Methodological Impacts on Microbiome Structure and Indicator Status in the Human Lower Respiratory Tract
- 1University of British Columbia
Abstract
Rationale: Lung cancer is the leading cause of cancer-related death globally, and rising incidence among traditionally low-risk individuals intensifies the need for improved early-detection methods that the lower-airway microbiome may inform. Objectives: To evaluate how respiratory tract sampling method shapes inferred microbiome structure, and whether bronchial brushing recovers a microbial community ecologically distinct from BAL and oral rinse. Methods: A pilot cohort of 33 participants (8 lung cancer, 25 non-cancer controls) underwent oral rinse, bilateral bronchoalveolar lavage (BAL), and bronchial brushing. Microbiome structure was characterized by small subunit ribosomal rRNA gene amplicon sequence variant (ASV) profiling, with indicator-species analysis and SPIEC-EASI correlation-network mapping used to identify ASVs associated with sample type or cancer status. Measurements and Main Results: Sampling method was the dominant variation axis. BAL communities closely resembled oral rinse (64 jointly indicative ASVs; none shared with brushing), whereas bronchial brushing yielded a distinct but low-biomass signal. After excluding host-derived sequences, a single brush-specific indicator ASV affiliated with Sphingomonadaceae was identified within a 17-node co-occurrence module. Cancer-status effects were not detectable, consistent with limited statistical power. Conclusions: Sampling method is the primary determinant of inferred lower respiratory microbiome structure. Bronchial brushing recovers a distinct but low-biomass signal that is obscured when BAL is used in isolation. However, overlap between this low-biomass signal with host and contaminant sequences indicates that confidently resolving a core microbiome will require deeper sequencing and dedicated contamination controls. These methodological findings directly inform the design of future lung cancer- and disease-association studies.
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BibTeX
@misc{noonan2026methodological,
title = {{Methodological Impacts on Microbiome Structure and Indicator Status in the Human Lower Respiratory Tract}},
author = {A. J. C. Noonan and R. Myers and R. J. McLaughlin and A. Nag and S. Chen and C. Bartolomeu and S. A. Borden and S. Lam and S. Hallam},
howpublished = {bioRxiv},
year = {2026},
month = aug,
doi = {10.64898/2026.08.02.742334},
url = {https://www.biorxiv.org/content/10.64898/2026.08.02.742334v2},
note = {CC BY-NC-ND},
}
RIS
TY - UNPB
TI - Methodological Impacts on Microbiome Structure and Indicator Status in the Human Lower Respiratory Tract
AU - A. J. C. Noonan
AU - R. Myers
AU - R. J. McLaughlin
AU - A. Nag
AU - S. Chen
AU - C. Bartolomeu
AU - S. A. Borden
AU - S. Lam
AU - S. Hallam
PY - 2026
DA - 2026/08/02
PB - bioRxiv
DO - 10.64898/2026.08.02.742334
UR - https://www.biorxiv.org/content/10.64898/2026.08.02.742334v2
AB - Rationale: Lung cancer is the leading cause of cancer-related death globally, and rising incidence among traditionally low-risk individuals intensifies the need for improved early-detection methods that the lower-airway microbiome may inform. Objectives: To evaluate how respiratory tract sampling method shapes inferred microbiome structure, and whether bronchial brushing recovers a microbial community ecologically distinct from BAL and oral rinse. Methods: A pilot cohort of 33 participants (8 lung cancer, 25 non-cancer controls) underwent oral rinse, bilateral bronchoalveolar lavage (BAL), and bronchial brushing. Microbiome structure was characterized by small subunit ribosomal rRNA gene amplicon sequence variant (ASV) profiling, with indicator-species analysis and SPIEC-EASI correlation-network mapping used to identify ASVs associated with sample type or cancer status. Measurements and Main Results: Sampling method was the dominant variation axis. BAL communities closely resembled oral rinse (64 jointly indicative ASVs; none shared with brushing), whereas bronchial brushing yielded a distinct but low-biomass signal. After excluding host-derived sequences, a single brush-specific indicator ASV affiliated with Sphingomonadaceae was identified within a 17-node co-occurrence module. Cancer-status effects were not detectable, consistent with limited statistical power. Conclusions: Sampling method is the primary determinant of inferred lower respiratory microbiome structure. Bronchial brushing recovers a distinct but low-biomass signal that is obscured when BAL is used in isolation. However, overlap between this low-biomass signal with host and contaminant sequences indicates that confidently resolving a core microbiome will require deeper sequencing and dedicated contamination controls. These methodological findings directly inform the design of future lung cancer- and disease-association studies.
ER -