Metal-organic framework-based aptasensor utilizing a novel electrochemiluminescence system for detecting acetamiprid residues in vegetables.

The work was based on N-(4-Aminobutyl)-N-ethylisoluminol (ABEI)-functionalized Fe-MIL-101 and gold nanoparticles (AuNPs) as sensing materials, and an electrochemiluminescence (ECL) aptasensor was constructed for detecting acetamiprid. As a metal-organic framework (MOF) material, Fe-MIL-101, was renowned for its unique three-dimensional network structure and efficient catalytic capability. ABEI, a common ECL reagent, was widely applied. ABEI was introduced into the Fe-MIL-101 structure as a luminescence functionalization reagent to form Fe-MIL-101@ABEI. This approach avoided limitations on the loading capacity of luminescent reagents imposed by modification and encapsulation methods. With character of excellent catalytic activity and ease of bioconjugation, AuNPs offered significant advantages in biosensing. Leveraging the reductive properties of ABEI, AuNPs were reduced around Fe-MIL-101@ABEI, resulting in the modified luminescent functionalized material denoted as Fe-MIL-101@ABEI@AuNPs. An aptamer was employed as a recognition element and was modified accordingly. The aptamer was immobilized on Fe-MIL-101@ABEI@AuNPs through gold-sulfur (Au-S) bonds. After capturing acetamiprid, the aptamer induced a decrease in the ECL signal intensity within the ABEI-hydrogen peroxide (H2O2) system, enabling the quantitative detection of acetamiprid. The aptasensor displayed remarkable stability and repeatability, featured a detection range of 1×10-3-1×102 nM, and had a limit of detection (LOD) of 0.3 pM (S/N=3), which underscored its substantial practical application potential.

Antimicrobial resistance and genetic relatedness of Salmonella serotypes isolated from food, asymptomatic carriers, and clinical cases in Shiyan, China.

Salmonella is a primary cause of foodborne diseases globally. Despite food contamination and clinical infections garnering substantial attention and research, asymptomatic Salmonella carriers, potential sources of infection, have been comparatively overlooked. In this study, we conducted a comparative analysis of serotype distribution, antimicrobial resistance phenotypes, and genetic profiles of archived Salmonella strains isolated from food (26), asymptomatic carriers (41), and clinical cases (47) in Shiyan City, China. Among the 114 Salmonella strains identified, representing 31 serotypes and 34 Sequence Types (STs), the most prevalent serovars included Typhimurium, Derby, Enteritidis, Thompson, and London, with the most predominant STs being ST11, ST40, ST26, ST34, and ST155. Antimicrobial resistance testing revealed that all strains were only sensitive to meropenem, with 74.6% showing antimicrobial resistance (AMR) and 53.5% demonstrating multidrug resistance (MDR). Strains resistant to five and six classes of antibiotics were the most common. Pearson's chi-square test showed no statistically significant difference in the occurrence of AMR (p = 0.105) or MDR (p = 0.326) among Salmonella isolates from the three sources. Our findings underscore associations and diversities among Salmonella strains isolated from food, asymptomatic carriers, and clinical patients, emphasizing the need for increased vigilance towards asymptomatic Salmonella carriers by authorities.

Advances in signal amplification strategies applied in pathogenic bacteria apta-sensing analysis-A review.

Pathogenic bacteria are primarily kinds of food hazards that provoke serious harm to human health via contaminated or spoiled food. Given that pathogenic bacteria continue to reproduce and expand once they contaminate food, pathogenic bacteria of high concentration triggers more serious losses and detriments. Hence, it is essential to detect low-dose pollution at an early stage with high sensitivity. Aptamers, also known as "chemical antibodies", are oligonucleotide sequences that have attracted much attention owing to their merits of non-toxicity, small size, variable structure as well as easy modification of functional group. Aptamer-based bioanalysis has occupied a critical position in the field of rapid detection of pathogenic bacteria. This is attributed to the unique advantage of using aptamers as recognition elements in signal amplification strategies. The signal amplification strategy is an effective means to improve the detection sensitivity. Some diverse signal amplification strategies emphasize the synthesis and assembly of nanomaterials with signal amplification capabilities, while others introduce various nucleic acid amplification techniques into the detection system. This review focuses on a variety of signal amplification strategies employed in aptamer-based detection approaches to pathogenic bacteria. Meanwhile, we provided a detailed introduction to the design principles and characteristics of signal amplification strategies, as well as the improvement of sensor sensitivity. Ultimately, the existing issues and development trends of applying signal amplification strategies in apta-sensing analysis of pathogenic bacteria are critically proposed and prospected. Overall, this review discusses from a new perspective and is expected to contribute to the further development of this field.

Fluorescent aptasensor mediated with multiple ssDNA for sensitive detection of acetamiprid in vegetables based on magnetic Fe3O4/C-assisted separation.

Acetamiprid (ACE) is a highly effective broad-spectrum insecticide, and its widespread use is potentially harmful to human health and environmental safety. In this study, magnetic Fe3O4/carbon (Fe3O4/C), a derivative of metal-organic framework MIL-101 (Fe), was synthesized by a two-step calcination method. And a fluorescent sensing strategy was developed for the efficient and sensitive detection of ACE using Fe3O4/C and multiple complementary single-stranded DNA (ssDNA). By using aptamer with multiple complementary ssDNA, the immunity of interference of the aptasensor was improved, and the aptasensor showed high selectivity and sensitivity. When ACE was present, the aptamer (Apt) combined with ACE. The complementary strand of Apt (Cs1) combined with two short complementary strands of Cs1, fluorophore 6-carboxyfluorescein-labeled complementary strand (Cs2-FAM) and the other strand Cs3. The three strands formed a double-stranded structure, and fluorescence would not be quenched by Fe3O4/C. In the absence of ACE, Cs2-FAM would be in a single-chain state and would be adsorbed by Fe3O4/C, and the fluorescence of FAM would be quenched by Fe3O4/C via photoelectron transfer. This aptasensor sensitively detected ACE over a linear concentration range of 10-1000 nM with a limit of detection of 3.41 nM. The recoveries of ACE spiked in cabbage and celery samples ranged from 89.49% to 110.76% with high accuracy.

Research progress on preparation methods and sensing applications of molecularly imprinted polymer-aptamer dual recognition elements.

The aptamer (Apt) and the molecularly imprinted polymer (MIP), as effective substitutes for antibodies, have received widespread attention from researchers because of their creation. However, the low stability of Apt in harsh detection environment and the poor specificity of MIP have hindered their development. Therefore, some researchers have attempted to combine MIP with Apt to explore whether the effect of "1 + 1 > 2" can be achieved. Since its first report in 2013, MIP-Apt dual recognition elements have become a highly focused research direction in the fields of biology and chemistry. MIP-Apt dual recognition elements not only possess the high specificity of Apt and the high stability of MIP in harsh detection environment, but also have high sensitivity and affinity. They have been successfully applied in medical diagnosis, food safety, and environmental monitoring fields. This article provides a systematic overview of three preparation methods for MIP-Apt dual recognition elements and their application in eight different types of sensors. It also provides effective insights into the problems and development directions faced by MIP-Apt dual recognition elements.

Clinical value of SLC12A9 for diagnosis and prognosis in colorectal cancer.

OBJECTIVE: The goal of the study is to assess the clinical value and the potential mechanism of SLC12A9 combing transcriptome and single cell sequencing data. METHODS: In this study, the expression level and the receiver operating characteristic curve analysis of SLC12A9 in CRC and normal tissue were analyzed in multiple data cohort. The standardized mean difference (SMD) calculation and the summary receiver operating characteristic (SROC) analysis were performed further to detect its diagnostic ability and expression level. KM survival analysis was performed to assess the prognosis value of SLC12A9. The expression level of SLC12A9 in different clinical characteristics was analyzed to explore the clinical value. Single cell data was studied to reveal the potential mechanism of SLC12A9. The correlation analysis of immunoinfiltration was performed to detect the potential immune cell related to SLC12A9. The nomogram was drawn to assess the probable mortality rate of CRC patient. RESULTS: We found that SLC12A9 was significantly up-regulated with the moderate diagnostic value in CRC. Patients with overexpressed SLC12A9 had a worse prognosis. SLC12A9 was related to Age, Pathologic N stage, Pathologic M stage, Lymphatic invasion and Pathologic stage (p < 0.05). The 1, 3 and 5-year survival rates of patient named TCGA-G4-6309 are 0.959, 0.897 and 0.827. PCR also showed that SLC12A9 was overexpressed in CRC comparing with normal tissue. CONCLUSION: In conclusion, our study comprehensively analyzed the clinical value of SLC12A9 and its potential mechanism, as well as immune cell infiltration, which may accelerate the diagnosis and improve the prognosis of CRC.

A novel electrochemical aptasensor based on core-shell nanomaterial labeling for simultaneous detection of acetamiprid and malathion.

At present, due to the coexistence of multiple pesticides in vegetables and the enhanced toxicity, a simultaneous detection method for multiple pesticides is urgently needed. In this work, two types of core-shell nanomaterials, Ag-Au core-shell nanoparticles (Ag@Au NPs) and Cu2O-Au core-shell nanoparticles (Cu2O@Au NPs), were synthesized and labeled with acetamiprid aptamer and malathion aptamer to prepare two novel electroactive signal probes, respectively. The two probes were hybridized on the surface of the electrode by the principle of base complementary pairing between the aptamers and the thiolated DNA oligonucleotide sequences, and a dual-signal electrochemical aptasensor for the simultaneous detection of acetamiprid and malathion was established by modified glassy carbon electrode (GCE). The limits of detection (LOD) were calculated to be 43.7 pg mL-1 for acetamiprid and 63.4 pg mL-1 for malathion. The aptasensor determined acetamiprid and malathion in spinach and rape with the recovery rates of 88.9%-112.5% and 98.0%-114.1%, respectively.

Novel Dual-Signal SiO2-COOH@MIPs Electrochemical Sensor for Highly Sensitive Detection of Chloramphenicol in Milk.

In view of the great threat of chloramphenicol (CAP) to human health and the fact that a few producers have illegally used CAP in the food production process to seek economic benefits in disregard of laws and regulations and consumer health, we urgently need a detection method with convenient operation, rapid response, and high sensitivity capabilities to detect CAP in food to ensure people's health. Herein, a molecularly imprinted polymer (MIP) electrochemical sensor based on a dual-signal strategy was designed for the highly sensitive analysis of CAP in milk. The NiFe Prussian blue analog (NiFe-PBA) and SnS2 nanoflowers were modified successively on the electrode surface to obtain dual signals from [Fe(CN)6]3-/4- at 0.2 V and NiFe-PBA at 0.5 V. SiO2-COOH@MIPs that could specifically recognize CAP were synthesized via thermal polymerization using carboxylated silica microspheres (SiO2-COOH) as carriers. When the CAP was adsorbed by SiO2-COOH@MIPs, the above two oxidation peak currents decreased at the same time, allowing the double-signal analysis. The SiO2-COOH@MIPs/SnS2/NiFe-PBA/GCE sensor used for determining CAP was successfully prepared. The sensor utilized the interactions of various nanomaterials to achieve high-sensitivity dual-signal detection, which had certain innovative significance. At the same time, the MIPs were synthesized using a surface molecular imprinting technology, which could omit the time of polymerization and elution and met the requirements for rapid detection. After optimizing the experimental conditions, the detection range of the sensor was 10-8 g/L-10-2 g/L and the limit of detection reached 3.3 × 10-9 g/L (S/N = 3). The sensor had satisfactory specificity, reproducibility, and stability, and was successfully applied to the detection of real milk samples.

Bipotential-resolved electrochemiluminescence biosensor based on Bi2S3@Au nanoflowers for simultaneous detection of Cd(II) and ampicillin in aquatic products.

In this paper, an electrochemiluminescence (ECL) biosensor was constructed using Bi2S3@Au nanoflowers as the based nanomaterial and Au@luminol and CdS QDs as independent ECL emission signal respectively. As the substrate of the working electrode, Bi2S3@Au nanoflowers improved the effective area of electrode and accelerated electron transfer rate between gold nanoparticles and aptamer, provided a good interface environment for the loading of luminescent materials. Then, the Au@luminol functionalized DNA2 probe was used as an independent ECL signal source under positive potential and recognized Cd(II), while the CdS QDs functionalized DNA3 probe was used as an independent ECL signal source under negative potential and recognized ampicillin. The simultaneous detection of Cd(II) and ampicillin in different concentrations are realized. This sensor not only has good selectivity and high sensitivity in real sample detection, but also open up a novel way to construct multi-target ECL biosensor for simultaneous detection.

Isolation of Bacteria Aptamers with Non-SELEX for the Development of a Highly Sensitive Colorimetric Assay Based on Dual Signal Amplification.

In this work, an aptamer against Escherichia coli is isolated via non-SELEX, which executes efficient selection by employing repetitive cycles of centrifugation-based partitioning, and the binding site of the aptamer on E. coli cell surfaces is inferred to be a membrane protein. Moreover, truncated sequence 2-17-2 with a higher affinity (Kd = 101.76 nM) is employed for highly sensitive colorimetric detection of bacteria based on the dual signal amplification strategy. When targets exist, the release of DNA 1 from the polymer activates a hybridization chain reaction (HCR) between DNA 1 and DNA 2, thereby inducing the aggregation of probe 1. Subsequently, DNA 3 dissociated from probe 1 as a linker DNA further assembles probe 2/3. In this system, two types of DNA@gold nanoparticles (AuNPs) coexist and successively aggregate AuNPs based on divergent triggering mechanisms. Under optimal conditions, the dual signal amplification strategy presents excellent sensitivity (10 CFU mL-1) and specificity, as well as the realization of real sample analysis.

Novel colorimetric aptasensor based on MOF-derived materials and its applications for organophosphorus pesticides determination.

For the visual detection of four organophosphorus pesticides (OPs), a colorimetric aptasensor was developed based on aptamer-mediated bimetallic metal-organic frameworks (MOFs) nano-polymers. Fe-Co magnetic nanoparticles (MNPs) and Fe-N-C nanozymes were prepared based on pyrolytic reaction, and were labeled with broad spectrum aptamers and complementary chains of organophosphorus pesticides respectively. The hybridization of aptamers and complementary chains led to the formation of nano-polymers. In the presence of target pesticides, they competed with complementary chains for aptamers on Fe-Co MNPs, resulting in a large number of Fe-N-C nanozymes signal labels being released into the supernatant. Fe-N-C nanozymes showed similar activity to peroxidase and catalyzed the 3,3',5,5'-tetramethylbenzidine-hydrogen peroxide (TMB-H2O2) color system to turn the solution blue-green under mild conditions. The magnetic probes had good selectivity and sensitivity, and were easily separated by magnetic absorption. The sensor functioned well under optimal conditions, demonstrating good stability and specificity for four pesticides: phorate, profenofos, isocarbophos and omethoate, and the detection limits of four pesticides were as low as 0.16 ng/mL, 0.16 ng/mL, 0.03 ng/mL and 1.6 ng/mL respectively, and the recovery rate of OPs residue in vegetable samples was satisfactory. The work described here provided a simple, rapid and sensitive way to construct a biosensor.

Ultrasensitive electrochemiluminescence aptasensor based on ABEI reduced silver nanoparticles for the detection of profenofos.

An ultrasensitive electrochemiluminescence (ECL) aptasensor for detection of profenofos was constructed by the reducibility and chemiluminescence property of N-(aminobutyl)-N-(ethylisoluminol) (ABEI). ABEI was used to reduce silver nitrate (AgNO3) to silver nanoparticles (AgNPs), which could be adsorbed on the lattice of graphene oxide (GO) to form ABEI-AgNPs-GO complex. This compound could achieve excellent luminescence. The aptamer (Apt) modified (5') by sulfhydryl groups could be immobilized on AgNPs to capture profenofos. When profenofos was present, the ECL signal of the aptasensor would be weakened. To further demonstrate the successful construction of the aptasensor, cyclic voltammetry tests were performed on an electrochemical workstation and an ECL analyzer, respectively. The standard curve and specificity experiment both showed that the sensor had the advantages of low limit of detection (LOD) and good specificity. Under the optimal conditions, the aptasensor had a good linear response for profenofos in the range of 1 × 10-1-1 × 104 ng/mL. It also had a LOD of 6.7 × 10-2 ng/mL and a correlation coefficient (R2) of 0.9991. The aptasensor had been successfully applied to the detection of profenofos in vegetables. The recovery range of the proposed ECL aptasensor was 98 % ~ 107.4 %.

Dual-Target Electrochemical Sensor Based on 3D MoS2-rGO and Aptamer Functionalized Probes for Simultaneous Detection of Mycotoxins.

A dual-target aptamer functionalized probes (DTAFP) was applied for the detection of aflatoxin B1 (AFB1) and zearalenone (ZEN) simultaneously, which has not been reported. Meanwhile, two functional materials for signal amplification of the DTAFP were synthesized: 1) a three-dimensional molybdenum disulfide-reduced graphene oxide (MoS2-rGO) as a favorable loading interface; 2) a double-probes gold nanoparticles (AuNPs) modified by Thionin (Thi) and 6-(Ferrocenyl) hexanethiol (FC6S) as distinguishable and non-interfering signals. Mycotoxins on the electrode surface release into solution under the function of the DTAFP, leading a reduction of the differential peak impulse in signal response. Under the optimum conditions, the aptasensor exhibited a detection range of 1.0 pg mL-1-100 ng mL-1 for AFB1 and ZEN, with no observable cross reactivity. In addition, the aptasensor performed excellent stability, reproducibility, specificity, and favorable recovery in the detection of edible oil. This work demonstrated a novel method for the construction of a simple, rapid, and sensitive aptasensor in the detection of multiple mycotoxins simultaneously.

Fermentation performance and metabolomic analysis of an engineered high-yield PUFA-producing strain of Schizochytrium sp.

The ω-3/long-chain polyunsaturated fatty acids (LC-PUFAs) play an important role in human health, but they cannot be synthesized in sufficient amounts by the human body. In a previous study, we obtained an engineered Schizochytrium sp. strain (HX-RS) by exchanging the acyltransferase (AT) gene, and it was able to co-produce docosahexaenoic acid and eicosapentaenoic acid. To investigate the mechanism underlying the increase of PUFA content in HX-RS, the discrepancies of fermentation performance, key enzyme activities and intracellular metabolites between HX-RS and its wild-type parent strain (WTS) were analyzed via fed-batch fermentation in 5-L bioreactors. The results showed that the cell dry weight (CDW) of HX-RS was higher than that of the WTS. Metabolomics combined with multivariate analysis showed that 4-aminobutyric acid, proline and glutamine are potential biomarkers associated with cell growth and lipid accumulation of HX-RS. Additionally, the shift of metabolic flux including a decrease of glyceraldehyde-3-phosphate content, high flux from pyruvate to acetyl-CoA, and a highly active glycolysis pathway were also found to be closely related to the high PUFA yield of the engineered strain. These findings provide new insights into the effects of exogenous AT gene expression on cell proliferation and fatty acid metabolism.

Influence of oxygen on the biosynthesis of polyunsaturated fatty acids in microalgae.

As one of the most important environmental factors, oxygen is particularly important for synthesis of n-3 polyunsaturated fatty acids (n-3 PUFA) in microalgae. In general, a higher oxygen supply is beneficial for cell growth but obstructs PUFA synthesis. The generation of reactive oxygen species (ROS) under aerobic conditions, which leads to the peroxidation of lipids and especially PUFA, is an inevitable aspect of life, but is often ignored in fermentation processes. Irritability, microalgal cells are able to activate a number of anti-oxidative defenses, and the lipid profile of many species is reported to be altered under oxidative stress. In this review, the effects of oxygen on the PUFA synthesis, sources of oxidative damage, and anti-oxidative defense systems of microalgae were summarized and discussed. Moreover, this review summarizes the published reports on microalgal biotechnology involving direct/indirect oxygen regulation and new bioreactor designs that enable the improved production of PUFA.

Transcriptomic Analysis of the Regulation of Lipid Fraction Migration and Fatty Acid Biosynthesis in Schizochytrium sp.

Schizochytrium sp. is the main source of docosahexaenoic acid-rich oil, which is widely used in food additive and pharmaceutical industry. In this study, using RNA-seq, comparative transcriptomic analyses were performed at four stages of DHA fermentation by Schizochytrium sp to get potential genes related to cell transition from cell growth to lipid accumulation and then to lipid turnover. 1406, 385, 1384 differently expressed genes were identified by comparisons in pairs of S2 vs S1, S3 vs S2 and S4 vs S3. Functional analysis revealed that binding and single-organism process might be involve in the cell transition from cell growth to lipid accumulation while oxidation-reduction process played an important role in the transition from lipid accumulation to lipid turnover. pfaC in the PKS pathway showed higher sensitivity to the environmental change, which might be the key regulator for enhancing PUFA biosynthesis in the future. Some other genes in signal transduction and cell transport were revealed to be related to lipid turnover, which would enrich the current knowledge regarding lipid metabolism and help to enhance the DHA production and enrich different lipid fractions by Schizochytrium in the future.