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Metagenomic nanopore sequencing using MinION to identify bacterial species and genes conferring antimicrobial resistance for early sepsis diagnosis using spiked human blood samples
University of Skövde, School of Bioscience.
2025 (English)Independent thesis Basic level (degree of Bachelor), 20 credits / 30 HE creditsStudent thesis
Abstract [en]

Sepsis is one of the life-threatening illnesses defined by tissue damage and organ failure caused by a dysregulated host response to infection. Blood culture is the golden standard and the first-line tool for detecting infections. It takes several days to identify pathogens, their sensitivity, and resistance to antibiotics. Early diagnosis of sepsis is thereby crucial to improving the patient's life. This study evaluates metagenomic sequencing using MinION and Flongle flow cells to detect bacterial species and antibiotic resistance genes. The DNA was extracted from the spiked whole human blood from a healthy individual with microbial DNA. This project focused on evaluation of the amount of spike-in to be used in the whole human blood, bacterial identification, and the detection of antimicrobial genes that confer antibiotic resistance. The obtained data from the nanopore sequencing was analysed using EPI2ME. The further analysis was performed using the BV-BRC taxonomic classification and metagenomic read mapping. The higher DNA concentration and consistent absorbance values were obtained from both DNA extraction kits. The results from nanopore sequencing showed the difference in classified reads and average sequence length depending on the DNA extraction kit as well as the flow cell used. The difference in the bacterial identification and antimicrobial genes identification could also be seen between the flow cells used. The results indicated that nanopore sequencing with the MinION flow cell could be more valuable in the early diagnosis of sepsis, but further research is required to make it more efficient and faster for sepsis treatment. Improving the detection of gram-positive bacteria and optimizing DNA extraction and nanopore sequencing protocols is needed for rapid and accurate identification of pathogens in the early stages of sepsis.

Place, publisher, year, edition, pages
2025. , p. 39
National Category
Infectious Medicine
Identifiers
URN: urn:nbn:se:his:diva-25561OAI: oai:DiVA.org:his-25561DiVA, id: diva2:1985323
Subject / course
Bioscience
Educational program
Molekylär biodesign 180 hp
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Examiners
Available from: 2025-07-23 Created: 2025-07-23 Last updated: 2025-09-29Bibliographically approved

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CiteExportLink to record
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