Predictors of persistent poor control and validation of ASSESS score : Longitudinal 5-year follow-up of severe asthma cohort

Funding Information: This research is supported by the Singapore Ministry of Health’s National Medical Research Council under its Transition Award Grant (grant no. MOH-001275-00 to P.Y.T.).Peer reviewe

Distinct 'Immuno-Allertypes' of Disease and High Frequencies of Sensitisation in Non-Cystic-Fibrosis Bronchiectasis

Rationale: Allergic sensitization is associated with poor clinical outcomes in asthma, chronic obstructive pulmonary disease, and cystic fibrosis; however, its presence, frequency, and clinical significance in non–cystic fibrosis bronchiectasis remain unclear. Objectives: To determine the frequency and geographic variability that exists in a sensitization pattern to common and specific allergens, including house dust mite and fungi, and to correlate such patterns to airway immune-inflammatory status and clinical outcomes in bronchiectasis. Methods: Patients with bronchiectasis were recruited in Asia (Singapore and Malaysia) and the United Kingdom (Scotland) (n = 238), forming the Cohort of Asian and Matched European Bronchiectasis, which matched recruited patients on age, sex, and bronchiectasis severity. Specific IgE response against a range of common allergens was determined, combined with airway immune-inflammatory status and correlated to clinical outcomes. Clinically relevant patient clusters, based on sensitization pattern and airway immune profiles (“immunoallertypes”), were determined. Measurements and Main Results: A high frequency of sensitization to multiple allergens was detected in bronchiectasis, exceeding that in a comparator cohort with allergic rhinitis (n = 149). Sensitization was associated with poor clinical outcomes, including decreased pulmonary function and more severe disease. “Sensitized bronchiectasis” was classified into two immunoallertypes: one fungal driven and proinflammatory, the other house dust mite driven and chemokine dominant, with the former demonstrating poorer clinical outcome. Conclusions: Allergic sensitization occurs at high frequency in patients with bronchiectasis recruited from different global centers. Improving endophenotyping of sensitized bronchiectasis, a clinically significant state, and a “treatable trait” permits therapeutic intervention in appropriate patients, and may allow improved stratification in future bronchiectasis research and clinical trials.Ministry of Education (MOE)Ministry of Health (MOH)National Medical Research Council (NMRC)Published versionSupported by the Singapore Ministry of Health’s National Medical Research Council under its Transition Award NMRC/TA/0048/2016 (S.H.C.) and Changi General Hospital Research grant CHF2016.03-P (T.B.L.). The work performed at NUS was supported by the Singapore Ministry of Education Academic Research Fund, SIgN, and National Medical Research Council grants N-154-000-038-001, R-154-000-404-112, R-154-000-553-112, R-154-000-565-112, R-154-000-630-112, R-154-000-A08-592, R-154-000-A27-597, SIgN-06-006, SIgN-08-020, and NMRC/1150/2008 (F.T.C.); J.D.C. is supported by the GSK/British Lung Foundation Chair of Respiratory Research

Clinical, immunological and microbiological phenotypes of chronic obstructive pulmonary disease (COPD) in ethnic Chinese populations in South-East Asia

Chronic Obstructive Pulmonary Disease (COPD) is of increasing prevalence across Asia. COPD in Asian populations remains under-researched, and most clinical trials of COPD therapeutics exclude Asians. Therefore, a major knowledge gap exists on whether specific clinical, immunological and microbiological phenotypes of COPD exist in Asians, with a particular lack of data evident in South-East Asia. Here, employing clinical, statistical, molecular and sequencing approaches, I identify unique clinical phenotypes of Asian COPD that associate with mortality including a novel ‘ex-tuberculosis’ group. Immunologically, fungal sensitisation relates to poor clinical outcomes in COPD, and environmental exposure serves as a key reservoir of fungal allergens. Distinct airway mycobiome profiles that vary by geographic region, and with association to frequent exacerbations and mortality are evident in South-East Asians with COPD. Collectively, my novel findings will improve risk stratification in Asians with COPD, and promote the development of specific clinical guidelines focused on Asian COPD.Doctor of Philosoph

Applying next-generation sequencing and multi-omics in chronic obstructive pulmonary disease

Microbiomics have significantly advanced over the last decade, driven by the widespread availability of next-generation sequencing (NGS) and multi-omic technologies. Integration of NGS and multi-omic datasets allow for a holistic assessment of endophenotypes across a range of chronic respiratory disease states, including chronic obstructive pulmonary disease (COPD). Valuable insight has been attained into the nature, function, and significance of microbial communities in disease onset, progression, prognosis, and response to treatment in COPD. Moving beyond single-biome assessment, there now exists a growing literature on functional assessment and host-microbe interaction and, in particular, their contribution to disease progression, severity, and outcome. Identifying specific microbes and/or metabolic signatures associated with COPD can open novel avenues for therapeutic intervention and prognosis-related biomarkers. Despite the promise and potential of these approaches, the large amount of data generated by such technologies can be challenging to analyze and interpret, and currently, there remains a lack of standardized methods to address this. This review outlines the current use and proposes future avenues for the application of NGS and multi-omic technologies in the endophenotyping, prognostication, and treatment of COPD.Ministry of Health (MOH)National Medical Research Council (NMRC)Published versionThis research is supported by the Singapore Ministry of Health’s National Medical Research Council under its Clinician Scientist Award (MOH-000710) (S.H.C) and the Open Fund-Individual Research Grant (MOH-000955) (S.H.C)

Clinical Aspergillus Signatures in COPD and Bronchiectasis

Pulmonary mycoses remain a global threat, causing significant morbidity and mortality. Patients with airways disease, including COPD and bronchiectasis, are at increased risks of pulmonary mycoses and its associated complications. Frequent use of antibiotics and corticosteroids coupled with impaired host defenses predispose patients to fungal colonization and airway persistence, which are associated with negative clinical consequences. Notably, Aspergillus species remain the best-studied fungal pathogen and induce a broad spectrum of clinical manifestations in COPD and bronchiectasis ranging from colonization and sensitization to more invasive disease. Next-generation sequencing (NGS) has gained prominence in the field of respiratory infection, and in some cases is beginning to act as a viable alternative to traditional culture. NGS has revolutionized our understanding of airway microbiota and in particular fungi. In this context, it permits the identification of the previously unculturable, fungal composition, and dynamic change within microbial communities of the airway, including potential roles in chronic respiratory disease. Furthermore, inter-kingdom microbial interactions, including fungi, in conjunction with host immunity have recently been shown to have important clinical roles in COPD and bronchiectasis. In this review, we provide an overview of clinical Aspergillus signatures in COPD and bronchiectasis and cover the current advances in the understanding of the mycobiome in these disease states. The challenges and limitations of NGS will be addressed

Optimisation and benchmarking of targeted amplicon sequencing for mycobiome analysis of respiratory specimens

(1) Background: Firm consensus has yet to be established in relation to taxonomic classification and primer choice in targeted amplicon sequencing of the mycobiome. While the nuclear ribosomal internal transcribed spacer (ITS) region are recognized as the formal fungal taxonomic barcode, appraisal of different ITS sub-regions and the influence of DNA extraction methods have not been comprehensively undertaken using human respiratory specimens. (2) Methods: We performed ITS analysis of respiratory (sputum) samples by assessing (a) the effect of alternate DNA extraction techniques and (b) an evaluation of four different ITS primer pairs (ITS1F and ITS2; ITS1-30F and ITS1-217R; gITS7ngs and ITS4ng; and Fseq and Rseq) on the mycobiome profiles generated for mock fungal communities and their respective clinical (airway) specimens. (3) Results: Primer pairs varied in their resulting ITS mycobiome profiles, suggesting that particular pairs may be more relevant for analysis of respiratory samples compared to others. Assessment of DNA extraction methods highlighted lower final DNA concentrations achieved by mechanical disruption compared to enzymatic lysis. However, despite lower yields, DNA liberated by mechanical lysis more readily yielded ITS bands with highest success in combination with the Fseq and Rseq primers. (4) Conclusion: Choice of extraction method, primers used, and sequencing approach are all important considerations in sequencing the mycobiome and should be tailored to sample type. A standardization of approach to mycobiome studies using respiratory specimens will permit more reliable comparisons between studies and improve our understanding of the role of fungi in the human airway.Published versio

Evaluation of droplet digital polymerase chain reaction (ddPCR) for the absolute quantification of Aspergillus species in the human airway

Background: Prior studies illustrate the presence and clinical importance of detecting Aspergillus species in the airways of patients with chronic respiratory disease. Despite this, a low fungal biomass and the presence of PCR inhibitors limits the usefulness of quantitative PCR (qPCR) for accurate absolute quantification of Aspergillus in specimens from the human airway. Droplet digital PCR (ddPCR) however, presents an alternative methodology allowing higher sensitivity and accuracy of such quantification but remains to be evaluated in head-to-head fashion using specimens from the human airway. Here, we implement a standard duplex TaqMan PCR protocol, and assess if ddPCR is superior in quantifying airway Aspergillus when compared to standard qPCR. Methods: The molecular approaches of qPCR and ddPCR were applied to DNA fungal extracts in n = 20 sputum specimens obtained from non-diseased (n = 4), chronic obstructive pulmonary disease (COPD; n = 8) and non-cystic fibrosis bronchiectasis (n = 8) patients where Aspergillus status was known. DNA was extracted and qPCR and ddPCR performed on all specimens with appropriate controls and head-to-head comparisons performed. Results: Standard qPCR and ddPCR were both able to detect, even at low abundance, Aspergillus species (Aspergillus fumigatus - A. fumigatus and Aspergillus terreus - A. terreus) from specimens known to contain the respective fungi. Importantly, however, ddPCR was superior for the detection of A. terreus particularly when present at very low abundance and demonstrates greater resistance to PCR inhibition compared to qPCR. Conclusion: ddPCR has greater sensitivity for A. terreus detection from respiratory specimens, and is more resistant to PCR inhibition, important attributes considering the importance of A. terreus species in chronic respiratory disease states such as bronchiectasis.Ministry of Health (MOH)Nanyang Technological UniversityNational Medical Research Council (NMRC)Published versionThis research is supported by the Singapore Ministry of Health’s National Medical Research Council under its Clinician-Scientist Individual Research Grant (MOH-000141, S.H.C.) and the NTU Integrated Medical, Biological and Environmental Life Sciences (NIMBELS), Nanyang Technological University, Singapore [NIM/03/2018] (S.H.C.). P.Y.T. is supported by Singapore Ministry of Health’s National Medical Research Council under its Research Training Fellowship (NMRC/Fellowship/0049/2017). L.L.Y.C. is supported by Lee Kong Chian School of Medicine Dean’s Postdoctoral Fellowship & Wong Peng Onn Postdoctoral Fellowship (2019)

Respiratory Mycoses in COPD and Bronchiectasis

Chronic obstructive pulmonary disease (COPD) and bronchiectasis represent chronic airway diseases associated with significant morbidity and mortality. Bacteria and viruses are commonly implicated in acute exacerbations; however the significance of fungi in these airways remains poorly defined. While COPD and bronchiectasis remain recognized risk factors for the occurrence of Aspergillus-associated disease including chronic and invasive aspergillosis, underlying mechanisms that lead to the progression from colonization to invasive disease remain uncertain. Nonetheless, advances in molecular technologies have improved our detection, identification and understanding of resident fungi characterizing these airways. Mycobiome sequencing has revealed the complex varied and myriad profile of airway fungi in COPD and bronchiectasis, including their association with disease presentation, progression, and mortality. In this review, we outline the emerging evidence for the clinical importance of fungi in COPD and bronchiectasis, available diagnostic modalities, mycobiome sequencing approaches and association with clinical outcomes.</p

The role of acute and chronic respiratory colonization and infections in the pathogenesis of COPD

COPD is a major global concern, increasingly so in the context of ageing populations. The role of infections in disease pathogenesis and progression is known to be important, yet the mechanisms involved remain to be fully elucidated. While COPD pathogens such as Haemophilus influenzae, Moraxella catarrhalis and Streptococcus pneumoniae are strongly associated with acute exacerbations of COPD (AECOPD), the clinical relevance of these pathogens in stable COPD patients remains unclear. Immune responses in stable and colonized COPD patients are comparable to those detected in AECOPD, supporting a role for chronic colonization in COPD pathogenesis through perpetuation of deleterious immune responses. Advances in molecular diagnostics and metagenomics now allow the assessment of microbe-COPD interactions with unprecedented personalization and precision, revealing changes in microbiota associated with the COPD disease state. As microbial changes associated with AECOPD, disease severity and therapeutic intervention become apparent, a renewed focus has been placed on the microbiology of COPD and the characterization of the lung microbiome in both its acute and chronic states. Characterization of bacterial, viral and fungal microbiota as part of the lung microbiome has the potential to reveal previously unrecognized prognostic markers of COPD that predict disease outcome or infection susceptibility. Addressing such knowledge gaps will ultimately lead to a more complete understanding of the microbe-host interplay in COPD. This will permit clearer distinctions between acute and chronic infections and more granular patient stratification that will enable better management of these features and of COPD.NMRC (Natl Medical Research Council, S’pore)MOH (Min. of Health, S’pore)Accepted versio

Understanding COPD-overlap syndromes

Introduction: Chronic obstructive pulmonary disease accounts for a large burden of lung disease. It can 'overlap' with other respiratory diseases including bronchiectasis, fibrosis and obstructive sleep apnea (OSA). While COPD alone confers morbidity and mortality, common features with contrasting clinical outcomes can occur in COPD 'overlap syndromes'. Areas covered: Given the large degree of heterogeneity in COPD, individual variation to treatment is adopted based on its observed phenotype, which in turn overlaps with features of other respiratory disease states such as asthma. This is coined asthma-COPD overlap syndrome ('ACOS'). Other examples of such overlapping clinical states include bronchiectasis-COPD ('BCOS'), fibrosis-COPD ('FCOS') and OSA-COPD ('OCOS'). The objective of this review is to highlight similarities and differences between the COPD-overlap syndromes in terms of risk factors, pathophysiology, diagnosis and potential treatment differences. Expert commentary: As a consequence of COPD overlap syndromes, a transition from the traditional 'one size fits all' treatment approach is necessary. Greater treatment stratification according to clinical phenotype using a precision medicine approach is now required. In this light, it is important to recognize and differentiate COPD overlap syndromes as distinct disease states compared to individual diseases such as asthma, COPD, fibrosis or bronchiectasis.Accepted versio