Untranslated Regions of a Segmented Kindia Tick Virus Genome Are Highly Conserved and Contain Multiple Regulatory Elements for Viral Replication.

Novel segmented tick-borne RNA viruses belonging to the group of Jingmenviruses (JMVs) are widespread across Africa, Asia, Europe, and America. In this work, we obtained whole-genome sequences of two Kindia tick virus (KITV) isolates and performed modeling and the functional annotation of the secondary structure of 5' and 3' UTRs from JMV and KITV viruses. UTRs of various KITV segments are characterized by the following points: (1) the polyadenylated 3' UTR; (2) 5' DAR and 3' DAR motifs; (3) a highly conserved 5'-CACAG-3' pentanucleotide; (4) a binding site of the La protein; (5) multiple UAG sites providing interactions with the MSI1 protein; (6) three homologous sequences in the 5' UTR and 3' UTR of segment 2; (7) the segment 2 3' UTR of a KITV/2017/1 isolate, which comprises two consecutive 40 nucleotide repeats forming a Y-3 structure; (8) a 35-nucleotide deletion in the second repeat of the segment 2 3' UTR of KITV/2018/1 and KITV/2018/2 isolates, leading to a modification of the Y-3 structure; (9) two pseudoknots in the segment 2 3' UTR; (10) the 5' UTR and 3' UTR being represented by patterns of conserved motifs; (11) the 5'-CAAGUG-3' sequence occurring in early UTR hairpins. Thus, we identified regulatory elements in the UTRs of KITV, which are characteristic of orthoflaviviruses. This suggests that they hold functional significance for the replication of JMVs and the evolutionary similarity between orthoflaviviruses and segmented flavi-like viruses.

The Viromes of Mosquitoes from the Natural Landscapes of Western Siberia.

The metagenomic analysis of mosquitoes allows for the genetic characterization of mosquito-associated viruses in different regions of the world. This study applied a metagenomic approach to identify novel viral sequences in seven species of mosquitoes collected from the Novosibirsk region of western Siberia. Using NGS sequencing, we identified 15 coding-complete viral polyproteins (genomes) and 15 viral-like partial sequences in mosquitoes. The complete sequences for novel viruses or the partial sequences of capsid proteins, hypothetical viral proteins, and RdRps were used to identify their taxonomy. The novel viral sequences were classified within the orders Tymovirales and Picornavirales and the families Partitiviridae, Totiviridae, Tombusviridae, Iflaviridae, Nodaviridae, Permutotetraviridae, and Solemoviridae, with several attributed to four unclassified RNA viruses. Interestingly, the novel putative viruses and viral sequences were mainly associated with the mosquito Coquillettidia richardii. This study aimed to increase our understanding of the viral diversity in mosquitoes found in the natural habitats of Siberia, which is characterized by very long, snowy, and cold winters.

The First Case of Zika Virus Disease in Guinea: Description, Virus Isolation, Sequencing, and Seroprevalence in Local Population.

The Zika virus (ZIKV) is a widespread mosquito-borne pathogen. Phylogenetically, two lineages of ZIKV are distinguished: African and Asian-American. The latter became the cause of the 2015-2016 pandemic, with severe consequences for newborns. In West African countries, the African lineage was found, but there is evidence of the emergence of the Asian-American lineage in Cape Verde and Angola. This highlights the need to not only monitor ZIKV but also sequence the isolates. In this article, we present a case report of Zika fever in a pregnant woman from Guinea identified in 2018. Viral RNA was detected through qRT-PCR in a serum sample. In addition, the seroconversion of anti-Zika IgM and IgG antibodies was detected in repeated blood samples. Subsequently, the virus was isolated from the C6/36 cell line. The detected ZIKV belonged to the African lineage, the Nigerian sublineage. The strains with the closest sequences were isolated from mosquitoes in Senegal in 2011 and 2015. In addition, we conducted the serological screening of 116 blood samples collected from patients presenting to the hospital of Faranah with fevers during the period 2018-2021. As a result, it was found that IgM-positive patients were identified each year and that the seroprevalence varied between 5.6% and 17.1%.

Novel Flavi-like virus in ixodid ticks and patients in Russia.

Novel Haseki tick virus (HSTV) was detected in ixodid ticks and patients in the Asian part of Russia. Sequencing of the genome fragments corresponding whole polyprotein and viral RdRp demonstrated that HSTV is genetically close to unclassified Flavi-like viruses. Phylogenetic analysis of HSTV sequences showed that these viruses were close to Bole tick virus 4 (BLTV 4), which was detected early in Asia, Europe, Africa and the Caribbean region. The organization of the genome predicts that HSTV and BLTV 4 may also be classified as putative new genera within Flaviviridae with enlarged Flavi-like positive-sense ssRNA viral genomes. Cases of HSTV putative human incidents after Ixodes persulcatus attack were discovered in hospital patients with tick-borne infections in Vladivostok (Russia). The illness was associated with 3-5 days of fever, accompanied by acute respiratory lesions. Mixed human tick-borne infections (TBIs) were also detected for these patients as dual or triple coinfections for tick-borne encephalitis virus, Borrelia spp., Anaplasma spp., and HSTV. Thus, it is necessary to study HSTV antibody tests, virus isolation, and surveillance for HSTV sequences in different species of ticks, different geographical regions and patients after tick attacks.

Ngari virus (Orthobunyavirus, Peribunyaviridae) in ixodid ticks collected from cattle in Guinea.

Ngari virus is a mosquito-borne virus belonging to the genus Orthobunyavirus (Peribunyaviridae family). This virus is pathogenic to humans and causes severe illness. Ngari virus is present in several African countries, including Madagascar. Here, we report the detection of Ngari virus in ixodid ticks collected from cows in Guinea. A tick survey was conducted in March-November of 2018 in six regions of Guinea. The sample comprised 710 pools, with a total of 2067 ticks belonging to five species collected from 197 cows. At the initial stage, we screened a subsample of tick pools of vector-borne viruses with a multiplex genus-specific primer panel. In the second stage of the study, we narrowed the search and screened all the samples by qPCR for the detection of Ngari virus. All positive samples were sequenced with primers flanking Ngari virus-specific fragments on the S and M segments. We found Ngari virus in 12 pools that were formed from engorged ticks collected from livestock in three villages of the Kindia and Kankan regions. Sequencing of the S and M segments confirmed that the detected viruses belong to Ngari virus, and the viruses were most similar to the strain Adrar, which was isolated in Mauritania. We detected viral RNA in ticks of the following species: Amblyomma variegatum, Rhipicephalus geigyi, and Rh. (Boophilus) spp. There is no evidence that ixodid ticks are competent vectors of the Ngari virus. Most likely, the ticks obtained the virus through blood from an infected host. The study of engorged ticks can be recommended as a simpler approach for the wide screening of the Ngari virus and subsequent testing of cattle and mosquitos in those locations where the PCR-positive ticks were collected.

Variability in the 3' untranslated regions of the genomes of the different tick-borne encephalitis virus subtypes.

Tick-borne encephalitis viruses (TBEVs) are usually divided into three major subtypes: European (TBEV-Eu), Siberian (TBEV-Sib) and Far Eastern (TBEV-FE). The TBEV-Eu strains have the longest genomes, and TBEV-FE strains have the smallest genomes. Changes in the variable region of the untranslated region (V3' UTR) play a major role in determining the viral genome length. Analyses of the 3' UTRs of the different subtypes of TBEV have revealed significant changes in the secondary structures of the V3' UTR of TBEV. More complex secondary structures of the V3' UTR regions are typical for TBEV-Eu. The Siberian strain Tomsk-PT122 was isolated from birds and has an unusual 3' UTR. Several short fragment (24-26 nucleotides) insertions derived from the viral E (2) and NS4a (1) genes have been found in the V3' UTR of Tomsk-PT122. Additionally, the length of the V3' UTR increases from 21 to 37 nucleotides during passages of the C11-13 strain of TBEV-Sib into PEK, 293 and Neuro-2a cells. The elongation of the V3' UTRs of Tomsk-PT122 and C11-13 is the first direct evidence of an intragenomic 3' UTR modification (insertion) for TBEV. Thus, the obtained results suggest that changing the length of the V3' UTR in the genome is typical for different TBEV subtypes and can play an essential role in effective TBEV replication in different host cells.

Adaptation of tick-borne encephalitis virus from human brain to different cell cultures induces multiple genomic substitutions.

The C11-13 strain from the Siberian subtype of tick-borne encephalitis virus (TBEV) was isolated from human brain using pig embryo kidney (PEK), 293, and Neuro-2a cells. Analysis of the complete viral genome of the C11-13 variants during six passages in these cells revealed that the cell-adapted C11-13 variants had multiple amino acid substitutions as compared to TBEV from human brain. Seven out of eight amino acids substitutions in the high-replicating C11-13(PEK) variant mapped to non-structural proteins; 13 out of 14 substitutions in the well-replicating C11-13(293) variant, and all four substitutions in the low-replicating C11-13(Neuro-2a) variant were also localized in non-structural proteins, predominantly in the NS2a (2), NS3 (6) and NS5 (3) proteins. The substitutions NS2a1067 (Asn → Asp), NS2a1168(Leu → Val) in the N-terminus of NS2a and NS31745(His → Gln) in the helicase domain of NS3 were found in all selected variants. We postulate that multiple substitutions in the NS2a, NS3 and NS5 genes play a key role in adaptation of TBEV to different cells.