16S rRNA sequencing of bronchoalveolar lavage fluid from high-weight broilers after nebulization of Ligilactobacillus salivarius

This study was designed to evaluate the effects of continuous nebulization of two Ligilactobacillus salivarius on the pulmonary microbial diversity of broilers. The two strains were isolated from high-yielding performance broilers, Ligilactobacillus salivarius GHL01 (designated BALF-A), derived from bronchoalveolar lavage fluid (BALF), and Ligilactobacillus salivarius GHL02 (designated Lung-A), isolated from lung tissue. A total of 1,000 one-day-old Arbor Acres (AA) broilers were randomly assigned to three experimental groups, a control group (a total of 200 pieces; 25 replicates, 8 birds per replicate) and two treatment groups receiving aerosolized Lung-A (a total of 320 pieces; 40 replicates, 8 birds per replicate) or BALF-A (a total of 320 pieces; 40 replicates, 8 birds per replicate). A bacterial suspension (the number of bacteria per milliliter is 10^9) was administered via aerosol using a medical nebulizer every three days, with each bird exposed for 20 minutes per session. At 42 days of age, 20 birds per group were randomly selected and humanely euthanized by cervical dislocation. Bronchoalveolar lavage fluid (BALF) was collected by instilling sterile, pre-chilled phosphate-buffered saline (PBS) into the trachea and retrieving the effluent. Approximately 2 mL of BALF per bird was obtained and stored for subsequent 16S rRNA amplicon sequencing to assess microbial community composition.