Elsevier

Chemosphere

Volume 237, December 2019, 124452
Chemosphere

Three chlorotoluene-degrading bacterial strains: Differences in biodegradation potential and cell surface properties

https://doi.org/10.1016/j.chemosphere.2019.124452Get rights and content

Highlights

  • Newly-isolated bacterial strains are efficient chlorotoluenes-degraders

  • Changes in the cell surface properties indicated the strains adaptation mechanisms

  • The isolated strains differed significantly in the cellular fatty acids profile

  • The studied strains have a great potential to be used in the chlorotoluenes removal

Abstract

Pollution of the environment with chlorinated aromatic compounds is a problem of increasing importance, which has stimulated the search for efficient methods for the remediation of contaminated soil and water. Additionally, for better understanding of the significance of bioavailability to biodegradation, investigation of the cell surface properties is necessary. Hence, this study concerns the properties and possible application, in chlorotoluene removal, of three newly isolated environmental bacterial strains from the genera Pseudomonas, Raoultella and Rahnella. The results show the differences in the biochemical profiles of the isolated strains, their cellular fatty acid composition and their hemolytic properties. However, all three strains exhibit high biodegradation potential, degrading not less than 60% of each monochlorotoluene isomer in 21-day experiments. What is more, observations of changes in the cell surface properties indicate the possible adaptation mechanisms of the strains that enable efficient biodegradation of hydrophobic pollutants such as monochlorotoluenes.

Introduction

Among all living organisms, it is the microorganisms, especially bacteria, that demonstrate the greatest ability to adapt to different conditions in various ecosystems. Specific bacterial strains inhabit extreme ecological niches, from hot springs to the water of lakes under glaciers (Chen et al., 2016; Poli et al., 2017). The adaptation of microorganisms also concerns the ability to use various organic compounds as carbon and energy sources. Because of their key role in the self-cleaning of the natural environment from toxic contaminants, the strains capable of biodegrading persistent organic pollutants attract a great deal of interest (Murínová and Dercová, 2014).

One of the most important groups of organic aromatic compounds of industrial significance are the chloroaromatics. Chlorotoluenes, chloronaphthalenes and chlorobenzene are used as solvents and substrates in the production of pesticides, dyes and disinfectants. It should be noted that global production of chlorotoluene isomers alone reached 130,000 tons in 2000 – of which 65,000 tons constituted a mixture of all three isomers – and that this production is increasing (Dobslaw and Engesser, 2012). The compounds, 2-chlorotoluene and 4-chlorotoluene, are precursors in the synthesis of active ingredients of drugs, pigments and cleaning agents (Rossberg et al., 2006).

Contact of living organisms with chlorotoluene isomers leads to weakening of the body, and breathing disorders in mammals (Rossberg et al., 2006). Moreover, chloroaromatic compounds may have toxic effects on microorganisms present in the environment (Lu et al., 2011). Therefore, their removal from the environment is extremely important. The use of biodegradation processes for this purpose not only permits removal of harmful substances from the environment, but also allows them to be transformed into non-toxic compounds or leads to their complete mineralization to inorganic compounds (Bhatt et al., 2007). Regardless of the metabolic pathway used by bacteria, one of the main factors limiting the rate of biodegradation is the bioavailability of contaminants to microorganism cells. In the field of environmental protection, bioavailability is understood as the quantity of a polluting chemical compound that can be collected and biodegraded by microorganisms. Hence, if the bioavailability of a compound is high in given conditions, its biodegradation is limited only by the rate of the biochemical reactions that make up the biodegradation pathway (Maier, 2000). In the biodegradation of aromatic compounds and their chlorinated derivatives, the most important factors include their hydrophobicity and low solubility in water. These parameters significantly limit the bioavailability of these compounds to cells. In addition, halogen derivatives of benzene and toluene undergo significant sorption on soil particles and sediments in water reservoirs, and their low rate of biodegradation leads to their accumulation (Jahan et al., 1999).

A relatively small proportion of naturally occurring microorganisms are able to degrade persistent organic pollutants. Bacterial species capable of biodegrading chlorinated aromatic compounds include certain strains from different genera, like Pseudomonas, Burkholderia (Field and Sierra-Alvarez, 2008), Alcaligenes (Field and Sierra-Alvarez, 2004), Achromobacter (Chaudhry and Chapalamadugu, 1991) or Comamonas (Duc, 2017). What is important, in the environment, e.g. soils, wetlands, river sediments, biodegradation is carried out by consortia of bacteria species mainly from described genera (Chen et al., 2015) Nevertheless, in these publications there is practically no information about biodegradation efficiency. Kaczorek et al. (2016) isolated the strain from Raoultella genera, which was able to use toluene and its three monochlorinated derivatives as only carbon and energy source. In turn, Rhodococcus sp. OCT 10 biodegraded only 2-chlorotoluene, but not other isomers, and additionally the biodegradation was observed only at high bacteria concentrations (Dobslaw and Engesser, 2012). The same authors described, that the used strain could be used in three-phase system in biofilters to remove gaseous 2-chlorotoluene (Dobslaw and Engesser, 2018). However, the majority of the above mentioned studies focused on metabolic pathways, but not on the cell surface properties and their impact on biodegradation efficiency. Cell surface properties are considered to have great impact on bioavailability and, consequently on the microbial uptake of hydrophobic compound (Kaczorek et al., 2016). Hence, the necessity of a deeper insight into the cell surface properties of bacterial strains degrading emerging pollutants, like chlorotoluene.

The research work reported in this paper contributes to respond to this necessity. The aim of the study presented was to characterize the properties of bacterial strains demonstrating good ability to biodegrade monochlorotoluene isomers from environmental samples. The first stage of the study included characterization of the isolated strains and assessment of their biodegradation potential. In addition, changes in cell surface properties were identified. These can significantly affect the bioavailability of biodegradable compounds to the cells and, as a consequence, the efficiency of biodegradation.

Section snippets

Chemicals and culture medium

The isomers of chlorotoluene (2-, 3- and 4-chlorotoluene) used in this study were purchased from AlfaAesar (United Kingdom). Other fine chemicals were of per analysis grade and were purchased from Sigma-Aldrich (Germany). Aqueous solutions were prepared with the use of deionized and ultrapurified MiliQ water. The culture medium was prepared according to Dobslaw and Engesser's method (Dobslaw and Engesser, 2012).

Bacterial strain isolation

In order to isolate bacterial strains capable of degrading chlorotoluenes, soil

Identification and biochemical profile

Isolated bacterial strains that were found to grow in the presence of chlorotoluene isomers as the only carbon and energy source were identified as Raoultella planticola SA2, Rahnella aquatilis DA2 and Pseudomonas sp. MChB (Fig. 1). Their nucleotide sequences have been deposited in the GeneBank database of NCBI (Table 1). All these strains are Gram-negative.

Environmental strains from the Raoutella, Pseudomonas and Rahnella genera often play an important role in processes of biodegradation of

Conclusions

Three strains capable of degrading chlorotoluene isomers – Raoultella planticola SA2, Rahnella aquatilis DA2 and Pseudomonas sp. MChB – were isolated from soil samples. The strain R. aquatilis DA2 was distinguished from the other two by causing alterations in the surface properties of the cells, which were an increased content of branched fatty acids, and the induced capacity for β-hemolysis. Despite these differences, all three strains demonstrated good ability to biodegrade the

Main findings

The results significantly increased the present knowledge about adaptation of bacteria strains to degradation of the hydrophobic and hazardous compounds.

Acknowledgments

This study was supported by the National Science Centre (Poland) under decision number DEC-2015/19/N/NZ9/02423.

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