Pseudomonas aeruginosa (PA) infection is the main cause of mortality in Cystic Fibrosis (CF) patients. Rapid and sensitive PA detection is pivotal to improve the prognosis. This thesis includes two parts. The first focuses on the development of a Real-time PCR protocol to detect dormant- non-culturable PA in sputum samples from CF patients (CFPA). 12 culture-positive (CP) and 29 culture-negative (CN) samples were analyzed by a qPCR targeting ecfX, 24% resulted positive (P), with bacterial counts (102 - 106 cells/ml) also higher than those obtained for CP samples. A significant association of culture-negative/PCR-positive samples from chronic patients with symptoms relapse (P=0.018) and following culture-positive samples (P=0.034) was found. These findings stress the value of qPCR approaches in monitoring P. aeruginosa colonization and foreseeing symptom recurrence in CF patients. In second part we investigated the epidemiological relatedness of CFPA. 77 CFPA and 19 PA from non-CF patients (NCFPA) were analysed by pulsed field gel electrophoresis (PFGE) profile, antibiotic resistance (AR) and mucous production. Population structure analysis showed an overall polyclonality, more evident among CF strains. 100% similarity was only found within each of 10 CFPA pairs from the same patient, and 95% between 2 CFPA isolates from different patients who never met each other in hospital, suggesting a common source of infection rather than cross-contamination. Among NCFPA four isolates from as many inpatients and two from as many outpatients showed the same pulsotype (H and M, respectively). AR was more common among CFPA and nonmucoid phenotype. In particular, a significant (P=0.016) association was found between gentamycin-resistance CFPA and nonmucoid phenotype. AR patterns were however highly variable, also among PA belonging to the same pulsotype; suggesting the lack of involvement of multiple antibiotic resistance (MAR) in cross-colonization.

Real-time PCR detection and PFGE typing of Pseudomonas aeruginosa from cystic fibrosis patients / Amiri, Mehdi. - (2016 Feb 17).

Real-time PCR detection and PFGE typing of Pseudomonas aeruginosa from cystic fibrosis patients

Amiri, Mehdi
2016-02-17

Abstract

Pseudomonas aeruginosa (PA) infection is the main cause of mortality in Cystic Fibrosis (CF) patients. Rapid and sensitive PA detection is pivotal to improve the prognosis. This thesis includes two parts. The first focuses on the development of a Real-time PCR protocol to detect dormant- non-culturable PA in sputum samples from CF patients (CFPA). 12 culture-positive (CP) and 29 culture-negative (CN) samples were analyzed by a qPCR targeting ecfX, 24% resulted positive (P), with bacterial counts (102 - 106 cells/ml) also higher than those obtained for CP samples. A significant association of culture-negative/PCR-positive samples from chronic patients with symptoms relapse (P=0.018) and following culture-positive samples (P=0.034) was found. These findings stress the value of qPCR approaches in monitoring P. aeruginosa colonization and foreseeing symptom recurrence in CF patients. In second part we investigated the epidemiological relatedness of CFPA. 77 CFPA and 19 PA from non-CF patients (NCFPA) were analysed by pulsed field gel electrophoresis (PFGE) profile, antibiotic resistance (AR) and mucous production. Population structure analysis showed an overall polyclonality, more evident among CF strains. 100% similarity was only found within each of 10 CFPA pairs from the same patient, and 95% between 2 CFPA isolates from different patients who never met each other in hospital, suggesting a common source of infection rather than cross-contamination. Among NCFPA four isolates from as many inpatients and two from as many outpatients showed the same pulsotype (H and M, respectively). AR was more common among CFPA and nonmucoid phenotype. In particular, a significant (P=0.016) association was found between gentamycin-resistance CFPA and nonmucoid phenotype. AR patterns were however highly variable, also among PA belonging to the same pulsotype; suggesting the lack of involvement of multiple antibiotic resistance (MAR) in cross-colonization.
17-feb-2016
P. aeruginosa, qPCR, cystic fybrosis,PFGE
File in questo prodotto:
File Dimensione Formato  
PhD thesis (Mehdi Amiri).pdf

Solo gestori archivio

Tipologia: Tesi di dottorato
Licenza d'uso: Non specificato
Dimensione 1.44 MB
Formato Adobe PDF
1.44 MB Adobe PDF   Visualizza/Apri   Richiedi una copia

I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.

Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11566/243098
 Attenzione

Attenzione! I dati visualizzati non sono stati sottoposti a validazione da parte dell'ateneo

Citazioni
  • ???jsp.display-item.citation.pmc??? ND
  • Scopus ND
  • ???jsp.display-item.citation.isi??? ND
social impact