Background: Infants born to HIV-infected mothers have an altered cellular immunity despite being HIV-infected themselves. It is known that the gut microbiome is crucial to immune development. Currently, 16S rRNA sequencing, the gold standard for characterising the microbiome, yields limited information on the function of the microbial community, predicted open reading frames might not be expressed under in vivo conditions.
Methods: Here, infant stool samples, both HIV-exposed and unexposed infants collected at birth and at 4-7days were analysed to characterise the metaproteome using mass spectrometry-based proteomics. Briefly, cold stored samples were snap-frozen and ground samples were suspended in organic buffer and precipitated proteinaceous material were subjected to in-solution trypsin digestion. Desalted tryptic peptides were analysed in triplicate on the nLC-MS/MS. An in-house developed pipeline, Metanovo, was used to construct a metaproteomic database using the Universal Reference Database and generated mass spectrometry data. The metaproteomic database was used for protein and organism identification in MaxQuant-suite.
Results: Using our unbiased approach, we identified 3943 protein groups for all 55 samples described. The microbial diversity changed dynamically from birth to 4-7 days after birth and the proportion of human proteins identified decreased as the microbial diversity evolved. Using this approach, we identified virus, parasite, bacteria and human proteins in a single sample. The number of proteins belonging to the Bifidobacteriaceae family were significantly different between HEU and HU infants at birth, 1 to 180 respectively. Furthermore, this bacterial family increased dramatically to 1800 proteins within the first week of birth. Members of the Bifidobacteriaceae family are important microbes that colonize the gut of humans and other animals during the early stages of life.
Conclusion: Here we demonstrated that using mass spectrometry-based proteomics approach we were able to construct a metaproteomic profile of the microbiota composition of infant stool samples.