Sarah Aherfi

Shan Virus: A New Mimivirus Isolated from the Stool of a Tunisian Patient with Pneumonia

Objective: Following the isolation of a Marseillevirus from the stool of a healthy young Senegalese and a Mimivirus from a Tunisian patient with pneumonia, we attempted to isolate other giant viruses of amoebae from a large human stool collection. Methods: During the period 2010-2011, a total of 1,605 stool samples, including 115 from Tunisian patients with pneumonia, were cultured on amoebae. We used a recently developed high-throughput isolation system to detect amoebae plaque lysis on agar plates; this method allows for the testing of 100 samples per plate per week. The giant virus was identified by sequencing of genes conserved in Megavirales. Results: A single giant virus, called Shan, was isolated from the stool of a Tunisian patient with pneumonia who responded poorly to antibiotics. This virus has an icosahedral shape typical of members of the family Mimiviridae and a size of 640 ± 10 nm. Phylogenetic analyses showed that Shan virus was classified as a member of Mimivirus lineage C that infects amoebae. Conclusion: Only one isolate was obtained in this study, suggesting that giant viruses of amoebae are rare in human stool. The isolation of Shan virus from a patient with pneumonia brings into question the etiological role of this virus and its subsequent release in stool.

First isolation of a Marseillevirus in the Diptera Syrphidae Eristalis tenax.

Objective: Giant viruses and amoebae are common in freshwater, where they can coexist with various insects. We screened insect larvae to detect giant viruses using a high-throughput method. Methods: We analyzed 86 Eristalis tenax larvae obtained from stagnant water reservoirs in Tunisia. The larvae were decontaminated and then dissected to remove internal parts for coculture with Acanthamoeba polyphaga. Genome sequencing of isolated viruses was performed on a 454 Roche instrument, and comparative genomics were performed. Results: One Marseillevirus, named Insectomime virus, was isolated. The genome assembly generated two scaffolds, which were 382,776 and 3,855 bp in length. Among the 477 identified predicted proteins, the best hit for 435 of the identified proteins was a Marseillevirus or Lausannevirus protein. Tunisvirus was the most closely related to Insectomime, with 446 orthologs. One Insectomime protein shared with Lausannevirus and Tunisvirus showed the highest similarity with a protein from an aphid. Conclusion: The isolation of a Marseillevirus from an insect expands the diversity of environments in which giant viruses have been isolated. The coexistence of larvae and giant viruses in stagnant water may explain the presence of the giant virus in the larva internal structures. This study illustrates the putative role of amoeba in lateral gene transfer not only between the organisms it phagocytoses, but also between organisms living in the same environment.

A Brazilian Marseillevirus Is the Founding Member of a Lineage in Family Marseilleviridae

In 2003, Acanthamoeba polyphaga mimivirus (APMV) was discovered as parasitizing Acanthamoeba. It was revealed to exhibit remarkable features, especially odd genomic characteristics, and founded viral family Mimiviridae. Subsequently, a second family of giant amoebal viruses was described, Marseilleviridae, whose prototype member is Marseillevirus, discovered in 2009. Currently, the genomes of seven different members of this family have been fully sequenced. Previous phylogenetic analysis suggested the existence of three Marseilleviridae lineages: A, B and C. Here, we describe a new member of this family, Brazilian Marseillevirus (BrMV), which was isolated from a Brazilian sample and whose genome was fully sequenced and analyzed. Surprisingly, data from phylogenetic analyses and comparative genomics, including mean amino acid identity between BrMV and other Marseilleviridae members and the analyses of the core genome and pan-genome of marseilleviruses, indicated that this virus can be assigned to a new Marseilleviridae lineage. Even if the BrMV genome is one of the smallest among Marseilleviridae members, it harbors the second largest gene content into this family. In addition, the BrMV genome encodes 29 ORFans. Here, we describe the isolation and genome analyses of the BrMV strain, and propose its classification as the prototype virus of a new lineage D within the family Marseilleviridae.

Liver transplantation for acute liver failure related to autochthonous genotype 3 hepatitis E virus infection.

Hepatitis E virus of genotype 3 (HEV-3) is an emerging cause of sporadic autochthonous acute hepatitis in Europe. Although spontaneous outcome of hepatitis E is usually favorable, fulminant liver failure has been described worldwide. In Europe, autochthonous hepatitis E associated with fulminant hepatic failure and leading to liver transplantation has been exceptionally reported. We report here four cases of fulminant and sub-fulminant hepatitis E proposed for liver transplantation in Marseille University hospitals between July 2006 and March 2010. HEV diagnosis relied on detection of anti-HEV IgM antibodies and HEV RNA in serum samples. All cases were men, with no travel history in hyperendemic areas. HEV sequence analyses revealed genotype 3 HEV in the four patients. Liver histology indicated severe acute hepatitis in all of them, pre-existing fibrosis being found in two cases. Two patients underwent liver transplantation, and the two other patients could not be transplanted due to septic complications and died. HEV testing should be performed for the initial evaluation of every acute liver failure regardless of the epidemiological and clinical context. With respect to the potentially fulminant evolution of HEV genotype 3 infections, treatment with ribavirin of severe acute hepatitis E should be considered.

First isolation of a giant virus from wild Hirudo medicinalis leech: Mimiviridae isolation in Hirudo medicinalis.

Giant viruses and amoebae are common in freshwater, where they can coexist with other living multicellular organisms. We screened leeches from the species Hirudo medicinalis for giant viruses. We analyzed five H. medicinalis obtained from Tunisia (3) and France (2). The leeches were decontaminated and then dissected to remove internal parts for co-culture with Acanthamoeba polyphaga. The genomes of isolated viruses were sequenced on a 454 Roche instrument, and a comparative genomics analysis was performed. One Mimivirus was isolated and the strain was named Hirudovirus. The genome assembly generated two scaffolds, which were 1,155,382 and 25,660 base pairs in length. Functional annotations were identified for 47% of the genes, which corresponds to 466 proteins. The presence of Mimividae in the same ecological niche as wild Hirudo may explain the presence of the mimivirus in the digestive tract of the leech, and several studies have already shown that viruses can persist in the digestive tracts of leeches fed contaminated blood. As leeches can be used medically and Mimiviruses have the potential to be an infectious agent in humans, patients treated with leeches should be surveyed to investigate a possible connection.

The expanding family Marseilleviridae

The family Marseilleviridae encompasses giant viruses that replicate in free-living Acanthamoeba amoebae. Since the discovery of the founding member Marseillevirus in 2007, 7 new marseilleviruses have been observed, including 3 from environmental freshwater, one from a dipteran, and two from symptom-free humans. Marseilleviruses have ≈250-nm-large icosahedral capsids and 346-386-kb-long mosaic genomes that encode 444-497 predicted proteins. They share a small set of core genes with Mimivirus and other large and giant DNA viruses that compose a monophyletic group, first described in 2001. Comparative genomics analyses indicate that the family Marseilleviridae currently includes three lineages and a pan-genome composed of ≈600 genes. Antibodies against marseilleviruses and viral DNA have been observed in a significant proportion of asymptomatic individuals and in the blood and lymph nodes of a child with adenitis; these observations suggest that these giant viruses may be blood borne and question if they may be pathogenic in humans.

Cedratvirus, a Double-Cork Structured Giant Virus, is a Distant Relative of Pithoviruses

Most viruses are known for the ability to cause symptomatic diseases in humans and other animals. The discovery of Acanthamoeba polyphaga mimivirus and other giant amoebal viruses revealed a considerable and previously unknown area of uncharacterized viral particles. Giant viruses have been isolated from various environmental samples collected from very distant geographic places, revealing a ubiquitous distribution. Their morphological and genomic features are fundamental elements for classifying them. Herein, we report the isolation and draft genome of Cedratvirus, a new amoebal giant virus isolated in Acanthamoeba castellanii, from an Algerian environmental sample. The viral particles are ovoid-shaped, resembling Pithovirus sibericum, but differing notably in the presence of two corks at each extremity of the virion. The draft genome of Cedratvirus—589,068 base pairs in length—is a close relative of the two previously described pithoviruses, sharing 104 and 113 genes with P. sibericum and Pithovirus massiliensis genomes, respectively. Interestingly, analysis of these viruses’ core genome reveals that only 21% of Cedratvirus genes are involved in best reciprocal hits with the two pithoviruses. Phylogeny reconstructions and comparative genomics indicate that Cedratvirus is most closely related to pithoviruses, and questions their membership in an enlarged putative Pithoviridae family.

Marseillevirus in lymphoma: a giant in the lymph node

The family Marseilleviridae is a new clade of giant viruses whose original member, marseillevirus, was described in 2009. These viruses were isolated using Acanthamoeba spp primarily from the environment. Subsequently, a close relative of marseillevirus was isolated from the faeces of a healthy young man, and others were detected in blood samples of blood donors and recipients and in a child with lymph node adenitis. In this Grand Round we describe the detection of marseillevirus by PCR, fluorescence in-situ hybridisation, direct immunofluorescence, and immunohistochemistry in the lymph node of a 30-year-old woman diagnosed with Hodgkins lymphoma, together with IgG antibodies to marseillevirus. A link with viruses and bacteria has been reported for many lymphomas. We review the literature describing these associations, the criteria used to consider a causal association, and the underlying mechanisms of lymphomagenesis. Our observations suggest that consideration should be given to marseillevirus infections as an additional viral cause or consequence of Hodgkins lymphoma, and that this hypothesis should be tested further.

The role of giant viruses of amoebas in humans.

Since 2003, dozens of giant viruses that infect amoebas (GVA), including mimiviruses and marseilleviruses, have been discovered. These giants appear to be common in our biosphere. From the onset, their presence and possible pathogenic role in humans have been serendipitously observed or investigated using a broad range of technological approaches, including culture, electron microscopy, serology and various techniques based on molecular biology. The link between amoebal mimiviruses and pneumonia has been the most documented, with findings that fulfill several of the criteria considered as proof of viral disease causation. Regarding marseilleviruses, they have been mostly described in asymptomatic persons, and in a lymph node adenitis. The presence and impact of GVA in humans undoubtedly deserve further investigation in medicine.

Hepatitis C virus NS3 protease genotyping and drug concentration determination during triple therapy with telaprevir or boceprevir for chronic infection with genotype 1 viruses, southeastern France

Telaprevir and boceprevir, the two first hepatitis C virus (HCV) NS3 protease inhibitors (PIs), considerably increase rates of sustained virologic response in association with pegylated interferon and ribavirin in chronic HCV genotype 1 infections. The 30 first patients treated by telaprevir or boceprevir including anti‐HCV therapies since 2011 in Marseille University hospitals, France, were monitored. HCV loads and plasmatic concentrations of telaprevir and boceprevir were determined on sequential blood samples. HCV NS3 protease gene population sequencing was performed at baseline of treatment and in case of treatment failure. Fifteen patients (including 7 co‐infected with HIV) received telaprevir and the other 15 patients (including 4 co‐infected with HIV) received boceprevir. At baseline, HCV NS3 protease from six patients harbored amino acid substitutions associated with PI‐resistance. Treatment failure occurred at week 12 for 7 patients. Amino acid substitutions associated with PI‐resistance were observed in six of these cases. HCV NS3 R155K and T54A/S mutants, all of genotype 1a, were found from four patients. Median (interquartile range) plasma concentrations were 3,092 ng/ml (2,320–3,525) for telaprevir and 486 ng/ml (265–619) for boceprevir. For HIV–HCV co‐infected patients, median concentrations were 3,162 ng/ml (2,270–4,232) for telaprevir and 374 ng/ml (229–519) for boceprevir. Plasma drug concentration monitoring revealed undetectable concentrations for two patients at week 4, and probable non‐adherence to therapy for another patient. These findings indicate that routine HCV NS3 protease sequencing and plasma PI concentration monitoring might be helpful to characterize cases of therapy failure, at a cost dramatically low compared to that of anti‐HCV therapy. J. Med. Virol. 86:1868–1876, 2014.