Biodegradation and metabolic pathway of fenvalerate by Citrobacter freundii CD-9

Abstract

Fenvalerate is a pyrethroid insecticide with rapid action, strong targeting, broad spectrum, and high efficiency. However, continued use of fenvalerate has resulted in its widespread presence as a pollutant in surface streams and soils, causing serious environmental pollution. Pesticide residues in the soil are closely related to food safety, yet little is known regarding the kinetics and metabolic behaviors of fenvalerate. In this study, a fenvalerate-degrading microbial strain, CD-9, isolated from factory sludge, was identified as Citrobacter freundii based on morphological, physio-biochemical, and 16S rRNA sequence analysis. Response surface methodology analysis showed that the optimum conditions for fenvalerate degradation by CD-9 were pH 6.3, substrate concentration 77 mg/L, and inoculum amount 6% (v/v). Under these conditions, approximately 88% of fenvalerate present was degraded within 72 h of culture. Based on high-performance liquid chromatography and gas chromatography-mass spectrometry analysis, ten metabolites were confirmed after the degradation of fenvalerate by strain CD-9. Among them, o-phthalaldehyde is a new metabolite for fenvalerate degradation. Based on the identified metabolites, a possible degradation pathway of fenvalerate by C. freundii CD-9 was proposed. Furthermore, the enzyme localization method was used to study CD-9 bacteria and determine that its degrading enzyme is an intracellular enzyme. The degradation rate of fenvalerate by a crude enzyme solution for over 30 min was 73.87%. These results showed that strain CD-9 may be a suitable organism to eliminate environmental pollution by pyrethroid insecticides and provide a future reference for the preparation of microbial degradation agents and environmental remediation.

Keywords: Biodegradation metabolites; Citrobacter freundii CD-9; Fenvalerate; Metabolic pathway; Response surface methodology.

Fig. 1

a Morphology of Citrobacter freundii cells by scanning electron microscopy. b Phylogenetic analysis of C. freundii CD-9 based on the 16S rRNA sequence. The numbers shown in parentheses are GenBank sequence accession numbers. The numbers at the nodes indicate bootstrap values from neighbor-joining analysis of 1000 resampled data sets. The bar represents sequence divergence

Fig. 2

Growth and fenvalerate degradation of C. freundii CD-9

Fig. 3

Three-dimensional plots of a quadratic model for fenvalerate degradation by strain CD-9

Fig. 4

Comparison chart of before and after fenvalerate degradation (high-performance liquid chromatography [HPLC])

Fig. 5

Gas chromatography-mass spectrometry (GC-MS) analysis of metabolites produced from fenvalerate degradation by strain CD-9. (C, F, H, I) Characteristic ions of compounds C, F, H, and I in GC-MS. Retention times of each compound were 14.732, 23.152, 25.408, and 28.825 min, respectively, and the compounds were identified as o-phthalaldehyde, 3-phenoxybenzaldehyde, dibutyl phthalate, and 3-phenoxybenzyl alcohol, respectively

Fig. 6

Proposed pathway for degradation of fenvalerate by strain CD-9