Genomic analysis of Burkholderia cenocepacia isolated from a liver abscess in a patient with cystic fibrosis.

Burkholderia cenocepacia complex is associated with high transmissibility, virulence, and poor prognosis in cystic fibrosis (CF) patients. However, extrapulmonary infections are rare. We investigated the genome of a B. cenocepacia IIIA isolated from a liver abscess in a Brazilian CF patient and compared it to strain J2315. The whole genome was sequenced, and contigs were annotated by Rapid Annotation using Subsystem Technology. The Pathosystems Resource Integration Center was used to map antimicrobial and virulence genes. The genomic island (GIs) analysis was performed using two prediction methods, and the presence of putative plasmids and insertion sequences (ISs) was investigated. The isolate was confirmed as B. cenocepacia IIIA to ST-28 (ET12 lineage). A total of 64 genes for antimicrobial resistance and 47 genes related to virulence were identified. Among the virulence factors, there was a predominance of factors related to the invasion mechanism, to the flagellar biosynthesis protein, and to the RNA polymerase sigma factor for flagellar operon (cdpA). Two IS families (IS3 and IS5) and only one plasmid were found. On average 56 GIs were predicted by at least one of the methods applied. Comparative analysis showed resistance mechanisms and virulence factors revealing invasive determinants used by B. cenocepacia IIIA (ET12) in the process of disease spread to other infection sites (extrapulmonary) of highly virulent strains in CF patients.

Decreasing the Cut-off Score Value of MALDI-ToF MS Increase the Identities of Burkholderia cepacia Complex Species.

Burkholderia cepacia complex (Bcc) comprises 24 related species genetically distinct, associated with high mortality in cystic fibrosis (CF) patients. Due to a high level of similarity among Bcc species, accurate identification has been problematic, and most conventional and automated phenotypic tests have shown low accuracy. We evaluated accuracy of MALDI-ToF MS decreasing the cut-off score value to distinguish Bcc species compared to recA gene sequencing. A total of 145 Bcc isolates were analyzed. B. vietnamiensis (41.37%), B. cenocepacia IIIA (23.44%), B. multivorans (20%), B. cenocepacia IIIB (11.03%), and B. contaminans (2.75%) among other species were identified by recA sequencing. MALDI-ToF MS identified 100% of Bcc isolates at the genus level and 53.1% at the species level. By decreasing cut-off values for ≥1.70, the correct identification at the species level increased to 74.5%. MALDI-ToF MS proved to be useful at the genus level identification, but it still requires improvements that allow more precise identification, requiring continuous updates and addition of new spectra to its database. A review of interpretative criteria is a field to be explored with a large collection of Bcc species.

Analysis of airway microbiota in adults from a Brazilian cystic fibrosis center.

The application of next-generation sequencing tools revealed that the cystic fibrosis respiratory tract is a polymicrobial environment. We have characterized the airway bacterial microbiota of five adult patients with cystic fibrosis during a 14-month period by 16S rRNA tag sequencing using the Illumina technology. Microbial diversity, estimated by the Shannon index, varied among patient samples collected throughout the follow-up period. The beta diversity analysis revealed that the composition of the airway microbiota was highly specific for each patient, showing little variation among the samples of each patient analyzed over time. The composition of the bacterial microbiota did not reveal any emerging pathogen predictor of pulmonary disease in cystic fibrosis or of its unfavorable clinical progress, except for unveiling the presence of anaerobic microorganisms, even without any established clinical association. Our results could potentialy help us to translate and develop strategies in response to the pathobiology of this disease, particularly because it represents an innovative approach for CF centers in Brazil.

Monitoring clinical and microbiological evolution of a cystic fibrosis patient over 26 years: experience of a Brazilian CF Centre.

BACKGROUND: Burkholderia cepacia complex is a group of opportunistic pathogens in cystic fibrosis (CF) patients believed to be associated with poor prognosis and patient-to-patient transmissibility. Little is known about clinical outcomes after B. vietnamiensis chronic colonization/infection. CASE PRESENTATION: A 33 yo male patient had diagnosis of CF by 7 yo, after recurrent pneumonia during infancy and lobectomy (left upper lobe) at 6 yo. Burkholderia cepacia complex (Bcc) was first isolated by 13 yo, and the patient fulfilled the criteria for chronic colonization by 15 yo. In the following 16 years (1997-2013), there was intermittent isolation of P. aeruginosa and continuous isolation of Bcc, identified as B. vietnamiensis. There was clinical and laboratorial stability for 16 years with annual rate of decline in forced expiratory volume in 1 s (FEV1) and forced vital capacity (FVC) of 1.61 and 1.35%, respectively. From 2013 to 2015, there was significant clinical and lung function deterioration: annual rate of decline in FEV1 and FVC was 3 and 4.1%, respectively while body mass index decreased from 18.1 to 17.1. Episodes of hemoptysis and respiratory exacerbations (with hospital admissions) became more frequent. CF related diabetes was diagnosed (fasting glycemia: 116 mg/dL, oral glucose tolerance test: 305 mg/dL). Because of the severity of the disease in the last years, in addition to traditional microbiological surveillance, microbiome analysis by next generation sequencing (NGS) was performed on respiratory secretions. The NGS showed that 97% of the sequencing data were attributed to genus Burkholderia. CONCLUSIONS: We report the case of a 33-year-old male CF patient known to have chronic infection with B. vietnamiensis who remained clinically stable for 16 years and presented recent clinical and laboratorial deterioration. Microbiome analysis of respiratory secretions was performed in 3 samples collected in 2014-2015. Clinical deterioration overlapped with cystic fibrosis-related diabetes and microbiome composition revealed no significant differences when compared microbiome results to culture dependent methods.