Degradation of alkanes and highly chlorinated benzenes, and production of biosurfactants, by a psychrophilic Rhodococcus sp. and genetic characterization of its chlorobenzene dioxygenase

Abstract

Rhodococcus sp. strain MS11 was isolated from a mixed culture. It displays a diverse range of metabolic capabilities. During growth on 1,2,4-trichlorobenzene, 1,2,4,5-tetrachlorobenzene (1,2,4,5-TeCB) and 3-chlorobenzoate stoichiometric amounts of chloride were released. It also utilized all three isomeric dichlorobenzenes and 1,2,3-trichlorobenzene as the sole carbon and energy source. Furthermore, the bacterium grew well on a great number of n-alkanes ranging from n-heptane to n-triacontane and on the branched alkane 2,6,10,14-tetramethylpentadecane (pristane) and slowly on n-hexane and n-pentatriacontane. It was able to grow at temperatures from 5 to 30 degrees C, with optimal growth at 20 degrees C, and could tolerate 6 % NaCl in mineral salts medium. Genes encoding the initial chlorobenzene dioxygenase were detected by using a primer pair that was designed against the alpha-subunit (TecA1) of the chlorobenzene dioxygenase of Ralstonia (formerly Burkholderia) sp. strain PS12. The amino acid sequence of the amplified part of the alpha-subunit of the chlorobenzene dioxygenase of Rhodococcus sp. strain MS11 showed >99 % identity to the alpha-subunit of the chlorobenzene dioxygenase from Ralstonia sp. strain PS12 and the parts of both alpha-subunits responsible for substrate specificity were identical. The subsequent enzymes dihydrodiol dehydrogenase and chlorocatechol 1,2-dioxygenase were induced in cells grown on 1,2,4,5-TeCB. During cultivation on medium-chain-length n-alkanes ranging from n-decane to n-heptadecane, including 1-hexadecene, and on the branched alkane pristane, strain MS11 produced biosurfactants lowering the surface tension of the cultures from 72 to </=29 mN m(-1). Glycolipids were extracted from the supernatant of a culture grown on n-hexadecane and characterized by (1)H- and (13)C-NMR-spectroscopy and mass spectrometry. The two major components consisted of alpha,alpha-trehalose esterified at C-2 or C-4 with a succinic acid and at C-2' with a decanoic acid. They differed from one another in that one 2,3,4,2'-trehalosetetraester, found in higher concentration, was esterified at C-2, C-3 or C-4 with one octanoic and one decanoic acid and the other one, of lower concentration, with two octanoic acids. The results demonstrate that Rhodococcus sp. strain MS11 may be well suited for bioremediation of soils and sediments contaminated for a long time with di-, tri- and tetrachlorobenzenes as well as alkanes.

Biosurfactants Production from Low Cost Substrate and Degradation of Diesel Oil by a Rhodococcus Strain

“…The analysis of fatty acids in the culture broth was carried out as described in [14]: after the fermentation is completed, the residual RSFO was removed using the conventional repeated decantation technique. Broth culture was acidified with conc.…”

“…A crude biosurfactant was extracted from the cell-free culture supernatant of R. erythropolis 16 LM-USTHB grown on residual sunflower frying oil, using the technique described in [14]; briefly, at the end of the cultivation on RSFO, and after the hydrophobic layer located at the surface has been removed, the culture medium was centrifuged at 12 000 g at 4°C. The supernatant was extracted by means of the organic solvent system CH 2 Cl 2 /CH 3 OH (2:1, v/v); the extract was concentrated under reduced pressure: resulting crude extract.…”

Diesel degradation and biosurfactant production by Gram-positive isolates

“…Their properties of interest include: (i) changing surface active phenomena, such as lowering of surface and interfacial tensions, (ii) wetting and penetrating actions, (iii) spreading, (iv) hydrophylicity and hydrophobicity actions, (v) microbial growth enhancement, (vi) metal sequestration and (vii) antimicrobial action (Kosaric, 2001). Among these biosurfactants, glycolipids are the most important group as they show a high surface or interfacial activity as well as pH and temperature stability, low toxicity and good biodegradability (Rapp and Gabriel-Jürgens, 2003). Amid the main types of biosurfactants produced by microorganisms, surfactin is one of the most well recognized products with commercial application (Menezes Bento et al, 2005).…”

Oil-Spill Bioremediation, Using a Commercial Biopreparation "MicroBak" and a Consortium of Plasmid-Bearing Strains "V&O" with Associated Plants

“…Previous research on biosurfactants produced by rhodococci indicated that the main signals in the mass-spectra of electron spray were induced by pseudo-molecular ions [M + Na +] with masses of 871.5 and 899.6 (Rapp & Gabriel-Jurgens, 2003;Tuleva et al, 2008). That corresponded to the homologous succinoyl trehalose lipids: dioctanoyl-decanoyl (848 Da) and octanoyl-didecanoyl (876 Da), differing by 28 Da, i.e.…”