Tobacco smoke mediated induction of sinonasal microbial biofilms

Abstract

Cigarette smokers and those exposed to second hand smoke are more susceptible to life threatening infection than non-smokers. While much is known about the devastating effect tobacco exposure has on the human body, less is known about the effect of tobacco smoke on the commensal and commonly found pathogenic bacteria of the human respiratory tract, or human respiratory tract microbiome. Chronic rhinosinusitis (CRS) is a common medical complaint, affecting 16% of the US population with an estimated aggregated cost of $6 billion annually. Epidemiologic studies demonstrate a correlation between tobacco smoke exposure and rhinosinusitis. Although a common cause of CRS has not been defined, bacterial presence within the nasal and paranasal sinuses is assumed to be contributory. Here we demonstrate that repetitive tobacco smoke exposure induces biofilm formation in a diverse set of bacteria isolated from the sinonasal cavities of patients with CRS. Additionally, bacteria isolated from patients with tobacco smoke exposure demonstrate robust in vitro biofilm formation when challenged with tobacco smoke compared to those isolated from smoke naïve patients. Lastly, bacteria from smoke exposed patients can revert to a non-biofilm phenotype when grown in the absence of tobacco smoke. These observations support the hypothesis that tobacco exposure induces sinonasal biofilm formation, thereby contributing to the conversion of a transient and medically treatable infection to a persistent and therapeutically recalcitrant condition.

Figure 1. Biofilm formation in bacteria obtained from endoscopically guided sinonasal cultures, following sham or tobacco smoke exposure.

Samples from patients evaluated in the outpatient clinic or in the operating room, who were found to have sinonasal mucopurulence were cultured. Samples were grown overnight and subjected to evaluation for biofilm forming capacity. A three hour tobacco smoke exposure (resultant from five cigarettes). Each isolate was performed in octuplet. A paired Student t-test was applied to compare smoke and sham exposed (* = p<0.01).

Figure 2. Tobacco biofilm index.

Data from figure 1A was normalized by creating a ratio of smoke to sham exposed biofilm formation. Value of <1 demonstrates biofilm inhibition while value >1 reflects biofilm induction.

Figure 3. Biofilm formation after repetitive smoke exposure.

Bacteria isolated from smoke naïve patients were subjected to daily sham or smoke exposure (5 cigarettes/3 h) and then subjected to the biofilm detection assay. Data is represented as the ratio of smoke exposure to sham exposure. *, #, and $ indicate significant differences (p<0.05) between day 1 and 2, day 1 and 3, and day 1 and 4, respectively. Indicate the number of replicates.

Figure 4. Smoke induced biofilm formation is reversible.

Bacteria isolated from smokers were grown for 4 days in the absence of tobacco smoke, before a single sham or smoke exposure (5 cigarettes/3 h) and subsequent biofilm detection assay. Indicate the number of replicates, and repeat the significance test performed, and the p values.