Abstract
Klebsiella pneumoniae infections affect infants and the immunocompromised, and the recent emergence of hypervirulent and multidrug-resistant K. pneumoniae lineages is a critical health care concern. Hypervirulence in K. pneumoniae is mediated by several factors, including the overproduction of extracellular capsule. However, the full details of how K. pneumoniae capsule biosynthesis is achieved or regulated are not known. We have developed a robust and sensitive procedure to identify genes influencing capsule production, density-TraDISort, which combines density gradient centrifugation with transposon insertion sequencing. We have used this method to explore capsule regulation in two clinically relevant Klebsiella strains, K. pneumoniae NTUH-K2044 (capsule type K1) and K. pneumoniae ATCC 43816 (capsule type K2). We identified multiple genes required for full capsule production in K. pneumoniae, as well as putative suppressors of capsule in NTUH-K2044, and have validated the results of our screen with targeted knockout mutants. Further investigation of several of the K. pneumoniae capsule regulators identified-ArgR, MprA/KvrB, SlyA/KvrA, and the Sap ABC transporter-revealed effects on capsule amount and architecture, serum resistance, and virulence. We show that capsule production in K. pneumoniae is at the center of a complex regulatory network involving multiple global regulators and environmental cues and that the majority of capsule regulatory genes are located in the core genome. Overall, our findings expand our understanding of how capsule is regulated in this medically important pathogen and provide a technology that can be easily implemented to study capsule regulation in other bacterial species.IMPORTANCE Capsule production is essential for K. pneumoniae to cause infections, but its regulation and mechanism of synthesis are not fully understood in this organism. We have developed and applied a new method for genome-wide identification of capsule regulators. Using this method, many genes that positively or negatively affect capsule production in K. pneumoniae were identified, and we use these data to propose an integrated model for capsule regulation in this species. Several of the genes and biological processes identified have not previously been linked to capsule synthesis. We also show that the methods presented here can be applied to other species of capsulated bacteria, providing the opportunity to explore and compare capsule regulatory networks in other bacterial strains and species.
Highlights
Klebsiella pneumoniae infections affect infants and the immunocompromised, and the recent emergence of hypervirulent and multidrug-resistant K. pneumoniae lineages is a critical health care concern
On the basis of this observation, we speculated that Klebsiella cells of different capsulation states could be separated by density-based centrifugation and that this method could be combined with transposon-directed insertion sequencing (TraDIS) to screen for capsule-regulating genes
Tests with K. pneumoniae NTUH-K2044 (Fig. 1A) and K. pneumoniae ATCC 43816 (K2, hypermucoid) and a noncapsulated Escherichia coli control showed that these strains differed in density and migrated to above 15%, above mbio.asm.org 3
Summary
Klebsiella pneumoniae infections affect infants and the immunocompromised, and the recent emergence of hypervirulent and multidrug-resistant K. pneumoniae lineages is a critical health care concern. We have developed a robust and sensitive procedure to identify genes influencing capsule production, density-TraDISort, which combines density gradient centrifugation with transposon insertion sequencing. We have used this method to explore capsule regulation in two clinically relevant Klebsiella strains, K. pneumoniae NTUH-K2044 (capsule type K1) and K. pneumoniae ATCC 43816 (capsule type K2). We have developed and applied a new method for genome-wide identification of capsule regulators Using this method, many genes that positively or negatively affect capsule production in K. pneumoniae were identified, and we use these data to propose an integrated model for capsule regulation in this species. More than 100 capsule locus types have been identified in K. pneumoniae [17], though the majority of hypervirulent K. pneumoniae isolates represent strains of capsule types K1 and K2 [12, 22]
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
