Isolation and Characterization of Pseudomonas spp. Strains That Efficiently Decompose Sodium Dodecyl Sulfate

Abstract

Due to their particular properties, detergents are widely used in household cleaning products, cosmetics, pharmaceuticals, and in agriculture as adjuvants tailoring the features of pesticides or other crop protection agents. The continuously growing use of these various products means that water soluble detergents have become one of the most problematic groups of pollutants for the aquatic and terrestrial environments. Thus it is important to identify bacteria having the ability to survive in the presence of large quantities of detergent and efficiently decompose it to non-surface active compounds. In this study, we used peaty soil sampled from a surface flow constructed wetland in a wastewater treatment plant to isolate bacteria that degrade sodium dodecyl sulfate (SDS). We identified and initially characterized 36 Pseudomonas spp. strains that varied significantly in their ability to use SDS as their sole carbon source. Five isolates having the closest taxonomic relationship to the Pseudomonas jessenii subgroup appeared to be the most efficient SDS degraders, decomposing from 80 to 100% of the SDS present in an initial concentration 1 g/L in less than 24 h. These isolates exhibited significant differences in degree of SDS degradation, their resistance to high detergent concentration (ranging from 2.5 g/L up to 10 g/L or higher), and in chemotaxis toward SDS on a plate test. Mass spectrometry revealed several SDS degradation products, 1-dodecanol being dominant; however, traces of dodecanal, 2-dodecanol, and 3-dodecanol were also observed, but no dodecanoic acid. Native polyacrylamide gel electrophoresis zymography revealed that all of the selected isolates possessed alkylsulfatase-like activity. Three isolates, AP3_10, AP3_20, and AP3_22, showed a single band on native PAGE zymography, that could be the result of alkylsulfatase activity, whereas for isolates AP3_16 and AP3_19 two bands were observed. Moreover, the AP3_22 strain exhibited a band in presence of both glucose and SDS, whereas in other isolates, the band was visible solely in presence of detergent in the culture medium. This suggests that these microorganisms isolated from peaty soil exhibit exceptional capabilities to survive in, and break down SDS, and they should be considered as a valuable source of biotechnological tools for future bioremediation and industrial applications.

Keywords: Pseudomonas sp.; SPME-GC-MS; alkyl sulfatase; biodegradation; biodiversity; sodium dodecyl sulfate; surface flow constructed wetland; xenobiotics.

FIGURE 1

Identification of bacteria potentially degrading sodium dodecyl sulfate (SDS). Comparison of growth profile of selected strains cultured on solidified 0.1X LB (A) or basal medium supplemented with 0.2% SDS (B). Differences in clear zone diameter are observed for each strain cultured on SDS-containing medium indicate a variety of degree of detergent metabolism.

FIGURE 2

Study of SDS degradation of selected isolates. (A) Correlation of growth rate and SDS degradation (decrease in substrate concentration [%]) after 24 h of incubation plotted for the 36 selected strains. The dotted line represents correlation. (B) Time course study of degree of SDS degradation of the top five selected strains. (C) Time course study of growth rate for the five most effective degraders selected during the experiment. The values are means of the three replicates, and the error bars indicate the standard deviations.

FIGURE 3

Biodiversity of AP3 soil sample based on bioinformatics analysis of deep sequencing of AP3MET library. Community composition at different taxonomic ranks: (A) phylum level. (B) 10 most represented genera in the soil sample.

FIGURE 4

Phylogenetic tree of selected isolates among chosen representatives of Pseudomonas. The phylogenetic tree was computed by Maximum Likelihood with 1,000 bootstrap replicates under an Tamura-Nei model. Examined isolates are marked with black triangles.

FIGURE 5

The effect of SDS concentration on the bacterial growth rates of selected strains reflected as optical density of cell cultures after 24 h of incubation. The values are means of the three replicates, and the error bars indicate the standard deviations. The dotted line represents the initial OD600 of each strain.

FIGURE 6

Drop plate assay for bacterial chemotaxis. Pictures of bacterial rings formed after 8 h incubation at 30°C. Rows represent selected strains’ chemotactic response toward glucose (I), SDS (II) and control plates without compound (III).

FIGURE 7

SPMS-GC-MS analysis of the SDS derivatives for selected strains after 24 h of incubation in minimal medium supplemented with 1 g/L SDS. The boxes indicate the peak positions for: 2-dodecanol and 3-dodecanol (green), dodecanal (blue), and dodecanol (red). MS-data (not-presented) from signals marked with asterisks were insufficient for certain confirmation of the presence of the compounds. However, the accordance of the retention times strongly suggests the presence of traces of these metabolites.

FIGURE 8

Native PAGE zymography of the five best SDS-degrading strains.