454 pyrosequencing analysis on faecal samples from a randomized DBPC trial of colicky infants treated with Lactobacillus reuteri DSM 17938

Stefan Roos, Johan Dicksved, Valentina Tarasco, Emanuela Locatelli, Fulvio Ricceri, Ulf Grandin, Francesco Savino, Stefan Roos, Johan Dicksved, Valentina Tarasco, Emanuela Locatelli, Fulvio Ricceri, Ulf Grandin, Francesco Savino

Abstract

Objective: To analyze the global microbial composition, using large-scale DNA sequencing of 16 S rRNA genes, in faecal samples from colicky infants given L. reuteri DSM 17938 or placebo.

Methods: Twenty-nine colicky infants (age 10-60 days) were enrolled and randomly assigned to receive either Lactobacillus reuteri (10(8) cfu) or a placebo once daily for 21 days. Responders were defined as subjects with a decrease of 50% in daily crying time at day 21 compared with the starting point. The microbiota of faecal samples from day 1 and 21 were analyzed using 454 pyrosequencing. The primers: Bakt_341F and Bakt_805R, complemented with 454 adapters and sample specific barcodes were used for PCR amplification of the 16 S rRNA genes. The structure of the data was explored by using permutational multivariate analysis of variance and effects of different variables were visualized with ordination analysis.

Results: The infants' faecal microbiota were composed of Proteobacteria, Firmicutes, Actinobacteria and Bacteroidetes as the four main phyla. The composition of the microbiota in infants with colic had very high inter-individual variability with Firmicutes/Bacteroidetes ratios varying from 4000 to 0.025. On an individual basis, the microbiota was, however, relatively stable over time. Treatment with L. reuteri DSM 17938 did not change the global composition of the microbiota, but when comparing responders with non-responders the group responders had an increased relative abundance of the phyla Bacteroidetes and genus Bacteroides at day 21 compared with day 0. Furthermore, the phyla composition of the infants at day 21 could be divided into three enterotype groups, dominated by Firmicutes, Bacteroidetes, and Actinobacteria, respectively.

Conclusion: L. reuteri DSM 17938 did not affect the global composition of the microbiota. However, the increase of Bacteroidetes in the responder infants indicated that a decrease in colicky symptoms was linked to changes of the microbiota.

Trial registration: ClinicalTrials.gov NCT00893711.

Conflict of interest statement

Competing Interests: The University of Turin and Swedish University of Agricultural Sciences both received a donation from the manufacturer of L. reuteri DSM 17938, BioGaia AB, Lund, Sweden. There are no further patents, products in development or marketed products to declare. This does not alter the authors' adherence to all the PLOS ONE policies on sharing data and materials.

References

    1. Wessel MA, Cobb JC, Jackson EB, Harris GS, Detwiler AC (1954) Paroxysmal fussing in infancy, sometimes called colic. Pediatrics 14: 421–435.
    1. Yalçin SS, Orün E, Mutlu B, Madendağ Y, Sinici I, et al. (2010) Why are they having infant colic? A nested case-control study. Paediatr Perinat Epidemiol 24: 584–596 doi:.
    1. Savino F (2007) Focus on infantile colic. Acta Paediatr 96: 1259–1264 doi:.
    1. Savino F, Tarasco V (2010) New treatments for infant colic. Curr Opin Pediatr 22: 791–797 doi:.
    1. Savino F, Cordisco L, Tarasco V, Calabrese R, Palumeri E, et al. (2009) Molecular identification of coliform bacteria from colicky breastfed infants. Acta Paediatr 98: 1582–1588 doi:.
    1. Rhoads JM, Fatheree NY, Norori J, Liu Y, Lucke JF, et al. (2009) Altered fecal microflora and increased fecal calprotectin in infants with colic. The Journal of Pediatrics 155: 823–828.e1 doi:.
    1. Savino F, Cresi F, Pautasso S, Palumeri E, Tullio V, et al. (2004) Intestinal microflora in breastfed colicky and non-colicky infants. Acta Paediatr 93: 825–829.
    1. Savino F, Cordisco L, Tarasco V, Locatelli E, Di Gioia D, et al. (2011) Antagonistic effect of Lactobacillus strains against gas-producing coliforms isolated from colicky infants. BMC Microbiol 11: 157 doi:.
    1. Pärtty A, Kalliomäki M, Endo A, Salminen S, Isolauri E (2012) Compositional development of Bifidobacterium and Lactobacillus microbiota is linked with crying and fussing in early infancy. PLoS ONE 7: e32495 doi:.
    1. Savino F, Cordisco L, Tarasco V, Palumeri E, Calabrese R, et al. (2010) Lactobacillus reuteri DSM 17938 in infantile colic: a randomized, double-blind, placebo-controlled trial. Pediatrics 126: e526–e533 doi:.
    1. Savino F, Pelle E, Palumeri E, Oggero R, Miniero R (2007) Lactobacillus reuteri (American Type Culture Collection Strain 55730) versus simethicone in the treatment of infantile colic: a prospective randomized study. Pediatrics 119: e124–e130 doi:.
    1. Szajewska H, Gyrczuk E, Horvath A (2012) Lactobacillus reuteri DSM 17938 for the Management of Infantile Colic in Breastfed Infants: A Randomized, Double-Blind, Placebo-Controlled Trial. The Journal of Pediatrics. doi:10.1016/j.jpeds.2012.08.004.
    1. Herlemann DP, Labrenz M, Jürgens K, Bertilsson S, Waniek JJ, et al. (2011) Transitions in bacterial communities along the 2000 km salinity gradient of the Baltic Sea. ISME J 5: 1571–1579 doi:.
    1. Arumugam M, Raes J, Pelletier E, Le Paslier D, Yamada T, et al. (2011) Enterotypes of the human gut microbiome. Nature 473: 174–180 doi:.
    1. De Filippo C, Cavalieri D, Di Paola M, Ramazzotti M, Poullet JB, et al. (2010) Impact of diet in shaping gut microbiota revealed by a comparative study in children from Europe and rural Africa. Proc Natl Acad Sci USA 107: 14691–14696 doi:.
    1. Andersson AF, Lindberg M, Jakobsson H, Bäckhed F, Nyrén P, et al. (2008) Comparative analysis of human gut microbiota by barcoded pyrosequencing. PLoS ONE 3: e2836 doi:.
    1. Cole JR, Wang Q, Cardenas E, Fish J, Chai B, et al. (2009) The Ribosomal Database Project: improved alignments and new tools for rRNA analysis. Nucleic Acids Res 37: D141–D145 doi:.
    1. Oksanen J, Blanchet GF, Kindt R, Legendre P, Minchin PR, et al.. (2012) vegan: Community Ecology Package. R package version 2.0–5. .
    1. Hammer O, Harper D, Ryan PD (2001) PAST: Paleontological Statistics Software Package for Education and Data Analysis. Palaeontologia Electronica 4: 1–9.
    1. Palmer C, Bik EM, DiGiulio DB, Relman DA, Brown PO (2007) Development of the human infant intestinal microbiota. PLoS Biol 5: e177 doi:.
    1. Adlerberth I, Wold AE (2009) Establishment of the gut microbiota in Western infants. Acta Paediatr 98: 229–238 doi:.
    1. Abrahamsson TRT, Jakobsson HEH, Andersson AFA, Björkstén BB, Engstrand LL, et al. (2012) Low diversity of the gut microbiota in infants with atopic eczema. J Allergy Clin Immunol 129: 434–432 doi:.
    1. Savino F, Castagno E, Bretto R, Brondello C, Palumeri E, et al. (2005) A prospective 10-year study on children who had severe infantile colic Vol. 94: 129–132 doi:.
    1. Koenig JE, Spor A, Scalfone N, Fricker AD, Stombaugh J, et al. (2011) Succession of microbial consortia in the developing infant gut microbiome. Proc Natl Acad Sci USA 108 Suppl 14578–4585 doi:.
    1. Shimizu K, Ogura H, Asahara T, Nomoto K, Morotomi M, et al. (2011) Gastrointestinal dysmotility is associated with altered gut flora and septic mortality in patients with severe systemic inflammatory response syndrome: a preliminary study. Neurogastroenterol Motil 23: 330–5–e157 doi:.
    1. Adlerberth I, Strachan DP, Matricardi PM, Ahrné S, Orfei L, et al. (2007) Gut microbiota and development of atopic eczema in 3 European birth cohorts. J Allergy Clin Immunol 120: 343–350 doi:.
    1. Fallani M, Young D, Scott J, Norin E, Amarri S, et al. (2010) Intestinal Microbiota of 6-week-old Infants Across Europe: Geographic Influence Beyond Delivery Mode, Breast-feeding, and Antibiotics. Journal of Pediatric Gastroenterology and Nutrition 51: 77–84 doi:.
    1. Jost T, Lacroix C, Braegger CP, Chassard C (2012) New insights in gut microbiota establishment in healthy breast fed neonates. PLoS ONE 7: e44595 doi:.
    1. Turroni F, Peano C, Pass DA, Foroni E, Severgnini M, et al. (2012) Diversity of bifidobacteria within the infant gut microbiota. PLoS ONE 7: e36957 doi:.
    1. Nicholson JK, Holmes E, Kinross J, Burcelin R, Gibson G, et al. (2012) Host-gut microbiota metabolic interactions. Science 336: 1262–1267 doi:.
    1. Preidis GA, Saulnier DM, Blutt SE, Mistretta T-A, Riehle KP, et al. (2012) Probiotics stimulate enterocyte migration and microbial diversity in the neonatal mouse intestine. FASEB J 26: 1960–1969 doi:.

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