CusS-CusR Two-Component System Mediates Tigecycline Resistance in Carbapenem-Resistant Klebsiella pneumoniae

Abstract

Background: The increase in carbapenem-resistant Klebsiella pneumoniae (CRKP), especially the emergence of tigecycline-resistant K. pneumoniae (KP), is a serious public health concern. However, the underlying mechanism of tigecycline resistance is unclear. In this study, we evaluated the role of the CusS-CusR two-component system (TCS), which is associated with copper/silver resistance, in tigecycline resistance in CRKP. Methods: Following the in vitro evolution of tigecycline-resistant KP, the minimum inhibitory concentrations of tigecycline were determined using the micro-broth dilution method. RNA sequencing and data analysis were performed to identify differentially expressed genes. Quantitative PCR (qPCR) was performed to verify the genes of interest. Genes associated with tigecycline resistance, such as ramR, tex (T), and tet (A), were detected by PCR, and then mutants were confirmed by sequencing. Additionally, the efflux pump-associated genes soxS, oqxA, oqxB, acrE, and acrF were also analyzed by qPCR. CusR was deleted and complemented by the suicide vector pKO3-Km plasmid and pGEM-T-easy plasmid, respectively. Results: Nine strains of KP were evaluated in our study. Strains A2 and A3 were evolved from A1, B2, and B3 were evolved from B1, and C2 and C3 were evolved from C1. The tigecycline minimum inhibitory concentration for A1, B1, and C1 was 0.5 µg/mL; that for A2, B2, and C3 was 16.0 µg/mL; and that for A3, B3, and C3 was 32.0 µg/mL. RNAsequencing and qPCR confirmed that the differentially expressed genes cusE, cusS, cusR, cusC, cusF, cusB, and cusA showed higher expression in C2 and C3 than in C1. Genes related to the efflux pump AcrAB-TolC showed higher expression in B2 and B3 than in B1. No mutants of ramR, tex (T), or tet (A) were detected. SoxS, oqxA, oqxB, acrE, and acrF did not show increased expression in any group. After deletion and

Molecular characterization of carbapenem-resistant Enterobacteriaceae and emergence of tigecycline non-susceptible strains in the Henan province in China: a multicentrer study

“…Meanwhile, the plasmid-mediated high-level TGC resistance gene tetX has recently emerged in animals and humans [10, 17]. Furthermore, the CusS-CusR two-component system is reportedly associated with CRKP resistance to TGC [18]. TGC resistance mechanisms have also been found to be caused by overexpression of tetL and tetM in Enterococcus faecium [19].…”

In-Situ Metatranscriptomic Analyses Reveal the Metabolic Flexibility of the Thermophilic Anoxygenic Photosynthetic Bacterium Chloroflexus aggregans in a Hot Spring Cyanobacteria-Dominated Microbial Mat

“…Blue-green cyanobacterial mat samples were collected from a small pool at 56 °C with slightly alkaline (pH 8.5–8.9) and sulfidic (46–138 µM) hot spring water [ 18 , 25 , 26 , 27 ] at Nakabusa Hot Springs, Nagano Prefecture, Japan (36°23′33″ N, 137°44′52″ E) [ 20 ]. Microbial mat samples of approximately 3 mm thickness with two distinct vertical layers, a green top layer and an orange-colored bottom layer ( Figure 1 ), were randomly collected in triplicate using a size 4 cork borer (8 mm diameter) as previously described [ 13 , 14 ].…”

“…RNA extraction from microbial mat samples was performed as previously described [ 18 ]. Briefly, 0.10–0.21 g wet weight samples were used for RNA extraction with an RNeasy PowerBiofilm Kit (Qiagen, Valencia, CA, USA) following the manufacturer’s protocol.…”

“…Microbial mats in the slightly alkaline, sulfidic Nakabusa Hot Springs have been intensively studied with regard to their microbial diversity and functions [ 13 , 14 , 15 , 16 , 17 , 18 , 19 , 20 , 21 ]. At water temperatures of 63–70 °C, olive-green microbial mats (“ Chloroflexus mats”) are dominated by C. aggregans [ 14 , 15 ], and oxygenic cyanobacteria are not found.…”

DNA-Binding and Transcription Activation by Unphosphorylated Response Regulator AgrR From Cupriavidus metallidurans Involved in Silver Resistance

“…The emergence of such organisms may impede the utilization of silver as antimicrobial agent, especially for medical applications. Furthermore, it has been shown that in some instances, increased metal resistance may be accompanied by an increase in antibiotic resistance ( McHugh et al, 1975 ; Choudhury and Kumar, 1998 ; Perron et al, 2004 ; Dieppois et al, 2012 ; Pal et al, 2017 ; Wang et al, 2017 ; Chen et al, 2019 ).…”