Bifidobacterium population analysis in the infant gut by direct mapping of genomic hybridization patterns: potential for monitoring temporal development and effects of dietary regimens

Abstract

A bifidobacterial mixed-species microarray platform was used in a proof-of-principle study to address the composition and development of bifidobacteria in DNA extracted from faecal samples. These were collected in a time-course of 2 years since birth and derived from human infants that were breastfed, standard formula-fed or received a prebiotic formula during their weaning period. A set of over 50 samples was analysed, testifying for the throughput of the designed platform for multiple genome hybridizations. The generated data revealed that faecal samples of breastfed infants contained a high abundance of genomic DNA homologous to Bifidobacterium breve. In contrast, faecal samples from standard formula-fed infants lacked detectable amounts of this B. breve DNA but contained genes with high similarity to B. longum. Remarkably, infants that received breastmilk and later a prebiotic formula consisting of a standard formula milk containing a mixture of specific galacto- and fructo-oligosaccharides, continued to harbour a B. breve-dominant faecal population. One infant that received standard formula in combination with the additional B. lactis Bb12 culture, contained significant amounts of faecal DNA belonging to Bb12 but only during the period of ingestion. The microarray platform showed sufficient sensitivity to analyse the B. breve group at the strain level. Overall, the B. breve populations observed in the faecal samples of the studied infants showed a stable composition over time and were unique per infant. In conclusion, our results show the applicability of comparative genome hybridization to study bifidobacterial populations in infant faecal samples without the use of any amplification step.

Figure 1

Schematic sample information overview. A selection of faecal samples collected during the study described by Bakker‐Zierikzee and colleagues (2005).

Figure 2

Heat map visualization of the microarray‐ and qPCR‐based screening of the Bifidobacterium composition of faecal samples of six infants who received three different diets (Fig. 1). Time‐course samples are indicated by the age of the infant (D = day, W = week, M = month). The microarray data are presented as the percentage of marker‐clones, per species or species group (Table 1), that give a significant hybridization. The percentages are visualized according to the colour‐bar above the heat map. The qPCR results are presented as DNA concentrations (nanograms of DNA per sample) of the different species and visualized according to the colour‐bar above the heat map. The asterisk ‘*’ indicates data not available.

Figure 3

Hierarchical clustering (Pearson correlation) of the hybridization signals of genomic DNA fragments within the B. breve marker set for all faecal samples in time‐course order. Signal intensities are indicated by a black‐white colour gradient. On the X‐axis the sample codes are presented (D = day; W = week; M = month). Samples from one infant are highlighted by the bars above the codes as indicated in Fig. 1.

Figure 4

Hybridization signal intensities and ‘best blastx hits’ (> 98% homology) of a subset of 11 B. breve markers that have been sequenced are visualized for each marker fragment. The sample order is similar to the order in Fig. 3. Marker fragments 1 and 2 can be described as B. breve core genome whereas marker fragments 3–11 show to be strain‐specific.