Search Results (56)

Acetamiprid (ACE) is a widely used neonicotinoid insecticide with potential health risks due to its persistence in the environment and bioaccumulation in plant-based foods. In this study, an improved dual-mode lateral flow immunochromatography assay (DMLFIA) was established for the sensitive detection of ACE residues in plant-based foods using Au@Ag@Pt NPs as dual-function signal probes. In the colorimetric mode (Mode 1), with the inhibition rate as the vertical coordinate and the ACE concentrations as the horizontal coordinate, the linear detection range of the colorimetric mode was 0.1–3000 μg/L with the detection limit of 0.08 μg/L. In the photothermal mode (Mode 2), the temperature values obtained via smartphones integrated thermal imager showed a linear relationship with 0.1–3000 μg/L of ACE, and the detection limit is 0.1 μg/L. The cross-reactivity rate of the developed DMLFIA with other structural analogs and functional substances was less than 0.39%. The recovery rate of the developed DMLFIA used in spiked plant-based foods was 81.1–99.4%, with relative standard deviations (RSD) of 1.3–10.7%. The detection results of this constructed DMLFIA showed good consistency with those obtained by HPLC and ELISA. The constructed DMLFIA provides a selective, sensitive and rapid detection tool for ACE in different scenarios in plant-based foods. Full article

Background/Objectives: Given the ongoing threat of antimicrobial resistance, the identification and characterization of multidrug-resistant isolates are essential. An increase in antimicrobial-resistant bacteria has been reported in Romania, but national data are still scarce. This study aimed to evaluate beta-lactamase-producing Gram-negative bacteria (GNB) isolated over two years at a Romanian nephrology hospital, while comparing carbapenemase detection phenotypic methods. Methods: Gram-negative bacterial isolates collected between January 2022 and May 2024 that met antimicrobial resistance screening criteria were evaluated. After identification, extensive disk diffusion antibiograms were performed, read, and interpreted, complemented by testing on cloxacillin/oxacillin-supplemented Mueller–Hinton agar. The colistin minimum inhibitory concentration (MIC) was not assessed, and aztreonam–avibactam was not tested for Enterobacterales. For non-fermenter GNB, the colistin MIC was determined. Phenotypic carbapenemase production tests were performed for all strains (BlueCarba Test, CIM, mCIM, zCIM, and rCIM). Carbapenemase detection immunochromatographic tests were performed for a set of strains. Results: Among the 397 evaluated strains, 335 (84.38%) were Enterobacterales and 62 (15.62%) non-fermenter GNB, showing high antimicrobial resistance levels. Of these, 188 (47.35%) were Klebsiella pneumoniae; 139/188 (73.93%) showed carbapenem resistance and carbapenemase production; 49/188 (26.06%) produced two carbapenemases; and 45/188 (23.93%) presented resistance to all tested antimicrobials. MALDI-TOF identified 28 KPC-producing K. pneumoniae strains. Lateral flow assays revealed NDM, VIM, KPC, and OXA-48-like enzymes in 48 of 56 tested Enterobacterales; 12/48 strains produced two carbapenemases. Of the 62 non-fermenter GNB, 33 were Pseudomonas spp. and 20 Acinetobacter baumannii; one Pseudomonas spp. was susceptible only to colistin and seven only to cefiderocol; four A. baumannii were susceptible only to colistin and three only to cefiderocol. Lateral flow assays detected VIM or IMP enzymes in 13/33 Pseudomonas spp. and OXA-23 and/or OXA-40/-58 enzymes in all 20 A. baumannii. Conclusions: Among the evaluated strains, many showed resistance to multiple antimicrobial classes. Furthermore, strains co-producing two carbapenemases were identified. Full article

Objectives: This study evaluated the diagnostic performance of the lateral flow immunochromatographic assay (RESIST-ACINETO, Coris BioConcept) for the rapid detection of the major carbapenemases in Acinetobacter baumannii. Methods: Blood culture isolates collected between 2014 and 2024 with meropenem MIC ≥ 4 mg/L were retrieved, re-identified by MALDI-TOF MS, and susceptibility was confirmed by broth microdilution. Carbapenemase genes (bla_OXA-23, bla_OXA-40, bla_NDM) were detected using multiplex PCR, which served as the reference standard. All isolates were tested using the RESIST ACINETO assay, and diagnostic accuracy parameters were calculated. Results: A total of 114 isolates were recovered and confirmed as A. baumannii. Among 93 carbapenem-non-susceptible isolates, 97.8% (91/93) were correctly identified by the assay. The test showed 99.1% sensitivity and 99.1% specificity, with most positive results appearing within 3–10 min. Two discrepant results were observed (one false positive, one false negative), while all meropenem-susceptible isolates tested negative. Conclusions: The RESIST ACINETO assay provides rapid, accurate detection of carbapenemases in A. baumannii, significantly reducing turnaround time compared with conventional workflows. Its performance supports integration into routine diagnostics to enhance timely resistance confirmation and infection-control interventions. Full article

Background: Fecal calprotectin (FC) is a well-established, non-invasive biomarker of intestinal inflammation and is widely used to differentiate inflammatory bowel disease (IBD) from functional gastrointestinal disorders. Although enzyme-linked immunosorbent assays (ELISA) remain the reference method, rapid immunochromatographic tests (ICTs) offer important operational advantages for point-of-care (POC) diagnostics. However, variability in analytical performance among available ICTs remains a concern. Objective: This study aimed to evaluate the diagnostic accuracy of the CerTest Calprotectin one-step card (CerTest Biotec S.L., Zaragoza, Spain) in comparison with the Actim^® Calprotectin lateral flow assay and the reference Calprest^® ELISA (Eurospital Diagnostics, Italy). Methods: A total of 128 fresh stool samples from patients clinically suspected of IBD were analyzed in parallel using all three assays. For the reference ELISA (Calprest^®), a cutoff value of >40 µg/g was applied according to the manufacturer’s instructions. For discrepant results between assays, a cutoff of 200 ng/mL (equivalent to 200 µg hCp/g stool) was employed for ELISA Calprest^® to resolve inconsistencies. The results of the lateral flow assays (CerTest^® Calprotectin ICT and Actim^® Calprotectin) were interpreted using their respective manufacturer-recommended thresholds. Diagnostic sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) were calculated using ELISA as the reference standard. Agreement between methods was assessed using Cohen’s κ coefficient. Results: Using ELISA, 47 of 128 samples (36.7%) exceeded the 40 µg/g cutoff. Compared with the Actim^® assay, the CerTest card demonstrated a sensitivity of 88.0% (95% CI: 75.7–95.5), a specificity of 100.0% (95% CI: 95.4–100), and a strong agreement (κ = 0.90). When compared with ELISA, the CerTest assay showed a sensitivity of 87.2% (95% CI: 74.3–95.2), a specificity of 96.3% (95% CI: 89.6–99.2), a PPV of 93.2%, an NPV of 93.2%, and a strong agreement (κ = 0.85). Conclusions: The CerTest Calprotectin one-step card provides a rapid and reliable detection of fecal calprotectin, demonstrating a high sensitivity and specificity that are comparable to both other lateral flow assays and the ELISA reference method. These findings support the use of rapid immunochromatographic testing as a valuable tool for preliminary screening and clinical decision-making in patients suspected of IBD, while acknowledging that histology remains the gold standard for definitive diagnosis. Full article

Potato virus X (PVX) and potato virus Y (PVY) are major pathogens that threaten seed potato quality and yield. To improve the efficiency of field screening, we developed monovalent PVX, monovalent PVY, and bivalent PVX/PVY nanozyme strips using Fe₃O₄ nanozymes as labels in a double-antibody sandwich lateral flow immunochromatographic assay. Western blot analysis demonstrated that four monoclonal antibodies (PVX 2, PVX 6, PVY 2, and PVY 5) specifically recognized their corresponding viral coat proteins. Specificity testing showed that the nanozyme strips reacted only with the target viruses and did not cross-react with other common potato viruses, including Potato virus A (PVA), Potato virus M (PVM), Potato virus S (PVS), and Potato leafroll virus (PLRV). The PVX nanozyme strip detected PVX-positive extracts diluted up to 10³-fold, the PVY nanozyme strip up to 10⁴-fold, and the bivalent strip detected PVX/PVY co-infected samples diluted up to 10³-fold. In addition, detection results by strips from 12 samples of plantlets in vitro were fully consistent with RT-PCR. These nanozyme strips provide rapid, simple, specific, and sensitive methods that can be stored at ambient temperature, enabling field surveys, warehouse screening, and on-site testing and supporting early detection of potato virus diseases. Full article

Aflatoxin B1 (AFB₁) is one of the most toxic fungal secondary metabolites. High-sensitivity and rapid detection of AFB₁ is crucial for safeguarding consumer health, reducing post-harvest food losses, and promoting agricultural trade. Here, we developed a magnetic enrichment–catalytic lateral flow immunochromatographic assay (E-C-LFIA) for quantitative AFB₁ detection. The approach couples immunomagnetic capture and enrichment with carboxylated magnetite (Fe₃O₄) nanozyme probes and post-assay peroxidase-like catalysis of the H₂O₂–TMB system to enhance colorimetric readout. Compared with conventional LFIA performed without magnetic enrichment or catalytic amplification, E-C-LFIA achieved a visual detection limit of 0.05 μg/L for AFB₁, corresponding to a 20-fold improvement in sensitivity. The quantitative limit of detection (LOD, 3σ) was 0.023 μg/L, representing a 14.8-fold improvement in sensitivity. The method was demonstrated for AFB₁ screening in representative cereal- and nut-based matrices (rice, corn and peanut). Overall, E-C-LFIA provides a sensitive, rapid, and equipment-light option for on-site AFB1 screening and offers a transferrable strategy for other small-molecule contaminants. Full article

Antibiotic stereoisomers as components of medicines are typically characterized by different biological activities. Because pharmaceuticals can include a racemic mixture of stereoisomers, monitoring of all forms is required. One contaminant of food products, antibiotic ofloxacin (OFL), as a chiral compound, has two enantiomers—the biologically active S-isomer and less active R-isomer. In this study, a sensitive immunochromatographic test system for simultaneous enantiospeсific detection of the two OFL isomers was developed for the first time. For this, polyclonal antibodies were produced, and conditions for a double lateral flow immunoassay (LFIA) were selected and optimized so that the cross-reactivity with another enantiomer was negligible. The LFIA was performed in a competitive format with gold nanoparticles as a label for secondary antibodies. The achieved LODs/cutoffs were 0.001/10 and 0.007/30 ng/mL for S-OFL and R-OFL detection, respectively; the assay procedure took only 15 min. A double LFIA was performed to detect S-OFL and R-OFL in milk with minimal sample pretreatment; the recoveries were 85–95%. The developed test system is an effective tool for the selective detection of both isomers of OFL, allowing for the avoidance of false negative results. This immunochromatographic approach can be promising for the control of other optically active food toxicants. Full article

Fumonisins are among the most prevalent mycotoxins in maize and maize-based products, posing significant food safety and public health risks due to their hepatotoxic, nephrotoxic, and potential carcinogenic effects. Given the strict regulatory limits set by the European Commission and Codex Alimentarius, the development of reliable, sensitive, and matrix–robust analytical methods remain a priority for routine monitoring in both food and feed systems. In this study, a reusable immuno-affinity purification methodology for the quantitative determination of fumonisin mycotoxins (FB1, FB2 and FB3) in foods and feeds (maize matrix) was developed. A single extraction protocol using 2% formic acid in water was employed, followed by cleanup with an immuno-affinity purification column and toxin elution by methanol/PBS (1:1, v/v). Detection and quantification of the mycotoxins was achieved by a normal phase ultra-high performance liquid chromatography coupled with electrospray ionisation triple quadrupole mass spectrometry (UHPLC/ESI-MS/MS). The chromatographic mobile phase utilised was a linear gradient of methanol/water containing 0.1% formic acid. The developed method has a limit of detection of 2.5 ng/g and a limit of quantification of 5 ng/g, all well below the European commission’s guidance values of 1000 ng/g for corn destined for human consumption and 800 ng/g for maize-based breakfast cereals and snacks. While the recovery rates of the method in this study ranged from 65–70% for the three fumonisin analogues in solutions, when tested in maize matrix, recoveries were markedly lower (~30%) due to pronounced matrix suppression. Good repeatability (standard deviation <10%) was achieved for all the fumonisin analogues. The developed method, although quick and effective in solvent systems, suffered limitations to its practical usage due to matrix suppression of the extracts derived from the immuno-affinity purification column, thus significantly reducing the application of the method in measuring fumonisin mycotoxins in food and feed samples. Overall, the method was effective in quantification of fumonisin mycotoxins in solvent solutions but not in food and feed matrices, thus necessitating further optimisation for practical usage. The performance of the developed method was compared to a commercial lateral flow immunochromatographic assay which proved to be better than the developed method in the quantification of toxins in food matrices, as the commercial lateral flow immunochromatographic assay outperformed the developed method in maize matrices. These findings highlight the need for matrix-based validation and further refinement of antibody stability to ensure robust application in regulatory monitoring of fumonisins using immunoaffinity purification methods. Full article

The spread of carbapenemase-producing Gram-negative bacteria poses a significant clinical challenge due to their association with severe Difficult-to-Treat nosocomial infections, as available therapies are drastically reduced. Rapid and accurate detection of carbapenemase-producing Gram-negative bacteria is critical for effective patient management, guiding appropriate antibiotic therapy, and implementing infection control measures to limit their dissemination within healthcare settings. Lateral flow immunoassays that detect the five main carbapenemases have become cornerstones in the fight against carbapenemase-producing Gram-negative bacteria. Carbapenemases evolve in response to antibiotic exposure, and therefore regular evaluation of these lateral flow immunoassays is crucial. Here, we have evaluated a novel assay, the KINVO assay (Medomics Medical Technology) and compared it to the Gold Standard of LFIAs for carbapenemase detection, the NG-TEST CARBA 5 assay (NG-Biotech) on a large panel of carbapenemase variants. The comparison between the two assays highlighted that both share key advantages such as rapidity and simplicity. However, NG-Test CARBA 5 demonstrated superior performance overall, particularly in accurately detecting IMP-type carbapenemases and the OXA-48 variant OXA-505. In contrast, the KINVO assay was more effective at detecting a broader range of KPC variants, including some that have lost carbapenem-hydrolyzing activity but gained resistance to ceftazidime/avibactam. If we consider these variants no longer as carbapenemases, and thus that they should not be detected, the NG-Test CARBA 5 performed better for KPC carbapenemase detection. Full article

The N-glycolylneuraminic acid (Neu5Gc), a major salivary acid molecule found on the cell surface of animals such as pigs, cows, and sheep, can be metabolically incorporated into the body through consumption of animal-derived foods like red meat. This leads to an immune response and chronic inflammation in individuals who do not naturally produce Neu5Gc, including humans and poultry, further increasing the risk of cancer. The trans-cleavage activity of Cas12a is activated by the recognition of the target aptamer by the crRNA, resulting in the cleavage of the dual-labeled probe. By combining this with immunochromatographic techniques, we established a chromatographic test strip assay that allows immediate on-site detection of Neu5Gc contamination in non-red meat samples devoid of Neu5Gc. Further optimization enabled specific detection within 25 min with a minimum detectable limit of 10 ng/mL. These analyses successfully detected the spiked samples and actual samples containing Neu5Gc. The developed lateral flow test strips based on aptamer-Cas12a can be utilized for detecting Neu5Gc contamination in non-red meat food products, animal bioproducts, and poultry feeds. Full article

Background: Carbapenem-resistant Enterobacterales infections are frequent in critically ill patients. Outbreaks caused by carbapenemase-producing Enterobacterales, in particular the New Delhi Metallo-beta-lactamase (NDM)-type carbapenemase-producing phenotype, are increasing in Italy. Unfortunately, the clinical impact of this new microorganism is still being defined, as well as the correlation between colonization and invasive infections. The aim of the study is to analyze factors related to the development of NDM infections in colonized patients and to evaluate their impact on patients’ outcome in high-complexity ICUs. Methods: All patients admitted to the General and Cardiac ICUs of ‘Città della Salute e della Scienza’ University Hospital in Turin (Italy) between January and August 2023 were enrolled. Microorganisms were examined by lateral flow immunochromatographic assays or molecular assays on weekly surveillance or clinically requested cultures. Antimicrobial susceptibility was determined by broth microdilution methods and interpreted according to EUCAST breakpoints. Results: Out of a total of 915 patients, 46 (5%) were positive for NDM-producing Enterobacterales and, among them, 13 (28%) developed an invasive infection. All patients were critical (SAPS II 40+/−13). The median times between ICU admission and colonization or infection were 6 and 16 days, respectively. Significant disparities emerged between colonized and infected patients regarding days of mechanical ventilation (1 vs. 28), ICU (7 vs. 39 days), and in-hospital (21 vs. 71 days) length of stay. Renal replacement treatment (OR 8.2461, p = 0.0173, 95% CI [1.3636–65.9114]) and surgery (OR 22.8747, p = 0.0149, CI95% [1.5986–1447.743]) seemed to impact the risk of developing infection. Six patients with invasive infection were treated with Cefiderocol and five with Ceftazidime/Avibactam and Aztreonam. In absence of early identification and appropriate treatment, patients may be at increased risk of colonization spread and potentially worse clinical outcomes. Conclusions: Early identification of the carbapenemase type is clinically relevant in critically ill patients with confirmed or suspected infection, as NDM production necessitates the use of specific agents for effective treatment. Full article

Nipah virus (NiV) and Hendra virus (HeV), which both belong to the genus henipavirus, are zoonotic pathogens that cause severe systemic, neurological, and/or respiratory disease in humans and a variety of mammals. Therefore, monitoring viral prevalence in natural reservoirs and rapidly diagnosing cases of henipavirus infection are critical to limiting the spread of these viruses. Current laboratory methods for detecting NiV and HeV include virus isolation, reverse transcription quantitative real-time PCR (RT-qPCR), and antigen detection via an enzyme-linked immunosorbent assay (ELISA), all of which require highly trained personnel and specialized equipment. Here, we describe the development of a point-of-care customized immunochromatographic lateral flow (ILF) assay that uses recombinant human ephrin B2 as a capture ligand on the test line and a NiV-specific monoclonal antibody (mAb) on the conjugate pad to detect NiV and HeV. The ILF assay detects NiV and HeV with a diagnostic specificity of 94.4% and has no cross-reactivity with other viruses. This rapid test may be suitable for field testing and in countries with limited laboratory resources. 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

Fumonisins, a class of mycotoxins predominantly produced by Fusarium species, represent a major threat to food safety and public health due to their widespread occurrence in staple crops including peanuts, wine, rice, sorghum, and mainly in maize and maize-based food and feed products. Although fumonisins occur in different groups, the fumonisin B series, particularly fumonisin B1 (FB1) and fumonisin B2 (FB2), are the most prevalent and toxic in this group of mycotoxins and are of public health significance due to the many debilitating human and animal diseases and mycotoxicosis they cause and their classification as by the International Agency for Research on Cancer (IARC) as a class 2B carcinogen (probable human carcinogen). This has made them one of the most regulated mycotoxins, with stringent regulatory limits on their levels in food and feeds destined for human and animal consumption, especially maize and maize-based products. Numerous countries have regulations on levels of fumonisins in foods and feeds that are intended to protect human and animal health. However, there are still gaps in knowledge, especially with regards to the molecular mechanisms underlying fumonisin-induced toxicity and their full impact on human health. Detection of fumonisins has been advanced through various methods, with immunological approaches such as Enzyme-Linked Immuno-Sorbent Assay (ELISA) and lateral flow immunoassays being widely used for their simplicity and adaptability. However, these methods face challenges such as cross-reactivity and matrix interference, necessitating the need for continued development of more sensitive and specific detection techniques. Chromatographic methods, including HPLC-FLD, are also employed in fumonisin analysis but require meticulous sample preparation and derivitization due to the low UV absorbance of fumonisins. This review provides a comprehensive overview of the fumonisin family, focusing on their biosynthesis, occurrence, toxicological effects, and levels of contamination found in foods and the factors affecting their presence. It also critically evaluates the current methods for fumonisin detection and quantification, including chromatographic techniques and immunological approaches such as ELISA and lateral flow immunoassays, highlighting the challenges associated with fumonisin detection in complex food matrices and emphasizing the need for more sensitive, rapid, and cost-effective detection methods. 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