Andras Bikov

Established methodological issues in electronic nose research: how far are we from using these instruments in clinical settings of breath analysis?

Electronic noses (e-noses) represent an easy and cheap method for exhaled volatile compound analysis. Various electronic noses are available which differ in material and thus analytical performance. In this review, we describe a wide range of electronic noses and summarize data on the methodological issues in electronic nose research. We also review studies which show the ability of electronic noses to distinguish pulmonary and extrapulmonary disorders from health.

A European Respiratory Society technical standard: exhaled biomarkers in lung disease

Breath tests cover the fraction of nitric oxide in expired gas (FENO), volatile organic compounds (VOCs), variables in exhaled breath condensate (EBC) and other measurements. For EBC and for FENO, official recommendations for standardised procedures are more than 10 years old and there is none for exhaled VOCs and particles. The aim of this document is to provide technical standards and recommendations for sample collection and analytic approaches and to highlight future research priorities in the field. For EBC and FENO, new developments and advances in technology have been evaluated in the current document. This report is not intended to provide clinical guidance on disease diagnosis and management. Clinicians and researchers with expertise in exhaled biomarkers were invited to participate. Published studies regarding methodology of breath tests were selected, discussed and evaluated in a consensus-based manner by the Task Force members. Recommendations for standardisation of sampling, analysing and reporting of data and suggestions for research to cover gaps in the evidence have been created and summarised. Application of breath biomarker measurement in a standardised manner will provide comparable results, thereby facilitating the potential use of these biomarkers in clinical practice. ERS technical standard: exhaled biomarkers in lung disease http://ow.ly/mAjr309DBOP

Evaluation of a partial genome screening of two asthma susceptibility regions using bayesian network based bayesian multilevel analysis of relevance.

Genetic studies indicate high number of potential factors related to asthma. Based on earlier linkage analyses we selected the 11q13 and 14q22 asthma susceptibility regions, for which we designed a partial genome screening study using 145 SNPs in 1201 individuals (436 asthmatic children and 765 controls). The results were evaluated with traditional frequentist methods and we applied a new statistical method, called Bayesian network based Bayesian multilevel analysis of relevance (BN-BMLA). This method uses Bayesian network representation to provide detailed characterization of the relevance of factors, such as joint significance, the type of dependency, and multi-target aspects. We estimated posteriors for these relations within the Bayesian statistical framework, in order to estimate the posteriors whether a variable is directly relevant or its association is only mediated. With frequentist methods one SNP (rs3751464 in the FRMD6 gene) provided evidence for an association with asthma (OR = 1.43(1.2–1.8); p = 3×10−4). The possible role of the FRMD6 gene in asthma was also confirmed in an animal model and human asthmatics. In the BN-BMLA analysis altogether 5 SNPs in 4 genes were found relevant in connection with asthma phenotype: PRPF19 on chromosome 11, and FRMD6, PTGER2 and PTGDR on chromosome 14. In a subsequent step a partial dataset containing rhinitis and further clinical parameters was used, which allowed the analysis of relevance of SNPs for asthma and multiple targets. These analyses suggested that SNPs in the AHNAK and MS4A2 genes were indirectly associated with asthma. This paper indicates that BN-BMLA explores the relevant factors more comprehensively than traditional statistical methods and extends the scope of strong relevance based methods to include partial relevance, global characterization of relevance and multi-target relevance.

Peripheral Th1/Th2/Th17/regulatory T-cell balance in asthmatic pregnancy

Asthma is a common chronic disease that may complicate pregnancy and a risk factor for complications; however, immunological mechanisms of the bilateral interactions between asthma and pregnancy are not fully understood. Healthy gestation is characterized by a sensitive balance of T(h)1/T(h)2/T(h)17/regulatory T (Treg) cells that may be altered in asthmatic pregnancy. The aim of this study was to describe the prevalence of these cell subsets in asthmatic compared with healthy pregnancy. The prevalence of T(h)1, T(h)2, T(h)17 and Treg lymphocytes was identified by cell surface and intracellular marker staining in blood samples of 24 healthy non-pregnant (HNP), 23 healthy pregnant (HP), 15 asthmatic non-pregnant (ANP) and 15 asthmatic pregnant (AP) women using flow cytometry. The T(h)1/T(h)2 cell ratio was decreased in both HP and ANP compared with HNP women; however, no further decrease was observed in the AP group. The T(h)17/Treg ratio was decreased in HP, but not in AP women, compared with HNP data. Healthy pregnancy increased Treg cell prevalence compared with HNP data (4.64% versus 2.98%; P < 0.05), and this pregnancy-induced elevation was absent in AP women (2.52% versus 4.64%; P < 0.05). T(h)17 cell prevalence was similar in the HP and HNP groups (2.78% versus 3.17%; P > 0.05). Asthma increased T(h)17 prevalence in non-pregnant patients (3.81% versus 3.17%; P < 0.05), and this asthma-specific increase of T(h)17 cell prevalence was also observed in AP patients (AP versus HP: 3.44% versus 2.78%; P < 0.05). The abnormal asthma-dependent T(h)17 elevation together with blunted Treg increase may play a role in the compromised immune tolerance characterizing asthmatic pregnancy.

Exhaled nitric oxide in pregnant healthy and asthmatic women

BACKGROUND Measurement of fractioned exhaled nitric oxide (FE(NO)) is useful for monitoring airway inflammation in asthma. Asthma is one of the most common diseases complicating pregnancy, and FE(NO) may be helpful for monitoring asthma in pregnancy. However, some physiological alterations of FE(NO) may be expected during healthy pregnancy due to vascular nitric oxide production. Until now no study assessed the level of FE(NO) in asthmatic pregnant patients. OBJECTIVE We aimed to assess the possible use and reproducibility of FE(NO) measurements in pregnant asthmatic women. We compared FE(NO) concentrations between four groups of subjects: healthy nonpregnant and pregnant females and asthmatic nonpregnant and pregnant patients. We also investigated the relationship between FE(NO) values and the level of asthma control in pregnant asthmatic patients. METHODS A total of 102 female subjects (35 healthy nonpregnant and 27 healthy pregnant females; 20 nonpregnant and 20 pregnant asthmatic women) were included in this cross-sectional study. Two FE(NO) measurements were performed in each subject using an electrochemical sensor based device (NIOX MINO, Aerocrine, Solna, Sweden). Data are given as median with range. RESULTS The repeatability of FE(NO) measurement was similar in pregnant and nonpregnant subjects. FE(NO) levels did not differ significantly between healthy pregnant versus nonpregnant subjects (16.0 [8, 31] vs. 16.0 [9, 35] ppb). FE(NO) levels were significantly increased in asthmatic women compared to healthy females (nonpregnant asthmatics: 38 [9, 54] ppb, p < 0.001 vs. healthy nonpregnant; pregnant asthmatic patients: 28 [10, 56] ppb; p < 0.05 vs. healthy pregnant). CONCLUSIONS FE(NO) level is not influenced by healthy pregnancy. In pregnant asthmatic patients FE(NO) level is elevated compared to healthy pregnant subjects and correlates with the level of asthma control. Further studies are required to assess the use of FE(NO) measurement to monitor asthma in this patient group.

Exhaled biomarker pattern is altered in children with obstructive sleep apnoea syndrome

OBJECTIVES Obstructive sleep apnoea syndrome (OSAS) is a common disorder in children, which is associated with enhanced inflammatory status. Inflammation-associated changes could be monitored by the assessment of exhaled biomarker profile. This study aimed to compare the exhaled biomarker profile in children with OSAS and habitual snorers. METHODS Eighteen children with OSAS (8 ± 2 years, mean ± SD) and ten non-OSAS subjects with habitual snoring (9 ± 2 years) were recruited. Exhaled breath was collected from the lower airways, processed using an electronic nose (E-nose) and analyzed off-line using principal component analysis, followed by discrimination analysis and logistic regression to build a receiver operating characteristic (ROC) curve. RESULTS Exhaled biomarker pattern of OSAS patients was discriminated from that of control subjects (p = 0.03, cross-validation accuracy: 64%), ROC curve analysis (area: 0.83) showed 78% sensitivity and 70% specificity. CONCLUSIONS The altered exhaled biomarker pattern in OSAS might reflect accelerated airway and/or systemic inflammation in diseased state. Breath pattern analysis by an E-nose can serve as a new tool to monitor inflammation in children with OSAS.

Exercise increases exhaled breath condensate cysteinyl leukotriene concentration in asthmatic patients

Background. Although the importance of cysteinyl leukotrienes (Cys-LTs) in exercise-induced bronchoconstriction (EIB) is supported by various sources of evidence, how the concentration of these mediators change during the development of EIB has not been investigated. Objectives. Our goal was to determine the effect of exercise on the concentration of airway Cys-LT in asthmatic patients by measuring Cys-LT in exhaled breath condensate (EBC). Methods. Seventeen atopic asthmatic patients with a previous history of EIB and six healthy volunteers were studied. Before and two times within 10 minutes after exercise challenge, FEV1 was measured and EBC was collected for Cys-LT measurement. Exhaled nitric oxide level, a marker of airway inflammation, was also determined at baseline. Results. Baseline Cys-LT level was higher in the asthmatic group versus healthy subjects (168 pg/mL /112–223/ vs. 77 pg/mL /36–119/, p = .03). EBC Cys-LT concentration increased in all asthmatic patients post-exercise (n = 17, p = .03), with the increase significantly greater in patients developing exercise-induced bronchospasm (n = 7, p = .03), whereas no change was observed in healthy controls (p = .59). The exercise-induced fall in FEV1 in asthmatics was related to the increase in EBC Cys-LT concentration (r = −0.40, p = .03). Conclusions. Our study shows that Cys-LT concentration of EBC is elevated minutes after physical exercise in asthmatic patients and strongly supports the concept that the release of this mediator is involved in the development of EIB.

Exhaled breath volatile alterations in pregnancy assessed with electronic nose

Context: Pregnancy-linked accelerated metabolism and oxidative stress may alter the exhaled volatile compound pattern (“breathprint”). Electronic noses can distinguish “breathprints” associated with different disorders. Objective: This is the first study assessing alterations in “breathprint” during gestation. Material and methods: 130 women participated in our study (78 pregnant vs. 52 non-pregnant). Breath samples were processed by an electronic nose and analyzed using principal component analysis. Results: Significant differences were found in exhaled breath pattern between pregnant and non-pregnant women (p = 0.001). Conclusion: Pregnancy-induced changes in exhaled gases need to be considered when pregnant women with respiratory disorders carry out breath tests.

Exhaled breath condensate pH decreases during exercise-induced bronchoconstriction

Exercise‐induced bronchoconstriction (EIB) is the temporary narrowing of the airways caused by physical exercise. Its exact pathophysiology is unclear; however, acute changes in airways pH may play a role. Exhaled breath condensate (EBC) pH was suggested as a surrogate indicator for airway acid–base status, but its value is also affected by volatile molecules and respiratory droplet dilution. The aim of the study was to assess changes in EBC pH during EIB.

Comparison of classification methods in breath analysis by electronic nose

Currently, many different methods are being used for pre-processing, statistical analysis and validation of data obtained by electronic nose technology from exhaled air. These various methods, however, have never been thoroughly compared. We aimed to empirically evaluate and compare the influence of different dimension reduction, classification and validation methods found in published studies on the diagnostic performance in several datasets. Our objective was to facilitate the selection of appropriate statistical methods and to support reviewers in this research area. We reviewed the literature by searching Pubmed up to the end of 2014 for all human studies using an electronic nose and methodological quality was assessed using the QUADAS-2 tool tailored to our review. Forty-six studies were evaluated regarding the range of different approaches to dimension reduction, classification and validation. From forty-six reviewed articles only seven applied external validation in an independent dataset, mostly with a case-control design. We asked their authors to share the original datasets with us. Four of the seven datasets were available for re-analysis. Published statistical methods for eNose signal analysis found in the literature review were applied to the training set of each dataset. The performance (area under the receiver operating characteristics curve (ROC-AUC)) was calculated for the training cohort (in-set) and after internal validation (leave-one-out cross validation). The methods were also applied to the external validation set to assess the external validity of the performance. Risk of bias was high in most studies due to non-random selection of patients. Internal validation resulted in a decrease in ROC-AUCs compared to in-set performance:  -0.15,-0.14,-0.1,-0.11 in dataset 1 through 4, respectively. External validation resulted in lower ROC-AUC compared to internal validation in dataset 1 (-0.23) and 3 (-0.09). ROC-AUCs did not decrease in dataset 2 (+0.07) and 4 (+0.04). No single combination of dimension reduction and classification methods gave consistent results between internal and external validation sets in this sample of four datasets. This empirical evaluation showed that it is not meaningful to estimate the diagnostic performance on a training set alone, even after internal validation. Therefore, we recommend the inclusion of an external validation set in all future eNose projects in medicine.