Bacterial Culture Underestimates Lung Pathogen Detection and Infection Status in Cystic Fibrosis

Abstract

Microbiological surveillance of airway secretions is central to clinical care in cystic fibrosis (CF). However, the efficacy of microbiological culture, the diagnostic gold standard for pathogen detection, has been increasingly questioned. Here we compared culture with targeted quantitative PCR (QPCR) for longitudinal detection of 2 key pathogens, Pseudomonas aeruginosa and Staphylococcus aureus. Prospectively collected respiratory samples taken from 20 pediatric and 20 adult CF patients over a period of 3-years were analyzed. Patients were eligible if considered free of chronic Pseudomonas infection within 12-months prior to start of study. QPCR revealed high levels of infection with both pathogens not apparent from culture alone. Pseudomonas and Staphylococcus were detected by culture on at least one sampling occasion in 12 and 29 of the patients, respectively. Conversely, both pathogens were detected in all 40 patients by QPCR. Classification of infection status also significantly altered in both pediatric and adult patients, where the number of patients deemed chronically infected with Pseudomonas and Staphylococcus increased from 1 to 28 and 9 to 34, respectively. Overall, Pseudomonas and Staphylococcus infection status classification changed respectively for 36 and 27 of all patients. In no cases did molecular identification lead to a patient being in a less clinically serious infection category. Pathogen detection and infection status classification significantly increased when assessed by QPCR in comparison to culture. This could have implications for clinical care of CF patients, including accuracy of infection diagnosis, relevant and timely antibiotic selection, antimicrobial resistance development, establishment of chronic infection, and cross-infection control. IMPORTANCE Chronic lung infection is the leading cause of morbidity and early mortality for people with cystic fibrosis (pwCF). Microbiological surveillance to detect lung pathogens is recommended as best practise in CF patient care. Here we studied pathogen detection in 40 pwCF over several years. We found that microbiological culture, the diagnostic gold standard, was significantly disparate to targeted culture-independent approaches for detection and determination of chronic infection status of two important pathogens in CF. Pathogen detection was significantly lower by culture and consequently infection status was also misclassified in most cases. In particular, the extent of chronic infection by both P. aeruginosa and S. aureus not realized with culture was striking. Our findings have implications for the development of infection and clinical care of pwCF. Future longitudinal studies with greater patient numbers will be needed to establish the full extent of the clinical implications indicated from this study.

Keywords: Pseudomonas aeruginosa; QPCR; Staphylococcus aureus; chronic infection; infection status; lung infection.

FIG 1

Flow diagram detailing patient selection process. Only patients who contributed ≥6 samples with contemporaneous diagnostic microbiology data (DM) were included in the final analyses. Using a modification of the Leeds criteria, patients were deemed to be chronically or intermittently colonized with a given pathogen if >50% or ≤50% of samples, respectively, over the 3-year study period were positive by diagnostic microbiology or targeted QPCR. A minimum of ≥6 samples was chosen as less samples would have increased the likelihood of misclassifying infection status.

FIG 2

Pathogen detection by conventional culture and molecular-based approaches in pediatric and adult cystic fibrosis patients. Given for each patient, is the percentage of respiratory samples over the 3-year study duration that were culture (orange) or QPCR (gray) positive for (A) Pseudomonas aeruginosa and (B) Staphylococcus aureus. In each instance, the dashed line denotes the threshold for chronic (>50%) or intermittent (≤50%) colonization. Numbers on the x axis represent individual patient study numbers.

FIG 3

Changes in pathogen infection status classification from when defined by diagnostic culture to then by molecular-based detection in pediatric and adult patients. Given are changes in infection status classification for (A) Pseudomonas aeruginosa and (B) Staphylococcus aureus in each of the children and adult CF patients. In each instance, colored lines represent individual patients.