Pseudomonas putida DOT-T1 was isolated after enrichment on minimal medium with 1% (vol/vol) toluene as the sole C source. The strain was able to grow in the presence of 90% (vol/vol) toluene and was tolerant to organic solvents whose log P(ow) (octanol/water partition coefficient) was higher than 2.3. Solvent tolerance was inducible, as bacteria grown in the absence of toluene required an adaptation period before growth restarted. Mg2+ ions in the culture medium improved solvent tolerance. Electron micrographs showed that cells growing on high concentrations of toluene exhibited a wider periplasmic space than cells growing in the absence of toluene and preserved the outer membrane integrity. Polarographic studies and the accumulation of pathway intermediates showed that the strain used the toluene-4-monooxygenase pathway to catabolyze toluene. Although the strain also thrived in high concentrations of m- and p-xylene, these hydrocarbons could not be used as the sole C source for growth. The catabolic potential of the isolate was expanded to include m- and p-xylene and related hydrocarbons by transfer of the TOL plasmid pWW0-Km.

The development of tolerance in Pseudomonas putida DOT-T1 to toluene and related highly toxic compounds involves short- and long-term responses. The short-term response is based on an increase in the rigidity of the cell membrane by rapid transformation of the fatty acid cis-9,10-methylene hexadecanoic acid (C17:cyclopropane) to unsaturated 9-cis-hexadecenoic acid (C16:1,9 cis) and subsequent transformation to the trans isomer. The long-term response involves in addition to the changes in fatty acids, alterations in the level of the phospholipid polar head groups: cardiolipin increases and phosphatidylethanolamine decreases. The two alterations lead to increased cell membrane rigidity and should be regarded as physical mechanisms that prevent solvent penetrance. Biochemical mechanisms that decrease the concentration of toluene in the cell membrane also take place and involve: (i) a solvent exclusion system and (ii) metabolic removal of toluene via oxidation. Mutants unable to carry out cis → trans isomerization of unsaturated lipids, that exhibit altered cell envelopes because of the lack of the OprL protein, or that are unable to exclude toluene from cell membranes are hypersensitive to toluene.

A bacterium, Pseudomonas sp. strain C1S1, able to grow on 2,4,6-trinitrotoluene (TNT), 2,4- and 2,6-dinitrotoluene, and 2-nitrotoluene as N sources, was isolated. The bacterium grew at 30 degrees C with fructose as a C source and accumulated nitrite. Through batch culture enrichment, we isolated a derivative strain, called Pseudomonas sp. clone A, which grew faster on TNT and did not accumulate nitrite in the culture medium. Use of TNT by these two strains as an N source involved the successive removal of nitro groups to yield 2,4- and 2,6-dinitrotoluene, 2-nitrotoluene, and toluene. Transfer of the Pseudomonas putida TOL plasmid pWW0-Km to Pseudomonas sp. clone A allowed the transconjugant bacteria to grow on TNT as the sole C and N source. All bacteria in this study, in addition to removing nitro groups from TNT, reduced nitro groups on the aromatic ring via hydroxylamine to amino derivatives. Azoxy dimers probably resulting from the condensation of partially reduced TNT derivatives were also found.

The goal of this work was to determine the chemical nature of the red pigment produced by Streptococcus agalactiae , which has been thought to be a carotene. We extracted the pigment with 0.1 M KOH and purified it by column chromatography on Sephadex LH. Data from elemental analysis and mass and nuclear magnetic resonance spectra lead us to propose the structure to be that of a new ornithine rhamno-polyene with 12 conjugated double bonds, to which we have assigned the trivial name granadaene.