Molecular identification of potential Th1/Th2 responses-modulating bacterial genes using suppression subtractive DNA hybridization

Background and objectivesWe characterized the immunomodulating potential of a number of lactobacilli isolated from an African fermented food by co-incubation with peripheral blood mononuclear cells (PBMCs). Two strains with different immune modulating properties were genetically compared by suppression subtractive hybridization (SSH).MethodsFrom 48 Lactobacillus strains isolated from Kimere, African fermented pearl millet dough, 10 were selected based on their bile salt tolerance. Their effects on secretion by PBMCs of the T-helper cells Th1- and Th2-cytokines IFN-γ and IL-4, respectively, in the presence or absence of staphylococcal enterotoxin A (SEA) were assessed. To study the genetic basis of different immune-modulating properties, a subtracted cDNA library for L. fermentum strains K1-Lb1 (Th1 inducer) and K8-Lb1 (Th1 and Th2 suppressor) was constructed using SSH. Finally, adhesion of these strains to hydrocarbons (relative hydrophobicity) and to human HT-29 colonic epithelial cell line was assessed.ResultsTwo strains, K1-Lb1 and K4-Lb6, induced basal IFN-γ secretion. Four strains, K1-Lb6, K6-Lb2, K7-Lb1, and K8-Lb1 diminished INF-γ secretion by SEA-stimulated PBMCs. All strains, except K1-Lb1, K2-Lb4, and K9-Lb3, inhibited SEA-stimulated IL-4 secretion. Comparing the genomes of K1-Lb1 and K8-Lb1 by SSH indicated that K1-Lb1 is able to synthetize polysaccharides, for the synthesis of which K1-Lb8 appears to lack enzymes. A difference in the hydrophobicity properties of the surfaces of both strains indicated that this has impact on their surface.ConclusionThe K1-Lb1-specific sequences encoding putative glycosyltransferases and enzymes for polysaccharides synthesis may account for the observed differences in immunomodulation and surface properties between the two strains and for mediating potential probiotic effects.