Fecal microbiota in premature infants prior to necrotizing enterocolitis

Abstract

Intestinal luminal microbiota likely contribute to the etiology of necrotizing enterocolitis (NEC), a common disease in preterm infants. Microbiota development, a cascade of initial colonization events leading to the establishment of a diverse commensal microbiota, can now be studied in preterm infants using powerful molecular tools. Starting with the first stool and continuing until discharge, weekly stool specimens were collected prospectively from infants with gestational ages ≤32 completed weeks or birth weights≤1250 g. High throughput 16S rRNA sequencing was used to compare the diversity of microbiota and the prevalence of specific bacterial signatures in nine NEC infants and in nine matched controls. After removal of short and low quality reads we retained a total of 110,021 sequences. Microbiota composition differed in the matched samples collected 1 week but not <72 hours prior to NEC diagnosis. We detected a bloom (34% increase) of Proteobacteria and a decrease (32%) in Firmicutes in NEC cases between the 1 week and <72 hour samples. No significant change was identified in the controls. At both time points, molecular signatures were identified that were increased in NEC cases. One of the bacterial signatures detected more frequently in NEC cases (p<0.01) matched closest to γ-Proteobacteria. Although this sequence grouped to the well-studied Enterobacteriaceae family, it did not match any sequence in Genbank by more than 97%. Our observations suggest that abnormal patterns of microbiota and potentially a novel pathogen contribute to the etiology of NEC.

Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1. Chao rarefaction diversity.

Chao diversity was calculated from sequence distribution A) one week before and B) within 72 hours of NEC diagnosis.

Figure 2. Unifrac diversity measures.

Principal component analysis (PCA) of overall diversity based on UniFrac (unweighted) metric A) one week before and B) within 72 hours of NEC diagnosis. Squares represent controls and circles represent cases. P1 is component 1 and P2 component 2.

Figure 3. Changes in proportion of bacterial phyla.

Proportions of the four major phyla in nine controls and nine cases one week before and within 72 hours of NEC diagnosis.

Figure 4. Differences in OTU abundance.

Heat map of selected OTUs at 98% similarity by subject A) one week before and B) within 72 hours of NEC diagnosis. Colors indicate the ratio for a particular OTU in a sample to the average ratio of this OTU in all samples. Dark red: >10, light red: >5 and 1.5 and Cronobacter muytjensii; EC: Escherichia coli; ES: Enterobacter sp. glo8; KP: Klebsiella pneumoniae; SE: Staphylococcus epidermidis.