Acute responses of bio-denitrification to short-term clopyralid exposure: Kinetic analysis and biological mechanisms

Abstract

The mass production and extensive application of clopyralid (CLP) have attracted much attention regarding its environmental effect in recent decades. However, studies on the acute responses and the underlying inhibition mechanisms of denitrifiers to CLP exposure are scarce. Herein, the crucial factors in denitrification were quantified through kinetics, metabolic activity analysis, and structural equation modeling (SEM). The acute response mechanisms of denitrifiers under short-term CLP exposure were further proposed according to the quantitative correlation of denitrification performance with C and N metabolism, ETS activity, and oxidative stress. The modified non-competitive inhibition model fitted well to describe the inhibitory characteristics of CLP on the nitrate reduction (R2 = 0.99), which was significantly inhibited with increasing CLP exposure concentrations. Short-term exposure to CLP could inhibit C and N metabolism and electron transport by inducing oxidative stress during denitrification. Quantitative PCR (qPCR) analysis demonstrated that CLP restrained the abundance of narG and nirS, suggesting that down-regulation of denitrification enzyme expression was highly related to the reduction in nitrogen conversion. Moreover, SEM indicated that ETS activity and NAR were the major factors in predicting NO3− reduction rate (DNR) and were mainly influenced by NADH and reactive oxygen species (ROS). Consistently, NADH and NIR were closely correlated with total nitrite accumulation (NIA). Short-term CLP exposure could affect denitrification performance by directly affecting microbial physicochemical properties or indirectly interfering with the abundance of denitrifying functional genes and metabolic activity through oxidative stress. This study provided new insights into acute responses of denitrifiers to short-term CLP exposure.