Microbial load perturbation model identifies commensal-dependent control of cough sensitivity in health and disease

The cough reflex is a fundamental airway defence mechanism regulated by interactions among epithelial, immune, and neuronal pathways. Recent evidence suggests that the low-biomass respiratory microbiome provides tonic signals essential for maintaining airway defence. Antibiotics (ATB) reduce microbial load in the airways, yet their impact on cough regulation under physiological and pathological conditions remains insufficiently understood. The aim of the present study is to investigate how ATB-induced perturbation of airway microbial load affects cough reflex sensitivity in naïve and allergen-sensitised airways, and to assess associated immune, cellular, and structural changes. Male and female Dunkin Hartley guinea pigs were studied under naïve or ovalbumin (OVA)-sensitised conditions. Animals received saline or sulfadoxine/trimethoprim pretreatment for 14 days. Cough was induced by inhalation of 0.4 M citric acid and quantified using whole-body plethysmography. Airway microbial load in bronchoalveolar lavage fluid (BALF) was measured by droplet digital PCR targeting the 16 S rRNA gene in naïve animals. Blood leukocyte counts, BALF cellularity and viability, and airway remodelling were assessed using automated cell analysis and histological staining (H&E and Sirius Red). ATB pretreatment significantly reduced airway microbial load in naïve animals, markedly suppressing cough counts and prolonging cough latency without major changes in blood cell counts or airway structure. In OVA-sensitised animals, cough latency was also significantly prolonged, whereas the number of provoked coughs showed a non-significant decrease. ATB treatment did not affect collagen deposition or peribronchiolar inflammatory infiltrates in either group. Immune responses were context-dependent: sensitised animals exhibited increased circulating neutrophils, eosinophils, and monocytes following ATB treatment, whereas naïve animals showed no systemic cellular changes. BALF cell viability decreased in naïve animals but increased in sensitised animals after ATB treatment. ATB-induced depletion of airway microbial load suppresses cough reflex sensitivity in both healthy and inflamed airways, independent of airway remodelling. These findings identify microbial-derived tonic signalling as a possible regulator of airway sensory excitability and demonstrate that immune effects of ATB depend on baseline inflammatory status. Excessive ATB use may therefore compromise airway defence in clinical practice.