Cystic fibrosis - an example of personalized and precision medicine.

Cystic fibrosis (CF) is a severe, monogenic, autosomal recessive disease caused by mutations in the CFTR (cystic fibrosis transmembrane regulator) gene, where disturbed chloride and bicarbonate transportation in epithelial cells results in a multiorgan disease with primarily pulmonary infections and pancreatic insufficiency. In 1968, the Copenhagen CF Center was established, and centralized care of CF patients with monthly control was introduced. Close monitoring and treatment of Pseudomonas lung infection as well as segregation of patients with different infection status improved the clinical outcome as well as survival. Prophylactic basic treatment as well as infection treatments follow specific algorithms. A variety of comorbidities have all along the pulmonary infection control necessitated personalized care, adjusted to the patients' phenotype. With the introduction of CFTR modulators, the treatment has shifted from prophylactic, symptomatic type toward a new era of precision medicine targeting the basic defect according to the patients' CFTR genotype. Future directions will focus on further improvement of the CFTR modulators and gene therapy, as well as modifier genes and CF phenotype.

Comparing the harmful effects of nontuberculous mycobacteria and Gram negative bacteria on lung function in patients with cystic fibrosis.

BACKGROUND: To better understand the relative effects of infection with nontuberculous mycobacteria and Gram negative bacteria on lung function decline in cystic fibrosis, we assessed the impact of each infection in a Danish setting. METHODS: Longitudinal registry study of 432 patients with cystic fibrosis contributing 53,771 lung function measures between 1974 and 2014. We used a mixed effects model with longitudinally structured correlation, while adjusting for clinically important covariates. RESULTS: Infections with a significant impact on rate of decline in %FEV1 were Mycobacterium abscessus complex with -2.22% points per year (95% CI -3.21 to -1.23), Burkholderia cepacia complex -1.95% (95% CI -2.51 to -1.39), Achromobacterxylosoxidans -1.55% (95% CI -2.21 to -0.90), and Pseudomonas aeruginosa -0.95% (95% CI -1.24 to -0.66). Clearing M. abscessus complex was associated with a change to a slower decline, similar in magnitude to the pre-infection slope. CONCLUSIONS: In a national population we have demonstrated the impact on lung function of each chronic CF pathogen. M. abscessus complex was associated with the worst impact on lung function. Eradication of M. abscessus complex may significantly improve lung function.

Denitrification by cystic fibrosis pathogens - Stenotrophomonas maltophilia is dormant in sputum.

OBJECTIVE: Chronic Pseudomonas aeruginosa lung infection is the most severe complication for cystic fibrosis (CF) patients. Infected endobronchial mucus of CF patients contains anaerobic zones mainly due to the respiratory burst of polymorphonuclear leukocytes. We have recently demonstrated ongoing denitrification in sputum from patients infected with P. aeruginosa. Therefore we aimed to investigate, whether the pathogenicity of several known CF pathogens is correlated to their ability to perform denitrification. METHODS: We measured denitrification with N(2)O microsensors in concert with anaerobic growth measurements by absorbance changes and colony counting in isolates from 32 CF patients chronically infected with the highly pathogenic bacteria P. aeruginosa, Achromobacter xylosoxidans, Burkholderia multivorans or the less pathogenic bacterium Stenotrophomonas maltophilia. Consumption of NO(3)(-) and NO(2)(-) was estimated by the Griess Assay. All isolates were assayed during 2 days of incubation in anaerobic LB broth with NO(3)(-) or NO(2)(-). PNA FISH staining of 16S rRNA was used to estimate the amount of ribosomes per bacterial cells and thereby the in situ growth rate of S. maltophilia in sputum. RESULTS: Supplemental NO(3)(-) caused increased production of N(2)O by P. aeruginosa, A. xylosoxidans and B. multivorans and increased growth for all pathogens. Growth was, however, lowest for S. maltophilia. NO(3)(-) was metabolized by all pathogens, but only P. aeruginosa was able to remove NO(2)(-). S. maltophilia had limited growth in sputum as seen by the weak PNA FISH staining. CONCLUSIONS: All four pathogens were able to grow anaerobically by NO(3)(-) reduction. Denitrification as demonstrated by N(2)O production was, however, not found in S. maltophilia isolates. The ability to perform denitrification may contribute to the pathogenicity of the infectious isolates since complete denitrification promotes faster anaerobic growth. The inability of S. maltophilia to proliferate by denitrification and therefore grow in the anaerobic CF sputum may explain its low pathogenicity in CF patients.

Nitrous oxide production in sputum from cystic fibrosis patients with chronic Pseudomonas aeruginosa lung infection.

Chronic lung infection by Pseudomonas aeruginosa is the major severe complication in cystic fibrosis (CF) patients, where P. aeruginosa persists and grows in biofilms in the endobronchial mucus under hypoxic conditions. Numerous polymorphonuclear leukocytes (PMNs) surround the biofilms and create local anoxia by consuming the majority of O2 for production of reactive oxygen species (ROS). We hypothesized that P. aeruginosa acquires energy for growth in anaerobic endobronchial mucus by denitrification, which can be demonstrated by production of nitrous oxide (N2O), an intermediate in the denitrification pathway. We measured N2O and O2 with electrochemical microsensors in 8 freshly expectorated sputum samples from 7 CF patients with chronic P. aeruginosa infection. The concentrations of NO3(-) and NO2(-) in sputum were estimated by the Griess reagent. We found a maximum median concentration of 41.8 µM N2O (range 1.4-157.9 µM N2O). The concentration of N2O in the sputum was higher below the oxygenated layers. In 4 samples the N2O concentration increased during the initial 6 h of measurements before decreasing for approximately 6 h. Concomitantly, the concentration of NO3(-) decreased in sputum during 24 hours of incubation. We demonstrate for the first time production of N2O in clinical material from infected human airways indicating pathogenic metabolism based on denitrification. Therefore, P. aeruginosa may acquire energy for growth by denitrification in anoxic endobronchial mucus in CF patients. Such ability for anaerobic growth may be a hitherto ignored key aspect of chronic P. aeruginosa infections that can inform new strategies for treatment and prevention.

Pseudomonas aeruginosa biofilms in the respiratory tract of cystic fibrosis patients.

The present study was undertaken to investigate the appearance and location of Pseudomonas aeruginosa in the cystic fibrosis (CF) lung and in sputum. Samples include preserved tissues of CF patients who died due to chronic P. aeruginosa lung infection prior to the advent of intensive antibiotic therapy, explanted lungs from 3 intensively treated chronically P. aeruginosa infected CF patients and routine sputum from 77 chronically P. aeruginosa infected CF patients. All samples were investigated microscopically using hematoxylin-eosin (HE), Gram and alcian-blue stain, PNA FISH and immunofluorescence for alginate.Investigation of the preserved tissues revealed that prior to aggressive antibiotic therapy, P. aeruginosa infection and destruction of the CF lung correlated with the occurrence of mucoid (alginate) bacteria present in aggregating structures surrounded by pronounced polymorphonuclear-leukocyte (PMN) inflammation in the respiratory zone (9/9). Non-mucoid bacteria were not observed here, and rarely in the conductive zone (1/9). However, in the explanted lungs, the P. aeruginosa aggregates were also mucoid but in contrast to the autopsies, they were very rare in the respiratory zone but abundant in the sputum of the conductive zone (3/3), which also contained abundances of PMNs (3/3). Non-mucoid and planktonic P. aeruginosa were also observed here (3/3).In conclusion, the present intensive antibiotic therapy of chronic P. aeruginosa infections, at the Copenhagen CF Centre, seems to restrain but not eradicate the bacteria from the conductive zone, whereas the remaining healthy respiratory zone appears to be protected, for a long period, from massive biofilm infection. This strongly suggests that the conductive zone serves as a bacterial reservoir where the bacteria are organized in mucoid biofilms within the mucus, protected against antibiotics and host defenses.

Validation of the Danish version of the revised cystic fibrosis quality of life questionnaire in adolescents and adults (CFQ-R14+).

BACKGROUND: Quality of life is an important parameter in the evaluation of quality and outcome of health care and treatment, especially in patients with chronic disorders. The aim of this study was to assess the validity and reliability of the Danish version of the revised disease-specific health-related quality of life questionnaire for adolescents and adults with cystic fibrosis (CFQ-R14+). METHODS: A total of 196 cystic fibrosis (CF) patients completed the CFQ-R14+ (response rate 71%). Forced expiratory volume in 1 s in percentage of predicted (FEV(1)%) and body mass index (BMI) were included as measures of health status. RESULTS: Internal consistency coefficients ranged from 0.54 to 0.95. Eight out of the twelve scales had alpha coefficients above 0.7. Test-retest correlations ranged from 0.42 to 0.88 and they were significant in eight scales. All the CFQ-R+14 scales except the digestive symptoms scale discriminated significantly (p<0.05) between patients with mild, moderate, and severe disease. Nine out of the twelve scales discriminated significantly (p<0.05) between nourished (BMI> or =19) and malnourished (BMI<19) patients. Significant differences between participants and non-responders were found for age, sex and FEV(1) (higher age, more males and lower FEV(1) among non-responders). All of the scales met standards for floor effects (<15% of the responders with the lowest score) but five of the scales failed to meet standards for ceiling effects (>15% of the responders with the highest score). CONCLUSION: The Danish CFQ-R14+ is a reliable and valid instrument for measuring the health-related quality of life in Danish adolescents and adults with CF, though with the exception from a few of its subscales.

Long-term azitromycin treatment of cystic fibrosis patients with chronic Pseudomonas aeruginosa infection; an observational cohort study.

BACKGROUND: In cystic fibrosis (CF), chronic endobronchial infection with Pseudomonas aeruginosa is a serious complication. Macrolides can increase lung function and weight in patients, and reduce exacerbations. METHODS: In 2001, we introduced long-term, low-dose azithromycin (AZ) treatment as an integral part of our routine treatment of these patients. Our study is an observational cohort study of all CF patients with chronic P. aeruginosa infection in our CF center comparing clinical parameters of the patients 12 months prior to treatment with the same values during 12 months of treatment. RESULTS: 45 patients (27 men, median age 29 years) completed 1-year treatment. Median weight increased from 63.1 kg in the pre-treatment period to 63.9 kg during treatment (p=0.01). Median slope of decline in lung function increased from pre-treatment FEV1 -4.1% and FVC -3.0% to +0.8% (p<0.001) and +1.6% (p=0.01), respectively. 90% of sputum samples contained mucoid P. aeruginosa before treatment, decreasing to 81% during treatment (p=0.003). Median CRP decreased from 6.2 mmol/l to 5.8 mmol/l (ns). CONCLUSION: Long-term, low-dose AZ treatment in adult CF patients with chronic P. aeruginosa infection is safe and reduces the decline in lung function, increases weight, and reduces the percentage of mucoid strains of P. aeruginosa in sputum samples.