Evolution and genetic diversity of the Spain23F-ST81 clone causing adult invasive pneumococcal disease in Barcelona (1990-2012).

OBJECTIVES: We aimed to analyse the clinical epidemiology and genetic diversity of invasive pneumococcal disease (IPD) episodes attributed to the Spain(23F)-ST81 (PMEN1) clone. METHODS: Fifty-eight (2.7%) of 2117 invasive pneumococci isolated from adult patients during the 1990-2012 period shared a PFGE pattern related to the PMEN1 clone. The genotype was confirmed by multilocus sequence typing. The pbp2x, pbp1a, pbp2b and pspA genes were PCR-amplified and sequenced. Polymorphisms in the pspC gene were identified by PCR restriction fragment length polymorphism. The presence of transposons with erythromycin and tetracycline resistance determinants was detected by PCR. RESULTS: The prevalence of the PMEN1 clone increased from 0.8% in 1991 to 6.2% in 2001, and decreased to 0% in 2010-12, concomitant with the introduction of the seven-valent pneumococcal conjugate vaccine for children. A total of 93.1% of patients had pneumonia, meningitis or peritonitis; 87.9% of patients had associated underlying diseases, mainly cancer, chronic obstructive pulmonary disease and diabetes. Two closely related sequence types (STs) (ST81, n = 52; ST85, n = 6) were detected, with different serotypes: 23F (n = 42), 19A (n = 9) and 19F (n = 6). All the isolates were resistant to penicillin, co-trimoxazole and chloramphenicol. All the isolates also shared the same pbp1a allele, whereas multiple alleles of pbp2b, pbp2x, pspA and pspC were detected. Of the isolates, 89.7% were tetracycline resistant and 60.3% (n = 35) were macrolide resistant, and resistance was associated with different Tn916-like transposons. CONCLUSIONS: Adult IPD caused by this clone was mainly detected in patients with underlying conditions, and genetic variability was observed among PMEN1 isolates collected in our area over the past 20 years.

Pneumococci can persistently colonize adult patients with chronic respiratory disease.

Streptococcus pneumoniae plays an important role in causing acute exacerbations in patients with chronic respiratory disease. However, few data are available regarding pneumococcal persistence in adult patients with chronic respiratory diseases. Fifty pneumococci recovered from sputum samples (1995 to 2010) from 13 adult patients with ≥ 3 episodes of acute exacerbation or pneumonia, with the same serotype and pulsed-field gel electrophoresis (PFGE) pattern, were studied. Multilocus sequence typing (MLST) loci, penicillin-binding protein (PBP) genes (pbp2x, pbp1a, pbp2b), and the quinolone-resistant determining regions (QRDRs) of parC, parE, and gyrA were PCR amplified and sequenced. The average time between the first and last episode was 582 days (standard deviation [SD], ± 362). All but two patients received multiple courses of ß-lactam treatment, and all persistent strains were resistant to penicillin; however, the PBP sequences were stable over time apart from one variable nucleotide in pbp2x, observed among pneumococci isolated from three patients. In contrast, 7/11 patients treated with fluoroquinolones had fluoroquinolone-resistant pneumococci. In three patients, the initially fluoroquinolone-susceptible strain developed resistance after fluoroquinolone therapy, and in the remaining four patients, the persistent strain was fluoroquinolone resistant from the first episode. QRDR changes involved in fluoroquinolone resistance were frequently observed in persistent strains after fluoroquinolone treatment; however, the PBP sequences and MLST genotypes of these strains were stable over time.

Antibiotic susceptibility and molecular epidemiology of nasopharyngeal pneumococci from Spanish children.

Nasopharyngeal pneumococci were collected from 635 Spanish children aged 6 months to 6 years attending four primary healthcare centres (n = 276) or two hospital emergency rooms (n = 359); 36% of the children had received >/=1 dose of pneumococcal conjugate vaccine (PCV7). Overall, the carriage rate of Streptococcus pneumoniae was 31%, with no significant differences in carriage rates according to setting. Colonization with vaccine serotypes was significantly associated with the absence of PCV7 immunization (29.4% vs. 5.9%, p <0.001). Forty-seven per cent of all isolates were penicillin- and/or erythromycin-non-susceptible; 13 international antibiotic-resistant clones were represented among non-susceptible pneumococci and were similarly distributed among vaccine and non-vaccine serotypes.

Epidemiology of candidemia: 3-year results from the emerging infections and the epidemiology of Iowa organisms study.

Bloodstream infections due to Candida species cause significant morbidity and mortality. Surveillance for candidemia is necessary to detect trends in species distribution and antifungal resistance. We performed prospective surveillance for candidemia at 16 hospitals in the State of Iowa from 1 July 1998 through 30 June 2001. Using U.S. Census Bureau and Iowa Hospital Association data to estimate a population denominator, we calculated the annual incidence of candidemia in Iowa to be 6.0 per 100,000 of population. Candida albicans was the most common species detected, but 43% of candidemias were due to species other than C. albicans. Overall, only 3% of Candida species were resistant to fluconazole. However, Candida glabrata was the most commonly isolated species other than C. albicans and demonstrated some resistance to azoles (fluconazole MIC at which 90% of the isolates tested are inhibited, 32 microg/ml; 10% resistant, 10% susceptible dose dependent). C. glabrata was more commonly isolated from older patients (P = 0.02) and caused over 25% of candidemias among persons 65 years of age or older. The investigational triazoles posaconazole, ravuconazole, and voriconazole had excellent in vitro activity overall against Candida species. C. albicans is the most important cause of candidemia and remains highly susceptible to available antifungal agents. However, C. glabrata has emerged as an important and potentially antifungal resistant cause of candidemia, particularly among the elderly.

The molecular epidemiology of penicillin-resistant Streptococcus pneumoniae in the United States, 1994-2000.

The genetic relatedness of 672 penicillin-resistant isolates of Streptococcus pneumoniae (PRSP) recovered during national surveillance studies conducted in the United States during the periods of 1994-1995, 1997-1998, and 1999-2000 was determined by use of pulsed-field gel electrophoresis (PFGE). Overall, 104 different PFGE types were elucidated. For all study periods combined, the 12 most prevalent PFGE types included >75% of all isolates, and 5 types were closely related to widespread clones (Spain(23F)-1, France(9V)-3, Spain(6B)-2, Tennessee(23F)-4, and Taiwan(19F)-14). From 1994-1995 to 1999-2000, 3 major PFGE types (not closely related to 16 recognized clones) increased in prevalence. Multidrug resistance was identified among 96%-100% of the isolates in 9 of 12 predominant PFGE types. The prevalence of erythromycin resistance increased within 4 major PFGE types. These observations support the hypothesis that the dominant factor in the emergence of PRSP in the United States during the 1990s has been human-to-human spread of relatively few clonal groups that harbor resistance determinants to multiple classes of antibiotics.

Antimicrobial resistance among clinical isolates of Streptococcus pneumoniae in the United States during 1999--2000, including a comparison of resistance rates since 1994--1995.

A total of 1,531 recent clinical isolates of Streptococcus pneumoniae were collected from 33 medical centers nationwide during the winter of 1999--2000 and characterized at a central laboratory. Of these isolates, 34.2% were penicillin nonsusceptible (MIC > or = 0.12 microg/ml) and 21.5% were high-level resistant (MIC > or = 2 microg/ml). MICs to all beta-lactam antimicrobials increased as penicillin MICs increased. Resistance rates among non-beta-lactam agents were the following: macrolides, 25.2 to 25.7%; clindamycin, 8.9%; tetracycline, 16.3%; chloramphenicol, 8.3%; and trimethoprim-sulfamethoxazole (TMP-SMX), 30.3%. Resistance to non-beta-lactam agents was higher among penicillin-resistant strains than penicillin-susceptible strains; 22.4% of S. pneumoniae were multiresistant. Resistance to vancomycin and quinupristin-dalfopristin was not detected. Resistance to rifampin was 0.1%. Testing of seven fluoroquinolones resulted in the following rank order of in vitro activity: gemifloxacin > sitafloxacin > moxifloxacin > gatifloxacin > levofloxacin = ciprofloxacin > ofloxacin. For 1.4% of strains, ciprofloxacin MICs were > or = 4 microg/ml. The MIC(90)s (MICs at which 90% of isolates were inhibited) of two ketolides were 0.06 microg/ml (ABT773) and 0.12 microg/ml (telithromycin). The MIC(90) of linezolid was 2 microg/ml. Overall, antimicrobial resistance was highest among middle ear fluid and sinus isolates of S. pneumoniae; lowest resistance rates were noted with isolates from cerebrospinal fluid and blood. Resistant isolates were most often recovered from children 0 to 5 years of age and from patients in the southeastern United States. This study represents a continuation of two previous national studies, one in 1994--1995 and the other in 1997--1998. Resistance rates with S. pneumoniae have increased markedly in the United States during the past 5 years. Increases in resistance from 1994--1995 to 1999--2000 for selected antimicrobial agents were as follows: penicillin, 10.6%; erythromycin, 16.1%; tetracycline, 9.0%; TMP-SMX, 9.1%; and chloramphenicol, 4.0%, the increase in multiresistance was 13.3%. Despite awareness and prevention efforts, antimicrobial resistance with S. pneumoniae continues to increase in the United States.

Comparison of results generated by serotyping, pulsed-field restriction analysis, ribotyping, and repetitive-sequence PCR used to characterize penicillin-resistant pneumococci from the United States.

One hundred forty-seven isolates of Streptococcus pneumoniae with high-level penicillin resistance collected during a national surveillance program in the United States were characterized by serotyping, pulsed-field restriction analysis, ribotyping, and repetitive-sequence (BOX element) PCR. The results generated by each method were compared by frequency of association to examine whether relationships existed between the various typing methods and statistically to determine association with the geographic source of the isolate or the age of the patient from whom the isolate was obtained. When the data were examined by pairwise analysis of individual strain classifications produced by each typing method, no statistically significant relationships between strain type, geographic location, or patient age were identified, suggesting that distinct clones of penicillin-resistant S. pneumoniae have been widely distributed throughout the United States. However, we did observed shared expression of two or three typing markers at a high frequency (>50%) among clusters of strains, indicating a certain level of concordance between the various typing methods used to classify penicillin-resistant S. pneumoniae.

Use of penicillin MICs to predict in vitro activity of other beta-lactam antimicrobial agents against Streptococcus pneumoniae.

Linear regression analysis was used to compare penicillin MICs determined with 3,129 recent clinical isolates of Streptococcus pneumoniae to MICs obtained with nine other beta-lactam antimicrobial agents. A strong correlation between penicillin MICs and those obtained with other beta-lactams was demonstrated. It may be possible to test penicillin and use MICs obtained with penicillin to predict MICs of other beta-lactam antimicrobials for Streptococcus pneumoniae.

Antimicrobial-drug use and changes in resistance in Streptococcus pneumoniae.

Resistance of Streptococcus pneumoniae to antimicrobial drugs is increasing. To investigate the relationship between antimicrobial use and susceptibility of S. pneumoniae isolates at 24 U.S. medical centers, we obtained data on outpatient antimicrobial-drug use for the regions surrounding 23 of these centers. We found an association between decreased penicillin susceptibility and use of beta-lactam antimicrobial drugs.

Molecular characterization of the beta-lactamases from clinical isolates of Moraxella (Branhamella) catarrhalis obtained from 24 U.S. medical centers during 1994-1995 and 1997-1998.

The beta-lactamases from 403 Moraxella (Branhamella) catarrhalis clinical isolates obtained during 1994-1995 and 1997-1998 U.S. multicenter surveillance studies were characterized by isoelectric focusing. The overall prevalences of the BRO-1 and BRO-2 enzymes among beta-lactamase-positive isolates were estimated to be 97.5 and 2.5%, respectively. The minimum inhibitory concentrations (MICs) of ampicillin for all BRO-2-producing isolates were

In vitro activity of ABT-773, a new ketolide, against recent clinical isolates of Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis.

The in vitro activity of ABT-773 was evaluated against Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis isolates. ABT-773 was the most active antimicrobial tested against S. pneumoniae. ABT-773 and azithromycin were equivalent in activity against H. influenzae and M. catarrhalis and more active than either clarithromycin or erythromycin.

Resistance among Streptococcus pneumoniae: patterns, mechanisms, interpreting the breakpoints.

Streptococcus pneumoniae is a frequent cause of respiratory tract infections. In the United States and worldwide, antimicrobial resistance of S pneumoniae has complicated the management of infections caused by this organism. In the United States, antimicrobial resistance with S pneumoniae has evolved almost entirely during the 1990s. Resistance currently exists at high rates with beta-lactams, macrolides, tetracyclines, chloramphenicol, and trimethoprim/sulfamethoxazole. Multiresistant strains--strains that are resistant to penicillin plus at least 2 other antimicrobial classes--are also increasing in prevalence. Fluoro-quinolone resistance remains at low levels in the United States. Control of the problem of antimicrobial resistance will require more judicious and appropriate use of antimicrobials, the development of new agents with novel targets of action, and strategies for preventing disease from occurring in the first place. In addition, the pursuit of an understanding of resistance mechanisms and pharmacodynamics as they relate to clinical outcome must be an ongoing effort, and that knowledge must be applied to the development of more effective approaches for the treatment of infections caused by S pneumoniae.

A 1997-1998 national surveillance study: Moraxella catarrhalis and Haemophilus influenzae antimicrobial resistance in 34 US institutions.

From November 1, 1997 to April 30, 1998, 726 Moraxella catarrhalis isolates and 1529 Haemophilus influenzae isolates were obtained from 34 medical centres throughout the United States. Rates of beta-lactamase production were 94.6% among M. catarrhalis and 31.1% among H. influenzae strains. Susceptibility rates of M. catarrhalis isolates to selected antimicrobial agents were greater than 99% for amoxycillin-clavulanate, cefixime, cefpodoxime, cefuroxime, cefaclor, loracarbef, clarithromycin, azithromycin, chloramphenicol and tetracycline, 97.8% for cefprozil, 50.4% for trimethoprim-sulphamethoxazole and 28.1% for ampicillin. Of the antimicrobials tested against H. influenzae, the only agents with susceptibility rates below 96% were loracarbef (87.6%), cefprozil (83.4%), cefaclor (82.7%), trimethoprim-sulphamethoxazole (67.3%) and ampicillin (64.7%). The clarithromycin susceptibility rate was 67.4% but this agent was not tested in the presence of its 14-OH metabolite.

Antimicrobial resistance with Streptococcus pneumoniae in the United States, 1997 98.

From November 1997 to April 1998, 1,601 clinical isolates of Streptococcus pneumoniae were obtained from 34 U.S. medical centers. The overall rate of strains showing resistance to penicillin was 29. 5%, with 17.4% having intermediate resistance. Multidrug resistance, defined as lack of susceptibility to penicillin and at least two other non-ss-lactam classes of antimicrobial drugs, was observed in 16.0% of isolates. Resistance to all 10 ss-lactam drugs examined in this study was directly related to the level of penicillin resistance. Penicillin resistance rates were highest in isolates from middle ear fluid and sinus aspirates of children ambulatory-care settings. Twenty-four of the 34 medical centers in this study had participated in a similar study 3 years before. In 19 of these 24 centers, penicillin resistance rates increased 2.9% to 39.2%. Similar increases were observed with rates of resistance to other antimicrobial drugs.

Prevalence of macrolide resistance mechanisms in Streptococcus pneumoniae isolates from a multicenter antibiotic resistance surveillance study conducted in the United States in 1994-1995.

Two main mechanisms of macrolide resistance have been described in erythromycin-resistant Streptococcus pneumoniae (ERSP): a ribosomal methylase, ErmAM, and a macrolide efflux pump, MefE. In this study, we examined the prevalence of these mechanisms in 114 clinical isolates of ERSP from a 30-center study conducted in the United States between November 1994 and April 1995. The isolates were screened by polymerase chain reaction for the presence of known macrolide resistance genes. Seventy (61%) ERSP contained the macrolide efflux gene (mefE), whereas 36 isolates (32%) contained the biosomal methylase gene (ermAM). Isolates that were ermAM-positive had constitutive macrolide resistance. The minimum inhibitory concentrations (for which 90% of isolates were susceptible) of clarithromycin for the efflux-positive strains were much lower than those for the ermAM-positive strains (4 microg/mL vs. >128 microg/mL, respectively). The efflux mechanism is the predominant form of macrolide resistance in the United States.

Simultaneous respiratory tract colonization by multiple strains of nontypeable haemophilus influenzae in chronic obstructive pulmonary disease: implications for antibiotic therapy.

Nontypeable Haemophilus influenzae often causes exacerbations of chronic obstructive pulmonary disease (COPD), and these exacerbations are frequently treated with oral antibiotics. The goals of this study were to determine the frequency of the simultaneous presence of multiple strains of H. influenzae in sputum and to measure the MICs of antibiotics for the isolates. In a prospective study, adults with COPD were seen monthly. Sputum cultures were obtained, and individual colonies were subjected to genomic DNA typing and MIC determinations. Multiple strains of H. influenzae were present simultaneously in the sputum of 26.3% of adults with COPD. In 64.5% of these, MICs of >/=1 antibiotic varied by >/=4-fold among the strains. Therefore, multiple strains of H. influenzae are frequently present simultaneously in the sputum of adults with COPD, and the antimicrobial susceptibility of different strains in the same sputum sometimes differs.

Assessment of laboratory performance with Streptococcus pneumoniae antimicrobial susceptibility testing in the United States: a report from the College of American Pathologists Microbiology Proficiency Survey Program.

OBJECTIVE: To assess the performance of clinical microbiology laboratories in the United States when conducting in vitro susceptibility tests with Streptococcus pneumoniae. METHODS: The results of a nationwide College of American Pathologists Proficiency Survey test sample, in which susceptibility testing of an isolate of S. pneumoniae was performed, were assessed with respect to precision and accuracy. RESULTS: Wide variability was noted among participating laboratories with both minimum inhibitory concentration procedures and disk diffusion susceptibility tests when both methods were applied to S. pneumoniae. Despite this high degree of variation, categorical interpretive errors were uncommon. Numerous laboratories reported results for antimicrobial agents that are not recommended by the National Committee for Clinical Laboratory Standards for tests with S. pneumoniae. CONCLUSIONS: Current susceptibility testing practices with S. pneumoniae in the United States indicate limited precision and a tendency for laboratories to test and report results obtained with antimicrobial agents of questionable therapeutic value against this organism. Continued efforts to standardize susceptibility testing of S. pneumoniae in the United States are warranted. In addition, modifications of existing interpretive criteria may be necessary.

Clonal relationships among high-level penicillin-resistant Streptococcus pneumoniae in the United States.

During the past 6 to 7 years, the problem of antimicrobial resistance in Streptococcus pneumoniae has grown dramatically in the United States. Currently, approximately 26.5% of pneumococcal isolates express intermediate levels of resistance to penicillin; approximately 17.5% are highly penicillin resistant. We studied whether clonal relationships exist among current isolates of high-level penicillin-resistant S. pneumoniae (PRSP) in the United States. One hundred forty-seven PRSP isolates recovered in a 30-center surveillance study in the United States during 1994-1995 were characterized with respect to serotype, antimicrobial susceptibility pattern, and pulsed-field gel electrophoresis (PFGE) profile. Only six serotypes were observed among the 147 PRSP isolates examined in this study: 6A, 6B, 9A, 14, 19F, and 23F. One hundred three (70.1%) of the 147 strains were characterized by one of only nine PFGE types; 76 (51.7%) of the 147 isolates were characterized by only four PFGE profiles. Currently in the United States, most PRSP strains are represented by relatively few clonal groups.