  Citation: Galaris, A.; Fanidis, D.; Stylianaki, E.-A.; Harokopos, V.; Kalantzi, A.-S.; Moulos, P.; Dimas, A.S.; Hatzis, P.; Aidinis, V. Obesity Reshapes the Microbial Population Structure along the Gut-Liver-Lung Axis in Mice. Biomedicines 2022, 10, 494. https://doi.org/10.3390/ biomedicines10020494 Academic Editor: Eugenia Bezirtzoglou Received: 3 January 2022 Accepted: 17 February 2022 Published: 19 February 2022 Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affil- iations. Copyright: © 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/). biomedicines Article Obesity Reshapes the Microbial Population Structure along the Gut-Liver-Lung Axis in Mice Apostolos Galaris 1,† , Dionysios Fanidis 1,† , Elli-Anna Stylianaki 1 , Vaggelis Harokopos 2 , Alexandra-Styliani Kalantzi 1 , Panagiotis Moulos 2 , Antigone S. Dimas 1 , Pantelis Hatzis 2 and Vassilis Aidinis 1, * 1 Institute of Bioinnovation, Biomedical Sciences Research Center Alexander Fleming, 16672 Athens, Greece; galaris@fleming.gr (A.G.); fanidis@fleming.gr (D.F.); stylianaki@fleming.gr (E.-A.S.); kalatzi@fleming.gr (A.-S.K.); dimas@fleming.gr (A.S.D.) 2 Institute for Fundamental Biomedical Research, Biomedical Sciences Research Center Alexander Fleming, 16672 Athens, Greece; harokopos@fleming.gr (V.H.); moulos@fleming.gr (P.M.); hatzis@fleming.gr (P.H.) * Correspondence: v.aidinis@fleming.gr † These authors contributed equally to this work. Abstract: The microbiome is emerging as a major player in tissue homeostasis in health and disease. Gut microbiome dysbiosis correlates with several autoimmune and metabolic diseases, while high-fat diets and ensuing obesity are known to affect the complexity and diversity of the microbiome, thus modulating pathophysiology. Moreover, the existence of a gut-liver microbial axis has been proposed, which may extend to the lung. In this context, we systematically compared the microbiomes of the gut, liver, and lung of mice fed a high-fat diet to those of littermates fed a matched control diet. We carried out deep sequencing of seven hypervariable regions of the 16S rRNA microbial gene to examine microbial diversity in the tissues of interest. Comparison of the local microbiomes indicated that lung tissue has the least diverse microbiome under healthy conditions, while microbial diversity in the healthy liver clustered closer to the gut. Obesity increased microbial complexity in all three tissues, with lung microbial diversity being the most modified. Obesity promoted the expansion of Firmicutes along the gut-liver-lung axis, highlighting staphylococcus as a possible pathologic link between obesity and systemic pathophysiology, especially in the lungs. Keywords: obesity; high-fat diet; microbiome; 16S rRNA; gut; liver; lung; comparative analysis; firmicutes; staphylococcus 1. Introduction The microbiome, the sum of commensal, symbiotic, and pathogenic organisms that populate animal bodies, is increasingly recognized as a major player in tissue homeostasis in health and disease [1], modulating a variety of host functions, including immunity and inflammation [2], as well as energy homeostasis and metabolism [3]. Changes in microbial population structure and the ensuing local or systemic effects can be induced by different environmental factors, most notably exposure to antibiotics and dietary changes, while the efficacy of various medications has been suggested to correlate with microbiome perturbations and vice versa [1]. Most microorganisms reside within the intestine. It is well established that the gut microbiome participates in multiple homeostatic functions essential for the host, including nutrient absorption and education of the immune system. Alterations in the composition and complexity of microbiomes can harm the health of an organism. Such alterations lead to dysbiosis and have been associated with autoimmune and metabolic diseases, mostly via secreted microbial metabolites [4]. Non-alcoholic fatty liver disease (NAFLD) has been linked to dysbiosis [5,6], highlighting a connection between gut microbiota and the liver, referred to as the gut-liver axis [7]. The gut and liver are in direct contact Biomedicines 2022, 10, 494. https://doi.org/10.3390/biomedicines10020494 https://www.mdpi.com/journal/biomedicines