Mobilization of the nonconjugative virulence plasmid from hypervirulent Klebsiella pneumoniae

Abstract

Background Klebsiella pneumoniae, as a global priority pathogen, is well known for its capability of acquiring mobile genetic elements that carry resistance and/or virulence genes. Its virulence plasmid, previously deemed nonconjugative and restricted within hypervirulent K. pneumoniae (hvKP), has disseminated into classic K. pneumoniae (cKP), particularly carbapenem-resistant K. pneumoniae (CRKP), which poses alarming challenges to public health. However, the mechanism underlying its transfer from hvKP to CRKP is unclear. Methods A total of 28 sequence type (ST) 11 bloodstream infection-causing CRKP strains were collected from Ruijin Hospital in Shanghai, China, and used as recipients in conjugation assays. Transconjugants obtained from conjugation assays were confirmed by XbaI and S1 nuclease pulsed-field gel electrophoresis, PCR detection and/or whole-genome sequencing. The plasmid stability of the transconjugants was evaluated by serial culture. Genetically modified strains and constructed mimic virulence plasmids were employed to investigate the mechanisms underlying mobilization. The level of extracellular polysaccharides was measured by mucoviscosity assays and uronic acid quantification. An in silico analysis of 2608 plasmids derived from 814 completely sequenced K. pneumoniae strains available in GenBank was performed to investigate the distribution of putative helper plasmids and mobilizable virulence plasmids. Results A nonconjugative virulence plasmid was mobilized by the conjugative plasmid belonging to incompatibility group F (IncF) from the hvKP strain into ST11 CRKP strains under low extracellular polysaccharide-producing conditions or by employing intermediate E. coli strains. The virulence plasmid was mobilized via four modes: transfer alone, cotransfer with the conjugative IncF plasmid, hybrid plasmid formation due to two rounds of single-strand exchanges at specific 28-bp fusion sites or homologous recombination. According to the in silico analysis, 31.8% (242) of the putative helper plasmids and 98.8% (84/85) of the virulence plasmids carry the 28-bp fusion site. All virulence plasmids carry the origin of the transfer site. Conclusions The nonconjugative virulence plasmid in ST11 CRKP strains is putatively mobilized from hvKP or E. coli intermediates with the help of conjugative IncF plasmids. Our findings emphasize the importance of raising public awareness of the rapid dissemination of virulence plasmids and the consistent emergence of hypervirulent carbapenem-resistant K. pneumoniae (hv-CRKP) strains.

Co-Occurrence of Rare ArmA-, RmtB-, and KPC-2–Encoding Multidrug-Resistant Plasmids and Hypervirulence iuc Operon in ST11-KL47 Klebsiella pneumoniae

“…In this study, the p1632-2 plasmid was typed as IncM2 plasmid and had an extremely high-conjugative frequency, higher than the other IncM2- armA plasmid pMS3802OXARMA studied before (10 −1 versus 10 −4 ) ( 25 ). As previous studies described ( 19 , 26 ), the oriT -positive plasmid could be cotransferred with the conjugative plasmid, and the p1632-1 plasmid ( oriT and relaxase protein positive) still holds the potential to be mobilized when p1632-2 is induced.…”

Escherichia fergusonii, an Underrated Repository for Antimicrobial Resistance in Food Animals

“…Plasmid conjugation experiments were performed on the mcr-1 -harboring E. fergusonii strains as described previously ( 34 ). A sodium azide-resistant E. coli strain J53 was used as the recipient.…”

The Role of Plasmid and Resistance Gene Acquisition in the Emergence of ST23 Multi-Drug Resistant, Hypervirulent Klebsiella pneumoniae

“…Recently, Xu et al ( 21 ) reported that virulence plasmid of ST11 hvKp could transfer via coexisting conjugative IncF plasmids using shared oriT regions. Additionally, a specific 28-bp fusion site located in the virulence plasmid of ST11 hvKp could assist with the virulence plasmid forming a hybrid plasmid with conjugative IncF plasmids and then be cotransferred.…”

“…It has been reported that ST11 CR-hvKp possessed five tandem repeats of the bla KPC-2 ::NTEKPC-Id fragment within the IncR plasmid was also mediated by the IS 26 ( 35 ). More importantly, with the help of the IncF plasmids, the nonconjugative virulence plasmid could achieve the mobilization among Enterobacterales strains ( 21 ). IS 26 was closely related to the IncFII plasmid replication among the ST23 K. pneumoniae , alarming that the transformation and conjugation among these organisms should be emphasized.…”

“…S1C), which is similar to Kelly’s study ( 17 ). Xu, et al ( 21 ) reported two mechanisms leading to the mobilization of the nonconjugative virulence plasmid of hvKp. The virulence plasmids containing oriT sequences identical to those of conjugative plasmids would achieve transfer with the help of the conjugative plasmids.…”