Tigecycline Susceptibility and the Role of Efflux Pumps in Tigecycline Resistance in KPC-Producing Klebsiella pneumoniae

Abstract

KPC-producing Klebsiella pneumoniae isolates have emerged as important pathogens of nosocomial infections, and tigecycline is one of the antibiotics recommended for severe infections caused by KPC-producing K. pneumoniae. To identify the susceptibility profile of KPC-producing K. pneumoniae to tigecycline and investigate the role of efflux pumps in tigecycline resistance, a total of 215 KPC-producing K. pneumoniae isolates were collected. The minimum inhibitory concentration (MIC) of tigecycline was determined by standard broth microdilution tests. Isolates showing resistance to tigecycline underwent susceptibility test with efflux pump inhibitors. Expression levels of efflux pump genes (acrB and oqxB) and their regulators (ramA, marA, soxS and rarA) were examined by real-time PCR, and the correlation between tigecycline MICs and gene expression levels were analysed. Our results show that the tigecycline resistance rate in these isolates was 11.2%. Exposure of the tigecycline-resistant isolates to the efflux pump inhibitor NMP resulted in an obvious decrease in MICs and restored susceptibility to tigecycline in 91.7% of the isolates. A statistically significant association between acrB expression and tigecycline MICs was observed, and overexpression of ramA was found in three tigecycline-resistant isolates, further analysis confirmed ramR mutations existed in these isolates. Transformation of one mutant with wild-type ramR restored susceptibility to tigecycline and repressed overexpression of ramA and acrB. These data indicate that efflux pump AcrAB, which can be up-regulated by ramR mutations and subsequent ramA activation, contributed to tigecycline resistance in K. pneumoniae clinical isolates.

Over expression of AdeABC and AcrAB-TolC efflux systems confers tigecycline resistance in clinical isolates of Acinetobacter baumannii and Klebsiella pneumoniae

“…However, low expression of soxS was observed in resistant isolates, and increased tigecycline MICs were correlated well with the overexpression of ramA. Although the overexpression of OqxAB efflux systems may also be also associated with tigecycline resistance (15) , no obvious overexpression of OqxB was observed in this study.…”

“…All experiments were performed in triplicate, and the average was calculated. The primers used for the aforementioned genes are listed in Table 1, as described in other studies (10) (11) (12) (13) (14) (15) . Two multidrug-susceptible strains of A. baumannii (AB21) and K. pneumoniae (KP18) with a tigecycline MIC of 1mg/L were used as the reference strains for the two microGrams (expression level = 1).…”

“…Additionally, 137 A. baumannii isolates and 47 K. pneumoniae isolates were resistant to tigecycline (MIC ≥ 4mg/L). As shown (10) rpoB-R ATTGCTTCATCTGCTGGTTG adeB-qPCR-F adeB AACGGACGACCATCTTTGAGTATT qRT-PCR (11) adeB-qPCR-R CAGTTGTTCCATTTCACGCATT adeJ-qPCR-F adeJ ATTGCACCACCAACCGTAAC qRT-PCR (11) adeJ-qPCR-R TAGCTGGATCAAGCCAGATA adeG-qPCR-F adeG TTCATCTAGCCAAGCAGAAG qRT-PCR (12) adeG-qPCR-R GTGTAGTGCCACTGGTTACT adeM-qPCR-F adeM GTAGGTGTAGGCTTATGGA qRT-PCR (13) adeM-qPCR-R GTACCGAAGTGACTGAAAT acrB-qPCR-F acrB AAACTTCGCCACTACGTCATA qRT-PCR (14) acrB-qPCR-R AGCTTAACGCCTCGATCAT oqxB-qPCR-F OqxB CGAAGAAAGACCTCCCTACCC qRT-PCR (15) oqxB-qPCR-R CGCCGCCAATGAGATACA ramA-qPCR-F ramA GATATCGCTCGCCATGC qRT-PCR (14) ramA-qPCR-R CTGTGGTTCTCTTTGCGGTAG soxS-qPCR-F soxS TACCTGCAGCGGATGTTC qRT-PCR (14) soxS-qPCR-R AAGGTTTGCTGCGAGACGTAG adeR-F adeR ATGTTTGATCATTCTTTTTCTTTTG Mutation detection (11) adeR-R TTAATTAACATTTGAAATATG adeS-F adeS ATGAAAAGTAAGTTAGGAATTAGTAAG Mutation detection (11) adeS-R TTAGTTATTCATAGAAATTTTTATG acrR-F acrR GCTAAGCTGCCTGAGAGCAT Mutation detection (14) acrR-R ATGCAAATGCCGGAGAATAC 16S rRNA-qPCR-F 16S rRNA GACGTACTCGCAGAATAAGC qRT-PCR (11) 16S rRNA-qPCR-R TTAGTCTTGCGACCGTACTC rrsE-qPCR-F rrsE GTCATCATGGCCCTTACGAG qRT-PCR (14) rrsE-qPCR-R ACTTTATGAGGTCCGCTTGCT qRT-PCR: quantitative real-time reverse transcription polymerase chain reaction; rRNA:. ribosomal ribonucleic acid.…”

In vitro Susceptibility of Multidrug-Resistant Klebsiella pneumoniae Isolates from an Egyptian Tertiary Care Hospital to Tigecycline and Colistin

“…Fortunately, tigecycline and colistin displayed a considerable activity against such strains where 87.5% and 81.25% of the carbapenemase-positive strains were susceptible to tigecycline and colistin, respectively. These findings are in a range similar to that reported by other investigators (He et al, 2015, andZheng et al, 2017).…”

Risk Factors, Outcomes, and Mechanisms of Tigecycline-Nonsusceptible Klebsiella pneumoniae Bacteremia

“…The RamA and AcrAB pathway and RarA, together with OqxAB pathways are implicated in tigecycline resistance in K. pneumoniae (12). Studies of tigecycline resistance mechanisms in K. pneumoniae usually include only a small number of isolates in the literature (12)(13)(14). Moreover, studies addressing both the clinical features of bacteremia and the tigecycline resistance mechanism have never been reported.…”