25Bacterial strain Pseudomonas sp. strain ADP is capable of metabolizing atrazine, a synthetic 26 herbicide, and uses atrazine as a sole nitrogen source for growth. The microbe completely 27 mineralizes the substrate in a catabolic pathway comprised of six enzymatic steps. All enzymes, 28 AtzA-AtzF, encoded by corresponding genes, AtzA-AtzF, are located on a self-transmissible 29 plasmid, pADP-1. (Souza, M. L., Wackett, L.P., and Sadowsky, M.J Appl. and Environ. 30 Microbiol. 64 (6): [2323][2324][2325][2326] 1998) RT-qPCR was used to differentiate gene expression in 31 atrazine-degrading genes in Pseudomonas sp. strain ADP cells grown as suspended cells and as 32 biofilms. Relative gene expression was also evaluated for biofilms grown at 25 o C, 30 o C, and 33 37 o C. Complementary atrazine kinetic data was collected using GC-MS for both modes of 34 growth and temperature variance. No significant difference in expression was observed for all 35 atrazine-degrading genes in biofilm-mediated cells relative to planktonic cells, suggesting neither 36 decreased or increased catabolic activity at the mRNA level. In contrasting experiments 37 concerning biofilm growth, expression was downregulated at 37 o C for genes AtzA, AtzB, and 38 AtzC and upregulated for genes AtzD, AtzE, AtzF, signifying Pseudomonas sp. strain ADP 39 biofilms catabolic activity may change in response to substantial temperature changes. Gradual 40 decreases in atrazine concentration were apparent in cells grown in shake flasks, while biofilm-41 mediated cells showed transient increases and decreases in reactor effluent. The complex 42 extracellular matrix components, quorum sensing, and genetic transfer may account for 43 accumulation and rapid degradation of atrazine. The data collected suggest biofilm-mediated 44 bioremediation may give insight into catabolic activity and atrazine degradation potential. 45 46 3 Importance 47 48Atrazine is the second most applied herbicide in the United States. It is applied to crops 49 including sorghum, corn, and sugarcane to prevent the growth of broad-leaved weeds. Once 50 used, it can permeate the soil and contaminate proximal groundwater sources, which provide 51 drinking water for over 90-million people. The Environmental Protection Agency sets the 52 maximum contaminant level at 3 parts per billion for atrazine in drinking water, however this is 53 frequently exceeded in rural regions which presents a public safety concern. Atrazine is an 54 endocrine disruptor compound and a suspected teratogen in humans and freshwater species, 55 respectively. This research is significant in evaluating the use an atrazine-degrading strain,
56Pseudomonas sp. strain ADP, grown in a biofilm mode of growth to increase the degradation 57 potential compared to suspended cells. Our results concerning expression and kinetics will aid 58 the development of biofilm reactors for ex situ bioremediation and understanding environmental 59 biofilms. 60 61 Introduction 62 63 Atrazine is a ubiquitous herbicide used to contro...