Accurate identification of members of the Burkholderia cepacia complex in cystic fibrosis sputum.

UNLABELLED: The Burkholderia cepacia complex (BCC) is a group of closely related species which includes opportunistic pathogens causing chronic respiratory infections in immunocompromised patients, or individuals affected by cystic fibrosis (CF). Other Burkholderia species causing infection in the CF population are Burkholderia gladioli and Burkholderia pseudomallei. Traditional phenotypic analyses have been demonstrated to be inadequate for reliable identifications of isolates of BCC and B. gladioli. A pan-genomic analysis approach was used to design species-specific probes for Burkholderia cenocepacia, B. cepacia, Burkholderia multivorans, Burkholderia vietnamiensis, Burkholderia ambifaria, Burkholderia dolosa, Burkholderia pyrrocinia and B. gladioli. Multiplex real-time PCR assay was developed and tested using sputum specimens collected from CF patients spiked with Burkholderia species. The assay exhibited 100% sensitivity for all eight target species and detected 10(2) to 10(3)  CFU ml(-1) when applied to spiked sputum. Our PCR assay resulted highly specific for each of the Burkholderia species tested, allowing discrimination among Burkholderia and non-Burkholderia pathogens. Analysis carried out on 200 sputa positive for the presence of Burkholderia revealed that PCR assay and recA sequencing were fully comparable for identification of Burkholderia at the level of species. SIGNIFICANCE AND IMPACT OF THE STUDY: Burkholderia cepacia complex (BCC) has a complex taxonomic organization and its identification is a challenge for microbiology laboratories. Nonidentification or misidentification of BCC isolates represent a problem in epidemiology and treatment of cystic fibrosis patients. The high specificity and sensitivity of the multiplex Real-time PCR assay developed in this study indicates its potential to be a rapid and reliable method for the detection of Burkholderia at the level of species from sputum samples of cystic fibrosis patients.

MALDI-TOF mass spectrometry and microsatellite markers to evaluate Candida parapsilosis transmission in neonatal intensive care units.

Recent studies on outbreaks of Candida showed an increased incidence of bloodstream infections in neonatal intensive care units (NICUs) caused by C. parapsilosis species, highlighting the need for the proper identification and epidemiology of these species. Several systems are available for molecular epidemiological and taxonomic studies of fungal infections: pulsed-field gel electrophoresis (PFGE) represents the gold standard for typing, but is also one of the most lengthy and expensive, while simple sequence repeats (SSRs) is based on polymerase chain reaction (PCR) amplification and is, therefore, faster. Only recently, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been used to identify and type microorganisms involved in nosocomial outbreaks. In our study, 19 strains of C. parapsilosis isolated from the blood cultures of neonates admitted to the University Hospital Federico II were genotyped by the amplification of eight SSR markers and by MALDI-TOF MS. Electrophoretic and spectrometric profile results were compared in order to identify similarities among the isolates and to study microevolutionary changes in the C. parapsilosis population. The discriminatory power and the unweighted pair group method with arithmetic mean (UPGMA) dendrograms generated were compared in order to evaluate the correlation of the groups established by the analysis of the clusters by both methods. Both methods were rapid and effective in highlighting identical strains and studying microevolutionary changes in the population. Our study evidenced that mass spectroscopy is a useful technique not only for the identification but also for monitoring the spread of strains, which is critical to control nosocomial infections.

Proton pump inhibitors as a risk factor for paediatric Clostridium difficile infection.

BACKGROUND: Proton pump inhibitors (PPIs) and H(2) receptor antagonists (H(2)RAs) may play an important role on the onset of Clostridium difficile-associated disease (CDAD) in adults. The impact of Clostridium difficile on children treated with gastric acid-suppressing agents remains unknown. AIM: To investigate the relationship between CDAD and exposure to acid suppressive therapy in hospitalized paediatric patients. METHODS: We reviewed the medical records of children, with a diagnosis of protracted diarrhoea and abdominal pain, whose stool was analysed for C. difficile toxins. We identified 68 patients with CDAD. For each patient, we randomly selected one control subjects with stool analysis negative for C. difficile. Comorbid illnesses, previous hospitalizations, antibiotics, corticosteroids, immunosuppressants and gastric acid suppressing exposures were recorded. RESULTS: The use of PPI was significantly higher in C. difficile positive group compared with C. difficile negative group [odds ratio (OR): = 4.5; 95% confidence interval (CI) = 1.4-14.4]. We also found a trend for the use of H(2)RAs in patients infected by C. difficile compared with C. difficile negative comparison group (OR: = 3.8; 95% CI = 0.7-18.9). CONCLUSIONS: Children exposed to PPIs therapy seem to be at higher risk for the development of Clostridium difficile-associated disease.