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. 2022 Aug 31;14(9):1933.
doi: 10.3390/v14091933.

Viral Metagenomics for the Identification of Emerging Infections in Clinical Samples with Inconclusive Dengue, Zika, and Chikungunya Viral Amplification

Juliana Vanessa Cavalcante Souza  1 , Hazerral de Oliveira Santos  1 , Anderson Brandão Leite  1   2 , Marta Giovanetti  3   4 , Rafael Dos Santos Bezerra  5 , Eneas de Carvalho  6 , Jardelina de Souza Todão Bernardino  6 , Vincent Louis Viala  6 , Rodrigo Haddad  7 , Massimo Ciccozzi  8 , Luiz Carlos Junior Alcantara  4 , Sandra Coccuzzo Sampaio  6 , Dimas Tadeu Covas  5   6 , Simone Kashima  5 , Maria Carolina Elias  6 , Svetoslav Nanev Slavov  5   6
Affiliations

Viral Metagenomics for the Identification of Emerging Infections in Clinical Samples with Inconclusive Dengue, Zika, and Chikungunya Viral Amplification

Juliana Vanessa Cavalcante Souza et al. Viruses. .

Abstract

Viral metagenomics is increasingly being used for the identification of emerging and re-emerging viral pathogens in clinical samples with unknown etiology. The objective of this study was to shield light on the metavirome composition in clinical samples obtained from patients with clinical history compatible with an arboviral infection, but that presented inconclusive results when tested using RT-qPCR. The inconclusive amplification results might be an indication of the presence of an emerging arboviral agent that is inefficiently amplified by conventional PCR techniques. A total of eight serum samples with inconclusive amplification results for the routinely tested arboviruses-dengue (DENV), Zika (ZIKV), and Chikungunya (CHIKV) obtained during DENV and CHIKV outbreaks registered in the state of Alagoas, Northeast Brazil between July and August 2021-were submitted to metagenomic next-generation sequencing assay using NextSeq 2000 and bioinformatic pipeline for viral discovery. The performed bioinformatic analysis revealed the presence of two arboviruses: DENV type 2 (DENV-2) and CHIKV with a high genome coverage. Further, the metavirome of those samples revealed the presence of multiple commensal viruses apparently without clinical significance. The phylogenetic analysis demonstrated that the DENV-2 genome belonged to the Asian/American genotype and clustered with other Brazilian strains. The identified CHIKV genome was taxonomically assigned as ECSA genotype, which is circulating in Brazil. Together, our results reinforce the utility of metagenomics as a valuable tool for viral identification in samples with inconclusive arboviral amplification. Viral metagenomics is one of the most potent methods for the identification of emerging arboviruses.

Keywords: Brazil; Chikungunya; RT-qPCR; dengue; emerging arboviruses; genotypes; metagenomics; virome.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Area of investigation and metagenomic analysis of the analyzed pool of samples. (A) Map representing the spatial area under investigation and position of state of Alagoas in Brazil. (B) Bar-plots illustrating the relative abundance of viral reads for the most presented viral agents in the metavirome of the tested patients with predominance of Chikungunya and Dengue viral reads and other commensal viruses, forming part of the normal metavirome. (C) Circular representation of the relative viral abundance; Chikungunya and Dengue viruses are highly present.
Figure 2
Figure 2
Phylogenetic analysis of the obtained complete Dengue and Chikungunya complete genomes. (A) Maximum likelihood tree showing that the new Dengue virus-2 genome sequences obtained in this study belonged to the Asian/American genotype and clustered together with other strains obtained from Brazil during the 2019–2020 DENV outbreak, especially strains that circulated in the São Paulo state. (B) Maximum likelihood tree analyzing complete Chikungunya genome sequence indicating that it belongs to the ECSA genotype, which is circulating in Brazil and especially the northeast, where the sample collection was performed.
See this image and copyright information in PMC

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