A comparative analysis of the intestinal metagenomes present in guinea pigs (Cavia porcellus) and humans (Homo sapiens).

BACKGROUND: Guinea pig (Cavia porcellus) is an important model for human intestinal research. We have characterized the faecal microbiota of 60 guinea pigs using Illumina shotgun metagenomics, and used this data to compile a gene catalogue of its prevalent microbiota. Subsequently, we compared the guinea pig microbiome to existing human gut metagenome data from the MetaHIT project. RESULTS: We found that the bacterial richness obtained for human samples was lower than for guinea pig samples. The intestinal microbiotas of both species were dominated by the two phyla Bacteroidetes and Firmicutes, but at genus level, the majority of identified genera (320 of 376) were differently abundant in the two hosts. For example, the guinea pig contained considerably more of the mucin-degrading Akkermansia, as well as of the methanogenic archaea Methanobrevibacter than found in humans. Most microbiome functional categories were less abundant in guinea pigs than in humans. Exceptions included functional categories possibly reflecting dehydration/rehydration stress in the guinea pig intestine. Finally, we showed that microbiological databases have serious anthropocentric biases, which impacts model organism research. CONCLUSIONS: The results lay the foundation for future gastrointestinal research applying guinea pigs as models for humans.

Xylo-oligosaccharides inhibit pathogen adhesion to enterocytes in vitro.

We previously reported that the non-digestible carbohydrates inulin and apple pectin promoted Listeria monocytogenes infection in guinea pigs, whereas xylo- and galacto-oligosaccharides (XOS and GOS), prevented infection by this pathogen. In the present study, mechanisms that could explain the previous in vivo observations were explored. Mixing bacterial cultures with XOS significantly (P < 0.05) decreased the ability of two out of three strains of L. monocytogenes to adhere to Caco-2 cells. Additionally, 2 h incubation with XOS followed by washing of the bacteria significantly (P < 0.05) decreased the ability of all three strains to adhere to Caco-2 cells. Consistently, expression of the adhesion-relevant genes inlA and lap was reduced by the presence of XOS. The observation that XOS inhibit the adhesion of Listeria to the intestinal epithelium in vitro may explain the reported preventive effect of XOS on Listeria infection in guinea pigs in vivo, while the preventive effect of GOS was not explicable by the assays chosen here.

Certain dietary carbohydrates promote Listeria infection in a guinea pig model, while others prevent it.

It has been proposed that dietary non-digestible carbohydrates can improve host resistance to intestinal infections by stimulating health-promoting bacteria in the gut. However, evidence from in vivo infection studies is scarce, particularly for gram-positive infections. We studied the effect of five non-digestible carbohydrates on the resistance of guinea pigs to Listeria monocytogenes infections. Animals were fed a diet supplemented with 10% xylooligosaccharides (XOS), galactooligosaccharides (GOS), inulin, apple pectin or polydextrose for three weeks before oral infection with a mixture of three different fluorescently labeled L. monocytogenes strains. Colonisation of L. monocytogenes in the intestine was determined by quantification of L. monocytogenes in faecal, ileal and caecal samples while translocation was determined by quantification of L. monocytogenes in mesenteric lymph nodes, spleen and liver. XOS and GOS significantly (P<0.05) improved the resistance of guinea pigs to L. monocytogenes, while inulin and apple pectin decreased the resistance (P<0.05). No significant effect on resistance to L. monocytogenes was seen after feeding with polydextrose. No difference in caecal weight or pH between the dietary groups was measured, except for a higher caecal weight and a lower caecal pH of animals fed with XOS, and a lower caecal pH for animals fed with polydextrose. In conclusion, this study shows for the first time that different non-digestible carbohydrates can have entirely different effects on the intestinal colonisation and translocation of a pathogenic bacterium.

Defining a role for Hfq in Gram-positive bacteria: evidence for Hfq-dependent antisense regulation in Listeria monocytogenes.

Small trans-encoded RNAs (sRNAs) modulate the translation and decay of mRNAs in bacteria. In Gram-negative species, antisense regulation by trans-encoded sRNAs relies on the Sm-like protein Hfq. In contrast to this, Hfq is dispensable for sRNA-mediated riboregulation in the Gram-positive species studied thus far. Here, we provide evidence for Hfq-dependent translational repression in the Gram-positive human pathogen Listeria monocytogenes, which is known to encode at least 50 sRNAs. We show that the Hfq-binding sRNA LhrA controls the translation and degradation of its target mRNA by an antisense mechanism, and that Hfq facilitates the binding of LhrA to its target. The work presented here provides the first experimental evidence for Hfq-dependent riboregulation in a Gram-positive bacterium. Our findings indicate that modulation of translation by trans-encoded sRNAs may occur by both Hfq-dependent and -independent mechanisms, thus adding another layer of complexity to sRNA-mediated riboregulation in Gram-positive species.

Identification of small Hfq-binding RNAs in Listeria monocytogenes.

The RNA-binding protein Hfq plays important roles in bacterial physiology and is required for the activity of many small regulatory RNAs in prokaryotes. We have previously shown that Hfq contributes to stress tolerance and virulence in the Gram-positive human pathogen Listeria monocytogenes. In the present study, we performed coimmunoprecipitations followed by enzymatic RNA sequencing to identify Hfq-binding RNA molecules in L. monocytogenes. The approach resulted in the discovery of three small RNAs (sRNAs). The sRNAs are conserved between Listeria species, but were not identified in other bacterial species. The initial characterization revealed a number of unique features displayed by each individual sRNA. The first sRNA is encoded from within an annotated gene in the L. monocytogenes EGD-e genome. Analogous to most regulatory sRNAs in Escherichia coli, the stability of this sRNA is highly dependent on the presence of Hfq. The second sRNA appears to be produced by a transcription attenuation mechanism, and the third sRNA is present in five copies at two different locations within the L. monocytogenes EGD-e genome. The cellular levels of the sRNAs are growth phase dependent and vary in response to growth medium. All three sRNAs are expressed when L. monocytogenes multiplies within mammalian cells. This study represents the first attempt to identify sRNAs in L. monocytogenes.