Optimization of colistin dosing regimen for cystic fibrosis patients with chronic Pseudomonas aeruginosa biofilm lung infections.

OBJECTIVE: The present study was performed to explore dosing regimens of colistin in patients of cystic fibrosis (CF) with Pseudomonas aeruginosa chronic biofilm lung infection. METHODS: Ten CF patients were involved. One dose colistimethate sodium (CMS) of 6 MIU (million international units) and 9 MIU were administered by intravenous infusion over 45 and 90 min. Venous blood was collected at different time points after the infusion of CMS. Pharmacokinetic parameters of colistin were calculated. Minimum inhibitory concentration for planktonic P. aeruginosa, minimum biofilm inhibitory concentration and minimum biofilm eradication concentration of P. aeruginosa were determined. Monte Carlo simulation was performed to determine the clinical probability of target attainment of different dosing regimens of colistin in CF patients. RESULTS: For 90 min (6 MIU), 45 min (6 MIU), and 45 min (9 MIU) intravenous infusion of colistin, Cmax was 8.9 ± 1.8, 15 ± 5.5, and 31.7 ± 5.3 µg/mL, respectively; Tmax was 1.2 ± 0.4, 0.7 ± 0.2, and 0.8 ± 0.2 h, respectively; AUCtot were 31 ± 3.8, 34 ± 10, and 135 ± 31mg · h/L, respectively; t1/2 was 2.1 ± 0.4, 2 ± 0.3, and 3.3 ± 0.4 h, respectively. MBIC and MBEC of colistin on biofilms at 24 h period treatment were 16-128 µg/mL for non-mucoid and mucoid biofilms of P. aeruginosa. For 90 min (6 MIU), 45 min (6 MIU) and 45 min iv infusion (9 MIU) with one dose colistin, PTA was 49.8%, 53.8%, 99.4% for planktonic infection, and 11.3%, 14.6%, 65.3%, respectively for biofilm infection. CONCLUSIONS: colistin treatment using 45 min iv infusion is better than 90 min iv infusion in this study. Colistin dosage of 9 MIU is better than 6 MIU on both planktonic and biofilm infections of P. aeruginosa in this study.

OligoG CF-5/20 Disruption of Mucoid Pseudomonas aeruginosa Biofilm in a Murine Lung Infection Model.

Biofilm growth is a universal survival strategy for bacteria, providing an effective and resilient approach for survival in an otherwise hostile environment. In the context of an infection, a biofilm provides resistance and tolerance to host immune defenses and antibiotics, allowing the biofilm population to survive and thrive under conditions that would destroy their planktonic counterparts. Therefore, the disruption of the biofilm is a key step in eradicating persistent bacterial infections, as seen in many types of chronic disease. In these studies, we used both in vitro minimum biofilm eradication concentration (MBEC) assays and an in vivo model of chronic biofilm infection to demonstrate the biofilm-disrupting effects of an alginate oligomer, OligoG CF-5/20. Biofilm infections were established in mice by tracheal instillation of a mucoid clinical isolate of Pseudomonas aeruginosa embedded in alginate polymer beads. The disruption of the biofilm by OligoG CF-5/20 was observed in a dose-dependent manner over 24 h, with up to a 2.5-log reduction in CFU in the infected mouse lungs. Furthermore, in vitro assays showed that 5% OligoG CF-5/20 significantly reduced the MBEC for colistin from 512 µg/ml to 4 µg/ml after 8 h. These findings support the potential for OligoG CF-5/20 as a biofilm disruption agent which may have clinical value in reducing the microbial burden in chronic biofilm infections.

Pharmacokinetics and pharmacodynamics of antibiotics in biofilm infections of Pseudomonas aeruginosa in vitro and in vivo.

Although progress on biofilm research has been obtained during the past decades, the treatment of biofilm infections with antibiotics remains a riddle. The pharmacokinetic (PK) and pharmacodynamic (PD) profiles of an antimicrobial agent provide important information helping to establish an efficient dosing regimen and to minimize the development of antimicrobial tolerance and resistance in biofilm infections. Unfortunately, most previous PK/PD studies of antibiotics have been done on planktonic cells, and extrapolation of the results on biofilms is problematic as bacterial biofilms differ from planktonic grown cells in the growth rate, gene expression, and metabolism. Here, we set up several protocols for the studies of PK/PD of antibiotics in biofilm infections of P. aeruginosa in vitro and in vivo. It should be underlined that none of the protocols in biofilms have yet been certificated for clinical use or proved useful for guidance of antibiotic therapy.

Regulation of pqs quorum sensing via catabolite repression control in Pseudomonas aeruginosa.

Bacteria have evolved a set of regulatory pathways to adapt to the dynamic nutritional environment during the course of infection. However, the underlying mechanism of the regulatory effects by nutritional cues on bacterial pathogenesis is unclear. In the present study, we showed that the Pseudomonas aeruginosa catabolite repression control protein regulates the Pseudomonas quinolone signal quorum sensing, which further controls synthesis of virulence factor pyocyanin, biofilm formation and survival during infection models. Our study suggests that deregulation of the catabolite repression by P. aeruginosa might enhance its fitness during cystic fibrosis infections.

High ß-lactamase levels change the pharmacodynamics of ß-lactam antibiotics in Pseudomonas aeruginosa biofilms.

Resistance to ß-lactam antibiotics is a frequent problem in Pseudomonas aeruginosa lung infection of cystic fibrosis (CF) patients. This resistance is mainly due to the hyperproduction of chromosomally encoded ß-lactamase and biofilm formation. The purpose of this study was to investigate the role of ß-lactamase in the pharmacokinetics (PK) and pharmacodynamics (PD) of ceftazidime and imipenem on P. aeruginosa biofilms. P. aeruginosa PAO1 and its corresponding ß-lactamase-overproducing mutant, PAΔDDh2Dh3, were used in this study. Biofilms of these two strains in flow chambers, microtiter plates, and on alginate beads were treated with different concentrations of ceftazidime and imipenem. The kinetics of antibiotics on the biofilms was investigated in vitro by time-kill methods. Time-dependent killing of ceftazidime was observed in PAO1 biofilms, but concentration-dependent killing activity of ceftazidime was observed for ß-lactamase-overproducing biofilms of P. aeruginosa in all three models. Ceftazidime showed time-dependent killing on planktonic PAO1 and PAΔDDh2Dh3. This difference is probably due to the special distribution and accumulation in the biofilm matrix of ß-lactamase, which can hydrolyze the ß-lactam antibiotics. The PK/PD indices of the AUC/MBIC and C(max)/MBIC (AUC is the area under concentration-time curve, MBIC is the minimal biofilm-inhibitory concentration, and C(max) is the maximum concentration of drug in serum) are probably the best parameters to describe the effect of ceftazidime in ß-lactamase-overproducing P. aeruginosa biofilms. Meanwhile, imipenem showed time-dependent killing on both PAO1 and PAΔDDh2Dh3 biofilms. An inoculum effect of ß-lactams was found for both planktonic and biofilm P. aeruginosa cells. The inoculum effect of ceftazidime for the ß-lactamase-overproducing mutant PAΔDDh2Dh3 biofilms was more obvious than for PAO1 biofilms, with a requirement of higher antibiotic concentration and a longer period of treatment.

In vivo pharmacokinetics/pharmacodynamics of colistin and imipenem in Pseudomonas aeruginosa biofilm infection.

Many Pseudomonas aeruginosa isolates from the airways of patients with cystic fibrosis (CF) are sensitive to antibiotics in susceptibility testing, but eradication of the infection is difficult. The main reason is the biofilm formation in the airways of patients with CF. The pharmacokinetics (PKs) and pharmacodynamics (PDs) of antimicrobials can reliably be used to predict whether antimicrobial regimens will achieve the maximum bactericidal effect against infections. Unfortunately, however, most PK/PD studies of antimicrobials have been done on planktonic cells and very few PK/PD studies have been done on biofilms, partly due to the lack of suitable models in vivo. In the present study, a biofilm lung infection model was developed to provide an objective and quantitative evaluation of the PK/PD profile of antimicrobials. Killing curves were set up to detect the antimicrobial kinetics on planktonic and biofilm P. aeruginosa cells in vivo. Colistin showed concentration-dependent killing, while imipenem showed time-dependent killing on both planktonic and biofilm P. aeruginosa cells in vivo. The parameter best correlated to the elimination of bacteria in lung by colistin was the area under the curve (AUC) versus MIC (AUC/MIC) for planktonic cells or the AUC versus minimal biofilm inhibitory concentration (MBIC; AUC/MBIC) for biofilm cells. The best-correlated parameter for imipenem was the time that the drug concentration was above the MIC for planktonic cells (T(MIC)) or time that the drug concentration was above the MBIC (T(MBIC)) for biofilm cells. However, the AUC/MIC of imipenem showed a better correlation with the efficacy of imipenem for biofilm infections (R(2) = 0.89) than planktonic cell infections (R(2) = 0.38). The postantibiotic effect (PAE) of colistin and imipenem was shorter in biofilm infections than planktonic cell infections in this model.

Polysaccharides serve as scaffold of biofilms formed by mucoid Pseudomonas aeruginosa.

Chronic lung infection by mucoid Pseudomonas aeruginosa is one of the major pathologic features in patients with cystic fibrosis. Mucoid P. aeruginosa is notorious for its biofilm forming capability and resistance to immune attacks. In this study, the roles of extracellular polymeric substances from biofilms formed by mucoid P. aeruginosa were investigated. Alginate is not an essential structure component for mucoid P. aeruginosa biofilms. Genetic studies revealed that Pel and Psl polysaccharides serve as essential scaffold and mediate macrocolony formation in mucoid P. aeruginosa biofilms. The Psl polysaccharide is more important than Pel polysaccharide in mucoid P. aeruginosa biofilm structure maintenance and phagocytosis resistance. The polysaccharides were further found to protect mucoid P. aeruginosa strain from host immune clearance in a mouse model of acute lung infection.

Pharmacokinetics/pharmacodynamics of colistin and imipenem on mucoid and nonmucoid Pseudomonas aeruginosa biofilms.

The time course of activity of colistin and imipenem against mucoid and nonmucoid Pseudomonas aeruginosa growing in a biofilm showed that compared with those for planktonic bacteria, the kinetics of colistin and imipenem retained the concentration- and time-dependent killing, respectively, but higher doses of antibiotics and longer dosing periods were required for biofilm eradication. Biofilms of mucoid P. aeruginosa were more difficult to eradicate than nonmucoid biofilms.

Effect of Meropenem on Biofilms of Pseudomonas aeruginosa lasR/rhlR Mutations in vitro / 中华医院感染学杂志

OBJECTIVE To understand the effects of meropenem on the biofilms of Pseudomonas aeruginosa lasR/rhlR mutations in vitro.METHODS The biofilms of PAEO1 and its lasR,rhlR,and lasR rhlR mutants were incubated in a polyvinyl chloride tube in 0.9% saline for three days,then were immersed in meropenem solution of 26 ?g/ml for 24 hours,and examined by scanning electron microscopy(SEM).RESULTS After three days incubation,PAEO1 biofilms showed a well-developed structure,however,the biofilms of PAEO1 lasR,rhlR,and lasR rhlR mutants were thin and poor developed;after 24 hours in meropenem solution,PAEO1 biofilm become rareness,whereas PAEO1 lasR,rhlR,and lasR rhlR mutants biofilms were almost destructed and only small pieces left.CONCLUSIONS lasR/rhlR Genes play probably an important role in the formation of P.aeruginosa biofilm and in the resistance to meropenem.

Effects of lasR/rhlR Mutation on Biofilm Formation of Pseudomonas aeruginosa in vitro / 中华医院感染学杂志

OBJECTIVE To understand the effects of lasR/rhlR mutations on the biofilm formation of Pseudomonas aeruginosa in vitro.METHODS The biofilms of PAEO1 and lasR,rhlR,and lasR rhlR mutants were incubated in a polyvinyl chloride tube in 0.9% saline and studied by means of scanning electronic microscopy(SEM) after three days′ incubation.RESULTS After 3 day′s incubation,PAEO1 biofilm showed a well-developed structure,whereas thin and poor developed biofilms were seen in lasR/rhlR mutant groups.CONCLUSIONS lasR/rhlR Genes play probably an important role in the formation of P.aeruginosa biofilm in vitro.

Effects of Quorum-sensing Systems on Chronic Pseudomonas aeruginosa Lung Infection in Rats / 中华医院感染学杂志

OBJECTIVE To understand the importance of quorum sensing(QS)system in Pseudomonas aeruginosa(PAE)biofilm infection in lungs,the pathogenic effects of the wild-type P.aeruginosa PAEO1 were compared with QS double mutants PAEO1 lasR rhlR and PAEO1 lasⅠ rhⅡ in vivo.METHODS Rats were challenged intratracheally with alginate embedded PAE strain PAEO1 and the mutants of QS in the concentration of 1?109 colony-forming units per milliliter(CFU/ml).Two weeks post intratracheal challenge with P.aeruginosa,parameters were evaluated.RESULTS Two weeks after challenge,significantly milder microscopic and macroscopic lung pathology(P