirulence. This study investigates a convergent K. pneumoniae strain, ST307, revealing a temperature-dependent switch to hypermucoviscosity above 37 {degrees}C. The findings showcase a correlation between increased temperature, hypermucoviscosity, enhanced attachment, and heightened in vivo mortality. Notably, a hybrid plasmid encoding carbapenemase and mucoid regulator genes was upregulated at elevated temperatures. The study sheds light on previously unexplored aspects of K. pneumoniae behavior, emphasizing its adaptability in response to changing environments. The identified temperature-associated regulatory mechanisms offer insights into the pathogens response to fever, contributing to our broader understanding of bacterial adaptation. This research contributes to addressing the global challenge of hypervirulent, drug-resistant K. pneumoniae strains, providing valuable implications for future treatment strategies.