Diagnosis Of Virus Research Articles

USAGE OF PARTICLE SWARM OPTIMIZATION IN DIGITAL IMAGES SELECTION FOR MONKEYPOX VIRUS PREDICTION AND DIAGNOSIS

Identifying skin diseases by using digital images of skin that are also automated, efficient, and accurate is critical for biomedical image analysis. Many researchers have developed numerous machine-learning techniques for the prediction and diagnosis of various diseases that help clinicians identify infections early and provide crucial data for virus management. In this work, we use the inherent attributes of Particle Swarm Optimization (PSO), such as exploration and exploitation, to identify images for monkeypox virus prediction and diagnosis. Alongside, monkeypox, chickenpox, smallpox, cowpox, measles, tomato flu, and normal skin images were all considered in this study for monkeypox virus prediction and diagnosis. We collect photos from the International Skin Imaging Collaboration (ISIC) for analysis and experimentation purposes. Finally, we compare the proposed model Particle Swarm Optimization- Monkeypox Virus (PSOMPX) for monkeypox virus identification with four distinct pre-trained deep learning models (e.g., VGG16, ResNet50, InceptionV3, and Ensemble). Then we use four performance evaluation metrics—accuracy, precision, recall, and F1 score—to evaluate the model and analyze the outcomes of experiments. The experimental results obtained through the PSOMPX model significantly outperform other models due to its numerous traits.

An oleanic acid decorated gold nanorod for highly efficient inhibition of hemagglutinin and visible rapid detection of the influenza virus

Accurate diagnosis and effective antiviral treatments are urgently needed for the prevention and control of flu caused by influenza viruses. In this study, a novel oleanic acid (OA) functionalized gold nanorod OA-AuNP was prepared through a convenient ligand-exchange reaction. As hemagglutinin (HA) on the viral surface binds strongly to the multiple OA molecules on the surface of the nanoparticle, the prepared OA-AuNP was found to exhibit potent antiviral activity against a wide range of influenza A virus strains. Furthermore, the change in color resulting from the specific binding between HA and OA and the resultant aggregation of the OA-AuNP can be visually observed or measured by UV–vis spectra with a detection limit of 2 and 0.18 hemagglutination units (HAU), respectively, which is comparable to the commercially available influenza colloid gold rapid diagnostic kits. These findings demonstrate the potential of the OA-AuNP for the development of novel multivalent antiviral conjugates and the diagnosis of influenza virus.

Optimization of Amplicon-Based Sequencing for Large-Scale Diagnostics of Known Pome Viruses and Viroids

The implementation of reliable methods for the simultaneous detection of multiple viruses is challenging in plant viral diagnosis. In the present study, we report the development and effectiveness of an optimized version of a robust multiplexed PCR (HiPlex) followed by simultaneous targeted amplicon sequencing to detect 14 viruses and 5 viroids of pome fruit trees. We used amplicon sequencing data from 1,400 pome fruit tree samples previously diagnosed with the HiPlex v.1.0, which included 290 primer pairs for pome viruses. The analysis enabled the identification of highly efficient and specific primer pairs to be assembled in a new HiPlex (dubbed HiPlex v.2.0). Additionally, newly designed primers were implemented for 30% of targeted viruses and viroids. Overall, 116 primer pairs were assembled into the HiPlex v.2.0, resulting in a reduction of the number of primers and multiplex PCR reactions needed by 60 and 50%, respectively. The new HiPlex also includes two nepoviruses, tomato ringspot virus and tobacco ringspot virus, showing flexibility at augmenting and expanding the number of targeted diagnosed viruses using this approach. This optimized set of primers increased the cost-effectiveness and sensitivity while reducing background amplification. Comparable results for virus and viroid detection were obtained using the HiPlex v.2.0 and individual RT-qPCRs, even at a 10,000-fold dilution, demonstrating similar sensitivity to standard PCR-based diagnostics. This new HiPlex-based amplicon sequencing represents a powerful, sensitive, and low-cost tool for large-scale screening of pome fruit trees for all reported viruses and viroids infecting these crops. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license .

Recent Advances in Lateral Flow Assays for Viral Protein Detection with Nanomaterial-Based Optical Sensors.

Controlling the progression of contagious diseases is crucial for public health management, emphasizing the importance of early viral infection diagnosis. In response, lateral flow assays (LFAs) have been successfully utilized in point-of-care (POC) testing, emerging as a viable alternative to more traditional diagnostic methods. Recent advancements in virus detection have primarily leveraged methods such as reverse transcription-polymerase chain reaction (RT-PCR), reverse transcription-loop-mediated isothermal amplification (RT-LAMP), and the enzyme-linked immunosorbent assay (ELISA). Despite their proven effectiveness, these conventional techniques are often expensive, require specialized expertise, and consume a significant amount of time. In contrast, LFAs utilize nanomaterial-based optical sensing technologies, including colorimetric, fluorescence, and surface-enhanced Raman scattering (SERS), offering quick, straightforward analyses with minimal training and infrastructure requirements for detecting viral proteins in biological samples. This review describes the composition and mechanism of and recent advancements in LFAs for viral protein detection, categorizing them into colorimetric, fluorescent, and SERS-based techniques. Despite significant progress, developing a simple, stable, highly sensitive, and selective LFA system remains a formidable challenge. Nevertheless, an advanced LFA system promises not only to enhance clinical diagnostics but also to extend its utility to environmental monitoring and beyond, demonstrating its potential to revolutionize both healthcare and environmental safety.

A sensitive fluorescence biosensor based on ligation-transcription and CRISPR/Cas13a-assisted cascade amplification strategies to detect the H1N1 virus.

We propose a sensitive H1N1 virus fluorescence biosensor based on ligation-transcription and CRISPR/Cas13a-assisted cascade amplification strategies. Products are generated via the hybridization of single-stranded DNA (ssDNA) probes containing T7 promoter and crRNA templates to a target RNA sequence using SplintR ligase. This generates large crRNA quantities in the presence of T7 RNA polymerase. At such crRNA quantities, ternary Cas13a, crRNA, and activator complexes are successfully constructed and activate Cas13a to enhance fluorescence signal outputs. The biosensor sensitively and specifically monitored H1N1 viral RNA levels down to 3.23 pM and showed good linearity when H1N1 RNA concentrations were 100 pM-1 µM. Biosensor specificity was also excellent. Importantly, our biosensor may be used to detect other viral RNAs by altering the sequences of the two probe junctions, with potential applications for the clinical diagnosis of viruses and other biomedical studies.

The role of viral diagnostic tests in respiratory tract infections: moving forward.

The increased knowledge on virology and the increased potential of their diagnostic has risen several relevant question about the role of an early viral diagnosis and potential early treatment on the management of respiratory tract infections (RTI). In order to further understand the role of viral diagnostic tests in the management of RTI, a panel of experts was convened to discuss about their potential role, beyond what had been agreed in Influenza. The objective of this panel was to define the plausible role of aetiologic viral diagnostic into clinical management; make recommendations on the potential expanded use of such tests in the future and define some gaps in the management of RTI. Molecular Infection Viral Diagnostic (mIVD) tests should be used in all adult patients admitted to Hospital with RTI, and in paediatric patients requiring admission or who would be referred to another hospital for more specialised care. The increased use of mIVD will not only reduce the inappropriate use of antibiotics so reducing the antibiotic microbe resistance, but also will improve the outcome of the patient if an aetiologic viral therapy can be warranted, saving resource requirements and improving patient flows. Implementing IVD testing in RTI has various organizational benefits as well, but expanding its use into clinical settings would need a cost-effectiveness strategy and budget impact assessment.

Diagnosis of bovine viral diarrhea virus: an overview of currently available methods.

Bovine viral diarrhea virus (BVDV) is the causative agent of bovine viral diarrhea (BVD), which results in significant economic losses in the global cattle industry. Fortunately, various diagnostic methods available for BVDV have been established. They include etiological methods, such as virus isolation (VI); serological methods, such as enzyme-linked immunosorbent assay (ELISA), immunofluorescence assay (IFA), and immunohistochemistry (IHC); molecular methods, such as reverse transcription-polymerase chain reaction (RT-PCR), real-time PCR, digital droplet PCR (ddPCR), loop-mediated isothermal amplification (LAMP), recombinase polymerase amplification (RPA), and CRISPR-Cas system; and biosensors. This review summarizes the current diagnostic methods for BVDV, discussing their advantages and disadvantages, and proposes future perspectives for the diagnosis of BVDV, with the intention of providing valuable guidance for effective diagnosis and control of BVD disease.

The CRISPR-Cas13a Gemini System for noncontiguous target RNA activation

Simultaneous multi-target detection and multi-site gene editing are two key factors restricting the development of disease diagnostic and treatment technologies. Despite numerous explorations on the source, classification, functional features, crystal structure, applications and engineering of CRISPR-Cas13a, all reports use the contiguous target RNA activation paradigm that only enables single-target detection in vitro and one-site gene editing in vivo. Here we propose a noncontiguous target RNA activation paradigm of Cas13a and establish a CRISPR-Cas13a Gemini System composed of two Cas13a:crRNA binary complexes, which can provide rapid, simultaneous, highly specific and sensitive detection of two RNAs in a single readout, as well as parallel dual transgene knockdown. CRISPR-Cas13a Gemini System are demonstrated in the detection of two miRNAs (miR-155 and miR-375) for breast cancer diagnosis and two small RNAs (EBER-1 and EBER-2) for Epstein-Barr virus diagnosis using multiple diagnostic platforms, including fluorescence and colorimetric-based lateral flow systems. We also show that CRISPR-Cas13a Gemini System can knockdown two foreign genes (EGFP and mCherry transcripts) in mammalian cells simultaneously. These findings suggest the potential of highly effective and simultaneous detection of multiple biomarkers and gene editing of multiple sites.

The use of a synthetic peptide based on the immunogenic epitope GP51 for the serological diagnosis of bovine leukemia virus

Diagnosis of leukemia remains one of the key problems in carrying out recreational activities in farms that are disadvantaged by this disease. In the Russian Federation, various methods are used to identify the causative agent of leukemia, including serological methods, one of which is enzyme immunoassay (ELISA). The principle of the indirect variant of ELISA is to identify a complex formed by an antigen immobilized on the surface of the wells of a polystyrene tablet and specific antibodies contained in blood serum samples or cattle milk. The present work is devoted to the study of the possibility of using a synthetic peptide, which is an amino acid sequence of epitopes of the gp51 protein of the leukemia virus, as an antigen for ELISA in the indication of the bovine leukemia virus. To develop the ELISA method, we designed and synthesized the antigen – synthetic peptide BLVPEP (NCKYSNQCGDQGSFYVNHQILFLHLK QCHGIFTLTWEIWNC). In order to best sensitize this peptide, the main parameters of the ELISA formulation were worked out. In particular, the optimal concentration of the peptide for sensitization of the tablet was determined - 1.0 micrograms/ml, blocking solution – 3% solution of skimmed milk powder, anti–species peroxidase conjugate - 1:40000, the reaction buffer of the interaction "antigen-antibody" - FSB-T (pH 7.3) was selected, the minimum titer of antibodies to the pathogen, detectable in the blood sera of cows with leukemia – 1:1600. The ELISA method presented in this paper can be used as a basis for creating a test system for detecting the causative agent of bovine leukemia and find wide application in veterinary practice for serological diagnostics.

Occurrence and Multiplex PCR Detection of Citrus Yellow Vein Clearing Virus in Korea.

Citrus yellow vein clearing virus (CYVCV) is a member of the Alphaflexiviridae family that causes yellow vein clearing symptoms on citrus leaves. A total of 118 leaf samples from nine regions of six provinces in Korea were collected from various citrus species in 2020 and 2021. Viral diagnosis using next-generation sequencing and reverse transcription polymerase chain reaction (RT-PCR) identified four viruses: citrus tristeza virus, citrus leaf blotch virus, citrus vein enation virus, and CYVCV. A CYVCV incidence of 9.3% was observed in six host plants, including calamansi, kumquat, Persian lime, and Eureka lemon. Among the citrus infected by CYVCV, only three samples showed a single infection; the other showed a mixed infection with other viruses. Eureka lemon and Persian lime exhibited yellow vein clearing, leaf distortion, and water-soak symptom underside of the leaves, while the other hosts showed only yellowing symptoms on the leaves. The complete genome sequences were obtained from five CYVCV isolates. Comparison of the isolates reported from the different geographical regions and hosts revealed the high sequence identity (95.2% to 98.8%). Phylogenetic analysis indicated that all the five isolates from Korea were clustered into same clade but were not distinctly apart from isolates from China, Pakistan, India, and Türkiye. To develop an efficient diagnosis system for the four viruses, a simultaneous detection method was constructed using multiplex RT-PCR. Sensitivity evaluation, simplex RT-PCR, and stability testing were conducted to verify the multiplex RT-PCR system developed in this study. This information will be useful for developing effective disease management strategies for citrus growers in Korea.

Comparing RT-PCR of Individual Samples with High-Throughput Sequencing of Pooled Plant Samples for Field-Level Surveillance of Viruses in Blackberry and Wild Rubus.

Blackberry production is increasing in the Southeastern United States with the availability of new cultivars. In addition to high production costs, growers are challenged by virus diseases. Blackberry yellow vein disease (BYVD) significantly limits blackberry production. BYVD is associated with the crinivirus blackberry yellow vein-associated virus in mixed infections with other viruses. The specific disease etiology and ecological factors underlying BYVD are not well understood and rely on the effective diagnosis of several viruses involved in the complex. In 2021, we collected samples from blackberry plants showing BYVD symptoms, asymptomatic blackberry plants, and wild Rosaceae spp. from nine farms across South Carolina, for a total of 372 individual plant samples. RNA from individual samples was isolated and pooled into sample groups (i.e., symptomatic, asymptomatic, and wild) from each farm for a total of 24 pooled samples. We sequenced the pooled RNA using Illumina and analyzed sequence profiles using the Virtool bioinformatics application. We also tested each plant for six viruses by reverse transcriptase PCR or reverse transcriptase quantitative PCR and compared plant (PCR)-level and field (high-throughput sequencing [HTS])-level data. Virtool detected 17 known viruses in the pooled samples, including 11 blackberry viruses. PCR testing was mostly consistent with HTS, with some notable disagreements for specific viruses. Our study demonstrates that HTS could be used as an efficient tool to detect viruses in bulked samples in blackberry fields, although limitations to using HTS for field-level surveillance are also discussed here.

Rapid Initiation of Antiretroviral Therapy Under the Treat-All Policy Reduces Loss to Follow-Up and Virological Failure in Routine Human Immunodeficiency Virus Care Settings in China: A Retrospective Cohort Study (2016-2022).

Following the World Health Organization's guidelines for rapid antiretroviral therapy (ART) initiation [≤7 days after human immunodeficiency virus (HIV) diagnosis], China implemented Treat-All in 2016 and has made significant efforts to provide timely ART since 2017. This study included newly diagnosed HIV adults from Tianjin, China, between 2016 and 2022. Our primary outcome was loss to follow-up (LTFU) at 12 months after enrollment. The secondary outcome was 12-month virological failure. The association between rapid ART and LTFU, as well as virological failure, was assessed via Cox regression and logistic regression. A total of 896 (19.1%) of 4688 participants received ART ≤7 days postdiagnosis. The rate of rapid ART has increased from 7.5% in 2016 to 33.3% by 2022. The rapid ART group had an LTFU rate of 3.3%, as opposed to 5.0% in the delayed group. The rapid ART group had a much reduced virological failure rate (0.6% vs. 1.8%). Rapid ART individuals had a reduced likelihood of LTFU [adjusted hazard ratio: 0.65, 95% confidence intervals (CI): 0.44-0.96] and virological failure (adjusted odds ratio: 0.35, 95% CI: 0.12-0.80). The real-world data indicated that rapid ART is practicable and beneficial for Chinese people with HIV, providing evidence for its widespread implementation and scaling up.

Molecular Diagnosis of Human Monkeypox Virus during 2022-23 Outbreak: Preliminary Evaluation of Novel Real-Time Qualitative PCR Assays.

In 2022-23, the human monkeypox virus (MPXV) caused a global outbreak in several non-endemic countries. Here, we evaluated the diagnostic performance of four real-time qualitative PCR assays for the laboratory diagnosis of mpox (monkeypox) monkeypox disease. From July to August 2022, 27 positive and 10 negative specimens (lesion, crust and exudate swabs) were tested in the laboratory of the Hygiene Unit of the San Martino Hospital (Genoa, Italy) by using home-made real-time PCR to detect MPXV generic G2R_G DNA. According to the manufacturer's instructions, we also retrospectively analyzed these specimens using RealCycler MONK-UX/-GX (Progenie Molecular), STANDARD M10 MPX/OPX (SD Biosensor), Novaplex MPXV (Seegene Inc.) and RealStar Orthopoxvirus PCR Kit 1.0 (Altona Diagnostics) assays, recognized as research-use-only tests. The diagnostic accuracy and sensitivity of these assays ranged from 97.3% (95% CI: 86.2-99.5%) to 100% (95% CI: 90.6-100%) and 96.3% (95% CI: 81.72-99.34%) to 100% (95% CI: 72.2-100%), respectively. The RealCycler MONK-UX and STANDARD M10 MPX/OPX did not detect one positive sample with a cycle threshold of 36. The overall specificity was 100% (95% CI: 72.2-100%), and Cohen's Kappa values ranged from 1 (95% CI: 0.67-1) to 0.93 (95% CI: 0.61-1). As they are highly accurate, reliable and user-friendly, these tests should be recommended for the routine or rapid laboratory discrimination of mpox from other rash illnesses.

Iterative In Silico Screening for Optimizing Stable Conformation of Anti-SARS-CoV-2 Nanobodies.

Nanobodies (Nbs or VHHs) are single-domain antibodies (sdAbs) derived from camelid heavy-chain antibodies. Nbs have special and unique characteristics, such as small size, good tissue penetration, and cost-effective production, making Nbs a good candidate for the diagnosis and treatment of viruses and other pathologies. Identifying effective Nbs against COVID-19 would help us control this dangerous virus or other unknown variants in the future. Herein, we introduce an in silico screening strategy for optimizing stable conformation of anti-SARS-CoV-2 Nbs. Firstly, various complexes containing nanobodies were downloaded from the RCSB database, which were identified from immunized llamas. The primary docking between Nbs and the SARS-CoV-2 spike protein receptor-binding domain was performed through the ClusPro program, with the manual screening leaving the reasonable conformation to the next step. Then, the binding distances of atoms between the antigen-antibody interfaces were measured through the NeighborSearch algorithm. Finally, filtered nanobodies were acquired according to HADDOCK scores through HADDOCK docking the COVID-19 spike protein with nanobodies under restrictions of calculated molecular distance between active residues and antigenic epitopes less than 4.5 Å. In this way, those nanobodies with more reasonable conformation and stronger neutralizing efficacy were acquired. To validate the efficacy ranking of the nanobodies we obtained, we calculated the binding affinities (∆G) and dissociation constants (Kd) of all screened nanobodies using the PRODIGY web tool and predicted the stability changes induced by all possible point mutations in nanobodies using the MAESTROWeb server. Furthermore, we examined the performance of the relationship between nanobodies' ranking and their number of mutation-sensitive sites (Spearman correlation > 0.68); the results revealed a robust correlation, indicating that the superior nanobodies identified through our screening process exhibited fewer mutation hotspots and higher stability. This correlation analysis demonstrates the validity of our screening criteria, underscoring the suitability of these nanobodies for future development and practical implementation. In conclusion, this three-step screening strategy iteratively in silico greatly improved the accuracy of screening desired nanobodies compared to using only ClusPro docking or default HADDOCK docking settings. It provides new ideas for the screening of novel antibodies and computer-aided screening methods.

Primary Care Screening Recommendations for People Living With Human Immunodeficiency Virus

Historically, a diagnosis of human immunodeficiency virus meant early death. However, advances in therapy have lengthened the life expectancy of people living with human immunodeficiency virus (PLWH), and primary care providers often manage their care. Human immunodeficiency virus infection increases the risk of developing coronary artery disease, cardiac arrhythmias, diabetes mellitus, osteoporosis, neuropathy, kidney disease, and cancer at a younger age. Consequently, PLWH require a distinct set of screening recommendations. Primary care providers should be aware of these recommendations to prevent debilitating sequelae and premature death associated with chronic diseases in PLWH.

Enhancing Point-of-Care Diagnosis of African Swine Fever Virus (ASFV) DNA with the CRISPR-Cas12a-Assisted Triplex Amplified Assay.

Accurate, ultrasensitive, and point-of-care (POC) diagnosis of the African swine fever virus (ASFV) remains imperative to prevent its spread and limit the losses incurred. Herein, we propose a CRISPR-Cas12a-assisted triplex amplified colorimetric assay for ASFV DNA detection with ultrahigh sensitivity and specificity. The specific recognition of recombinase aided amplification (RAA)-amplified ASFV DNA could activate the Cas12a/crRNA/ASFV DNA complex, leading to the digestion of the linker DNA (bio-L1) on magnetic beads (MBs), thereby preventing its binding of gold nanoparticles (AuNPs) network. After magnetic separation, the release of AuNPs network comprising a substantial quantity of AuNPs could lead to a discernible alteration in color and significantly amplify the plasmonic signal, which could be read by spectrophotometers or smartphones. By combining the RAA, CRISPR/Cas12a-assisted cleavage, and AuNPs network-mediated colorimetric amplification together, the assay could detect as low as 0.1 copies/μL ASFV DNA within 1 h. The assay showed an accuracy of 100% for the detection of ASFV DNA in 16 swine tissue fluid samples, demonstrating its potential for on-site diagnosis of ASFV.

Understanding the impact of women's correct risk perception on human immunodeficiency virus diagnosis: Insights from South Africa.

South African women have the highest burden of HIV infections globally. We investigated the temporal trends and the impact of key factors associated with HIV diagnosis among a nationally representative cohort of South African women. Total of 24,657 women who participated in the National HIV, Behaviour and Health Surveys conducted from 2002-to-2017. Despite decades-long prevention efforts, we observed a significant increase in HIV prevalence over time (22% in 2002 to 21% in 2005-2008 and 29% in 2012-2017). Overall, 46% of the women living with HIV were not aware of their risk of HIV with age-specific disparities. Our findings revealed compelling evidence between HIV seropositivity and high HIV risk-perception (adjusted Odds Ratio (aOR):1.47 to 3.29) which increased overtime and exceeded the other factors. At a population-level, 45% of the HIV diagnoses were exclusively associated with women who believed they were at risk of HIV in 2012-2017. Women who reported using condoms at last sexual act were also at increased risk of HIV infection, with a population attributable risk of 18% (2002) to 21% (2012-2017). There is an urgent need for culturally, socially, and linguistically appropriate prevention and awareness campaigns with realistic, non-confrontational messages.

Crimean-Congo Hemorrhagic Fever: A Systematic Review

Crimean-Congo hemorrhagic fever is an predominatingly lethal viral contagion qualified in around thirty countries, and it has the roughly inclusive geographic allocation of the medically significant tick borne viral infections. Human become infected by tick bites, through pulverize infected ticks, next to contact with a case of Crimean-Congo hemorrhagic fever pending the acute stage of infection, or through handle with tissues or blood from viremic kine. Clinical features mostly display a spectacular progression recognized by myalgia, fever and bleeding. The best method used for diagnosis of Crimean-Congo hemorrhagic fever virus by real time polymerase chain reaction.

An Integrated Sample Referral System for HIV Viral Load and Early Infant Diagnosis in North-Western Province, Zambia-A Retrospective Cross-Sectional Study.

Zambia's adult HIV prevalence is high at 11% and faces challenges in achieving UNAIDS 95-95-95 targets for HIV, with a national viral load suppression of 86.2% falling short of the required 95%. North-Western Province has the lowest viral load suppression at 77.5%. Our study investigated the role of an integrated sample referral system in optimizing HIV viral load coverage and Early Infant Diagnosis turnaround time in the province. Using electronic data from the DISA Laboratory Information System and Smartcare, a retrospective cross-sectional analysis was conducted, involving 160,922 viral load and Early Infant Diagnosis results. The chi-square test and multiple linear regression were used for analysis. Following the implementation of the integrated sample referral system, viral load coverage consistently increased monthly (p < 0.001), Early Infant Diagnosis turnaround time improved by 47.7%, and sample volume increased by 25%. The study identifies associations between various factors and testing outcomes. These findings demonstrate improvements in viral load coverage and the Early Infant Diagnosis turnaround time and suggest targeting modifiable factors to further optimize the referral system. We recommend continued strengthening of the referral system and more deliberate demand-creation implementation strategies.

Digital Surface-Enhanced Raman Spectroscopy-Lateral Flow Test Dipstick: Ultrasensitive, Rapid Virus Quantification in Environmental Dust.

This study introduces a novel surface-enhanced Raman spectroscopy (SERS)-based lateral flow test (LFT) dipstick that integrates digital analysis for highly sensitive and rapid viral quantification. The SERS-LFT dipsticks, incorporating gold-silver core-shell nanoparticle probes, enable pixel-based digital analysis of large-area SERS scans. Such an approach enables ultralow-level detection of viruses that readily distinguishes positive signals from background noise at the pixel level. The developed digital SERS-LFTs demonstrate limits of detection (LODs) of 180 fg for SARS-CoV-2 spike protein, 120 fg for nucleocapsid protein, and 7 plaque forming units for intact virus, all within <30 min. Importantly, digital SERS-LFT methods maintain their robustness and their LODs in the presence of indoor dust, thus underscoring their potential for accurate and reliable virus diagnosis and quantification in real-world environmental settings.