Characterization of two novel propachlor degradation pathways in two species of soil bacteria

Abstract

Propachlor (2-chloro-N-isopropylacetanilide) is an acetamide herbicide used in preemergence. In this study, we isolated and characterized a soil bacterium, Acinetobacter strain BEM2, that was able to utilize this herbicide as the sole and limiting carbon source. Identification of the intermediates of propachlor degradation by this strain and characterization of new metabolites in the degradation of propachlor by a previously reported strain of Pseudomonas (PEM1) support two different propachlor degradation pathways. Washed-cell suspensions of strain PEM1 with propachlor accumulated N-isopropylacetanilide, acetanilide, acetamide, and catechol. Pseudomonas strain PEM1 grew on propachlor with a generation time of 3.4 h and a Ks of 0.17 +/- 0.04 mM. Acinetobacter strain BEM2 grew on propachlor with a generation time of 3.1 h and a Ks of 0.3 +/- 0.07 mM. Incubations with strain BEM2 resulted in accumulation of N-isopropylacetanilide, N-isopropylaniline, isopropylamine, and catechol. Both degradative pathways were inducible, and the principal product of the carbon atoms in the propachlor ring was carbon dioxide. These results and biodegradation experiments with the identified metabolites indicate that metabolism of propachlor by Pseudomonas sp. strain PEM1 proceeds through a different pathway from metabolism by Acinetobacter sp. strain BEM2.

FIG. 1

Kinetics of propachlor metabolism by propachlor-grown cells. (A) Acinetobacter sp. strain BEM2. (B) Pseudomonas sp. strain PEM1. Linear regression analysis of the data in a Hanes plot (insert) revealed the K_s values shown in Table 1.

FIG. 2

Utilization of propachlor by Acinetobacter sp. strain BEM2 (A) and by Pseudomonas sp. strain PEM1 (B) and appearance of the metabolic intermediates. Metabolites were measured by GC analysis as described in Materials and Methods. Propachlor (•), N-isopropylacetanilide (▩), N-isopropylaniline (), isopropylamine (), acetanilide (), and acetamide (░⃞) concentrations are shown.

FIG. 3

Schematic pathways proposed for the degradation of propachlor. Metabolites IIa and IIIa were specifically produced by Acinetobacter sp. strain BEM2, and metabolites IIb and IIIb were produced by Pseudomonas sp. strain PEM1. Metabolites I and IV were produced by both strains during propachlor degradation. Metabolite I (N-isopropylacetanilide) was identified in a previous publication (12). Chemical designations: IIa, N-isopropylaniline; IIIa, isopropylamine; IIb, acetanilide; IIIb, acetamide; IV, catechol.