Analysis of intestinal flora development in breast-fed and formula-fed infants by using molecular identification and detection methods

Background: An obvious difference between breast-fed and formula-fed newbor n infants is the development of the intestinal flora, considered to be of i mportance for protection against harmful micro-organisms and for the matura tion of the intestinal immune system. In this study, novel molecular identi fication methods were used to verify the data obtained by traditional cultu re methods and to validate the culture independent fluorescent in situ hybr idization (FISH) technique, Methods: From each of six breast-fed and six formula-fed newborn infants, s ix fecal samples were obtained during the first 20 days of life. The microb ial compositions of the samples were analyzed by culturing on specific medi a and by FISH, by using specific 16S rRNA-targeted oligonucleotide probes. The colonies growing on the media were identified by random amplified polym orphic DNA pattern analysis and by polymerase chain reaction amplification and subsequent analysis of the 16S rRNA gene. Results: Molecular identification of the colonies showed that the selective media are insufficiently selective and unsuitable for quantitative analyse s. Qualitative information from the culturing results combined with the dat a obtained by the FISH technique revealed initial colonization in all infan ts of a complex (adult-like) flora. After this initial colonization, a sele ction of bacterial strains began in all infants, in which Bifidobacterium s trains played an important role. In all breast-fed infants, bifidobacteria become dominant, whereas in most formula-fed infants similar amounts of Bac teroides and bifidobacteria (similar to 40%) were found. The minor componen ts of the fecal samples from breast-fed infants were mainly lactobacilli an d streptococci; samples from formula-fed infants often contained staphyloco cci, Escherichia coli, and clostridia. Conclusions: This study confirms the differences in development of intestin al flora between breast-fed and formula-fed infants. The results obtained f rom the FISH technique were consistent. Although the repertoire of probes f or this study was not yet complete, the FISH technique will probably become the method of reference for future studies designed to develop breast-fed- like intestinal flora in formula-fed infants. (C) 2000 Lippincott Williams & Wilkins, Inc.