Polyphasic characterisation of Burkholderia cepaciacomplex species isolated from children with cystic fibrosis.

Fernando José Vicenzi,Carlos Antônio Riedi,Jussara Kasuko Palmeiro,Nelson Augusto Rosario-Filho,Libera Maria Dalla-Costa,Helena Aguilar Peres Homem de Mello de Souza,Marcelo Pillonetto

doi:10.1590/0074-02760150314

Abstract

Cystic fibrosis (CF) patients with Burkholderia cepacia complex (Bcc) pulmonary infections have high morbidity and mortality. The aim of this study was to compare different methods for identification of Bcc species isolated from paediatric CF patients. Oropharyngeal swabs from children with CF were used to obtain isolates of Bcc samples to evaluate six different tests for strain identification. Conventional (CPT) and automatised (APT) phenotypic tests, polymerase chain reaction (PCR)-recA, restriction fragment length polymorphism-recA, recAsequencing, and matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) were applied. Bacterial isolates were also tested for antimicrobial susceptibility. PCR-recA analysis showed that 36 out of the 54 isolates were Bcc. Kappa index data indicated almost perfect agreement between CPT and APT, CPT and PCR-recA, and APT and PCR-recA to identify Bcc, and MALDI-TOF and recAsequencing to identify Bcc species. The recAsequencing data and the MALDI-TOF data agreed in 97.2% of the isolates. Based on recA sequencing, the most common species identified were Burkholderia cenocepacia IIIA (33.4%),Burkholderia vietnamiensis (30.6%), B. cenocepaciaIIIB (27.8%), Burkholderia multivorans (5.5%), and B. cepacia (2.7%). MALDI-TOF proved to be a useful tool for identification of Bcc species obtained from CF patients, although it was not able to identify B. cenocepacia subtypes.

Highlights

Cystic fibrosis (CF) pathophysiology leads to an increased susceptibility to respiratory infections (Drevinek & Mahenthiralingam 2010)
Identification of one bacterial isolate did not agree by different techniques: Bordetella spp by conventional phenotypic tests (CPT), Cupriavidus pauculus by automated phenotypic tests (APT), and as B. cepacia by both recA sequencing method and MALDI-TOF
One strain identified as B. vietnamiensis by recA sequencing was identified as B. cenocepacia by MALDI-TOF (Table I)

Summary

Introduction

Cystic fibrosis (CF) pathophysiology leads to an increased susceptibility to respiratory infections (Drevinek & Mahenthiralingam 2010). CF patients with Bcc infection have high rates of morbidity and mortality (Drevinek & Mahenthiralingam 2010, LiPuma 2010). Bcc species comprise phenotypically indistinguishable microorganisms with high genetic similarity (> 97.5%) that was determined using 16S rDNA sequencing (Coenye et al 2001, Vandamme & Dawyndt 2011). The discriminatory power of recA sequencing method is better than 16S rDNA sequencing analysis because recA has enough nucleotide variability to enable differentiation of the main Bcc species isolated from CF patients (Mahenthiralingam et al 2000). The aim of this study was to compare different methods for identification of Bcc species isolated from oropharyngeal swabs of paediatric CF patients

Objectives

Results

Conclusion