Ana Fernández-Olmos

MALDI-TOF MS improves routine identification of non-fermenting Gram negative isolates from cystic fibrosis patients.

Identification of non-fermenting Gram-negative bacteria (NFGNB) from cystic fibrosis (CF) patients is often limited. A collection of stored NFGNB isolates (n=182) recovered from CF patients over a 15 year period was examined. The routinely reported identification during this period was compared with that obtained by MALDI-TOF MS. Isolates giving discrepant identification at the genus level were further analyzed by 16S rDNA sequencing. The MALDI-TOF MS system identified 94% of the isolates, including Burkholderia cepacia and Pandoraea spp. isolates, the latter previously misidentified as other NFGNB by conventional microbiological methods. Lack of identification by MALDI-TOF MS was associated with the absence of entries in the database.

In vitro prevention of Pseudomonas aeruginosa early biofilm formation with antibiotics used in cystic fibrosis patients

The ability of antibiotics used in bronchopulmonary infections in cystic fibrosis (CF) patients to prevent Pseudomonas aeruginosa early biofilm formation was studied using a biofilm microtitre assay with 57 non-mucoid P. aeruginosa isolates (44 first colonisers and 13 recovered during the initial intermittent colonisation stage) obtained from 35 CF patients. Minimum biofilm inhibitory concentrations (BICs) of levofloxacin, ciprofloxacin, imipenem, ceftazidime, tobramycin, colistin and azithromycin were determined by placing a peg lid with a formed biofilm onto microplates containing antibiotics. A modification of this protocol consisting of antibiotic challenge during biofilm formation was implemented in order to determine the biofilm prevention concentration (BPC), i.e. the minimum concentration able to prevent biofilm formation. The lowest BPCs were for fluoroquinolones, tobramycin and colistin and the highest for ceftazidime and imipenem. The former antibiotics had BPCs identical to or only slightly higher than their minimum inhibitory concentrations (MICs) determined by standard Clinical and Laboratory Standards Institute (CLSI) microdilution and were also active on formed biofilms as reflected by their low BIC values. In contrast, ceftazidime and imipenem were less effective for prevention of biofilm formation and on formed biofilms. In conclusion, the new BPC parameter determined in non-mucoid P. aeruginosa isolates recovered during early colonisation stages in CF patients supports early aggressive antimicrobial treatment guidelines in first P. aeruginosa-colonised CF patients.

Emergence of a mutL Mutation Causing Multilocus Sequence Typing–Pulsed-Field Gel Electrophoresis Discrepancy among Pseudomonas aeruginosa Isolates from a Cystic Fibrosis Patient

ABSTRACT A multilocus sequence type (MLST) shift (from ST242 to ST996) was detected in Pseudomonas aeruginosa isolates with a uniform pulsed-field gel electrophoresis (PFGE) pattern obtained from a chronically colonized patient. MLST mutational change involved the mutL gene with the consequent emergence of a hypermutable phenotype. This observation challenges the required neutrality of mutL as an appropriate marker in MLST and alerts researchers to the limitations of MLST-only-based population studies in chronic infections under constant antibiotic selective pressure.

Population Structure and Antimicrobial Susceptibility of Both Nonpersistent and Persistent Pseudomonas aeruginosa Isolates Recovered from Cystic Fibrosis Patients

ABSTRACT Seventy-six Pseudomonas aeruginosa isolates recovered from chronically (n = 18) and nonchronically (n = 18) colonized cystic fibrosis (CF) patients (2002 to 2009) were grouped in separate polyclonal populations. International CF epidemic clones were not identified, but the high-risk clone ST274, also found circulating in Spanish hospitals, was present. Persistent isolates were more resistant to antibiotics than nonpersistent isolates.

Clinical and Microbiological Features of a Cystic Fibrosis Patient Chronically Colonized with Pandoraea sputorum Identified by Combining 16S rRNA Sequencing and Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry

ABSTRACT Clonal isolates identified as various nonfermentative Gram-negative bacilli over a 5-year period from sputum cultures of a 30-year-old cystic fibrosis patient were successfully reidentified as Pandoraea sputorum by combining 16S rRNA sequencing and matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS). Decreased lung function improved after 1 year of azithromycin and inhaled 7%-hypertonic saline treatment.

Unorthodox long‐term aerosolized ampicillin use for methicillin‐susceptible Staphylococcus aureus lung infection in a cystic fibrosis patient

Staphylococcus aureus is a significant cause of pulmonary colonization in cystic fibrosis (CF) patients. The optimal strategy of therapy in chronically infected patients with this pathogen is not yet established. We report a successful long‐term aerosolized ampicillin treatment of a 14‐year‐old girl with chronic symptomatic S. aureus lung infection. Pediatr Pulmonol. 2009; 44:512–515.

Evaluation of Cobas Ampliprep Automated Nucleic Acid Extraction for Genotyping Human Papillomavirus (HPV) by the Roche linear array HPV Test

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Clinical features of a cystic fibrosis patient chronically colonized with Pandoraea pnomenusa identified using MALDI-TOF mass spectrometry

461* Clinical features of a cystic fibrosis patient chronically colonized with Pandoraea pnomenusa identified using MALDI-TOF mass spectrometry A. Fernandez-Olmos1, M.I. Morosini1, M. Garcia-Castillo1, L. Garcia-Garcia1, A. Lamas2, R. Canton1, L. Maiz2. 1Hosp. Univ. Ramon y Cajal, Servicio de Microbiologia, CIBER Epidemiologia y Salud Publica, Madrid, Spain; 2Hosp. Univ. Ramon y Cajal, Unidad Fibrosis Quistica, Madrid, Spain