Chl1, an ATP-Dependent DNA Helicase, Inhibits DNA:RNA Hybrids Formation at DSB Sites to Maintain Genome Stability in S. pombe.

As an ATP-dependent DNA helicase, human ChlR1/DDX11 (Chl1 in yeast) can unwind both DNA:RNA and DNA:DNA substrates in vitro. Studies have demonstrated that ChlR1 plays a vital role in preserving genome stability by participating in DNA repair and sister chromatid cohesion, whereas the ways in which the biochemical features of ChlR1 function in DNA metabolism are not well understood. Here, we illustrate that Chl1 localizes to double-strand DNA break (DSB) sites and restrains DNA:RNA hybrid accumulation at these loci. Mutation of Chl1 strongly impairs DSB repair capacity by homologous recombination (HR) and nonhomologous end-joining (NHEJ) pathways, and deleting RNase H further reduces DNA repair efficiency, which indicates that the enzymatic activities of Chl1 are needed in Schizosaccharomyces pombe. In addition, we found that the Rpc37 subunit of RNA polymerase III (RNA Pol III) interacts directly with Chl1 and that deletion of Chl1 has no influence on the localization of Rpc37 at DSB site, implying the role of Rpc37 in the recruitment of Chl1 to this site.

A fluorometric method for aptamer-based simultaneous determination of two kinds of the fusarium mycotoxins zearalenone and fumonisin B1 making use of gold nanorods and upconversion nanoparticles.

An aptamer-based assay for the determination of two different kinds of fusarium mycotoxins, i.e., zearalenone (ZEN) and fumonisin B1 (FB1), is presented. Based on the inner filter effect (IFE) strategy, the contents of ZEN and FB1 can be simultaneously quantified. It is making use of 65-nm gold nanorods (AuNRs), 20-nm upconversion nanoparticles (UCNPs), fluorescence dyes, and DNA sequences. In the absence of ZEN and FB1, the UCNPs and AuNRs associate through DNA sequences. Due to IFE effect, weak fluorescence signals are collected. In the presence of ZEN or FB1, UCNPs and AuNRs become unstable and partially separate from each other. This results in the recovery of fluorescence signals. Under 980-nm laser excitation, the logarithmic values of fluorescence signal intensities at 606 nm and 753 nm gradually increase with the concentration of ZEN and FB1 in the ranges 0.05-100 µg L-1 (the coefficient of determination is 0.997) and 0.01-100 ng L-1 (the coefficient of determination is 0.986), respectively. The limits of detection (LOD) of the fabricated assay for ZEN and FB1 are 0.01 µg L-1 and 0.003 ng L-1, respectively. The proposed method has a high selectivity over other competitive mycotoxins, including aflatoxin B1, ochratoxin A, patulin and ochratoxin B. The applicability of the assay was evaluated in the determination of ZEN and FB1 contents in spiked corn samples. The average recoveries ranged from 89.9 to 106.6%. This result confirms the practicality of this method. Graphical abstract Schematic representation of an aptamer-based fluorometric method for simultaneous determination of two kinds of the fusarium mycotoxins zearalenone and fumonisin B1.

Aptamer and gold nanorod-based fumonisin B1 assay using both fluorometry and SERS.

An aptamer-based assay is presented for the determination of fumonisin B1 (FB1). It is bimodal in that both surface-enhanced Raman spectroscopy (SERS) and fluorometry are applied for quantitation. It makes use of platinum-coated gold nanorod (AuNR) and DNA sequences. The complementary DNA of aptamer (cDNA) against FB1 is immobilized on the surface of AuNR. The aptamer of FB1 modified with Cy5.5 are complementarily hybridized with cDNA. In the absence of FB1, the aptamer and its cDNA associate. In this situation, strong SERS and weak fluorescence signals are obtained. In the presence of FB1, the aptamer disassociates with its cDNA and binds the target. As the concentration of FB1 increases, the SERS and fluorescence signal intensities of the mixture are gradually decreased and increased, respectively. Under optimized conditions, the SERS signal at 1366 cm-1 decreases linearly in the 10-500 pg mL-1 concentration range with the calibration equation of y = 1997lgx-594 (the coefficient of determination is 0.998). The fluorescence signal at 670 nm increases linearly in the 10-250 pg mL-1 concentration range with the calibration equation of y = 500lgx-383 (the coefficient of determination is 0.991). The assay was applied to the determination of FB1 contents in spiked corn samples. The average recoveries ranged from 92 to 107%, confirming the practicality of this method. The results obtained by this assay are in good agreement with that of LC-MS/MS method. Graphical abstractSchematic illustration of a bimodal aptasensor based on surface enhanced Raman scattering (SERS) and fluorescence change for the detection of fumonisin B1 (FB1).

Trimer-based aptasensor for simultaneous determination of multiple mycotoxins using SERS and fluorimetry.

An aptasensor is reported for the detection of three different kinds of mycotoxins, i.e., zearalenone (ZEN), ochratoxin A (OTA), and fumonisin B1 (FB1). Based on fluorescence resonance energy transfer effect (FRET) and surface-enhanced Raman scattering (SERS), the levels of ZEN, FB1, and OTA can be simultaneously determined. Under 980-nm and 650-nm laser excitation, the logarithmic values of fluorescence signal intensities at 543 nm and 670 nm are slowly increased as the concentrations of ZEN and OTA vary from 0.1 ng mL-1 and 0.05 ng mL-1 to 100 ng mL-1 and 25 ng mL-1, respectively. For FB1, under 980-nm laser excitation, the logarithmic value of SERS signal intensity at 1567 cm-1 gradually increases with the concentration of FB1 in the range 0.05-200 pg mL-1 (R2 = 0.996). The detection limits of the proposed assay for ZEN, OTA, and FB1 are 0.03 ng mL-1, 0.01 ng mL-1, and 0.02 pg mL-1, respectively. The selectivity experiment results indicate this assay possesses a high selectivity over other commonly encountered mycotoxins. The average recoveries range from 90 to 107%, revealing satisfactory application potential of the proposed assay. The developed aptasensor will bring bright prospects for research in the field of multiplexed mycotoxine detection. Graphical Abstract Schematic representation of an aptamer-based assay for multiple mycotoxins determination.

A fluorometric assay for staphylococcal enterotoxin B by making use of platinum coated gold nanorods and of upconversion nanoparticles.

An aptamer based fluorometric assay is presented for fast and accurate detection of staphylococcal enterotoxin B (SEB). It is making use of platinum-coated gold nanorods (AuNR@Pt) and upconversion nanoparticles (UCNPs). The aptamer against SEB is immobilized on AuNR@Pt while the complementary DNA fragment of SEB aptamer is immobilized on UCNPs. As the concentration of SEB increases, the fluorescence of the satellite assembly (AuNR@Pt-UCNPs) is gradually restored. Under the optimized conditions, fluorescence (best measured at excitation/emission wavelengths of 980/543 nm) linearly increases in the 2.0-400 pg·mL-1 SEB concentration range. The limit of detection is as low as 0.9 pg·mL-1 (at an S/N of 3), significantly lower than existing methods. The method was applied to the determination of SEB in spiked milk samples. The average recoveries ranged from 91.2% to 104.6%, confirming the practicality of this method. Graphical abstract Schematic illustration of a fluorometric assay based on inner filter effect (IFE) between platinum coated gold nanorods (AuNR@Pt) and upconversion nanoparticles (UCNPs) for the determination of staphylococcal enterotoxin B (SEB).

A bimodal (SERS and colorimetric) aptasensor for the detection of Pseudomonas aeruginosa.

An aptamer based assay is described for the determination of Pseudomonas aeruginosa (P. aeruginosa). It is bimodal in that both surface enhanced Raman spectroscopy (SERS) and colorimetry are applied for quantitation. The aptamer against P. aeruginosa and its corresponding complementary DNA fragment (cDNA) were conjugated to two kinds of differently sized gold nanoparticles (AuNPs). The 30 nm AuNPs carrying the aptamer are used as color signal probes, while the cDNA-15 nm AuNP conjugates serve as SERS signalling probes. In the absence of P. aeruginosa, the two probes assemble to form the duplex structure. When the probes are exposed to P. aeruginosa, the aptamer dissociates from its cDNA and binds the target. After centrifugation, the SERS signal from the supernatant decreased, due to the decreased electromagnetic effect. On addition of 3,3',5,5'-tetramethylbenzidine (TMB) and hydrogen peroxide, a relatively rapid catalytic reaction between horseradish peroxidase (linked to the aptamer on the 30-nm AuNPs) and TMB occurred, resulting in the formation of a green color with an absorption peak at 640 nm. The reliability of the approach was validated by detecting different levels of P. aeruginosa in spiked tap water and chicken meat samples. The average recoveries ranged from 88% to 112%, confirming the practicality of this method. In our perception, this dual mode aptasensor paves the way for accurate and reliable determination of P. aeruginosa. Conceivably, the method has a wide scope in that it may be extended to the determination of various other species for which respective aptamers are available. Graphical abstract Schematic illustration of a bimodal aptasensor based on surface enhanced Raman scattering (SERS) and color change for the detection of Pseudomonas aeruginosa.

Intelligent food tag: A starch-anthocyanin-based pH-sensitive electrospun nanofiber mat for real-time food freshness monitoring.

A starch-based nanofiber mat was prepared for real-time monitoring of food freshness for the first time. UV-vis results showed that roselle anthocyanins (RS) conferred a wide pH sensing range on the nanofiber mat. The prepared nanofiber mats demonstrated good color visibility (total color difference value (ΔE) increased to 56.4 ± 0.7) and a reversible response (within 120 s). Scanning electron microscopy and Fourier transform infrared spectroscopy results suggested that the nanofibers had smooth surfaces without beaded fibers and that RS was well embedded into the nanofibers. The introduction of RS improved the thermal stability of the nanofibers. Color stability tests revealed that the nanofibers exhibited excellent color stability (maximum change ΔE = 1.57 ± 0.03) after 14 days of storage. Pork and shrimp freshness tests verified that the nanofibers could effectively reflect the dynamic freshness of pork and shrimp. Nontoxic, degradable and responsive characteristics make the pH-sensitive nanofiber mat a smart food label with great application potential.

Improving structural and functional properties of starch-catechin-based green nanofiber mats for active food packaging by electrospinning and crosslinking techniques.

The hydrophilic and low mechanical properties limited the application of starch-based films. In this work, a hydrophobic starch-based nanofiber mat was first successfully prepared from aqueous solution at room temperature by using electrospinning and glutaraldehyde (GTA) vapor phase crosslinking techniques for active packaging applications. Catechin (CAT) was immobilized in the nanofibers by electrospinning, resulting in higher thermal stability (Tdmax = 315.23 °C), antioxidant (DPPH scavenging activity = 94.31 ± 2.70 %) and antimicrobial (inhibition zone diameter = 15.6 ± 0.3 mm) of the fibers, which further demonstrated hydrogen bonding and electrostatic interaction between CAT and fibers. Nanofibers after GTA vapor phase crosslinking exhibited enhanced hydrophobicity (water contact angle: 15.6 ± 1.5° â†’ 93.5 ± 2.3°) and mechanical properties (tensile strength: 1.82 ± 0.06 MPa â†’ 7.64 ± 0.24 MPa, elastic modulus: 19.35 ± 0.63 MPa â†’ 45.34 ± 0.51 MPa). The results demonstrated that preparation of starch-based electrospun nanofiber mats in aqueous system at room temperature overcame the challenges of organic solvent pollution and thermosensitive material encapsulation, while GTA vapor phase crosslinking technique improved the hydrophobicity and mechanical properties of nanofiber mats, which facilitated the application of starch-based materials in the field of packaging.

Organic solvent-free starch-based green electrospun nanofiber mats for curcumin encapsulation and delivery.

Developing green and efficient methods for the delivery of active food substances is a sustained demand for food scientists and industries. In this work, for the first time, we prepared a curcumin (CUR)-loaded starch-based fast-dissolving nanofiber by electrospinning technology. This green nanofiber was obtained by incorporating CUR with octenyl succinic anhydride starch (OSA) and pullulan (PUL) matrix using pure water as the solvent. To overcome the poor water-solubility and bioavailability of CUR, hydroxypropyl-beta-cyclodextrin (HPßCD) was used to form inclusion complexes. Phase solubility test results showed that by introducing HPßCD, the water-solubility of CUR was obviously improved. The prepared electrospun nanofibers were systematically characterized through scanning electron microscopy (SEM), X-ray diffraction (XRD), proton nuclear magnetic resonance spectroscopy (1H NMR), Fourier transform infrared spectroscopy (FT-IR), differential scanning calorimetry (DSC), thermogravimetric analysis (TGA), encapsulation efficiency testing, solubility testing and antioxidant activity testing. The results demonstrated that CUR was well encapsulated into HPßCD and OSA/PUL/CUR-HPßCD electrospun nanofibers with fine morphology and fast-dissolving character were successfully prepared. It is worth noting that the whole process and raw materials were green, suggesting that the prepared fast-dissolving nanofiber has great application potential in the food and pharmaceutical fields.

Chl1 coordinates with H3K9 methyltransferase Clr4 to reduce the accumulation of RNA-DNA hybrids and maintain genome stability.

A genome-wide analysis in Schizosaccharomyces pombe indicated that double-deletion mutants of Chl1 and histone H3K9 methyltransferase complex factors are synthetically sick. Here, we show that loss of Chl1 increases the accumulation of RNA-DNA hybrids at pericentromeric dg and dh repeats in the absence of the H3K9 methyltransferase Clr4, which leads to genome instability, including more severe defects in chromosome segregation and increased chromatin accessibility. Localization of Chl1 at pericentromeric regions depends on a subunit of replication protein A (RPA), Ssb1. In wild-type (WT) cells, transcriptionally repressed heterochromatin prevents the formation of RNA-DNA hybrids. When Clr4 is deleted, dg and dh repeats are highly transcribed. Then Ssb1 associates with the displaced single-stranded DNA (ssDNA) and recruits Chl1 to resolve the RNA-DNA hybrids. Together, our data suggest that Chl1 coordinates with Clr4 to eliminate RNA-DNA hybrids, which contributes to the maintenance of genome integrity.

Improving the sensitivity of lateral flow immunoassay for Salmonella typhimurium detection via flow-rate regulation.

Application of the traditional immunochromatographic assay (ICGA) has been limited by its poor sensitivity. The objective of this study was to increase the sensitivity of the traditional ICGA. A dual-mode ICGA (D-M ICGA) was developed by combining a nanozyme-assisted signal-amplification strategy with a magnetic-nanoparticle-based flow-speed-control strategy. Salmonella typhimurium can be detected simultaneously based on color and magnetic signals in the detection area of the D-M ICGA strip. The calculated limits of detection of 50 cfu·mL-1 and 75 cfu·mL-1 in the color and magnetic modes, respectively, were approximately 1000 times lower than those of the traditional ICGA. The selectivity and practical applicability of the D-M ICGA were also confirmed in this study. The results prove that the D-M ICGA is an assay that could be used for Salmonella typhimurium detection and can be easily adapted to detect other pathogenic bacteria.

Establishment of a dual mode immunochromatographic assay for Campylobacter jejuni detection.

Campylobacter jejuni (C. jejuni) is considered one of the most common cause of human gastroenteritis. Aiming to detect C. jejuni in food products rapidly and sensitively, a dual mode lateral flow assay, based on the peroxidase mimicking and surface enhanced Raman scattering (SERS) enhancement properties of platinum coated gold nanorods (AuNR@Pt), was developed in this study. Under color mode and SERS mode, the proposed assay showed good linear response in the range of 102-106 cfu/mL and 102-5 × 106 cfu/mL with limits of detection of 75 cfu/mL and 50 cfu/mL, respectively (S/N = 3). Furthermore, the reliability of the dual-readout lateral flow assay (LFA) was successfully demonstrated by the application on milk samples, in which the recoveries ranged from 89.33% to 107.62%. Overall, the immunochromatographic assay developed in this work is promising and has good chance to be employed for sensitive detection of C. jejuni in food products.

Ultrasensitive detection of microcystin-LR with gold immunochromatographic assay assisted by a molecular imprinting technique.

A simple and highly sensitive lateral flow immunochromatographic assay (LFICA) towards microcystin-LR (MC-LR) was proposed in this study. Molecular imprinting technique was combined with enzyme assisted colorimetric method to substantially enhance the sensitivity of traditional LFICA. The target, i.e., MC-LR molecular, was first separated from the complex matrix by molecularly-imprinted polymers (MIPs). Then, the obtained MC-LR was detected by LFICA based on the blue color generated by an enzyme-substrate reaction. Using the proposed assay, under optimal conditions, the determination of MC-LR was doable in a wide linear range from 0.1 ng/mL to 100 ng/mL with a detection limit of 0.04 ng/mL. In addition, the reliability of the developed method was validated by determining MC-LR contents in real samples. The overall results suggested that the neoteric LFICA method possess great potential in sensitive detection of MC-LR.

A novel SERS-based aptasensor for ultrasensitive sensing of microcystin-LR.

We developed a novel aptasensor based on surface enhanced Raman spectroscopy (SERS) and applied it for highly sensitive detection of microcystin-LR (MC-LR). In this work, MC-LR aptamer and its corresponding complementary DNA fragments (cDNA) were conjugated to gold nanoparticles (AuNPs) and magnetic nanoparticles (MNPs), respectively. Then, MC-LR aptamer-AuNPs and cDNA-MNPs conjugates were used as signal probes and capture probes, respectively. The proposed assay exhibited a linearity range from 0.01 to 200 ng/mL with the limit of detection (LOD) of 0.002 ng/mL. In addition, the reliability of the novel approach was validated by detecting different levels of MC-LR spiked in tap water samples. Overall, the novel aptasensor paves a new way for rapid and accurate determination of MC-LR and can be referred to detect other hazardous substances in water products.

Rapid detection of ß-conglutin with a novel lateral flow aptasensor assisted by immunomagnetic enrichment and enzyme signal amplification.

A simple, rapid and economic lateral flow immunochromatographic assay (LFICA) was designed for ultrasensitive detection of ß-conglutin. Instead of antibodies and gold nanoparticles (AuNPs) used in conventional LFICA, a cognate aptamer duo, binding to ß-conglutin and Fe3O4@Au core-shell nanoparticles, was applied in this study. An enzyme signal amplification strategy was used to enhance sensitivity. In addition, a new magnetic enrichment strategy was employed to further enhance sensitivity of the assay, slowing down movement of the capture probe (i.e., Fe3O4@Au nanostructures) using an external magnetic field. The novel LFICA assay can be completed within 20 min and achieved a detection limit of 8 fM, a thousand-times lower than similar assays without magnetic focusing. Overall, our results demonstrated the potential for the proposed LFICA sensor in rapid detection of ß-conglutin without any special analytical expertise or instrumentations.