Transcriptomic analysis of induced resistance to polymyxin in carbapenem-resistant Enterobacter cloacae complex isolate carrying mcr-9.

OBJECTIVES: Polymyxins are currently the last-resort treatment against multi-drug resistant Gram-negative bacterial infections, but plasmid-mediated mobile polymyxin resistance genes (mcr) threaten its efficacy, especially in carbapenem-resistant Enterobacter cloacae complex (CRECC). The objective of this study was to provide insights into the mechanism of polymyxin-induced bacterial resistance and the effect of overexpression of mcr-9. METHODS: The clinical strain CRECC414 carrying the mcr-9 gene was treated with a gradient concentration of polymyxin. Subsequently, the broth microdilution was used to determine the minimum inhibitory concentration (MIC) and RT-qPCR was utilized to assess mcr-9 expression. Transcriptome sequencing and whole genome sequencing (WGS) was utilized to identify alterations in strains resulting from increased polymyxin resistance, and significant transcriptomic differences were analysed alongside a comprehensive examination of metabolic networks at the genomic level. RESULTS: Polymyxin treatment induced the upregulation of mcr-9 expression and significantly elevated the MIC of the strain. Furthermore, the WGS and transcriptomic results revealed a remarkable up-regulation of arnBCADTEF gene cassette, indicating that the Arn/PhoPQ system-mediated L-Ara4N modification is the preferred mechanism for achieving high levels of resistance. Additionally, significant alterations in bacterial gene expression were observed with regards to multidrug efflux pumps, oxidative stress and repair mechanisms, cell membrane biosynthesis, as well as carbohydrate metabolic pathways. CONCLUSION: Polymyxin greatly disrupts the transcription of vital cellular pathways. A complete PhoPQ two-component system is a prerequisite for polymyxin resistance of Enterobacter cloacae, even though mcr-9 is highly expressed. These findings provide novel and important information for further investigation of polymyxin resistance of CRECC.

In vitro and in vivo activity of ceftazidime/avibactam and aztreonam alone or in combination against mcr-9, serine- and metallo-ß-lactamases-co-producing carbapenem-resistant Enterobacter cloacae complex.

PURPOSE: Enterobacteriaceae carrying mcr-9, in particularly those also co-containing metallo-ß-lactamase (MBL) and TEM type ß-lactamase, present potential transmission risks and lack adequate clinical response methods, thereby posing a major threat to global public health. The aim of this study was to assess the antimicrobial efficacy of a combined ceftazidime/avibactam (CZA) and aztreonam (ATM) regimen against carbapenem-resistant Enterobacter cloacae complex (CRECC) co-producing mcr-9, MBL and TEM. METHODS: The in vitro antibacterial activity of CZA plus ATM was evaluated using a time-kill curve assay. Furthermore, the in vivo interaction between CZA plus ATM was confirmed using a Galleria mellonella (G. mellonella) infection model. RESULTS: All eight clinical strains of CRECC, co-carrying mcr-9, MBL and TEM, exhibited high resistance to CZA and ATM. In vitro time-kill curve analysis demonstrated that the combination therapy of CZA + ATM exerted significant bactericidal activity against mcr-9, MBL and TEM-co-producing Enterobacter cloacae complex (ECC) isolates with a 100% synergy rate observed in our study. Furthermore, in vivo survival assay using Galleria mellonella larvae infected with CRECC strains co-harboring mcr-9, MBL and TEM revealed that the CZA + ATM combination significantly improved the survival rate compared to the drug-treatment alone and untreated control groups. CONCLUSION: To our knowledge, this study represents the first report on the in vitro and in vivo antibacterial activity of CZA plus ATM against CRECC isolates co-harboring mcr-9, MBL and TEM. Our findings suggest that the combination regimen of CZA + ATM provides a valuable reference for clinicians to address the increasingly complex antibiotic resistance situation observed in clinical microorganisms.

Activities of aztreonam in combination with several novel ß-lactam-ß-lactamase inhibitor combinations against carbapenem-resistant Klebsiella pneumoniae strains coproducing KPC and NDM.

Isolates coproducing serine/metallo-carbapenems are a serious emerging public health threat, given their rapid dissemination and the limited number of treatment options. The purposes of this study were to evaluate the in vitro antibacterial activity of novel ß-lactam-ß-lactamase inhibitor combinations (BLBLIs) against carbapenem-resistant Klebsiella pneumoniae (CRKP) coproducing metallo-ß-lactamase and serine-ß-lactamase, and to explore their effects in combination with aztreonam, meropenem, or polymyxin in order to identify the best therapeutic options. Four CRKP isolates coproducing K. pneumoniae carbapenemase (KPC) and New Delhi metallo-ß-lactamase (NDM) were selected, and a microdilution broth method was used to determine their susceptibility to antibiotics. Time-kill assay was used to detect the bactericidal effects of the combinations of antibiotics. The minimum inhibitory concentration (MIC) values for imipenem and meropenem in three isolates did not decrease after the addition of relebactam or varbobactam, but the addition of avibactam to aztreonam reduced the MIC by more than 64-fold. Time-kill assay demonstrated that imipenem-cilastatin/relebactam (ICR) alone exerted a bacteriostatic effect against three isolates (average reduction: 1.88 log10 CFU/mL) and ICR combined with aztreonam exerted an additive effect. Aztreonam combined with meropenem/varbobactam (MEV) or ceftazidime/avibactam (CZA) showed synergistic effects, while the effect of aztreonam combined with CZA was inferior to that of MEV. Compared with the same concentration of aztreonam plus CZA combination, aztreonam/avibactam had a better bactericidal effect (24 h bacterial count reduction >3 log10CFU/mL). These data indicate that the combination of ATM with several new BLBLIs exerts powerful bactericidal activity, which suggests that these double ß-lactam combinations might provide potential alternative treatments for infections caused by pathogens coproducing-serine/metallo-carbapenems.

Comparison of MRI and CT Scan for the Detection of Liver Cancer.

OBJECTIVE: The objective of the paper was to compare the value of CT and MRI in the diagnosis of primary carcinoma of the liver. METHODS: A retrospective analysis was performed on 132 cases of suspected primary liver carcinoma. CT and MRI diagnosis were performed and pathological results were compared to determine the diagnostic value of the two methods. RESULTS: 96 cases were diagnosed as primary liver carcinoma by pathological examination after operation. The total detection rate of 96 lesions through MRI was 93.75%, while 84.38% through CT (P<0.05). For lesions with a <3 cm diameter, the CT detection rates of lesions in the plain, arterial, portal, and equilibrium phases were 52.94%, 73.53%, 58.82%, and 58.82% respectively. For lesions with a diameter ≥ 3 cm, the CT detection rate was 80.65 %, 93.55%, 85.48%, and 83.87%, respectively (P<0.05). For lesions with <3cm diameter, the MRI detection rates of lesions in the T1WI, T2WI, LAVA arterial phase, LAVA portal phase, and LAVA balance phase were 61.76%, 76.47%, 88.24%, 79.41%, and 52.94%, respectively, and for lesions with ≥3cm diameter, the detection rates of MRI were 77.42%, 87.10%, 91.94%, 90.32%, and 90.32%, respectively, and the detection rate of lesions with ≥3cm diameter in the balance phase of LAVA was higher (P<0.05). Taking pathological results as the gold standard, the sensitivity of diagnosing primary liver carcinoma through CT is 81.25%, specificity is 75.00%, accuracy is 79.55%, the positive predictive value is 89.66%, the negative predictive value is 60.00%, and the values of the same parameters for the MRI are 93.75. %, 86.11%, 91.67%, 94.74%, and 83.78% respectively. CONCLUSION: Both CT and MRI have diagnostic value for primary liver carcinoma. The comparison showed that MRI has a higher diagnostic value and higher detection rate for small lesions. However, the actual process of diagnosis cannot rely solely on MRI, and a comprehensive combination of diagnosis methods will be effective.

High frequency of rejections in HIV-positive recipients of kidney transplantation: a single center prospective trial.

BACKGROUND: This is a single institution report of the incidence of combined acute antibody-mediated rejection (ABMR) + acute cellular rejection (ACR) [mixed rejection] in HIV (+) kidney transplant recipients. METHODS: We prospectively enrolled 92 HIV (+) patients who received a kidney transplant between 2001 and 2009. There were three cohorts: no rejection [n=26], ACR [n=53], and mixed rejections (ABMR and ACR) [n=13]. Immunosuppression comprised of basiliximab, cyclosporine, sirolimus, and steroid minimization. Fisher exact tests for categorical variables, t test for continuous variables, and Kaplan-Meier estimates were used to describe events. RESULTS: Mixed rejections were seen in all 13 HIV (+) kidney transplant recipients (14%) with a median time to ABMR of 48 days. Acute cellular rejection occurred in 28% at 1 month and 55% at 12 months. eGFR was lower for recipients who experienced ABMR versus those experiencing ACR and those never experiencing rejection up to 3 years (14 ± 9.4 vs 19 ± 3.3 vs 29 ± 7.3 mL/min, respectively). Kaplan-Meier showed that graft survival up to 9 years was worse in recipients experiencing mixed rejection. Suboptimal donors with terminal creatinine greater than 2.5 mg/dL was associated with increased incidence of mixed rejections versus cellular rejections and no rejection (42% vs 17% vs. 8%, respectively). CONCLUSIONS: Our single center study showed a relatively higher incidence of mixed rejection compared with that reported for non-HIV transplant recipients. A donor terminal serum creatinine greater than 2.5 mg/dL predicted mixed rejection, which was associated with poor outcomes. Donor selection and optimization of immunosuppression may be critical in these patients.