An Insect Nidovirus Emerging from a Primary Tropical Rainforest

Tropical rainforests show the highest level of terrestrial biodiversity and may be an important contributor to microbial diversity. Exploitation of these ecosystems may foster the emergence of novel pathogens. We report the discovery of the first insect-associated nidovirus, tentatively named Cavally virus (CAVV). CAVV was found with a prevalence of 9.3% during a survey of mosquito-associated viruses along an anthropogenic disturbance gradient in Côte d’Ivoire. Analysis of habitat-specific virus diversity and ancestral state reconstruction demonstrated an origin of CAVV in a pristine rainforest with subsequent spread into agriculture and human settlements. Virus extension from the forest was associated with a decrease in virus diversity (P < 0.01) and an increase in virus prevalence (P < 0.00001). CAVV is an enveloped virus with large surface projections. The RNA genome comprises 20,108 nucleotides with seven major open reading frames (ORFs). ORF1a and -1b encode two large proteins that share essential features with phylogenetically higher representatives of the order Nidovirales, including the families Coronavirinae and Torovirinae, but also with families in a basal phylogenetic relationship, including the families Roniviridae and Arteriviridae. Genetic markers uniquely conserved in nidoviruses, such as an endoribonuclease- and helicase-associated zinc-binding domain, are conserved in CAVV. ORF2a and -2b are predicted to code for structural proteins S and N, respectively, while ORF3a and -3b encode proteins with membrane-spanning regions. CAVV produces three subgenomic mRNAs with 5′ leader sequences (of different lengths) derived from the 5′ end of the genome. This novel cluster of mosquito-associated nidoviruses is likely to represent a novel family within the order Nidovirales

Re-assessing the diversity of negative strand RNA viruses in insects.

The spectrum of viruses in insects is important for subjects as diverse as public health, veterinary medicine, food production, and biodiversity conservation. The traditional interest in vector-borne diseases of humans and livestock has drawn the attention of virus studies to hematophagous insect species. However, these represent only a tiny fraction of the broad diversity of Hexapoda, the most speciose group of animals. Here, we systematically probed the diversity of negative strand RNA viruses in the largest and most representative collection of insect transcriptomes from samples representing all 34 extant orders of Hexapoda and 3 orders of Entognatha, as well as outgroups, altogether representing 1243 species. Based on profile hidden Markov models we detected 488 viral RNA-directed RNA polymerase (RdRp) sequences with similarity to negative strand RNA viruses. These were identified in members of 324 arthropod species. Selection for length, quality, and uniqueness left 234 sequences for analyses, showing similarity to genomes of viruses classified in Bunyavirales (n = 86), Articulavirales (n = 54), and several orders within Haploviricotina (n = 94). Coding-complete genomes or nearly-complete subgenomic assemblies were obtained in 61 cases. Based on phylogenetic topology and the availability of coding-complete genomes we estimate that at least 20 novel viral genera in seven families need to be defined, only two of them monospecific. Seven additional viral clades emerge when adding sequences from the present study to formerly monospecific lineages, potentially requiring up to seven additional genera. One long sequence may indicate a novel family. For segmented viruses, cophylogenies between genome segments were generally improved by the inclusion of viruses from the present study, suggesting that in silico misassembly of segmented genomes is rare or absent. Contrary to previous assessments, significant virus-host codivergence was identified in major phylogenetic lineages based on two different approaches of codivergence analysis in a hypotheses testing framework. In spite of these additions to the known spectrum of viruses in insects, we caution that basing taxonomic decisions on genome information alone is challenging due to technical uncertainties, such as the inability to prove integrity of complete genome assemblies of segmented viruses

No Evidence of Gouléako and Herbert Virus Infections in Pigs, Côte d’Ivoire and Ghana

A recent report suggested that 2 novel bunyaviruses discovered in insects in Côte d’Ivoire caused lethal disease in swine in South Korea. We conducted cell culture studies and tested serum from pigs exposed to mosquitoes in Côte d’Ivoire and Ghana and found no evidence for infection in pigs

Taxonomy of the family Arenaviridae and the order Bunyavirales : update 2018

In 2018, the family Arenaviridae was expanded by inclusion of 1 new genus and 5 novel species. At the same time, the recently established order Bunyavirales was expanded by 3 species. This article presents the updated taxonomy of the family Arenaviridae and the order Bunyavirales as now accepted by the International Committee on Taxonomy of Viruses (ICTV) and summarizes additional taxonomic proposals that may affect the order in the near future.Peer reviewe

Identifikation und Charakterisierung einer neuen Virusfamilie (<em>Mesoniviridae</em>) und erstmaliger Nachweis der Virusordnung <em>Nidovirales</em> in Insekten

Im Rahmen dieser Promotionsarbeit wurden erstmalig Viren aus der Ordnung Nidovirales in Insekten nachgewiesen, die molekularbiologisch und taxonomisch charakterisiert wurden. In tropischen Moskitos wurden fünf bisher unbekannte Viren entdeckt, die Cavally-Virus (CavV), Hana-Virus (HanaV), Méno-Virus (MénoV), Nsé-Virus (NséV) und Moumo-Virus benannt wurden. CavV, HanaV, MénoV und NséV wurden in Aedes albopictus Insektenzellen isoliert und das Genom der Viren wurde sequenziert. Die Genomgröße der neuen Viren lag mit 20 kb zwischen den „großen“ (Corona- und Roniviridae) und „kleinen“ Nidoviren (Arteriviridae). Phylogenetisch bilden die neuen Viren eine Klade zwischen den Virusfamilien Coronaviridae und den Roniviridae und etablieren eine neue, eigenständige Virusfamilie, die Mesoniviridae genannt wurde. Die umhüllten, mit Glykoproteinen bedeckten Virionen der Mesoniviren sind sphärisch und besitzen einen Durchmesser von ca. 120 nm. Wie bei allen Nidoviren codieren die 5’-terminalen zwei Drittel des positiv-orientierten RNA-Genoms für ein Replikase-Polyprotein, welches durch einen ribosomalen „frameshift“ gebildet wird. Innerhalb des Replikase-Polyproteins befinden sich die funktionellen, konservierten Domänen: drei Transmembrandomänen (TM1, 2 und 3), die eine Chemomotrypsin-ähnliche Serin-Protease (3CLpro) umschließen, gefolgt von der ribosomalen „frameshift“-Stelle (RFS), der RNA-abhängigen RNA-Polymerase (RdRp), dem Zink-Finger ähnlichen Motiv (Z), der 5’-3’-Helikase (Hel), einer 3’-5’-Exoribonuklease (ExoN), einer Guanin-N7-Methyltransferase (NMT) und einer Ribose-2’-O-Methyltransferase (OMT). Das 3’-terminale Drittel des Mesonivirus-Genoms codiert für die drei Strukturproteine („spike“, S; Nukleocapsid, N und Membranprotein, M), die mittels Proteinsequenzierung und Western Blot in den Viruspartikeln nachgewiesen wurden. Das „spike“ Glykoprotein kommt dabei in drei Varianten (S, S1 und S2) und das Membranprotein in verschieden glykosylierten Formen vor. Mesoniviren nutzen für die Generierung ihrer zwei 5’- und 3’-coterminaler, subgenomischer mRNAs einen diskontinuierlichen Transkriptionsmechanismus. Dieser Mechanismus ähnelt dem anderer Nidoviren, wobei Mesoniviren als einzige zwei transkriptionsregulierende Sequenzen verwenden. Antikörper gegen CavV reagieren auch gegen HanaV, MénoV und NséV und es wird vermutet, dass alle Viren vermutlich eigene Spezies darstellen und alle zum Genus Alphamesonivirus gehören. Die Viren konnten in verschiedenen Vertebratenzellen nicht vermehrt werden und gehören damit wahrscheinlich nicht zur Gruppe der durch Arthropoden übertragenen Viren. Diese Arbeit gibt fundamentale Einblicke in gemeinsame und differenzierende biologische Eigenschaften der Mesoniviridae im Vergleich zu anderen Virusfamilien der Ordnung Nidovirales. Zusätzlich wird der mögliche Ursprung der Nidoviren in Arthropoden verdeutlicht. Weiterführende Studien werden helfen, die Ökologie, den Wirtstropismus und eventuell eine medizinische oder ökonomische Relevanz der Mesoniviren zu entschlüsseln

Mosquito community composition shapes virus prevalence patterns along anthropogenic disturbance gradients

Previously unknown pathogens often emerge from primary ecosystems, but there is little knowledge on the mechanism behind. Most studies analyzing the influence of land-use change on pathogen emergence focus on a single host-pathogen system and often observe contrary effects. We studied virus diversity and prevalence patterns in natural and disturbed ecosystems using a multi-host and multi-taxa approach. We detected 331 viral sequences pertaining to 49 viruses of ten RNA-virus families. Highest host and virus diversity was observed in pristine and intermediately disturbed habitats. The majority of the viruses was detected with low prevalence. However, nine viruses were found frequently of which five viruses increased in prevalence from pristine to disturbed habitats, in congruence with the dilution effect hypothesis. Interestingly, the observed increased prevalence of these five viruses in disturbed habitats was not caused by higher host infection rates but by increased host abundance, an effect tentatively named abundance effect

No Evidence of Gouléako and Herbert Virus Infections in Pigs, Côte d’Ivoire and Ghana

A recent report suggested that 2 novel bunyaviruses discovered in insects in Côte d’Ivoire caused lethal disease in swine in South Korea. We conducted cell culture studies and tested serum from pigs exposed to mosquitoes in Côte d’Ivoire and Ghana and found no evidence for infection in pigs

Detection of Two Highly Diverse Peribunyaviruses in Mosquitoes from Palenque, Mexico

The Peribunyaviridae family contains the genera Orthobunyavirus, Herbevirus, Pacuvirus, and Shangavirus. Orthobunyaviruses and pacuviruses are mainly transmitted by blood-feeding insects and infect a variety of vertebrates whereas herbeviruses and shangaviruses have a host range restricted to insects. Here, we tested mosquitoes from a tropical rainforest in Mexico for infections with peribunyaviruses. We identified and characterized two previously unknown viruses, designated Baakal virus (BKAV) and Lakamha virus (LAKV). Sequencing and de novo assembly of the entire BKAV and LAKV genomes revealed that BKAV is an orthobunyavirus and LAKV is likely to belong to a new genus. LAKV was almost equidistant to the established peribunyavirus genera and branched as a deep rooting solitary lineage basal to herbeviruses. Virus isolation attempts of LAKV failed. BKAV is most closely related to the bird-associated orthobunyaviruses Koongol virus and Gamboa virus. BKAV was successfully isolated in mosquito cells but did not replicate in common mammalian cells from various species and organs. Also cells derived from chicken were not susceptible. Interestingly, BKAV can infect cells derived from a duck species that is endemic in the region where the BKAV-positive mosquito was collected. These results suggest a narrow host specificity and maintenance in a mosquito-bird transmission cycle