Protective effect of LAB and yeast isolated from kefir against Shigella invasion

FERNANDEZ M; BOLLA P; KAKISU E.; DE URRAZA P.J; DE ANTONI G.L

Tucuman

Congreso; III Simposio Internacional de Bacterias Lácticas (BAL); 2009

CERELA

Shigella strains cause bacillary dysentery in humans by invading epithelial cells of the colon. This mechanism has been described as essential for Shigella pathogenesis. In vitro models have been used to study cell invasion. In our country the most frequent species identified are S. flexneri and S. sonnei. It is known the use of probiotics to prevent or treat a variety of infectious diseases caused by oral and enteric pathogens (Rolfe RD. 2000). Lactic acid bacteria and yeast are the most common microorganisms used as probiotics. The aim of this work is to study the ability of some bacteria and yeasts isolated from kefir to reduce the invasive capacity of Shigella isolates in vitro. In this work Lactobacillus plantarum (CIDCA83114), Lactobacillus kefir (CIDCA8348), Lactococcus lactis subsp. lactis (CIDCA8221), Kluyveromyces marxianus (CIDCA 8154) and Saccharomyces serevisiae (CIDCA 8112) were used. Some of these strains have been demonstrated in previous works to have a protective effect against other intestinal pathogens. In addition, two clinical isolates of Shigella were used; one of these was S. flexneri and the other S. sonnei. To assay the protective role of the potentially probiotic strains, Hep-2 cells were preincubated with kefir isolates, for one hour. After washing, an overnight culture of approximately 108 cfu/ml was inoculated onto confluent HEp-2 cell monolayers and allowed to invade the cells for 2.5 hs incubation period. The cell monolayers were washed, and DMEM medium containing 100 mg/ml of gentamicin was added for an additional 90 min incubation period to kill extracellular bacteria. The cell monolayers were washed and then lysed with water. The bacteria released from the epithelial cells were plate count on nutrient agar. In addition, Hep-2 cells were grown in presence of a glass cover slip and, after the invasion period, analyzed by scanning electron microscopy. We found the highest protective effect of S. flexneri and S. sonnei invasion when we used the Lactobacillus plantarum, with a diminution of two logs in the invasion of both Shigella isolates. A lower protective effect was found when we used the strains of Lactococcus lactis subsp. lactis and Saccharomyces serevisiae, but in these cases protection was found only with S. sonnei isolate. We could not find a significant protective effect when Hep-2 cells were preincubated with Lactobacillus kefir and Kluyveromyces marxianus. Scanning electron microscopy confirms the results. In conclusion we have two bacteria and a yeast potentially probiotic that reduce the invasive capacity of Shigella. The highest invasion inhibition was found with the Lactobacillus plantarum (CIDCA83114) and this bacteria is the only strain assayed that is able to reduce the invasion of the S. flexneri isolate.