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Monitoring Perinatal Gut Microbiota in Mouse Models by Mass Spectrometry Approaches: Parental Genetic Background and Breastfeeding Effects

View Article: PubMed Central - PubMed

ABSTRACT

At birth, contact with external stimuli, such as nutrients derived from food, is necessary to modulate the symbiotic balance between commensal and pathogenic bacteria, protect against bacterial dysbiosis, and initiate the development of the mucosal immune response. Among a variety of different feeding patterns, breastfeeding represents the best modality. In fact, the capacity of breast milk to modulate the composition of infants’ gut microbiota leads to beneficial effects on their health. In this study, we used newborn mice as a model to evaluate the effect of parental genetic background (i.e., IgA-producing mice and IgA-deficient mice) and feeding modulation (i.e., maternal feeding and cross-feeding) on the onset and shaping of gut microbiota after birth. To investigate these topics, we used either a culturomic approach that employed Matrix Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry (MS), or bottom–up Liquid Chromatography, with subsequent MSMS shotgun metaproteomic analysis that compared and assembled results of the two techniques. We found that the microbial community was enriched by lactic acid bacteria when pups were breastfed by wild-type (WT) mothers, while IgA-deficient milk led to an increase in the opportunistic bacterial pathogen (OBP) population. Cross-feeding results suggested that IgA supplementation promoted the exclusion of some OBPs and the temporary appearance of beneficial species in pups fed by WT foster mothers. Our results show that both techniques yield a picture of microbiota from different angles and with varying depths. In particular, our metaproteomic pipeline was found to be a reliable tool in the description of microbiota. Data from these studies are available via ProteomeXchange, with identifier PXD004033.

No MeSH data available.


Metaproteomics analysis at the genus level by workflow B. (A) A histogram representing MGM relative abundance at the genus level, from Balb/c and Rag2ko baby mice biopsy contents at days 3, 7, and 14. (B) Pie charts comparing the MGM content in Rag2ko and Balb/c mice with that of their respective, cross-fed offspring at days 3 and 7. Phyla are identified by color shades, specifically blue for Firmicutes and orange for Proteobacteria.
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Figure 3: Metaproteomics analysis at the genus level by workflow B. (A) A histogram representing MGM relative abundance at the genus level, from Balb/c and Rag2ko baby mice biopsy contents at days 3, 7, and 14. (B) Pie charts comparing the MGM content in Rag2ko and Balb/c mice with that of their respective, cross-fed offspring at days 3 and 7. Phyla are identified by color shades, specifically blue for Firmicutes and orange for Proteobacteria.

Mentions: We obtained taxonomic charts, employing two different approaches (workflows A and B) for each taxonomic level (Supplementary Table S3). The two representations showed the same distribution of taxa at a generic taxonomic level (e.g., phylum, order), but when considering more specific levels in the lineage (e.g., family, genus, and species), Firmicutes were almost exclusively represented by the Lactobacillus genus, while there was a large number of different genera belonging to Proteobacteria, sharing a considerable portion of the genome (e.g., Escherichia/Shigella). In the present work, the outcomes of the two counting procedures showed a comparable trend in the evolution of the MGM in mice pups, and we thus chose to use the more restrictive one (workflow B), which was also more similar in concept to the LCA approach (Figure 3; Supplementary Figure S1).


Monitoring Perinatal Gut Microbiota in Mouse Models by Mass Spectrometry Approaches: Parental Genetic Background and Breastfeeding Effects
Metaproteomics analysis at the genus level by workflow B. (A) A histogram representing MGM relative abundance at the genus level, from Balb/c and Rag2ko baby mice biopsy contents at days 3, 7, and 14. (B) Pie charts comparing the MGM content in Rag2ko and Balb/c mice with that of their respective, cross-fed offspring at days 3 and 7. Phyla are identified by color shades, specifically blue for Firmicutes and orange for Proteobacteria.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5036385&req=5

Figure 3: Metaproteomics analysis at the genus level by workflow B. (A) A histogram representing MGM relative abundance at the genus level, from Balb/c and Rag2ko baby mice biopsy contents at days 3, 7, and 14. (B) Pie charts comparing the MGM content in Rag2ko and Balb/c mice with that of their respective, cross-fed offspring at days 3 and 7. Phyla are identified by color shades, specifically blue for Firmicutes and orange for Proteobacteria.
Mentions: We obtained taxonomic charts, employing two different approaches (workflows A and B) for each taxonomic level (Supplementary Table S3). The two representations showed the same distribution of taxa at a generic taxonomic level (e.g., phylum, order), but when considering more specific levels in the lineage (e.g., family, genus, and species), Firmicutes were almost exclusively represented by the Lactobacillus genus, while there was a large number of different genera belonging to Proteobacteria, sharing a considerable portion of the genome (e.g., Escherichia/Shigella). In the present work, the outcomes of the two counting procedures showed a comparable trend in the evolution of the MGM in mice pups, and we thus chose to use the more restrictive one (workflow B), which was also more similar in concept to the LCA approach (Figure 3; Supplementary Figure S1).

View Article: PubMed Central - PubMed

ABSTRACT

At birth, contact with external stimuli, such as nutrients derived from food, is necessary to modulate the symbiotic balance between commensal and pathogenic bacteria, protect against bacterial dysbiosis, and initiate the development of the mucosal immune response. Among a variety of different feeding patterns, breastfeeding represents the best modality. In fact, the capacity of breast milk to modulate the composition of infants’ gut microbiota leads to beneficial effects on their health. In this study, we used newborn mice as a model to evaluate the effect of parental genetic background (i.e., IgA-producing mice and IgA-deficient mice) and feeding modulation (i.e., maternal feeding and cross-feeding) on the onset and shaping of gut microbiota after birth. To investigate these topics, we used either a culturomic approach that employed Matrix Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry (MS), or bottom–up Liquid Chromatography, with subsequent MSMS shotgun metaproteomic analysis that compared and assembled results of the two techniques. We found that the microbial community was enriched by lactic acid bacteria when pups were breastfed by wild-type (WT) mothers, while IgA-deficient milk led to an increase in the opportunistic bacterial pathogen (OBP) population. Cross-feeding results suggested that IgA supplementation promoted the exclusion of some OBPs and the temporary appearance of beneficial species in pups fed by WT foster mothers. Our results show that both techniques yield a picture of microbiota from different angles and with varying depths. In particular, our metaproteomic pipeline was found to be a reliable tool in the description of microbiota. Data from these studies are available via ProteomeXchange, with identifier PXD004033.

No MeSH data available.