3.5.5S
MOLECULAR APPROACHES FOR ELUCIDATING THE IN SITU ACTIVITIES OF BIOLOGICAL CONTROL AGENTS AND THE NATURE OF BACTERIAL HABITATS IN THE RHIZOSPHERE


JE LOPER, VO STOCKWELL and MD HENKELS

US Department of Agriculture, Agricultural Research Service, Horticultural Crops Research Laboratory, 3420 N.W. Orchard Ave., Corvallis, Oregon, 97330, USA

Certain strains of bacteria suppress plant pathogenic bacteria and fungi in the rhizosphere through processes of resource competition or antibiosis. These processes are not likely to occur uniformly in time or space; instead, they undoubtedly take place only in those microhabitats where environmental factors favor their occurrence, and during periods in the bacterial life cycle when their genetic determinants are expressed. Molecular approaches are now available to assess the chemical nature of habitats that bacteria occupy in the rhizosphere, to determine if resource availability to microorganisms is influenced by their coinhabitants in the rhizosphere, and to assess the activities and physiological status of bacteria inhabiting the rhizosphere.

It is widely recognized that bacteria inhabiting rhizosphere soil are present in microcolonies where concentrations of nutrients and microbial metabolites may vary greatly from those in bulk soil solutions. Numerous biotic and abiotic factors influence the immediate chemical environment of bacterial cells in the rhizosphere, and reporter gene systems provide powerful methods for assessing biologically-meaningful levels of specific compounds or elements in microhabitats occupied by bacteria. A biological sensor constructed by fusing the ice nucleation reporter gene to an iron-regulated promoter has been useful in elucidating temporal alterations in iron availability to Pseudomonas fluorescens in the rhizosphere [1]. Reporter gene systems have also provided some of the first glimpses into signalling that occurs between bacterial coinhabitants of the rhizosphere [3]. Recent evidence indicates that bacteria recognize compounds, (i.e. N-acyl-homoserine lactone autoinducers or ferric-siderophore complexes), produced by their coinhabitants in the rhizosphere and respond through alterations in their own patterns of gene expression [3, Henkels and Loper, unpublished). Our understanding of communication that occurs between bacteria in natural habitats is yet undeveloped, but further studies investigating these phenomena promise to illuminate the importance of microbial community context in the activities of biological control agents.

Rhizosphere bacteria exist in a commensal relationship with plants, utilizing amino acids, sugars and other organic compounds exuded by roots. Nevertheless, the rhizosphere can be a hostile environment, in which bacterial population size is restricted by nutrient limitation or by exposure to environmental and physiological stresses. Recent evidence indicates that strains of P. fluorescens exist, for at least a portion of their lives in the rhizosphere, in a physiological state approximating the stationary phase of the bacterial life cycle [2]. Studies utilizing molecular approaches are providing insights into the physiological status and metabolic activities of bacteria inhabiting the rhizosphere.

References
1. Loper JE, Henkels, MD, 1997. Applied and Environmental Microbiology 63,99-105.
2. Sarniguet A, Kraus J, Henkels MD, Muehichen AM, Loper JE, 1995. Proceedings of the National Academy of Sciences (USA) 92, 12255-12259.
3. Wood DW, Gong F, Daykin MM, Williams P, Pierson LS, 1997. Journal of Bacteriology 179,7663-7670.