Microbes play a decisive role in respiratory health by both directly infecting the respiratory tract and exacerbating respiratory tract diseases such as chronic obstructive pulmonary disease (COPD), cystic fibrosis (CF), and asthma. While opportunistic pathogens—pathobionts—frequently colonize the upper respiratory tract asymptomatically, invasion to the lungs causes pneumonia due to the ensuing immune response. Bacterial-viral co-infections, such as S. pneumoniae co-infection with influenza A virus, can result in worsened disease due to overwhelming inflammation. In contrast, commensal bacteria from both the gut and upper respiratory tract play important roles in protecting against respiratory tract infection. This collection will highlight methods tailored to investigating host-pathogen interactions in the respiratory tract. These include methods to interrogate the immune response to respiratory tract colonization and infection by bacterial, viral and fungal pathogens. Approaches to study respiratory tract co-infections and the role of the microbiota in maintaining respiratory tract immune homeostasis are also encouraged. Small animal models are critical for investigating dynamic relationships between microbes and respiratory tract immunity. In such models, pathogen burdens quantified by growth assays or qPCR can be paired with techniques to analyze the immune response including flow cytometry, ELISA, qRT-PCR, and immunohistochemistry. Cell culture or ex vivo models, including primary immune cell cultures and opsonophagocytosis assays, permit analysis of specific immune cells and signaling pathways involved in pathogen clearance. Both whole systems and reductive approaches are important for progressing our understanding of the host-pathogen dynamic in the respiratory tract.