The Relationship Between Bacterial Denitrification by SIR Inhibition Method and Denitrifying Bacteria Populations by Real-Time PCR in Louisiana Coastal Swamp Forest Sediment Under Various Redox Conditions.

The Relationship between Bacterial Denitrification by SIR Inhibition Method and Denitrifying Bacteria Populations by Real-time PCR in Louisiana Coastal Swamp Forest Sediment under Various Redox Conditions

 

Dong Cheol Seo¹, Hee Sung Bae², Ronald D. Delaune^1,*

 

¹Department of Oceanography and Coastal Sciences, School of the Coast and Environment, Louisiana State University, Baton Rouge, LA 70803, USA,

²Soil and Water Science Department, Institute of Food and Agricultural Sciences, University of Florida, 2169 McCarty Hall, Gainesville, FL 32611, USA.

Abstract

Bacterial denitrification rates and denitrifying bacteria populations were determined under a range of redox conditions in sediment from a Louisiana Mississippi River deltaic plain swamp forest used for wastewater treatment. Sediment was incubated in microcosms at 6 Eh levels (–200, –100, 0, +100, +250 and +400 mV) ranging from strongly reducing to moderately oxidizing conditions. Denitrification was determined using the substrate-induced respiration (SIR) inhibition and acetylene inhibition methods. The densities of denitrification genes coding for nitrite reductase (nirK and nirS) in sediment were quantified by real-time PCR. At Eh values of +250 mV and +400 mV, denitrification by bacteria was 1.46–1.59% of total denitrification, and the number of gene copies per gram of DNA in sediment were 1.67 x 10⁶ ~ 4.53 x 10⁶ for nirS, and 1.22 x 10⁸ ~ 2.74 x 10⁸ for nirK, respectively. At Eh –200 mV, denitrification rate of bacteria was 64.9% of total denitrification, and the number of gene copies per gram of DNA in sediment were 7.12 x 10⁵ for nirS, and 4.95 x 10⁷ for nirK, respectively. At Eh values between –100 to +100 mV, denitrification by bacteria was 53.0–51.1% of total denitrification, and the number of gene copies per gram of DNA in sediment were 1.31 x 10⁵~ 1.43 x 10⁶ for nirS, and 1.19 x 10⁷ ~ 1.37 x 10⁸ for nirK, respectively. Therefore, bacteria were responsible for most of the denitrification under strongly reducing conditions.

*Corresponding Author. Ronald D. DeLaune, Department of Oceanography and Coastal Sciences, School of the Coast and Environment, Louisiana State University, Baton Rouge, LA 70803, USA. Tel.: +1 225 578 6421; Fax: +1 225 578 6423; E-mail: rdelaun@lsu.edu.

Keywords: denitrifying bacteria, real-time PCR, nirK, nirS, sediment redox potential (Eh), swamp forest.