Lactobacillus characterization and effects on oral biofilm composition

............................................................................................................................... v SAMMANFATTNING PÅ SVENSKA.................................................................................. vii LIST OF PUBLICATIONS..................................................................................................... ix ABBREVIATIONS................................................................................................................... x INTRODUCTION.................................................................................................................... 1 The human gastro-intestinal (GI) microbiota........................................................................ 1 Development of the GI microbiota......................................................................................... 2 Factors influencing infant colonization.................................................................................. 4 The genus Lactobacillus.......................................................................................................... 6 Establishment and colonization .................................................................................. 7 Lactobacillus taxonomy................................................................................................ 8 Lactobacillus identification techniques........................................................................ 9 Lactobacillus as a probiotic.......................................................................................... 10 Saliva and biofilm formation................................................................................................... 13 Human milk and milk fat globule membrane ....................................................................... 14 AIMS.......................................................................................................................................... 16 MATERIALS AND METHODS............................................................................................. 17 Study populations and study designs...................................................................................... 17 Bacteria and fungus strains..................................................................................................... 19 Saliva, mucosa and dental biofilm sampling ......................................................................... 19 Epithelial cells......................................................................................................................... 19 Test products........................................................................................................................... 19 Lactobacilli identification........................................................................................................ 20 Culture-dependent methods......................................................................................... 20 Culture-independent methods...................................................................................... 21 Evaluation of probiotic traits ................................................................................................. 22 Growth inhibition ......................................................................................................... 22 Adhesion ....................................................................................................................... 22 Adhesion to human epithelial cells............................................................................... 23 Mutans streptococci (MS) levels in saliva............................................................................... 24 Oral microbiota diversity......................................................................................................... 24 Human microbe identification microarray................................................................... 24 454-Pyrosequencing ..................................................................................................... 26 Statistical analyses................................................................................................................... 28 RESULTS.................................................................................................................................. 29 Studies on the oral microbiota in infants (I, II) ..................................................................... 29 Cohort characteristics ................................................................................................... 29 Lactobacilli in the oral cavity by feeding mode............................................................. 29 Lactobacillus species in breastfed infants.................................................................... 30 Potential probiotic properties of L.gasseri .................................................................. 30 Clustering of infants by feeding mode associated microbiota...................................... 31 Microbiota in breastfed versus formula-fed infants..................................................... 32 Studies on L. reuteri effects in adults (III, IV)....................................................................... 33 L.reuteri treatment and viable lactobacilli................................................................... 33 Persistence of L. reuteri DSM 17938 and PTA 5289.................................................... 33 Delayed regrowth of MS when L. reuteri persists........................................................ 33 454-Pyrosequencing determined microbiota............................................................... 34 Individuality in oral microbiota composition............................................................... 35 Effects of L. reuteri exposure on oral microbiota composition................................... 35 DISCUSSION........................................................................................................................... 36 CONCLUSIONS AND FUTURE PERSPECTIVES........................................................... 44 ACKNOWLEDGMENTS........................................................................................................ 46 REFERENCES.......................................................................................................................... 49