Co-occurrence of Klebsiella variicola and Klebsiella pneumoniae Both Carrying bla_KPC from a Respiratory Intensive Care Unit Patient

Abstract

Objective

The aim of this study was to use whole-genome sequencing to characterize Klebsiella pneumoniae SKp2F and Klebsiella variicola SKv2E, both carrying bla_KPC, co-isolated from the same sputum specimen.

Methods

Antimicrobial susceptibility testing was performed using microbroth dilution. Biofilm formation was determined by crystal violet staining and virulence was measured by a serum killing assay. Whole-genome sequencing of SKp2F and SKv2E was performed using an Illumina sequencer and the genetic characteristics were analyzed by computer.

Results

SKp2F and SKv2E were sensitive only to tigecycline and polymyxin among the tested antibiotics. The biofilm-forming ability of SKv2E is stronger than that of SKp2F. The grades of serum resistance of SKp2F and SKv2E are 4 and 3. MLST analysis of the 6,115,610 bp and 5,403,687 bp of SKv2E and SKp2F showed associations with ST1615 and ST631, respectively. SKv2E carried 13 resistance genes (bla_KPC-2, bla_TEM-1A, bla_LEN17, aadA16, arr-3, qnrB4, oqxA/B, dfrA27, sul1, tetD, fosA, qacEΔ1) and SKp2F carried 23 (bla_KPC-2, bla_CTX-M-3, bla_TEM-1B, bla_CTX-M-65, bla_SHV-27, aac(6ʹ)-IIa, rmtB, arr-3, aph(3ʹ)-Ia, aadA16, qnrS1, aac(6ʹ)-Ib-cr, qnrB91, oqxA/B, mph(A), tet(A), fosA, dfrA27, and two copies of qacEΔ1-sul1). Most of them were carried by various mobile genetic elements, such as IncFIB(K)/IncFII(K)/IncFII(Yp), IncFII(K) plasmid, Tn6338, and In469. Both SKv2E and SKp2F carried a large number of virulence factors, including type 1 and 3 fimbriae, capsule, aerobactin (iutA), ent siderophore (entABCDEFS, fepABCDGfes), and salmochelin (iroE/iroEN). SKv2E also carried type IV pili (pilW), fimbrial adherence (steB, stfD), and capsule biosynthesis gene (glf).

Conclusion

bla_KPC-2-carrying K. variicola and K. pneumoniae, which carried multiple resistance genes, virulence factors, and highly similar mobile genetic elements, were identified from the same specimen, indicating that clinical samples may carry multiple bacteria. We should avoid misidentification, and bear in mind that resistance genes carrying mobile genetic elements can be transmitted or integrated between bacteria in the same host.

Keywords:

• Klebsiella variicola
• Klebsiella pneumoniae
• carbapenem-resistant Enterobacteriaceae
• CRE
• bla _KPC

Nucleotide Sequence Accession Numbers

These Whole Genome Shotgun projects have been deposited in DDBJ/EMBL/GenBank under the sequence accession numbers JAHRXK000000000 and JAHRXL000000000 for Klebsiella pneumoniae strain SKp2F and Klebsiella variicola strain SKv2E, respectively.

Ethical Approval

This study was conducted after agreement from the local ethics committee (no. 20180309059) and with the patient’s informed consent.

Disclosure

The authors have no conflicts of interest to declare.