Search Results (60)

Background/Objectives: The COVID-19 pandemic has highlighted the urgent need for rapid, accurate, and accessible diagnostic testing to effectively manage and contain the spread of SARS-CoV-2. RT-PCR is widely recognized as the gold standard for SARS-CoV-2 detection due to its high sensitivity and specificity. However, RT-PCR testing requires specialized laboratory equipment, highly trained personnel, and extended processing times, which limits its feasibility for large-scale screening and point-of-care applications. This study aims to systematically evaluate the diagnostic performance of RT-PCR and a colloidal gold immunochromatographic assay (GICA). Methods: By comparing these two methods, we seek to determine a GICA’s effectiveness as a complementary or alternative diagnostic tool, particularly in resource-limited settings and scenarios requiring rapid, large-scale testing. We assessed the following key clinical parameters: sensitivity, specificity, NPV, PPV, and accuracy. Additionally, we investigated the correlation between GICA signal intensity and RT-PCR Ct values using regression analysis, receiver operating characteristic curve analysis, and the calculated area under the curve. Results: Our findings indicate that while RT-PCR exhibits superior sensitivity, GICA results demonstrate a strong correlation with RT-PCR results and provide a rapid, cost-effective alternative for SARS-CoV-2 detection. Unlike RT-PCR, which requires extensive resources and prolonged turnaround times, a GICA delivers results within 20 min, making it a viable option for decentralized testing and real-time public health interventions. Conclusions: These results suggest that a GICA can serve as a complementary diagnostic tool alongside RT-PCR, particularly in resource-limited settings and high-throughput screening scenarios. By integrating GICAs into broader testing strategies, healthcare systems can enhance early detection efforts, improve accessibility to diagnostics, and strengthen pandemic response measures. Full article

The development of rapid, sensitive and cost-effective lateral flow assays is crucial for the detection of mycotoxins, ideally at the point-of-care level. This study presents the design and optimization of a competitive lateral flow assay based on gold nanoparticles (AuNPs) for the detection of zearalenone in food samples. Beginning with the synthesis and functionalization of gold nanoparticles, it proceeds to compare the immobilization of antibodies using chemical conjugation and physical adsorption binding strategies, upon optimizing parameters including the pH, antibody concentration and blocking conditions to enhance the stability of the prepared bioconjugates. The bioconjugates are characterized using UV–visible absorption spectroscopy and dynamic light scattering to monitor changes in the spectra and hydrodynamic size of AuNPs upon the addition of antibodies. The assessment of these bioconjugates is based on their ability to bind and manifest a color, developed due to nanoparticle binding with the test zone on the strip with the toxin–protein conjugate. The lateral flow immunochromatographic assay (LFIA) strips are then prepared by dispensing a control line (IgG) and test line (toxin–protein conjugate) on a nitrocellulose membrane using a lateral flow strip dispenser. The sensitivity of the LFIA strips is evaluated after standardizing the conditions by varying the concentration of zearalenone in the spiked samples and optimizing the running buffer solution. The limit of detection and limit of quantification under optimized conditions are determined to be 0.7 ng/mL and 2.37 ng for zearalenone-spiked samples. Furthermore, the mean pixel intensity and RGB values are plotted against the concentration of zearalenone, which can be used in a colorimetric smartphone-based application for the quantification of the amount of mycotoxin in the sample. Full article

Feline leukemia virus (FeLV) and Feline immunodeficiency virus (FIV) are globally prevalent retroviral pathogens that pose significant health risks to domestic cats. This study aimed to compare the diagnostic performance of two point-of-care—the immunochromatographic assay (ICA) and the RNase hybridization-assisted amplification (RHAM) test kit—against reverse transcription quantitative polymerase chain reaction (RT-qPCR), the current gold standard for FeLV and FIV detection. For FeLV detection, ICA demonstrated a sensitivity of 86.89%, specificity of 96.55%, accuracy of 90.00%, and precision of 98.15%, while for FIV detection, the assay showed a sensitivity of 75.86%, specificity of 88.52%, accuracy of 84.44%, and precision of 75.86%. In contrast, the RHAM test exhibited superior performance, with FeLV detection sensitivity of 93.44%, specificity of 98.28%, accuracy of 94.44%, and precision of 98.28%. For FIV detection, RHAM demonstrated a sensitivity of 75.86%, specificity of 100%, accuracy of 92.22%, and precision of 100%. Additionally, the RHAM assay significantly reduced detection time compared to RT-qPCR, enabling expedited clinical decision-making, alleviating laboratory workload, and lowering diagnostic costs. These benefits are particularly relevant in veterinary settings with limited access to PCR-based diagnostics, where the RHAM assay represents a rapid, reliable, and resource-efficient alternative for FeLV and FIV detection. Full article

Background: The alarming increase in carbapenemase-producing Enterobacterales is a matter of grave public health concern. The most ubiquitous carbapenemases, Klebsiella pneumoniae carbapenemase (KPC)-, New Delhi metallo-β-lactamase (NDM)-, and oxacillinase (OXA-48)-like enzymes, belong to the Ambler molecular classes A, B, and D, respectively. KPC- and OXA-48-like enzymes have a serine-based hydrolytic mechanism, while NDMs are metallo-β-lactamases that contain zinc in the active site. For the judicious use of reserve drugs and promoting antimicrobial stewardship, timely detection of carbapenemases is essential. While molecular tools are the gold standard for the detection of these enzymes, many laboratories have limited access to them. This study focused on evaluating in-house tools and commercial phenotypic tests for the detection of OXA-48-, KPC-, and NDM-like enzymes in K. pneumoniae, the predominant extremely drug-resistant pathogen in Oman. Methods: In total, 80 GeneXpert/PCR-confirmed (40 OXA-48 and 20 KPC and NDM each) and 37 whole-genome-sequenced (25 OXA-232 and 6 KPC-2, plus NDM-1 and NDM-5) K. pneumoniae were subjected to screening by temocillin (30 μg disk) (MAST Diagnostica, Germany) and D71C (MASTDISCS^®). Isolates resistant to temocillin (<11 mm) and D71C were subjected to four tests: an in-house tool (OXA-48 disk test) and three commercial phenotypic tests: (i) the MASTDISCS^® Combi (D72C) (MAST Group Ltd., Bootle, UK); (ii) the MASTDISCS^® Combi (D73C) (MAST Group Ltd., UK); and (iii) an immunochromatographic assay (ICT), which is the KPC/IMP/NDM/VIM/OXA-48 Combo test kit (Medomics, China), for the detection of OXA-48-, KPC-, and NDM-like carbapenemases. Results: Temocillin exhibited good sensitivity and specificity (100% and 97.50%) compared to D71C (70% and 100%). Among the confirmatory tests, the in-house OXA-48 disk test had 92.50% sensitivity and 100% specificity, while the commercial MAST DISC tests D72C, D73C, and ICT had 97.50%, 95.00%, and 100% sensitivity and 100%, 91.67%, and 95% specificity, respectively. Conclusions: The temocillin disk test is a good screening tool. With high sensitivity and specificity, ease of performance, short turnaround time, and low cost, we recommend the ICT format for routine diagnostic use. In resource-constrained centers, the OXA-48 disk test is an excellent alternative with high sensitivity and specificity. Full article

Actinobacillus pleuropneumoniae (APP) is a bacterial pathogen causing porcine pleuropneumonia, causing great economic loss to the global pig industry. Although natural apxIV contributes to the prevention and control of porcine pleuropneumonia, its isolation poses a great challenge, and recombinant soluble apxIV proteins tend to carry large molecular weight tags. The traditional serologic methods tend not to accurately detect the apxIV-partially deleted vaccine (GDV). In this study, we screened the soluble protein apxIVA N2 (756 bp) from six apxIV-truncated proteins and applied it to the enzyme-linked immunosorbent assay (ELISA) and colloidal gold immunochromatographic strip for detecting the samples vaccinated with APP GDV. The results indicate that N2 was close to the natural apxIV protein in terms of structure and function as it only contained a single His (0.86 kDa) tag and a single S (2 kDa) tag. Among the six candidate proteins, N2 exhibited the best performance in distinguishing APP-infected samples from those vaccinated with the APP GDV. Both ELISA and colloidal gold immunochromatographic strips based on this protein exhibited an excellent performance in detecting and distinguishing wild-strain-infected samples from those vaccinated with the subunit vaccine or the GDV. In addition, three monoclonal antibodies against different antigenic epitopes were identified using these truncated proteins. Our studies are of great significance for further research on APP, the differential diagnosis of wild strains and vaccine strains, and pig control breeding, exhibiting a broad application prospect in the on-site diagnosis of APP, particularly in remote areas lacking detection instruments and professionals. Full article

Pesticide residues still pose a risk to human health. With the rapid development of rapid testing technology, the levels of different types of pesticide residues in agricultural products can be identified in a shorter period; thus, the safety of food can be guaranteed. However, the effectiveness of commercially available testing products has yet to be evaluated. In this study, colloidal gold immunochromatographic qualitative testing products manufactured by 34 companies were tested for their assay performance on Emamectin Benzoate, Isocarbophos, and fipronil with standardized cowpea samples. The results indicated that most of the evaluated products were identified as having ‘passed’. Most pesticide residue rapid test immunoassay products can be considered ideal means for testing certain pesticide residues. However, further evaluation of pesticide residue rapid test immunoassay products is needed, as detection technologies are still developing. Full article

A highly sensitive lateral flow immunoassay (LFIA) for imidacloprid, a widely used neonicotinoid insecticide, has been developed. The LFIA realizes the indirect coupling of anti-imidacloprid antibodies and gold nanoparticle (GNP) labels directly in the course of the assay. For this purpose, the common GNPs conjugate with anti-imidacloprid antibodies and are changed into a combination of non-modified, anti-imidacloprid antibodies, and the GNPs conjugate with anti-species antibodies. The given approach provides the possibility of selecting independent concentrations of GNPs and anti-imidacloprid antibodies to obtain the influence of minimal imidacloprid concentrations in the samples on the formation of detected, labeled immune complexes. A comparative study of imidacloprid LFIAs with common and indirect antibody–label coupling was implemented. The second variant reduced the limit of detection (LOD) of imidacloprid 20 times, reaching 0.2 ng/mL and 0.002 ng/mL for visual and instrumental detection, respectively, thus surpassing the existing LFIAs for imidacloprid. The developed highly sensitive LFIA was tested for imidacloprid detection in freshly squeezed fruits and berries without any additional sample preparation. The imidacloprids revealed were in the range of 75–97% for grape, 75–85% for orange, and 86–97% for apple samples. The time of the testing was 15 min. Full article

The livestock industry uses ofloxacin, an antibiotic, to prevent several animal diseases; however, the overdose of ofloxacin used in animal farming treatments may appear in food products and cause some adverse human health effects. Hence, there is an immediate need to develop a method suitable for on site large-scale detection of ofloxacin residues in animal-derived foods. This study aimed to prepare a monoclonal antibody with high sensitivity and affinity for ofloxacin by re-synthesizing the ofloxacin hapten and synthesizing the corresponding complete antigen. The IC₅₀ of the enzyme-linked immunosorbent assay (ic-ELISA) was 0.13 ng/mL, and the detection limit was 0.033 ng/mL. The visual detection limit of the established colloidal gold immunochromatographic test strip, for the visual detection of actual samples, was 1 ng/g. In summary, this work establishes a rapid detection method of ofloxacin residues on the basis of colloidal gold immunochromatography that is suitable for actual detection. Full article

Gatifloxacin (GAT), an antibiotic belonging to the fluoroquinolone (FQ) class, is a toxicant that may contaminate food products. In this study, a method of ultrasensitive immunochromatographic detection of GAT was developed for the first time. An indirect format of the lateral flow immunoassay (LFIA) was performed. GAT-specific monoclonal antibodies and labeled anti-species antibodies were used in the LFIA. Bimetallic core@shell Au@Ag nanoparticles (Au@Ag NPs) were synthesized as a new label. Peroxidase-mimic properties of Au@Ag NPs allowed for the catalytic enhancement of the signal on test strips, increasing the assay sensitivity. A mechanism of Au@Ag NPs-mediated catalysis was deduced. Signal amplification was achieved through the oxidative etching of Au@Ag NPs by hydrogen peroxide. This resulted in the formation of gold nanoparticles and Ag⁺ ions, which catalyzed the oxidation of the peroxidase substrate. Such “chemical enhancement” allowed for reaching the instrumental limit of detection (LOD, calculated by Three Sigma approach) and cutoff of 0.8 and 20 pg/mL, respectively. The enhanced assay procedure can be completed in 21 min. The enhanced LFIA was tested for GAT detection in raw meat samples, and the recoveries from meat were 78.1–114.8%. This method can be recommended as a promising instrument for the sensitive detection of various toxicants. Full article

Actinidia chlorotic ringspot-associated virus (AcCRaV, Emaravirus actinidiae) is prevalent in Chinese kiwifruit, leading to substantial yield reduction. The intricate nature of symptoms presents diagnostic challenges, underscoring the necessity for a rapid and accurate detection method that facilitates effective control. In this investigation, AcCRaV isolates from key kiwi-producing regions in Sichuan province were collected and analyzed, with representative strains chosen as experimental materials. Primers targeting the nucleoprotein gene of AcCRaV were designed, and their codon usage was optimized to enhance performance. Various serological methods utilizing polyclonal antibodies were developed, including ELISA, dot immunobinding assay, and AcCRaV-specific gold immunochromatographic bands (AcCRaV-GICS). Field samples exhibited high specificity and sensitivity when tested using these methods. Furthermore, the results obtained from a large number of field samples are consistent with those derived from RT-PCR analysis, further validating the applicability of our approach. A detection method capable of handling a large volume of field samples infected with AcCRaV is currently lacking; thus, our system construction provides an important reference for addressing this gap. Full article

Salmonella typhimurium and Listeria monocytogenes are relevant foodborne bacterial pathogens which may cause serious intoxications and infectious diseases in humans. In this study, a sensitive immunochromatographic analysis (ICA) for the simultaneous detection of these two pathogens was developed. For this, test strips containing two test zones with specific monoclonal antibodies (MAb) against lipopolysaccharides of S. typhimurium and L. monocytogenes and one control zone with secondary antibodies were designed, and the double-assay conditions were optimized to ensure high analytical parameters. Prussian blue nanoparticles (PBNPs) were used as nanozyme labels and were conjugated with specific MAbs to perform a sandwich format of the ICA. Peroxidase-mimic properties of PBNPs allowed for the catalytic amplification of the colorimetric signal on test strips, enhancing the assay sensitivity. The limits of detection (LODs) of Salmonella and Listeria cells were 2 × 10² and 7 × 10³ cells/mL, respectively. LODs were 100-fold less than those achieved due to the ICA based on the traditional gold label. The developed double ICA was approbated for the detection of bacteria in cow milk samples, which were processed by simple dilution by buffer before the assay. For S. typhimurium and L. monocytogenes, the recoveries from milk were 86.3 ± 9.8 and 118.2 ± 10.5% and correlated well with those estimated by the enzyme-linked immunosorbent assay as a reference method. The proposed approach was characterized by high specificity: no cross-reactivity with other bacteria strains was observed. The assay satisfies the requirements for rapid tests: a full cycle from sample acquisition to result assessment in less than half an hour. The developed ICA has a high application potential for the multiplex detection of other foodborne pathogens. Full article

Lomefloxacin (LOM), an antibiotic crucial for preventing various animal diseases in animal husbandry, can pose serious health risks when found in excessive amounts in meat products. The development of highly specific and sensitive colloidal gold immunochromatographic test strips is essential for the accurate detection of this class of antibiotics. Our study utilized a monoclonal antibody (mAb) assay and immunochromatographic strips to detect lomefloxacin residues in meat products. The results showed minimal cross-reactivity with other structural analogs, with a maximum half inhibitory concentration (IC₅₀) of 0.93 ng/mL and a linear range of 0.38 to 2.3 ng/mL for the indirect competitive enzyme-linked immunosorbent assay (ic-ELISA). The recovery of LOM was 80% to 120%, with an average coefficient of variation below 5%. The immunochromatographic strip test results showed a visual detection limit of 2.5 ng/g, meeting the market requirements for the test. This study highlights the significance of specific and sensitive testing methods for detecting lomefloxacin, ensuring consumers’ safety and health. Full article

Acute hepatopancreatic necrosis disease (AHPND) poses a significant threat to shrimp aquaculture worldwide, necessitating the accurate and rapid detection of the pathogens. However, the increasing number of Vibrio species that cause the disease makes diagnosis and control more difficult. This study focuses on developing a monoclonal antibody against the Photorhabdus insect-related (Pir) toxin B (PirB), a pivotal virulence factor in AHPND-causing Vibrio, and establishing a colloidal gold immunochromatographic assay for the enhanced early diagnosis and monitoring of AHPND. Monoclonal antibodies targeting PirB were developed and utilized in the preparation of colloidal-gold-labeled antibodies for the immunochromatographic assay. The specificity and sensitivity of the assay were evaluated through various tests, including antibody subclass detection, affinity detection, and optimal labeling efficiency assessment. The developed PirB immunochromatographic test strips exhibited a good specificity, as demonstrated by the positive detection of AHPND-causing Vibrio and negative results for non-AHPND-causing Vibrio. The study highlights the potential of the developed monoclonal antibody and immunochromatographic assay for the effective detection of AHPND-causing Vibrio. Further optimization is needed to enhance the sensitivity of the test strips for improved practical applications in disease prevention and control in shrimp aquaculture. Full article

Foodborne illnesses can be infectious and dangerous, and most of them are caused by bacteria. Some common food-related bacteria species exist widely in nature and pose a serious threat to both humans and animals; they can cause poisoning, diseases, disabilities and even death. Rapid, reliable and cost-effective methods for bacterial detection are of paramount importance in food safety and environmental monitoring. Polymerase chain reaction (PCR), lateral flow immunochromatographic assay (LFIA) and electrochemical methods have been widely used in food safety and environmental monitoring. In this paper, the recent developments (2013–2023) covering PCR, LFIA and electrochemical methods for various bacterial species (Salmonella, Listeria, Campylobacter, Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli)), considering different food sample types, analytical performances and the reported limit of detection (LOD), are discussed. It was found that the bacteria species and food sample type contributed significantly to the analytical performance and LOD. Detection via LFIA has a higher average LOD (24 CFU/mL) than detection via electrochemical methods (12 CFU/mL) and PCR (6 CFU/mL). Salmonella and E. coli in the Pseudomonadota domain usually have low LODs. LODs are usually lower for detection in fish and eggs. Gold and iron nanoparticles were the most studied in the reported articles for LFIA, and average LODs were 26 CFU/mL and 12 CFU/mL, respectively. The electrochemical method revealed that the average LOD was highest for cyclic voltammetry (CV) at 18 CFU/mL, followed by electrochemical impedance spectroscopy (EIS) at 12 CFU/mL and differential pulse voltammetry (DPV) at 8 CFU/mL. LOD usually decreases when the sample number increases until it remains unchanged. Exponential relations (R² > 0.95) between LODs of Listeria in milk via LFIA and via the electrochemical method with sample numbers have been obtained. Finally, the review discusses challenges and future perspectives (including the role of nanomaterials/advanced materials) to improve analytical performance for bacterial detection. Full article

Japanese encephalitis virus (JEV) causes acute encephalitis in humans and is of major public health concern in most Asian regions. Dogs are suitable sentinels for assessing the risk of JEV infection in humans. A neutralization test (NT) or an enzyme-linked immunosorbent assay (ELISA) is used for the serological detection of JEV in dogs; however, these tests have several limitations, and, thus, a more convenient and reliable alternative test is needed. In this study, a colloidal gold immunochromatographic strip (ICS), using a purified recombinant EDIII protein, was established for the serological survey of JEV infection in dogs. The results show that the ICSs could specifically detect JEV antibodies within 10 min without cross-reactions with antibodies against other canine viruses. The test strips could detect anti-JEV in serum with dilution up to 640 times, showing high sensitivity. The coincidence rate with the NT test was higher than 96.6%. Among 586 serum samples from dogs in Shanghai examined using the ICS test, 179 (29.98%) were found to be positive for JEV antibodies, and the high seropositivity of JEV in dogs in China was significantly correlated with the season and living environment. In summary, we developed an accurate and economical ICS for the rapid detection of anti-JEV in dog serum samples with great potential for the surveillance of JEV in dogs. Full article