No abstract

The influence of iron on the entry of Listeria monocytogenes into Caco-2 cells was studied. Iron availability was found to modify the surface hydrophobicity and protein profile of L. monocytogenes, with the result that cell invasion strongly increased upon bacterial growth in iron-rich medium. The enhanced invasive capability of iron-overloaded L. monocytogenes cells correlates to the higher-level expression of the inlAB virulence genes, which were positively iron regulated at the transcriptional level.

The growth of seven Thiobacillus ferrooxidans strains was trested on various solid media prepared according to previously published procedures in comparison with a series of new formulations (TSMs, Thiobacillus solid media) set up in our laboratory. These media differed in the type of gelling agent and in the concentration of phosphate and ferrous ions. It was determined that the established formulations brought qualitatively and quantitatively unsatisfactory results whereas, among the new media, TSM 1 produced quantitative yields of T. ferrooxidans colonies. The TSM 1 supports the growth of T. ferrooxidans strains and can be recommended for the estimation of cell numbers in liquid cultures and for the isolation of single clones. There was also a noticeable similarity in the morphology of colonies formed by different Ts ferrooxidans strains.The chemoautotrophic bacterium Thiobacillus ferrooxidans is able to obtain energy from the oxidation of ferrous iron and/or inorganic sulfur compounds. An important feature of this extremophile is its ability to grow and reproduce readily at the lower extremes of the pH scale (1.5-2.0) (3).Studies of the biochemistry and physiology of this microorganism have been stimulated by the ever increasing development of the bioleaching processes that are used industrially to recover metals from ores and to desulphurize coal (2,11). In spite of this interest in T, ferrooxidans, and its genetic improvement for biomining operations (15) much work has still to be done to select efficient strains for making the leaching systems commercially feasible. The lack of studies of the genetic manipulation of this bacterium is mainly concerned with the difficulty of applying * Address reprint requests to: Dr .

Shift of three Thiobacillus fi'rrooxidans strains from Fe(ll) to S" or thiosulphate liquid medium caused distinctive changes in the outer membrane protein profile. In addition to a new 55-kDa protein which was synthesized only in the presence of sulphur compounds, a higher expression of a 47-kDa protein was observed. This latter protein appeared to be constitutively synthesized, since it was detectable in small amounts even in tile presence of ferrous iron as sole energy source, but its expression was greatly enhanced when elemental sulphur or thiosulphate were present in the growth medium.

Thiobacillus ferrooxidans is a gram-negative, acidophilic, chemolithotrophic bacterium oxidizing ferrous to ferric ions and reduced sulfur compounds to sulfate. These unusual physiological features enable this bacterium to play an important role in the leaching of metals from low grade ores (8). The metabolism of T. ferrooxidans is fairly well known, while studies on its genetic organization started only a few years ago and are still in progress. Plasmid occurrence in this species has been shown since 1980 (2) but no plasmid function has been conclusively identified.In this research we focused our attention on plasmids from T. ferrooxidans in view of the flexibility of manipulation of these DNA molecules, which could make possible an improvement in the leaching capabilities of industrially used strains by inserting into them suitable plasmid determinants from other sources.Twelve strains of T. ferrooxidans isolated from different parts of the world (Table 1) have been examined for plasmid content and metal-resistance with the aim of correlating plasmid profiles with relevant phenotypic traits.To improve yields and to obtain reproducible results in the extraction of plasmids from Ts ferrooxidans we modified the alkaline extraction method of plasmid DNA conceived by Birnboim in 1983 (1). Bacterial cultures in late stationary phase (96-120 h old) grown in 9K liquid medium (7) were filtered on Whatman N1 paper to remove iron precipitates. Bacteria were harvested by centrifugation (5,000 x g, 20 min, 4°C) and washed twice in 1 mM H2SO4 and once in TE buffer (10 mM Tris-HC1 pH 7.5, 1 mM EDTA pH 8). The lysozyme solution was substituted by * Address reprint requests to: I)r . M. Polidoro,