Bacteria Assessment of Soil Samples in Houston Watersheilds; Impact of Heavy Metal, and Stress Responses in an Eukaryotic Co-Culture System.

Houston has a complex watershed in which bayous intersect one another making the city prone to flooding, as evidenced by the 2017 Hurricane Harvey flood. We sought to evaluate bacterial population dynamics in Houston watershed soils pre- and post-Hurricane Harvey; additionally, we evaluated population dynamics in neighboring, downstream bayous ~ 1 year later in the summer and winter of 2018. This study quantified bacterial loads for pre-Hurricane Harvey (June 2017) and post-hurricane Harvey (November 2017) soil samples, as well as competitive samples from one year later [summer (June 2018) and winter (November) 2018]. Unexpectedly, bayous closer to Houston’s densely populated urban core, including Buffalo, Halls, Mustang, and Horsepen Bayous, had significantly higher enteric bacterial loads during the winter than the summer. Perhaps this was due to water flow rate changes or proximity to wastewater treatment plants. Following bacterial load determination, isolated colonies were identified using biochemical tests and DNA sequencing of the 16S ribosomal DNA region. Additionally, we employed next generation metagenomic sequencing of 16S rDNA, capturing both culturable and unculturable organisms. The phyla Proteobacteria, Actinobacteria, Bacteriodetes and Firmicutes were found to be dominant in our metagenomic analysis and are human gut bacteria. Some opportunistic bacterial Proteobacteria pathogens identified in our metabolomic analysis were Serratia marcenscens, Pseudomonas mendocina, Pseudomonas fulva, and Pseudomonas putida. To investigate the effects of heavy metal exposures, an environmentally isolated Serratia marsescens and its reference strain were exposed to Pb, Zn and manganese and subsequent oxidative stress responses and biofilm production were measured. Additionally, heavy metal exposures were characterized in gut and lung cell co-culture models using CCD-841, HT29 and BEAS-2B cell lines. Additionally, MTT assays were performed to determine cell cytotoxicity in bacterial cultures containing 10, 50, and 100 μg/ml of Zn and Pb and 100, 500 and 1000 μg/ml of Mn. Interestingly, the environmentally isolated S. marsescens produced increased biofilm and was more resistant to oxidative stress in the presence of Zn and Pb than its reference strain. Perhaps this was due to environmental adaptations of the environmental isolate. To our knowledge, this is the first study that compares Houston-area bacterial populations before and after a major flooding event. Taken together, Hurricane Harvey likely contributed to a redistribution of enteric bacteria, as there was a significant increase in the enteric population of Buffalo and Halls Bayous following the Hurricane Harvey flooding event. Similarly, our 2018 winter data set followed the same trend, as significant increases were seen in the enteric populations of Horsepen, Mustang, and Cypress Creek watershed soils. Further, the environmentally isolated S. marcescens was better able to withstand environmental stressors than the reference strain and the lung cell line was more susceptible to lead treatment by a decrease in viability of about 2.0-fold when compared with the gut cell lines. Also, the cell co-culture infection with the environmental isolate in the presence of various metal toxicants respond differently to its environmental reference strain and a significant decrease (p\u3c 0.05) in bacteria proliferation at various time points for the cell lines were observed