Calcium carbonate precipitation, a widespread phenomenon among bacteria, has been investigated due to its wide range of scientific and technological implications. Nevertheless, little is known of the molecular mechanisms by which bacteria foster calcium carbonate mineralization. In our laboratory, we are studying calcite formation byBacillus subtilis, in order to identify genes involved in the biomineralization process. A previous screening of UV mutants and of more than one thousand mutants obtained from the EuropeanB. subtilisFunctional Analysis project allowed us to isolate strains altered in the precipitation phenotype. Starting from these results, we focused our attention on a cluster of five genes (lcfA,ysiA,ysiB,etfB, andetfA) called thelcfAoperon. By insertional mutagenesis, mutant strains carrying each of the five genes were produced. All of them, with the exception of the strain carrying the mutatedlcfAoperon, were unable to form calcite crystals. By placing transcription under IPTG (isopropyl-β-d-thiogalactopyranoside) control, the last gene,etfA, was identified as essential for the precipitation process. To verify cotranscription in thelcfAoperon, reverse transcription-PCR experiments were performed and overlapping retrocotranscripts were found comprising three adjacent genes. The genes have putative functions linked to fatty acid metabolism. A link between calcium precipitation and fatty acid metabolism is suggested.

Chemical extraction, water retting, microbiological and enzymatic methods were applied on entire nettle stalks and/or unretted decorticated fiber of a selected fiber nettle clone. Morphological and mechanical properties and chemical composition were then determined on fiber samples. The first interesting result concerned the good degree of separation between fibers and shives obtained by mechanical scutching applied on stalks stored for 1 year, probably resulting from natural retting processes occurring during the storage. Microbiological retting (anaerobic plus aerobic bacteria) of entire stalks and/or unretted decorticated fiber produced fibers with a higher quality than water retting. Both enzymes used (Viscozyme ^® L and Pectinex^® Ultra SP-L), improved fiber quality if EDTA was added. The enzyme vat retting gave good results on both water-retted fibers and unretted decorticated fibers, while the spray enzyme treated fibers usually displayed thicker diameter, lower cellulose content and, for Viscozyme ^® L, lower strength values, without differences between the two storage methods used after enzyme application.