Conjugal opines secreted by crown gall tumours induce strains of Agrobacterium tumefaciens that are donors of Ti plasmids to produce a diffusible conjugation factor. This enhances the conjugal transfer efficiency of the Ti plasmid in other strains of A. tumefaciens. This factor behaves as a secondary messenger, transmitting the environmental information to tra genes. Here we report the use of spectrometry to show that this factor is identical to synthetic N-(beta-oxo-octan-1-oyl)-L-homoserine lactone and confirm that the synthetic compound is biologically active. N-(Hexan-1-oyl)-L-homoserine lactone has also been detected. A closely related molecule, N-(beta-oxo-hexan-1-oyl)-L-homoserine lactone, autoinduces bioluminescence in the distantly related bacterium, Vibrio fischeri. N-Acyl-homoserine lactones thus seem to be conserved molecules in which the length and nature of the lipophilic acyl chain determines the biological function to be regulated. Mutants that do not produce the factor fail to conjugate unless supplied with it in the induction medium (our unpublished data). These data indicate that the conjugation factor is an autoinducer and a key signal molecule in the conjugation system of A. tumefaciens. It is, to our knowledge, the first example of a second messenger molecule in a bacterial conjugation system.

Conjugal transfer of the nopaline-type Agrobacterium Ti plasmid pTiC58 is regulated by a trasriptional activator, TraR, and a diffusible signal molecule, conjugation factor (CF). CF is a member of a family of substituted homoserine lactones (HSLs) that act as coinducers for regulating gene expression in diverse Gram-negative bacteria by a mechanism called antoinduction. In Vibrio fischeur HSL production is conferred by the luxI gene. Homologues of this gene are responsible for HSL production by other Gram-negative bacteria. A gene that we call tral, conferring production of material with CF activity, was localized to a 1-kb region at the upstream end of tra of pTiC58. Spectroscopy showed that the activity was authentic CF. Sequence analysis showed that Otn could encode a protein of 211 amino acids, TraI, that is related to the proteins responsible for HSL production by other bacteria. A second, partial open reading frame immiatel downstream of tral could encode a protein related to TrbB of plasmid RP4, which is required for conjugal transer. Transcription of tnal and of the downstream tra3 genes requires TraR and CF and initiates from the tral promoter. The results show that tnd is responsible for CF production, that it is the first gene of the n3 operon, and that expression of this operon is regulated by autoinduction.

Organic materials responsible for water-repellency in some Australian soils were extracted with an amphiphilic mixture of iso-propanol/l5.7 M ammonia (7 : 3, v : v) in a Soxhlet apparatus, after which the water-repellent soils were rendered wettable. The successful extraction by an organic solvent system indicates that the bulk of hydrophobicity in these soils is not covalently linked to the surface of the sand. The extracted materials restored hydrophobicity on acid washed sands or ignited sands at levels comparable to the original soils.Spectroscopic and chromatographic examination of the extracted materials indicated that both free and esterified long-chain, 1G32 carbon atom, fatty acids were present with a bimodal distribution showing maxima at C,, and CZ2. The I3C-NMR and infrared spectra of the most hydrophobic extract suggest that hydrophobicity is caused by molecules with extensive polymethylene chains. Calculations with model compounds indicate that at least a close packed monolayer is required before measurable hydrophobicity can be detected with the molarity of ethanol droplet penetration test.

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