Smartphone-based pathogen diagnosis in urinary sepsis patients

Abstract

Background: There is an urgent need for rapid, sensitive, and affordable diagnostics for microbial infections at the point-of-care. Although a number of innovative systems have been reported that transform mobile phones into potential diagnostic tools, the translational challenge to clinical diagnostics remains a significant hurdle to overcome.

Methods: A smartphone-based real-time loop-mediated isothermal amplification (smaRT-LAMP) system was developed for pathogen ID in urinary sepsis patients. The free, custom-built mobile phone app allows the phone to serve as a stand-alone device for quantitative diagnostics, allowing the determination of genome copy-number of bacterial pathogens in real time.

Findings: A head-to-head comparative bacterial analysis of urine from sepsis patients revealed that the performance of smaRT-LAMP matched that of clinical diagnostics at the admitting hospital in a fraction of the time (~1 h vs. 18-28 h). Among patients with bacteremic complications of their urinary sepsis, pathogen ID from the urine matched that from the blood - potentially allowing pathogen diagnosis shortly after hospital admission. Additionally, smaRT-LAMP did not exhibit false positives in sepsis patients with clinically negative urine cultures.

Interpretation: The smaRT-LAMP system is effective against diverse Gram-negative and -positive pathogens and biological specimens, costs less than $100 US to fabricate (in addition to the smartphone), and is configurable for the simultaneous detection of multiple pathogens. SmaRT-LAMP thus offers the potential to deliver rapid diagnosis and treatment of urinary tract infections and urinary sepsis with a simple test that can be performed at low cost at the point-of-care. FUND: National Institutes of Health, Chan-Zuckerberg Biohub, Bill and Melinda Gates Foundation.

Keywords: Smartphone-based pathogen diagnosis; Urinary diagnostic test; Urinary sepsis; Urinary tract infection.

Graphical abstract

Fig. 1

SmaRT-LAMP direct specimen testing of urine from sepsis patients. (a) Assay schematic for smaRT-LAMP, which entails sample collection, bacterial cell lysis/reagent addition, and real-time analysis via the smartphone. (b) Schematic of workflow for the Bacticount app, which analyzes fluorescence data collected continuously from multiple samples through the phone's camera (left panel), and then uses these data to automatically determine the genome copy-number of bacterial pathogens in real time (right panel).

Fig. 2

SmaRT-LAMP quantification of ST with performance equivalent to a benchtop laboratory qPCR instrument. (a–d) Normalized representative traces and T_t values of ST CFU in buffer at concentrations of 10¹⁻10⁵ CFU/mL using qPCR-LAMP and smaRT-LAMP; (e, f), Percentage of total samples amplified at each concentration using qPCR-LAMP or smaRT-LAMP (21 samples/concentration).

Fig. 3

SmaRT-LAMP quantification of ST in spiked diverse biological specimens. (a–f) T_t values for ST CFU in murine blood, urine, and feces using qPCR-LAMP and smaRT-LAMP. (g, h) corresponding representative traces (2–2 × 10⁴ CFU/reaction); NC, no cell. (I, J) Percentage of total pathogen samples amplifying at each concentration using smaRT-LAMP.

Fig. 4

SmaRT-LAMP quantitation of diverse pathogens in spiked murine whole blood and human donor urine. (a–e) Tt values for SPN, SE, EC, YP (2–2 × 10⁴ CFU/reaction); SA (4 × 10¹–2 × 10⁴ CFU/reaction). (f) Percentage of total pathogen samples amplifying at each concentration in smaRT-LAMP. (g, h) Representative traces and T_t values for EC in spiked human donor urine (2–2 × 10⁴ CFU/reaction). (i) Percentage of total EC samples amplifying at each concentration in smaRT-LAMP (≥ 10 samples/concentration).

Fig. 5

SmaRT-LAMP detection and quantitation of Salmonella in whole blood of septic mice. (a–c). Mice were orally infected with ST via gastric intubation at a dose of 2 × 10⁷ cells and whole blood was sampled at days 6 (pre-sepsis), 8 (sepsis), and 10 (severe sepsis) post-infection. CFU were determined by qPCR-LAMP (closed boxes), smaRT-LAMP (open boxes), and direct colony count (circles). n = 14 mice.