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Composition of intestinal microbiota in immune-deficient mice kept in three different housing conditions.

Thoene-Reineke C, Fischer A, Friese C, Briesemeister D, Göbel UB, Kammertoens T, Bereswill S, Heimesaat MM - PLoS ONE (2014)

Bottom Line: Abundance of commensals constituting the intestinal microbiota (IM) affects the immune system and predisposes to a variety of diseases, including intestinal infections, cancer, inflammatory and metabolic disorders.Overall, differences in IM composition were subtle.Housing-dependent and immune-deficiency mediated changes in intestinal microbiota composition were rather subtle but may nevertheless impact immunopathology in experimental models.

View Article: PubMed Central - PubMed

Affiliation: Forschungseinrichtung für Experimentelle Medizin, Charité - University Medicine Berlin, Berlin, Germany.

ABSTRACT

Background: Abundance of commensals constituting the intestinal microbiota (IM) affects the immune system and predisposes to a variety of diseases, including intestinal infections, cancer, inflammatory and metabolic disorders. Housing conditions determine the IM and can hence influence the immune system. We analyzed how both variables affect the IM of four immune-compromized mouse lines kept under different housing conditions.

Methodology/principal findings: We investigated the IM composition in mice by quantitative 16S rRNA RT-PCR analysis of the main fecal bacterial groups (Enterobacteriaceae, enterococci, lactobacilli, bifidobacteria, Bacteroides/Prevotella (BP) spp., Clostridium leptum and coccoides groups). Mice were homozygous (HO) or heterozygous (HE) for a targeted inactivating mutation of either the IFN-γ Receptor (R), IFN-γ, Rag1 or IL-4 genes. Overall, differences in IM composition were subtle. However, in the SPF-barrier, total eubacterial loads were higher in Rag1 HE versus Rag1 HO mice as well as in IFN-γR HE versus IFN-γR HO and WT animals. Although absent in WT mice, bifidobacterial loads were higher in HO and HE IFN-γ and Rag1 as well as IL-4 HO mice. Furthermore, BP was slightly lower in HO and HE IFN-γR and IFN-γ mice as well as in IL-4 HO mice as compared to WT controls. Interestingly, IM compositions were comparable in WT mice when kept in individual ventilated cages (IVC) or open cages (OC). IFN-γ HO and HE mice, however, had higher enterobacteria and BP loads, but lacked bifidobacteria when kept in OC versus IVC, as was the case in HO and HE Rag1 mice. In addition, Rag1 HO mice harbored higher clostridial loads when housed in OC as compared to IVC. Unexpectedly, lactobacilli levels were higher in IFN-γR mice when kept in OC versus IVC.

Conclusion/significance: Housing-dependent and immune-deficiency mediated changes in intestinal microbiota composition were rather subtle but may nevertheless impact immunopathology in experimental models.

No MeSH data available.


Related in: MedlinePlus

Commensal intestinal microbiota composition (total eubacterial load, Enterobacteriaceae and enterococci) of mice kept under SPF conditions.After twelve weeks of housing under SPF conditions, 16S rRNA of main intestinal bacterial groups was quantified by qRT-PCR in fecal samples derived from homozygous (HO; filled symbols) and heterozygous (HE; open symbols) mice deficient for IFN-γ Receptor (IFN-γ R; circles), IFN-γ (diamonds), Rag1 (triangle), or IL-4 (hexagons). Wildtype C57BL/6 (WT, black squares) served as controls. (A) Individual total eubacterial loads, and quantitative abundance of (B) Enterobacteriaceae and (C) enterococci are expressed in gene numbers per ng DNA. Medians (black bars), levels of significance (P-values) determined by the Mann-Whitney-U test and numbers of analyzed animals (in parentheses) are indicated.
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pone-0113406-g001: Commensal intestinal microbiota composition (total eubacterial load, Enterobacteriaceae and enterococci) of mice kept under SPF conditions.After twelve weeks of housing under SPF conditions, 16S rRNA of main intestinal bacterial groups was quantified by qRT-PCR in fecal samples derived from homozygous (HO; filled symbols) and heterozygous (HE; open symbols) mice deficient for IFN-γ Receptor (IFN-γ R; circles), IFN-γ (diamonds), Rag1 (triangle), or IL-4 (hexagons). Wildtype C57BL/6 (WT, black squares) served as controls. (A) Individual total eubacterial loads, and quantitative abundance of (B) Enterobacteriaceae and (C) enterococci are expressed in gene numbers per ng DNA. Medians (black bars), levels of significance (P-values) determined by the Mann-Whitney-U test and numbers of analyzed animals (in parentheses) are indicated.

Mentions: First, we investigated the commensal intestinal microbiota composition in homozygous and heterozygous mice of different genotypes kept in open cages in the breeding facility (a barrier access restricted to animal care personnel). The hygiene status of this facility is specific pathogen free according to FELASA standards 2002 [16]. Fecal samples were derived from 12-weeks-old wildtype (WT) mice, IFN-γR+/− (HE) and IFN-γR−/− (HO) mice, IFN-γ+/− (HE) and IFN-γ−/− (HO) mice, Rag1+/− (HE) and Rag1−/− (HO) mice, as well as IL-4+/− (HE) and IL-4−/− (HO) mice and subjected to 16S rRNA analysis of the most prevalent commensal intestinal bacterial groups by quantitative RT-PCR. The total eubacterial loads were slightly higher (mean difference approximately one order of magnitude) in IFN-γ+/− (HE) and Rag-1+/− (HE) mice as compared to the respective HO mice as well as in IFN-γ+/− (HE) versus WT control animals (p<0.01, p<0.005, and p<0.005, respectively; Fig. 1A). However, eubacterial 16S rRNA levels did not differ between mice of the remaining genotypes. Analysis of specific bacterial groups revealed that all mice harbored comparable 16S rRNA gene levels of Enterobacteriaceae, enterococci, and lactobacilli (Fig. 1B, C; Fig. 2A). Whereas bifidobacteria were virtually absent in WT and IFN-γR−/− (HO) mice, bifidobacterial 16S rRNA levels were significantly higher in IFN-γ+/− (HE), Rag1−/− (HO), Rag1+/− (HE), and IL-4−/− (HO) mice as compared to WT controls (mean differences up to 3.5 orders of magnitude; p<0.005–0.0005; Fig. 2B). Interestingly, bifidobacterial loads were higher in IFN-γ+/− (HE) mice as compared to IFN-γ−/− (HO) mice (p<0.05), but lower in IL-4+/− (HE) versus IL-4−/− (HO) animals (p<0.01) (Fig. 2B). Moreover, WT control mice displayed approximately one log higher fecal Bacteroides/Prevotella spp. loads as compared to IFN-γR−/− (HO), IFN-γR+/− (HE), IFN-γ−/− (HO), IFN-γ+/− (HE), and IL-4−/− (HO) mice (p<0.05–0.005; Fig. 3A), whereas IL-4+/− (HE) mice harbored one order of magnitude higher Bacteroides/Prevotella spp. 16S rRNA in the feces as compared to IL-4−/− (HO) mice (p<0.05; Fig. 3A).


Composition of intestinal microbiota in immune-deficient mice kept in three different housing conditions.

Thoene-Reineke C, Fischer A, Friese C, Briesemeister D, Göbel UB, Kammertoens T, Bereswill S, Heimesaat MM - PLoS ONE (2014)

Commensal intestinal microbiota composition (total eubacterial load, Enterobacteriaceae and enterococci) of mice kept under SPF conditions.After twelve weeks of housing under SPF conditions, 16S rRNA of main intestinal bacterial groups was quantified by qRT-PCR in fecal samples derived from homozygous (HO; filled symbols) and heterozygous (HE; open symbols) mice deficient for IFN-γ Receptor (IFN-γ R; circles), IFN-γ (diamonds), Rag1 (triangle), or IL-4 (hexagons). Wildtype C57BL/6 (WT, black squares) served as controls. (A) Individual total eubacterial loads, and quantitative abundance of (B) Enterobacteriaceae and (C) enterococci are expressed in gene numbers per ng DNA. Medians (black bars), levels of significance (P-values) determined by the Mann-Whitney-U test and numbers of analyzed animals (in parentheses) are indicated.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4234647&req=5

pone-0113406-g001: Commensal intestinal microbiota composition (total eubacterial load, Enterobacteriaceae and enterococci) of mice kept under SPF conditions.After twelve weeks of housing under SPF conditions, 16S rRNA of main intestinal bacterial groups was quantified by qRT-PCR in fecal samples derived from homozygous (HO; filled symbols) and heterozygous (HE; open symbols) mice deficient for IFN-γ Receptor (IFN-γ R; circles), IFN-γ (diamonds), Rag1 (triangle), or IL-4 (hexagons). Wildtype C57BL/6 (WT, black squares) served as controls. (A) Individual total eubacterial loads, and quantitative abundance of (B) Enterobacteriaceae and (C) enterococci are expressed in gene numbers per ng DNA. Medians (black bars), levels of significance (P-values) determined by the Mann-Whitney-U test and numbers of analyzed animals (in parentheses) are indicated.
Mentions: First, we investigated the commensal intestinal microbiota composition in homozygous and heterozygous mice of different genotypes kept in open cages in the breeding facility (a barrier access restricted to animal care personnel). The hygiene status of this facility is specific pathogen free according to FELASA standards 2002 [16]. Fecal samples were derived from 12-weeks-old wildtype (WT) mice, IFN-γR+/− (HE) and IFN-γR−/− (HO) mice, IFN-γ+/− (HE) and IFN-γ−/− (HO) mice, Rag1+/− (HE) and Rag1−/− (HO) mice, as well as IL-4+/− (HE) and IL-4−/− (HO) mice and subjected to 16S rRNA analysis of the most prevalent commensal intestinal bacterial groups by quantitative RT-PCR. The total eubacterial loads were slightly higher (mean difference approximately one order of magnitude) in IFN-γ+/− (HE) and Rag-1+/− (HE) mice as compared to the respective HO mice as well as in IFN-γ+/− (HE) versus WT control animals (p<0.01, p<0.005, and p<0.005, respectively; Fig. 1A). However, eubacterial 16S rRNA levels did not differ between mice of the remaining genotypes. Analysis of specific bacterial groups revealed that all mice harbored comparable 16S rRNA gene levels of Enterobacteriaceae, enterococci, and lactobacilli (Fig. 1B, C; Fig. 2A). Whereas bifidobacteria were virtually absent in WT and IFN-γR−/− (HO) mice, bifidobacterial 16S rRNA levels were significantly higher in IFN-γ+/− (HE), Rag1−/− (HO), Rag1+/− (HE), and IL-4−/− (HO) mice as compared to WT controls (mean differences up to 3.5 orders of magnitude; p<0.005–0.0005; Fig. 2B). Interestingly, bifidobacterial loads were higher in IFN-γ+/− (HE) mice as compared to IFN-γ−/− (HO) mice (p<0.05), but lower in IL-4+/− (HE) versus IL-4−/− (HO) animals (p<0.01) (Fig. 2B). Moreover, WT control mice displayed approximately one log higher fecal Bacteroides/Prevotella spp. loads as compared to IFN-γR−/− (HO), IFN-γR+/− (HE), IFN-γ−/− (HO), IFN-γ+/− (HE), and IL-4−/− (HO) mice (p<0.05–0.005; Fig. 3A), whereas IL-4+/− (HE) mice harbored one order of magnitude higher Bacteroides/Prevotella spp. 16S rRNA in the feces as compared to IL-4−/− (HO) mice (p<0.05; Fig. 3A).

Bottom Line: Abundance of commensals constituting the intestinal microbiota (IM) affects the immune system and predisposes to a variety of diseases, including intestinal infections, cancer, inflammatory and metabolic disorders.Overall, differences in IM composition were subtle.Housing-dependent and immune-deficiency mediated changes in intestinal microbiota composition were rather subtle but may nevertheless impact immunopathology in experimental models.

View Article: PubMed Central - PubMed

Affiliation: Forschungseinrichtung für Experimentelle Medizin, Charité - University Medicine Berlin, Berlin, Germany.

ABSTRACT

Background: Abundance of commensals constituting the intestinal microbiota (IM) affects the immune system and predisposes to a variety of diseases, including intestinal infections, cancer, inflammatory and metabolic disorders. Housing conditions determine the IM and can hence influence the immune system. We analyzed how both variables affect the IM of four immune-compromized mouse lines kept under different housing conditions.

Methodology/principal findings: We investigated the IM composition in mice by quantitative 16S rRNA RT-PCR analysis of the main fecal bacterial groups (Enterobacteriaceae, enterococci, lactobacilli, bifidobacteria, Bacteroides/Prevotella (BP) spp., Clostridium leptum and coccoides groups). Mice were homozygous (HO) or heterozygous (HE) for a targeted inactivating mutation of either the IFN-γ Receptor (R), IFN-γ, Rag1 or IL-4 genes. Overall, differences in IM composition were subtle. However, in the SPF-barrier, total eubacterial loads were higher in Rag1 HE versus Rag1 HO mice as well as in IFN-γR HE versus IFN-γR HO and WT animals. Although absent in WT mice, bifidobacterial loads were higher in HO and HE IFN-γ and Rag1 as well as IL-4 HO mice. Furthermore, BP was slightly lower in HO and HE IFN-γR and IFN-γ mice as well as in IL-4 HO mice as compared to WT controls. Interestingly, IM compositions were comparable in WT mice when kept in individual ventilated cages (IVC) or open cages (OC). IFN-γ HO and HE mice, however, had higher enterobacteria and BP loads, but lacked bifidobacteria when kept in OC versus IVC, as was the case in HO and HE Rag1 mice. In addition, Rag1 HO mice harbored higher clostridial loads when housed in OC as compared to IVC. Unexpectedly, lactobacilli levels were higher in IFN-γR mice when kept in OC versus IVC.

Conclusion/significance: Housing-dependent and immune-deficiency mediated changes in intestinal microbiota composition were rather subtle but may nevertheless impact immunopathology in experimental models.

No MeSH data available.


Related in: MedlinePlus