A survey of fungal microbiota in airways of healthy volunteer subjects from Puglia (Apulia), Italy.

BACKGROUND: The human respiratory tract represents the major portal of entry for numerous microorganisms, primarily those occurring as airborne particles such as viral and bacterial entities, or fungal spores. Microorganism characteristics coupled with the local host immune response will determine whether they will be cleared or adhere and colonize the airways leading to acute or chronic pulmonary disease. Like bacteria, fungi can cause severe lung diseases, but their infection rates are much lower. The lung microbiota is commonly sampled using relatively invasive bronchoscopic procedures. Exhaled breath condensate (EBC) collection offers a potentially less invasive alternative for lung microbiota sampling. This study tries to determine the composition of fungal communities in a cohort of healthy adult volunteer subjects from Puglia (Apulia), Italy. METHODS: Fungi diversity in 27 EBC samples collected from Italian adult volunteers was investigated using conventional microbiological culturing and DNA sequencing approach. RESULTS: Ten tested subjects (37,03%) turned out to present fungi in the EBC. We observed complex fungal communities, in which more than 10% of the isolated species are represented by Aspergillus sydowii (14,8%) and Cladosporium spp (11,11%). Three subjects that showed fungal presence in EBC have been diagnosed with a respiratory disease. CONCLUSIONS: We present a survey of an important scientific field in its early stages that is fungal contamination of airways of healthy subjects in a small geographic area. Furthermore, we interpreted our results to highlight the potential role of fungi in the context of respiratory diseases.

Analysis of the fungal microbiome in exhaled breath condensate of patients with asthma.

BACKGROUND: The presence of virus and bacteria in the airways of subjects with asthma is common and seems to be associated with a deterioration due to the disease. The microbiologic study of airways in asthma is foreseen by guidelines with induced sputum that is often ineffective and contraindicated in severe asthma. AIM: To analyze the fungal microbiome in the exhaled breath condensate (EBC) of subjects with asthma by evaluating a possible correlation with anthropometric and asthma severity data. METHODS: We enrolled 47 consecutive subjects with asthma (28 with atopic asthma and 19 with nonatopic asthma) and 20 controls. Enrolled subjects underwent EBC and sputum collection. Fungal microbiome was assessed by culture on EBC and sputum samples by using Czapek yeast extract agar. RESULTS: A fungal colonization in the EBC of 70% of enrolled subjects with asthma was detected (none detected in the controls). An overlap of fungal microbiome in EBC and sputum was observed (100% of overlap). Fungal colonization was higher in subjects without atopic, obesity, and severe and uncontrolled asthma. CONCLUSION: When considering the high morbidity and mortality of patients with severe asthma in whom we found an important fungal airways colonization, we support the use of the analysis of exhaled fungal microbiome in these subjects.

Mitochondrial DNA alteration in obstructive sleep apnea.

BACKGROUND: Obstructive Sleep Apnea (OSAS) is a disease associated with the increase of cardiovascular risk and it is characterized by repeated episodes of Intermittent Hypoxia (IH) which inducing oxidative stress and systemic inflammation. Mitochondria are cell organelles involved in the respiratory that have their own DNA (MtDNA). The aim of this study was to investigate if the increase of oxidative stress in OSAS patients can induce also MtDNA alterations. METHODS: 46 OSAS patients (age 59.27 ± 11.38; BMI 30.84 ± 3.64; AHI 36.63 ± 24.18) were compared with 36 control subjects (age 54.42 ± 6.63; BMI 29.06 ± 4.7; AHI 3.8 ± 1.10). In blood cells Content of MtDNA and nuclear DNA (nDNA) was measured in OSAS patients by Real Time PCR. The ratio between MtDNA/nDNA was then calculated. Presence of oxidative stress was evaluated by levels of Reactive Oxygen Metabolites (ROMs), measured by diacron reactive oxygen metabolite test (d-ROM test). RESULTS: MtDNA/nDNA was higher in patients with OSAS than in the control group (150.94 ± 49.14 vs 128.96 ± 45.8; p = 0.04), the levels of ROMs were also higher in OSAS subjects (329.71 ± 70.17 vs 226 ± 36.76; p = 0.04) and they were positively correlated with MtDNA/nDNA (R = 0.5, p < 0.01). CONCLUSIONS: In OSAS patients there is a Mitochondrial DNA damage induced by the increase of oxidative stress. Intermittent hypoxia seems to be the main mechanism which leads to this process.

Searching for Inflammatory and Oxidative Stress Markers Capable of Clustering Severe Asthma.

OBJECTIVE: Asthma inflammation may feature an imbalance between oxidative stress and antioxidant defenses. Oxidative stress induces propagation of airways inflammation and corticosteroid insensitivity contributing to poor asthma control, and frequent severe acute exacerbations. This study assessed inflammation and oxidative stress in severe asthmatic subjects and evaluated the possible correlations between inflammatory and oxidative stress markers investigated and asthma severity. MATERIAL AND METHOD: Fifty-three patients with severe asthma, 11 patients with mild-moderate asthma and 12 healthy subjects were enrolled and underwent fractional exhaled nitric oxide (FENO) analysis and blood and sputum count cell collection. The content of mitochondrial DNA (MtDNA) and nuclear DNA (nDNA) was measured in exhaled breath condensate (EBC) by Real Time PCR and the ratio between MtDNA/nDNA was calculated. We detected MtDNA/nDNA in the EBC of severe asthmatics. RESULTS: We found higher exhaled MtDNA/nDNA in severe asthmatics respectively compared to mild-moderate ones and to healthy controls (10.4±2.2 vs 7.9±2.5, p<0.05 and 10.4±2.2 vs 6.51±0.21, p<0.05). The level of exhaled MtDNA/nDNA was significantly higher in Non-T2 endotype severe asthmatics than T2 (14.07±10. 8 vs 6.5±5.5, p<0.05). CONCLUSION: Oxidative stress marker (MtDNA/nDNA) is increased significantly with asthma severity and may be useful for endotyping severe asthma.

Viral colonization in exhaled breath condensate of lung cancer patients: Possible role of EBV and CMV.

INTRODUCTION: Today, an increasing interest is being addressed to the viral etiology of lung tumors. As a consequence, research efforts are currently being directed to the identification of the new viruses involved in lung carcinogenesis toward which the screening programs could be directed. OBJECTIVES: The aim of this study was to investigate the airways colonization by the Epstein-Barr virus (EBV) and Citomegalovirus (CMV) in patients affected by lung cancer using, as a respiratory non-invasive sample, the exhaled breath condensate (EBC). METHODS: About 70 lung-cancer patients and 40 controls were enrolled. All subjects underwent bronchial brushing and EBC collection. EBV-DNA and CMV-DNA were evaluated in both samples by real-time PCR assay. RESULTS: They were able to detect EBV and CMV in the EBC. An increase of the EBV positivity in non-small cell lung cancer (NSCLC) patients compared with controls and of the CMV in advanced stages of lung cancer were observed. The association of the positivity of the cytology and the CMV test (in EBC or brushing) slightly increased the sensitivity of malignant diagnosis. CONCLUSION: EBV and CMV resulted detectable in the EBC. In consideration of the potential involvement of these viruses in lung cancer, which was confirmed in this study, future studies in this direction were supported.

Searching for Inflammatory and Oxidative Stress Markers Capable of Clustering Severe Asthma / Búsqueda de marcadores de estrés inflamatorio y oxidativo capaces de agrupar asma grave

Objective: Asthma inflammation may feature an imbalance between oxidative stress and antioxidant defenses. Oxidative stress induces propagation of airways inflammation and corticosteroid insensitivity contributing to poor asthma control, and frequent severe acute exacerbations. This study assessed inflammation and oxidative stress in severe asthmatic subjects and evaluated the possible correlations between inflammatory and oxidative stress markers investigated and asthma severity.Material and method: Fifty-three patients with severe asthma, 11 patients with mild-moderate asthma and 12 healthy subjects were enrolled and underwent fractional exhaled nitric oxide (FENO) analysis and blood and sputum count cell collection. The content of mitochondrial DNA (MtDNA) and nuclear DNA (nDNA) was measured in exhaled breath condensate (EBC) by Real Time PCR and the ratio between MtDNA/nDNA was calculated. We detected MtDNA/nDNA in the EBC of severe asthmatics.Results: We found higher exhaled MtDNA/nDNA in severe asthmatics respectively compared to mild-moderate ones and to healthy controls (10.4±2.2 vs 7.9±2.5, p<0.05 and 10.4±2.2 vs 6.51±0.21, p<0.05). The level of exhaled MtDNA/nDNA was significantly higher in Non-T2 endotype severe asthmatics than T2 (14.07±10. 8 vs 6.5±5.5, p<0.05).Conclusion: Oxidative stress marker (MtDNA/nDNA) is increased significantly with asthma severity and may be useful for endotyping severe asthma. (AU)

Objetivo: La inflamación en el asma puede presentar un desequilibrio entre el estrés oxidativo y las defensas antioxidantes. El estrés oxidativo induce la propagación de la inflamación de las vías aéreas y la insensibilidad a los corticosteroides, lo que contribuye a un control deficiente del asma y a frecuentes exacerbaciones agudas graves. Este estudio evaluó la inflamación y el estrés oxidativo en sujetos asmáticos graves y estudió las posibles correlaciones entre los marcadores de estrés inflamatorio y oxidativo investigados y la gravedad del asma.Material y método: Se incluyó a 53 pacientes con asma grave, 11 pacientes con asma leve a moderada y 12 sujetos sanos, a los que se les realizó un análisis de fracción exhalada de óxido nítrico (FeNO) y un recuento celular del esputo y de sangre. Se midió el contenido de ADN mitocondrial (ADNmt) y ADN nuclear (ADNn) en el condensado de aire exhalado (CAE) mediante PCR en tiempo real y se calculó la ratio ADNmt/ADNn. Detectamos ADNmt/ADNn en el CAE de los asmáticos graves.Resultados: Encontramos unos niveles más altos de ADNmt/ADNn exhalados en los asmáticos graves en comparación con los leves moderados y los controles sanos (respectivamente, 10,4±2,2 frente a 7,9±2,5, p<0,05 y 10,4±2,2 frente a 6,51±0,21, p<0,05). El nivel de ADNmt/ADNn exhalado fue significativamente mayor en los asmáticos graves de endotipo no-T2 que en los T2 (14,07±10,8 frente a 6,5±5,5, p<0,05).Conclusión: El marcador de estrés oxidativo (ADNmt/ADNn) aumenta significativamente con la gravedad del asma y puede ser útil para endotipar el asma grave. (AU)

Is the exhaled breath temperature in lung cancer influenced by airways neoangiogenesis or by inflammation?

Recently the exhaled breath temperature (EBT) was seen to increase in non-small cell lung cancer and was subsequently proposed as a possible non-invasive tool for its diagnosis. The need for further studies that confirm the previous findings and support the potential scope of this method underlies the main aim of this study that seeks to explore the pathogenic mechanisms determining the EBT in lung cancer. We enrolled 44 consecutive patients with a radiological suspicion of lung cancer and ten healthy non-smoker volunteers, after which their EBT was measured. On the same day, the subjects underwent breath condensate collection for the measurement of leukotriene (LTB)-4 and of the vascular endothelial growth factor (VEGF), the former being a marker of airways inflammation and the latter of neoangiogenesis. We confirmed the presence of a higher EBT in lung cancer patients compared to the controls. The multiple linear regression model showed that the exhaled VEGF was the only predictor of elevations of EBT. In conclusion, it can be stated that for the first time in this study, we have shown that EBT is higher in subjects with lung cancer and that the airways angiogenesis drives the increase in EBT in lung cancer. Moreover, the study suggests the potential for the use of EBT in monitoring the lung cancer progression, although the implementation of more in-depth studies to verify this result is recommended.

A fish-specific transposable element shapes the repertoire of p53 target genes in zebrafish.

Transposable elements, as major components of most eukaryotic organisms' genomes, define their structural organization and plasticity. They supply host genomes with functional elements, for example, binding sites of the pleiotropic master transcription factor p53 were identified in LINE1, Alu and LTR repeats in the human genome. Similarly, in this report we reveal the role of zebrafish (Danio rerio) EnSpmN6_DR non-autonomous DNA transposon in shaping the repertoire of the p53 target genes. The multiple copies of EnSpmN6_DR and their embedded p53 responsive elements drive in several instances p53-dependent transcriptional modulation of the adjacent gene, whose human orthologs were frequently previously annotated as p53 targets. These transposons define predominantly a set of target genes whose human orthologs contribute to neuronal morphogenesis, axonogenesis, synaptic transmission and the regulation of programmed cell death. Consistent with these biological functions the orthologs of the EnSpmN6_DR-colonized loci are enriched for genes expressed in the amygdala, the hippocampus and the brain cortex. Our data pinpoint a remarkable example of convergent evolution: the exaptation of lineage-specific transposons to shape p53-regulated neuronal morphogenesis-related pathways in both a hominid and a teleost fish.