Effect of magnetic ion exchange (MIEX®) on removal of emerging organic contaminants.

In this study, the removal of nine emerging organic contaminants was investigated by using anion exchange resins. The selected compounds were carbamazepine, atrazine, simazine, estrone, bisphenol A, methylparaben, ethylparaben, propylparaben and butylparaben. Two different magnetic anionic exchanger resins were tested: MIEX® DOC and MIEX® GOLD. The optimal resin dose (40 mL/L) and contact time (20 min) had been previously determined. Once these optimum parameters were set, the effect of the initial concentration of contaminants on the removal efficiency of the contaminants by the resins was studied. The study was carried out using mono and multicomponent systems, with distilled water and natural waters, to which contaminants had been previously added, in order to evaluate the competitive and matrix effects. Results showed that the average removal percentages obtained with the MIEX® DOC resin were: 51%, 61%, 68% and 80% for methyl-, ethyl-, propyl-, and butylparaben, respectively. For bisphenol A the result was similar, i.e., 66%, whereas for the rest of the compounds studied, removal efficiencies lower than 15% were obtained. The MIEX® GOLD resin achieved lower elimination rates than the MIEX® DOC resin in all cases.

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.

Expression and characterization of the gD protein of HSV-2 fused to the tetramerization domain of the transcription factor p53.

The highly immunogenic glycoprotein D (gD) of herpes simplex virus type 2 (HSV-2) is a very important element for entry of this virus into host cells. These characteristics have made this protein a very interesting HSV-2 subunit vaccine candidate. Despite efforts to prevent genital herpes using gD-based subunit vaccines, to date, clinical trials using this antigen have failed. Therefore, using a small animal model, we sought to determine if a tetramerized truncated form of gD subunit vaccine, produced by recombinant baculovirus infected insect larvae, would elicit better protection against genital herpes than a monomeric gD-2 subunit vaccine. Three out of 5 mice immunized with the tetramerized antigen produced in a baculovirus expression vector system, survived a lethal challenge with a wild type HSV-2 strain (for more than 3 weeks after challenge). In contrast, all the mice (5) immunized with the truncated protein, produced by the same methodology, died within 2 weeks after challenge. These results suggest that multimerization (increasing the structural complexity) of the truncated gD antigen might be more likely protective than the monomer form. Also the use of an alternative cost-efficient eukaryotic expression system is described.

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.

Emerging infectious endocarditis due to Scedosporium prolificans: a model of therapeutic complexity.

Scedosporium prolificans is an emerging agent for severe infections. Although among the dematiaceous fungi Scedosporium is the most frequently isolated in blood cultures, Scedosporium endocarditis is rarely reported. We show herein a patient with acute leukaemia who developed S. prolificans endocarditis. Twelve cases were found in an extensive review of the English literature. In six cases (46%), there was predisposing heart conditions such as a prosthetic valve or an intracavitary device. Only 4 patients (31%) were immunocompromised hosts with haematologic neoplasia, solid-organ transplantation or acquired immunodeficiency syndrome (AIDS). Exposure to Scedosporium was observed in immunocompetent patients who developed infection while in the community. Scedosporium endocarditis occurred on both sides of the heart. Systemic and pulmonary emboli and other metastatic complications were seen in all of these patients. The overall mortality was 77% and, specifically, all of the immunocompromised hosts and 6 out of 7 patients with mitral or aortic valve endocarditis died. Patients with right-sided endocarditis associated with a removable intracardiac device exhibited a better prognosis. Scedosporium endocarditis, although still rare, is an emerging infection with an ominous prognosis. At the present time, valve replacement or the removal of cardiac devices plus combined antifungal treatment may offer the best possibility of cure.

Construction and properties of a recombinant pseudorabies virus with tetracycline-regulated control of immediate-early gene expression.

A study was carried out to determine whether altering the control of expression of the IE180 gene of pseudorabies virus (PRV), by replacing the IE180 promoter with the tetracycline-responsive promoter (Ptet), affects virus replication and virulence. This PRV-BT90 mutant virus was constructed by complementation and recombination in Hela Tet-Off cells. The virus yield produced by infection of Hela Tet-Off cells with PRV-BT90 was similar to that of the parental virus vBecker2. Viral replication of PRV-BT90 was reduced in Vero cells as reflected by a reduction of virus yield and plating efficiency compared to vBecker2. PRV-BT90 plaque formation in Hela Tet-Off cells was inhibited in the presence of doxycycline, whereas vBecker2 plaque formation was not affected. Subcutaneous infection of mice with the two viruses revealed a LD(50) higher than 10(6) TCID(50) for the PRV-BT90 mutant virus while the LD(50) was 178 TCID(50) for the vBecker2 parental virus.

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.

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.

Immunological properties of a DNA plasmid encoding a chimeric protein of herpes simplex virus type 2 glycoprotein B and glycoprotein D.

A DNA plasmid containing a chimeric sequence encoding both herpes simplex virus type 2 (HSV-2) glycoprotein B (gB) and glycoprotein D (gD) external domains (pcgDB) was used to immunize BALB/c mice against genital HSV-2 infection. To determine the efficacy of this vaccine, groups of mice immunized with the pcgDB plasmid were compared with animals immunized with plasmids corresponding to the individual proteins (pcgBt or pcgDt), administered separately or in combination (pcgBt + pcgDt). We studied the response of the different mouse groups to viral challenge by analyzing clinical disease (vaginitis), serum antibody levels, as well as lymphoproliferative responses and cytokine production by spleen cells. Increased IFN-gamma levels correlated with prolonged survival in mice immunized with the plasmid pcgDB, relative to mice immunized with plasmids coding for the individual proteins alone or in combination. Our results show that immunization with the plasmid encoding the chimeric protein is advantageous over separate proteins. These findings may have important implications for the development of multivalent DNA vaccines against HSV and other complex pathogens.

Antimicrobial resistance among clinical isolates of Streptococcus pneumoniae isolated in four southern European countries (ARISE project) from adult patients: results from the cefditoren surveillance program.

From four southern European countries (Spain, Italy, Portugal, and Greece) 877 Streptococcus pneumoniae isolates were recovered from adult patients with respiratory tract infections between September 2000 and March 2001. The antimicrobial susceptibility to 11 antibiotics was determined in a central laboratory. Penicillin resistance was high in Greece (47.1%) and Spain (25.1%) but much lower in Portugal (7.9%) and Italy (4.8%). On the other hand, erythromycin resistance was high in Italy (38.5%) and Spain (36.2%) with no statistical difference with Greece (29.4%) but reaching significance (p <0.01) with Portugal (15.7%). Resistance to levofloxacin was low (1.5%) but present in Spanish and Italian isolates. Cefditoren, a new cephem antibiotic tested, was the most potent compound (MIC90 = 0.5 microg/ml) followed by levofloxacin and cefotaxime (MIC90 = 1 microg/ml). Given the high rates of penicillin and macrolide resistance reported, there is an evident need for new drugs and continued antimicrobial surveillance of S. pneumoniae.

Nosocomial enterococcal endocarditis: a serious hazard for hospitalized patients with enterococcal bacteraemia.

OBJECTIVES: Enterococci are a major leading cause of infectious endocarditis and also a common cause of hospital-acquired bacteraemia, which is not believed to represent a serious hazard for the endocarditis. The incidence and risk factors for infectious endocarditis in patients with hospital-acquired enterococcal bacteraemia is determined. METHODS: Prospective analysis of 116 patients with enterococcal bacteraemia admitted to medical or surgical wards of a tertiary-care, university affiliated hospital during a period of 5 years. Echocardiography was performed when indicated by clinical criteria. RESULTS: Seventy-five (61.4%) episodes were hospital-acquired and 47 (38.5%) were community-acquired. Most patients had one or more underlying chronic diseases and major abdominal (58.6%) or genitourinary (38.6%) surgery. Seventeen patients (14.6%) developed enterococcal endocarditis. By univariate analysis the risk factors associated with endocarditis were community-acquired infection (P 0.012); monomicrobial bacteraemia (P 0.006); three or more positive blood cultures (P < 0.001); underlying valvulopathy (P < 0.001); presence of a prosthetic valve (P < 0.001) and age (P 0.012). Six patients (8%) developed nosocomial endocarditis. In this group of patients, three or more positive blood cultures (P < 0.01), bacteraemia as a result of Enterococcus faecalis (P 0.007); underlying valvulopathy (P < 0.001) and presence of a prosthetic valve (P < 0.001) were associated with endocarditis. By logistic regression, the presence of underlying valvulopathy and three or more positive blood cultures were associated with endocarditis (OR 21.0; CI 95% 1.65-26.9; P 0.019). CONCLUSIONS: The risk of developing infectious endocarditis in patients with hospital-acquired enterococcal bacteraemia is significant. Patients with underlying valvulopathy and three or more positive blood cultures with E. faecalis are prone to nosocomial enterococcal endocarditis.

A comparison between disk diffusion and microdilution for susceptibility testing of Mycobacterium fortuitum complex.

We evaluated a disk diffusion method using Mueller-Hinton agar for susceptibility testing of Mycobacterium fortuitum complex organisms. Ninety-five strains were tested both by broth microdilution and disk diffusion. Global results showed good correlation for all antimicrobials except for clarithromycin and erythromycin. However, when the results were analyzed according to species, correlation was poor except for a few antimicrobials. The analysis of the resistant/susceptible results was good for all the antimicrobials tested except azithromycin and erythromycin. In conclusion, the disk diffusion technique could be useful as a screening technique for some antibiotics, but the results must be confirmed by using an accepted reference technique.

Mycobacterium tuberculosis bacteremia in a university hospital.

SETTING: Patients with blood cultures positive for Mycobacterium tuberculosis between 1988 and 1999. OBJECTIVE: To study the clinical and microbiological characteristics of patients with tuberculous bacteremia, including data about evolution and management. DESIGN: Retrospective review of the clinical charts and microbiological records of patients with culture-proven tuberculous bacteremia between 1988-1999. RESULTS: During the study period, 19 patients with culture-proven M. tuberculosis bacteremia were detected (1.42 isolates/patient, 4.7% of all patients with blood cultures for mycobacteria). Four patients were non-infected with the human immunodeficiency virus and 15 were HIV-infected. In four patients blood was the only positive sample. Five patients were diagnosed simultaneously with tuberculosis and HIV infection. Only 13 had a temperature higher than 37.5 degrees C. Most patients had symptoms or signs of respiratory tract involvement, and 11 patients died (10 from tuberculosis). The average time for detection of positive blood cultures was 33.25 days for lysis-centrifugation cultures and 26.46 days for BACTEC cultures. The incidence of M. tuberculosis bacteremia remained stable during the study period. CONCLUSIONS: Although blood cultures are useful for definitive diagnosis of disseminated tuberculosis, the long incubation times made them of limited usefulness in the clinical management of patients. Mortality remains high in these patients.