Abstract
Sepsis is a serious bloodstream infection where the immunity of the host body is compromised, leading to organ failure and death of the patient. In early sepsis, the concentration of bacteria is very low and the time of diagnosis is very critical since mortality increases exponentially with every hour after infection. Common culture-based methods fail in fast bacteria determination, while recent rapid diagnostic methods are expensive and prone to false positives. In this work, we present a sepsis kit for fast detection of bacteria in whole blood, here achieved by combining selective cell lysis and a sensitive colorimetric approach detecting as low as 103 CFU/mL bacteria in less than 5 h. Homemade selective cell lysis buffer (combination of saponin and sodium cholate) allows fast processing of whole blood in 5 min while maintaining bacteria alive (100% viability). After filtration, retained bacteria on filter paper are incubated under constant illumination with the electrochromic precursors, i.e., ferricyanide and ferric ammonium citrate. Viable bacteria metabolically reduce iron(III) complexes, initiating a photocatalytic cascade toward Prussian blue formation. As a proof of concept, we combine this method with antibiotic susceptibility testing to determine the minimum inhibitory concentration (MIC) using two antibiotics (ampicillin and gentamicin). Although this kit is used to demonstrate its applicability to sepsis, this approach is expected to impact other key sectors such as hygiene evaluation, microbial contaminated food/beverage, or UTI, among others.
Highlights
Sepsis is a serious bloodstream infection where the immunity of the host body is compromised, leading to organ failure and death of the patient
We present a novel sepsis kit, where a selective cell lysis-based sample preparation method is combined with a highly sensitive photochemical reaction capable of reporting the presence of bacterial infection in blood samples within 5 h for 103 colony forming units (CFU)/mL bacterial concentration through a simple color change
We demonstrate this by initially characterizing the selective cell lysis buffer to rupture blood cells in 5 to 10 min, without compromising bacterial integrity and activity (i.e,. 100% viable)
Summary
Sepsis is a serious bloodstream infection where the immunity of the host body is compromised, leading to organ failure and death of the patient. 6 million people die of sepsis in the world every year, 500 thousand of them are newborn kids, and this condition is responsible for 1 in 10 maternal deaths This high mortality is mostly associated with the difficulty of diagnosing sepsis at its early stages: initial bacterial concentration does not exceed 100 colony forming units (CFU)/mL3 and every hour of delay in the diagnostic and treatment increases the mortality of patients up to 10%.4. Better performances are obtained with sepsis diagnosis kits currently available in the market, e.g., IRIDICA, SeptiFast, SeptiTest, or U-them These kits combine lysis buffers, for fast blood sample pretreatment and DNA extraction, with PCR analysis, enabling the detection of bacterial DNA, and diagnosing sepsis within 4 and 8 h. The presence of live bacteria can be detected after 10−15 h of incubation
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