Amplification-free detection of SARS-CoV-2 with CRISPR-Cas13a and mobile phone microscopy.

The December 2019 outbreak of a novel respiratory virus, SARS-CoV-2, has become an ongoing global pandemic due in part to the challenge of identifying symptomatic, asymptomatic, and pre-symptomatic carriers of the virus. CRISPR diagnostics can augment gold-standard PCR-based testing if they can be made rapid, portable, and accurate. Here, we report the development of an amplification-free CRISPR-Cas13a assay for direct detection of SARS-CoV-2 from nasal swab RNA that can be read with a mobile phone microscope. The assay achieved ∼100 copies/µL sensitivity in under 30 min of measurement time and accurately detected pre-extracted RNA from a set of positive clinical samples in under 5 min. We combined crRNAs targeting SARS-CoV-2 RNA to improve sensitivity and specificity and directly quantified viral load using enzyme kinetics. Integrated with a reader device based on a mobile phone, this assay has the potential to enable rapid, low-cost, point-of-care screening for SARS-CoV-2.

Low-Field, Low-Cost, Point-of-Care Magnetic Resonance Imaging.

Low-field magnetic resonance imaging (MRI) has recently experienced a renaissance that is largely attributable to the numerous technological advancements made in MRI, including optimized pulse sequences, parallel receive and compressed sensing, improved calibrations and reconstruction algorithms, and the adoption of machine learning for image postprocessing. This new attention on low-field MRI originates from a lack of accessibility to traditional MRI and the need for affordable imaging. Low-field MRI provides a viable option due to its lack of reliance on radio-frequency shielding rooms, expensive liquid helium, and cryogen quench pipes. Moreover, its relatively small size and weight allow for easy and affordable installation in most settings. Rather than replacing conventional MRI, low-field MRI will provide new opportunities for imaging both in developing and developed countries. This article discusses the history of low-field MRI, low-field MRI hardware and software, current devices on the market, advantages and disadvantages, and low-field MRI's global potential.

Critical Advances for Democratizing Ultrasound Diagnostics in Human and Veterinary Medicine.

The democratization of ultrasound imaging refers to the process of making ultrasound technology more accessible. Traditionally, ultrasound imaging has been predominately used in specialized medical facilities by trained professionals. Advancements in technology and changes in the health-care landscape have inspired efforts to broaden the availability of ultrasound imaging to various settings such as remote and resource-limited areas. In this review, we highlight several key factors that have contributed to the ongoing democratization of ultrasound imaging, including portable and handheld devices, recent advancements in technology, and training and education. Examples of diagnostic point-of-care ultrasound (POCUS) imaging used in emergency and critical care, gastroenterology, musculoskeletal applications, and other practices are provided for both human and veterinary medicine. Open challenges and the future of POCUS imaging are presented, including the emerging role of artificial intelligence in technology development.

Clinical and laboratory diagnosis of Mayaro virus (MAYV): Current status and opportunities for further development.

The recent outbreaks related to Mayaro virus (MAYV) infection in the Americas have brought this neglected virus as a potential threat to global public health. Given the range of symptoms that can be associated with MAYV infection, it can be challenging to diagnose individuals based on clinical signs, especially in countries with simultaneous circulation of other mosquito-borne viruses, such as dengue virus (DENV) and chikungunya virus (CHIKV). With this challenge in mind, laboratory-based diagnosis assumes a critical role in the introduction of measures to help prevent virus dissemination and to adequately treat patients. In this review, we provide an overview of the clinical features reported in infected patients and currently available laboratory tools that are used for MAYV diagnosis, discussing their advances, advantages, and limitations to apply in the field. Moreover, we explore novel point-of-care (PoC) diagnostic platforms that can provide de-centralised diagnostics for use in areas with limited laboratory infrastructure.

Comparable performance of antigen-detecting rapid test by healthcare worker-collected and self-collected swabs for SARS-CoV-2 diagnostic: A systematic review and meta-analysis.

Usage of self-screening tests has become increasingly relevant in public health perspective for early detection of SARS-CoV-2 infection in the transitioning era of the COVID-19 pandemic into an endemic. This study was designed to compare the diagnostic accuracy of self-conducted and health professional-conducted SARS-CoV-2 rapid antigen tests (Ag-RDTs) and whether the sample was taken from anterior nasal or nasal mid-turbinate. Eligible comparative Ag-RDTs accuracy studies were retrieved from electronic databases systematically, in accordance with PRISMA. Selected studies were assessed for risk of bias using QUADAS-2 and QUADAS-C. In total, we selected five out of 1952 studies retrieved using the keywords. The overall sensitivity for the self-collected nasal swab method and healthcare worker-collected nasopharyngeal swab method was 79% (95% CI 68-87; I2  = 62%) and 83% (95% CI 75-89; I2  = 32%), respectively, which was not statistically different (p = 0.499). Nasal mid-turbinate swabs have a significantly higher sensitivity compared to anterior nasal swabs (p < 0.01). Both sampling methods represent high and comparable specificity values of 98% (95% CI 97-99; I2  = 0%) and 99% (95% CI 98-99; I2  = 0%). Positive predictive value (range 90%-99%) and negative predictive value (range 87%-98%) were equivalent for both methods. Our findings indicated the accuracy of self-collected Ag-RDT on nasal swabs was comparable to those performed by healthcare worker-collected on nasopharyngeal swabs. Self-collected Ag-RDT could be considered as a transmission prevention method in the transition of COVID-19 pandemic.

Smartphone-based low-cost and rapid quantitative detection of urinary creatinine with the Tyndall effect.

This research aims to develop a robust and quantitative method for measuring creatinine levels by harnessing the enhanced Tyndall effect (TE) phenomenon. The envisioned sensing assay is designed for practical deployment in resource-limited settings or homes, where access to advanced laboratory facilities is limited. Its primary objective is to enable regular and convenient monitoring of renal healthcare, particularly in cases involving elevated creatinine levels. The creatinine sensing strategy is achieved based on the aggregation of gold nanoparticles (AuNPs) triggered via the direct crosslinking reaction between creatinine and AuNPs, where an inexpensive laser pointer was used as a handheld light source and a smartphone as a portable device to record the TE phenomenon enhanced by the creatinine-induced aggregation of AuNPs. After evaluation and optimization of parameters such as AuNP concentrations and TE measurement time, the subsequent proof-of-concept experiments demonstrated that the average gray value change of TE images was linearly related to the logarithm of creatinine concentrations in the range of 1-50 µM, with a limit of detection of 0.084 µM. Meanwhile, our proposed creatinine sensing platform exhibited highly selective detection in complex matrix environments. Our approach offers a straightforward, cost-effective, and portable means of creatinine detection, presenting an encouraging signal readout mechanism suitable for point-of-care (POC) applications. The utilization of this assay as a POC solution exhibits potential for expediting timely interventions and enhancing healthcare outcomes among individuals with renal health issues.

Point-of-care high-sensitivity cardiac troponin in suspected acute myocardial infarction assessed at baseline and 2†h.

BACKGROUND AND AIMS: Strategies to assess patients with suspected acute myocardial infarction (AMI) using a point-of-care (POC) high-sensitivity cardiac troponin I (hs-cTnI) assay may expedite emergency care. A 2-h POC hs-cTnI strategy for emergency patients with suspected AMI was derived and validated. METHODS: In two international, multi-centre, prospective, observational studies of adult emergency patients (1486 derivation cohort and 1796 validation cohort) with suspected AMI, hs-cTnI (Siemens Atellica® VTLi) was measured at admission and 2 h later. Adjudicated final diagnoses utilized the hs-cTn assay in clinical use. A risk stratification algorithm was derived and validated. The primary diagnostic outcome was index AMI (Types 1 and 2). The primary safety outcome was 30-day major adverse cardiac events incorporating AMI and cardiac death. RESULTS: Overall, 81 (5.5%) and 88 (4.9%) patients in the derivation and validation cohorts, respectively, had AMI. The 2-h algorithm defined 66.1% as low risk with a sensitivity of 98.8% [95% confidence interval (CI) 89.3%-99.9%] and a negative predictive value of 99.9 (95% CI 99.2%-100%) for index AMI in the derivation cohort. In the validation cohort, 53.3% were low risk with a sensitivity of 98.9% (95% CI 92.4%-99.8%) and a negative predictive value of 99.9% (95% CI 99.3%-100%) for index AMI. The high-risk metrics identified 5.4% of patients with a specificity of 98.5% (95% CI 96.6%-99.4%) and a positive predictive value of 74.5% (95% CI 62.7%-83.6%) for index AMI. CONCLUSIONS: A 2-h algorithm using a POC hs-cTnI concentration enables safe and efficient risk assessment of patients with suspected AMI. The short turnaround time of POC testing may support significant efficiencies in the management of the large proportion of emergency patients with suspected AMI.

Bimetallic Nanozyme: A Credible Tag for In Situ-Catalyzed Reporter Deposition in the Lateral Flow Immunoassay for Ultrasensitive Cancer Diagnosis.

The lateral flow immunoassay (LFIA) is a sought-after point-of-care testing platform, yet the insufficient sensitivity of the LFIA limits its application in the detection of tumor biomarkers. Here, a colorimetric signal amplification method, bimetallic nanozyme-mediated in situ-catalyzed reporter deposition (BN-ISCRD), was designed for ultrasensitive cancer diagnosis. The bimetallic nanozyme used, palladium@iridium core-shell nanoparticles (Pd@Ir NPs), had ultrahigh enzyme-like activity, which was further explained by the electron transfer of Pd@Ir NPs and the change in the Gibbs free energy during catalysis through density functional theory calculations. With gastric cancer biomarkers pepsinogen I and pepsinogen II as model targets, this assay could achieve a cutoff value of 10 pg/mL, which was 200-fold lower than that without signal enhancement. The assay was applied to correctly identify 8 positive and 28 negative clinical samples. Overall, this BN-ISCRD-based LFIA showed great merits and potential in the application of ultrasensitive disease diagnosis.

Ultrasensitive and Rapid Detection of Procalcitonin via Waveguide-Enhanced Nanogold-Linked Immunosorbent Assay for Early Sepsis Diagnosis.

Sepsis, a life-threatening inflammatory response, demands economical, accurate, and rapid detection of biomarkers during the critical "golden hour" to reduce the patient mortality rate. Here, we demonstrate a cost-effective waveguide-enhanced nanogold-linked immunosorbent assay (WENLISA) based on nanoplasmonic waveguide biosensors for the rapid and sensitive detection of procalcitonin (PCT), a sepsis-related inflammatory biomarker. To enhance the limit of detection (LOD), we employed sandwich assays using immobilized capture antibodies and detection antibodies conjugated to gold nanoparticles to bind the target analyte, leading to a significant evanescent wave redistribution and strong nanoplasmonic absorption near the waveguide surface. Experimentally, we detected PCT for a wide linear response range of 0.1 pg/mL to 1 ng/mL with a record-low LOD of 48.7 fg/mL (3.74 fM) in 8 min. Furthermore, WENLISA has successfully identified PCT levels in the blood plasma of patients with sepsis and healthy individuals, offering a promising technology for early sepsis diagnosis.

Toward the Scalable, Rapid, Reproducible, and Cost-Effective Synthesis of Personalized Nanomedicines at the Point of Care.

Organic nanoparticles are used in nanomedicine, including for cancer treatment and some types of COVID-19 vaccines. Here, we demonstrate the scalable, rapid, reproducible, and cost-effective synthesis of three model organic nanoparticle formulations relevant to nanomedicine applications. We employed a custom-made, low-cost fluid mixer device constructed from a commercially available three-dimensional printer. We investigated how systematically changing aqueous and organic volumetric flow rate ratios determined liposome, polymer nanoparticle, and solid lipid nanoparticle sizes, size distributions, and payload encapsulation efficiencies. By manipulating inlet volumes, we synthesized organic nanoparticles with encapsulation efficiencies approaching 100% for RNA-based payloads. The synthesized organic nanoparticles were safe and effective at the cell culture level, as demonstrated by various assays. Such cost-effective synthesis approaches could potentially increase the accessibility to clinically relevant organic nanoparticle formulations for personalized nanomedicine applications at the point of care, especially in nonhospital and low-resource settings.

Point of care testing for hepatitis C in the priority settings of mental health, prisons and drug & alcohol facilities - the PROMPt Study.

BACKGROUND: A barrier to hepatitis C virus (HCV) cure is conventional testing. The aim of this study was to evaluate the effect of HCV antibody and RNA point-of-care-testing (POCT) on testing rates, linkage to care, treatment and acceptability of testing in three priority settings in Australia. METHODS: Participants were enrolled in an interventional cohort study at a reception prison, inpatient mental health service (MHS), and inpatient alcohol and other drug (AOD) unit-between October 2020 and December 2021. HCV POCT was performed using SD Bioline HCV antibody fingerstick test and a reflexive Xpert® HCV Viral Load Fingerstick test using capillary blood samples. A retrospective audit of HCV testing and treatment data was performed at each site for the preceding 12-month period to generate a historical control. RESULTS: 1,549 participants received a HCV antibody test with 17% (264/1,549) receiving a positive result, of which 21% (55/264) tested HCV RNA positive. Across all settings the rate of testing per year significantly increased between the historical controls and the study intervention period by three-fold (RR:2.57 95% CI: 2.32, 2.85) for HCV antibody testing and four-fold (RR:1.62; 95% CI:1.31, 2.01) for RNA testing. Treatment uptake was higher during the POCT intervention (86%, 47/55; P=0.010) compared to the historical controls (61%, 27/44). CONCLUSIONS: This study demonstrated across three settings that the use of HCV antibody and RNA POCT increased testing rates, treatment uptake linkage to care. The testing model was highly acceptable for most participants. CLINICAL TRIAL REGISTRATION: ACTRN-12621001578897.

Point-of-care ultrasound in infectious diseases: Current insights and future perspectives.

Point-of-Care Ultrasound (POCUS) is a safe, non-invasive technique performed at the patient's bedside, providing immediate results to the operator. It complements physical examination and facilitates clinical decision-making. In infectious diseases, POCUS is particularly valuable, offering an initial assessment in cases of suspected infection. It often leads to an early tentative diagnosis enabling the prompt initiation of antimicrobial treatment without the delay associated with traditional radiology. POCUS provides direct visualization of affected organs, assists in evaluating fluid balance and facilitates various interventions, all while reducing patient discomfort. For infectious disease specialists, becoming proficient in POCUS is a critical future challenge, requiring dedicated training for effective utilization.

Upconversion Nanoparticles Based Sensing: From Design to Point-of-Care Testing.

Rare earth-doped upconversion nanoparticles (UCNPs) have achieved a wide range of applications in the sensing field due to their unique anti-Stokes luminescence property, minimized background interference, excellent biocompatibility, and stable physicochemical properties. However, UCNPs-based sensing platforms still face several challenges, including inherent limitations from UCNPs such as low quantum yields and narrow absorption cross-sections, as well as constraints related to energy transfer efficiencies in sensing systems. Therefore, the construction of high-performance UCNPs-based sensing platforms is an important cornerstone for conducting relevant research. This work begins by providing a brief overview of the upconversion luminescence mechanism in UCNPs. Subsequently, it offers a comprehensive summary of the sensors' types, design principles, and optimized design strategies for UCNPs sensing platforms. More cost-effective and promising point-of-care testing applications implemented based on UCNPs sensing systems are also summarized. Finally, this work addresses the future challenges and prospects for UCNPs-based sensing platforms.

Kiwi-Inspired Rational Nanoarchitecture with Intensified and Discrete Magneto-Fluorescent Functionalities for Ultrasensitive Point-of-Care Immunoassay.

Fluorescent lateral flow immunoassays (FLFIA) is a well-established rapid detection technique for quantitative analysis. However, achieving accurate analysis of biomarkers at the pg mL-1 level using FLFIA still poses challenges. Herein, an ultrasensitive FLFIA platform is reported utilizing a kiwi-type magneto-fluorescent silica nanohybrid (designated as MFS) that serves as both a target-enrichment substrate and an optical signal enhancement label. The spatially-layered architecture comprises a Fe3O4 core, an endocarp-fibers like dendritic mesoporous silica, seed-like quantum dots, and a kiwi-flesh like silica matrix. The MFS demonstrates heightened fluorescence brightness, swift magnetic response, excellent size uniformity, and dispersibility in water. Through liquid-phase capturing and fluorescence-enhanced signal amplification, as well as magnetic-enrichment sample amplification and magnetic-separation noise reduction, the MFS-based FLFIA is successfully applied to the detection of cardiac troponin I that achieved a limit of detection at 8.4 pg mL-1, tens of times lower than those of previously published fluorescent and colorimetric lateral flow immunoassays. This work offers insights into the strategic design of magneto-fluorescent synergetic signal amplification on LFIA platform and underscores their prospects in high-sensitive rapid and on-site diagnosis of biomarkers.

Electrochemical Paper-Based Microfluidics: Harnessing Capillary Flow for Advanced Diagnostics.

Electrochemical paper-based microfluidics has attracted much attention due to the promise of transforming point-of-care diagnostics by facilitating quantitative analysis with low-cost and portable analyzers. Such devices harness capillary flow to transport samples and reagents, enabling bioassays to be executed passively. Despite exciting demonstrations of capillary-driven electrochemical tests, conventional methods for fabricating electrodes on paper impede capillary flow, limit fluidic pathways, and constrain accessible device architectures. This account reviews recent developments in paper-based electroanalytical devices and offers perspective by revisiting key milestones in lateral flow tests and paper-based microfluidics engineering. The study highlights the benefits associated with electrochemical sensing and discusses how the detection modality can be leveraged to unlock novel functionalities. Particular focus is given to electrofluidic platforms that embed electrodes into paper for enhanced biosensing applications. Together, these innovations pave the way for diagnostic technologies that offer portability, quantitative analysis, and seamless integration with digital healthcare, all without compromising the simplicity of commercially available rapid diagnostic tests.

At-home testing for respiratory viruses: a minireview of the current landscape.

The landscape of at-home testing using over-the-counter (OTC) tests has been evolving over the last decade. The United States Food and Drug Administration Emergency Use Authorization rule has been in effect since the early 2000s, and it was widely employed during the severe acute respiratory syndrome coronavirus 2 pandemic to authorize antigen and nucleic acid detection tests for use in central laboratories as well as OTC. During the pandemic, the first at-home tests for respiratory viruses became available for consumer use, which opened the door for additional respiratory virus OTC tests. Concerns may exist regarding the public's ability to properly collect samples, perform testing, interpret results, and report results to public health authorities. However, favorable comparison studies between OTC testing and centralized laboratory test results suggest that OTC testing may have a place in healthcare, and it is likely here to stay. This mini-review of OTC tests for viral respiratory diseases will briefly cover the regulatory and reimbursement environment, current OTC test availability, as well as the advantages and limitations of OTC tests.

Q3 lab-on-chip real-time PCR for the diagnosis of Leishmania infantum infection in dogs.

Leishmaniasis is a vector-borne disease caused by many Leishmania spp. which infect humans and other mammalian hosts. Leishmania infantum is the main agent of canine leishmaniasis (CanL) whose diagnosis is usually confirmed by serological and molecular tests. This study aimed to evaluate the clinical and analytical sensitivities of a lab-on-chip (LOC) real-time PCR applied on the portable Q3-Plus V2 platform (Q3 qPCR) in the detection of L. infantum. The Q3 qPCR performance was assessed by comparing the quantification cycle (Cq) values with those obtained using the qPCR run on a CFX96 Real-Time System (CFX96 qPCR). A total of 173 DNA samples (extracted from bone marrow, lymph node, blood, buffy coat, conjunctival swab, and skin) as well as 93 non-extracted samples (NES) (bone marrow, lymph node, blood, and buffy coat) collected from dogs were tested with both systems. Serial dilutions of each representative DNA and NES sample were used to assess the analytical sensitivity of the Q3 qPCR system. Overlapping Cq values were obtained with the Q3 qPCR and CFX96 qPCR, both using DNA extracted from L. infantum promastigotes (limit of detection, <1 promastigote per milliliter) and from biological samples as well as with NES. However, the Q3 qPCR system showed a higher sensitivity in detecting L. infantum in NES as compared with the CFX96 qPCR. Our data indicate that the Q3 qPCR system could be a reliable tool for detecting L. infantum DNA in biological samples, bypassing the DNA extraction step, which represents an advance in the point-of-care diagnostic of CanL.

A simple and sensitive test for Candida auris colonization, surveillance, and infection control suitable for near patient use.

Candida auris is a multidrug-resistant fungal pathogen with a propensity to colonize humans and persist on environmental surfaces. C. auris invasive fungal disease is being increasingly identified in acute and long-term care settings. We have developed a prototype cartridge-based C. auris surveillance assay (CaurisSurV cartridge; "research use only") that includes integrated sample processing and nucleic acid amplification to detect C. auris from surveillance skin swabs in the GeneXpert instrument and is designed for point-of-care use. The assay limit of detection (LoD) in the skin swab matrix was 10.5 and 14.8 CFU/mL for non-aggregative (AR0388) and aggregative (AR0382) strains of C. auris, respectively. All five known clades of C. auris were detected at 2-3-5× (31.5-52.5 CFU/mL) the LoD. The assay was validated using a total of 85 clinical swab samples banked at two different institutions (University of California Los Angeles, CA and Wadsworth Center, NY). Compared to culture, sensitivity was 96.8% (30/31) and 100% (10/10) in the UCLA and Wadsworth cohorts, respectively, providing a combined sensitivity of 97.5% (40/41), and compared to PCR, the combined sensitivity was 92% (46/50). Specificity was 100% with both clinical (C. auris negative matrix, N = 31) and analytical (non-C. auris strains, N = 32) samples. An additional blinded study with N = 60 samples from Wadsworth Center, NY yielded 97% (29/30) sensitivity and 100% (28/28) specificity. We have developed a completely integrated, sensitive, specific, and 58-min prototype test, which can be used for routine surveillance of C. auris and might help prevent colonization and outbreaks in acute and chronic healthcare settings. IMPORTANCE: This study has the potential to offer a better solution to healthcare providers at hospitals and long-term care facilities in their ongoing efforts for effective and timely control of Candida auris infection and hence quicker response for any potential future outbreaks.

A Lateral Flow Assay Based on Streptavidin-biotin Amplification System with Recombinase Polymerase Amplification for Rapid and Quantitative Detection of Salmonella enteritidis.

Salmonella constitutes a prevalent alimentary pathogen, instigating zoonotic afflictions. Consequently, the prompt discernment of Salmonella in sustenance is of cardinal significance. Lateral flow assays utilizing colorimetric methodologies adequately fulfill the prerequisites of point-of-care diagnostics, however, their detection threshold remains elevated, generally permitting only qualitative discernment, an impediment to the preliminary screening of nascent pathogens. In response to this conundrum, we propose a lateral flow diagnostic predicated upon a streptavidin-biotin amplification system with recombinase polymerase amplification engineered for the expeditious and quantitative discernment of Salmonella enteritidis. Trace nucleic acids within a sample undergo exponential amplification via recombinase polymerase amplification to a level discernable, constituting the initial signal amplification. Subsequently, along the test line (T-line) of the lateral flow strip, the chromatic signal undergoes augmentation by securing a greater quantity of AuNPs through the magnification capacity of the streptavidin-biotin mechanism, affecting the second signal amplification. Quantitative results are procured via smartphone capture and transferred to computer software for precise calculation of the targeted quantity. The lateral flow strip exhibits a LOD at 19.41 CFU/mL for cultured S. enteritidis. The RSD of three varying concentrations were respectively 3.74 %, 5.96 %, and 4.25 %.

Inherited disorders of hemoglobin: A review of old and new diagnostic methods.

The genetic regulation of hemoglobin is complex and there are a number of genetic abnormalities that result in clinically important hemoglobin disorders. Here, we review the molecular pathophysiology of hemoglobin disorders and review both old and new methods of diagnosing these disorders. Timely diagnosis of hemoglobinopathies in infants is essential to coordinate optimal life-saving interventions, and accurate identification of carriers of deleterious mutations allows for genetic counseling and informed family planning. The initial laboratory workup of inherited disorders of hemoglobin should include a complete blood count (CBC) and peripheral blood smear, followed by carefully selected tests based on clinical suspicion and available methodology. We discuss the utility and limitations of the various methodologies to fractionate hemoglobin, including cellulose acetate and citrate agar hemoglobin electrophoresis, isoelectric focusing, high-resolution high-performance liquid chromatography, and capillary zone electrophoresis. Recognizing that most of the global burden of hemoglobin disorders exists in low- and middle-income countries, we review the increasingly available array of point-of-care-tests (POCT), which have an increasingly important role in expanding early diagnosis programs to address the global burden of sickle cell disease, including Sickle SCAN, HemoTypeSC, Gazelle Hb Variant, and Smart LifeLC. A comprehensive understanding of the molecular pathophysiology of hemoglobin and the globin genes, as well as a clear understanding of the utility and limitations of currently available diagnostic tests, is essential in reducing global disease burden.