Exploring the interaction between the human microbiota and infections

The microbiota is a living ecosystem that is influenced by a variety of host and environmental factors. Distinct microbiota colonizes various body sites, such as the gastrointestinal system and vaginal tract, corresponding to the unique microenvironment. A healthy gut microbiota contains a stable, balanced, and highly diverse reservoir of microbes. Commensal microorganisms co-evolved with the host have conferred pathogen colonization resistance. Lactobacillus species usually dominate the vaginal microbiota of majority healthy women. The vaginal microbiota has also been linked to sexually transmitted diseases, such as the human papillomavirus (HPV) infection. MicroRNA expression was found to be associated with microbiota composition. We studied the interactions of microbiota gut and vagina with metabolites, miRNA expression, and HPV infection in our investigations. In vitro three- dimensional (3D) cell-culture methods were also analyzed and developed for mechanistic investigations of the microbiota-host interaction.

Study I is a cross-sectional study investigating the microbiota features in HPV-related diseases. We defined the HPV-related microbial composition in a high-vaccination coverage population of 345 young Swedish women. The associations of microbial composition and the infection of 27 HPV types were analyzed. HPV infection, especially for the infection of oncogenic HPV types, was characterized by a higher microbial alpha-diversity with non- Lactobacillus-dominant vaginal microbiota composition. The prevalence of bacterial vaginosis-associated bacteria (BV AB), Sneathia, Prevotella, and Megasphaera were significantly higher in women with HPV infection than in uninfected individuals.

Study II investigated the associations among miRNA, HPV infection, and vaginal microbiota composition. A global miRNA expression increase was identified in non-Lactobacillus- dominate women compared with Lactobacillus-dominated women. The top two differently expressed miRNA, miR-23a-3p and miR-130a-3p, showed a prediction accuracy of > 97% for distinguishing Lactobacillus-dominated and non- Lactobacillus-dominated samples. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of the target genes of miR-23a-3p and miR-130a-3p found that many pathways involved in cancer and infection were enriched, such as the mitogen-activated protein kinase (MAPK) signaling pathway, and the phosphatidylinositol 3-kinase (PI3K)-Akt signaling pathway.

Study III explored the interactions between commensal bacteria and pathogens. We cocultured Salmonella with commensal bacteria isolations from an anaerobically cultivated human intestinal microflora (ACHIM). Commensal bacteria isolations belonging to the Clostridium and Eubacterium family showed significant inhibition of Salmonella growth. Following metabolite extraction and liquid chromatography-mass spectrometry analysis of the commensal bacteria and Salmonella metabolites in the coculture system of the commensal bacteria and Salmonella, adenine and adenosine were significantly higher in the coculture systems that inhibited Salmonella growth compared with the expressive systems. Functional assays of metabolite activity further validated the inhibition effect of adenine and adenosine on the growth of Salmonella and other antibiotic-resistant pathogens.

Study IV established an anaerobic gut 3D model to study bacteria-host interactions. The Caco2 cells showed good cell viability and formed intestinal villi-like structures in our model. Anaerobic culture for 12 hours did not show a significant effect on the viability of cells, with only six genes differently expressed between cells cultured under anaerobic and aerobic conditions in RNA sequencing. RNA sequencing of Salmonella infected Caco2 cells cocultured in anaerobic conditions showed a large set of genes differentially expressed compared to aerobic conditions with the pathways associated with cell cycle, homologous recombination, and DNA replication. This supports our gut 3D model could be used for investigating host-microbe direct interactions under the anaerobic condition, which is essential for obligate anaerobic gut microbes.

List of scientific papers

I. LIQIN CHENG*, Johanna Norenhag*, Yue O. O. Hu*, Nele Brusselaers, Emma Fransson, Andreas Ährlund-Richter, Unnur Guðnadóttir, Pia Angelidou, Yinghua Zha, Marica Hamsten, Ina Schuppe-Koistinen, Matts Olovsson, Lars Engstrand, and Juan Du. Vaginal microbiota and human papillomavirus infection among young Swedish women. NPJ Biofilms Microbiomes. 2020 Oct 12;6(1):39. *Shared authorship.
https://doi.org/10.1038/s41522-020-00146-8

II. LIQIN CHENG, Dominika Ka zmierczak, Johanna Norenhag, Marica Hamsten, Emma Fransson, Ina Schuppe-Koistinen, Matts Olovsson, Lars Engstrand, Per Hydbring* and Juan Du*. A microRNA gene panel predicts the vaginal microbiota composition. mSystems. 2021 May 4;6(3):e00175-21. *Shared authorship.
https://doi.org/10.1128/mSystems.00175-21

III. LIQIN CHENG*, Mário S. P. Correia*, Shawn M. Higdon, Enrique Joffre, Åsa Sjöling, Fredrik Boulund, Elisabeth Norin, Lars Engstrand, Daniel Globisch* and Juan Du*. Commensal gut microbes and their metabolites inhibit bacterial pathogens growth. *Shared authorship. [Submitted]

IIV. LIQIN CHENG, Tingting Liu, Qiong Liu, Liming Lian, Guosheng Tang, Luis Santiago Mille, Fabricio R. García, Lars Engstrand, Yu Shrike Zhang* and Juan Du*. A 3D-bioprinted gut anaerobic model to study bacteria-host interactions. *Shared authorship. [Submitted]