Rogelio Martín

Resistance to Penicillin and Cephalosporin and Mortality from Severe Pneumococcal Pneumonia in Barcelona, Spain

BACKGROUND Penicillin-resistant strains of Streptococcus pneumoniae are now found worldwide, and strains with resistance to cephalosporin are being reported. The appropriate antibiotic therapy for pneumococcal pneumonia due to resistant strains remains controversial. METHODS To examine the effect of resistance to penicillin and cephalosporin on mortality, we conducted a 10-year, prospective study in Barcelona of 504 adults with culture-proved pneumococcal pneumonia. RESULTS Among the 504 patients, 145 (29 percent) had penicillin-resistant strains of S. pneumoniae (minimal inhibitory concentration [MIC] of penicillin G, 0.12 to 4.0 micrograms per milliliter), and 31 patients (6 percent) had cephalosporin-resistant strains (MIC of ceftriaxone or cefotaxime, 1.0 to 4.0 micrograms per milliliter). Mortality was 38 percent in patients with penicillin-resistant strains, as compared with 24 percent in patients with penicillin-sensitive strains (P = 0.001). However, after the exclusion of patients with polymicrobial pneumonia and adjustment for other predictors of mortality, the odds ratio for mortality in patients with penicillin-resistant strains was 1.0 (95 percent confidence interval, 0.5 to 1.9; P = 0.84). Among patients treated with penicillin G or ampicillin, the mortality was 25 percent in the 24 with penicillin-resistant strains and 19 percent in the 126 with penicillin-sensitive strains (P = 0.51). Among patients treated with ceftriaxone or cefotaxime, the mortality was 22 percent in the 59 with penicillin-resistant strains and 25 percent in the 127 with penicillin-sensitive strains (P = 0.64) The frequency of resistance to cephalosporin increased from 2 percent in 1984-1988 to 9 percent in 1989-1993 (P = 0.002). Mortality was 26 percent in patients with cephalosporin-resistant S. pneumoniae and 28 percent in patients with susceptible organisms (P = 0.89). Among patients treated with ceftriaxone or cefotaxime, mortality was 22 percent in the 18 with cephalosporin-resistant strains and 24 percent in the 168 with cephalosporin-sensitive organisms (P = 0.64). CONCLUSIONS Current levels of resistance to penicillin and cephalosporin by S. pneumoniae are not associated with increased mortality in patients with pneumococcal pneumonia. Hence, these antibiotics remain the therapy of choice for this disease.

Epidemiology of Invasive Pneumococcal Disease among Adult Patients in Barcelona Before and After Pediatric 7-Valent Pneumococcal Conjugate Vaccine Introduction, 1997–2007

BACKGROUND A dramatic decrease in the incidence of invasive pneumococcal disease (IPD) was observed among children and adults in the United States after the introduction of the 7-valent pneumococcal conjugate vaccine (PCV7). Little is known about the incidence of IPD after PCV7 licensure in Europe. The objective of this study was to examine changes in the prevalence of IPD among adults in the PCV7 era. METHODS We undertook a prospective study involving adults with IPD who required hospital admission in the southern area of Barcelona, Spain. Three periods were studied: the pre-PCV7 period (1997-2001), the early PCV7 period (2002-2004), and the late PCV7 period (2005-2007). RESULTS A total of 1007 episodes of IPD were observed. Rates of IPD among adults increased from 13.9 to 14.6 episodes per 100,000 population between the pre-PCV7 period and the early PCV7 period (P = .6) and then to 19.55 episodes per 100,000 population in the late PCV7 period (P < .001). The rates of IPD among adults due to non-PCV7 serotypes increased from 8.4 to 9.7 episodes per 100,000 population between the pre-PCV7 period and the early PCV7 period (P = .15) and then to 15.3 episodes per 100,000 population in the late PCV7 period (P < .001); IPD due to PCV7 serotypes decreased from 5.6 to 4.9 episodes per 100,000 population between the pre-PCV7 period and the early PCV7 period (P = .3), then to 4.3 episodes per 100,000 population in the late PCV7 period (P = .056). Among people aged > or = 65 years, IPD due to PCV7 serotypes decreased from 19.5 to 14.6 episodes per 100,000 population between the pre-PCV7 period and the early PCV7 period (P = .13), then to 12.3 episodes per 100,000 population in the late PCV7 period (P = .02). A decrease in the prevalence of antibioticresistant pneumococci in the late PCV7 period was associated with a decrease in the prevalence of multidrugresistant PCV7 clones (Spain(23F)-ST81, Spain(6B)-ST90, and ST88(19F)) and an increase in the prevalence of non-PCV7 antibiotic-susceptible clones (ST306(1), ST191(7F), ST989(12F), and ST433(22F)). CONCLUSIONS Rates of IPD among adults increased in Barcelona in the late PCV7 period, coinciding with a clonal expansion of non-PCV7 serotypes. In contrast, rates of IPD caused by PCV7 serotypes decreased among people aged > or = 65 years, which suggests the development of a herd immunity.

Rapid Detection of Mycobacterium tuberculosis Complex and Rifampin Resistance in Smear-Negative Clinical Samples by Use of an Integrated Real-Time PCR Method

ABSTRACT Sixty-four of 85 (75.3%) smear-negative respiratory (n = 78) and nonrespiratory (n = 7) samples with positive cultures of Mycobacterium tuberculosis complex (MTC) were detected by the GeneXpert system using the Xpert MTB/RIF assay (GX). In addition, GX found rpoB mutations in all six of the rifampin-resistant strains detected. The test was negative in 20 culture-negative and 20 nontuberculous culture-positive samples (100% specificity). GX offers high potential for the diagnosis of tuberculosis due to its capacity for direct detection of MTC, its rapidity, and its simplicity.

In vitro activities of 22 beta-lactam antibiotics against penicillin-resistant and penicillin-susceptible viridans group streptococci isolated from blood.

A total of 410 strains of viridans group streptococci isolated consecutively from blood were tested by the microdilution method for in vitro susceptibility to 22 beta-lactam antibiotics. One hundred thirty-eight strains (33.6%) were resistant to penicillin with a MIC range of 0.25 to 8 micrograms/ml. MICs of all beta-lactam agents tested were higher for penicillin-resistant strains than for susceptible strains. These antibiotics were classified into three groups according to their in vitro activities (MICs at which 50 and 90% of the isolates are inhibited). Beta-Lactams of the first group (these included imipenem, cefpirome, FK-037, cefditoren, cefotaxime, ceftriaxone, and cefepime) showed activities higher than or similar to that of penicillin against penicillin-resistant viridans group streptococci. However, 80% of highly penicillin-resistant Streptococcus mitis organisms required cefotaxime and ceftriaxone MICs of > or = 2 micrograms/ml (range, 2 to 16 micrograms/ml). Beta-Lactams of the second group (cefpodoxime, ampicillin, amoxicillin-clavulanate, piperacillin, and cefuroxime) showed lower activities than penicillin. Finally, antibiotics of the third group (cephalothin, oxacillin, ceftazidime, cefixime, cefaclor, cefetamet, cefadroxil, cephalexin, and ceftibuten) showed poor in vitro activities. Therefore, some of the beta-lactam agents included in the first group could be an acceptable alternative in the treatment of serious infections due to strains highly resistant to penicillin, although clinical experience is needed.

Molecular characterization of macrolide- and multidrug-resistant Streptococcus pyogenes isolated from adult patients in Barcelona, Spain (1993–2008)

OBJECTIVES The increase in erythromycin resistance among Streptococcus pyogenes isolates is a cause for concern. We analysed trends in macrolide resistance, phenotypes, genotypes, resistance determinants and transposons among erythromycin-resistant S. pyogenes isolates collected from adults in a Barcelona hospital (1993-2008). METHODS Antibiotic susceptibility was studied by microdilution. Molecular typing was performed by PFGE, emm typing and multilocus sequence typing (MLST). Macrolide resistance genes and those related to the Tn916 family of transposons were detected by PCR. RESULTS Ninety-nine (18.3%) of 541 isolates were erythromycin resistant. Erythromycin resistance rates progressively increased from 0% (0/24) in 1993-1994 to 34.2% (50/146, P < 0.001) in 2001-2004, then falling to 7.4% (8/108, P = 0.02) in 2007-2008. Sixty-six erythromycin-resistant isolates were available for molecular studies. Of these, 26 had an M phenotype [mef(A)] and 40 had an MLS(B) phenotype [erm(B), n = 33; and erm(TR), n = 7]. Among M-phenotype isolates, the most frequent genotypes (88.5%) were emm4-ST39, emm6-ST382 and emm75-ST49, whereas genotypes emm11-ST403, emm28-ST52 and emm25-ST350 accounted for 72.5% of MLS(B)-phenotype isolates. Twenty-five isolates harboured both erm(B) and tet(M) genes related to the Tn916 family of transposons, Tn6002 being the most frequent. Ten isolates (10.1%) were ciprofloxacin non-susceptible, related to the emm6-ST382 clone with a ParC S79A change. CONCLUSIONS The peak of macrolide resistance rates among S. pyogenes observed in the 2001-2004 period was associated with an increase in the MLS(B) phenotype caused by the spread of emm11-ST403 and emm28-ST52 clones harbouring transposons of the Tn916 family. However, erythromycin resistance rates decreased significantly in the 2007-2008 period.

Effectiveness of an Integrated Real-Time PCR Method for Detection of the Mycobacterium tuberculosis Complex in Smear-Negative Extrapulmonary Samples in an Area of Low Tuberculosis Prevalence

ABSTRACT Early extrapulmonary tuberculosis (EPTB) diagnosis is particularly difficult. Among 108 smear-negative extrapulmonary samples showing a positive culture for Mycobacterium tuberculosis complex (43 body fluids and 65 nonliquid specimens), 63 (58.3%) were positive with the Xpert MTB/RIF assay (GX). GX sensitivity was quite low for samples from sterile locations (especially for pleural fluids: 26.9%) but high for some nonliquid samples, like abscess aspirates (76.5%). In summary, GX may be a useful tool to be considered for EPTB diagnosis.

Relevance of in vitro antimicrobial susceptibility of Brucella melitensis to relapse rate in human brucellosis.

The in vitro susceptibility of Brucella melitensis was examined vis-a-vis the clinical outcome in 75 patients with brucellosis. The initial MICs for Brucella isolates from patients who relapsed and from those who did not were similar. Furthermore, the MICs for isolates from patients whose infections relapsed were no different from those for original isolates. Our results clearly showed that neither initial nor subsequent antibiotic susceptibility plays a role in the likelihood of relapse of patients with brucellosis.

Serotypes, Clones, and Mechanisms of Resistance of Erythromycin-Resistant Streptococcus pneumoniae Isolates Collected in Spain

ABSTRACT The aim of this study was to analyze the distributions of antibiotic susceptibility patterns, serotypes, phenotypes, genotypes, and macrolide resistance genes among 125 nonduplicated erythromycin-resistant Streptococcus pneumoniae clinical isolates collected in a Spanish point prevalence study. The prevalence of resistance to macrolides in this study was 34.7%. Multiresistance (to three or more antimicrobials) was observed in 81.6% of these strains. Among 15 antimicrobials studied, cefotaxime, moxifloxacin, telithromycin, and quinupristin-dalfopristin were the most active drugs. The most frequent serotypes of erythromycin-resistant isolates were 19F (25%), 19A (17%), 6B (12%), 14 (10%), and 23F (10%). Of the 125 strains, 109 (87.2%) showed the MLSB phenotype [103 had the erm(B) gene and 6 had both erm(B) and mef(E) genes]. Sixteen (12.8%) strains showed the M phenotype [14 with mef(E) and 2 with mef(A)]. All isolates were tested by PCR for the presence of the int, xis, tnpR, and tnpA genes associated with conjugative transposons (Tn916 family and Tn917). Positive detection of erm(B), tet(M), int, and xis genes related to the Tn916 family was found in 77.1% of MLSB phenotype strains. In 16 strains, only the tndX, erm(B), and tet(M) genes were detected, suggesting the presence of Tn1116, a transposon recently described for Streptococcus pyogenes. Five clones, namely, Sweden15A-25, clone19F ST87, Spain23F-1, Spain6B-2, and clone19A ST276, accounted for half of the MLSB strains. In conclusion, the majority of erythromycin-resistant pneumococci isolated in Spain had the MLSB phenotype, belonged to multiresistant international clones, and carried the erm(B), tet(M), xis, and int genes, suggesting the spread of transposons of the Tn916 family.

Association of quantitative cytomegalovirus antigenemia with symptomatic infection in solid organ transplant patients

A prospective virologic follow-up of solid organ transplant patients was designed to determine the usefulness of antigenemia and viremia as virologic markers for the diagnosis of cytomegalovirus (CMV) infections, and also for monitoring CMV disease and therapy control. A total of 629 blood samples from 127 patients (60 liver, 47 kidney, and 20 heart transplant recipients) were studied by tube and shell vial cultures, and by antigenemia assay. This later was carried out by an indirect immunofluorescent assay method for formalin-fixed cytospin slides containing 2 x 10(5) leukocytes, using a monoclonal antibody directed against the CMV pp65 antigen. CMV was detected by at least one of the three methods in 238 specimens (37.8%) from a total of 63 patients. The antigenemia assay was positive in 215 (90.3% of positive samples). A total of 94 samples were detected only by this marker, which occurred either in samples with low positive counts (70.2% with antigenemia counts < 10 positive cells/10(5) leukocytes) or in specimens from treated patients. There were 30 episodes of CMV disease in 23 patients. Antigenemia was positive in all these episodes, 27 of them with counts > 20 positive cells/10(5) leukocytes. With this cut-off, positive and negative predictive values for symptomatic CMV infection were 100% and 97.2%, respectively. The antigenemia assay is a rapid, sensitive, specific, and early marker of CMV infection in transplantees. Cultures became negative with antiviral therapy while remaining antigenemia detectable. There was an association between highest quantitative antigenemia test results and clinical symptoms in our patients. In its quantitative version, the assay is useful to detect symptomatic infection and appears to be a helpful tool in managing patients at risk and in guiding antiviral therapy.

Comparative In Vitro Activities of Linezolid, Telithromycin, Clarithromycin, Levofloxacin, Moxifloxacin, and Four Conventional Antimycobacterial Drugs against Mycobacterium kansasii

ABSTRACT Mycobacterium kansasii is one of the most pathogenic and frequent nontuberculous mycobacteria isolated from humans. Patients with adverse drug reactions, resistant isolates, or suboptimal response require alternative treatment regimens. One hundred forty-eight consecutive clinical isolates of M. kansasii were tested for antimicrobial susceptibilities by the BACTEC 460 system (NCCLS) with two different inoculation protocols, one conventional and one alternative. In the alternative protocol, the inoculum 12B vial was incubated until the growth index was between 250 and 500. Four conventional antimycobacterial drugs (isoniazid, rifampin, streptomycin, and ethambutol) were studied with standard critical concentrations. The in vitro activities of linezolid, telithromycin, clarithromycin, levofloxacin, and moxifloxacin were determined by measuring radiometric MICs. All isolates tested were identified as M. kansasii genotype I and were resistant to isoniazid at a concentration of 0.4 μg/ml. One hundred twenty isolates (81.1%) were inhibited by 1 μg of isoniazid per ml. A high level of resistance to isoniazid (>10 μg/ml) was observed in six isolates (4.1%). Only five strains (3.4%) were resistant to rifampin (>1 μg/ml). All isolates studied were susceptible to streptomycin and ethambutol. The MICs at which 90% of the isolates were inhibited (in micrograms per milliliter) were as follows: linezolid, 1 (range, ≤0.25 to 2); telithromycin, >16 (range, 4 to >16); clarithromycin, 0.5 (range, ≤0.03 to 1); levofloxacin, 0.12 (range, 0.12 to 0.25); and moxifloxacin, 0.06 (range, ≤0.06 to 0.12). The susceptibility testing results with both inoculation protocols showed perfect correlation. In conclusion, all M. kansasii isolates showed decreased susceptibility to isoniazid, but resistance to rifampin was infrequent. Quinolones, especially moxifloxacin, were the most active antimicrobial agents tested, followed by clarithromycin. Linezolid also showed good activity against these microorganisms, but telithromycins in vitro activity was poor.