The objectives are: the development and evaluation of alternative vector systems without antibiotic resistance genes as selectable markers, and of suicide 
systems for the biological containment of soil microbial inoculants; risk assessment on the release of soil microorganisms altered in symbiotically relevant 
traits and evaluation of strains, recovered from inoculated fields, for genetic and phenotypic properties that may be altered since inoculation (computer 
assisted analysis).   
First year objectives of the project were:  
development of vector and reporter systems; exploit the natural selection method to obtain various Rhizobium and Bradyrhizobium strains defective in 
thymidilate synthase, evaluate the performance of thy vector in the rhizosphere under greenhouse and field conditions, and construction of inducible reporter 
systems.  
Construction of a suicide system for R meliloti; evaluate the performance of the R meliloti thy in the rhizosphere under greenhouse and field conditions, 
and genetically manipulate the Lactococcus lactis thy cartridge in preparation for a conditional expression system.  
Evaluation of genetically modified inoculum strains; during inoculant manufacturing process and at microcosm and greenhouse level.  
Evaluation of natural occurring populations; to use pyrolysis mass spectrometry (PYMS) to look for distances between isolates of Bradyrhizobium japonicum 
isolated from soybean plants grown in Italy, to study isolates from Italy using deoxyribonucleic acid (DNA) hybridization, to look at populations of Rhizobium 
in soil differing characteristics and from different geographic characteristics and from different geographic regions, and to look at the effect of factors 
such as manipulating the water table on survival.  
  
In work to date the thy system has proved highly effective in ensuring stable maintenance of the plasmid under field conditions. This was found to be the case 
even though the thy mutant strain was found to revert at a rather high frequency. This demonstrates that the pressure for the thy host strain to revert to a wild 
type. The lac and lac/Hg{r} cassettes provide a tool which is based on a catabolic inducible reporter system without antibiotic resistance markers. The expression 
levels, comparable or higher than those obtainable with tac promoter, allow unambiguous detection of the genetically modified microbe. The project is continuing.