μSTRAW: In-Field Detection of <i>Escherichia coli</i>

Fast detection of <i>Escherichia coli</i> is required in multiple OneHealth settings, from bathing water to urinary tract infections (urine) and septicemia (i.e., blood infections). Current methods for bacteria enumeration are limited to lab-based conventional microbiology, requiring 24-48 h. Nucleic acid testing represents an alternative, with high specificity and a broad range of targets. However, a scalable and affordable method for portable DNA extraction is missing, limiting point-of-care applications. To address that gap, we introduced μSTRAW, a compact gravity-driven microfluidic device for portable DNA extraction, which directly interfaces with a commercial, portable LAMP instrument. Using 45 mm-long, 10-bored 200 μm i.d. microcapillary strips and a magnetic beads (MB)-based extraction kit, MB were captured and efficiently washed before DNA elution. μSTRAW offered simultaneous detection from 8 samples, and the usage of MB, lysis buffer, and proteinase K was reduced by 91, 83, and 65% compared to the manufacturer's protocol. Integration with portable LAMP showed a low limit of detection (LLoD) in the range of 1000-5000 CFU/mL in PBS within 120-140 min, which improved to 50-100 CFU/mL using a pre-concentration with a 0.22 μm-pore-size syringe filter. μSTRAW also yielded an LLoD of 100-1000 CFU/mL in synthetic urine, whereas in sheep blood, we observed an LLoD as low as 10-23 CFU/mL upon 5 h of incubation in LB broth, which was therefore within the common CFU/mL found for bloodstream infections. The proposed approach is portable, affordable, scalable, and potentially transformative for a range of global OneHealth applications, especially bathing waters and wastewater.