Temporal trends of serotypes from invasive pneumococcal disease (IPD) in Spain from 1979 to September 2007 under antibiotic and vaccine pressure were analyzed. A significant trend in pneumococcal conjugate 7-valent vaccine (PCV7) serotypes (except serotype 4) was found, whereby the prevalence increased from the early 1980s and decreased in the 2000s for all but serotype 23F, which began decreasing in the late 1980s. Among the major non-PCV7 serotypes, a significant decrease was observed for serotypes 1, 5, and 7F in the 1980s. From the late 1990s, serotypes 1, 5, 6A, 7F, and 19A increased significantly, while serotypes 3 and 8 showed similar but nonsignificant trends over time. The incidence of IPD cases was 10.7/100,000 for the period 1996 to 2006, with reporting coverage ranging from 18% to 43%. A significant decrease in IPD incidence due to PCV7 serotypes was observed, while the incidence of non-PCV7 serotypes increased, with the consequence that there was no clear pattern in the overall incidence of IPD. Penicillin nonsusceptibility was correlated with the proportion of PCV7 serotypes. Erythromycin nonsusceptibility increased in association with long-half-life macrolide consumption and then decreased in 2004 to 2007. The increase in PCV7 serotypes and antibiotic nonsusceptibility related to antibiotic consumption in the 1980s and 1990s was reversed in the 2000s, probably as a result of PCV7 immunization. The decrease in IPD incidence due to PCV7 serotypes was mirrored by an increase in that of non-PCV7 serotypes. The impact of various preventive/therapeutic strategies on pneumococcal evolution is serotype dependent, and the dynamics remain unpredictable.

Pediatric parapneumonic empyema (PPE) has been increasing in several countries including Spain. Streptococcus pneumoniae is a major PPE pathogen; however, antimicrobial pretreatment before pleural fl uid (PF) sampling frequently results in negative diagnostic cultures, thus greatly underestimating the contribution of pneumococci, especially pneumococci susceptible to antimicrobial agents, to PPE. The study aim was to identify the serotypes and genotypes that cause PPE by using molecular diagnostics and relate these data to disease incidence and severity. A total of 208 children with PPE were prospectively enrolled; blood and PF samples were collected. Pneumococci were detected in 79% of culture-positive and 84% of culture-negative samples. All pneumococci were genotyped by multilocus sequence typing. Serotypes were determined for 111 PPE cases; 48% were serotype 1, of 3 major genotypes previously circulating in Spain. Variance in patient complication rates was statistically signifi cant by serotype. The recent PPE increase is principally due to nonvaccine serotypes, especially the highly invasive serotype 1. P leural effusions occur in at least 40% of children hospitalized with bacterial pneumonia. Occasionally, the infectious agent invades the pleura to cause pediatric paraneumonic empyema (PPE) (1), characterized by the presence of pus. Although rarely associated with fatalities in industrialized countries, PPE often results in prolonged hospitalization and surgical intervention, and patients are at risk for serious and long-lasting illness (2,3).An increasing incidence of PPE has been reported in several countries since the mid-1990s (2-6), but it is not clear why. Streptococcus pneumoniae is the most frequently found microorganism in most recent reports. However, conventional microbiologic culture methods have low sensitivity, usually because of antimicrobial pretreatment before sterile-site sampling. Consequently, the contribution of antimicrobial drug-susceptible serotypes might be higher than reported estimates. Molecular and antigen detection-based techniques, including direct molecular typing of culture-negative pleural fl uid (PF) samples (7), can be useful adjuncts in defi ning the contributory role of different microorganisms and pneumococcal serotypes to PPE etiology (4,8).Our study's goal was to prospectively investigate the molecular epidemiology of pneumococcal PPE among children admitted to 3 of the largest tertiary-care pediatric hospitals in Spain. There were 4 objectives: 1) identify the serotypes and multilocus sequence typing (MLST) genotypes causing PPE and determine whether a temporal change in the circulating genotypes could explain the recent increase; 2) determine whether the causal genotypes were only associated with PPE or also caused other invasive pneumococcal disease (IPD) in the same population, or were carried by healthy children; 3) compare serotypes and genotypes recovered from northern and southern Spain in the context of regional differences in 7-valent pneumococcal conjugat...

Pneumococcal parapneumonic empyema is an increasingly common complication in children. Conventional microbiological cultures indicate bacterial causes in as few as 8% of cases; therefore, there is a vital need for new molecular methods of detection and diagnosis. The development and clinical evaluation of real-time PCR-based assays to detect the pneumococcal capsular wzg gene of all serotypes tested are reported here, and 24 of them have been identified in clinical specimens. Using real-time PCR assays with highly specific TaqMan MGB probes that target DNA sequences within the capsular polysaccharide gene cluster, it was possible to differentiate serotypes 1, 3, 5, 4, 6A, 6B, 7F/A, 8, 9V/A/N/L, 14, 15B/C, 18C/B, 19A, 19F/B/C, 23F and 23A. These assays showed high sensitivity (five to ten pneumococcal DNA equivalents) and they were validated with 175 clinical isolates of known serotypes. The clinical value of this approach was demonstrated by analysis of 88 culture-negative pleural fluids from children diagnosed with parapneumonic empyema in three Spanish hospitals. Pneumococcal DNA was detected in 87.5% of pleural fluids, and serotypes 1, 7F and 3 were responsible for 34.3%, 16.4% and 11.9%, respectively, of cases of parapneumonic empyema in children. Such molecular methods are critical for the diagnosis of invasive pneumococcal disease and continued epidemiological surveillance in order to monitor serotype vaccine effectiveness.

Hand, foot and mouth disease (HFMD) is a childhood illness frequently caused by genotypes belonging to the enterovirus A species, including coxsackievirus (CV)-A16 and enterovirus (EV)-71. Between 2010 and 2012, several outbreaks and sporadic cases of HFMD occurred in different regions of Spain. The objective of the present study was to describe the enterovirus epidemiology associated with HFMD in the country. A total of 80 patients with HFMD or atypical rash were included. Detection and typing of the enteroviruses were performed directly in clinical samples using molecular methods. Enteroviruses were detected in 53 of the patients (66%). CV-A6 was the most frequent genotype, followed by CV-A16 and EV-71, but other minority types were also identified. Interestingly, during almost all of 2010, CV-A16 was the only causative agent of HFMD but by the end of the year and during 2011, CV-A6 became predominant, while CV-A16 was not detected. In 2012, however, both CV-A6 and CV-A16 circulated. EV-71 was associated with HFMD symptoms only in three cases during 2012. All Spanish CV-A6 sequences segregated into one major genetic cluster together with other European and Asian strains isolated between 2008 and 2011, most forming a particular clade. Spanish EV-71 strains belonged to subgenogroup C2, as did most of the European sequences circulated. In conclusion, the recent increase of HFMD cases in Spain and other European countries has been due to a larger incidence of circulating species A enteroviruses, mainly CV-A6 and CV-A16, and the emergence of new genetic variants of these viruses.