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. 2024 Dec:350:199464.
doi: 10.1016/j.virusres.2024.199464. Epub 2024 Sep 20.

Development of targeted whole genome sequencing approaches for Crimean-Congo haemorrhagic fever virus (CCHFV)

Jake D'Addiego  1 , Sonal Shah  2 , Ayşe Nur Pektaş  3 , Bi Nnur Köksal Bağci  4 , Murtaza Öz  5 , Sasha Sebastianelli  6 , Nazif Elaldı  7 , David J Allen  8 , Roger Hewson  9
Affiliations

Development of targeted whole genome sequencing approaches for Crimean-Congo haemorrhagic fever virus (CCHFV)

Jake D'Addiego et al. Virus Res. 2024 Dec.

Abstract

Crimean-Congo haemorrhagic fever (CCHF) is the most prevalent human tick-borne viral disease, with a reported case fatality rate of 30 % or higher. The virus contains a tri-segmented, negative-sense RNA genome consisting of the small (S), medium (M) and large (L) segments encoding respectively the nucleoprotein (NP), the glycoproteins precursor (GPC) and the viral RNA-dependent RNA polymerase (RDRP). CCHFV is one of the most genetically diverse arboviruses, with seven distinct lineages named after the region they were first reported in and based on S segment phylogenetic analysis. Due to the high genetic divergence of the virus, a single targeted tiling PCR strategy to enrich for viral nucleic acids prior to sequencing is difficult to develop, and previously we have developed and validated a tiling PCR enrichment method for the Europe 1 genetic lineage. We have developed a targeted, probe hybridisation capture method and validated its performance on clinical as well as cell-cultured material of CCHFV from different genetic lineages, including Europe 1, Europe 2, Africa 2 and Africa 3. The method produced over 95 % reference coverages with at least 10x sequencing depth. While we were only able to recover a single complete genome sequence from the tested Europe 1 clinical samples with the capture hybridisation protocol, the data provides evidence of its applicability to different CCHFV genetic lineages. CCHFV is an important tick-borne human pathogen with wide geographical distribution. Environmental as well as anthropogenic factors are causing increased CCHFV transmission. Development of strategies to recover CCHFV sequences from genetically diverse lineages of the virus is of paramount importance to monitor the presence of the virus in new areas, and in public health responses for CCHFV molecular surveillance to rapidly detect, diagnose and characterise currently circulating strains.

Keywords: CCHFV; Next-generation sequencing, Targeted enrichment; Probe hybridisation capture.

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Conflict of interest statement

Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Figures

Fig. 1
Fig. 1
RNA copies/ml of serum (black dots) and time to sampling (days, grey line) from patients' symptoms onset.
Fig. 2
Fig. 2
Percentage of mapped sequencing reads over total reads (thousands) for (A) tiling PCR enrichment and (B) probe hybridization capture. Genetic segments are colour-coded in blue (S), orange (M) and grey (L). Circular markers represent clinical samples; triangular markers represent cell-cultured viruses.
Fig. 3
Fig. 3
Reference coverage and sequencing depth for (A) probe hybridisation capture and (B) tiling PCR enrichment with Illumina sequencing technology on a CCHFV Europe 1 positive clinical serum sample (sample 4).
Fig. 4
Fig. 4
Reference coverages (%) for the S, M and L segments at 10X and 100X sequencing depths for tiling PCR (blue) and hybridisation capture (orange) on a CCHFV Europe 1 positive clinical sample (sample 4).
Fig. 5
Fig. 5
Maximum Likelihood phylogeny of the S, M and L CCHFV partial genome segments. Enrichment strategy in the generated consensus name is indicated by “amp” and “hyb” for the tiling PCR and capture hybridisation respectively.
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References

    1. Arab-Bafrani Z., Jabbari A., Mostakhdem H.M, et al. Identification of the crucial parameters regarding the efficacy of ribavirin therapy in Crimean-Congo haemorrhagic fever (CCHF) patients: a systematic review and meta-analysis. J. Antimicrob. Chemother. 2019;74(12):3432–3439. doi: 10.1093/jac/dkz328. [published Online First: 2019/08/02] - DOI - PubMed
    1. Au C.H., Ho D.N., Kwong A., et al. BAMClipper: removing primers from alignments to minimize false-negative mutations in amplicon next-generation sequencing. Sci. Rep. 2017;7(1):1567. doi: 10.1038/s41598-017-01703-6. [published Online First: 2017/05/10] - DOI - PMC - PubMed
    1. Bente D.A., Forrester N.L., Watts D.M., et al. Crimean-Congo hemorrhagic fever: history, epidemiology, pathogenesis, clinical syndrome and genetic diversity. Antiviral Res. 2013;100(1):159–189. doi: 10.1016/j.antiviral.2013.07.006. [published Online First: 2013/08/03] - DOI - PubMed
    1. Bernard C., Joly K.C, Rakotoarivony I., et al. Detection of Crimean-Congo haemorrhagic fever virus in Hyalomma marginatum ticks, southern France, May 2022 and April 2023. Euro Surveill. 2024;29(6) doi: 10.2807/1560-7917.ES.2024.29.6.2400023. [published Online First: 2024/02/09] - DOI - PMC - PubMed
    1. Bolger A.M., Lohse M., Usadel B. Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics. 2014;30(15):2114–2120. doi: 10.1093/bioinformatics/btu170. [published Online First: 2014/04/04] - DOI - PMC - PubMed

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