Sarah Temmam

Massilia Virus, A Novel Phlebovirus (Bunyaviridae) Isolated from Sandflies in the Mediterranean

A new virus was isolated from three independent pools of Phlebotomus perniciosus sandflies (Diptera; Psychodidae) trapped in two regions of southeastern France, located 90 miles apart. Microscopic, antigenic and genetic analyses indicate that this novel virus belongs to the genus Phlebovirus in the family Bunyaviridae. The new virus is designated Massilia virus since the first isolate was obtained from sandflies collected in the suburban area of Marseille. The complete genome sequence was determined and used to compare the genetic and phylogenetic relationships of Massilia virus with other phleboviruses. Genetic and antigenic properties were employed to address whether or not Massilia virus should be considered a new species within the genus, or a member of a previously recognized species. Cerebrospinal fluid specimens, collected from local patients with central nervous system infections during the previous four-year period were tested for the presence of Massilia virus RNA, but gave negative results. In conclusion, Massilia virus is proposed as a member of the Sandfly fever Naples virus complex; its public health importance has yet to be determined.

Describing the silent human virome with an emphasis on giant viruses.

Viruses are the most abundant obligate intracellular entities in our body. Until recently, they were only considered to be pathogens that caused a broad array of pathologies, ranging from mild disease to deaths in the most severe cases. However, recent advances in unbiased mass sequencing techniques as well as increasing epidemiological evidence have indicated that the human body is home to diverse viral species under non-pathological conditions. Despite these studies, the description of the presumably healthy viral flora, i.e. the normal human virome, is still in its infancy regarding viral composition and dynamics. This review summarizes our current knowledge of the human virome under non-pathological conditions.

Viral Metagenomics on Animals as a Tool for the Detection of Zoonoses Prior to Human Infection

Many human viral infections have a zoonotic, i.e., wild or domestic animal, origin. Several zoonotic viruses are transmitted to humans directly via contact with an animal or indirectly via exposure to the urine or feces of infected animals or the bite of a bloodsucking arthropod. If a virus is able to adapt and replicate in its new human host, human-to-human transmissions may occur, possibly resulting in an epidemic, such as the A/H1N1 flu pandemic in 2009. Thus, predicting emerging zoonotic infections is an important challenge for public health officials in the coming decades. The recent development of viral metagenomics, i.e., the characterization of the complete viral diversity isolated from an organism or an environment using high-throughput sequencing technologies, is promising for the surveillance of such diseases and can be accomplished by analyzing the viromes of selected animals and arthropods that are closely in contact with humans. In this review, we summarize our current knowledge of viral diversity within such animals (in particular blood-feeding arthropods, wildlife and domestic animals) using metagenomics and present its possible future application for the surveillance of zoonotic and arboviral diseases.

Viral metagenomics: are we missing the giants?

Amoeba-infecting giant viruses are recently discovered viruses that have been isolated from diverse environments all around the world. In parallel to isolation efforts, metagenomics confirmed their worldwide distribution from a broad range of environmental and host-associated samples, including humans, depicting them as a major component of eukaryotic viruses in nature and a possible resident of the human/animal virome whose role is still unclear. Nevertheless, metagenomics data about amoeba-infecting giant viruses still remain scarce, mainly because of methodological limitations. Efforts should be pursued both at the metagenomic sample preparation level and on in silico analyses to better understand their roles in the environment and in human/animal health and disease.

Characterization of Viral Communities of Biting Midges and Identification of Novel Thogotovirus Species and Rhabdovirus Genus

More than two thirds of emerging viruses are of zoonotic origin, and among them RNA viruses represent the majority. Ceratopogonidae (genus Culicoides) are well-known vectors of several viruses responsible for epizooties (bluetongue, epizootic haemorrhagic disease, etc.). They are also vectors of the only known virus infecting humans: the Oropouche virus. Female midges usually feed on a variety of hosts, leading to possible transmission of emerging viruses from animals to humans. In this context, we report here the analysis of RNA viral communities of Senegalese biting midges using next-generation sequencing techniques as a preliminary step toward the identification of potential viral biohazards. Sequencing of the RNA virome of three pools of Culicoides revealed the presence of a significant diversity of viruses infecting plants, insects and mammals. Several novel viruses were detected, including a novel Thogotovirus species, related but genetically distant from previously described tick-borne thogotoviruses. Novel rhabdoviruses were also detected, possibly constituting a novel Rhabdoviridae genus, and putatively restricted to insects. Sequences related to the major viruses transmitted by Culicoides, i.e., African horse sickness, bluetongue and epizootic haemorrhagic disease viruses were also detected. This study highlights the interest in monitoring the emergence and circulation of zoonoses and epizooties using their arthropod vectors.

Host-Associated Metagenomics: A Guide to Generating Infectious RNA Viromes.

Background Metagenomic analyses have been widely used in the last decade to describe viral communities in various environments or to identify the etiology of human, animal, and plant pathologies. Here, we present a simple and standardized protocol that allows for the purification and sequencing of RNA viromes from complex biological samples with an important reduction of host DNA and RNA contaminants, while preserving the infectivity of viral particles. Principal Findings We evaluated different viral purification steps, random reverse transcriptions and sequence-independent amplifications of a pool of representative RNA viruses. Viruses remained infectious after the purification process. We then validated the protocol by sequencing the RNA virome of human body lice engorged in vitro with artificially contaminated human blood. The full genomes of the most abundant viruses absorbed by the lice during the blood meal were successfully sequenced. Interestingly, random amplifications differed in the genome coverage of segmented RNA viruses. Moreover, the majority of reads were taxonomically identified, and only 7–15% of all reads were classified as “unknown”, depending on the random amplification method. Conclusion The protocol reported here could easily be applied to generate RNA viral metagenomes from complex biological samples of different origins. Our protocol allows further virological characterizations of the described viral communities because it preserves the infectivity of viral particles and allows for the isolation of viruses.

First molecular detection of Rickettsia africae in ticks from the Union of the Comoros

BackgroundRickettsia africae is the agent of African tick bite fever, a disease transmitted by ticks in sub-Saharan Africa. In Union of the Comoros, a recent study reported the presence of a Rickettsia africae vector but no information has been provided on the circulation of the pathogenic agent in this country.MethodsTo evaluate the possible circulation of Rickettsia spp. in Comorian cattle, genomic DNA was extracted from 512 ticks collected either in the Union of the Comoros or from animals imported from Tanzania and subsequently tested for Rickettsia infection by quantitative PCR.ResultsRickettsia africae was detected in 90% (60/67) of Amblyomma variegatum, 1% (1/92) of Rhipicephalus appendiculatus and 2.7% (8/296) of Rhipicephalus (Boophilus) microplus ticks collected in the Union of the Comoros, as well as in 77.14% (27/35) of Amblyomma variegatum ticks collected from imported cattle. Partial sequences of both bacterial gltA and ompA genes were used in a phylogenetic analysis revealing the presence of several haplotypes, all included within the Rickettsia africae clade.ConclusionsOur study reports the first evidence of Rickettsia africae in ticks collected from the Union of the Comoros. The data show a significant difference of infection rate of Rickettsia africae infected ticks between the Islands, with maximum rates measured in Grande Comore Island, sheltering the main entry port for live animal importation from Tanzania. The high infection levels reported herein indicate the need for an in-depth assessment of the burden of rickettsioses in the Union of the Comoros, especially among those at risk of infection, such as cattle herders.

Screening for Viral Pathogens in African Simian Bushmeat Seized at A French Airport.

Summary Illegal bushmeat traffic is an important threat to biodiversity conservation of several endangered species and may contribute to the emergence and spread of infectious diseases in humans. The hunting, manipulation and consumption of wildlife‐based products, especially those of primate origin, may be a threat to human health; however, few studies have investigated the role of bushmeat trade and consumption as a potential source of human infections to date. In this study, we report the screening of viral pathogens in African simian game seized by French customs at Toulouse Blagnac Airport. Epifluorescence microscopy revealed the presence of virus‐like particles in the samples, and further metagenomic sequencing of the DNA and RNA viromes confirmed the presence of sequences related to the Siphoviridae, Myoviridae and Podoviridae bacteriophage families; some of them infecting bacterial hosts that could be potentially pathogenic for humans. To increase the sensitivity of detection, twelve pan‐generic PCRs targeting several viral zoonoses were performed, but no positive signal was detected. A large‐scale inventory of bacteria, viruses and parasites is urgently needed to globally assess the risk for human health of the trade, manipulation and consumption of wildlife‐related bushmeat.

Protocol for Generating Infectious RNA Viromes from Complex Biological Samples

This chapter proposes a simple, standardized protocol for generating RNA viromes from complex host-associated biological samples of various origins. Compared to other existing protocols to generate RNA viromes, this protocol preserves the infectivity of viral particles and allows for downstream applications such as viral characterization and isolation tests.