Domestic plant food loss and waste in the United States: Environmental footprints and mitigation strategies.

The United States (U.S.) aims to reduce half of food loss and waste (FLW) by 2030. To achieve this goal, the public, academic, and political attentions on FLW have been increasing, and a series of actions have been implemented. However, the actions lack consideration on the categorical priority of FLW mitigation in relation to environmental footprints. In this article, we compare the FLW of three main plant food categories (i.e., grains, vegetables, and fruits) and their water and carbon footprints during 1970-2017. The vegetable FLW doubled during the period, reaching 3.39 × 1010 kg in 2017, which was 5- and 2-fold higher than the FLW of grains and fruits, respectively. The FLW of vegetables, grains, and fruits contributed 29%, 47%, and 24% to the total blue water wasted through FLW. The total carbon dioxide emissions generated by plant FLW were contributed by vegetables with 50%, grains with 31%, and fruits with 19%. Canonical correspondence analysis indicates that vegetable FLW had a higher positive correlation with urbanization, household incomes, gross domestic product, and high-income population than grain FLW, whereas fruit FLW was not influenced by these socioeconomic factors. Therefore, we suggest that the FLW mitigation should be prioritized on vegetables. Specific strategies include local food sourcing, shortening food miles, building food belts, and developing controlled-environment agriculture. Our data-based comparisons provide valuable insights into food policy improvement for achieving the 2030 reduction goal of the U.S., but the insights could be improved by considering the influences of foods imported from other nations.

Coupled Effects of Pore Water Velocity and Soil Heterogeneity on Bacterial Transport: Intact vs. Repacked Soils.

Transport of pathogenic bacteria from land surface to groundwater is largely influenced by rainfall intensity and geochemical and structural heterogeneities of subsurface sediments at different depths. It has been assumed that the change in rainfall intensity has different effects on bacterial transport as a function of soil depth. In this study, repacked and intact column systems were used to investigate the influences of pore water velocity on the transport of Escherichia coli 652T7 through a loamy soil collected from varying soil depths. The soils differed in geochemical properties and soil structures. The concentrations of bacteria in soil and liquid samples were measured using plate counting method. The breakthrough percentages of E. coli 652T7 increased with pore water velocity at each depth in both intact and disturbed soils. Among the different soil depths, the largest velocity effect was observed for the transport through the top soil (0-5 cm) of both disturbed and intact soil profiles. This depth-dependent effect of pore water velocity was attributed to down gradients of soil organic matter (SOM) and iron oxide contents with depth because SOM and iron oxides were favorable for bacterial attachment on soil surfaces. In addition, less bacteria broke through the disturbed soil than through the intact soil at the same depth, and the pore water velocity effect was stronger with the disturbed than intact soils. Specifically, the maximum C/C0 (i.e., ratio of effluent to influent concentration) doubled (i.e., from 0.36 to 0.76) in the 0-5 cm intact soil columns and tripled (i.e., from 0.16 to 0.43) in the 0-5 cm repacked soil columns. This structure-dependent effect of pore water velocity was attributed to larger pore tortuosity and a narrower range of pore sizes in the disturbed soil than in the intact soil. These findings suggest that change in pore water velocity could trigger bacterial remobilization especially in surface soils, where more bacteria are retained relative to deep soils.

Improvements in Smartphone and Night Vision Imaging Technologies Enable Low Cost, On-Site Assays of Bioluminescent Cells.

Technologies enabling on-site environmental detection or medical diagnostics in resource-limited settings have a strong disruptive potential compared to current analytical approaches that require trained personnel in laboratories with immobile, resource intensive instrumentation. Handheld devices, such as smartphones, are now routinely produced with CPUs, RAM, wireless data transfer capabilities, and high-resolution complementary metal oxide semiconductor (CMOS) cameras capable of supporting the capture and processing of bioluminescent signals. In theory, combining the capabilities of these devices with continuously bioluminescent human cell-based bioreporters would allow them to replicate the functionality of more expensive, more complex, and less flexible platforms while supporting human-relevant conclusions. In this work, we compare the performance of smartphone (CMOS) and night vision (image intensifier) devices with in vivo (CCD camera), and in vitro (photomultiplier tube) laboratory instrumentation for monitoring signal dynamics from continuously bioluminescent human cellular models under toxic, stable, and induced expression scenarios. All systems detected bioluminescence from cells at common plating densities. While the in vivo and in vitro systems were more sensitive and detected signal dynamics representing cellular health changes earlier, the night vision and smartphone systems also detected these changes with relatively similar coefficients of variation and linear detection capabilities. The smartphone system did not detect transcriptional induction. The night vision system did detect transcriptional activation, but was less sensitive than the in vivo or in vitro systems and required a stronger induction before the change could be resolved.

Screening for androgen agonists using autonomously bioluminescent HEK293 reporter cells.

Due to the public health concerns of endocrine-disrupting chemicals, there is an increasing demand to develop improved high-throughput detection assays for enhanced exposure control and risk assessment. A substrate-free, autobioluminescent HEK293ARE/Gal4-Lux assay was developed to screen compounds for their ability to induce androgen receptor (AR)-mediated transcriptional activation. The assay was validated against a group of 40 recommended chemicals and achieved an overall 87.5% accuracy in qualitatively classifying positive and negative AR agonists. The HEK293ARE/Gal4-Lux assay was demonstrated as a suitable tool for Tier 1 AR agonist screening. By eliminating exogenous substrate, this assay provided a significant advantage over traditional reporter assays by enabling higher-throughput screening with reduced testing costs while maintaining detection accuracy.

Real-time tracking of stem cell viability, proliferation, and differentiation with autonomous bioluminescence imaging.

BACKGROUND: Luminescent reporter proteins are vital tools for visualizing cells and cellular activity. Among the current toolbox of bioluminescent systems, only bacterial luciferase has genetically defined luciferase and luciferin synthesis pathways that are functional at the mammalian cell temperature optimum of 37 °C and have the potential for in vivo applications. However, this system is not functional in all cell types, including stem cells, where the ability to monitor continuously and in real-time cellular processes such as differentiation and proliferation would be particularly advantageous. RESULTS: We report that artificial subdivision of the bacterial luciferin and luciferase pathway subcomponents enables continuous or inducible bioluminescence in pluripotent and mesenchymal stem cells when the luciferin pathway is overexpressed with a 20-30:1 ratio. Ratio-based expression is demonstrated to have minimal effects on phenotype or differentiation while enabling autonomous bioluminescence without requiring external excitation. We used this method to assay the proliferation, viability, and toxicology responses of iPSCs and showed that these assays are comparable in their performance to established colorimetric assays. Furthermore, we used the continuous luminescence to track stem cell progeny post-differentiation. Finally, we show that tissue-specific promoters can be used to report cell fate with this system. CONCLUSIONS: Our findings expand the utility of bacterial luciferase and provide a new tool for stem cell research by providing a method to easily enable continuous, non-invasive bioluminescent monitoring in pluripotent cells.

Repetitive Detection of Aromatic Hydrocarbon Contaminants with Bioluminescent Bioreporters Attached on Tapered Optical Fiber Elements.

In this study, we show the repetitive detection of toluene on a tapered optical fiber element (OFE) with an attached layer of Pseudomonas putida TVA8 bioluminescent bioreporters. The bioluminescent cell layer was attached on polished quartz modified with (3-aminopropyl)triethoxysilane (APTES). The repeatability of the preparation of the optical probe and its use was demonstrated with five differently shaped OFEs. The intensity of measured bioluminescence was minimally influenced by the OFE shape, possessing transmittances between 1.41% and 5.00%. OFE probes layered with P. putida TVA8 were used to monitor liquid toluene over a two-week period. It was demonstrated that OFE probes layered with positively induced P. putida TVA8 bioreporters were reliable detectors of toluene. A toluene concentration of 26.5 mg/L was detected after <30 min after immersion of the probe in the toluene solution. Additional experiments also immobilized constitutively bioluminescent cells of E. coli 652T7, on OFEs with polyethyleneimine (PEI). These OFEs were repetitively induced with Lauria-Bertani (LB) nutrient medium. Bioluminescence appeared 15 minutes after immersion of the OFE in LB. A change in pH from 7 to 6 resulted in a decrease in bioluminescence that was not restored following additional nutrient inductions at pH 7. The E. coli 652T7 OFE probe was therefore sensitive to negative influences but could not be repetitively used.

High-Throughput Analysis of Endocrine-Disrupting Compounds Using BLYES and BLYAS Bioluminescent Yeast Bioassays.

Bioluminescent yeast assays BLYES and BLYAS are whole-cell bioassays that utilize genetically modified Saccharomyces cerevisiae bioreporters to detect estrogenic and androgenic activities, respectively. The bioreporter strains chromosomally express human estrogen receptor alpha (BLYES) or androgen receptor (BLYAS) and contain a reporter plasmid expressing the complete bacterial luciferase gene cassette (luxCDABE) under the control of an estrogen- or androgen-responsive promoter. Exposure to endocrine-disrupting compounds activates the receptor which subsequently turns on the expression of the reporter genes, resulting in dose-dependent bioluminescence (i.e., light) emission. These yeast whole-cell bioassays provide rapid, cost-effective, and high-throughput detection of endocrine-disrupting activities in environmental samples. This protocol will provide a detailed description of the standard assay procedures as well as a framework for data analysis.

Continuous and Real-Time In Vivo Autobioluminescent Imaging in a Mouse Model.

In vivo small animal bioluminescent imaging has become an indispensable technique for interrogating the localization, health, and functionality of implanted cells within the complex environment of a living organism. However, this task can be daunting for even the most experienced researchers because it requires multiple animal handling steps and produces differential output signal characteristics in response to a number of experimental design variables. The recent emergence of autobioluminescent cells, which autonomously and continuously produce bioluminescent output signals without external stimulation, has the potential to simplify this process, reduce variability by removing human-induced error, and improve animal welfare by reducing the number of required needlesticks per procedure. This protocol details the implantation and imaging of autobioluminescent cells within a mouse model to demonstrate how cells implanted from a single injection can be imaged repeatedly across any post-implantation timescale without the need for further human-animal interaction or signal activation steps. This approach provides a facile means to continuously monitor implanted cellular output signals in real-time for extended time periods.

Improving Estrogenic Compound Screening Efficiency by Using Self-Modulating, Continuously Bioluminescent Human Cell Bioreporters Expressing a Synthetic Luciferase.

A synthetic bacterial luciferase-based autobioluminescent bioreporter, HEK293ERE/Gal4-Lux, was developed in a human embryonic kidney (HEK293) cell line for the surveillance of chemicals displaying endocrine disrupting activity. Unlike alternative luminescent reporters, this bioreporter generates bioluminescence autonomously without requiring an external light-activating chemical substrate or cellular destruction. The bioreporter's performance was validated against a library of 76 agonistic and antagonistic estrogenic endocrine disruptor chemicals and demonstrated reproducible half maximal effective concentration (EC50) values meeting the U.S. Environmental Protection Agency (EPA) guidelines for Tier 1 endocrine disrupting chemical screening assays. For model compounds, such as the estrogen receptor (ER) agonist 17ß-estradiol, HEK293ERE/Gal4-Lux demonstrated an EC50 value (7.9 × 10-12 M) comparable to that of the current EPA-approved HeLa-9903 firefly luciferase-based estrogen receptor transcription assay (4.6 × 10-12 M). Screening against an expanded array of common ER agonists likewise produced similar relative effect potencies as compared with existing assays. The self-initiated autobioluminescent signal of the bioreporter permitted facile monitoring of the effects of endocrine disrupting chemicals, which decreased the cost and hands-on time required to perform these assays. These characteristics make the HEK293ERE/Gal4-Lux bioreporter potentially suitable as a high-throughput human cell-based assay for screening estrogenic activity.

Surface-Adsorbed Contaminants Mediate the Importance of Chemotaxis and Haptotaxis for Bacterial Transport Through Soils.

Chemotaxis and haptotaxis are important biological mechanisms that influence microbial movement toward concentrated chemoattractants in mobile liquids and along immobile surfaces, respectively. This study investigated their coupled effect, as induced by naphthalene (10 mg L-1), on the transport and retention of two pollutant-degrading bacteria, Pseudomonas fluorescens 5RL (Pf5RL) and Pseudomonas stutzeri DQ1 (PsDQ1), in quartz sand and natural soil. The results demonstrated that PsDQ1 was not chemotactic, whereas Pf5RL was chemotactic at 25°C but not at 4°C due to the restricted movement. In a quartz sand column, haptotaxis did not play a role in increasing the transport of Pf5RL as compared with chemotaxis. Compared with a naphthalene-free soil column, Pf5RL broke through naphthalene-presaturated soil columns to reach a stable effluent concentration 0.5 pore volumes earlier due to advective chemotaxis occurring behind the plume front in the bulk solution. Pf5RL also demonstrated greater retention (e.g., a doubled rate of attachment and a one-third smaller breakthrough percentage) due to along-surface haptotaxis and near-surface chemotaxis occurring in less mobile water near the soil surface. However, both chemotaxis and haptotaxis were weakened when Pf5RL co-transported with naphthalene due to reduced adsorption of naphthalene on the soil. This study suggests that surface adsorption of naphthalene can mediate the relative importance of advective chemotaxis (facilitating initial breakthrough), near-surface chemotaxis (increasing bacterial collision), and haptotaxis (increasing bacterial residence time).

A rapid and reagent-free bioassay for the detection of dioxin-like compounds and other aryl hydrocarbon receptor (AhR) agonists using autobioluminescent yeast.

An autonomously bioluminescent Saccharomyces cerevisiae BLYAhS bioreporter was developed in this study for the simple and rapid detection of dioxin-like compounds (DLCs) and aryl hydrocarbon receptor (AhR) agonists. This recombinant yeast reporter was based on a synthetic bacterial luciferase reporter gene cassette (lux) that can produce the luciferase as well as the enzymes capable of self-synthesizing the requisite substrates for bioluminescent production from endogenous cellular metabolites. As a result, bioluminescent signal production is generated continuously and autonomously without cell lysis or exogenous reagent addition. By linking the expression of the autobioluminescent lux reporter cassette to AhR activation via the use of a dioxin-responsive promoter, the S. cerevisiae BLYAhS bioreporter emitted a bioluminescent signal in response to DLC exposure in a dose-responsive manner. The model dioxin, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), could be detected within 4 h with a half maximal effective concentration (EC50) of ~ 8.1 nM and a lower detection limit of 500 pM. The autobioluminescent response of BLYAhS to other AhR agonists, including 2,3,7,8-tetrachlorodibenzofuran (TCDF), polychlorinated bisphenyl congener 126 (PCB-126) and 169 (PCB-169), 1,2,3,6,7,8-hexachlorodibenzo-p-dioxin (HxCDD), 1,2,3,4,6,7,8-heptachlorodibenzo-p-dioxin (HpCDD), benzo[a]pyrene (BaP), and ß-naphthoflavone (bNF), were also characterized in this study. The non-destructive and reagent-free nature of the BLYAhS reporter assay facilitated near-continuous, automated signal acquisition without additional hands-on effort and cost, providing a simple and cost-effective method for rapid DLC detection.

Co-Cultured Continuously Bioluminescent Human Cells as Bioreporters for the Detection of Prodrug Therapeutic Impact Pre- and Post-Metabolism.

Modern drug discovery workflows require assay systems capable of replicating the complex interactions of multiple tissue types, but that can still function under high throughput conditions. In this work, we evaluate the use of substrate-free autobioluminescence in human cell lines to support the performance of these assays with reduced economical and logistical restrictions relative to substrate-requiring bioluminescent reporter systems. The use of autobioluminescence was found to support assay functionality similar to existing luciferase reporter targets. The autobioluminescent assay format was observed to correlate strongly with general metabolic activity markers such as ATP content and the presence of reactive oxygen species, but not with secondary markers such as glutathione depletion. At the transcriptional level, autobioluminescent dynamics were most closely associated with expression of the CYP1A1 phase I detoxification pathway. These results suggest constitutively autobioluminescent cells can function as general metabolic activity bioreporters, while pairing expression of the autobioluminescent phenotype to detoxification pathway specific promoters could create more specific sensor systems.

A Real-Time Non-invasive Auto-bioluminescent Urinary Bladder Cancer Xenograft Model.

PURPOSE: The study was to develop an auto-bioluminescent urinary bladder cancer (UBC) xenograft animal model for pre-clinical research. PROCEDURE: The study used a humanized, bacteria-originated lux reporter system consisting of six (luxCDABEfrp) genes to express components required for producing bioluminescent signals in human UBC J82, J82-Ras, and SW780 cells without exogenous substrates. Immune-deficient nude mice were inoculated with Lux-expressing UBC cells to develop auto-bioluminescent xenograft tumors that were monitored by imaging and physical examination. RESULTS: Lux-expressing auto-bioluminescent J82-Lux, J82-Ras-Lux, and SW780-Lux cell lines were established. Xenograft tumors derived from tumorigenic Lux-expressing auto-bioluminescent J82-Ras-Lux cells allowed a serial, non-invasive, real-time monitoring by imaging of tumor development prior to the presence of palpable tumors in animals. CONCLUSIONS: Using Lux-expressing auto-bioluminescent tumorigenic cells enabled us to monitor the entire course of xenograft tumor development through tumor cell implantation, adaptation, and growth to visible/palpable tumors in animals.

The Expanding Toolbox of In Vivo Bioluminescent Imaging.

In vivo bioluminescent imaging (BLI) permits the visualization of engineered bioluminescence from living cells and tissues to provide a unique perspective toward the understanding of biological processes as they occur within the framework of an authentic in vivo environment. The toolbox of in vivo BLI includes an inventory of luciferase compounds capable of generating bioluminescent light signals along with sophisticated and powerful instrumentation designed to detect and quantify these light signals non-invasively as they emit from the living subject. The information acquired reveals the dynamics of a wide range of biological functions that play key roles in the physiological and pathological control of disease and its therapeutic management. This mini review provides an overview of the tools and applications central to the evolution of in vivo BLI as a core technology in the preclinical imaging disciplines.

The Repetitive Detection of Toluene with Bioluminescence Bioreporter Pseudomonas putida TVA8 Encapsulated in Silica Hydrogel on an Optical Fiber.

Living cells of the lux-based bioluminescent bioreporter Pseudomonas putida TVA8 were encapsulated in a silica hydrogel attached to the distal wider end of a tapered quartz fiber. Bioluminescence of immobilized cells was induced with toluene at high (26.5 mg/L) and low (5.3 mg/L) concentrations. Initial bioluminescence maxima were achieved after >12 h. One week after immobilization, a biofilm-like layer of cells had formed on the surface of the silica gel. This resulted in shorter response times and more intensive bioluminescence maxima that appeared as rapidly as 2 h after toluene induction. Considerable second bioluminescence maxima were observed after inductions with 26.5 mg toluene/L. The second and third week after immobilization the biosensor repetitively and semiquantitatively detected toluene in buffered medium. Due to silica gel dissolution and biofilm detachment, the bioluminescent signal was decreasing 20-32 days after immobilization and completely extinguished after 32 days. The reproducible formation of a surface cell layer on the wider end of the tapered optical fiber can be translated to various whole cell bioluminescent biosensor devices and may serve as a platform for in-situ sensors.

Biological toxicity of cellulose nanocrystals (CNCs) against the luxCDABE-based bioluminescent bioreporter Escherichia coli 652T7.

The aim of this study was to evaluate the biological toxicity of cellulose nanocrystals (CNCs) using the constitutively bioluminescent luxCDABE-based bioreporter Escherichia coli 652T7. The effects of CNCs on E. c oli 652T7 biotoxicity were investigated at different CNC concentrations, reaction times, and IC50 values. CNC toxicity was also compared with and without ultrasonic dispersion to establish dispersibility effects. The results demonstrated that CNCs were not significantly toxic at concentrations at or below 250 mg/L. At concentrations higher than 300 mg/L, toxicity increased linearly as CNC concentrations increased up to 2000 mg/L. IC50 calculations demonstrated an increase in cytotoxicity as CNC exposure times increased, and elevated dispersibility of the CNCs were shown to increase cytotoxicity effects. These results suggest that CNCs can impact microbial populations if elevated concentration thresholds are met.

Microbial availability of mercury: effective detection and organic ligand effect using a whole-cell bioluminescent bioreporter.

A luxCDABE-based genetically engineered bacterial bioreporter (Escherichia coli ARL1) was used to detect bioavailable ionic mercury (Hg(II)) and investigate the effects of humic acids and ethylenediaminetetraacetic acid (EDTA) on the bioavailability of mercury in E. c oli. Results showed that the E. c oli ARL1 bioreporter was sensitive to mercury, with a detection limit of Hg(II) of 0.5 µg/L and a linear dose/response relationship up to 2000 µg Hg(II)/L. Humic acids and EDTA decreased the Hg(II)-induced bioluminescent response of strain ARL1, suggesting that the two organic ligands reduced the bioavailability of Hg(II) via complexation with Hg(II). Compared with traditional chemical methods, the use of E. c oli ARL1 is a cost-effective, rapid, and reliable approach for measuring aqueous mercury at very low concentrations and thus has potential for applications in field in situ monitoring.

Real-time toxicity and metabolic activity tracking of human cells exposed to Escherichia coli O157:H7 in a mixed consortia.

Escherichia coli O157:H7 is a significant human pathogen that is continually responsible for sickness, and even death, on a worldwide scale. While the pathology of E. coli O157:H7 infection has been well studied, the effect of it's multiple resulting cytotoxic mechanisms on host metabolic activity has not been well characterized. To develop a more thorough understanding of these effects, several bioluminescence assays were evaluated for their ability to track both toxicity and host metabolic activity levels in real-time. The use of continuously autobioluminescent human cells was determined to be the most favorable method for tracking these metrics, as its self-sufficient autobioluminescent phenotype was unaffected by the presence of the infecting bacteria and its signal could be measured without cellular destruction. Using this approach, it was determined that infection with as few as 10 CFU of E. coli O157:H7 could elicit cytotoxic effects. Regardless of the initial infective dose, an impact on metabolic expression was not observed until bacterial populations reached levels between 5 × 10(5) and 1 × 10(6) (R(2) = 0.933), indicating that a critical bacterial infection level must be reached prior to the onset of cytotoxic effects. Supporting this hypothesis, it was found that cells displaying infection-mediated metabolic activity reductions could recover to wild type metabolic activity levels if the infecting bacteria were removed prior to cell death. These results indicate that rapid treatment of E. coli O157:H7 infection could serve to limit host metabolic impact and reduce overall host cell death.

Detection of organic compounds with whole-cell bioluminescent bioassays.

Natural and manmade organic chemicals are widely deposited across a diverse range of ecosystems including air, surface water, groundwater, wastewater, soil, sediment, and marine environments. Some organic compounds, despite their industrial values, are toxic to living organisms and pose significant health risks to humans and wildlife. Detection and monitoring of these organic pollutants in environmental matrices therefore is of great interest and need for remediation and health risk assessment. Although these detections have traditionally been performed using analytical chemical approaches that offer highly sensitive and specific identification of target compounds, these methods require specialized equipment and trained operators, and fail to describe potential bioavailable effects on living organisms. Alternatively, the integration of bioluminescent systems into whole-cell bioreporters presents a new capacity for organic compound detection. These bioreporters are constructed by incorporating reporter genes into catabolic or signaling pathways that are present within living cells and emit a bioluminescent signal that can be detected upon exposure to target chemicals. Although relatively less specific compared to analytical methods, bioluminescent bioassays are more cost-effective, more rapid, can be scaled to higher throughput, and can be designed to report not only the presence but also the bioavailability of target substances. This chapter reviews available bacterial and eukaryotic whole-cell bioreporters for sensing organic pollutants and their applications in a variety of sample matrices.

Expression of a humanized viral 2A-mediated lux operon efficiently generates autonomous bioluminescence in human cells.

BACKGROUND: Expression of autonomous bioluminescence from human cells was previously reported to be impossible, suggesting that all bioluminescent-based mammalian reporter systems must therefore require application of a potentially influential chemical substrate. While this was disproven when the bacterial luciferase (lux) cassette was demonstrated to function in a human cell, its expression required multiple genetic constructs, was functional in only a single cell type, and generated a significantly reduced signal compared to substrate-requiring systems. Here we investigate the use of a humanized, viral 2A-linked lux genetic architecture for the efficient introduction of an autobioluminescent phenotype across a variety of human cell lines. METHODOLOGY/PRINCIPAL FINDINGS: The lux cassette was codon optimized and assembled into a synthetic human expression operon using viral 2A elements as linker regions. Human kidney, breast cancer, and colorectal cancer cell lines were both transiently and stably transfected with the humanized operon and the resulting autobioluminescent phenotype was evaluated using common imaging instrumentation. Autobioluminescent cells were screened for cytotoxic effects resulting from lux expression and their utility as bioreporters was evaluated through the demonstration of repeated monitoring of single populations over a prolonged period using both a modified E-SCREEN assay for estrogen detection and a classical cytotoxic compound detection assay for the antibiotic Zeocin. Furthermore, the use of self-directed bioluminescent initiation in response to target detection was assessed to determine its amenability towards deployment as fully autonomous sensors. In all cases, bioluminescent measurements were supported with traditional genetic and transcriptomic evaluations. CONCLUSIONS/SIGNIFICANCE: Our results demonstrate that the viral 2A-linked, humanized lux genetic architecture successfully produced autobioluminescent phenotypes in all cell lines tested without the induction of cytotoxicity. This autobioluminescent phenotype allowed for repeated interrogation of populations and self-directed control of bioluminescent activation with detection limits and EC50 values similar to traditional reporter systems, making the autobioluminescent cells amenable to automated monitoring and significantly reducing the time and cost required to perform bioluminescent workflows.