Contents lists available at ScienceDirect Microbial Pathogenesis journal homepage: www.elsevier.com/locate/micpath Sporulation on blood serum increases the virulence of Mucor circinelloides J. Alberto Patiño-Medina a,1 , David Vargas-Tejeda a,1 , Marco I. Valle-Maldonado a,b , Viridiana Alejandre-Castañeda a , Irvin E. Jácome-Galarza b , Javier Villegas-Moreno a , Rosa E. Nuñez-Anita c , Martha I. Ramírez-Díaz a , Rafael Ortiz-Alvarado d,2 , Victor Meza-Carmen a,*,2 a Instituto de Investigaciones Químico Biológicas, Universidad Michoacana de San Nicolás de Hidalgo, 58030, Morelia, Michoacán, Mexico b Departamento de Biología Molecular, Laboratorio Estatal de Salud Pública del Estado de Michoacán, 58279, Morelia, Michoacán, Mexico c Facultad de Medicina Veterinaria y Zootecnia, Universidad Michoacana de San Nicolás Hidalgo, 58130, Morelia, Michoacán, Mexico d Facultad de Químico Farmacobiología, Universidad Michoacana de San Nicolás de Hidalgo, 58240, Morelia, Michoacán, Mexico ARTICLE INFO Keywords: Mucor circinelloides Fungal sporulation Virulence Blood serum ABSTRACT Mucor circinelloides is an opportunistic human pathogen that is used to study mucormycosis, a rare but lethal infection in susceptible immunosuppressed patients. However, the virulence characteristics of this pathogen have not been fully elucidated. In this study, sporangiospores (spores) produced on YPG medium supplemented with native blood serum increased the virulence of M. circinelloides compared with spores produced on YPG supplemented with denatured blood serum or on YPG alone. The spores produced from YPG supplemented with native blood serum increased nematode death and led to significant increases in interleukin (IL)-6, IL-1β, macrophage inhibitory protein-2, and tumour necrosis factor α mRNA levels in liver and lung tissues from infected diabetic mice compared with those in tissues from animals infected with spores produced in the pre- sence of YPG supplemented with denatured blood serum or of YPG alone. Moreover, spores produced from cultures supplemented with native blood serum showed increased germination rates and longer hyphae com- pared with other spores. The spores produced in YPG supplemented with native blood serum also enhanced resistance to stress factors and H 2 O 2 and increased thermotolerance compared with spores produced under other conditions. In addition, spores produced in presence of blood serum increased the ability of the pathogen to survive in the presence of macrophages. Taken together, our results showed that these factors were important features for fungal virulence in humans and suggested that thermolabile components in the blood serum may induce M. circinelloides virulence. 1. Introduction Mucormycosis is a fungal opportunistic infection that is lethal in certain populations, particularly those at high risk, such as patients with ketoacid diabetes [1] and transplant recipients [2]. Mucormycosis is named based on the ethylogical agents that belong to the fungal order of Mucorales; the genus Rhizopus is the most common genus causing mucormycosis, followed by Lichtheimia, Apophysomyces, Rhizomucor, Mucor, and Cunninghamella [3]. The global incidence of mucormycosis is increasing, with rates 34% and 28% higher in Europe and the Americas, respectively, during the period from January 2000 through January 2017 [4]. This fungal infection is considered the third most important disease among patients with haematological conditions in Europe [5]. The worldwide incidence is 1.23 per 100,000 persons [6], although the low sensitivity of diagnostic tests could underestimate the true incidence of mucormycosis [3]. Mucor circinelloides has been used to elucidate the basic virulence mechanisms involved in mucormycosis [7–10]. Moreover, M. circi- nelloides is the most well-studied biological model among Mucorales organisms, and many molecular manipulation tools have been devel- oped for this fungus [11–13]. Fungal interactions with the host can enhance some genetic traits in fungi to increase virulence; for example, in Bauveria bassiana, con- secutive spore propagation cycles through malaria mosquitoes (Anopheles coluzzii) leads to selection of variants with increased viru- lence rates [14]. A similar effect was observed in the ascomycete https://doi.org/10.1016/j.micpath.2019.103737 Received 4 May 2019; Received in revised form 6 August 2019; Accepted 8 September 2019 * Corresponding author. Laboratorio de Diferenciación Celular, Instituto de Investigaciones Químico Biológicas, Universidad Michoacana de San Nicolás de Hidalgo, Ciudad Universitaria, 58030, Morelia, Michoacán, Mexico. E-mail address: victor_meza2004@yahoo.com.mx (V. Meza-Carmen). 1 J. Alberto Patiño-Medina and David Vargas-Tejeda contributed equally to this work. 2 Rafael Ortiz-Alvarado and Victor Meza-Carmen are the senior authors. Microbial Pathogenesis 137 (2019) 103737 Available online 09 September 2019 0882-4010/ © 2019 Elsevier Ltd. All rights reserved. T