A systematic, deep sequencing-based methodology for identification of mixed-genotype hepatitis C virus infections

Olmstead, Andrea D.; Montoya, Vincent; Chui, Celia; Dong, Winnie; Joy, Jeffrey B.; Tai, Vera; Poon, Art F. Y.; Nguyen, Thuy; Brumme, Chanson J.; Martinello, Marianne; Matthews, Gail V.; Harrigan, P. Richard; Dore, Gregory J.; Applegate, Tanya; Grebely, Jason; Howe, Anita Y.M.

doi:10.1016/j.meegid.2019.01.016

Cited by 7 publications

(5 citation statements)

References 49 publications

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“…NGS identified novel subtypes in 9/1038 (0.9%) samples. The low frequency of recombinants and mixtures (3/1038, 0.3% each) confirms findings in the literature [15][16][17][18].…”

Section: Discussionsupporting

confidence: 90%

Clinical evaluation of a Hepatitis C Virus whole-genome sequencing pipeline for genotyping and resistance testing

Bradshaw

Bibby

Manso

et al. 2022

Clinical Microbiology and Infection

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This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

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“…NGS identified novel subtypes in 9/1038 (0.9%) samples. The low frequency of recombinants and mixtures (3/1038, 0.3% each) confirms findings in the literature [15][16][17][18].…”

Section: Discussionsupporting

confidence: 90%

Clinical evaluation of a Hepatitis C Virus whole-genome sequencing pipeline for genotyping and resistance testing

Bradshaw

Bibby

Manso

et al. 2022

Clinical Microbiology and Infection

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“…It should be mentioned that with standard PCR and Sanger sequencing, parasites at much higher parasitemia (quantity) may be amplified preferentially, masking the fact that other parasites are present. In order to overcome this problem in future studies, deep amplicon sequencing would provide a possibility to accurately determine the percentage of two or more pathogen lineages in samples with mixed infections (Olmstead et al 2019).…”

Section: Discussionmentioning

confidence: 99%

High prevalence and genetic diversity of Haemoproteus columbae (Haemosporida: Haemoproteidae) in feral pigeons Columba livia in Cape Town, South Africa

et al. 2019

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In this study, we explore blood parasite prevalence, infection intensity, and co-infection levels in an urban population of feral pigeons Columba livia in Cape Town. We analyze the effect of blood parasites on host body condition and the association between melanin expression in the host's plumage and parasite infection intensity and co-infection levels. Relating to the haemosporidian parasite itself, we study their genetic diversity by means of DNA barcoding (cytochrome b) and show the geographic and host distribution of related parasite lineages in pigeons worldwide. Blood from 195 C. livia individuals was collected from April to June 2018. Morphometric measurements and plumage melanism were recorded from every captured bird. Haemosporidian prevalence and infection intensity were determined by screening blood smears and parasite lineages by DNA sequencing. Prevalence of Haemoproteus spp. was high at 96.9%. The body condition of the hosts was negatively associated with infection intensity. However, infection intensity was unrelated to plumage melanism. The cytochrome b sequences revealed the presence of four Haemoproteus lineages in our population of pigeons, which show high levels of co-occurrence within individual birds. Three lineages (HAECOL1, COLIV03, COQUI05) belong to Haemoproteus columbae and differ only by 0.1% to 0.8% in the cytochrome b gene. Another lineage (COLIV06) differs by 8.3% from the latter ones and is not linked to a morphospecies, yet. No parasites of the genera Leucocytozoon and Plasmodium were detected.

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“…A recent study comparing three different deep sequencing methods was able to determine the presence of mixed infections in 1 out of 143 samples analyzed, even when many reads detected at 1–2% mapped to different genotypes. However, most of those reads did not pass the mixed-infection cut-off criteria [ 42 ]. Our NGS results indeed showed nucleotide changes present in very low frequencies (1–1.9%), which were later confirmed by cloning (sample 1021, see Figure 1 ), which highlights the importance of combining both methods for accurately detecting mixed infections characterized by a large disparity in the abundance of the major and minor genotypes.…”

Section: Discussionmentioning

confidence: 99%

“…Next-generation sequencing methods offer a promising alternative to traditional sequencing approaches in the study of viral genetic diversity. These advanced techniques provide a comprehensive view of viral populations, enabling the detection of mixed infections and the identification of recombinant variants, even when present as minority or low-frequency variants [ 39 , 40 , 41 , 42 ].…”

Section: Introductionmentioning

confidence: 99%

Mixed Infections Unravel Novel HCV Inter-Genotypic Recombinant Forms within the Conserved IRES Region

Echeverría,

Gámbaro,

Beaucourt

et al. 2024

Viruses

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Hepatitis C virus (HCV) remains a significant global health challenge, affecting millions of people worldwide, with chronic infection a persistent threat. Despite the advent of direct-acting antivirals (DAAs), challenges in diagnosis and treatment remain, compounded by the lack of an effective vaccine. The HCV genome, characterized by high genetic variability, consists of eight distinct genotypes and over ninety subtypes, underscoring the complex dynamics of the virus within infected individuals. This study delves into the intriguing realm of HCV genetic diversity, specifically exploring the phenomenon of mixed infections and the subsequent detection of recombinant forms within the conserved internal ribosome entry site (IRES) region. Previous studies have identified recombination as a rare event in HCV. However, our findings challenge this notion by providing the first evidence of 1a/3a (and vice versa) inter-genotypic recombination within the conserved IRES region. Utilizing advanced sequencing methods, such as deep sequencing and molecular cloning, our study reveals mixed infections involving genotypes 1a and 3a. This comprehensive approach not only confirmed the presence of mixed infections, but also identified the existence of recombinant forms not previously seen in the IRES region. The recombinant sequences, although present as low-frequency variants, open new avenues for understanding HCV evolution and adaptation.

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A systematic, deep sequencing-based methodology for identification of mixed-genotype hepatitis C virus infections

Cited by 7 publications

References 49 publications

Clinical evaluation of a Hepatitis C Virus whole-genome sequencing pipeline for genotyping and resistance testing

Clinical evaluation of a Hepatitis C Virus whole-genome sequencing pipeline for genotyping and resistance testing

High prevalence and genetic diversity of Haemoproteus columbae (Haemosporida: Haemoproteidae) in feral pigeons Columba livia in Cape Town, South Africa

Mixed Infections Unravel Novel HCV Inter-Genotypic Recombinant Forms within the Conserved IRES Region

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