Diarrhoea is prevalently widespread in developing countries and can lead to death. This disease could be caused by a prerequisite attachment of Enterotoxigenic Escherichia coli (ETEC) on human intestinal epithelial cells and followed by toxin production. Several antibiotics have been developed to reduce diarrhoeal severity. However, the use of antibiotics could also disrupt the balance of gut microbiota. This research found that tempeh fermented with the addition of Lactobacillus plantarum culture could decrease the adhesion of ETEC to the intestinal epithelial cells. This study measured the ability of L. plantarum to produce exopolysaccharide (EPS) and examined the effect of L. plantarum addition to tempeh, particularly its effect on anti-adhesion activity against ETEC. By using yeast agglutination assay to observe anti-adhesion activity, the data showed that soybean tempeh fermented with the addition of L. plantarum at OD600 of 0.5 had an increase of anti-adhesion bioactivity against ETEC nearly three times compared to tempeh without the addition of L. plantarum culture. Moreover, the exclusion test showed that EPS from L. plantarum could act as a preventive agent against ETEC adhesion to eukaryotic cells. Our finding indicated that the addition of lactic acid bacteria culture can be considered to augment the anti-adhesion bioactivity in soybean tempeh against ETEC.