Rapid Diagnostic Platform Research Articles

Development of a Recombinase Polymerase Amplification-Coupled CRISPR/Cas12a Platform for Rapid Detection of Antimicrobial-Resistant Genes in Carbapenem-Resistant Enterobacterales

The worldwide spread of carbapenemase-producing Enterobacterales (CPE) represents a significant threat owing to the high mortality and morbidity rates. Traditional diagnostic methods are often too slow and complex for rapid point-of-care testing. Therefore, we developed a recombinase polymerase amplification (RPA)-coupled CRISPR/Cas12a system (RCCS), a rapid, accurate, and simple diagnostic platform for detecting antimicrobial-resistant genes. The RCCS detected carbapenemase genes (blaKPC and blaNDM) within 50 min, including 10 min for DNA extraction and 30–40 min for RCCS reaction (a 20 min RPA reaction with a 10–20-min CRISPR/Cas12a assay). Fluorescence signals obtained from the RCCS platform were visualized using lateral-flow test strips (LFSs) and real-time and endpoint fluorescence. The LFS clearly displayed test lines while detecting carbapenemase genes. Furthermore, the RCCS platform demonstrated high sensitivity by successfully detecting blaKPC and blaNDM at the attomolar and picomolar levels, respectively. The accuracy of the RCCS platform was validated with clinical isolates of Klebsiella pneumoniae and Escherichia coli; a 100% detection accuracy was achieved, which has not been reported when using conventional PCR. Overall, these findings indicate that the RCCS platform is a powerful tool for rapid and reliable detection of carbapenemase-encoding genes, with significant potential for implementation in point-of-care settings and resource-limited environments.

Evaluation of a near-patient SARS-CoV-2 novel rapid diagnostic platform.

The goal of this study is to test a novel device and methodology based on the "Pebble" platform and real-time quantitative colorimetric loop-mediated isothermal amplification (qcLAMP) during SARS-CoV-2 detection using crude samples and extracted RNA. The new method employs an inexpensive lightweight device aimed toward rapid point-of-care testing. An extensive evaluation was performed consisting of 1,693 clinical samples across five independent clinical testing centers. Positive colorimetric results were observed within 20 minutes of testing. At a 20-minute time-to-positive cut-off, the specificity is 98.5% with a diagnostic accuracy of 91.9%, compared to qPCR assays. Our findings indicate that the SARS-CoV-2 qcLAMP diagnostic assay in conjunction with the Pebble device is ideal for point-of-care/near-patient testing.IMPORTANCEHere, we describe our analyses and validation of a novel real-time quantitative colorimetric loop-mediated isothermal amplification (qcLAMP) device, available under the name "Pebble" and associated SARS-CoV-2 diagnostic qcLAMP assay for clinical diagnostic use. The analyses were performed in five independent testing sites across Europe using clinical samples from the associated clinical sites and support the use of "pebble" and associated kit in the diagnostic environment.

Lab-in-a-Vial Rapid Test for Internet of Things-Embedded Point-of-Healthcare Protein Biomarker Detection in Bodily Fluids.

Amateurs often struggle with detecting and quantifying protein biomarkers in body fluids due to the high expertise required. This study introduces a Lab-in-a-Vial (LV) rapid diagnostic platform, featuring hydrangea-like platinum nanozymes (PtNH), for rapid, accurate detection and quantification of protein biomarkers on-site within 15min. This method significantly enhances detection sensitivity for various biomarkers in body fluids, surpassing traditional methods such as enzyme-linked immunosorbent assays (ELISA) and lateral flow assays (LFA) by ≈250 to 1300 times. The LV platform uses a glass vial coated with specific bioreceptors such as antigens or antibodies, enabling rapid in vitro evaluation of disease risk from small fluid samples, similar to a personal ELISA-like point-of-care test (POCT). It overcomes challenges in on-site biomarker detection, allowing both detection and quantification through a portable wireless spectrometer for healthcare internet of things (H-IoT). The platform's effectiveness and adaptability are confirmed using IgG/IgM antibodies from SARS-CoV-2 infected patients and nuclear matrix protein (NMP22) from urothelial carcinoma (UC) patients as biomarkers. These tests demonstrated its accuracy and flexibility. This approach offers vast potential for diverse disease applications, provided that the relevant protein biomarkers in bodily fluids are identified.

Metagenomic sequencing in the diagnosis of resistance to antimicrobials

The purpose of the article is to analyze the literature on antimicrobial resistance and the metagenomic sequencing method. In total, 32 articles published in the years 2019–2024 and some previously published classic works were analyzed. Antimicrobial resistance (AMR) is one of the most serious threats to public health, animal health and environmental well-being globally, generating high costs. Metagenomic sequencing allows the detection of all genes in a sample, providing a complete picture of AMR. By sequencing it is possible to assess the location and transmissibility of the AMR genes. As surveillance networks and metagenomic sequencing, as one of the rapid diagnostic platforms, are implemented globally, it will be possible to detect and limit infectious outbreaks at a much earlier stage, thus saving lives and substantially reducing costs. In the near future, metagenomic sequencing will no longer be a luxury but a necessity as we engage in the ongoing fight against infectious diseases.

Clinical Utility of Blood Culture Identification 2 Panel in Flagged Blood Culture Samples from the Intensive Care Unit of a Tertiary Care Hospital.

The availability of rapid diagnostic platforms for positive blood cultures has accelerated the speed at which the clinical microbiology laboratory can identify the causative organism and facilitate early appropriate antimicrobial therapy. There is a paucity of data regarding the clinical utility of the blood culture identification 2 (BCID2) panel test and its correlation with phenotypic drug susceptibility testing (DST) in flagged blood culture bottles from intensive care units (ICUs) in countries such as India, which have high rates of multidrug-resistant gram-negative bacteria (MDR-GNB). We conducted a retrospective observational study in a tertiary care ICU on 200 patients above 18 years of age in whom a BCID2 test was ordered when blood cultures flagged positive. We found 99% concordance between BCID2 and cultures in the identification of bacteria and yeasts and 96.5% concordance between phenotypic and genotypic DST. Furthermore, BCID2 was available about 1.5 days earlier than conventional ID and DST and played a key role in tailoring antimicrobials in 82.5% of the patients. Polymyxin-based therapy was discontinued earlier after an empiric dose in 138 patients (69%) based on BCID2 reports. In critically ill patients with monomicrobial bacteremia, BCID2 rapidly identifies bacteria and antimicrobial resistance (AMR) genes and is significantly faster than conventional culture and sensitivity testing. Antibiotics were escalated in more than a third of patients and de-escalated in almost a fifth on the same day. We recommend that all ICUs routinely incorporate the test in their antibiotic decision-making process and in antimicrobial stewardship. Vineeth VK, Nambi PS, Gopalakrishnan R, Sethuraman N, Ramanathan Y, Chandran C, et al. Clinical Utility of Blood Culture Identification 2 Panel in Flagged Blood Culture Samples from the Intensive Care Unit of a Tertiary Care Hospital. Indian J Crit Care Med 2024;28(5):461-466.

The Role and Value of Professional Rapid Testing of Acute Respiratory Infections (ARIs) in Europe: A Special Focus on the Czech Republic, Poland, and Romania.

This review aims to explore the role of professional diagnostic rapid testing of acute respiratory infections (ARIs), especially COVID-19 and influenza, ensuring proper disease management and treatment in Europe, and particularly in Czech Republic, Poland, and Romania. The paper was constructed based on a review of scientific evidence and national and international policies and recommendations, as well as a process of validation by four experts. The development of new testing technologies, treatment options, and increased awareness of the negative multidimensional impact of ARI profiles transformed differential diagnosis into a tangible and desirable reality. This review covers the following topics: (1) the multidimensional impact of ARIs, (2) ARI rapid diagnostic testing platforms and their value, (3) the policy landscape, (4) challenges and barriers to implementation, and (5) a set of recommendations illustrating a path forward. The findings indicate that rapid diagnostic testing, including at the point of care (POC), can have a positive impact on case management, antimicrobial and antibiotic stewardship, epidemiological surveillance, and decision making. Integrating this strategy will require the commitment of governments and the international and academic communities, especially as we identified room for improvement in the access and expansion of POC rapid testing in the focus countries and the inclusion of rapid testing in relevant policies.

Establishment and application of a rapid molecular diagnostic platform for the isothermal visual amplification of group B Streptococcus based on recombinase polymerase.

With growing concerns about Group B streptococcal (GBS) infections and their adverse effects on perinatal pregnancies, including infection, premature delivery, neonatal septicemia, and meningitis, it is urgent to promote GBS screening at all pregnancy stages. The purpose of this study is to establish a device-independent, fast, sensitive, and visual GBS detection method. Taking advantage of the characteristics of the recombinase polymerase isothermal amplification (RPA), the activity of the nfo nuclease cleavage base analog (tetrahydrofuran, THF) site, and the advantages of visual reading of the lateral flow chromatography strip (LFS), a GBS detection method was developed. This method focused on the conservative region of the Christie-Atkins-Munch-Petersen factor encoded by the cfb gene, a virulence gene specific to GBS. Two forward primers, two biotin-labeled reverse primers, and one fluorescein isothiocyanate (FITC)-labeled and C3spacer-blocked probe were designed. The study involved optimizing the primer pair and probe combination, determining the optimal reaction temperature and time, evaluating specificity, analyzing detection limits, and testing the method on 87 vaginal swabs from perinatal pregnant women. The results showed that the visual detection method of GBS-RPA-LFS, using the cfb-F1/R2/P1 primer probe, could detect GBS within 15 min at the temperature ranging from 39°C to 42°C. Furthermore, the method specifically amplified only GBS, without cross-reacting with pathogens like Lactobacillus iners, Lactobacillus crispatus, Candida albicans, Listeria monocytogenes, Yersinia enterocolitica, Klebsiella Pneumoniae, Enterobacter cloacae, Citrobacter freundii, Vibrio alginolyticus, Vibrio parahaemolyticus, Salmonella typhimurium, Staphylococcus aureus, Pseudomonas aeruginosa, or Trichomonas vaginalis. It could detect a minimum of 100 copies per reaction. In clinical 98 samples of vaginal swabs from pregnant women, the agreement rate between the GBS-RPA-LFS method and TaqMan real-time fluorescence quantification method was 95.92%. In conclusion, this study successfully established a combined RPA and LFS GBS in situ detection platform, with short reaction time, high sensitivity, high specificity, portability, and device independence, providing a feasible strategy for clinical GBS screening.

VIDIIA Hunter diagnostic platform: a low-cost, smartphone connected, artificial intelligence-assisted COVID-19 rapid diagnostics approved for medical use in the UK.

Introduction: Accurate and rapid diagnostics paired with effective tracking and tracing systems are key to halting the spread of infectious diseases, limiting the emergence of new variants and to monitor vaccine efficacy. The current gold standard test (RT-qPCR) for COVID-19 is highly accurate and sensitive, but is time-consuming, and requires expensive specialised, lab-based equipment. Methods: Herein, we report on the development of a SARS-CoV-2 (COVID-19) rapid and inexpensive diagnostic platform that relies on a reverse-transcription loop-mediated isothermal amplification (RT-LAMP) assay and a portable smart diagnostic device. Automated image acquisition and an Artificial Intelligence (AI) deep learning model embedded in the Virus Hunter 6 (VH6) device allow to remove any subjectivity in the interpretation of results. The VH6 device is also linked to a smartphone companion application that registers patients for swab collection and manages the entire process, thus ensuring tests are traced and data securely stored. Results: Our designed AI-implemented diagnostic platform recognises the nucleocapsid protein gene of SARS-CoV-2 with high analytical sensitivity and specificity. A total of 752 NHS patient samples, 367 confirmed positives for coronavirus disease (COVID-19) and 385 negatives, were used for the development and validation of the test and the AI-assisted platform. The smart diagnostic platform was then used to test 150 positive clinical samples covering a dynamic range of clinically meaningful viral loads and 250 negative samples. When compared to RT-qPCR, our AI-assisted diagnostics platform was shown to be reliable, highly specific (100%) and sensitive (98-100% depending on viral load) with a limit of detection of 1.4 copies of RNA per µL in 30min. Using this data, our CE-IVD and MHRA approved test and associated diagnostic platform has been approved for medical use in the United Kingdom under the UK Health Security Agency's Medical Devices (Coronavirus Test Device Approvals, CTDA) Regulations 2022. Laboratory and in-silico data presented here also indicates that the VIDIIA diagnostic platform is able to detect the main variants of concern in the United Kingdom (September 2023). Discussion: This system could provide an efficient, time and cost-effective platform to diagnose SARS-CoV-2 and other infectious diseases in resource-limited settings.

Toehold switch plus signal amplification enables rapid detection.

Recent world events have led to an increased interest in developing rapid and inexpensive clinical diagnostic platforms for viral detection. Here, the development of a cell-free toehold switch-based biosensor, which does not require upstream amplification of target RNA, is described for the detection of RNA viruses. Toehold switches were designed to avoid interfering secondary structure in the viral RNA binding region, mutational hotspots, and cross-reacting sequences of other coronaviruses. Using these design criteria, toehold switches were targeted to a low mutation region of the SARS-CoV-2 genome nonstructural protein 2 (nsp2). The designs were tested in a cell-free system using trigger RNA based on the viral genome and a highly sensitive fluorescent reporter gene, mNeonGreen. The detection sensitivity of our best toehold design, CSU 08, was in the low picomolar range of target (trigger) RNA. To increase the sensitivity of our cell-free biosensor to a clinically relevant level, we developed a modular downstream amplification system that utilizes toehold switch activation of tobacco etch virus (TEV) protease expression. The TEV protease cleaves a quenched fluorescent reporter, both increasing the signal fold change between control and sample and increasing the sensitivity to a clinically relevant low femtomolar range for target RNA detection.

Highly Uniform Self-Assembly of Gold Nanoparticles by Butanol-Induced Dehydration and Its SERS Applications in SARS-CoV-2 Detection.

We report the development of a reproducible and highly sensitive surface-enhanced Raman scattering (SERS) substrate using a butanol-induced self-assembly of gold nanoparticles (AuNPs) and its application as a rapid diagnostic platform for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The butanol-induced self-assembly process was used to generate a uniform assembly of AuNPs, with multiple hotspots, to achieve high reproducibility. When an aqueous droplet containing AuNPs and target DNAs was dropped onto a butanol droplet, butanol-induced dehydration occurred, enriching the target DNAs around the AuNPs and increasing the loading density of the DNAs on the AuNP surface. The SERS substrate was evaluated by using Raman spectroscopy, which showed strong electromagnetic enhancement of the Raman signals. The substrate was then tested for the detection of SARS-CoV-2 using SERS, and a very low limit of detection (LoD) of 3.1 × 10-15 M was obtained. This provides sufficient sensitivity for the SARS-CoV-2 screening assay, and the diagnostic time is significantly reduced as no thermocycling steps are required. This study demonstrates a method for the butanol-induced self-assembly of AuNPs and its application as a highly sensitive and reproducible SERS substrate for the rapid detection of SARS-CoV-2. The results suggest the potential of this approach for developing rapid diagnostic platforms for other biomolecules and infectious diseases.

Rapid Diagnostic Platform for Personalized Vitamin B6 Detection in Erythrocytes via PLP Cofactor Mimics.

Personalized assessment of vitamin levels in point-of-care (POC) devices is urgently needed to advance the recognition of diseases associated with malnutrition and unbalanced diets. We here introduce a diagnostic platform, which showcases an easy and rapid readout of vitamin B6 (pyridoxal phosphate, PLP) levels in erythrocytes as a first step toward a home-use POC. The technology is based on fluorescent probes, which bind to PLP-dependent enzymes (PLP-DEs) and thereby indirectly report their occupancy with endogenous B6. For example, low vitamin levels result in high probe binding, yielding a strong signal and vice versa. Antibodies against signature human PLP-DEs were immobilized on microarrays to capture probe labeled enzymes for fluorescent detection. Calibrating the system with defined B6 levels revealed a concentration-depended readout as well as sufficient sensitivity for its detection in erythrocytes. To account for individual differences in protein expression, a second antibody was used to normalize protein abundance. This sandwiched assay correctly reported relative B6 levels in human erythrocyte samples, as confirmed by classical laboratory diagnostics. In principle, the platform layout can be easily expanded to other crucial vitamins beyond B6 via an analogous probe strategy.

Solid-Phase Optical Sensing Techniques for Sensitive Virus Detection.

Viral infections can pose a major threat to public health by causing serious illness, leading to pandemics, and burdening healthcare systems. The global spread of such infections causes disruptions to every aspect of life including business, education, and social life. Fast and accurate diagnosis of viral infections has significant implications for saving lives, preventing the spread of the diseases, and minimizing social and economic damages. Polymerase chain reaction (PCR)-based techniques are commonly used to detect viruses in the clinic. However, PCR has several drawbacks, as highlighted during the recent COVID-19 pandemic, such as long processing times and the requirement for sophisticated laboratory instruments. Therefore, there is an urgent need for fast and accurate techniques for virus detection. For this purpose, a variety of biosensor systems are being developed to provide rapid, sensitive, and high-throughput viral diagnostic platforms, enabling quick diagnosis and efficient control of the virus's spread. Optical devices, in particular, are of great interest due to their advantages such as high sensitivity and direct readout. The current review discusses solid-phase optical sensing techniques for virus detection, including fluorescence-based sensors, surface plasmon resonance (SPR), surface-enhanced Raman scattering (SERS), optical resonators, and interferometry-based platforms. Then, we focus on an interferometric biosensor developed by our group, the single-particle interferometric reflectance imaging sensor (SP-IRIS), which has the capability to visualize single nanoparticles, to demonstrate its application for digital virus detection.

Ultra‐Fast Photonic Digital Polymerase Chain Reaction based on N‐Heterocyclic Carbene Self‐Assembled Monolayer

AbstractA molecular diagnosis of the respiratory syncytial virus (RSV) without bulky and expensive instrumentation is of great importance for the early detection and prevention in a fast‐spreading pandemic. However, the current representative diagnostic methods have the limitation of being time‐consuming, cost, the processing time for polymerase chain reaction (PCR), and inaccurate for lateral flow assay (LFA), representatively. Herein, an integrated photonic digital PCR (dPCR) is developed with high‐velocity photonic scanner for in situ fluorescence detection by introducing the N‐heterocyclic carbene self‐assembled monolayer‐based Au film to prevent the quenching effect. The on‐site rapid molecular diagnostic platform shows the driving of 40 cycles in under 8 min and fluorescence scanning in under 7 min, resulting in a total analysis time within 15 min. In particular, the technology clearly demonstrates the classification of SARS‐CoV‐2 patients and healthy controls (99% in sensitivity, 98.6% in specificity, and 96.4% in accuracy with RdRp gene), comparing with standard RT‐qPCR. This platform can be utilized for prompt point‐of‐care molecular diagnostics in early diagnosis and large‐scale prevention of next pandemic spreading for upcoming infectious diseases and for the distinction diagnosis with other RSV.

Enhancement in the limit of detection of lab‐on‐chip microfluidic devices using functional nanomaterials

AbstractTechnological developments in recent years have witnessed a paradigm shift towards lab‐on‐chip devices for various diagnostic applications. Lab‐on‐chip technology integrates several functions typically performed in a large‐scale analytical laboratory on a small‐scale platform. These devices are more than the miniaturized versions of conventional analytical and diagnostic techniques. The advances in fabrication techniques, material sciences, surface modification strategies, and their integration with microfluidics and chemical and biological‐based detection mechanisms have enormously enhanced the capabilities of these devices. The minuscule sample and reagent requirements, capillary‐driven pump‐free flows, faster transport phenomena, and ease of integration with various signal readout mechanisms make these platforms apt for use in resource‐limited settings, especially in developing and underdeveloped parts of the world. The microfluidic lab‐on‐a‐chip technology offers a promising approach to developing cost‐effective and sustainable point‐of‐care testing applications. Numerous merits of this technology have attracted the attention of researchers to develop low‐cost and rapid diagnostic platforms in human healthcare, veterinary medicine, food quality testing, and environmental monitoring. However, one of the major challenges associated with these devices is their limited sensitivity or the limit of detection. The use of functional nanomaterials in lab‐on‐chip microfluidic devices can improve the limit of detection by enhancing the signal‐to‐noise ratio, increasing the capture efficiency, and providing capabilities for devising novel detection schemes. This review presents an overview of state‐of‐the‐art techniques for integrating functional nanomaterials with microfluidic devices and discusses the potential applications of these devices in various fields.

Multipurpose advanced split T7 promoter-based transcription amplification for ultrasensitive molecular diagnostics

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.

Colorimetric detection of SARS‐CoV‐2 variants with paper‐based analytical devices

Colorimetric detection of SARS‐CoV‐2 variants with paper‐based analytical devices

A novel highly sensitive compilation-detachment fluorescence sensing strategy based on RNA-cleavage DNAzyme for MDA-MB-231 breast cancer biomarker determination.

Herein, we designed a novel and highly sensitive fluorescence multicomponent detachable platform for MDA-MB-231 breast cancer cell detection as a model. The RNA cleavage DNAzyme was used as a central operator of the multicomponent probe through which compilation and induced detachment of probe was done. During the compilation step, the dsDNA-Sybr green 1 complexes on gold nanoparticles (GNP@dsDNA@SG1) were assembled. The intercalated Sybr green in the DNA structure has been used as an amplified signal generator on one site of DNAzyme and magnetic nanoparticles (MNP) act as a biological carrier and probe collector on the opposite side. The enzyme activator co-factor (MDA-MB-231 cell cytoplasmic protein) provokes the activation of the catalytic core of enzyme sequence in the DNAzyme molecule, followed by cleavage reaction in the substrate sequence and releasing GNP@ dsDNA@SG1 into the solution. The results indicate that the Sybr green emission fluorescence (520 nm) increases with the increment of MDA-MB-231 protein concentration in the linear dynamic range of 8.10 × 10-2 to 1.95 ng ml-1 (0.77 × 10-3-0.019 cell ml-1) with a detection limit (LOD) of 1/72 × 10-2 pg ml-1 under optimal conditions. The proposed immunosensor has great potential in developing ultrasensitive and rapid diagnostic platforms.

968. The Impact of Film Array Detection of High-risk and Resistant Pathogens on the Time to Optimal Antibiotic Therapy at a Community Health System

Abstract Background The importance of effective antibiotic therapy for the treatment of bloodstream infections has been well described. Rapid diagnostic testing platforms like the BioFire® FilmArray® blood culture identification (BCID) have the potential to decrease time to effective antibiotics when acted on appropriately. The Cone Health stewardship team has collaborated with clinical pharmacists and infectious diseases physicians to define a process to optimize antibiotic therapy. This process involves microbiology calling a pharmacist with newly positive blood cultures, and the pharmacist notifies the attending physician with the results and evidence-based treatment recommendations. We transitioned from BCID to BCID2 in August, 2021. As CTX-M (marker for extended spectrum beta-lactamase production) was not a target on BCID, we were interested in how the addition of this marker could improve care in this subset of patients. Procedure for Positive Blood Culture Workup and Notification Treatment Recommendations for Selected Organisms Methods From August 2021 - March 2022, the times to administration of effective and optimal antibiotics were evaluated in patients with the following BCID2 organism and resistance gene targets identified: S. aureus, P. aeruginosa, CTX-M, KPC, OXA-48 like, IMP, VIM, NDM, and vancomycin resistant enterococci. An effective antibiotic was defined as an antibiotic with in vitro activity to the organism, and an optimal antibiotic was defined as the preferred agent based on institutional guidelines. For CTX-M isolates, we compared effective and optimal times to a historical control of ceftriaxone resistant enterobacterales and analyzed by student’s t-test. Results The combined mean time from BCID2 to administration of effective antibiotics = 1.2 hours (range 0-7.9 hours) and to optimal therapy =7.6 hours (range 0 – 113.8 hours). Prior to the BCID2 transition, patients found to be growing ceftriaxone resistant enterobacterales had an average time to optimal antibiotic administration of 17.7 hours, which improved to 2.8 hours after BCID2 implementation (p=0.0041). Time to Effective and Optimal Antibiotics after implementation of BCID2 (August 2021 – March 2022) Time to Effective and Optimal Antibiotics for Ceftriaxone Resistant Enterobacterales Prior to BCID2 Implementation (January 2021 - July 2021) Conclusion BioFire® FilmArray® coupled with antibiotic stewardship team support can help optimize time to effective and optimal antibiotic therapy in bloodstream infection. The addition of the CTX-M target to BCID2 was a helpful addition to the panel which allowed for decreased time to optimal antibiotic therapy. Disclosures All Authors: No reported disclosures.

Integrated microdroplet array platform with temperature controller and micro-stirring for ultra-fast SARS-CoV-2 detection

The outbreak of COVID-19 has created a huge challenge to global health systems. Experience in fighting the epidemic shows that the development of a rapid and sensitive POCT diagnostic platform for SARS-CoV-2 that can be deployed in situ is crucial to contain the outbreak. Here, we have developed a portable microdroplet detection platform that integrated temperature controller and micro-stirring for high-throughput and ultrafast COVID-19 diagnosis. Such a device uses a p-n junction (PN junction) as the temperature controller to adjust the temperature in a single microdroplet independently and precisely, ensuring the amplification of reverse transcription loop-mediated isothermal amplification (RT-LAMP). Meanwhile, the platform incorporates an ultrasonic micro-stirring unit, greatly increasing the interaction between RT-LAMP molecules and accelerating the amplification. The results show good linearity over a wide linear range (1 to 105 copies/μL) and low LOD (0.48 copy/μL). Our method reports in only 6.1 min for high-viral load samples, and combines with sample preparation, the total detection process could be done within 30 min. Such a portable and fully integrated microdroplet molecular diagnostic platform is a promising tool for point-of-care diagnosis of COVID-19 and other infectious diseases in resource-limited settings.

Ultrasensitive electrochemical detection of hepatitis b virus surface antigen based on hybrid nanomaterials

This study investigates a sandwich-type electrochemical immunosensor for the determination of Hepatitis B Virus Surface Antigen (HBsAg) as the biological target. Capture antibody (Ab1) was immobilized on the synthesized magnetic nanoparticles coated with silica and APTES (MNP@SiO2@APTES). A reporter antibody (Ab2) was assembled on gold nanoparticle-Thionine (GNP), which made use of Th as a redox probe for electrochemical detection. Each gold nanoparticle holds a lot of thionine molecules on its surface. Therefore, after the formation of a sandwich-type probe a magnified electrical signal is achieved. On the other hand, a magnet has been implemented in the electrode. Thus, Magnetic nanoparticle (MNP) acts as the probe collector and fixator component on the electrode's surface, assuring a strong and prolonged interaction of the ultimate sandwich-type probe with the electrode. Under optimal conditions, the current signal increased with the increment of HBsAg concentration in the linear dynamic range of 0.001 fg/ml-2 ng/ml with a detection limit (LOD) of 0.3ag/ml (S/N = 3). Furthermore, the assembled immunosensor successfully quantified the HBsAg concentration in serum. The successful results imply the high potential of the proposed immunosensor in developing ultrasensitive and rapid diagnostic platforms.