Yvonne Yau

Risk factors and outcomes among children admitted to hospital with pandemic H1N1 influenza

Background: Limited data are available on disease characteristics and outcomes of children with 2009 pandemic influenza A(H1N1) virus infection (pandemic H1N1 influenza) who have required hospital admission. Methods: We reviewed the charts of 58 children with pandemic H1N1 influenza admitted to a large pediatric hospital in Ontario, Canada, between May 8 and July 22, 2009. We compared risk factors, severity indicators and outcomes of these children with those of 200 children admitted with seasonal influenza A during the previous 5 years (2004/05 to 2008/09). Results: Children with pandemic H1N1 influenza were significantly older than those with seasonal influenza (median age 6.4 years v. 3.3 years). Forty-six (79%) of the children with pandemic H1N1 influenza had underlying medical conditions; of the other 12 who were previously healthy, 42% were under 2 years of age. Children admitted with pandemic H1N1 influenza were significantly more likely to have asthma than those with seasonal influenza (22% v. 6%). Two children had poorly controlled asthma, and 6 used inhaled medications only intermittently. The median length of stay in hospital was 4 days in both groups of children. Similar proportions of children required admission to the intensive care unit (21% of those with pandemic H1N1 influenza and 14% of those with seasonal influenza) and mechanical ventilation (12% and 10% respectively). None of the children admitted with pandemic H1N1 influenza died, as compared with 1 (0.4%) of those admitted with seasonal influenza. Interpretation: Pandemic H1N1 influenza did not appear to cause more severe disease than seasonal influenza A. Asthma appears to be a significant risk factor for severe disease, with no clear relation to severity of asthma. This finding should influence strategies for vaccination and pre-emptive antiviral therapy.

Effect of pulmonary exacerbations on long-term lung function decline in cystic fibrosis

It is unknown what proportion of long-term lung function decline in cystic fibrosis (CF) is explained by pulmonary exacerbations. The aim of this study was to determine how exacerbations requiring hospitalisation contribute to the course of CF lung disease. This was a retrospective cohort study. The primary outcome was the rate of decline of forced expiratory volume in 1 s (FEV1) % predicted. Out of 851 subjects, 415 (48.8%) subjects had ≥1 exacerbation. After adjustment for confounders, the annual rate of FEV1 decline in those without an exacerbation was 1.2% per yr (95% CI 1.0–1.5), compared with 2.5% per yr (95% CI 2.1–2.8) in those with an exacerbation. The proportion of overall FEV1 decline associated with ≥1 exacerbation was 52% (95% CI 35.0–68.9). For a given number of exacerbations, the annual rate of FEV1 decline was greatest in subjects with ≤6 months between exacerbations. Half of FEV1 decline seen in CF patients was associated with pulmonary exacerbations. Time between exacerbations, specifically ≤6 months between exacerbations, plays an important contribution to overall lung function decline. These findings support using time to next exacerbation as a clinical end-point for CF trials.

Lung microbiota across age and disease stage in cystic fibrosis

Understanding the significance of bacterial species that colonize and persist in cystic fibrosis (CF) airways requires a detailed examination of bacterial community structure across a broad range of age and disease stage. We used 16S ribosomal RNA sequencing to characterize the lung microbiota in 269 CF patients spanning a 60 year age range, including 76 pediatric samples from patients of age 4–17, and a broad cross-section of disease status to identify features of bacterial community structure and their relationship to disease stage and age. The CF lung microbiota shows significant inter-individual variability in community structure, composition and diversity. The core microbiota consists of five genera - Streptococcus, Prevotella, Rothia, Veillonella and Actinomyces. CF-associated pathogens such as Pseudomonas, Burkholderia, Stenotrophomonas and Achromobacter are less prevalent than core genera, but have a strong tendency to dominate the bacterial community when present. Community diversity and lung function are greatest in patients less than 10 years of age and lower in older age groups, plateauing at approximately age 25. Lower community diversity correlates with worse lung function in a multivariate regression model. Infection by Pseudomonas correlates with age-associated trends in community diversity and lung function.

Stenotrophomonas maltophilia in cystic fibrosis: serologic response and effect on lung disease.

RATIONALE Stenotrophomonas maltophilia is one of the more common multidrug-resistant organisms isolated from the respiratory tract of patients with cystic fibrosis (CF), but the effect of chronic S. maltophilia infection on CF lung disease is unknown. OBJECTIVES To determine the impact of chronic S. maltophilia infection on lung disease in CF. METHODS We developed a serologic assay specific for S. maltophilia and in a cross-sectional study, measured serum antibodies to S. maltophilia in patients with CF to determine if a definition of chronic S. maltophilia isolation based on culture results corresponded to an immunologic response (serologic study). We then used this validated definition to examine the effect of chronic S. maltophilia on the severity of lung disease in a retrospective cohort study using the Toronto CF Database from 1997-2008 (cohort study). MEASUREMENTS AND MAIN RESULTS Serum antibody levels to S. maltophilia were measured in 179 patients with CF. Patients with chronic S. maltophilia had significantly higher mean antibody levels to S. maltophilia flagellin (P < 0.0001) and whole cell (P = 0.0004) compared with patients with intermittent or no S. maltophilia. The cohort study included 692 patients with an average follow-up of 8.3 years. In an adjusted log linear model, patients with chronic S. maltophilia infection had a significantly increased risk of pulmonary exacerbation requiring hospitalization and antibiotics compared with patients who had never had S. maltophilia (relative risk = 1.63; P = 0.0002). CONCLUSIONS Chronic S. maltophilia infection in patients with CF is associated with a specific immune response to this organism and is an independent risk factor for pulmonary exacerbations.

Chronic Stenotrophomonas maltophilia infection and mortality or lung transplantation in cystic fibrosis patients

BACKGROUND Chronic Stenotrophomonas maltophilia infection is an independent risk factor for severe pulmonary exacerbations in cystic fibrosis (CF) patients. The goal of this study was to determine the effect of chronic S. maltophilia infection on mortality and the need for lung transplantation in a longitudinal study of children and adults with CF. METHODS This was a cohort study of CF patients from the Hospital for Sick Children and St Michaels Hospital (Toronto, Canada) from 1997 to 2008. A Cox Regression model was used to estimate the hazard ratio (HR) to time of death or lung transplantation adjusting for age, gender, genotype, pancreatic status, CF related diabetes (CFRD), forced expiratory volume in 1 s (FEV1), body mass index, number of pulmonary exacerbations, Pseudomonas aeruginosa, Burkholderia cepacia complex, Aspergillus and chronic S. maltophilia infection. RESULTS A total of 687 patients were followed over the 12 year study period; 95 patients underwent a lung transplantation (of which 26 died) and an additional 49 patients died (total 144 events). In a Cox Regression model adjusting for baseline FEV1, baseline infection with B. cepacia complex (HR 1.72, 95% CI 1.09-2.71) and baseline chronic S. maltophilia infection (HR 2.80, 95% CI 1.65-4.76) were significantly associated with death or lung transplant. However, in a time-varying model, infection with B. cepacia complex and chronic S. maltophilia infection were no longer significant. CONCLUSIONS Baseline chronic S. maltophilia infection is associated with an almost three-fold increased risk of death or lung transplant in CF patients. It is still unclear, however, whether chronic S. maltophilia infection is simply a marker of severity of disease and ultimate mortality or whether it is causally related to disease progression.

Chronic Stenotrophomonas maltophilia infection and exacerbation outcomes in cystic fibrosis

BACKGROUND Chronic Stenotrophomonas maltophilia infection is a risk factor for pulmonary exacerbation in cystic fibrosis (CF) but its impact on subsequent clinical outcomes is unknown. The aim of this study was to determine the effect of chronic S. maltophilia infection and associated antimicrobial therapy on the recovery of forced expiratory lung volume in 1s (FEV(1)) following pulmonary exacerbation. METHODS This was a retrospective cohort study of patients with CF followed at The Hospital for Sick Children and St. Michaels Hospital from 1997 to 2008. The primary outcome was the difference in FEV(1) percent predicted from baseline to follow up after a pulmonary exacerbation. Secondary outcomes for the effect of antimicrobial therapy included time to subsequent exacerbation. RESULTS There were 1667 pulmonary exacerbations in 440 CF patients. Patients with chronic S. maltophilia infection did not recover their baseline FEV(1) following 31% of exacerbations and had an overall mean FEV(1) decline of 1.84% predicted after exacerbation. Older (p=0.02), female (p=0.02) patients with lower BMI z score (p=0.002) and Burkholderia cepacia complex infection (p=0.005), but not chronic S. maltophilia infection (p=0.86), had a greater decrease in follow up FEV(1)% pred compared to baseline. The number of days of antibiotic therapy against S. maltophilia during a pulmonary exacerbation was not associated with a significant difference in the FEV(1) recovery (p=0.69) or with a longer time to subsequent pulmonary exacerbation (p=0.56). CONCLUSIONS Although CF patients experience a significant decline in lung function following exacerbation, chronic S. maltophilia infection and associated antimicrobial therapy do not affect subsequent lung function recovery.

Phenotypic diversity within a Pseudomonas aeruginosa population infecting an adult with cystic fibrosis

Chronic airway infections caused by Pseudomonas aeruginosa contribute to the progression of pulmonary disease in individuals with cystic fibrosis (CF). In the setting of CF, within-patient adaptation of a P. aeruginosa strain generates phenotypic diversity that can complicate microbiological analysis of patient samples. We investigated within- and between- sample diversity of 34 phenotypes among 235 P. aeruginosa isolates cultured from sputum samples collected from a single CF patient over the span of one year, and assessed colony morphology as a screening tool for predicting phenotypes, including antimicrobial susceptibilities. We identified 15 distinct colony morphotypes that varied significantly in abundance both within and between sputum samples. Substantial within sample phenotypic heterogeneity was also noted in other phenotypes, with morphotypes being unreliable predictors of antimicrobial susceptibility and other phenotypes. Emergence of isolates with reduced susceptibility to β-lactams was observed during periods of clinical therapy with aztreonam. Our findings confirm that the P. aeruginosa population in chronic CF lung infections is highly dynamic, and that intra-sample phenotypic diversity is underestimated if only one or few colonies are analyzed per sample.

Role of respiratory viruses in pulmonary exacerbations in children with cystic fibrosis

Abstract Background The role of respiratory viruses in cystic fibrosis (CF) exacerbations is incompletely understood. Methods Cross-sectional study of CF children with a pulmonary exacerbation. Mid-turbinate swabs were tested by a direct immunofluorescent antibody assay and a multiplex PCR panel (ResPlex II v2.0, Qiagen). Resplex II was also applied to sputum or throat swab samples. Pulmonary function tests and quality of life and severity scores were recorded. Sputum cell counts, bacterial density and cytokines were measured. Results 26/43 (60.5%) subjects tested positive for at least one respiratory virus by any diagnostic method applied to any sample type. Virus-positive patients were younger (p =0.047), more likely to be male (p =0.029), and had higher CF clinical severity (p =0.041) and lower quality of life (physical) scores (p =0.023) but similar IL-8, neutrophil percentage and elastase levels. Conclusions Compared to non-viral exacerbations, viral-related exacerbations were associated with worse severity and quality of life scores but similar pulmonary inflammation.

Selective Sweeps and Parallel Pathoadaptation Drive Pseudomonas aeruginosa Evolution in the Cystic Fibrosis Lung

ABSTRACT Pulmonary infections caused by Pseudomonas aeruginosa are a recalcitrant problem in cystic fibrosis (CF) patients. While the clinical implications and long-term evolutionary patterns of these infections are well studied, we know little about the short-term population dynamics that enable this pathogen to persist despite aggressive antimicrobial therapy. Here, we describe a short-term population genomic analysis of 233 P. aeruginosa isolates collected from 12 sputum specimens obtained over a 1-year period from a single patient. Whole-genome sequencing and antimicrobial susceptibility profiling identified the expansion of two clonal lineages. The first lineage originated from the coalescence of the entire sample less than 3 years before the end of the study and gave rise to a high-diversity ancestral population. The second expansion occurred 2 years later and gave rise to a derived population with a strong signal of positive selection. These events show characteristics consistent with recurrent selective sweeps. While we cannot identify the specific mutations responsible for the origins of the clonal lineages, we find that the majority of mutations occur in loci previously associated with virulence and resistance. Additionally, approximately one-third of all mutations occur in loci that are mutated multiple times, highlighting the importance of parallel pathoadaptation. One such locus is the gene encoding penicillin-binding protein 3, which received three independent mutations. Our functional analysis of these alleles shows that they provide differential fitness benefits dependent on the antibiotic under selection. These data reveal that bacterial populations can undergo extensive and dramatic changes that are not revealed by lower-resolution analyses. IMPORTANCE Pseudomonas aeruginosa is a bacterial opportunistic pathogen responsible for significant morbidity and mortality in cystic fibrosis (CF) patients. Once it has colonized the lung in CF, it is highly resilient and rarely eradicated. This study presents a deep sampling examination of the fine-scale evolutionary dynamics of P. aeruginosa in the lungs of a chronically infected CF patient. We show that diversity of P. aeruginosa is driven by recurrent clonal emergence and expansion within this patient and identify potential adaptive variants associated with these events. This high-resolution sequencing strategy thus reveals important intraspecies dynamics that explain a clinically important phenomenon not evident at a lower-resolution analysis of community structure. Pseudomonas aeruginosa is a bacterial opportunistic pathogen responsible for significant morbidity and mortality in cystic fibrosis (CF) patients. Once it has colonized the lung in CF, it is highly resilient and rarely eradicated. This study presents a deep sampling examination of the fine-scale evolutionary dynamics of P. aeruginosa in the lungs of a chronically infected CF patient. We show that diversity of P. aeruginosa is driven by recurrent clonal emergence and expansion within this patient and identify potential adaptive variants associated with these events. This high-resolution sequencing strategy thus reveals important intraspecies dynamics that explain a clinically important phenomenon not evident at a lower-resolution analysis of community structure.

Analysis of the Cystic Fibrosis Lung Microbiota via Serial Illumina Sequencing of Bacterial 16S rRNA Hypervariable Regions

The characterization of bacterial communities using DNA sequencing has revolutionized our ability to study microbes in nature and discover the ways in which microbial communities affect ecosystem functioning and human health. Here we describe Serial Illumina Sequencing (SI-Seq): a method for deep sequencing of the bacterial 16S rRNA gene using next-generation sequencing technology. SI-Seq serially sequences portions of the V5, V6 and V7 hypervariable regions from barcoded 16S rRNA amplicons using an Illumina short-read genome analyzer. SI-Seq obtains taxonomic resolution similar to 454 pyrosequencing for a fraction of the cost, and can produce hundreds of thousands of reads per sample even with very high multiplexing. We validated SI-Seq using single species and mock community controls, and via a comparison to cystic fibrosis lung microbiota sequenced using 454 FLX Titanium. Our control runs show that SI-Seq has a dynamic range of at least five orders of magnitude, can classify >96% of sequences to the genus level, and performs just as well as 454 and paired-end Illumina methods in estimation of standard microbial ecology diversity measurements. We illustrate the utility of SI-Seq in a pilot sample of central airway secretion samples from cystic fibrosis patients.