# Viromes of two migratory birds as support for identifying the influence of molecular biology and bioinformatics settings

S. Tirera, A. Salmier, D. Donato, Jean-Marc Frigério, S. Thérond, V. Lacoste, B. de Thoisy, Alain Franc, A. Lavergne

► To cite this version:

S. Tirera, A. Salmier, D. Donato, Jean-Marc Frigério, S. Thérond, et al.. Viromes of two migratory birds as support for identifying the influence of molecular biology and bioinformatics settings. 23. International BioInformatics Workshop on Virus Evolution and Molecular Epidemiology, Aug 2018, Berlin, Germany. 10.1093/ve/vez002.069. hal-02735266

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**A70** Viromes of two migratory birds as support for identifying the influence of molecular biology and bioinformatics settings

S. Tirera,¹ A. Salmier,¹ D. Donato,¹ J. M. Frigerio,² S. Therond,³
V. Lacoste,¹ B. de Thoisy,¹ A. Franc,² and A. Lavergne¹

¹LIVH, Institut Pasteur de la Guyane, Cayenne, French Guiana,

²INRA, UMR BioGeCo, Cestas, France and ³CNRS - IDRIS, Orsay, France

Thanks to high-throughput sequencing (HTS) and dedicated
bioinformatics tools, it is possible to study all viruses present in a
sample with an unprecedented detection of viral diversity. Applied
to wildlife and humans, HTS becomes a powerful means to
understand viral emerging infectious diseases. BLAST searches to
find remote homolog sequences after an assembly of shotgun-
generated sequences are often used to analyze virome datasets.
However, this process might be biased by molecular biology
approaches (use of whole genome/transcriptome amplification)
and bioinformatics assembly that can induce chimeric sequences
formation. Our objective was to identify confounding factors
(molecular biology and bioinformatics) that can bias virome
composition. Hence, we studied the viromes from cloacal swab
samples of two migratory birds. Viral DNA and RNA were
extracted using three different methods (easymag, phenol, and

trizol), followed by whole genome amplification (WGA)/WTA
amplifications, before Illumina MiSeq sequencing. After trimming
and assembly, sequences underwent BLAST (BLASTn and BLASTx).
At the end of the first BLAST (n/x), some portions of sequences
were uncovered and unassigned. We, then, implemented a novel
recursive split-resubmit python program that searched for
homologs of > 50 base pairs (bp), uncovering parts for a better
exploitation of the datasets. Thirty-eight known viral families
were detected in our samples. Circoviridae, Parvoviridae, and
Microviridae were found in the majority of the results from
easymag and Phenol extracted samples, while trizol samples
resulted in a majority of Picornaviridae and Coronaviridae. These
results show that the viromes are of different composition
depending on extraction methods. Most of the viral sequences
were predominantly identified at the blastx step reflecting a high
level of divergence with known viral sequences. Compared to a
unique BLAST search, our resubmission pipeline allowed the
assignment of up to 23 per cent of the viral sequences. Most of
these sequences belonged to taxa already observed at the first
blast step and few viral sequences of different families were
detected. Knowing the complementarities between extraction
methods and a better identification of viral sequences using our
pipeline, the next step will be to get a precise position of each viral
genomic segment in phylogenetic distance-based tree that will
give a more accurate representation of viral diversity.