Phytoremediation is a cost-effective and sustainable technology used to clean up pollutants from soils and waters through the use of plant species. Indeed, plants are naturally capable of absorbing metals and degrading organic molecules. However, in several cases, the presence of contaminants causes plant suffering and limited growth. In such situations, thanks to the production of specific root exudates, plants can engage the most suitable bacteria able to support their growth according to the particular environmental stress. These plant growth-promoting rhizobacteria (PGPR) may facilitate plant growth and development with several beneficial effects, even more evident when plants are grown in critical environmental conditions, such as the presence of toxic contaminants. For instance, PGPR may alleviate metal phytotoxicity by altering metal bioavailability in soil and increasing metal translocation within the plant. Since many of the PGPR are also hydrocarbon oxidizers, they are also able to support and enhance plant biodegradation activity. Besides, PGPR in agriculture can be an excellent support to counter the devastating effects of abiotic stress, such as excessive salinity and drought, replacing expensive inorganic fertilizers that hurt the environment. A better and in-depth understanding of the function and interactions of plants and associated microorganisms directly in the matrix of interest, especially in the presence of persistent contamination, could provide new opportunities for phytoremediation.

Three differently metabolically engineered strains, 2 single PHA ) and Hup ) mutants and one double PHA ) /Hup ) mutant, of the purple nonsulfur photosynthetic bacterium Rhodobacter sphaeroides RV, were constructed to improve a light-driven biohydrogen production process combined with the disposal of solid food wastes. These phenotypes were designed to abolish, singly or in combination, the competition of H 2 photoproduction with polyhydroxyalkanoate (PHA) accumulation by inactivating PHA synthase activity, and with H 2 recycling by abolishing the uptake hydrogenase enzyme. The performance of these mutants was compared with that of the wild-type strain in laboratory tests carried out in continuously fed photobioreactors using as substrates both synthetic media containing lactic acid and media from the acidogenic fermentation of actual fruit and vegetable wastes, containing mainly lactic acid, smaller amounts of acetic acia, and traces of higher volatile acids. With the lactic acid-based synthetic medium, the single Hup ) and the double PHA ) / Hup ) mutants, but not the single PHA ) mutant, exhibited increased rates of H 2 photoproduction, about one third higher than that of the wild-type strain. With the food-waste-derived growth medium, only the single Hup ) mutant showed higher rates of H 2 production, but all 3 mutants sustained a longer-term H 2 photoproduction phase than the wild-type strain, with the double mutant exhibiting overall the largest amount of H 2 evolved. This work demonstrates the feasibility of single and multiple gene engineering of microorganisms to redirect their metabolism for improving H 2 photoproduction using actual waste-derived substrates.

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A large operon-type structure has been located between the g/tA and citB loci on the Bacillus subtilis chromosome. On the basis of the analysis of the 25 kb sequenced so far, it potentially encodes at least three large proteins which contain structural motifs associated with the subunits of all characterized peptide synthases. The amino acid recognition specificity of this new peptide synthase is discussed in the light of sequence homology with other synthases.Keywords : Bacillus subtilis, peptide synthase operon, racemase Micro-organisms produce a large number of peptides, generally as secondary metabolites, which have many important activities varying from antibacterial to antiviral or antifungal, and from anticarcinogenic to immunosuppressive. Regardless of their function, these peptides are synthesized either using the usual ribosomal system or through the action of complex multi-enzyme systems known as peptide synthases.It is now clear that although their products can vary considerably in length and amino acid composition, all peptide synthases studied so far share remarkable similarity in their structural organization and mechanism of action (Kleinkauf & von Dohren, 1990; Cosmina e t al., 1993). They are in fact organized into structural domains, each of which is responsible for the recognition and binding of a specific amino acid. Peptide synthesis takes place through subsequent reactions of thioester cleavage and amide bond formation.Bacillzrs szxbtilis produces a number of peptides and lipopeptides, both linear and cyclic (Zuber etal., 1993). Of these, only surfactin has been thoroughly characterized at the genetic level (Cosmina e t al., 1993; van Sinderen e t al., 1993). From gene sequence analysis and a variety of biochemical studies, it was found that surfactin synthase is a multi-subunit enzyme complex in which there are seven structural domains, corresponding to the number of amino acids present in surfactin.In this communication we report the identification in the B. mbtilis chromosome of a large operon encoding a new peptide synthase.The 15.4 kb fragment present in A clone AB21 (Fig. l) (Kunst & Devine, 1991), two additional A clones (A1A and A4B) were identified by D N A : DNA hybridization, the inserts of which overlap with the chromosomal fragment carried by AB21 (Fig. 1). Finally, an additional 4.3 kb overlapping fragment was isolated by chromosome walking (Glaser et a/., 1993) after cloning the AB21 HindIII-AcyI fragment into the suicide vector pDIA5304. Altogether, the three A clones and the pDIA5304 derivative contained an uninterrupted chromosomal region of approximately 25 kb.The identity of the 25 kb region carried by the A and plasmid vectors with the corresponding chromosomal area was confirmed by both restriction enzyme and Southern blot analyses. In particular, when AB21, A4B and A1A were digested with EcoRI and NotI, fragments of identical size were identified on an agarose gel. Furthermore, the large 3.0 kb EcoRI fragment of the three clones ( Fig. 1) hybridized with an intern...

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