| Title: |
Rapid Nanopore Sequencing of Positive Blood Cultures Using Automated Benzyl-Alcohol Extraction Improves Time-Critical Sepsis Management |
| Authors: |
Chi-Sheng Tai; Hsing-Yi Chung; Tai-Han Lin; Chih-Kai Chang; Cherng-Lih Perng; Po-Shiuan Hsieh; Hung-Sheng Shang; Ming-Jr Jian |
| Source: |
Antibiotics ; Volume 14 ; Issue 10 ; Pages: 1001 |
| Publisher Information: |
Multidisciplinary Digital Publishing Institute |
| Publication Year: |
2025 |
| Collection: |
MDPI Open Access Publishing |
| Subject Terms: |
pathogen detection; sepsis; benzyl alcohol extraction; automation; Oxford Nanopore |
| Description: |
Background/Objective: Timely identification of bloodstream pathogens is critical for sepsis management; however, PCR inhibitors such as sodium polyanetholesulfonate (SPS) in blood culture broth compromise nucleic acid recovery and long read sequencing. We assessed whether coupling a benzyl alcohol SPS-removal step to the fully automated LabTurbo AIO extractor improves Oxford Nanopore-based pathogen detection. Methods: Thirteen positive blood culture broths were pre-treated with benzyl alcohol and divided: half volumes were purified on the LabTurbo AIO; paired aliquots underwent manual QIAamp extraction. DNA purity was evaluated by NanoDrop and Qubit. Barcoded libraries were sequenced on MinION R9.4.1 flow cells for 6 h. Results: Automated eluates showed a median A260/A280 of 1.92 and A260/A230 of 1.96, versus 1.80 and 1.48 for manual extracts. The automated workflow generated 1.69 × 106 total reads compared with 3.9 × 105 reads for manual extraction. The median N50 read length increased from 5.9 kb to 8.7 kb, and the median proportion of reads classified to species increased from 62% to 84%. The hands-on time was |
| Document Type: |
text |
| File Description: |
application/pdf |
| Language: |
English |
| Relation: |
https://dx.doi.org/10.3390/antibiotics14101001 |
| DOI: |
10.3390/antibiotics14101001 |
| Availability: |
https://doi.org/10.3390/antibiotics14101001 |
| Rights: |
https://creativecommons.org/licenses/by/4.0/ |
| Accession Number: |
edsbas.B4953FA |
| Database: |
BASE |