Efflux Pump, the Masked Side of ß-Lactam Resistance in Klebsiella pneumoniae Clinical Isolates

International audienceBACKGROUND: Beta-lactamase production and porin decrease are the well-recognized mechanisms of acquired beta-lactam resistance in Klebsiella pneumoniae isolates. However, such mechanisms proved to be absent in K. pneumoniae isolates that are non susceptible to cefoxitin (FOX) and susceptible to amoxicillin+clavulanic acid in our hospital. Assessing the role of efflux pumps in this beta-lactam phenotype was the aim of this study. METHODOLOGY/FINDINGS: MICs of 9 beta-lactams, including cloxacillin (CLX), and other antibiotic families were tested alone and with an efflux pump inhibitor (EPI), then with both CLX (subinhibitory concentrations) and EPI against 11 unique bacteremia K. pneumoniae isolates displaying the unusual phenotype, and 2 ATCC strains. CLX and EPI-dose dependent effects were studied on 4 representatives strains. CLX MICs significantly decreased when tested with EPI. A similar phenomenon was observed with piperacillin+tazobactam whereas MICs of the other beta-lactams significantly decreased only in the presence of both EPI and CLX. Thus, FOX MICs decreased 128 fold in the K. pneumoniae isolates but also 16 fold in ATCC strain. Restoration of FOX activity was CLX dose-dependent suggesting a competitive relationship between CLX and the other beta-lactams with regard to their efflux. For chloramphenicol, erythromycin and nalidixic acid whose resistance was also due to efflux, adding CLX to EPI did not increase their activity suggesting differences between the efflux process of these molecules and that of beta-lactams. CONCLUSION: This is the first study demonstrating that efflux mechanism plays a key role in the beta-lactam susceptibility of clinical isolates of K. pneumoniae. Such data clearly evidence that the involvement of efflux pumps in beta-lactam resistance is specially underestimated in clinical isolates

Invasive actinomycosis: surrogate marker of a poor prognosis in immunocompromised patients

Objectives: Actinomycosis is a rare disease favored by disruption of the mucosal barrier. In order to investigate the impact of immunosuppression on outcome we analyzed the most severe cases observed in patients hospitalized in three tertiary care centers. Methods: We reviewed all cases of proven invasive actinomycosis occurring over a 12-year period (1997 to 2009) in three teaching hospitals in the Paris area. Results: Thirty-three patients (16 male) were identified as having an invasive actinomycosis requiring hospitalization. The diagnosis was made by microbiological identification in 26 patients, pathological examination in eight patients, and by both methods in one. Twenty patients (61%) were immunocompromised. Actinomycosis localization was abdominal or pelvic in 17 patients, thoracic in 11, cervicofacial in three, and neurological in two. Twenty patients (61%) underwent surgery. All strains were susceptible to amoxicillin. All patients were treated with a beta-lactam antibiotic, for a median length of 82 days. Twenty-eight patients (85%) were considered as cured. Overall mortality at hospital discharge was 21% (7/33). Mortality was higher in immunocompromised patients (7/20; 21%) compared to non-immunocompromised patients (0/13) (p = 0.027). However, six of seven deaths were directly related to the underlying disease. Conclusions: Actinomycosis is a cause of severe infection in immunocompromised patients and a surrogate marker of a poor prognosis in this specific populatio

Early Diagnosis of Extrapulmonary Tuberculosis by a New Procedure Combining Broth Culture and PCR▿

The diagnosis of extrapulmonary tuberculosis is difficult because of the paucibacillary nature of these infections. We developed a culture-enhanced PCR assay combining a preliminary step of broth culture in BacT/Alert MP bottles with the subsequent detection of Mycobacterium tuberculosis using the GenoType Mycobacteria Direct test. First, the procedure was applied to 10-fold-diluted suspensions of M. tuberculosis prepared in vitro. These experiments showed that a 15-day incubation time was required to detect bacilli in the suspension, with the lowest inoculum size yielding a single colony on Lowenstein-Jensen slants. The efficacy of culture-enhanced PCR at day 15 was subsequently evaluated with 225 nonrespiratory specimens from 189 patients with suspected tuberculosis. All these specimens were smear negative, and 31 (13.8%) from 27 patients were culture positive. The result of culture-enhanced PCR at day 15 was consistent with final culture results in all specimens tested. Compared to culture results, the sensitivity, specificity, positive predictive value, and negative predictive value were 100%. Four patients with a negative culture and a negative PCR result were diagnosed as having tuberculosis on the basis of histological findings or therapeutic response. When using a positive diagnosis of tuberculosis as a gold standard, the sensitivity, specificity, positive predictive value, and negative predictive value were 88.6%, 100%, 100%, and 97.9%, respectively. These results indicate that culture-enhanced PCR is a highly sensitive and specific method for the early detection of M. tuberculosis in extrapulmonary specimens

Successful treatment of meningococcal bacteremia using oral doxycycline: A case report

International audienceWe report the case of an 18-year-old immunocompetent man who presented to the hospital with fever, headaches, and arthromyalgia, which progressed to include an erythematous rash. He had a history of a tick bite 72 h earlier. The diagnosis of rickettsial infection was suspected and a course of doxycycline was initiated for a total of 5 days. His evolution was rapidly favorable under treatment, with resolution of the symptoms within 24 h. Blood cultures came back positive for Neisseria meningitidis serotype B, indicating an authentic purpura fulminans. Purpura fulminans is a medical emergency, a syndrome of intravascular thrombosis characterized by a very rapid evolution that requires early recognition and specific treatment. It is commonly described in the young and healthy patient and has high mortality and morbidity. Common bacteria mainly associated with purpura fulminans are Meningococcus spp., Pneumococcus spp., and Staphylococcus spp

Efficacy of ceftazidime-avibactam in various combinations for the treatment of experimental osteomyelitis due to Klebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae

International audienceBackground: : Optimal treatment of carbapenemase-producing Enterobacterales (CPE) bone infections is poorly defined. This study evaluated the efficacy of the novel beta-lactam-beta-lactamase inhibitor—ceftazidime-avibactam (CAZ-AVI)—with different antibiotic combinations in an experimental model of CPE osteomyelitis. Methods: : KPC-99YC is a clinical strain of Klebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae with intermediate susceptibility to meropenem (MIC 4 mg/L), gentamicin (MIC 0.25 mg/L), colistin (MIC 0.25 mg/L), fosfomycin (MIC 4 mg/L) and ceftazidime-avibactam (MIC 1 mg/L). Time-kill curves were performed at 4x MIC. Osteomyelitis was induced in rabbits by tibial injection of 2×108 CFU of KPC-99YC. Six groups started treatment 14 days later for 7 days: control, colistin, CAZ-AVI, CAZ-AVI plus gentamicin, CAZ-AVI plus colistin and CAZ-AVI plus fosfomycin. Antibiotic dosages were selected to simulate plasma concentrations obtained in humans. Treatment was evaluated according to bone cultures quantified in log10 CFU. Results: : In vitro, CAZ-AVI plus colistin or gentamicin were rapidly bactericidal in contrast with CAZ-AVI plus fosfomycin. In vivo, compared with controls, colistin alone (P = 0.045) and CAZ-AVI alone or in combination significantly lowered bone bacterial counts (P < 0.001). Bone sterilisation was achieved in 67% and 100% of animals with combinations of CAZ-AVI plus colistin or gentamicin (P = 0.001 and P < 0.001, respectively) whereas other treatments were no different from controls. CAZ-AVI plus gentamicin provided greater bone bacterial reduction than CAZ-AVI plus colistin (P = 0.033). No CAZ-AVI-resistant strains emerged in treated rabbits, regardless of combination. Conclusions: : CAZ-AVI plus gentamicin was the best effective combination therapy. Combinations with CAZ-AVI appear to be a promising treatment of KPC-producing Klebsiella pneumoniae osteomyelitis

Efficacy of ceftazidime-avibactam in various combinations for the treatment of experimental osteomyelitis due to Klebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae

International audienceBackground: : Optimal treatment of carbapenemase-producing Enterobacterales (CPE) bone infections is poorly defined. This study evaluated the efficacy of the novel beta-lactam-beta-lactamase inhibitor—ceftazidime-avibactam (CAZ-AVI)—with different antibiotic combinations in an experimental model of CPE osteomyelitis. Methods: : KPC-99YC is a clinical strain of Klebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae with intermediate susceptibility to meropenem (MIC 4 mg/L), gentamicin (MIC 0.25 mg/L), colistin (MIC 0.25 mg/L), fosfomycin (MIC 4 mg/L) and ceftazidime-avibactam (MIC 1 mg/L). Time-kill curves were performed at 4x MIC. Osteomyelitis was induced in rabbits by tibial injection of 2×108 CFU of KPC-99YC. Six groups started treatment 14 days later for 7 days: control, colistin, CAZ-AVI, CAZ-AVI plus gentamicin, CAZ-AVI plus colistin and CAZ-AVI plus fosfomycin. Antibiotic dosages were selected to simulate plasma concentrations obtained in humans. Treatment was evaluated according to bone cultures quantified in log10 CFU. Results: : In vitro, CAZ-AVI plus colistin or gentamicin were rapidly bactericidal in contrast with CAZ-AVI plus fosfomycin. In vivo, compared with controls, colistin alone (P = 0.045) and CAZ-AVI alone or in combination significantly lowered bone bacterial counts (P < 0.001). Bone sterilisation was achieved in 67% and 100% of animals with combinations of CAZ-AVI plus colistin or gentamicin (P = 0.001 and P < 0.001, respectively) whereas other treatments were no different from controls. CAZ-AVI plus gentamicin provided greater bone bacterial reduction than CAZ-AVI plus colistin (P = 0.033). No CAZ-AVI-resistant strains emerged in treated rabbits, regardless of combination. Conclusions: : CAZ-AVI plus gentamicin was the best effective combination therapy. Combinations with CAZ-AVI appear to be a promising treatment of KPC-producing Klebsiella pneumoniae osteomyelitis

Efficacy of ceftazidime-avibactam in various combinations for the treatment of experimental osteomyelitis due to Klebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae

International audienceBackground: : Optimal treatment of carbapenemase-producing Enterobacterales (CPE) bone infections is poorly defined. This study evaluated the efficacy of the novel beta-lactam-beta-lactamase inhibitor—ceftazidime-avibactam (CAZ-AVI)—with different antibiotic combinations in an experimental model of CPE osteomyelitis. Methods: : KPC-99YC is a clinical strain of Klebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae with intermediate susceptibility to meropenem (MIC 4 mg/L), gentamicin (MIC 0.25 mg/L), colistin (MIC 0.25 mg/L), fosfomycin (MIC 4 mg/L) and ceftazidime-avibactam (MIC 1 mg/L). Time-kill curves were performed at 4x MIC. Osteomyelitis was induced in rabbits by tibial injection of 2×108 CFU of KPC-99YC. Six groups started treatment 14 days later for 7 days: control, colistin, CAZ-AVI, CAZ-AVI plus gentamicin, CAZ-AVI plus colistin and CAZ-AVI plus fosfomycin. Antibiotic dosages were selected to simulate plasma concentrations obtained in humans. Treatment was evaluated according to bone cultures quantified in log10 CFU. Results: : In vitro, CAZ-AVI plus colistin or gentamicin were rapidly bactericidal in contrast with CAZ-AVI plus fosfomycin. In vivo, compared with controls, colistin alone (P = 0.045) and CAZ-AVI alone or in combination significantly lowered bone bacterial counts (P < 0.001). Bone sterilisation was achieved in 67% and 100% of animals with combinations of CAZ-AVI plus colistin or gentamicin (P = 0.001 and P < 0.001, respectively) whereas other treatments were no different from controls. CAZ-AVI plus gentamicin provided greater bone bacterial reduction than CAZ-AVI plus colistin (P = 0.033). No CAZ-AVI-resistant strains emerged in treated rabbits, regardless of combination. Conclusions: : CAZ-AVI plus gentamicin was the best effective combination therapy. Combinations with CAZ-AVI appear to be a promising treatment of KPC-producing Klebsiella pneumoniae osteomyelitis

A mobile DNA laboratory for forensic science adapted to coronavirusSARS-CoV-2 diagnosis

International audienceThe Forensic Science Institute of the French "Gendarmerie Nationale" (IRCGN™) developed in 2015 an ISO 17025 certified mobile DNA laboratory for genetic analyses. This Mobil'DNA laboratory is a fully autonomous and adaptable mobile laboratory to perform genetic analyses in the context of crime scenes, terrorism attacks or disasters.To support the hospital taskforce in Paris during the peak of the COVID-19 epidemic, we adapted this mobile genetic laboratory to perform high-throughput molecular screening for coronavirus SARS-CoV-2 by real-time PCR. We describe the adaptation of this Mobil'DNA lab to assist in Coronavirus SARS-CoV-2 diagnosis

Short Antibiotic Treatment Duration for Osteomyelitis Complicating Pressure Ulcers: A Quasi-experimental Study

International audienceBackground. Osteomyelitis-complicating pressure ulcers are frequent among patients with spinal cord injuries (SCIs), and the optimal management is unknown. In our referral center, the current management is debridement and flap coverage surgeries, followed by a short antibiotic treatment. We aimed to evaluate patients’ outcomes a year after surgery. Methods. We performed a quasi-experimental retrospective before/after study on SCI patients with presumed osteomyelitis associated with perineal pressure ulcers. We included all patients who underwent surgery with debridement and flap covering, followed by effective antibiotic treatment, between May 1, 2016, and October 30, 2020. The effective antimicrobial treatment duration included the 10 days leading up to January 1, 2018 (before period), and the 5 to 7 days after (after period). We also compared the efficacy of 5–7-day vs 10-day antibiotic treatment and performed uni- and multivariable analyses to identify factors associated with failure. Results. Overall, 415 patients were included (77.6% male patients; mean age ± SD, 53.0 ± 14.4 years). Multidrug-resistant organisms (MDROs) were involved in 20.7% of cases. Favorable outcomes were recorded in 69.2% of cases: 117/179 (65.3%) in the 10-day treatment group vs 169/287 (71.9%) in the 5–7-day treatment group (P = .153). The only factor associated with failure in the multivariate analysis was a positive culture from suction drainage (odds ratio, 1.622; 95% CI, 1.005–2.617; P = .046). Effective treatment duration >7 days and intraoperative samples negative for MDROs were not associated with better outcomes (P = .153 and P = .241, respectively). Conclusions. A treatment strategy combining surgical debridement and flap covering, followed by 5 to 7 days of effective antibiotic treatment seems safe