Use molecular tools to determine ability of probiotics to hinder pathogens.

The success of applying probiotic cultures to limit the colonization of foodborne pathogens in the gastrointestinal tracts of food animals ensures the continued use of such cultures. But it is essential to develop a way to accurately identify and quantify these organisms during their commercial production, as well as during their in vivo colonization after they are administered to animals.

Also needed are ways to assess the expression of key metabolic processes that occur during the establishment of the probiotic culture, as well as its ability to limit a pathogen's colonization. Traditional techniques are used to study individual probiotic gastrointestinal organisms, such as selective plating to identify specific nutritional groups. But the requirement for strict anaerobiosis for the anaerobic members of these cultures can complicate the handling and preparation of samples.

Immunological methods can circumvent some of these problems. But they are limited to assessing functionality. Using molecular tools enables researchers to obtain the genetic diversity of the microflora as well as data on their gene activity. Such methods permit us to study the individual components of microbes, fluxes in the diversity of the microorganisms, their spatial distribution and the expression of specific microbial genes within a community of probiotics. These techniques involve using both DNA- and RNA-targeted probes, gene amplification protocols and mRNA analysis.

If further improvements of a culture's effectiveness and additional applications are to be attainable, we need new methods that assess expression of key metabolic processes that occur during the establishment of the probiotic culture, as well as its ability to limit foodborne pathogen colonization. Applying molecular techniques can help overcome the need to cultivate organisms before their detection.

Still, there are some issues to be resolved. Of primary concern is a practical way to acquire samples for molecular analysis. Sampling approaches based on an understanding of the environmental microbiology associated with housing livestock appear more feasible than sampling individual animals on farms. Direct sampling of aerosols from poultry houses has detected pathogens both by cultural means and gene amplification. This approach might couple with molecular detection of individual probiotic bacteria as they proliferate in the livestock house environment.

Future priorities will include: the use of real-time samples instead of spiked samples to determine efficacies of molecular techniques; the development of a cost-effective and efficient sampling plan that can detect specific probiotic organisms; and the use of round-robin laboratory evaluations to determine the true reproducibility and applicability of sampling protocols and assays.

Further information. Suresh Pillai, Institute of Food Science and Engineering, Texas A&M University, 418D Kleberg Center, 2472 TAMU, College Station, TX 77843; phone: [phone]; fax: [phone]; email: [e-mail].