Efecto de bacteriocinas sobre cepas sensibles y resistentes de Listeria monocytogenes

IBARGUREN, CAROLINA; RAYA. RAÚL; APELLA, M.CRISTINA; AUDISIO, M. CARINA

Tucumán, Argentina

Simposio; II Simposio Internacional de Bacterias Lácticas y Primer Encuentro red BAL; 2006

Centro de Referencias para Lactobacilos

Listeria monocytogenes is a potential pathogen for older people, children, pregnant women, immune deficient persons and AIDS patients. The most common or commercially accepted bacteriocin used to inhibit Listeria in food products is nisin. However, some researchers have already informed of L. monocytogenes  strains “spontaneously” resistant to this molecule and to some bacteriocins of class IIa. Enterococcus faecium Mori1, isolated from a honey sample, synthesizes bacteriocin/s with strong anti-Listeria effect. In this work, the genetic characterization of these compounds and their action on L. monocytogenes sensitive and resistant strains are presented. The structural genes of the bacteriocin/s were amplified by the specific PCR technique in samples of DNA isolated from the strain selected using the following specific primers: EntA, EntB, EntP, Ent31 y EntL50. Two different cultures of L. monocytogenes were analyzed: strain 99/287s (sensitive to bacteriocins assayed) and strain 99/287 MoriR, a clone spontaneously resistant to the compounds mentioned before. This clone originated as a satellite colony in the inhibition halo when analizing bacteriocin production by the well-diffusion assay. The antagonistic effect of bacteriocin on different L. monocytogenes strains was studied by means of the microplaque technique where a cell-free supernatant (CFS) from the producer strain was put in direct contact with a 14 h culture of different indicator strains, and of mixed cultures of the producer and different indicator strains. Viable cells number of Listeria was determined on BHI agar plaques in the first case and selective agar for Listeria in the second case. The plaques were incubated at 37ºC for 24 h. The bacteriocins studied are highly termostable (121ºC for 15 min), active without previous purification and they may be synthesized even at low temperatures (4ºC). The products of the amplification of bacteriocin structural genes evidenced that Ent. faecium synthesizes enterocins A and B. Viability of the Listeria resistant strain was decreased due to the enterocins but to a lesser extent than that of the sensitive strain. After 24 h of contact between a Listeria cell suspension (8.95 log CFU mL-1) and Ent. faecium CFS, a drop of 3 units in the log number of viable cells per mL was observed. However, the number of Listeria viable cells (6.05 log UFC mL-1) diminished respectively to 3.60 and 5.43 log UFC mL-1 for the sensible and resistant strains after 24 h in contact with a culture of the producer strain. This result suggests a different action mode of the enterocins, bactericidal for the sensible strain and bacteriostatic for the resistant clone.