Clinical, biological and genome-wide comparison of carbapenem-resistant Klebsiella pneumoniae with susceptibility transformation to polymyxin B during therapy.

OBJECTIVES: The emergence of carbapenem-resistant Klebsiella pneumoniae (CRKP) is a major clinical concern, and polymyxin B (PMB) is a 'last resort' antibiotic for its treatment. Understanding the effects of drug susceptibility transformation in CRKP-infected patients undergoing PMB treatment would be beneficial to optimize PMB treatment strategies. METHODS: We retrospectively collected data from patients infected with CRKP and treated with PMB from January 2018 to December 2020. CRKPs were collected before and after PMB therapy, and patients were classified into the 'transformation' group (TG) and 'non-transformation' group (NTG) by the shift of susceptibility to PMB. We compared clinical characteristics between these groups, and further analysed the phenotypic and genome variation of CRKP after PMB susceptibility transformation. RESULTS: A total of 160 patients (37 in the TG and 123 in the NTG) were included in this study. The duration of PMB treatment before PMB-resistant K. pneumoniae (PRKP) appearance in TG was even longer than the whole duration of PMB treatment in NTG (8 [8] vs. 7 [6] days; p 0.0496). Compared with isogenic PMB-susceptible K. pneumoniae (PSKP), most PRKP strains had missense mutations in mgrB (12 isolates), yciC (10 isolates) and pmrB (7 isolates). The competition index of 82.4% (28/34) of PRKP/PSKP pairs was <67.6% (23/34), and 73.5% (25/34) of PRKP strains showed a higher 7-day lethality in Galleria mellonella and a greater ability to resist complement-dependent killing than their corresponding PSKP, respectively. CONCLUSION: Low dose with longer PMB treatment durations may be associated with the emergence of polymyxin resistance. The evolution of PRKP is predominantly mediated by an accumulation of mutations, including those in mgrB, yciC, and pmrB. Lastly, PRKP exhibited reduced growth and increased virulence compared with parental PSKP.

Cefepime, not Piperacillin/Tazobactam use, for empirical treatment of bloodstream infections caused by Enterobacter spp.: Results from a population pharmacokinetic/pharmacodynamic analysis.

OBJECTIVE: There is a paucity of published data to evaluate the efficacy and safety of imipenem, cefepime and piperacillin/tazobactam dosing regimens against bloodstream infections caused by Klebsiella aerogenes (BSIs-Kae) and Enterobacter cloacae complex (BSIs-Ecc) in patients with various degrees of renal function. METHODS: Pathogens were isolated from China's blood bacterial resistant investigation network. The dosing regimens of imipenem, cefepime and piperacillin were simulated with intermittent infusion and extended infusion. Monte Carlo simulation was performed to calculate the probability of target attainment and a cumulative fraction of response (CFR) against BSIs-Kae/Ecc. RESULTS: In total, 203 BSIs-Kae, and 785 BSIs-Ecc were isolated from the surveillance network. Imipenem showed the highest in vitro activity against BSIs-Kae/Ecc, followed by cefepime (85%) and piperacillin/tazobactam (70-80%). The MIC90 values of imipenem, cefepime and piperacillin/tazobactam aginst BSIs-Kae and BSIs-Ecc were 1/1 mg/L, 16/16 mg/L, and 64/128 mg/L, respectively. The simulation results showed imipenem achieved the highest CFRs in patients with normal or decreased renal function, with values of 91-99%, followed by FEP (88-96%), without risk of excessive dosing. However, the intermittent and extended dosing regimens of piperacillin/tazobactam were unlikely to provide adequate exposure for empirical management of BSIs-Kae/Ecc (CFRs, 50-80%), regardless of renal function. Besides, the traditional intermittent piperacillin/tazobactam dosing regimens were highly likely to contribute to suboptimal therapeutic exposure when MIC was close to clinical breakpoints. CONCLUSIONS: Cefepime, not piperacillin/tazobactam, can be a reasonable carbapenem-sparing option in empirically treating BSIs-Kae/Ecc.

Optimal Empiric Polymyxin B Treatment of Patients Infected with Gram-Negative Organisms Detected Using a Blood Antimicrobial Surveillance Network in China.

BACKGROUND: Few pharmacodynamics studies to date have evaluated the efficacy and safety of polymyxin B (PMB) in treating patients with bloodstream infections (BSIs) in China. METHODS: Patients with BSIs were identified using an antimicrobial surveillance network, and their pathogens were isolated. Patients were treated with a loading dose of PMB followed by either a weight-based or weight-independent maintenance dose. Monte Carlo simulation was utilized to calculate the probability of target attainment (PTA) and cumulative fraction of response (CFR) against Gram-negative organisms in patients with normal or decreased renal function. RESULTS: A total of 10,066 Gram-negative organisms, including 5500 Escherichia coli (Eco), 2519 Klebsiella pneumoniae (Kpn), 501 Acinetobacter baumannii (Aba), were isolated from patients with BSIs. Although these strains were highly resistant to carbapenem, they remained susceptible to PMB. Among patients with renal impairment (mean CrCL, 42 mL/min), a PMB 2.5 mg/kg loading dose followed by a maintenance dose of 60 mg q12h reached ≥90% PTA against isolates with an MIC of 2 mg/L, with a low risk of toxicity. Among patients with normal renal function (mean CrCL, 123 mL/min), all simulated regimens showed PTAs of 25-80%. A weight-based loading dose followed by either a weight-based or weight-independent maintenance dose showed a promising CFR, especially in patients with renal impairment, with CFRs ≥90% against carbapenem-resistant Eco, Kpn, and Aba. Simulated regimens showed a disappointing CFR (<80%) against carbapenem-resistant Pae in patients with normal renal function. Based on the optimal balance of efficacy and toxicity, a fixed maintenance dose of 60 mg q12h among patients with renal impairment yielded a CFR similar to regimens based on total body weight and was associated with a probability of toxicity (12.5%) significantly lower than that of simulations based on total body weight. Among patients with normal renal function, a weight-based maintenance dose of 1.25 mg/kg q12h achieved a higher CFR than a fixed maintenance dose, without significantly increasing toxicity. CONCLUSION: A 2.5 mg/kg loading dose of PMB is optimal, regardless of renal function. A fixed maintenance dose of 60 mg q12h is recommended for empirical treatment of patients with renal impairment infected with Eco, Kpn, and Aba, whereas a weight-based maintenance dose of 1.25 mg/kg is recommended for patients with normal renal function.

Optimal empiric treatment for KPC-2-producing Klebsiella pneumoniae infections in critically ill patients with normal or decreased renal function using Monte Carlo simulation.

BACKGROUND: Limited clinical studies describe the pharmacodynamics of fosfomycin (FOS), tigecycline (TGC) and colistin methanesulfonate (CMS) in combination against KPC-producing Klebsiella pneumoniae (KPC-Kp). Population pharmacokinetic models were used in our study. Monte Carlo simulation was conducted to calculate probability of target attainment (PTA) and cumulative fraction of response (CFR) of each agent alone and in combination against KPC-Kp in patients with normal or decreased renal function. RESULTS: The simulated regimen of FOS 6 g q8h reached ≥90% PTA against a MIC of 64 mg/L in patients with normal renal function. For patients with renal impairment, FOS 4 g q8h could provide sufficient antimicrobial coverage against a MIC of 128 mg/L. And increasing the daily dose could result to the cut-off value to 256 mg/L in decreased renal function. For TGC, conventional dosing regimens failed to reach 90% PTA against a MIC of 2 mg/L. Higher loading and daily doses (TGC 200/400 mg loading doses followed by 100 mg q12h/200 mg q24h) were needed. For CMS, none achieved 90% PTA against a MIC of 2 mg/L in normal renal function. Against KPC-Kp, the regimens of 200/400 mg loading dose followed by 100 q12h /200 mg q24h achieved > 80% CFRs regardless of renal function, followed by CMS 9 million IU loading dose followed by 4.5/3 million IU q12h in combination with FOS 8 g q8h (CFR 75-91%). CONCLUSIONS: The use of a loading dose and high daily dose of TGC and CMS in combination with FOS can provide sufficient antimicrobial coverage against critically ill patients infected with KPC-Kp.

A retrospective analysis of Pseudomonas aeruginosa bloodstream infections: prevalence, risk factors, and outcome in carbapenem-susceptible and -non-susceptible infections.

Background: Pseudomonas aeruginosa (PA) is a leading cause of nosocomial infections, and carbapenem non-susceptible strains are a major threat to patient safety. Methods: A single center, retrospective comparative analysis of carbapenem-non-susceptible PA (CnSPA) and carbapenem-susceptible PA (CSPA) bloodstream infections (BSIs) was conducted between January 1, 2007, and December 31, 2016. Prevalence and risk factors associated with CnSPA BSIs were examined. Results: The study enrolled 340 patients with PA BSIs; 30.0% (N = 101) of patients had CnSPA. High APACHE II scores (≥15), central venous catheterization, and delayed application of appropriate definitive therapy were independently associated with higher risk of mortality in PA BSIs. Multivariate analysis revealed that respiratory disease and exposure to carbapenems within the previous 90 days to onset of BSI were independent risk factors for acquisition of CnSPA BSIs. Overall all-cause 30-day mortality associated with PA BSIs was 26.8% (91/340). In addition, mortality was higher in patients with CnSPA than in those with CSPA (37.6% vs. 22.2%, respectively; P = 0.003). Corticosteroid therapy and delayed receipt of effective definitive therapy were independent risk factors for death from CnSPA BSIs. Conclusion: Increased incidence of CnSPA BSIs was observed during the study period, with higher mortality seen in patients with these infections. Respiratory disease and exposure to carbapenems were independent risk factors for development of CnSPA BSIs. Appropriate definitive therapy reduced mortality rates. BLBLIs were as effective as carbapenems as a treatment for PA BSIs.

Comparison of Tigecycline or Cefoperazone/Sulbactam therapy for bloodstream infection due to Carbapenem-resistant Acinetobacter baumannii.

Background: We retrospectively analyzed the effect of tigecycline and cefoperazone/sulbactam therapies on the prognosis of patients with carbapenem-resistant Acinetobacter baumannii bloodstream infection (CRAB-BSI). Methods: CRAB-BSI patients receiving tigecycline therapy or cefoperazone/sulbactam therapy between January 2012 and December 2017 was enrolled, and strict exclusion criteria were followed. The 28-day mortality of patients was analyzed. The impact of cefoperazone/sulbactam therapy on prognosis was evaluated using Cox multivariate regression analysis. The 28-day mortality of patients receiving cefoperazone/sulbactam monotherapy and cefoperazone/sulbactam-based combination therapy was also compared. Results: Three hundred forty eight patients with CRAB-BSI were enrolled in the study. Two hundred ten patients were included after applying the exclusion criteria. Of these, 135 patients received tigecycline therapy and 75 patients received cefoperazone/sulbactam therapy. The 28-day mortality of patients in the latter group was, significantly lower than that of the tigecycline group [29.3% vs. 51.9%; P = 0.001]. Cox multivariate regression analysis revealed that cefoperazone/sulbactam therapy exerted a protective effect on the prognosis of patients [hazard ratio 0.566, 95% confidence interval (0.342-0.940); P = 0.028]. Kaplan-Meier survival curve analysis indicated that the 28-day mortality of patients receiving cefoperazone/sulbactam therapy was lower than that of patients receiving cefoperazone/sulbactam monotherapy, but the difference was not significant (22.2% vs. 40%; P = 0.074). However, the mortality of patients receiving cefoperazone/sulbactam with imipenem/cilastatin was significantly lower than that of patients receiving cefoperazone/sulbactam monotherapy (P = 0.048). Conclusions: Patients treated with cefoperazone/sulbactam therapy had a better clinical outcome. The mortality of patients receiving cefoperazone/sulbactam with imipenem/cilastatin seems to be the lowest.

Study protocol for One Health data collections, analyses and intervention of the Sino-Swedish integrated multisectoral partnership for antibiotic resistance containment (IMPACT).

INTRODUCTION: To effectively minimise the emergence and dissemination of antibiotic resistant bacteria, a holistic One Health approach is called for. The Sino-Swedish Integrated Multisectoral Partnership for Antibiotic Resistance Containment is a cross-sectoral and integrated project on antibiotic resistance, conducted in Shandong Province in China. This paper outlines the overall study protocol for the project. To our knowledge, this is the first research programme aiming to take a true holistic approach across multiple sectors simultaneously in China, and the first to incorporate both antibiotic use and infection prevention and control in addition to antibiotic resistance patterns. The project aims to address gaps in current knowledge and seeks to improve the situation through a system-wide intervention. By using a One Health approach we can address important research questions that individual discipline investigations are unable to. The results obtained should thus more closely reflect the world in which human health, animal health and the environment are inextricably and intimately interlinked. METHODS AND ANALYSIS: Both quantitative and qualitative studies are included for households from 12 villages, their surrounding environment and a tertiary care hospital in a nearby town. The studies include analyses of antibiotic consumption for humans and pigs; qualitative and quantitative data on perceptions, knowledge and attitudes; faecal carriage of extended spectrum ß-lactamase and carbapenemase-producing Enterobacteriaceae from pigs and humans, and occurrence in household drinking water, surface water, waste water and clinical bacterial isolates from the hospital. Carriage of methicillin-resistant Staphylococcus aureus in humans, household pigs and clinical bacterial isolates is also investigated. Furthermore, potential inter-relationships between these sources are analysed. A multifaceted One Health intervention is designed and implemented in 6 of the 12 villages. Repeated and continuous data collections take place over 2 years, where the repeated data collection is performed after 1 year of intervention. Comparisons are made between intervention and control villages, before and after the intervention. ETHICS: Ethics approval was obtained from the first Affiliated Hospital, College of Medicine, Zhejiang University, China, reference number 2015#185 and 2015#283.

Effects of calcium Ionophore A23187 on the apoptosis of hepatic stellate cells stimulated by transforming growth factor-ß1.

BACKGROUND: Our previous study showed that during in vitro experiments changes in calcium concentration were associated with apoptosis. We presumed that the calcium ion might play a role as intermediate messenger for apoptosis-related genes. No such evidence has been reported in the literature. Here, we investigate the effect of calcium ionophore A23187 on the apoptosis of rat hepatic stellate cells (HSCs) stimulated by transforming growth factor-ß1 (TGF-ß1) to explore the mechanism of apoptosis through the endoplasmic reticulum stress pathway. METHODS: The apoptotic rate was determined using flow cytometry. The changes in Ca2+ level in HSCs were examined with laser confocal microscopy. The expressions of caspase-12 GRP78 and caspase-9 were assayed via western blot. RESULTS: The respective apoptosis rates for the blank group, the TGF-ß1 group and the TGF-ß1 + low, medium and high dose calcium ionophore A23187 groups were 3.40 ± 0.10%, 1.76 ± 0.12%, 5.86 ± 0.31%, 11.20 ± 0.48% and 15.08 ± 0.75%, with significant differences between the groups (p < 0.05). The concentration of Ca2+and the expression of the GRP78, caspase-9 and caspase-12 proteins significantly increased with increasing calcium ionophore A23187 doses (p < 0.05). CONCLUSION: Calcium ionophore A23187 increased intracellular Ca2+ and activated endoplasmic reticulum stress, which promoted HSC apoptosis.

Epidemiology and characteristics of antimicrobial resistance in China.

A comprehensive surveillance system for bacterial resistance in tertiary hospitals has been established in China that involves tertiary hospitals in distinct regions nationwide, enabling the collection of a large amount of antimicrobial surveillance data. Antimicrobial resistance in China has become a serious healthcare problem, with high resistance rates of most common bacteria to clinically important antimicrobial agents. Methicillin-resistant S. aureus, ESBL-producing Enterobacteriaceae and carbapenem-resistant Acinetobacter baumannii represent more than 50% of microbial isolates. Additionally, bacterial resistance to fluoroquinolones, macrolides and third-generation cephalosporins is of serious concern. The molecular epidemiology and resistance mechanisms of the antimicrobial strains in China exhibited regional specificity, as well as the influence of dissemination of international clonal complexes. The molecular characteristics of MRSA, ESBL- and carbapenemase-producing Enterobacteriaceae, and macrolide-resistant gram-positive Streptococci in China were significantly different from those in other countries and regions, while S. pneumoniae serotypes appear to have been affected by the global spread of prevalent clones in other parts of the world. Moreover, important antimicrobial resistant bacteria such as community-acquired-MRSA, multidrug-resistant P. aeruginosa and extensive-resistant A. baumannii, and the antimicrobial resistance in primary healthcare and outpatient setting should be intensely monitored and investigated in the future.

Intervention study of the association of antibiotic utilization measures with control of extended-spectrum beta-lactamase (ESBL)-producing bacteria.

We investigated the effects of replacing third-/fourth-generation cephalosporins with piperacillin-tazobactam on the rate of acquisition of extended spectrum beta-lactamase-producing Klebsiella pneumoniae and Escherichia coli by patients hospitalized in a Department of Respiratory Medicine. This 9-month, prospective, non-controlled, intervention study comprised two phases: a 3-month pre-intervention phase (Phase I) and a 6-month intervention phase (Phase II), during which the use of third-/forth-generation cephalosporins was restricted and replaced by piperacillin-tazobactam. Rectal swabs were obtained within 24 h after admission (baseline screening), weekly, and 48 h before discharge during Phase I and the last 3 months of Phase II (Phase IIb). Swabs were tested for E. coli and K. pneumoniae, and extended spectrum beta-lactamase production was detected with the double disc test. Use of third/fourth-generation cephalosporins decreased by 63.0% and 100%, respectively; while the use of piperacillin-tazobactam increased by 28-fold. The rate of acquisition of extended spectrum beta-lactamase-producing E. coli and K. pneumoniae together in rectal swab specimens decreased in Phase IIb as compared with Phase I (19.5% vs 29.5%). Few rectal swab specimens were positive for extended spectrum beta-lactamases-producing K. pneumoniae, and no substantial decrease in the rate of its acquisition was observed.

Design and evaluation of sifuvirtide, a novel HIV-1 fusion inhibitor.

Enfuvirtide (T20) is the first and only HIV-1 fusion inhibitor approved for clinical use, but it can easily induce drug resistance limiting its practical application. A novel anti-HIV peptide, termed sifuvirtide, was designed based on the three-dimensional structure of the HIV-1 gp41 fusogenic core conformation. Here we report its in vitro anti-HIV potency, its mechanism of action, as well as the results from Phase Ia clinical studies. We demonstrated that sifuvirtide inhibited HIV-1-mediated cell-cell fusion in a dose-dependent manner and exhibited high potency against infections by a wide range of primary and laboratory-adapted HIV-1 isolates from multiple genotypes with R5 or X4 phenotypes. Notably, sifuvirtide was also highly effective against T20-resistant strains. Unlike T20, sifuvirtide could efficiently block six-helix bundle formation in a dominant negative fashion. These results suggest that sifuvirtide has a different mechanism of action from that of T20. Phase Ia clinical studies of sifuvirtide (FS0101) in 60 healthy individuals demonstrated good safety, tolerability, and pharmacokinetic profiles. A single dose regimen (5, 10, 20, 30, and 40 mg) by subcutaneous injection once daily at abdominal sites was well tolerated without serious adverse events. Pharmacokinetic studies of single and multiple administration of sifuvirtide showed that its decay half-lives were 20.0 +/- 8.6 h and 26.0 +/- 7.9 h, respectively. In summary, sifuvirtide has potential to become an ideal fusion inhibitor for treatment of HIV/AIDS patients, including those with HIV-1 strains resistant to T20.

Effects of drug serum of anti-fibrosis I herbal compound on calcium in hepatic stellate cell and its molecular mechanism.

AIM: To investigate the effects of anti-fibrosis I herbal compound on intracellular Ca(2+) in activated hepatic stellate cell (HSC) and to try to survey its molecular mechanism in treatment and prevention of hepatic fibrosis and portal hypertension. METHODS: The activated HSC line was plated on small glass cover slips in 24 wells culture dishes at a density of 5X10(6) /mL, and incubated in RPMI-1640 media for 24 h. After the cells were loaded with Fluo-3/AM, intracellular Ca(2+) was measured with laser scanning confocal microscopy (LSCM). The dynamic changes of intracellular Ca(2+), stimulated by carbon tetrachloride, TGF-beta(1) antibody and the drug serum of anti-fibrosis I herbal compound and under orthogonal design were determined by LSCM. The effect of anti-fibrosis I herbal compound on intracellular Ca(2+) was observed before and after the addition of TGF-(1) antibody. RESULTS: The intracellular Ca(2+) were significantly different in different dosage of carbon tetrachloride anti-fibrosis I formula drug serum, TGF-beta(1) antibody and different turn of these substance, but their interval time between CCl(4) and TGF-beta(1) antibody, CCl(4) and anti-fibrosis I drug serum had no influence on intracellular Ca(2+). The result showed intracellular Ca(2+) wasn't significantly different between rat serum without anti-fibrosis I and untreated group. After carbon tetrachloride stimulation, intracellular Ca(2+) of activated HSC increased significantly when the dosage of CCl(4) from 5 to 15 mmol/L, however, decreased significantly after stimulation by 5-20 microg/mL TGF-beta(1) antibody or 5-20 mL/L drug serum. Moreover, before and after the addition of TGF-beta(1) antibody, intracellular Ca(2+) was significantly different. These results suggested that the molecular mechanism was independent of blocking TGF-beta(1) effects. CONCLUSION: Anti-fibrosis I herbal compound may treat hepatic fibrosis and decrease portal hypertension by inhibiting activated HSC contractility through decrease of intracellular Ca(2+).

Therapeutic effects and molecular mechanisms of anti-fibrosis herbs and selenium on rats with hepatic fibrosis.

AIM: To study the therapeutic effects of anti-fibrosis herbs and selenium on hepatic fibrosis induced by carbon tetrachloride (CCl(4)) in rats and the underlining molecular mechanisms. METHODS: Fifty-three Wistar rats were randomly divided into: normal control group, model control group, colchicine group, anti-fibrosis herbs group (AF group) and anti-fibrosis herbs plus selenium group (AS group). The last four groups were administered with CCl(4) at the beginning of experiment to induce hepatic fibrosis. Then colchicine, anti-fibrosis herbs and selenium were used to treat them. The normal control group and the model control group were given normal saline at the same time. At the end of the 6(th) week, rats in each group were sacrificed. Blood and tissue specimens were taken. Serum indicators (ALT, AST, HA, LN) were determined and histopathological changes were graded. Lymphocyte CD(4) and CD(8) were examined by flow cytometry. Expression of TGF-beta(1) and NF-kappaB was detected by immunohistochemistry and expression of TGF-beta(1) mRNA was detected by semi-quantified RT-PCR. RESULTS: Histological grading showed much a smaller degree of hepatic fibrogenesis in AS group and AF group than that in colchicine group and model control group. The serum content of ALT, AST, HA and LN in AF group and AS group were significantly lower than that in colchicine group (ALT: 65.8+/-26.5, 67.3+/-18.4 and 96.2+/-20.9 in AF, AS and colchicine groups respectively; AST: 150.8+/-34.0, 154.6+/-27.3 and 215.8+/-24.6 respectively; HA: 228+/-83, 216+/-58 and 416+/-135 respectively; LN: 85.9+/-15.0, 80.6+/-18.6 and 106.3+/-14.2 respectively) (P<0.05). The level of CD(4) and CD(4)/CD(8) ratio in AF group and AS group was significantly higher that those in cochicine group (CD(4): 50.8+/-3.8, 52.6+/-3.4 and 40.2+/-2.1 in AF, AS and colchicine groups respectively; CD(4)/CD(8) ratio: 1.45, 1.46 and 1.26, respectively (P<0.05). The expression level of NF-kappaB and TGF-beta(1) in the liver tissues of AF and AS treatment groups was markedly decreased compared with that in cochicine group, and TGF-beta(1) mRNA was also markedly decreased (1.07+/-0.31 and 0.98+/-0.14 vs 2.34+/-0.43, P<0.05). CONCLUSION: Anti-fibrosis herbs and selenium have beneficial effects on hepatic fibrosis in rats by enhancing immunity and inhibiting NF-kappaB and TGF-beta(1) expressions.