Abstract
Cystic fibrosis patients are highly susceptible to infections with non-tuberculous mycobacteria. Especially Mycobacterium abscessus infections are common but reliable diagnosis is hampered by non-specific clinical symptoms and insensitive mycobacterial culture. In the present study we established novel methods for rapid detection and immune characterization of Mycobacterium abscessus infection in cystic fibrosis patients. We performed Mycobacterium abscessus specific DNA-strip- and quantitative PCR-based analyses of non-cultured sputum samples to detect and characterize Mycobacterium abscessus infections. Concomitantly in vitro T-cell reactivation with purified protein derivatives (PPDs) from different mycobacterial species was used to determine Mycobacterium abscessus specific T-cell cytokine expression of infected cystic fibrosis patients. Four of 35 cystic fibrosis patients (11.4%) were Mycobacterium abscessus culture positive and showed concordant DNA-strip-test results. Quantitative PCR revealed marked differences of mycobacterial burden between cystic fibrosis patients and during disease course. Tandem-repeat analysis classified distinct Mycobacterium abscessus strains of infected cystic fibrosis patients and excluded patient-to-patient transmission. Mycobacterium abscessus specific T-cells were detected in the blood of cystic fibrosis patients with confirmed chronic infection and a subgroup of patients without evidence of Mycobacterium abscessus infection. Comparison of cytokine expression and phenotypic markers revealed increased proportions of CD40L positive T-cells that lack Interleukin-2 expression as a marker for chronic Mycobacterium abscessus infections in cystic fibrosis patients. Direct sputum examination enabled rapid diagnosis and quantification of Mycobacterium abscessus in cystic fibrosis patients. T-cell in vitro reactivation and cytokine expression analyses may contribute to diagnosis of chronic Mycobacterium abscessus infection.
Highlights
Mutations on both alleles of the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) are the genetic cause of Cystic Fibrosis (CF), the most common single-gene caused disease in Caucasians [1]
Mycobacterium (M.) avium complex (MAC) and M. abscessus complex (MABSC) are the non-tuberculous mycobacterial species most commonly detected in the sputum of CF patients [6]
We analyzed the sensitivity of DNA-strip test in sputum samples by establishing a MABSC-specific rpoB-Quantitative PCR (qPCR)
Summary
Mutations on both alleles of the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) are the genetic cause of Cystic Fibrosis (CF), the most common single-gene caused disease in Caucasians [1]. Non-tuberculous mycobacteria are rarely pathogenic for immunocompetent individuals but frequently colonize vulnerable pulmonary epithelia of CF patients [3]. Mycobacterium (M.) avium complex (MAC) and M. abscessus complex (MABSC) are the non-tuberculous mycobacterial species most commonly detected in the sputum of CF patients [6]. MABSC has been found to be the most frequent ‘rapid growing’ human pathogenic mycobacterial species [8]. Most intriguingly MABSC is able to cause persistent lung disease characterized by development of caseous lesions, a hallmark of human tuberculosis [9]. Evidence for direct transmission of MABSC between CF patients has been found [10]. The possibility of direct transmission renders early detection and characterization of MABSC in CF patients crucial
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