A Whole-Cell Bacterial Biosensor for Blood Markers Detection in Urine

Abstract

The early detection of blood in urine (hematuria) can play a crucial role in the treatment of serious diseases (e.g., infections, kidney disease, schistosomiasis, and cancer). Therefore, the development of low-cost portable biosensors for blood detection in urine has become necessary. Here, we designed an ultrasensitive whole-cell bacterial biosensor interfaced with an optoelectronic measurement module for heme detection in urine. Heme is a red blood cells (RBCs) component that is liberated from lysed cells. The bacterial biosensor includes Escherichia coli cells carrying a heme-sensitive synthetic promoter integrated with a luciferase reporter (luxCDABE) from Photorhabdus luminescens. To improve the bacterial biosensor performance, we re-engineered the genetic structure of luxCDABE operon by splitting it into two parts (luxCDE and luxAB). The luxCDE genes were regulated by the heme-sensitive promoter, and the luxAB genes were regulated by either constitutive or inducible promoters. We examined the genetic circuit's performance in synthetic urine diluent supplied with heme and in human urine supplied with lysed blood. Finally, we interfaced the bacterial biosensor with a light detection setup based on a commercial optical measurement single-photon avalanche photodiode (SPAD). The whole-cell biosensor was tested in human urine with lysed blood, demonstrating a low-cost, portable, and easy-to-use hematuria detection with an ON-to-OFF ratio of 6.5-fold for blood levels from 5 × 10 4 to 5 × 10 5 RBC per mL of human urine.

Engineering Coupled Consortia-Based Biosensors using Incoherent Feedforward Loops

“…In this study, we utilized the stability output IFFL circuit to regulate individual consortia within bacterial biosensors to detect clinical biomarkers such as Heme and Lactate. Meanwhile, the luxCDE genes encode three enzymes (reductase, transferase, and synthetase) that are crucial for providing the substrate for the luminescence reaction [19], [38]. For the Heme-responsive strain, the Lactococcus lactis Heme-responsive transcriptional repressor HrtR and an outer-membrane transporter ChuA are expressed under constitutive promoters ProD and PJ23107, respectively, which are located on a medium-copy-number plasmid…”

“…However, the '01' configuration, which represents the presence of Heme only, displays a response similar to the '11' configuration in both shape and signal intensity, indicating that Heme alone can elicit a response nearly equivalent to that of both inducers combined. The decline in signal (after 20 hours) intensity over time is primarily attributed to an intrinsic characteristic of the heme biosensor [19]. Furthermore, variation in the initial time point at which the signal begins to decrease is influenced by the design of the biosensor and the interaction of the bacterial consortium, which can affect biosensor performance.…”

“…Bacterial biosensors have become indispensable, offering versatility from environmental monitoring to disease diagnosis [16], [19]. A significant trend in this field is the simultaneous detect multiple targets, providing comprehensive outcomes [20], [21], [22].…”

Whole‐Cell P450 Biocatalysis Using Engineered Escherichia coli with Fine‐Tuned Heme Biosynthesis

“…The heme‐responsive biosensor (HrtR regulator and its binding site hrtO from Lactococcus lactis ) has been used to design an E. coli whole‐cell biosensor for detecting heme in urine. [ 22 ] Additionally, heme‐responsive biosensors and CRISPRi have been used to regulate the transcription of several heme biosynthesis genes ( hemB , hemC , and hemH ) in E. coli . [ 23 ] However, this system should be modified to provide sufficient heme in the cells overexpressing P450 genes.…”

Comparative Analysis of Cell Metabolic Activity Sensing by Escherichia coli rrnB P1-lux and Cd Responsive-Lux Biosensors: Time-Resolved Experiments and Mechanistic Modelling

“…Whole-cell bacterial sensors are now well-established tools among the arsenal of instruments used by the scientific community for detecting and quantifying chemicals (e.g., drugs [ 1 , 2 ], toxins [ 3 ], quorum sensing signaling molecules [ 4 ], siderophores [ 5 ]), pollutants (e.g., metals [ 6 ], organics [ 7 ]) or microbial pathogens [ 8 ] in biological media [ 9 ] or environmental samples [ 10 , 11 ], and for addressing the toxicity of contaminants [ 12 ]. They offer an elegant alternative to conventional physicochemical methods (e.g., extraction, chromatography) due to their cost, ease of use, specificity and sensivity [ 13 ].…”