Resumo

Introduction: L. monocytogenes (LM) is Gram-positive bacterium able to tolerate many adverse conditions found in foods and can cause severe foodborne infections in immunocompromised people and in pregnant women. Objective: To evaluate LM isolates for the ability to adhere and invade Caco-2 cells under adverse conditions. Methods: A total of 10 isolates of LM from different sources were grown in Brain Heart Infusion Broth (BHI) at 37°C/24h and at 5°C/72h and also under the following conditions: a) BHI plus 2% sucrose; b) BHI plus 5% NaCl; c) BHI plus 0.3% Oxgall; d) BHI pH 4.5 (adjusted with HCl) and e) BHI plus 50% (v/v) of extract from bacteriocin-producing cultures of Lactobacillus sakei 1, Leuconostoc mesenteroides 11A or Enterococcus faecium 130 incubated at 37°C/24h. Aliquots of 100µl were transferred to tissue culture plate wells with confluent Caco-2 cells (10⁵cells/well) loaded with 900 µl incomplete RPMI medium. For adhesion assay, cultures were incubated under 5% CO2 at 37°C/30 min, trypsinized and plated on Tryptic Soy agar supplemented with 6g/l of yeast extract (TSAye) for LM enumeration. For invasion assay, cultures were incubated under 5% CO2 at 37°C/3 hours, treated for 1.5h with 250 µl (250 µg/ml) gentamicin, lysed with 0.1% (vol/vol) Triton X-100 solution and surface plated on TSAye for enumeration of LM. Results and discussion: LM isolates were able to adhere and invade Caco-2 cells under adverse conditions. Isolates grown in BHI plus Oxgall showed lower ability to adhere and invade Caco-2 cells. Cultivation in BHI pH 4.5 (adjusted with HCl) and BHI incubated at 5°C/72h demonstrated significant influence in LM invasiveness. The isolate LM H-2 demonstrated the highest invasion rates in all conditions studied and no correlation was observed with adhesion. Conclusion: This study showed that conditions found in food and the host are important to modulate the adhesion and invasion processes of the LM in eukaryotic cells.