The objective of this study was to develop an appropriate method for detecting specific bacteriocin-mediated inhibition. This method could be used to determine the effectiveness of known bacteriocin-producing lactic acid bacteria against foodborne pathogens. Lactococcus lactis subsp. lactis 11454, Pediococcus pentosaceous FBB61, and P. pentosaceous FBB63-DG2 were evaluated for their ability to inhibit a variety a foodborne pathogens. Of the three procedures used, the agar spot method was chosen as the most reproducible and reliable assay method. After optimization of the plate agar media composition, and the time and temperature of the assay, the three bacteriocin-producing strains were screened against a broad range of Gram-positive and Gram-negative foodborne pathogens. The three bacteriocin-producing organisms inhibited the following Gram-positive foodborne pathogens: Bacillus cereus, Clostridium perfringens. Staphylococcus aureus, and 22 representatives of 5 species of Listeria including L. monocytogenes, L. ivanovii, L. innocua, L. seeligeri, and L. welshimeri. Additionally, strain 11454 slightly inhibited several Gram-negative foodborne pathogens, including Aeromonas hydrophila AH2, Escherichia coli 0157:H7, Vibrio cholerae 851, and V. parahaemolyticus A865957. Gram-negative organisms not inhibited by any of the bacteriocin-producing organisms included Campylobacter jejuni, several serovars of Salmonella enteriditis, and species of Yersinia and Vibrio.

Recent advances in agricultural biotechnology have highlighted the need for experimental evidence and sound scientific judgment to assess the benefits and risks to society. Nutrition scientists and other animal biologists need a balanced understanding of the issues to participate in this assessment. To date most modifications to crop plants have benefited producers. Crops have been engineered to decrease pesticide and herbicide usage, protect against stressors, enhance yields and extend shelf life. Beyond the environmental benefits of decreased pesticide and herbicide application, consumers stand to benefit by development of food crops with increased nutritional value, medicinal properties, enhanced taste and esthetic appeal. There remains concern that these benefits come with a cost to the environment or increased risk to the consumer. Most U.S. consumers are not aware of the extent that genetically modified foods have entered the marketplace. Consumer awareness of biotechnology seems to have increased over the last decade, yet most consumers remain confused over the science. Concern over the impact on the safety of the food supply remains low in the United States, but is substantially elevated in Europe. Before a genetically engineered crop is introduced into commerce it must pass regulatory scrutiny by as many as four different federal regulatory bodies to ensure a safe food supply and minimize the risk to the environment. Key areas for more research are evaluation of the nutritional benefits of new crops, further investigation of the environmental impact, and development of better techniques to identify and track genetically engineered products.

The extracellular enzyme activities ofStreptococcus mutans 6715 that synthesize glucans from sucrose were concentrated and partially purified by ammonium sulfate precipitation and gel permeation column chromatography. Polyacrylamide gel analysis demonstrated that all of the major proteins precipitated by ammonium sulfate were quantitatively recovered in the high-molecularweight, enzyme-containing aggregates found in the void volume of the gel column. Anion-exchange column chromatography was used to fractionate the aggregates into preparations, a and /3, which produced water-insoluble and water-soluble glucans, respectively. Polyacrylamide gel analysis showed that a and ,3 contained unique proteins and dextransucrase (EC 2.4.1.5) activities. Studies on the time course of glucan synthesis by a demonstrated that this enzyme preparation contained dextranase activity, which partially degraded nascent alcohol-insoluble glucan into alcohol-soluble products that were subsequently reincorporated into insoluble product. The ,3 enzyme preparation contained no detectable dextranase activity. Mixing experiments in the absence of primer dextran demonstrated that the dextranase activity present in a could modify glucan production by /. CsCl density gradient analysis of product glucans demonstrated that exogenous primer dextrans were used as acceptor molecules by both the a and /8 enzyme preparations, and that water-soluble glucans synthesized by /8 could be converted into water-insoluble glucans by a.It is proposed that the structural heterogeneity of the native glucans produced from sucrose by S. mutans is a result of the concerted action of glucan-forming dextransucrases and endohydrolytic dextranase activity. 31). A significant effect of the presence of exogenous dextranase (Spicaria violaceae) during glucan synthesis by S. mutans dextransucrase on the quantity of a-(1--+3) linkages in the product glucans has been reported (7). Thus, the endogenous S. mutans dextranase may con-637 on July 16, 2020 by guest http://iai.asm.org/ Downloaded from cates the position of the bovine serum albumin standard run in an adjacent lane of the gel.

The effect of nonfermented dairy products containing yogurt or acidophilus cultures on lactose utilization by lactose-maldigesting humans was investigated. Yogurt and acidophilus milk containing 10(7) or 10(8) of Streptococcus thermophilus and Lactobacillus bulgaricus, or Lactobacillus acidophilus, respectively, were prepared using commercially processed 2% low fat milk. Immediately following inoculation, products were refrigerated. Lactose maldigestion was monitored by measuring breath hydrogen excretion at hourly intervals for 8 h following consumption of 400 ml of each test meal containing approximately 20 g of lactose. The yogurt milk containing 10(8) cfu/ml was shown to contain significant concentrations of microbial beta-galactosidase (EC 3.2.1.23; approximately 3 U/ml), which remained stable for at least 14 d at refrigerator temperatures. Breath hydrogen peaks were delayed and significantly lower (approximately 20 ppm at 5 to 7 h) than control values (approximately 70 ppm at 4 h), and intolerance symptoms were eliminated in all subjects. Yogurt milk containing 10(7) cfu/ml demonstrated intermediate breath hydrogen values and was marginally significantly different from control values. Lactobacillus acidophilus strains with varying resistance to bile and total beta-galactosidase-producing potential were also tested. Only one strain, LA-1, which demonstrated low bile resistance and intermediate beta-galactosidase activity, was capable of significantly decreasing breath hydrogen values when 10(8) cfu/ml of milk was consumed.

The effect of dextranases (EC 3.2.1.11) from the oral isolates Actinomyces israelii and Bacteroides ochraceus on water-insoluble glucan production by the Streptococcus mutans dextransucrase (EC 2.4.1.5) and sucrose-dependent adherence to smooth glass surfaces by S. mutans was studied. Collection on membrane filters of water-insoluble polysaccharides synthesized from radioactive sucrose was used to demonstrate the marked sensitivity of insoluble glucan formation to the presence of dextranase. Concentrations of A. israelii dextranase as low as 0.002 U/ml inhibited insoluble glucan formation by 60%. Similar results were obtained with the B. ochraceus enzyme. An assay for sucrose-stimulated adherence of S. mutans to smooth surfaces involved attachment of radioactively labeled nongrowing cells to the bottom of glass scintillation vials. This facile and sensitive assay was utilized to demonstrate that sucrose-dependent adherence was affected by low levels of dextranase from either A. israelii or B. ochraceus. Enzyme at 0.005 U/ml reduced adherence of S. mutans by 80%. Treatment of S. mutans cells previously attached to glass with low concentrations of the dextranases resulted in removal of 50 to 60% of the bacteria. These results indicate that dextranase-producing oral bacteria may affect sucrose-dependent colonization of S. mutans on the tooth surface and offer a possible explanation for both the difficulties involved in implanting this bacterium into the human mouth and the limited intraoral transmission of S. mutans from one tooth surface to another. Recent studies in these laboratories (39) demonstrated that a significant proportion of the human dental plaque bacterial flora is capable of producing dextran-degrading enzymes. These bacteria were shown to be a heterogeneous mixture of cell types with varying morphological and biochemical properties. One representative isolate which produces an extracellular endohydrolytic dextranase activity has been identified as Actinomyces israelii (39; R. Staat and C.