Diagnosis of spacer-associated infection using quantitative cultures from sonicated antibiotics-loaded spacers: implications for the clinical outcome.

Recent studies showed that a positive microbiological result from sonication of the PMMA spacer was associated with poor outcome of patients, but no quantitative analysis has yet been performed. For this purpose, a prospective analysis of 50 spacers (46 patients) was performed. All spacers were processed according to a previously described protocol, including centrifugation and quantitative culture. Clinical data and outcome were also analysed. A statistical relationship between the results of the cultures and the outcome of the patient was assessed. Sixteen patients were diagnosed with spacer-associated infection. Thirteen out of 50 spacers gave a positive culture. Nine of 13 presented with growth of an organism not isolated in the first-stage cultures, and in 7 out of 13 the organisms count was high (>10,000 CFU/ml). We have detected a significant statistical relationship between poor outcome and positive cultures, high colony counts, isolation of different organisms, positive periprosthetic cultures and spacer-associated infection. The detection in a sonicated, antibiotic-loaded PMMA spacer of organisms other than those isolated in the first surgical samples or high colony counts of any organisms is diagnostic with regard to spacer-associated infection.

Prolonged incubation time does not increase sensitivity for the diagnosis of implant-related infection using samples prepared by sonication of the implants.

We have designed a prospective study to evaluate the usefulness of prolonged incubation of cultures from sonicated orthopaedic implants. During the study period 124 implants from 113 patients were processed (22 osteosynthetic implants, 46 hip prostheses, 54 knee prostheses, and two shoulder prostheses). Of these, 70 patients had clinical infection; 32 had received antibiotics at least seven days before removal of the implant. A total of 54 patients had sonicated samples that produced positive cultures (including four patients without infection). All of them were positive in the first seven days of incubation. No differences were found regarding previous antibiotic treatment when analysing colony counts or days of incubation in the case of a positive result. In our experience, extending incubation of the samples to 14 days does not add more positive results for sonicated orthopaedic implants (hip and knee prosthesis and osteosynthesis implants) compared with a conventional seven-day incubation period.

Risk factors and clinical significance of invasive infections caused by levofloxacin-resistant Streptococcus pneumoniae.

PURPOSE: Fluoroquinolones are recommended for the treatment of pneumonia. The recognition of risk factors for invasive levofloxacin-resistant Streptococcus pneumoniae is important for the design of treatment. METHODS: A retrospective review of cases of invasive pneumococcal infections in adults was undertaken. Epidemiologic data, predisposing factors, clinical variables, and outcome were recorded from previously established protocols. Antimicrobial susceptibility was determined by disk diffusion and the Etest method. Serotyping was performed by latex agglutination and Quellung reaction. RESULTS: Twenty patients with infection caused by levofloxacin-resistant pneumococci [minimum inhibitory concentration (MIC) ≥2 µg/ml] were compared with 102 patients harboring levofloxacin-susceptible strains; 80% of levofloxacin-resistant pneumococci were resistant to ≥3 antibiotics but susceptible to penicillin. Most levofloxacin-resistant strains (80%) belonged to serotype 8. In comparison, only 8% of levofloxacin-susceptible pneumococci belonged to serotype 8. In the multivariate analysis, residence in public shelters [odds ratio (OR) 26.13; p 0.002], previous hospitalization (OR 61.77; p < 0.001), human immunodeficiency virus (HIV) infection (OR 28.14; p = 0.009), and heavy smoking (OR 14.41; p = 0.016) were associated with an increased risk of infection by levofloxacin-resistant pneumococci. Mortality caused by levofloxacin-resistant and levofloxacin-susceptible pneumococci was 35 and 14%, respectively. Among HIV-positive individuals infected with levofloxacin-resistant pneumococci 44% died, but only 12.5% of HIV-positive patients with levofloxacin-susceptible strains died. CONCLUSIONS: We observed the emergence of serotype 8 as the main cause of invasive disease caused by levofloxacin-resistant S. pneumoniae. HIV-positive patients seem to be prone to infection caused by multidrug-resistant serotype 8 and have a high mortality rate.

Sonication of intramedullary nails: clinically-related infection and contamination.

BACKGROUND AND AIM: Sonication is currently considered the best procedure for microbiological diagnosis of implant-related osteoarticular infection, but studies in nail-related infections are lacking. The study aim was to evaluate implant sonication after intramedullary nail explantation, and relate it to microbiological cultures and clinical outcome. PATIENTS AND METHODS: A study was performed in two University Hospitals from the same city. Thirty-one patients with implanted nails were prospectively included, whether with clinical infection (8 cases) or without (23 cases). Retrieved nails underwent sonication according a previously published protocol. The clinical and microbiological outcome patient was related to the presence of microorganisms in the retrieved implant. RESULTS: Positive results appeared in 15/31 patients (9 with polymicrobial infections) almost doubling those clinically infected cases. The most commonly isolated organisms were Staphylococcus epidermidis (19.2 %) and Staphylococcus aureus (15.4 %). A significant relationship was found between the presence of positive cultures and previous local superficial infection (p=0.019). The presence of usual pathogens was significantly related to clinical infection (p=0.005) or local superficial infection (p=0.032). All patients with positive cultures showed pain diminution or absence of pain after nail removal (15/15), but this only occurred in 8 (out of 16) patients with negative cultures. CONCLUSIONS: In patients with previously diagnosed infection or local superficial infection, study of the hardware is mandatory. In cases where pain or patient discomfort is observed, nail sonication can help diagnose the implant colonization with potential pathogens that might require specific treatment to improve the final outcome.

Detection of lfrA and tap efflux pump genes among clinical isolates of non-pigmented rapidly growing mycobacteria.

This study was performed to detect LfrA and Tap efflux pumps among clinical isolates of non-pigmented rapidly growing mycobacteria (NPRGM). Gene detection was performed using polymerase chain reaction (PCR) with specific primers designed for each gene. Susceptibility of the strains to doxycycline, tigecycline and ciprofloxacin was analysed using the broth microdilution reference technique. In total, 166 clinical isolates were included in the study. The lfrA gene was detected in four strains (2.4%), comprising two strains of Mycobacterium chelonae (6.7% of this species), one Mycobacterium fortuitum (1.1%) and one Mycobacterium mucogenicum (14.3%). The tap gene was detected in 109 strains (65.7%), comprising 3 Mycobacterium abscessus (33.3%), 12 M. chelonae (40%), 75 M. fortuitum (84.3%), 2 Mycobacterium mageritense (40%), 15 Mycobacterium peregrinum (68.2%), 1 Mycobacterium alvei and 1 Mycobacterium porcinum; no strains of M. mucogenicum were tap-positive. No differences between tap-positive and -negative strains were observed for resistance to doxycycline (Fisher's exact test, P=0.055). lfrA is rare among clinical isolates of NPRGM, whilst tap is found more commonly. No correlation was detected between the presence of the efflux pumps and resistance to quinolones or tetracyclines.

Biofilm development by clinical strains of non-pigmented rapidly growing mycobacteria.

The relationship between clinical significance of non-pigmented, rapidly growing mycobacteria (NPRGM), in vitro biofilm development and sliding motility was evaluated in this study. One hundred and sixty-eight clinical strains of NPRGM were included. Forty-one of these were clinically significant isolates. Biofilm was formed by 123 strains. Seventy-six biofilm-positive and 25 biofilm-negative strains showed sliding motility. There was a relationship between clinical significance and biofilm development (p <0.000,001), sliding motility (p 0.0037) and species (p <0.000,001). No relationship was found between motility and biofilm development. The ability to develop biofilm is a characteristic that can have importance in the development of infections caused by NPRGM.

Prevalence of erm methylase genes in clinical isolates of non-pigmented, rapidly growing mycobacteria.

The aim of this study was to determine the frequency of erm genes coding for macrolide resistance among clinical isolates of non-pigmented rapidly growing mycobacteria (NPRGM) and to evaluate their importance in phenotypic resistance. Broth microdilution susceptibility testing was performed for all NPRGM tested. A PCR assay with consensus primers was used to evaluate the presence of erm genes among the 167 clinical isolates studied, which belonged to nine species of NPRGM; erm genes were detected in all nine species and 109 strains were erm-positive. The highest percentage of erm-positive isolates was found among Mycobacterium mageritense (100%) and the lowest among Mycobacterium mucogenicum (14%). The MICs of macrolides were found to be lower for erm-negative isolates (MIC(90): 2 mg/L) than for erm-positive isolates (MIC(90): 16 mg/L), although in some cases high MICs were found for erm-negative isolates. The finding that erm methylases are present in the majority of the species of NPRGM analysed in this study is not in agreement with conventional susceptibility studies. It therefore appears necessary to use a combination therapy to treat infections caused by NPRGM.

In vitro activity of tigecycline and 10 other antimicrobials against clinical isolates of the genus Corynebacterium.

We studied the in vitro activity of tigecycline and 10 other commonly used antibiotics against 135 clinical isolates of non-diphtheria Corynebacterium spp. using the Etest system. Tigecycline minimum inhibitory concentrations for 50% and 90% of the organisms (MIC(50) and MIC(90) values, respectively, in mg/L) were: Corynebacterium urealyticum, 0.094 and 0.125; Corynebacterium amycolatum, 0.125 and 2; Corynebacterium jeikeium, 0.094 and 0.75; Corynebacterium coyleae, 0.064 and 0.064; Corynebacterium striatum, 0.064 and 1; Corynebacterium aurimucosum, 0.094 and 0.125; and Corynebacterium afermentans, 0.064 and 0.094. The activities of all other antimicrobials were variable, with good activity of glycopeptides, linezolid, quinupristin/dalfopristin and daptomycin and with resistance to macrolides in a high number of isolates. Tigecycline is a good alternative for the therapy of infections caused by non-diphtheria corynebacteria.

In vitro activities of tigecycline and 10 other antimicrobials against nonpigmented rapidly growing mycobacteria.

We evaluated the in vitro activities of tigecycline and 10 other antibiotics against clinical isolates of nonpigmented rapidly growing mycobacteria. Fifteen collection strains and 165 clinical isolates were included in the study. Tigecycline showed the highest activity among all antibiotics studied: all the strains were inhibited by 1 mg/liter.

Epidemiology of infections due to nonpigmented rapidly growing mycobacteria diagnosed in an urban area.

The objective was to determine the incidence, clinical significance, and epidemiology of the isolates of nonpigmented, rapidly growing mycobacteria (NPRGM) in Madrid, Spain. Patients with new isolates of NPRGM during 2005 were selected prospectively for review of clinical charts. Clinical significance was analyzed according internationally accepted criteria. Randomly amplified polymorphic DNA (RAPD) was used for the genotyping of the isolates. NPRGM were identified in 70 patients (1.51 cases/100,000 inhabitants). The species were M. abscessus (in 5 patients), M. chelonae (in 9), M. fortuitum (in 40), M. peregrinum (in 9), M. mageritense (in 5), M. mucogenicum (in 2), and M. alvei (in 1 patient). The isolates were clinically significant in 17 cases (24.3%, 0.39 cases/100,000 inhabitants): in 4 cases of M. abscessus, in 5 of M. chelonae, and in 9 of M. fortuitum. Only 10.7% of the respiratory isolates were significant, whereas 75% of the nonrespiratory ones were significant (p < 0.001). RAPD analysis showed no relationship among the 74 strains available for the study. No characteristic resistance pattern could be found, although 4 strains appeared to be resistant to amikacin. Significant isolates were mainly nonrespiratory ones. The most significant species was M. abscessus. No relationship between the various isolates was detected, ruling out interhuman transmission between these cases.

The isolation of Corynebacterium coyleae from clinical samples: clinical and microbiological data.

Fourteen Corynebacterium coyleae isolates were recovered from 12 in-patients during a 5-years period. In six patients, the isolates were considered as clinically significant, three definite (sepsis), two probable (sepsis and soft tissue infection), and one possible (post-transfusional bacteremia). In the remaining 6 patients (all neonatal bacteremias), there was not enough data for considering the isolates as clinical significant. API Coryne identified all isolates as C. jeikeium, while Biolog GP2 correctly identified 7 out of the 14 isolates. Definitive identification was achieved in all isolates by the sequencing of a fragment of 724 to 1423 pb of 16S rDNA. Successive isolations from two patients presented identical random amplified polymorphic DNA (RAPD) profiles. All of the isolates were in-vitro-sensitive to beta-lactams, gentamicin, rifampin, tetracycline, vancomycin, linezolid, and resistant to clindamycin. Resistance to erythromycin occurred in 83.3% of isolates, all of them presenting phenotype cMLS and harboring the gene ermX.

Clinical significance and epidemiology of non-pigmented rapidly growing mycobacteria in a university hospital.

OBJECTIVES: To study the clinical significance and epidemiology of Non-pigmented rapidly growing mycobacteria (NPRGM) during a 13-year period. METHODS: We performed a retrospective study of patients with isolates of NPRGM to evaluate their clinical significance. We also analyzed the strains using Randomly Amplified Polymorphic DNA (RAPD) analysis to evaluate the relationship between strains. RESULTS: Between 1990 and 2003, 65 patients had an isolate of NPRGM. Twenty of them were considered significant (19 cases) or doubtful (1 case). Many cases were skin and soft tissue infections. Six cases were foreign-body related. All the patients recovered with antibiotic therapy and removal of the foreign body. All the patients were apparently unrelated, despite 56.9% of the isolates were detected between 1995 and 1997. RAPD analysis was performed on 43 strains, and showed only a cluster of two Mycobacterium chelonae isolates. Both of them were related with contamination of a laboratory reactive, and were considered non-significant. CONCLUSION: In our hospital, almost one-third of the isolates of NPRGM were significant, being this percentage higher for skin and soft tissue isolates. Patients were cured with antibiotic therapy, but the removal of foreign bodies appeared to be necessary for a good outcome. A minor pseudo-outbreak was detected. No predominant strain was detected.

Pseudo-outbreak of Mycobacterium gordonae: usefulness of randomly amplified polymorphic DNA analysis to assess the clonality of the isolates.

Mycobacterium gordonae was detected in 18 of 21 clinical samples processed during the same day from patients with clinical suspicion of tuberculosis. Randomly amplified polymorphic DNA (RAPD) analysis revealed that all the isolates generated an identical pattern with each of the five primers used, and that these patterns were different from those of epidemiologically non-related isolates of M. gordonae. M. gordonae was not detected in the distilled water used for the procedures, and following replacement of the commercial products and sterilisation of home-made reagents, no more isolates belonging to the same clone of M. gordonae were detected.

Antimicrobial susceptibility of Haemophilus influenzae, Haemophilus parainfluenzae and Moraxella catarrhalis isolated from adult patients with respiratory tract infections in four southern European countries. The ARISE project.

Over a 7-month period in 2000-2001, 1213 Haemophilus influenzae, 112 Haemophilus parainfluenzae and 142 Moraxella catarrhalis isolates were recovered from adult patients with respiratory tract infections. Patients were from four southern European countries (Spain, Italy, Portugal and Greece). The antimicrobial susceptibility of the isolates to 11 antibiotics was determined in a central laboratory. The most active drugs on the basis of MICs were levofloxacin, cefditoren, cefotaxime, cefpodoxime and amoxicillin/clavulanate. MICs > or = 2 mg/l for amoxicillin were found in 19.5, 28.6, and 75.4% of H. influenzae, H. parainfluenzae and M. catarrhalis isolates, respectively. Isolates of H. influenzae and H. parainfluenzae with reduced susceptibility or that were fully resistant to amoxicillin/clavulanate, cefuroxime and clarithromycin were detected (0.2-1.8%) as well as M. catarrhalis resistant to clarithromycin (0.7%). Regular surveys of resistance patterns for antimicrobial agents are necessary.