Daily Respiratory Research Analysis
Three impactful respiratory studies span stewardship, models, and diagnostics: long-term azithromycin in HIV-associated chronic lung disease reduced gut microbial diversity with effects persisting after cessation; a human precision-cut lung slice acid injury–repair model enables translational discovery and drug testing; and an extraction-free CRISPR workflow detects SARS-CoV-2 from dry swabs and viral transport media, offering rapid, low-cost testing.
Summary
Three impactful respiratory studies span stewardship, models, and diagnostics: long-term azithromycin in HIV-associated chronic lung disease reduced gut microbial diversity with effects persisting after cessation; a human precision-cut lung slice acid injury–repair model enables translational discovery and drug testing; and an extraction-free CRISPR workflow detects SARS-CoV-2 from dry swabs and viral transport media, offering rapid, low-cost testing.
Research Themes
- Antibiotic stewardship and microbiome effects in chronic lung disease
- Translational human ex vivo models of lung injury and repair
- Rapid, extraction-free CRISPR diagnostics for respiratory viruses
Selected Articles
1. Effect of long-term azithromycin treatment on gut microbial diversity in children and adolescents with HIV-associated chronic lung disease.
In a double-blind, placebo-controlled trial of children and adolescents with HIV-associated chronic lung disease, once-weekly azithromycin for 48 weeks reduced gut bacterial alpha-diversity and altered the abundance of 27 genera compared with placebo. Network connectivity among genera was lower with azithromycin, and Campylobacter depletion persisted 6 months post-cessation. These findings highlight durable microbiome impacts of long-term macrolide therapy.
Impact: This randomized trial quantifies sustained gut microbiome perturbations from long-term azithromycin in a high-risk respiratory population, informing antibiotic stewardship in chronic lung disease.
Clinical Implications: Clinicians should balance exacerbation reduction against potential long-lasting microbiome disruption when considering long-term azithromycin in pediatric HIV-associated chronic lung disease, monitor for consequences (e.g., dysbiosis, resistance), and prioritize stewardship and individualized duration.
Key Findings
- After 48 weeks, bacterial alpha-diversity was significantly lower in the azithromycin group versus placebo.
- Twenty-seven bacterial genera showed differential abundance between groups at 48 weeks.
- Microbial interconnectivity networks were more interconnected in placebo than in azithromycin at 48 weeks.
- Correlations between top differentially abundant genera and plasma biomarkers seen at inclusion were no longer significant at 48 weeks.
- Campylobacter depletion persisted 6 months after stopping azithromycin.
Methodological Strengths
- Double-blind, placebo-controlled randomized design with longitudinal sampling at 0, 48, and 72 weeks
- Use of 16S rRNA sequencing and multiplex plasma biomarker profiling
- Multisite recruitment (Zimbabwe and Malawi), enhancing external validity within target population
Limitations
- Microbiome assessed from rectal swabs, which may not fully represent stool microbiota
- 16S rRNA profiling limits taxonomic resolution and lacks functional/metagenomic data
- Generalizability beyond children/adolescents with HIV-associated chronic lung disease is uncertain
- Antimicrobial resistance outcomes were not directly measured
Future Directions: Integrate metagenomics and resistome profiling, assess clinical consequences of dysbiosis, and test stewardship-informed dosing/duration strategies to mitigate microbiome disruption while preserving respiratory benefits.
BACKGROUND: HIV-associated chronic lung disease (HCLD) is common in children and adolescents growing up with HIV. The use of azithromycin (AZM) reduces the rate of acute respiratory exacerbations in this population, however, impact of this treatment on the gut microbiota and associations with blood-derived inflammatory markers have not been studied. METHODS: Children and adolescents with HCLD in Harare, Zimbabwe and Blantyre, Malawi were recruited in a double-blind, placebo-controlled trial of once-weekly AZM or placebo for 48 weeks (BREATHE trial, NCT02426112). Rectal swabs were collected at inclusion (N = 346), 48 weeks (treatment end, N = 313), and 72 weeks (six months after treatment cessation, N = 244). The bacterial composition of fecal swabs was determined using 16S rRNA gene sequencing. Plasma biomarkers at inclusion and 48 weeks were measured using Luminex multiplex bead assay. FINDINGS: At 48 weeks, bacterial α-diversity was significantly lower in the AZM group, with 27 bacterial genera showing differential abundance between the study groups. The placebo group exhibited higher interconnectivity between bacterial genera at 48 weeks compared to the AZM group. Correlations between the top seven differentially abundant genera and biomarkers observed at inclusion were no longer significant at 48 weeks in both groups. Depletion of Campylobacter persisted for six months after cessation of AZM treatment. INTERPRETATION: Long-term AZM treatment in HCLD patients affects their gut bacterial composition at least 6 months after its cessation. The consequences of reduced bacterial diversity, such as altered interaction with the immune system and risk of resistance, need further investigation to understand how to optimise gut health during long-term antibiotic treatments. FUNDING: The study was funded by the Norwegian Research Council and Helse Nord (HNF 1387-17).
2. A human PCLS model of lung injury and repair for discovery and pharmaceutical research.
Investigators adapted the acid injury–repair model to human precision-cut lung slices, creating a translational ex vivo platform. Acid injury did not alter overall proliferation but significantly increased alveolar type II/progenitor markers (proSP-C, HTII) in injured regions, and multiple repair-relevant cell types (lipofibroblasts, endothelial cells) were identifiable. The model mimics heterogeneous injury and supports mechanistic and drug screening studies.
Impact: Provides a human tissue-based injury–repair system bridging in vitro and in vivo models, enabling mechanistic studies and preclinical screening in authentic 3D lung microenvironments.
Clinical Implications: While not immediately practice-changing, this human ex vivo platform can accelerate development and validation of therapeutics aimed at enhancing lung repair and may improve translational fidelity of preclinical findings.
Key Findings
- Human precision-cut lung slices were successfully adapted to the acid injury–repair (AIR) model (hAIR).
- Acid injury did not change overall proliferation (Ki67) but increased the proportion of proSP-C and HTII-positive cells in injured regions.
- Repair-relevant non-epithelial populations (lipofibroblasts and endothelial cells) were identified and tracked within hAIR.
- The model establishes adjacent injured and uninjured regions, mimicking heterogeneous lung injury patterns.
Methodological Strengths
- Use of human resected lung tissue and 3D precision-cut lung slices with localized injury
- Multiplex cell marker immunostaining (Ki67, proSP-C, HTII, ADRP, ERG) enabling cell-type–specific analyses
- Parallel viability assessment (MTT, Live/Dead) to support tissue integrity over culture
Limitations
- Short culture timeframe (48 h) captures early repair only; longer-term dynamics not assessed
- Acid injury may not represent all clinical injury etiologies (e.g., infection, mechanical, toxin)
- Donor number and variability were not specified, limiting generalizability and statistical inference
Future Directions: Extend culture duration and functional readouts, incorporate diverse injury stimuli, quantify donor variability, and apply the platform for preclinical testing of pro-repair therapeutics.
BACKGROUND: The Acid Injury and Repair (AIR) model is an ex-vivo model of lung injury and repair, that was previously established using mouse precision-cut lung slices (PCLS). The AIR model provides a bridge between the current in -vitro and in-vivo models to study the effects of lung injury in 3D lung tissue slices. Here, we show that the AIR model can be adapted for use in human tissue as a translational model for discovery research and drug screening. METHODS: To generate PCLS, resected human lung tissue was coated with alginate hydrogel to form an artificial pleura. Lung tissue was inflated by point injecting 3% agarose, followed by generation of 450-500 µM thick slices of tissue. An isolated area of each slice was injured by brief application of 0.1 M hydrochloric acid. AIR-PCLS were then washed and cultured for 48 h before immunostaining to assess proliferating cells (Ki67) alveolar type II/progenitor cell markers (HTII, proSP-C), lipofibroblasts (ADRP) and endothelial cells (ERG). Viability of PCLS was assessed by both MTT assay and Live/Dead staining. RESULTS: We show that levels of proliferation do not change in response to acid injury. However, there is a significant increase in the percentage of proSP-C and HTII positive cells in the injured regions of AIR-PCLS. We also identify non-epithelial cell populations; lipofibroblasts and endothelial cells in human AIR-PCLS, to demonstrate that other repair relevant cell types can be identified and tracked in the human AIR (hAIR model). CONCLUSIONS: The hAIR model is an effective ex-vivo tool to study early mechanisms of lung repair following injury. By establishing an area of injured tissue adjacent to uninjured tissue, this model mimics the heterogenous pattern of lung injury frequently present in lung diseases. The hAIR model will facilitate mechanistic studies of human lung repair and provides a valuable pre-clinical model for drug testing.
3. RNA extraction-free CRISPR-based SARS-CoV-2 detection in viral transport medium and dry swab-a comparative analysis: Short title: Extraction-free CRISPR-based Covid detection.
An extraction-free workflow combining proteinase K/heat treatment, RT-LAMP, and CRISPR detected SARS-CoV-2 directly from both viral transport medium and dry swabs. Detection was more efficient for dry swabs than for VTM. This approach can reduce cost and turnaround time and may be adaptable to other respiratory pathogens.
Impact: Demonstrates a practical, extraction-free CRISPR diagnostic workflow across common sample types, addressing throughput and accessibility constraints during respiratory outbreaks.
Clinical Implications: Laboratories could implement extraction-free CRISPR workflows, especially for dry swabs, to expand rapid testing capacity in low-resource settings; however, formal validation of diagnostic accuracy versus RT-PCR is needed prior to adoption.
Key Findings
- SARS-CoV-2 was detectable without RNA extraction from both dry swabs and two commercial VTM preparations using proteinase K/heat, RT-LAMP, and CRISPR.
- Detection efficiency was higher for dry swab samples than for VTM samples.
- The workflow is presented as adaptable to other respiratory diseases.
Methodological Strengths
- Direct comparison of two common specimen types (dry swab vs VTM) within an extraction-free workflow
- Use of orthogonal amplification (RT-LAMP) and CRISPR detection to enhance sensitivity
- Simple sample pre-processing (proteinase K and heat) enabling low-cost implementation
Limitations
- Diagnostic performance metrics (sensitivity/specificity) relative to RT-PCR were not fully quantified
- Sample size and patient characteristics were not specified, limiting inference
- Only two VTM brands were tested, so generalizability to other media is unknown
Future Directions: Undertake large-scale clinical validation across diverse VTMs and respiratory pathogens, define analytical limits of detection, and integrate into point-of-care platforms.
CRISPR-Cas-based methods have shown high efficacy in detecting SARS-CoV-2. Amidst the Covid-19 pandemic, numerous studies have explored SARS-CoV-2 detection methods without the need for RNA extraction, aiming to reduce cost and processing time. Here, we assessed a CRISPR-based SARS-CoV-2 detection method's ability to detect the virus in viral transport medium (VTM). Swabs obtained from Covid-19 positive patients and stored in two Indian brands of VTM were examined alongside dry swab samples. The samples underwent proteinase -K treatment followed by heat incubation. The released nucleic acids were tested by RT-LAMP and CRISPR-based detection. We conclude that SARS-CoV-2 can be detected in VTM of two commercial preparations as well as dry swab samples without RNA extraction and purification. COVID detection was found to be more efficient for dry swab samples compared to VTM samples. The work flow described in this paper can be extended to other respiratory diseases.