Abstract
Objective: The aim of this study was to develop a practical and efficient method for isolation and characterization of the enterobacterial microbiota of infant fecal samples. It could be used for reliable comparisons between individual microflora in different neonates, and in the follow-up of the succession in the floral developments in various nutritional conditions. Methods: A combination of traditional plate culture and different enrichment procedures carried out in a portable microbe enrichment unit (PMEU) was studied. The strains were isolated by plate culture both before and after enrichment. The isolated strains were identified by biochemical tests and the phenotypes of the isolates were compared by the PhenePlate™ technique. To obtain a picture of the formation of intestinal flora during the first year two follow-up samples were collected at the ages of 3 and 12 months from eight Finnish infants (one girl and seven boys). Results: Altogether 21 and 46 enterobacterial isolates were detected in samples taken at the ages of 3 and 12 months, respectively; 7/21 (33%) and 24/46 (52%) isolates were not detected by direct plating but only after enrichment. This material was supposed to demonstrate the differences between the different methodological approaches. Conclusion: There were age-dependent changes in the species proportions of different subpopulations of Enterobacteriaceae detected from the infantile fecal microbiota at different times. The statistically significant difference in the number of isolations with and without enrichment step suggests that the use of enrichment is highly recommendable and the PMEU equipment and method have now proven suitable for that purpose, at least for screening and monitoring the infant fecal flora. The method offers more representative, quicker, and relatively practical tools for obtaining an overall picture of the enterobacterial microbiota of the fecal samples and its variations, thus also giving an idea of the microbial communities in the gastrointestinal tract.
Acknowledgments
We express our gratitude to the Päivikki and Sakari Sohlberg Foundation, National Graduate School of Clinical Investigation and Foundation for Paediatric Research for financial support of the study. We are grateful to Professor Atte von Wright and Professor Raimo Voutilainen from the University of Kuopio for useful discussions and advice. We are also grateful to our laboratory assistant Tanja Auraniemi for her skillful work.
Declaration of interest: The study has been supported by National Graduate School of Clinical Investigation and Foundation for Paediatric Research from Finland by grants to J.P. and by Päivikki and Sakari Sohlberg Foundation by grants to O.V. - E.H. is the inventor of the PMEU and the founder of Finnoflag Oy. He has been involved in multiple scientific projects applying PMEU e.g. at medical, environmental and industrial fields. The authors alone are responsible for the content and writing of the paper.