Microorganism-Driven 2,4-D Biodegradation: Current Status and Emerging Opportunities

Abstract

The herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) has been widely used around the world in both agricultural and non-agricultural fields due to its high activity. However, the heavy use of 2,4-D has resulted in serious environmental contamination, posing a significant risk to non-target organisms, including human beings. This has raised substantial concerns regarding its impact. In addition to agricultural use, accidental spills of 2,4-D can pose serious threats to human health and the ecosystem, emphasizing the importance of prompt pollution remediation. A variety of technologies have been developed to remove 2,4-D residues from the environment, such as incineration, adsorption, ozonation, photodegradation, the photo-Fenton process, and microbial degradation. Compared with traditional physical and chemical remediation methods, microorganisms are the most effective way to remediate 2,4-D pollution because of their rich species, wide distribution, and diverse metabolic pathways. Numerous studies demonstrate that the degradation of 2,4-D in the environment is primarily driven by enzymatic processes carried out by soil microorganisms. To date, a number of bacterial and fungal strains associated with 2,4-D biodegradation have been isolated, such as Sphingomonas, Pseudomonas, Cupriavidus, Achromobacter, Ochrobactrum, Mortierella, and Umbelopsis. Moreover, several key enzymes and genes responsible for 2,4-D biodegradation are also being identified. However, further in-depth research based on multi-omics is needed to elaborate their role in the evolution of novel catabolic pathways and the microbial degradation of 2,4-D. Here, this review provides a comprehensive analysis of recent progress on elucidating the degradation mechanisms of the herbicide 2,4-D, including the microbial strains responsible for its degradation, the enzymes participating in its degradation, and the associated genetic components. Furthermore, it explores the complex biochemical pathways and molecular mechanisms involved in the biodegradation of 2,4-D. In addition, molecular docking techniques are employed to identify crucial amino acids within an alpha-ketoglutarate-dependent 2,4-D dioxygenase that interacts with 2,4-D, thereby offering valuable insights that can inform the development of effective strategies for the biological remediation of this herbicide.

Keywords: 2,4-D; biodegradation; degradation pathways; herbicide; metabolites; molecular mechanisms.

Figure 1

The chemical structures of 2,4-D and its derivants. 2,4-D: 2,4-dichlorophenoxyacetic acid; 2,4-D butyl ester: butyl 2,4-dichlorophenoxyacetate; 2,4-D isobutyl ester: isobutyl 2,4-dichlorophenoxyacetate; MCPA: 2-methyl-4-chlorophenoxyacetic acid. They all belong to the phenoxyalkanoic acid herbicide group.

Figure 2

The proposed mechanism and pattern of action of the auxin herbicide 2,4-D at concentrations exceeding the optimal levels within weeds [21,22,23]. Upon 2,4-D’s activation of the ethylene synthesis process via the 1-aminocyclopropane-1-carboxylic acid (ACC, a precursor to ethylene biosynthesis) pathway, cyanide builds up as a side product. Under such circumstances, elevated levels of ethylene can impede plant growth, and cyanide can be detrimental to the plants’ health. Within the bud tissue, the gene for 9-cis-epoxycarotenoid dioxygenase (NCED), a key enzyme that mediates the regulation of abscisic acid (ABA) biosynthesis in plants, is overexpressed, triggering the generation of ABA. This hormone triggers the closure of the stomata, thereby limiting the plant’s transpiration process. Concurrently, there is an excessive production of reactive oxygen species (ROS). Furthermore, ABA, together with ethylene, promotes leaf senescence, impairs chloroplast function, and undermines the structural integrity of the vascular system. As a consequence, this may result in tissue dehydration and decomposition, culminating in the demise of the plant.

Figure 3

The occurrence of 2,4-D in the environment and its fate. A very small fraction of 2,4-D reaches the target weeds, while the remainder is released into the soil and water environments. As it flows with runoff and groundwater, it poses a threat to non-target flora and fauna, and more seriously, it create notable hazards for human health. Decomposition and metabolism by soil microorganisms is the primary method for the transformation of residual 2,4-D in the natural environment.

Figure 4

The 2,4-D degradation pathways proposed in Cupriavidus necator JMP134 [88,113,129].

Figure 5

The 2,4-D degradation pathways proposed in Azotobacter chroococcum [131,132,133].

Figure 6

The 2,4-D degradation pathways proposed in Aspergillus niger [120,121,122].

Figure 7

A neighbor-joining phylogenetic tree constructed based on the amino acid sequences of alpha-ketoglutarate-dependent 2,4-D dioxygenases from Bradyrhizobium sp. strain HW13 and other microbial strains (available from the NCBI nonredundant protein sequence database and the UniProtKB/Swiss-Prot database). The phylogenetic neighbor-joining tree was constructed using MEGA (version 11) software with 1000 bootstraps. Consensus sequences of every protein were aligned with ClustalW and trimmed to the same sizes. The number below the branch represents the branch length, which is calculated from the p-distance to show the evolutionary distance between different samples. The bar represents a 0.10 amino acid difference per site. The code preceding each species name is the NCBI accession number for the 2,4-D dioxygenase.

Figure 8

Prediction of the protein structure and a molecular docking model of an alpha-ketoglutarate-dependent 2,4-D dioxygenase (GenBank Accession Number 499491759) from Burkholderiaceae bacteria. (a) Cartoon structure of the 2,4-D dioxygenase; (b) Ramachandran plot of the 2,4-D dioxygenase; (c) Surface structure of the 2,4-D dioxygenase; (d) Molecular docking result. Arg 266 and Arg 267 as key amino acids in the 2,4-D dioxygenase.