Application of bioinformatics methods to identify viral infections with impact on hemotherapy

Author(s)

Rafael dos Santos Bezerra

• Document type: Master's Dissertation
• Press: Ribeirão Preto.
• Institution: Universidade de São Paulo (USP). Faculdade de Medicina de Ribeirão Preto (PCARP/BC)
• Defense date: 2021-07-16

Examining board members

Svetoslav Nanev Slavov; Antonio Charlys da Costa; José Eduardo Levi

• Advisor: Svetoslav Nanev Slavov

Abstract

With the introduction in the laboratory practice of new sequencing technologies, like second and third generation techniques, the time and cost of the sequencing has drastically reduced while the generated data has increased significantly. In this way, the field of application of these new techniques has expanded incliding analysis of the microbial communities of clinical and environmental samples known as metagenomics. Virome is an important area of metagenomics where the focus is concentrated on the viruses found in these samples. Among the possible applications of virome analysis is the identification of emerging and re-emerging viruses which can impact the area of hemotherapy. To investigate these agents, we applied virome techniques in blood units obtained from donors who related with Post-Donation Disease Reports (PDDR) (including symptoms like fever, rash, conjunctivitis, retro-orbital pain, arthralgia, myalgia, diarrhea, vomiting, and nausea among others). We also applied viral metagenomics to blood donations obtained from high-risk blood donors from Northern and Southern Brazil. To do this, a high-performance bioinformatics pipeline was implemented to identify emerging and re- emerging viruses. The implementation was carried out in BASH script, DOCKER (containers) and in-house developed PERL scripts. The main stages of the pipeline include quality control of the sequences with the FastQC software, cleaning of low-quality sequences and poly-x tails by Trimmomatic and AfterQC, genome assembly by SPAdes software and finally sequence taxonomy using Kraken2 and Diamond. The phylodynamic analyzes of the most important viruses found were performed using the IQ-TREE software and the BEAST package. Among our results from PDDR samples we can highlight the first report in the literature of the Influenza A virus (H2N3) in plasma samples from blood donors. We also identified the dengue virus and Parvovirus B19 (both not included in the route testing of the blood collection services). Interestingly, in samples from North Brazil, we identified in high-risk blood donors the scarcely studied studied Human Gemykibivirus-2, which has already been related to severe clinical conditions and HIV co-infections. Another interesting finding in these samples was the Merkell cell polyomavirus with high sequence numbers (which may indicate a viremic state). In Brazil, comprehensive studies on infections which can impact transfusion safety have not been performed and the real impact of the emerging viruses on the hemotherapy processes is underestimated and little known. However, some challenges in this area must be overcame, and among them we can mention the bioinformatics analysis of the generated data, which often requires high computational resources and specialized labor. Therefore, our study examines in extensive manner the impact of emerging viruses in the area of hemotherapy using the newest sequencing generation and implementing an efficient bioinformatics pipeline for virome analysis. (AU)

• FAPESP's process: 19/08528-0 - Application of bioinformatic procedures for identification of the impact of the viral infections in hemotherapy
• Grantee: Rafael dos Santos Bezerra
• Support Opportunities: Scholarships in Brazil - Master