In vivo assessment of antimicrobial activity of enterocin CRL35 and Enterococcus mundtii CRL35 in a murine model of pregnancy-associated Listeria monocytogenes infection

SAAVEDRA, L.; SALVUCCI, E.; HEBERT, E.M.; HARO, C.; VIGNOLO, G.; SESMA, F.

Conferencia; Gut Micro Ecology (GME) 2010 - The International Scientific Conference on Gastro-Intestinal Microbial Ecology.; 2010

Listeria monocytogenes has been recognized as a foodborne pathogen that can be acquired mainly by the consumption of refrigerated and ready-to-eat products. Pregnant individuals are among the most susceptible population for listeriosis where the consequences may result in miscarriage and fetal death.Enterococcus mundtii CRL35 produces enterocin CRL35, a subclass IIa bacteriocin active against Listeria. Previous studies demonstrated that combinations of enterocin CRL 35 with certain antibiotics strongly increase their activity leading to marked reduction of their doses. These results suggest a potential application of such combinations in the medicine or pharmaceutical fields. As a first step in this direction we set up a murine model of listeriosis to test the effectiveness of enterocin CRL35 and its producer strain.Methods: E.mundtii CRL35 was grownfor 16 h at 37ºC in LAPTg broth and enterocin CRL35 was purified (PE) from this culture supernatant by our standard lab procedure. A murine model of pregnancy associated Listeria infection was achieved. Five to six week-old female BALB/c mice were orally infected with L. monocytogenes FBUNT at day 14 and sacrificed at day 17 of pregnancy. Mice were randomly divided in five groups that received: a) L .monocytogenes FBUNT (5 x 109 CFU); b) L. monocytogenes FBUNT (5 x 109 CFU) plus a treatment of 65 μg of PE per animal every 12 h; c) E mundtii CRL35 (2 x 109 CFU) and 5 h later L. monocytogenes FBUNT (5 x 109 CFU); d) PBS; e) 65μg of PE every 12 h. Mice were anaesthetized and killed after blood collection by cardiac puncture. Liver, spleen and fetus were aseptically taken, homogenized and plated on Listeria selective agar (Oxoid) and Brain Heart Infusion (BHI). Plates were incubated at 37ºC for 48 h. Viable counts were expressed as log colony forming units (CFU) per gram of organ. Changes in weight, food and fluid intake as well as hematologic parameters were also analyzed. All animal protocols were preapproved by the CERELA Animal Protection Committee and all experiments complied with the current laws of ArgentinaResults: The administration of a single dose of L. monocytogenes FBUNT resulted in the translocation of the pathogen to liver and spleen although it was not detected in fetus. Bacteriological analysis demonstrated that the oral administration of enterocin CRL35 as well as the bacteriocinogenic strain markedly reduced the CFU count of Listeria cells in liver and spleen. However, preventive administration (3 days) of E. mundtii CRL35 resulted in unspecific translocation of other members of gut microbiota. None of the groups showed any remarkable changes of hematological parameters, food and fluid intake.Conclusion: These results evidenced that enterocin CRL35 might be a good candidate for pharmacological or medical applications. Although more studies need to be done in order to pursue this issue.