Available online at www.sciencedirect.com ScienceDirect Mimivirus inaugurated in the 21st century the beginning of a reclassification of viruses 3,4 1,2,4 3 Vikas Sharma , Philippe Colson , Pierre Pontarotti and 1,2 Didier Raoult Mimivirus and other giant viruses are visible by light to microbes [3–5]. Accordingly, these agents were named microscopy and bona fide microbes that differ from other ultraviruses, or inframicrobes, and, eventually, viruses viruses and from cells that have a ribosome. They can be [1 ]. During the 1910–1920s, viruses became increasingly defined by: giant virion and genome sizes; their complexity, established as small entities that need living cells to repli- with the presence of DNA and mRNAs and dozens or hundreds cate; Rickettsia and Chlamydia, also intracellular parasites, of proteins in virions; the presence of translation-associated definitively turned out not being viruses [6,7]. During the components; a mobilome including (pro)virophages (and a 1930–1940s, the first electron micrographs of virions were defence mechanism, named MIMIVIRE, against them) and obtained [8] and the eclipse period of virus replication was transpovirons; their monophyly; the presence of the most discovered [9]. Then, during the 1950s, the virus concept archaic protein motifs they share with cellular organisms but was unravelled by A. Lwoff, based mainly on negative not other viruses; a broader host range than other viruses. criteria [1 ]. Lwoff defined viruses as potentially patho- These features show that giant viruses are specific, genic strictly intracellular entities, which have either DNA autonomous, biological entities that warrant the creation of a or RNA, multiply in the form of their genetic material, are new branch of microbes. unable to grow and divide, and are devoid of energy Addresses production enzymes. Hence, viruses were considered as 1 Aix-Marseille Univ., URMITE UM 63 CNRS 7278 IRD 198 INSERM simple cell parasites consisting of a nucleic acid enclosed in U1095, 27 boulevard Jean Moulin, 13385 Marseille Cedex 05, France 2 a symmetric protein shell, the capsid [1 ,7], and were, IHU Me´ diterrane´ e Infection, Assistance Publique-Hoˆ pitaux de further, also shown to lack ribosomes [10]. Marseille, Centre Hospitalo-Universitaire Timone, Poˆ le des Maladies Infectieuses et Tropicales Clinique et Biologique, Fe´ de´ ration de Bacte´ riologie-Hygie` ne-Virologie, 264 rue Saint-Pierre, 13385 Marseille Cedex 05, France Mimivirus challenges the definition of viruses 3 Aix-Marseille Univ., Ecole Centrale de Marseille, I2M UMR 7373, CNRS During the last 12 years, six new or putative families of e´ quipe Evolution Biologique et Mode´ lisation, Marseille, France giant viruses have been discovered through co-culture isolation, by inoculating environmental and human sam- Corresponding author: Raoult, Didier ([email protected]) 4 Contributed equally. ples on amoebas. Mimivirus was the pioneer of this viral group [11 ,12 ]. Visibility under a light microscope and Current Opinion in Microbiology 2016, 31:16–24 the Gram positivity of this virus, isolated in 1992 from This review comes from a themed issue on Virus reclassification cooling tower water, misled researchers into considering it as a bacterium. It was eventually revealed in 2003 to Edited by Didier Raoult and Jonatas Abrahao harbour a 0.5-mm-large icosahedral capsid and a 1.2- megabase pair (Mbp)-large genome with 1.000 genes [12 ]. The discovery of Mimivirus led several groups to http://dx.doi.org/10.1016/j.mib.2015.12.010 search for other giant viruses using amoeba co-culture. # 1369-5274/ 2015 The Authors. Published by Elsevier Ltd. This is an Subsequently, isolations of Marseillevirus [13], Pandor- open access article under the CC BY-NC-ND license avirus spp. [14,15], Pithovirus sibericum [16], faustoviruses (http://creativecommons.org/licenses/by-nc-nd/4.0/). [17] and Mollivirus sibericum [18] confirmed the fruitful- ness of this culture strategy. All these viruses were discovered in Marseille, France, by two different teams. The concept and definition of viruses during Moreover, the first virophage (a Mimivirus-infecting vi- the 19th and 20th centuries rus) was also identified in this city. Strikingly, these For a long time, the concept of ‘virus’ was muddled viruses were isolated through strategies (co-culture on (Figure 1). The term ‘virus’ initially designated any Acanthamoeba polyphaga, or Vermamoeba vermiformis for infectious agent [1 ]. During the 19th century, Pasteur faustoviruses) implemented to grow microbes, and dis- and Roux considered the rabies agent as a microbe, covered by bacteriologists [19]. although it was invisible under a light microscope [2]. Between 1886 and 1898, the foundations of virology were These giant amoeba viruses were linked through phylo- laid, with the discovery of causative agents of tobacco genomics to other double-stranded DNA viruses includ- mosaic and foot-and-mouth diseases, which were ultra- ing poxviruses, asfarviruses, asco-/irido-viruses, and filterable and invisible under light microscopy, in contrast phycodnaviruses, which were formerly the largest viral Current Opinion in Microbiology 2016, 31:16–24 www.sciencedirect.com Virus reclassification Sharma et al. 17 Figure 1 Virus = ‘poison’ = infectious agent 1898 ‘Contagium vivum fluidum’ Viruses of eukaryotes Ultrafilterable, invisible under a light microscope not a microbe ‘Ultraviruses’, ‘inframicrobes’, ’small microbes’ Viruses Mycoplasma sp. Strictly intracellular Viruses Small strictly intracellular bacteria: Chlamydia spp. Rickettsia spp. 1913 First cultivation of a virus 1915-17 Viruses of bacteria 1938 First electron micrographs of viruses 1937-1943 Discovery of eclipse phase of the viral replicative cycle Characterization of ribosomes 1950s 1957 Virus concept : 1959 • potentially pathogenic entities The term ‘capsid is coined • strictly intracellular • have a single type of nucleic acid (DNA or RNA) • multiply in the form of their genetic material • are unable to grow and divide • are devoid of a system of enzymes for energy production Microbes No ribosome, ribosomal protein or RNA 1974 Viruses of archeae Chlamydia spp. Rickettsia spp. 1976 Carsonella rudii Complete viral genome Hodgkinia cicadicola sequencing Mycoplasma spp. 2003 Giant viruses Mimivirus discovery Giruses (Giant) ‘Megavirales’ TRUC 2008 ‘Traditional’ viruses Virophages Virophage discovery Capsid Encoding Organisms Organisms replicating strictly intracellularly, with an eclipse phase, and devoid of ribosomal material Current Opinion in Microbiology Schematic of a brief history of virus naming and definition. The usage and significance of the term ‘virus’ changed over time. The definition of what a virus is evolved in different steps according to new discoveries and technologies. Giant amoeba viruses share numerous features with small intracellular microbes and stand apart from ‘traditional’ viruses, whose definition was mainly founded by Lwoff during the 1950s. www.sciencedirect.com Current Opinion in Microbiology 2016, 31:16–24 18 Virus reclassification representatives and were shown in 2001 to share a set of ba viruses. The other breaks may suggest that the diver- 41 core conserved genes and grouped under the name of sity of giant amoeba viruses is greater that currently nucleocytoplasmic large DNA viruses (NCLDV) apprehended. [12 ,20 ,21 ]. Then maximum-likelihood reconstruc- tion of the evolution of these viruses mapped a set of Complexity 50 genes on their putative ancestor [22 ]. In 2012, it Giant viruses are more complex than ‘traditional’ viruses was proposed to classify giant amoeba viruses and in terms of their nucleic acid and protein content. Thus, NCLDV families in a new viral order, Megavirales, as in contrast with most other viruses, megaviruses harbour these viruses have a common origin and virion architec- both DNA and RNA, which includes messenger RNAs ture and share major biological characteristics, such as and transfer RNAs [23 ,30]. In addition, proteomics replication within viral factories [23 ]. The term ‘Girus’ identified dozens or hundreds of proteins inside giant was also coined to designate these megaviruses, to un- virions, some of which are involved in transcription and derline their intermediate status between small parasitic translation, and a substantial proportion of which are prokaryotes and standard viruses [24]. Successive isola- hypothetical proteins [14,16–18,30]. These messenger tions of new Megavirales representatives continued to RNAs and proteins may facilitate the first steps in the challenge previously established viral hallmark features replicative cycle and make giant viruses far less depen- and definitions. Simultaneously, the discovery of new dent on their host for replication than other viruses. giant viruses highlighted their diversity and ubiquity on earth and, for some, their presence in humans and, Presence of components of translation consistently, megaviruses related sequences were The discovery of Mimivirus revealed the presence of detected in environmental and human metagenomes translation factors including a peptide chain release factor [25,26]. The remarkable features of giant amoeba viruses eRF1, a GTP-binding elongation factor eF-Tu, two challenged the virus paradigm and fuelled debates on the translation initiation factors, SUI1 and 4E, and four evolution, origin and the definition of viruses aminoacyl-tRNA synthetases, some of which were shown [12 ,27 ,28 ]. In particular, their gene repertoire was to be functional