Winnie Dejonghe

Diversity of Transconjugants that Acquired Plasmid pJP4 or pEMT1 after Inoculation of a Donor Strain in the A- and B-horizon of an Agricultural Soil and Description of Burkholderia hospita sp. nov. and Burkholderia terricola sp. nov.

We examined the diversity of transconjugants that acquired the catabolic plasmids pJP4 or pEMT1, which encode degradation of 2,4-dichlorophenoxyacetic acid (2,4-D), in microcosms with agricultural soil inoculated with a donor strain (Dejonghe, W., Goris, J., El Fantroussi, S., Höfte, M., De Vos, P., Verstraete, W., and Top, E. M. Appl. Environ. Microbiol. 2000, p. 3297-3304). Using repetitive element PCR fingerprinting, eight different rep-clusters and six separate isolates could be discriminated among 95 transconjugants tested. Representative isolates were identified using 16S rDNA sequencing, cellular fatty acid analysis, whole-cell protein analysis and/or DNA-DNA hybridisations. Plasmids pJP4 and pEMT1 appeared to have a similar transfer and expression range, and were preferably acquired and expressed in soil by indigenous representatives of Ralstonia and Burkholderia. Two rep-clusters were shown to represent novel Burkholderia species, for which the names Burkholderia hospita sp. nov. and Burkholderia terricola sp. nov. are proposed. When easily degradable carbon sources were added together with the plasmid-bearing donor strain, also a significant proportion of Stenotrophomonas maltophilia isolates were found. The transconjugant collections isolated from A- (0-30 cm depth) and B-horizon (30-60 cm depth) soil were similar, except for B. terricola transconjugants, which were only isolated from the B-horizon.

Diversity of 3‐chloroaniline and 3,4‐dichloroaniline degrading bacteria isolated from three different soils and involvement of their plasmids in chloroaniline degradation

Attempts were made to isolate 3-chloroaniline (3-CA) and 3,4-dichloroaniline (3,4-DCA) degrading bacteria from the A- and B-horizon of three different soils. A variety of 3-CA degrading bacteria was obtained from all soils, whereas 3,4-DCA degrading strains were only isolated from one soil. Amongst the 3-CA and 3,4-DCA degraders, two belong to the gamma-Proteobacteria and seven to the beta-Proteobacteria. Of the latter group, five are members of the family of the Comamonadaceae. Interestingly, all isolates contained an IncP-1beta plasmid. These plasmids could be divided into four major groups based on restriction digest patterns. While all plasmids that were detected in the isolates, except one, encode total degradation of 3-CA, no indigenous plasmid that codes for total degradation of 3,4-DCA was found. This is the first study that reports the presence of diverse transferable plasmids that encode mineralisation of 3-CA in different 3-CA degrading species.

Proteomic study of linuron and 3,4-dichloroaniline degradation by Variovorax sp WDL1: evidence for the involvement of an aniline dioxygenase-related multicomponent protein

A proteomic approach was used to explore the metabolism of the phenylurea herbicide linuron and 3,4-dichloroaniline (3,4-DCA) in Variovorax sp. WDL1. This bacterium grows on linuron as sole source of carbon, nitrogen and energy, while it transiently accumulates 3,4-DCA as a metabolite. Differential protein expression analysis of Variovorax sp. WDL1 grown in a heterotrophic medium in the presence and absence of linuron or 3,4-DCA was conducted using 2-D PAGE. Selected up- and downregulated proteins were identified with nanoLC-ESI-MS/MS. In the 3,4-DCA-supplemented culture, upregulation of several proteins showing high amino acid sequence similarity to different components of the multicomponent aniline dioxygenase in aniline-degrading Proteobacteria was observed. For one of the components, multiple variant proteins were detected, suggesting that strain WDL1 harbors several copies of the aniline dioxygenase (AD) gene cluster which are simultaneously expressed in the presence of 3,4-DCA. A number of unidentifiable proteins, which were upregulated in the linuron- and/or 3,4-DCA-supplemented cultures, might represent up to now uncharacterized proteins with a role in linuron and/or 3,4-DCA degradation in strain WDL1. In addition, several stress-related proteins were differentially expressed.