BackgroundChronic obstructive pulmonary disease (COPD) is characterized by chronic inflammation and progressive decline in pulmonary function. Neutrophil-to-lymphocyte ratio (NLR), YKL-40 and calprotectin are biomarkers of inflammation and predict mortality in patients with different inflammatory diseases. We aimed to investigate the correlation between levels of these three biomarkers and neutrophil granulocyte and lymphocyte count in patients with moderate to very severe COPD stratified by use of systemic glucocorticoids. Furthermore, we studied the ability of these biomarkers to predict all-cause mortality.Methods386 patients with moderate to very severe COPD were followed prospectively for 10 years. Patients were divided into two groups according to systemic glucocorticoid use at baseline. Correlations between biomarkers were assessed by Spearman’s Rho, and mortality was evaluated in uni- and multivariate Cox regression analyses with hazard ratios (HR) and 95% confidence intervals (CI).ResultsPlasma calprotectin was positively correlated with neutrophil granulocyte count and NLR. No significant association was found between plasma YKL-40 and the cellular biomarkers, irrespective of glucocorticoid treatment. In the group not treated with systemic glucocorticoids, plasma calprotectin [HR 1.002 (95% CI 1.000 – 1.004)], NLR [HR 1.090 (1.036 – 1.148)] and lymphocyte count [HR 0.667 (0.522 – 0.851)] were significantly associated with higher mortality. In the group treated with systemic glucocorticoids, higher plasma YKL-40 was significantly associated with mortality in univariate Cox regression analysis [HR 1.006 (1.003 – 1.008)].ConclusionsCalprotectin was related to neutrophil granulocyte count and NLR in patients with moderate to very severe COPD in stable phase and not in treatment with systemic glucocorticoids. Lymphopenia, higher plasma calprotectin and higher NLR were independent predictors of increased all-cause mortality in this group. Our data also suggests that treatment with systemic glucocorticoids has a significant impact on the ability of inflammatory biomarkers to predict all-cause mortality.Trial registrationClinicalTrials.gov NCT00132860.

BackgroundRecent studies have found vitamin D (25-OHD) deficiency and insufficiency to be common among patients with COPD. Serum level of 25-OHD seems to correlate to pulmonary function, COPD disease staging, and increased susceptibility to respiratory infections. We wanted to investigate whether vitamin D deficiency or insufficiency was associated with mortality rate in patients suffering from advanced COPD.Methods25-OHD serum levels were measured in 462 patients suffering from moderate to very severe COPD. Patients were stratified into three groups according to serum levels of 25-OHD. Outcome measure was mortality in a 10 year follow-up period. Kaplan-Meier curves (KM) were plotted and mortality hazard ratios (HR) were calculated using Cox Proportional Hazard regression (Cox PH).ResultsSerum 25-OHD deficiency and insufficiency were prevalent. We were unable to demonstrate any association between baseline serum levels of 25-OHD and mortality rate. We found an association between mortality and age [HR 1.05 (CI 95%: 1.03–1.06)], Charlson score [HR 1.49 (CI 95%: 1.06–2.09)], increasing neutrophil count [HR 1.05 (CI 95%: 1.02–1.09)], severe [HR 1.41 (CI 95%: 1.06–1.86)]/very severe COPD [HR 2.19 (CI 95%: 1.58–3.02)] and a smoking history of more than 40 pack years [HR 1.27 (CI 95%: 1.02–1.70)].ConclusionsSerum level of 25-OHD does not seem to be associated with mortality rate, suggesting no or only a minor role of 25-OHD in disease progression in patients with moderate to very severe COPD.

Acanthamoeba is a free-living amoeba of extensive genetic diversity. It may cause infectious keratitis (IK), which can also be caused by bacteria, fungi, and viruses. High diagnostic sensitivity is essential to establish an early diagnosis of Acanthamoeba-associated keratitis. Here, we investigated the applicability of next-generation sequencing (NGS)-based ribosomal gene detection and differentiation (16S-18S) compared with specific real-time PCR for detection of Acanthamoeba. Two hundred DNAs extracted from corneal scrapings and screened by Acanthamoeba-specific real-time PCR were analyzed using an in-house 16S-18S NGS assay. Of these, 24 were positive using specific real-time PCR, 21 of which were positive using the NGS assay. Compared with real-time PCR; the specificity and sensitivity of the NGS assay were 100% and 88%, respectively. Genotypes identified by the NGS assay included T4 (n = 19) and T6 (n = 2). Fungal and bacterial species of potential clinical relevance were identified in 31 of the samples negative for Acanthamoeba, exemplified by Pseudomonas aeruginosa (n = 11), Moraxella spp. (n = 6), Staphylococcus aureus (n = 2), Fusarium spp. (n = 4), and Candida albicans (n = 1). Conclusively, the 16S-18S assay was slightly less sensitive than real-time PCR in detecting Acanthamoeba-specific DNA in corneal scrapings. Robust information on genotype was provided by the NGS assay, and other pathogens of potential clinical relevance were identified in 16% of the samples negative for Acanthamoeba. NGS-based detection of ribosomal genes in corneal scrapings could be an efficient screening method for detecting non-viral causes of IK, including Acanthamoeba.

BackgroundChronic obstructive pulmonary disease (COPD) is hallmarked by inflammatory processes and a progressive decline of lung function. YKL-40 is a potential biomarker of inflammation and mortality in patients suffering from inflammatory lung disease, but its prognostic value in patients with COPD remains unknown. We investigated whether high plasma YKL-40 was associated with increased mortality in patients with moderate to very severe COPD.MethodsFour hundred and ninety-three patients with moderate to very severe COPD were followed prospectively for up to 10 years. Patients were divided into two groups according to plasma YKL-40: concentration higher than the 75th percentile for age-matched healthy subjects (i.e. high levels) and normal levels. Outcome was overall survival (OS) and was evaluated in uni- and multivariate proportional hazards Cox regression analyses and adjusted for factors affecting mortality.ResultsMedian plasma YKL-40 was increased in patients with COPD (81 ng/ml, p < 0.001) compared to healthy subjects (40 ng/ml). Patients with high plasma YKL-40 had a hazard ratio (HR) of 1.42 (95% CI: 1.15–1.75, p = 0.001) for all-cause mortality. Multivariate analysis showed that YKL-40 (HR 1.38; 95% CI: 1.11–1.72, p = 0.004), age (HR 1.05; 95% CI: 1.03–1.06, p < 0.0001), Severe COPD (HR 1.35; 95 CI: 1.03-1.76, p = 0.03) very severe COPD (HR 2.19; 95% CI: 1.60 - 2.99 < 0.0001), neutrophil granulocyte count (HR 1.05; 95% CI: 1.01-1.08, p = 0.01), and a smoking history of > 40 years (HR 1.38; 95% CI: 1.11-1.71, p = 0.003) were independent prognostic markers of OS.ConclusionHigh plasmaYKL-40 is associated with increased mortality in patients with moderate to very severe COPD, suggesting a role for YKL-40 as a potential biomarker of mortality in this patient group.Trial registrationClinicalTrials.gov: NCT00132860.

Calprotectin comprises more than 45% of the cytosolic content of neutrophil granulocytes. Because pathogenesis, disease activity and disease progression in COPD are believed to be partly dependent of neutrophil driven inflammation we decided to investigate whether plasma level of calprotectin (p-calprotectin) was associated with all-cause mortality in patients with COPD. We measured p-calprotectin in blood samples from 460 patients with moderate to very severe COPD in stable phase. Patients were stratified into three groups according to p-calprotectin level. Outcome measure was all-cause mortality. Analyses were adjusted for factors known to influence mortality using a Cox regression analysis. We found a time dependent correlation between p-calprotectin levels and mortality during the first 5 years of follow-up. Increasing levels of p-calprotectin were associated with concomitant increases in mortality from HR 1.56 (CI 95%: 1.03 -2.38) at calprotectin between 100 -200 ng/ml to HR 2.02 (CI 95%: 1.27-3.19) at calprotectin >200 ng/ml. P-calprotectin could be a useful marker of all-cause mortality in patients suffering from moderate to very severe COPD.