ABSTRACT
During the COVID-19 pandemic, point-of-care genetic testing (POCT) devices were used for on-time and on-site detection of the virus, which helped to prevent and control the spread of the pandemic. Smartphones, which are widely used electronic devices with many functions, have the potential to be used as a molecular diagnostic platform for universal healthcare monitoring. Several integrated diagnostics platforms for the real-time and end-point detection of COVID-19 were developed using the functions of smartphones, such as the operating system, power, sound, camera, data storage, and display. These platforms use the 5V output power of smartphones, which can be amplified to power a micro-capillary electrophoresis system or a thin-film heater, and the CMOS camera of smartphones can capture the color change during a colorimetric loop-mediated isothermal amplification test and detect fluorescence signals. Smartphones can also be used with self-written web-based apps to enable automatic and remote pathogen analysis on POCT platforms. Our lab developed a handheld micro-capillary electrophoresis device for end-point detection of SARS-CoV-2, as well as an integrated smartphone-based genetic analyzer for the qualitative and quantitative colorimetric detection of foodborne pathogens with the help of a custom mobile app. © 2023 SPIE.
ABSTRACT
An accurate, convenient, and rapid diagnostic platform, which can be applied in facility-limited or point-of-care (POC) settings, is essential to help prevent the spread of infectious diseases and enable the most effective treatment to be selected. In this study, we describe the development of a new isothermal molecular diagnostic system named multipurpose advanced split T7 promoter-based transcription amplification (MASTER) for the rapid and ultrasensitive detection of various pathogens containing single-stranded RNA and double-stranded DNA. MASTER produces a large number of RNA amplicons in the presence of target pathogens, which generate fluorescence or colorimetric signals based on light-up RNA aptamers or lateral flow assays. Implementing MASTER at 37 °C for<1 h achieved the detection of a single copy per reaction without cross-reactivity. Moreover, the testing of 40 clinical samples revealed that MASTER exhibited excellent accuracy with 100% sensitivity and specificity for SARS-CoV-2 diagnosis. Furthermore, a one-pot MASTER system capable of accelerating practical applications was demonstrated, indicating that the MASTER system is a promising platform for the effective surveillance of various pathogens. © 2023 Elsevier B.V.
ABSTRACT
In the ongoing COVID-19 pandemic, sensitive and rapid on-site detection of the SARS-CoV-2 coronavirus has been one of crucial objectives. A point-of-care (PoC) device called PATHPOD for quick, on-site detection of SARS-CoV-2 employing a real-time reverse-transcription loop-mediated isothermal amplification (RT-rLAMP) reaction on a polymer cartridge. The PATHPOD consists of a standalone device (weighing under 1.2 kg) and a cartridge, and can identify 10 distinct samples and 2 controls in less than 50 minutes. The PATHPOD PoC system is fabricated and clinically validated for the first time in this work © 2023 IEEE.
ABSTRACT
Here, we developed a copper sulfate (CuSO4)-initiated diphenylamine (DPA)-based colorimetric strategy coupled with loop-mediated isothermal amplification (LAMP) for rapid detection of two critical contagious pathogens, SARS-CoV-2 and Enterococcus faecium. To detect the DNA, acid hydrolysis of LAMP amplicons was executed, enabling the development of a blue color. In the LAMP amplicons, the bond between the purines and deoxyribose is extremely labile. It can be broken using 70% sulfuric acid followed by phosphate group elimination, which generates a highly active keto aldehyde group. CuSO4 plays an imperative role inducing DPA to rapidly react with the keto aldehyde group, producing an intense blue color within 5 min. Moreover, low quantities such as 103 copies μL-1 of SARS-CoV-2 RNA and 102 CFU mL-1 of E. faecium were successfully detected, revealing the advantages of the introduced method. To confirm practical applicability, multiplex detection of pathogens was performed using a foldable microdevice comprising reaction and detection zones. Various reactions such as DNA extraction, LAMP, and acid hydrolysis occurred in the reaction zone. Then, colorimetric reagents (DPA, CuSO4, and ethylene glycol) contained in the detection zone were mixed with the keto aldehyde group by simply folding the microdevice, which was heated at 65 °C for 5 min for colorimetric detection. An intense blue color was developed where the target DNA was present. These results indicate that the method proposed in this study is highly suitable for point-of-care applications, especially in resource-limited settings for the rapid detection of harmful pathogens. © 2023 American Chemical Society.
ABSTRACT
In recent years, the emergence of COVID-19 has created disastrous health effects worldwide. Doxycycline, a member of the tetracycline group, has been prescribed as a treatment companion for attending this catastrophe. Due to extensive use and high solubility, a significant amount of un-metabolized doxycycline has been found to reach water bodies within a short time, and consumption of this water may lead to the development of fatal resistance in organisms and create health problems. Therefore, it has become necessary to develop suitable technologies from a geoenvironmental point of view to remove these unwanted antibiotics from wastewater. In this context, locally obtainable silty-sandy soil was explored as a low-cost material in a constructed wetland with Chrysopogon zizanioides (vetiver sp.) for phytoremediation to mitigate doxycycline spiked wastewater. The obtained soil hydraulic conductivity was 1.63×10-7 m/s. Batch adsorption tests conducted on silty-sandy soil, vetiver leaf, and vetiver root provided maximum removal efficiencies of 90%, 72%, and 80% percent, respectively, at optimal sorbent doses of 10 g/L, 17 g/L, and 16 g/L, and contaminant concentrations of 25 mg/L, 20 mg/L, and 23 mg/L, with a 30-min time of contact. The Freundlich isotherm was the best fit, indicative of sufficient sorption capacity of all the adsorbents for doxycycline. The best match in the kinetic research was pseudo-second-order kinetics. A one dimensional vertical column test with the used soil on doxycycline revealed a 90% breakthrough in 24 h for a soil depth of 30 mm. Studies on a laboratory-scale wetland and numerically modeled yielded removal of around 92% by the selected soil and about 98% combined with Chrysopogon zizanioides for 25 mg/L of initial doxycycline concentration, which is considered quite satisfactory. Simulated results matched the laboratory tests very well. The study is expected to provide insight into remedies for similar practical problems. © 2023 American Society of Civil Engineers.
ABSTRACT
Here, we developed a copper sulfate (CuSO4)-initiated diphenylamine (DPA)-based colorimetric strategy coupled with loop-mediated isothermal amplification (LAMP) for rapid detection of two critical contagious pathogens, SARS-CoV-2 and Enterococcus faecium. To detect the DNA, acid hydrolysis of LAMP amplicons was executed, enabling the development of a blue color. In the LAMP amplicons, the bond between the purines and deoxyribose is extremely labile. It can be broken using 70% sulfuric acid followed by phosphate group elimination, which generates a highly active keto aldehyde group. CuSO4 plays an imperative role inducing DPA to rapidly react with the keto aldehyde group, producing an intense blue color within 5 min. Moreover, low quantities such as 103 copies μL-1 of SARS-CoV-2 RNA and 102 CFU mL-1 of E. faecium were successfully detected, revealing the advantages of the introduced method. To confirm practical applicability, multiplex detection of pathogens was performed using a foldable microdevice comprising reaction and detection zones. Various reactions such as DNA extraction, LAMP, and acid hydrolysis occurred in the reaction zone. Then, colorimetric reagents (DPA, CuSO4, and ethylene glycol) contained in the detection zone were mixed with the keto aldehyde group by simply folding the microdevice, which was heated at 65 °C for 5 min for colorimetric detection. An intense blue color was developed where the target DNA was present. These results indicate that the method proposed in this study is highly suitable for point-of-care applications, especially in resource-limited settings for the rapid detection of harmful pathogens. © 2023 American Chemical Society
ABSTRACT
Background: The novel COVID-19 outbreak has infected human population all around the world. Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) diagnosis in a rapid manner remains challenging for health care professionals. Currently, RT-qPCR technique is extensively practiced in SARS-CoV-2 diagnosis and is considered as gold standard. The constraints of RT-qPCR, high cost and need for trained technician, longer detection time, highlighted the need for alternate healthcare diagnostic approaches. They follow the WHO assured standard and offer the health-care sector optimism. One of them is the Loop Mediated isothermal amplification system (LAMP). There is no need for costly equipment like thermal cycler since LAMP assay is performed at a fixed temperature. It can also be implemented as a point of care testing device. RT-LAMP is one of the extensively used isothermal amplification system in pathogen diagnostics.Aims: The current study aims to validate and standardize RT-LAMP assay for rapid diagnosis of SARS-CoV-2 in both lab and field conditions. The reactions can be carried out using a heating vessel including the use of a water bath and end-point detection by colorimetry. A rising middle ground of tiny, more portable technology, that provides most of the capability at less cost and time.Methods and Results: 20 Samples were taken from COVID-19 positive patients. RNA extraction from COVID-19 samples was followed up by one-step reverse transcription and loop-mediated isothermal amplification (LAMP). LAMP primers were designed to amplify the conserved regions of SARS-COV-2 specific genes. The target regions for primer design were selected after genome-wide sequence alignment of SARS-CoV-2 strains isolated in various regions of the world i.e., Europe, Africa, Asia, and North America. RT-LAMP assays were performed at the specific incubation temperature (60°C) for 50 minutes. Assay was optimized as per consumable compatibility, COVID template integrity, primer concentration, template concentration, primer ratio, testing time etc. Sensitivity and specificity of the assay was elucidated. Finally, different end-point analysis i.e., Agarose Gel Electrophoresis and Colorimetry have been used to interpret the results.Conclusion: RT-LAMP assay has shown to be a quick and accurate diagnostic method that can be put to use for SARS-CoV-2 detection in laboratories and Point-of- Care settings. © 2022 IEEE.
ABSTRACT
Because disinfectants have been essential during the COVID-19 pandemic, the global demand for benzalkonium chlorides (BACs) has significantly increased. BACs can inactivate coronaviruses, but are known as toxic. In this study, we investigated the adsorption mechanisms of BAC12, BAC14, and BAC16 in water using powdered activated carbon (PAC). The effects of the reaction time, pH, and temperature on the adsorption kinetics of BACs were examined. The adsorption reaction followed pseudo-second-order kinetics, and better fitted to the Langmuir isotherm than the Freundlich isotherm. The best adsorption of BACs was achieved at neutral pH conditions. Thermodynamic analysis revealed that adsorption of BACs onto PAC is a spontaneous and endothermic process. Competitive adsorption experiments revealed that BACs with longer alkyl chains were adsorbed more effectively onto PAC than shorter alkyl chain BACs, implying that, while the electrostatic interaction is an important adsorption mechanism for BAC12, van der Waals interaction plays a more important role during the adsorption of BAC14 and BAC16. Finally, we observed the partial detoxification (69%) BAC in adsorption treated water with PAC using a Microtox test. © 2022 Korean Society of Environmental Engineers.
ABSTRACT
Rolling Circle Amplification (RCA) has shown significant potential for pathogen diagnostics providing high specificity and sensitivity combined with relatively low temperature (<37 °C) isothermal amplification. In the context of the ongoing COVID-19 pandemic, we report the development of an RCA-based method allowing direct detection of SARS-CoV-2 RNA in microfluidics. The viral RNA was hybridized to biotinylated oligos and L-probes in solution, enriched in a microchannel and subsequently amplified in situ using padlock probes against the L-probes. This method allowed the detection of 1x103 viral copies/μL within 90 minutes of amplification, demonstrating an alternative approach to current isothermal amplification methods. © 2021 MicroTAS 2021 - 25th International Conference on Miniaturized Systems for Chemistry and Life Sciences. All rights reserved.
ABSTRACT
The demand for scalable, rapid and sensitive COVID-19 diagnostics is particularly pressing at present to help contain the spread of infection and prevent overwhelming the capacity of health systems. While high-income countries have managed to rapidly expand diagnostic capacities, such is not the case in resource-limited settings of low- to medium-income countries. We report the development of an integrated modular centrifugal microfluidic platform costing less than 250 USD to perform loop-mediated isothermal amplification (LAMP) of viral RNA directly from heat-inactivated nasopharyngeal swab samples. The platform was validated with a panel of 131 nasopharyngeal swab samples collected from symptomatic COVID-19 patients. © 2021 MicroTAS 2021 - 25th International Conference on Miniaturized Systems for Chemistry and Life Sciences. All rights reserved.
ABSTRACT
Silica fibers and capillaries offer opportunities for compact integration of optics with microfluidics while adding advantages such as;flexibility within a high aspect ratio format, uniaxial arrangements, and measurement-at-a-distance. Here, we describe droplet microfluidics-based nucleic acid detection of SARS-CoV-2 in a lab-in-a-fiber platform. The fiber component integrates three modules with key functions: droplet generation, incubation, and fluorescence detection. Within the scope of this work, we developed the component specifically to target the quantification of SARS-CoV-2 viral RNA through reverse-transcription loop-mediated isothermal amplification (RT-LAMP). The all-fiber component could successfully generate uniform droplets and differentiate pre-amplified positive LAMP reaction from negative sample. © 2021 MicroTAS 2021 - 25th International Conference on Miniaturized Systems for Chemistry and Life Sciences. All rights reserved.
ABSTRACT
The result readouts of loop-mediated isothermal amplification (LAMP) still remain challenging because current techniques require bulky equipment and could not give clear visualization. In this study, we developed a paper device to integrate LAMP and a novel strategy for power-free and naked-eye readout of result relied on polydopamine aggregation. The introduced paper device was used to detect DNA extracted from Escherichia coli O157:H7 (E. coli O157:H7), Enterococcus faecium (E. faecium), and SARS-CoV-2 plasmid. © 2021 MicroTAS 2021 - 25th International Conference on Miniaturized Systems for Chemistry and Life Sciences. All rights reserved.
ABSTRACT
This paper presents a newly developed microfluidic flow control theory for autonomous sample dispensing into an array of reaction microchambers. The theoretical predictions for the possible dispensing number and maximum flow rate were validated by comparison to experimental results. Moreover, we successfully demonstrated the rapid genetic detection of multiple infectious viruses including SARS-CoV-2 in fabricated polydimethylsiloxane (PDMS)-based microfluidic devices based on the loop-mediated isothermal amplification (LAMP) method. © 2021 MicroTAS 2021 - 25th International Conference on Miniaturized Systems for Chemistry and Life Sciences. All rights reserved.
ABSTRACT
Early and accurate detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) at the point-of-care (POC) is crucial for reducing the transmission of coronavirus disease 2019 (COVID-19). To address this need, we have developed a valve-enabled lysis, paper-based RNA enrichment, and RNA amplification device (VLEAD) for detecting SARSCoV-2. We have combined VLEAD with a smart coffee mug for sample preparation, nucleic acid isothermal amplification, and colorimetric detection using a smartphone camera or a naked eye. VLEAD enables two critical functions required for POC testing: sample preparation and detection. Since the reagents can be pre-packaged in the device, all operational steps can be carried out at POC. We have demonstrated the functions of the device by analyzing samples spiked with heat-inactivated SARSCoV-2, which were obtained from BEI Resources. This rapid and highly sensitive POC platform for SARS-CoV-2 detection has a potential to help reduce COVID-19 transmission. Copyright © 2021 by ASME
ABSTRACT
In developing an effective monitoring program for the wastewater surveillance of SARS-CoV-2 ribonucleic acid (RNA), the importance of sampling methodology is paramount. Passive sampling has been shown to be an effective tool to detect SARS-CoV-2 RNA in wastewater. However, the adsorption characteristics of SARS-CoV-2 RNA on passive sampling material are not well-understood, which further obscures the relationship between wastewater surveillance and community infection. In this work, adsorption kinetics and equilibrium characteristics were evaluated using batch-adsorption experiments for heat-inactivated SARS-CoV-2 (HI-SCV-2) adsorption to electronegative filters. Equilibrium isotherms were assessed or a range of total suspended solids (TSS) concentrations (118, 265, and 497 mg L-1) in wastewater, and a modeled q(max) of 7 X 10(3) GU cm(-2) was found. Surrogate adsorption kinetics followed a pseudo-first-order model in wastewater with maximum concentrations achieved within 24 h. In both field and isotherm experiments, equilibrium behavior and viral recovery were found to be associated with wastewater and eluate TSS. On the basis of the results of this study, we recommend a standard deployment duration of 24-48 h and the inclusion of eluate TSS measurement to assess the likelihood of solids inhibition during analysis.