Biology and Control of Human Pathogens on Fresh Produce
Location: Produce Safety and Microbiology Research
Title: Bacterial communities in urban aerosols collected with wetted-wall cyclonic samplers and seasonal fluctuations of live and culturable airborne bacteria
Submitted to: Journal of Environmental Monitoring
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: November 21, 2011
Publication Date: February 12, 2012
Citation: Ravva, S.V., Hernlem, B.J., Sarreal, C.Z., Mandrell, R.E. 2012. Bacterial communities in urban aerosols collected with wetted-wall cyclonic samplers and seasonal fluctuations of live and culturable airborne bacteria. Journal of Environmental Monitoring. 14(2):473-481. doi: 10.1039/clem10573d.
Interpretive Summary: Major outbreaks associated with produce indicate that pre-harvest contamination has occurred in the field, so it is critical to identify sources of pathogens in the environment and interventions for minimizing them. Animal manure is a potential pathogen reservoir, and transport of these pathogens through aerosols to crops grown in close proximity to dairy operations to croplands in California cannot be ignored. Since contamination of ‘ready to eat’ produce cannot be washed off, on-site prevention of contamination is vital. Any such attempts require an understanding of the biological and environmental factors that regulate the proliferation of pathogens during their transport from animal reservoirs to produce grown in proximity to livestock operations. We have worked on developing improved detection methods for pathogens, and on understanding how pathogens survive and re-grow in dairy environments. Our long-term goals are to develop on-site prevention and control of pathogens at the source prior to contaminating produce we consume.
This study evaluated a flow cytometry linked culture method to monitor and characterize live bacteria and pathogens transported through atmosphere. Live cells from airborne particles were collected in liquid instead of the harsher impaction on filters known to decrease viability. The detection of culturable pathogens from bacterial communities in airborne particles indicates a potential for applying this technique for monitoring the environmental transport of pathogens. This method complements the currently used non-culture based molecular methods to characterize the community diversity in the atmosphere.
The airborne transmission of bacterial pathogens from point sources (e.g. ranches, waste treatment facilities) to areas of food production (farms) has been suspected. However, there have been few studies monitoring the incidence, transport and viability of bacteria in aerosols. We monitored the numbers of culturable bacteria in urban aerosols on 21 separate days during a 9 month period using high volume cyclonic samplers at an elevation of 6 m above the ground level. Culturable bacteria in aerosols fluctuated from 3 CFU to 6 million CFU/L of air per hour and correlated significantly with changes in seasonal temperatures, but not with humidity or wind speed. Concentrations of viable bacteria determined by fluorescence staining and flow cytometry correlated significantly with culturable bacteria. Members of the phylum Proteobacteria constituted 98% of the bacterial community, which was characterized using 16S rRNA gene sequencing using DNA from aerosols. Aquabacterium sp., previously characterized from aquatic environments, represented 63% of all clones and the second most common were Burkholderia sp; these are ubiquitous in nature and some species are known to be pathogenic. Whole genome amplification prior to sequencing resulted in a substantial decrease in species diversity as compared to characterizing culturable bacteria sorted by flow cytometry based on scatter signals. Although 27 isolated colonies were characterized, we were able to culture 38% of bacteria characterized by sequencing of DNA clone libraries. The whole genome amplification method amplified DNA preferentially from Phyllobacterium myrsinacearum, a minor member of the bacterial communities, whereas Variovorax paradoxus dominated the cultured organisms.