Funding Information This study was supported by a PhD grant to ST from the Fund for Scientific Research-Flanders, Belgium (FWO-Vlaanderen) and the UHasselt Methusalem project 08M03VGRJ.
Potential for plant growth promotion by a consortium of stress-tolerant 2,4-dinitrotoluene-degrading bacteria: isolation and characterization of a military soil
Article first published online: 28 JAN 2014
© 2014 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.
This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.
Volume 7, Issue 4, pages 294–306, July 2014
How to Cite
Thijs, S., Weyens, N., Sillen, W., Gkorezis, P., Carleer, R. and Vangronsveld, J. (2014), Potential for plant growth promotion by a consortium of stress-tolerant 2,4-dinitrotoluene-degrading bacteria: isolation and characterization of a military soil. Microbial Biotechnology, 7: 294–306. doi: 10.1111/1751-7915.12111
- Issue published online: 5 JUN 2014
- Article first published online: 28 JAN 2014
- Manuscript Accepted: 17 DEC 2013
- Manuscript Received: 29 JUL 2013
- Scientific Research-Flanders
- UHasselt Methusalem. Grant Number: 08M03VGRJ
Fig. S1. Photographs of the investigated field soils at the military training range in the North-East of Belgium. Left, the bare 2,4-DNT contaminated soil with bomb remnants and detonation container. On the left side, the adjacent grassland soil colonized by Molinia caerulea and Agrostis tenuis, photograph was taken in November 2009.
Fig. S2. Scatterplot of Biolog EcoPlate responses (average net area under the curve for each of the 31 carbon sources) comparing the grassland soil winter (GS-w), 2,4-DNT contaminated soil summer (DS-s) and winter (DS-w) inoculum metabolic profiles to that obtained for the grassland soil summer (GS-s) inoculum (plotted on x-axis).
Fig. S3. Microscopic images of bacteria containing PHB inclusions using the lipid Sudan stain and saffranin counterstain. The bacteria strains are (A) Rhodanobacter sp. HC50 and (B) Pseudomonas sp. HC94. (C) negative control, bacterium with no lipid granules inclusions. The blue dots in the cytoplasma are the lipid-granules, indicated with the arrows. (Nikon Eclipse 80i, oil-immersion objective)
Fig. S4. Clear halos appearing on the opaque white plates indicate bacterial phosphorous solubilisation. A phosphate-buffer washed suspension of bacteria, pre-grown in rich medium was inoculated into the holes. All strains were tested in triplicate.
Fig. S5. Chemotactic response and flagella stain of Ralstonia sp. HC90. (A) Chemotactic response towards aspartic acid and (B) 2,4-DNT. (C) Negative control, lack of chemotaxis of heat-killed cells towards 2,4-DNT. Red arrows indicate the chemotaxis rings. Plates were scanned with a flatbed scanner. (D) Leifson flagella stain of Ralstonia sp. HC90, black arrow indicates flagella. Photograph was taken with oil-immersion objective (1500×).
Fig. S6. Photographs of the Arabidopsis thaliana plants on VAPS-plates (A) control; (B) UHS3 inoculated control; (C) exposed to 2,4-DNT and (D) exposed to 2,4-DNT with UHS3 inoculation, grown at a density of 5 plants per plate. The photograph was taken 9d after transfer of the seedlings when root length measures were taken, root hair pictures and the shoots were collected for reisolation of the inoculated bacteria.
Fig. S7. UPGMA dendrogram showing the relationships among the bacteria isolated from A. thaliana shoot tissues and the inoculated strains from consortium UHS3.
Table S1. Physicochemical characteristics of the 2,4-DNT contaminated soil and adjacent grassland soil. Nitro-aromatic concentrations in the samples were measured with HPLC and are expressed in mg kg −1 dry soil with standard error. EC: Electric Conductivity; 2,4-dinitrotoluene (2,4-DNT); 2,6-dinitrotoluene (2,6-DNT); 1,3-dinitrobenzene (DNB); 1,3,5-trinitrobenzene (TNB); nitrobenzene (NB); 2-amino-4-nitrotoluene (2-A-4NT); 4-amino-2-nitrotoluene (4-A-2NT). ND = not detected (< 0.01 mg l−1 ). DS-s: 2,4-DNT contaminated soil summer; DS-w: 2,4-DNT contaminated soil winter; GS-s: grassland soil summer; GS-w: grassland soil winter.
Table S2. Fingerprint of the inocula tested with Ecoplates. The absorbance data of the Biolog Ecoplates were blanked to the water containing well and then converted into Boolean ones, considering positive (1) the substrates for which the blanked absorbance data > 0.10 and negative (0) the substrates for which the blanked absorbance data < 0.10. DS-s: 2,4-DNT contaminated soil summer; DS-w: 2,4-DNT contaminated soil winter; GS-s: grassland soil summer; GS-w: grassland soil winter.
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