Viruses 2021, 13, 2178. https://doi.org/10.3390/v13112178 www.mdpi.com/journal/viruses Article Macrophages and Monocytes: “Trojan Horses” in COVID‐19 Elena Percivalle 1 , Josè Camilla Sammartino 1 , Irene Cassaniti 1, *, Eloisa Arbustini 2 , Mario Urtis 2 , Alexandra Smirnova 2 , Monica Concardi 2 , Cristina Belgiovine 3 , Alessandro Ferrari 1 , Daniele Lilleri 1 , Antonio Piralla 1 and Fausto Baldanti 1,4 1 Molecular Virology Unit, Microbiology and Virology Department, Fondazione IRCCS Policlinico San Matteo, 27100 Pavia, Italy; e.percivalle@smatteo.pv.it (E.P.); jose.sammartino@iusspavia.it (J.C.S.); alessandro.ferrari04@universitadipavia.it (A.F.); d.lilleri@smatteo.pv.it (D.L.); a.piralla@smatteo.pv.it (A.P.); fausto.baldanti@unipv.it (F.B.) 2 Transplant Research Area and Centre for Inherited Cardiovascular Diseases, Department of Medical Sciences and Infectious Diseases, Fondazione IRCCS Policlinico San Matteo, 27100 Pavia, Italy; e.arbustini@smatteo.pv.it (E.A.); m.urtis@smatteo.pv.it (M.U.); a.smirnova@smatteo.pv.it (A.S.); m.concardi@smatteo.pv.it (M.C.) 3 Humanitas Clinical and Research Center—IRCCS, 20089 Milan, Italy; Cristina.Belgiovine@humanitasresearch.it 4 Department of Clinical, Surgical, Diagnostics and Pediatric Sciences, University of Pavia, 27100 Pavia, Italy * Correspondence: i.cassaniti@smatteo.pv.it Abstract: We aimed to explore whether variants of SARS‐CoV‐2 (Chinese‐derived strain (D614, lineage A), Italian strain PV10734 (D614G, lineage B.1.1) and Alpha strain (lineage B.1.1.7)) were able to infect monocytes (MN) and monocyte‐derived macrophages (MDM) and whether these infected cells may, in turn, be vectors of infection. For this purpose, we designed an in vitro study following the evolution of MN and MDM infection at different time points in order to confirm whether these cells were permissive for SARS‐CoV‐2 replication. Finally, we investigated whether, regardless of viral replication, the persistent virus can be transferred to non‐infected cells permissive for viral replication. Thus, we co‐cultured the infected MN/MDM with permissive VERO E6 cells verifying the viral transmission. This is a further in vitro demonstration of the important role of MN and MDM in the dissemination of SARS‐CoV‐2 and evolution of the COVID‐19 disease. Keywords: SARS‐CoV‐2; Trojan horse; VERO E6 cells 1. Introduction Beta‐coronaviruses are associated with human diseases, and, in the last two decades, the emergence of Middle East Respiratory Syndrome (MERS‐CoV) and Severe Acute Respiratory Syndrome (SARS‐CoV‐1) viruses, with a zoonotic origin, were connected to two outbreaks of severe respiratory diseases in 2012 and 2003, respectively [1]. In late 2019, a novel coronavirus disease (COVID‐19) caused by the new beta‐coronavirus Severe Acute Respiratory Syndrome 2 virus (SARS‐CoV‐2) was reported in Wuhan (Hubei province, China). COVID‐19 was declared as a pandemic by the World Health Organization on 11 March 2020 [2]. Despite the paramount scientific effort in the dissection of the pathogenetic mechanisms of COVID‐19, virologic and immunologic factors triggering severe disease in SARS‐CoV‐2‐infected subjects are not completely defined. In addition, while it is now evident that COVID‐19 is a multi‐organ disease [3], the virus dissemination mechanisms are not fully elucidated. A common denominator of all infected organs in COVD‐19 disease is the expression of angiotensin‐converting enzyme 2 (ACE2), which was identified as the binding receptor for the spike viral glycoprotein [4] that allowed viral internalization and replication in the host cells. However, little attention has been given to ACE‐2 expression in the immune system; indeed also, monocytes and macrophages [5] express this receptor, making them Citation: Percivalle, E.; Sammartino, J.C.; Cassaniti, I.; Arbustini, E.; Urtis, M.; Smirnova, A.; Concardi, M.; Belgiovine, C.; Ferrari, A.; Lilleri, D.; Piralla, A. Macrophages and Monocytes: “Trojan Horses” in COVID‐19. Viruses 2021, 13, 2178. https://doi.org/10.3390/v13112178 Academic Editor: Oliver Schildgen Received: 8 September 2021 Accepted: 25 October 2021 Published: 28 October 2021 Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. Copyright: © 2021 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 (http://creativecommons.org/licenses /by/4.0/).