Ana M. Suárez

Research note: unsuitability of the MRS medium for the screening of hydrogen peroxide-producing lactic acid bacteria.

The effect of MRS broth on the stability of hydrogen peroxide (H2O2) has been studied. Known concentrations (1–100 μg ml−1) of H2O2 were prepared in distilled water, phosphate buffer (pH 7·0) and MRS broth (pH 6·2 and 3·9). H2O2 was very stable in aqueous and buffer solutions but it was rapidly degraded in MRS broth (pH 3·9). The presence of H2O2 in MRS broth (pH 6·2) could not be detected.

Generation of polyclonal antibodies against a chemically synthesized N‐terminal fragment of the bacteriocin pediocin PA‐1

Six mice were immunized intraperitoneally (i.p.) with a chemically synthesized 9‐mer fragment (PH1) designed from the N‐terminal part of the bacteriocin pediocin PA‐1 and conjugated to keyhole limpet haemocyanin (KLH). After three doses of the immunogen had been administered, serum‐specific antibodies were detected by a competitive direct ELISA. Myeloma cells were injected i.p. into mice in order to obtain ascites polyclonal antibodies. Although four mice developed ascites, only mouse 2 had detectable specific antibodies in the ascites fluid. The serum and ascites antibodies were specific for PH1 but they did not recognize the whole pediocin PA‐1 molecule. This is the first attempt to generate antibodies against bacteriocins with a chemically synthesized oligopeptide as immunogen. This approach still remains attractive for detection, quantification, mode of action studies and purification of bacteriocins, especially those for which the purification process is difficult or inefficient at present.

Use of Genetic and Immunological Probes for Pediocin PA-1 Gene Detection and Quantification of Bacteriocin Production in Pediococcus acidilactici Strains of Meat Origin

PCR amplification analysis with pediocin PA-1 (pedA1) specific primers, a gene probe covering the pedA1 structural gene, and an immunological probe based on anti-peptide antibodies with predetermined specificity for pedA1, have been evaluated for pedA1 structural gene detection and quantification of bacteriocin production in Pediococcus acidilactici strains of meat origin. PCR products were obtained from colonies and purified plasmid DNA. The gene probe was used in dot-blot and colony-blot hybridization analysis. The production of pedA1 was evaluated by a non-competitive indirect ELISA (NCI-ELISA), a competive indirect ELISA (CI-ELISA), a competitive direct ELISA (CD-ELISA), and by immunoblotting. The approaches taken in the generation and performance of the reported probes could be useful for the rapid detection and identification of other bacteriocin-producing lactic acid bacteria (LAB) from many substrates and reservoirs and for quantification of bacteriocin production in food and environmental isolates and heterologous hosts.