Analysis of the Role of Host Genetics in Shaping Diversity of the Murine Lung Microbiota

Although lungs were long considered as sterile, the presence and importance of microbes inhabiting the lungs are now widely recognized. Using the 15th generation (“G15”) of a mouse advanced intercross line (AIL) population, we performed QTL mapping in an effort to identify host genes that influence lung microbes and may play a role in lung functioning and disease susceptibility. The lung microbiota of the G15 mouse population was examined through bacterial 16S rRNA gene amplicon sequencing, whereby Lactobacillus was identified as the most abundant genus. High-precision droplet digital PCR (ddPCR) was adapted in order to refine the dataset, remove contaminants, and quantify the absolute load of the candidate genera Lactobacillus and Pelomonas. The association between host genetic loci and bacterial traits was determined using a QTL linkage mapping approach on the relative abundances of the core microbiota as well as the absolute load data of the two candidate genera. This analysis yielded candidate host genes associated with various lung diseases. For example, two candidate genes, Il-10 and Mk2, were detected from mapping Lactobacillus load and were especially interesting as both are involved in regulating inflammatory responses. As such, these genes are known to be involved in lung diseases such as lung cancer, cystic fibrosis, and allergic airway inflammation. To test for potential interaction and feedback between these genes and Lactobacillus, their gene expression levels were measured using ddPCR, which revealed a significant negative correlation between Lactobacillus load and Mk2 expression. A follow-up study was performed with Il-10 knockout (KO) mice, but the initial results revealed no significant differences in lung microbiota composition according to Il-10 genotype. Finally, a candidate gene analysis was performed in wild type versus B4galnt2 knock out mice, as this blood group related gene was previously shown to influence the gut microbiota, and is also predicted to be expressed in the lung. This analysis also failed to detect an influence of host genotype. Through this in-depth study of the murine lung microbiota, we generated novel working hypotheses for the impact of candidate bacteria and disease genes on host inflammatory responses in the lung, which will be explored in future work.