Extreme prematurity for humans is associated with major acute and chronic morbidity and mortality including sepsis, meningitis, necrotizing enterocolitis and growth failure.  The origins of many of these processes are presumed to arise from the gut-associated microbiota.  Molecular determination of the metagenome in these infants will serve as a platform on which to understand how medical practices change the primary succession of the microbiota to promote health and disease.

We sought to determine the metagenome of the extremely low birth weight infant (ELBW; <1000 g).  We hypothesized that ELBW infants have a phylogenetically-restricted gut microbiota dominated by the Firmicutes. 

Stool samples were collected from 2 infants between 3-4 weeks of life (gestational age 25 5/7 wks/birth weight 980g; 25 1/7 wks/birth weight 560g).  The infants received 7 and 10 days of antibiotics, respectively. Both received human breast milk.  The low abundance of extractable DNA limited direct analysis; DNA was subjected to whole genome amplification (WGA) followed by 454 (Roche) shotgun sequencing.  The sequences were analyzed by BLAST protocols and subsystems analysis against the SEED database using the metagenomic server, MG-RAST.

Results/Conclusions

From the samples, 9.2E5 and 1.1E6 sequences were obtained (average reads, 370 nt and 394 nt, respectively).  Of the unique sequences, 56.6% and 58.2% aligned with the SEED database (maximum E scores 1E-5).  In each sample, the Firmicutes predominated, representing 99.4% and 99.2% of the total matched sequences.  At the Phylum level, 0.48%/0.45% and 0.69%/0.07% of the sequences matched within the Proteobacteria and Bacteroidetes, respectively.  For one infant, Staphylococcaceae was the dominant family (92.9% of all bacterial sequences).  Species level analysis suggested the majority arose from the ELBW pathogen S. epidermidis.  In the same infant sample, 0.86% of the sequences originated from the commensal-pathogen family Enterococcaceae.   In contrast, Enterococcaceae dominated the other infant’s microbiota (81.9%) while Staphylococceae sequences were 12.9% of the total.   The metagenomes of each infant had evidence of basal diversity, including fungal, human DNA viral, and bacteriophage sequences among a spectrum of other bacterial genomic and plasmid sequences. 

Preliminary analysis of the metagenome of the ELBW infant demonstrates a mono-predominant microbiota at the Family level.  However, low level diversity is evident by the fourth week of life despite extreme prematurity and medical management including caesarian section delivery, prolonged antibiotic administration, delayed feeding, and environmental isolation.  Future analysis of inter-individual and temporal variation will provide yet further details of the primary succession in the ELBW infant.