Identifying aerosolized cyanobacteria in the human respiratory tract: A proposed mechanism for cyanotoxin-associated diseases

Abstract

Cyanobacteria produce harmful toxins that have been associated with several acute conditions and chronic human diseases, like gastroenteritis, non-alcoholic liver disease, and amyotrophic lateral sclerosis. Aerosol from waterbodies appears to be a likely mechanism for exposure. We conducted a study of human biospecimens focused on the cyanobacterial aerosilization process by evaluating the extent to which cyanobacteria can invade the human respiratory tract. Our study suggests that humans routinely inhale aerosolized cyanobacteria, which can be harbored in the nostrils and the lungs. Using PCR, cyanobacteria were found at high frequencies in the upper respiratory tract (92.20%) and central airway (79.31%) of our study subjects. Nasal swabs were not predictive of bronchoalveolar lavage (BAL) when detecting inhaled cyanobacteria. Interestingly, we found no evidence that time of year was a significant factor for cyanobacteria positivity (BAL cytology p = 1.0 and PCR p = 1.0); (nasal swab cytology p = 0.051 and PCR p = 0.65). Additionally, we found that proximity to a waterbody was not a significant factor for cyanobacteria positivity in BAL and nasal swabs collected during cyanobacteria bloom season [May-October] (p = 0.46 and p = 0.38). These data suggest that cyanobacteria exposure may be a prevalent and chronic phenomenon not necessarily restricted to waterbodies alone. Sources of indoor exposure warrant future investigation. Given the widespread prevalence of cyanobacterial exposure in the airway, investigation of the aerosol spread of cyanotoxins, more specifically, is warranted. Our findings are consistent with the hypothesis that aerosol is a significant route for cyanobacteria exposure, and thus a likely route of transmission for cyanotoxin-associated human diseases.

Keywords: Aerosol exposure; Cyanobacteria; Environmental toxin; Human health; Respiration.

Figure 1.. Representative sample preparation of normal human BAL cells at a concentration of ~5,500 cells/mL.

This sample was identified as PCR+ for cyanobacteria. (a) Giemsa stain (purple/ pink) was used to label nucleated human lavage cells. (b) Two cyanobacteria cells expressing phycocyanobilin pigments are seen (10x, 50um scale) using an Olympus IX73 Inverted Fluorescence Microscope. A phycocyanobilin cube with the excitation wavelength of 572nm and emission wavelength of 640nm was used. Images were enhanced to increase cell visibility using a standard procedure applied to all images.

Figure 2.. Control microbes.

Preparations (at ~5,500 cells/mL) were made for the two control microbes. (a)/ (b) Positive Control: brightfield microscopy shows a population of Microcystis sp. cells and fluorescence microscopy depicts these cells globally expressing phycocyanobilin pigments (c)/ (d) Negative Control: brightfield microscopy shows a population of Escherichia coli K-12 strain cells while fluorescence microscopy do not show any cells expressing phycocyanobilin pigments. Performed using an Olympus IX73 Inverted Fluorescence Microscope (10x, 50um scale). A phycocyanobilin cube with the excitation wavelength of 572nm and emission wavelength of 640nm was used. Images were enhanced to increase cell visibility using a standard procedure applied to all images.

Figure 3.. Representative sample preparation of a nasal swab smear.

This sample was identified as PCR+ for cyanobacteria (a) Giemsa stain (purple/ pink) was used to label nucleated cells. (b) A singular cyanobacteria cell expressing phycocyanobilin pigments is seen (10x, 50um scale) using an Olympus IX73 Inverted Fluorescence Microscope. A phycocyanobilin cube with the excitation wavelength of 572nm and emission wavelength of 640nm was used. Images were enhanced to increase cell visibility using a standard procedure applied to all images.

Figure 4.. May - Oct Samples vs Distance to Nearest Waterbody:

(a) BAL: Two-samples independent t-test was used to determine if cyanobacteria positive BAL samples collected during May-October came from subjects who lived a closer mean log distance to the nearest waterbody. (b) Nasal Swabs: Two-samples independent T-test was used to determine if cyanobacteria positive nasal swab samples collected during May-October came from subjects who lived a closer mean log distance to the nearest waterbody.