Analysis of the Bacterial Communities Present in Lungs of Patients with Cystic Fibrosis from American and British Centers

Molecular Microbiology Research Laboratory, Pharmaceutical Science Research Division, King's College London, 150 Stamford Street, Franklin-Wilkins Building, London, SE1 9NH, United Kingdom.
Journal of clinical microbiology (Impact Factor: 4.23). 11/2010; 49(1):281-91. DOI: 10.1128/JCM.01650-10
Source: PubMed

ABSTRACT The aim of this study was to determine whether geographical differences impact the composition of bacterial communities present in the airways of cystic fibrosis (CF) patients attending CF centers in the United States or United Kingdom. Thirty-eight patients were matched on the basis of clinical parameters into 19 pairs comprised of one U.S. and one United Kingdom patient. Analysis was performed to determine what, if any, bacterial correlates could be identified. Two culture-independent strategies were used: terminal restriction fragment length polymorphism (T-RFLP) profiling and 16S rRNA clone sequencing. Overall, 73 different terminal restriction fragment lengths were detected, ranging from 2 to 10 for U.S. and 2 to 15 for United Kingdom patients. The statistical analysis of T-RFLP data indicated that patient pairing was successful and revealed substantial transatlantic similarities in the bacterial communities. A small number of bands was present in the vast majority of patients in both locations, indicating that these are species common to the CF lung. Clone sequence analysis also revealed that a number of species not traditionally associated with the CF lung were present in both sample groups. The species number per sample was similar, but differences in species presence were observed between sample groups. Cluster analysis revealed geographical differences in bacterial presence and relative species abundance. Overall, the U.S. samples showed tighter clustering with each other compared to that of United Kingdom samples, which may reflect the lower diversity detected in the U.S. sample group. The impact of cross-infection and biogeography is considered, and the implications for treating CF lung infections also are discussed.

  • [Show abstract] [Hide abstract]
    ABSTRACT: Sputum obtained from patients with cystic fibrosis (CF) is highly viscous and often heterogeneous in bacterial distribution. Adding dithiothreitol (DTT) is the standard method for liquefaction prior to processing sputum for molecular detection assays. To determine if DTT treatment homogenizes bacterial distribution within sputum, we measured the difference in mean total bacterial abundance and abundance of Burkholderia multivorans between aliquots of DTT-treated sputum samples with and without a mechanical homogenization (MH) step using a high speed dispersing element. Additionally, we measured the effect of MH on bacterial abundance. We found a significant decrease in the difference between the means of aliquots which subjected to only DTT treatment and those which included a MH step (all bacteria, p = 0.04; B. multivorans, p = 0.05). There was no significant effect of MH on bacterial abundance in sputum. Although our results are from a single CF patient, they indicate that mechanical homogenization increases the homogeneity of bacteria in sputum.
    Journal of clinical microbiology 04/2014; 52(7). DOI:10.1128/JCM.00487-14 · 4.23 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: In critically ill patients, the development of pneumonia results in significant morbidity, mortality, and additional healthcare costs. Accurate and rapid identification of the microbial pathogens in patients with pulmonary infections could lead to targeted antimicrobial therapy with potentially less adverse effects and lower costs. Major advances in next-generation sequencing (NGS) allow for culture-independent identification of pathogens. The present study used NGS of essentially full-length PCR-amplified 16S ribosomal DNA from the bronchial aspirates of intubated patients with suspected pneumonia. The results from 61 patients demonstrated that sufficient DNA could be obtained from 72% of samples, 44% of which (27 samples) yielded PCR amplimers suitable for NGS. Out of 27 sequenced samples, only 20 had bacterial culture growth, while microbiological and NGS identification of bacteria coincided in 17 (85%) of these samples. Despite the lack of bacterial growth in 7 samples that yielded amplimers and were sequenced, the NGS identified a number of bacterial species in these samples. Overall, a significant diversity of bacterial species was identified from the same genus as the predominant cultured pathogens. The number of NGS-identifiable bacterial genera was consistently higher than identified by standard microbiological methods. As technical advances reduce the processing and sequencing times, NGS-based methods will ultimately be able to provide clinicians with rapid, precise, culture-independent identification of bacterial, fungal, and viral pathogens and their antimicrobial sensitivity profiles.
    Journal of Clinical Microbiology 08/2014; 52(11). DOI:10.1128/JCM.01678-14 · 4.23 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Cystic fibrosis is characterised by chronic polymicrobial infection and inflammation in the airways of patients. Antibiotic treatment regimens, targeting recognised pathogens, have substantially contributed to increased life expectancy of patients with this disease. Although the emergence of antimicrobial resistance and selection of highly antibiotic-resistant bacterial strains is of major concern, the clinical relevance in cystic fibrosis is yet to be defined. Resistance has been identified in recognised cystic fibrosis pathogens and in other bacteria (eg, Prevotella and Streptococcus spp) detected in the airway microbiota, but their role in the pathophysiology of infection and inflammation in chronic lung disease is unclear. Increased antibiotic resistance in cystic fibrosis might be attributed to a range of complex factors including horizontal gene transfer, hypoxia, and biofilm formation. Strategies to manage antimicrobial resistance consist of new antibiotics or localised delivery of antimicrobial agents, iron sequestration, inhibition of quorum-sensing, and resistome analysis. Determination of the contributions of every bacterial species to lung health or disease in cystic fibrosis might also have an important role in the management of antibiotic resistance.
    The Lancet 08/2014; 384(9944):703-13. DOI:10.1016/S0140-6736(14)61137-5 · 39.21 Impact Factor

Full-text (2 Sources)

Available from
May 19, 2014