A capillary-based centrifugal indicator equipped with in situ pathogenic bacteria culture for fast antimicrobial susceptibility testing.

Antimicrobial resistance has become a major global health threat due to the misuse and overuse of antibiotics. Rapid, affordable, and high-efficiency antimicrobial susceptibility testing (AST) is among the effective means to solve this problem. Herein, we developed a capillary-based centrifugal indicator (CBCI) equipped with an in situ culture of pathogenic bacteria for fast AST. The bacterial incubation and growth were performed by macro-incubation, which seamlessly integrated the capillary indicator. Through simple centrifugation, all the bacterial cells were confined at the nanoliter-level capillary column. The packed capillary column height could linearly reflect the bacterial count, and the minimum inhibitory concentration (MIC) was determined based on the difference in the column height between the drug-added groups and the control group. The AST results could easily be determined by the naked eye or smartphone imaging. Thus, the CBCI realized the combination of macro-bacterial incubation and early micro assessment, which accelerated the phenotypic AST without complex microscopic counting or fluorescent labelling. The whole operation process was simple and easy to use. AST results could be determined for E. coli ATCC strains within 3.5 h, and the output results for clinical samples were consistent with the hospital reports. We expect this AST platform to become a useful tool in limiting antimicrobial resistance, especially in remote/resource-limited areas or in underdeveloped countries.

A Wide-Band Magnetoelectric Sensor Based on a Negative-Feedback Compensated Readout Circuit.

Magnetoelectric (ME) sensors cannot effectively detect broadband magnetic field signals due to their narrow bandwidth, and existing readout circuits are unable to vary the bandwidth of the sensors. To expand the bandwidth, this paper introduces a negative-feedback readout circuit, fabricated by introducing a negative-feedback compensation circuit based on the direct readout circuit of the ME sensor. The negative-feedback compensation circuit contains a current amplifier, a feedback resistor, and a feedback coil. For this purpose, a Metglas/PVDF/Metglas ME sensor was prepared. Experimental measurements show that there is a six-fold difference between the maximum and minimum values of the ME voltage coefficients in the 6-39 kHz frequency band for the ME sensor without the negative-feedback compensation circuit when the sensor operates at the optimal bias magnetic field. However, the ME voltage coefficient in this band remains stable, at 900 V/T, after the charge amplification of the direct-reading circuit and the negative-feedback circuit. In addition, experimental results show that this negative-feedback readout circuit does not increase the equivalent magnetic noise of the sensor, with a noise level of 240 pT/√Hz in the frequency band lower than 25 kHz, 63 pT/√Hz around the resonance frequency of 30 kHz, and 620 pT/√Hz at 39 kHz. This paper proposes a negative-feedback readout circuit based on the direct readout circuit, which greatly increases the bandwidth of ME sensors and promotes the widespread application of ME sensors in the fields of broadband weak magnetic signal detection and DBS electrode positioning.

Bent-Capillary-Centrifugal-Driven Monodisperse Droplet Generator with Its Application for Digital LAMP Assay.

We developed a bent-capillary-centrifugal-driven (BCCD) monodisperse droplet generator, which could achieve a perfect combination of driving and segmentation for the dispersed phase only using a rotating bent capillary immersed in the continuous phase (mineral oil). The sample could flow continuously to the bent-capillary outlet to form the droplet precursors, which were segmented into homogeneous droplets in the continuous phase. Through the investigation of influence factors on droplet size and stability, we found that the droplet size could be conveniently controlled by the rotational speed of the bent capillary. The droplet volumes could be adjusted with the range from 34 pL to 1 µL, and the coefficient variations (CVs) were less than 3%. Meanwhile, the BCCD droplet generator could realize the controllable droplet output with a high-efficiency sample utilization of 99.75 ± 1.15%, which offered a significant advantage in reducing the waste of precious samples in the droplet generation process. We validated this system with a digital loop-mediated isothermal amplification (dLAMP) assay for the absolute quantification of Mycobacterium tuberculosis complex nucleic acids. The results demonstrated that the BCCD droplet generator was easy to build, was of low cost, and was convenient to operate, as well as avoided sample loss and cross-contamination by coupling with a 96-well plate. Overall, the present platform, as a simple chip-free droplet generator, will provide an especially valuable droplet generation solution for biochemical applications based on droplets.

In silico analyses and experimental validation of the MHC class-I restricted epitopes of Ebolavirus GP.

Ebolavirus (EBOV) causes an extremely high mortality and prevalence disease called Ebola virus disease (EVD). There is only one glycoprotein (GP) on the virus particle surface, which mediates entry into the host cell. Major histocompatibility complex (MHC) class-I restricted cluster of differentiation 8 (CD8+) T cell responses are important antiviral immune responses. Therefore, it is of great importance to understand EBOV GP-specific MHC class-I restricted epitopes within immunogenicity. In this study, computational approaches were employed to predict the dominant MHC class-I molecule epitopes of EBOV GP for mouse H2 and major alleles of human leukocyte antigen (HLA) class-I supertypes. Our results yielded 42 dominant epitopes in H2 haplotypes and 301 dominant epitopes in HLA class-I haplotypes. After validation by enzyme-linked immunospot (ELISpot) assay, in-depth analyses to ascertain their nature of conservation, immunogenicity, and docking with the corresponding MHC class-I molecules were undertaken. Our study predicted MHC class-I restricted epitopes that may aid the advancement of anti-EBOV immune responses. An integrated strategy of epitope prediction, validation and comparative analyses was postulated, which is promising for epitope-based immunotherapy development and application to viral epidemics.

Cytogenetic Characterization and Molecular Marker Development for a Wheat-T. boeoticum 4Ab (4B) Disomic Substitution Line with Stripe Rust Resistance.

Triticum boeoticum (2n = 2x = 14, AbAb) is an important relative of wheat. This species tolerates many different types of environmental stresses, including drought, salt, and pathogenic infection, and is lower in dietary fiber and higher in antioxidants, protein (15 to 18%), lipids, and trace elements than common wheat. However, the gene transfer rate from this species to common wheat is low, and few species-specific molecular markers are available. In this study, the wheat-T. boeoticum substitution line Z1889, derived from a cross between the common wheat cultivar Crocus and T. boeoticum line G52, was identified using multicolor fluorescence in situ hybridization, multicolor genomic in situ hybridization, and a 55K single-nucleotide polymorphism array. Z1889 was revealed to be a 4Ab (4B) substitution line with a high degree of resistance to stripe rust pathogen strains prevalent in China. In addition, 22 4Ab chromosome-specific molecular markers and 11 T. boeoticum genome-specific molecular markers were developed from 1,145 4Ab chromosome-specific fragments by comparing the sequences generated by specific-length amplified fragment sequencing, with an efficiency of up to 55.0%. Furthermore, the specificity of these markers was verified in four species containing the Ab genome. These markers not only can be used for the detection of the 4Ab chromosome but also provide a basis for molecular marker-assisted, selection-based breeding in wheat.

Identification of the Solid Stem Suppressor Gene Su-TdDof in Synthetic Hexaploid Wheat Syn-SAU-117.

Lodging is one of the most important factors affecting the high and stable yield of wheat worldwide. Solid-stemmed wheat has higher stem strength and lodging resistance than hollow-stemmed wheat does. There are many solid-stemmed varieties, landraces, and old varieties of durum wheat. However, the transfer of solid stem genes from durum wheat is suppressed by a suppressor gene located on chromosome 3D in common wheat, and only hollow-stemmed lines have been created. However, synthetic hexaploid wheat can serve as a bridge for transferring solid stem genes from tetraploid wheat to common wheat. In this study, the F1, F2, and F2:3 generations of a cross between solid-stemmed Syn-SAU-119 and semisolid-stemmed Syn-SAU-117 were developed. A single dominant gene, which was tentatively designated Su-TdDof and suppresses stem solidity, was identified in synthetic hexaploid wheat Syn-SAU-117 by using genetic analysis. By using bulked segregant RNA-seq (BSR-seq) analysis, Su-TdDof was mapped to chromosome 7DS and flanked by markers KASP-669 and KASP-1055 within a 4.53 cM genetic interval corresponding to 3.86 Mb and 2.29 Mb physical regions in the Chinese Spring (IWGSC RefSeq v1.1) and Ae. tauschii (AL8/78 v4.0) genomes, respectively, in which three genes related to solid stem development were annotated. Su-TdDof differed from a previously reported solid stem suppressor gene based on its origin and position. Su-TdDof would provide a valuable example for research on the suppression phenomenon. The flanking markers developed in this study might be useful for screening Ae. tauschii accessions with no suppressor gene (Su-TdDof) to develop more synthetic hexaploid wheat lines for the breeding of lodging resistance in wheat and further cloning the suppressor gene Su-TdDof.

Neutrophil-to-Lymphocyte Ratio Predicts the Severity of Incarcerated Groin Hernia.

BACKGROUND The incarceration of a segment of bowel within a groin hernia can result in intestinal strangulation if hernia treatment is delayed. Once intestinal strangulation occurs, a bowel resection may be required, and there is an overall increased risk for postoperative complications. The aim of this study was to identify biomarkers to predict the severity of an incarcerated groin hernia. MATERIAL AND METHODS We retrospectively evaluated the records of 95 patients with incarcerated groin hernias who underwent emergency surgical correction of the hernias. The need for a bowel resection was regarded as an indicator of severity in incarcerated groin hernia patients. The patients were divided into 2 groups: patients with bowel resection surgery and patients without bowel resection surgery. RESULTS We discovered that leukocyte count (leukocyte count ≥10×10³/mm³), neutrophil-to-lymphocyte ratio (NLR, NLR ≥11.5), presentation of bowel obstruction, and duration of incarceration (duration of incarceration ≥26 h) were significantly associated with bowel resection in incarcerated groin hernia patients by using the chi-square test. Factors such as leukocyte count, NLR, presentation of bowel obstruction, and duration of incarceration were analyzed using multivariate logistic regression analysis. We found that NLR, presentation of bowel obstruction, and duration of incarceration were independently and significantly related to bowel resection in incarcerated groin hernia patients. CONCLUSIONS An elevated NLR can serve as a biomarker for the prediction of severity of incarcerated groin hernias. Additionally, incarcerated groin hernia patients who present with bowel obstruction or with duration of intestinal incarceration longer than 26 h have an increased risk for bowel resection.

Chromosome-level assembly and gene annotation of Decapterus maruadsi genome using Nanopore and Hi-C technologies.

Decapterus maruadsi is one of the representative offshore fish in the Western Pacific. Since the last century, it has become a commercially valuable marine fishery species in the Western Pacific region. Despite its high economic value, there is still a lack of high-quality reference genome of D. maruadsi in germplasm resource evaluation research. Here we report a chromosome-level reference genome of D. maruadsi based on Nanopore sequencing and Hi-C technologies. The whole genome was assembled through 169 contigs with a total length of 723.69 Mb and a contig N50 length of 24.67 Mb. By chromosome scaffolding, 23 chromosomes with a total length of 713.58 Mb were constructed. In addition, a total of 199.49 Mb repetitive elements, 33,515 protein-coding genes, and 6,431 ncRNAs were annotated in the reference genome. This reference genome of D. maruadsi will provide a solid theoretical basis not only for the subsequent development of genomic resources of D. maruadsi but also for the formulation of policies related to the protection of D. maruadsi.

Bismuth Ferrite-Based Lead-Free High-Entropy Piezoelectric Ceramics.

Piezoelectric ceramics, as essential components of actuators and transducers, have captured significant attention in both industrial and scientific research. The "entropy engineering" approach has been demonstrated to achieve excellent performance in lead-based materials. In this study, the "entropy engineering" approach was employed to introduce the morphotropic phase boundary (MPB) into the bismuth ferrite (BF)-based lead-free system. By employing this strategy, a serial of novel "medium to high entropy" lead-free piezoelectric ceramics were successfully synthesized, namely (1-x)BiFeO3-x(Ba0.2Sr0.2Ca0.2Bi0.2Na0.2)TiO3 (BF-xBSCBNT, x = 0.15-0.5). Our investigation systematically examined the phase structure, domain configuration, and ferroelectric/piezoelectric properties as a function of conformational entropy. Remarkable performances with a largest strain of 0.50% at 100 kV/cm, remanent polarization ∼40.07 µC/cm2, coercive field ∼74.72 kV/cm, piezoelectric coefficient ∼80 pC/N, and d33* ∼500 pm/V were achieved in BF-0.4BSCBNT ceramics. This exceptional performance can be attributed to the presence of MPB, coexisting rhombohedral and cubic phases, along with localized nanodomains. The concept of high-entropy lead-free piezoelectric ceramics in this study provides a promising strategy for the exploration and development of the next generation of lead-free piezoelectric materials.

A graphene oxide-assisted protein immobilization paper-tip immunosensor with smartphone and naked eye readout for the detection of okadaic acid.

BACKGROUND: Okadaic acid (OA), as a diarrhetic shellfish poisoning, can increase the risk of acute carcinogenic or teratogenic effects for the ingestion of OA contaminated shellfish. At present, much effort has been made to graft immunoassay onto a paper substrate to make paper-based sensors for rapid and simple detection of shellfish toxin. However, the complicated washing steps and low protein fixation efficiency on the paper substrate need to be further addressed. RESULTS: A novel paper-tip immunosensor for detecting OA was developed combined with smartphone and naked eye readout. The trapezoid paper tip was consisted of quantitative and qualitative detection zones. To improve the OA antigen immobilization efficiency on the paper substrate, graphene oxide (GO)-assisted protein immobilization method was introduced. Meanwhile, Au nanoparticles composite probe combined with the lateral flow washing was developed to simplify the washing step. The OA antigen-immobilized zone, as the detection zone Ⅰ, was used for quantitative assay by smartphone imaging. The paper-tip front, as the detection zone Ⅱ, which could qualitatively differentiate OA pollution level within 45 min using the naked eye. The competitive immunoassay on the paper tip exhibited a wide linear range for detecting OA (0.02-50 ng∙mL-1) with low detection limit of 0.02 ng∙mL-1. The recovery of OA in spiked shellfish samples was in the range of 90.3 %-113.%. SIGNIFICANCE: These results demonstrated that the proposed paper-tip immunosensor could provide a simple, low-cost and high-sensitivity test for OA detection without the need for additional large-scale equipment or expertise. We anticipate that this paper-tip immunosensor will be a flexible and versatile tool for on-site detecting the pollution of marine products.

Designing of a functional paper-tip substrate for sensitive surface-enhanced Raman spectroscopy (SERS) detection.

A simple and flexible fabrication method of paper SERS substrate was developed by nanoparticles (NPs) droplet self-assembly at the paper tip with a temperature gradient (PTTG). We turned the drawback of the coffee ring effect into an effective way of preparing paper SERS substrate. When the NPs droplets were continuously dripped onto the PTTG, NPs were densely and uniformly distributed at the paper-tip front based on the combination of gravity and the coffee ring effect, which could achieve 91.2-fold improvement of SERS performance compared to a flat filter paper. Meanwhile, the analytes could also be enriched at the paper-tip front, which could achieve 9.3-fold signal enhancement compared to the paper-tip tail. Thus, the PTTG realized an excellent signal amplification for SERS detection. The paper-tip SERS substrate combined with a portable Raman spectrometer yielded an excellent analytical enhancement factor of 1.15 × 105 with the detection limit of 10 nM Rhodamine 6G (R6G). The whole fabrication procedure was completed within 2 h, and the paper-tip substrate showed a satisfactory substrate-to-substrate reproducibility with a relative standard deviation (RSD) of 5.13% (n = 10). It was successfully applied for quantitatively detecting real samples of oxytetracycline and malachite green with recoveries of 83.84-105.25% (n = 3). Meanwhile, we further evaluated the SERS performance of the PTTG using a laboratory-based Raman spectrometer, and it could realize the detection as low as 10 pM R6G. The proposed paper-tip substrate would offer a promising potential application for the on-site SERS analysis of food safety and environmental health.

Corrigendum: Integration: gospel for immune bioinformatician on epitope-based therapy.

[This corrects the article DOI: 10.3389/fimmu.2023.1075419.].

Molecular cytogenetic identification and nutritional composition evaluation of newly synthesized Triticum turgidum-Triticum boeoticum amphiploids (AABBAbAb).

Triticum boeoticum Boiss. (AbAb, 2n = 2x = 14) is a wheat-related species with the blue aleurone trait. In this study, 18 synthetic Triticum turgidum-Triticum boeoticum amphiploids were identified, which were derived from crosses between T. boeoticum and T. turgidum. Three probes (Oligo-pTa535, Oligo-pSc119.2, and Oligo-pTa713) for multicolor fluorescence in situ hybridization (mc-FISH) were combined with genomic in situ hybridization (GISH) to identify chromosomal composition. Seven nutritional indices (anthocyanins, protein, total essential amino acids TEAA, Fe, Zn, Mn and Cu) were measured, and the nutritional components of 18 synthetic amphiploids were comprehensively ranked by principal component analysis (PCA). The results showed that all three synthetic amphiploids used for cytological identification contained 42 chromosomes, including 14 A, 14 B, and 14 Ab chromosomes. The average anthocyanin content was 82.830 µg/g to 207.606 µg/g in the whole meal of the 17 blue-grained lines (Syn-ABAb-1 to Syn-ABAb-17), which was obviously higher than that in the yellow-grained line Syn-ABAb-18 (6.346 µg/g). The crude protein content was between 154.406 and 180.517 g/kg, and the EAA content was 40.193-63.558 mg/g. The Fe, Zn, Mn and Cu levels in the 17 blue-grained lines were 60.55 to 97.41 mg/kg, 60.55-97.41 mg/kg, 35.11 to 65.20 mg/kg and 5.74 to 7.22 mg/kg, respectively, which were higher than those in the yellow-grained line. The contribution of the first three principal components reached 84%. The first principal component was mainly anthocyanins, Fe, Zn and Mn. The second principal component contained protein and amino acids, and the third component contained only Cu. The top 5 Triticum turgidum-Triticum boeoticum amphiploids were Syn-ABAb-11, Syn-ABAb-17, Syn-ABAb-5, Syn-ABAb-8 and Syn-ABAb-4. These amphidiploids exhibited the potential to serve as candidates for hybridization with common wheat, as indicated by comprehensive score rankings, toward enhancing the nutritional quality of wheat.

Aptamer-based signal amplification strategies coupled with microchips for high-sensitivity bioanalytical applications: A review.

With their advantages in specificity, high stability and easy screening, aptamers are becoming increasingly popular recognition elements for biosensor platforms. At the same time, microchips as the new analytical detection platforms have achieved significant growth in the past decades. At present, with the intersection of aptamer and microfluidic technology, aptamer-based high-sensitivity bioanalysis on microchips exhibits a great application potential in biomedical science and environmental fields. In this review, we highlight the recent progress in high-sensitivity bioanalytical applications based on aptamer signal amplification strategies on microchips. Furthermore, the main challenges in the practical application are discussed, and the development in the future is prospected.

A smartphone-assisted microarray immunosensor coupled with GO-based multi-stage signal amplification strategy for high-sensitivity detection of okadaic acid.

Okadaic acid (OA) is one of the main virulence factors of diarrheal shellfish toxins (DSP), which can cause acute carcinogenic or teratogenic effects after ingestion of contaminated shellfish. Therefore, high-sensitivity and fast detection of OA is a key to preventing the occurrence of safety accidents. In this paper, we effectively established a smartphone-assisted microarray immunosensor combined with an indirect competitive ELISA (iELISA) for quantitative colorimetric detection of OA. To further improve the detection sensitivity and match the smartphone imaging, a novel graphene oxide (GO) composite probe was developed to realize the multi-stage signal amplification. The system exhibited a wide linear range for the detection of OA (0.02-33.6 ng ·mL-1) with low detection limit of 0.02 ng ·mL-1. The recovery of OA in spiked shellfish samples was in the range of 80%-103.5%, which indicates the good applicability of this biosensor. The whole detection system has advantages of simplicity, low cost, high sensitivity and portability, which is expected to be a powerful alternative tool for on-site detecting and early warning of the pollution of marine products.

Target-induced gold nanoparticles colorimetric sensing coupled with aptamer for rapid and high-sensitivity detecting kanamycin.

Herein we report a novel colorimetric sensing strategy for the detection of kanamycin (kana) based on target-induced gold nanoparticles (AuNPs) coupled with aptamers. Aptamer-functionalized AuNPs, as the colorimetric probe, showed a distinct red shift with addition of kana, which avoiding the tedious and unnecessary additive-induced process. To study the interaction between kana and AuNPs and the effects of the specific aptamer adsorption, a series of experiments including UV-vis absorbance and surface enhanced Raman spectroscopy (SERS) were performed. Based on the results, a new alternative view is proposed that kana can directly induce the aggregation of aptamer-wrapped AuNPs, attributed to the co-adsorption of kana and aptamer on the surface of AuNPs. The proposed colorimetric sensing exhibited high selectivity and sensitivity for kanamycin assay with a wide linear range from 10.0 nM to 4.0 µM, and the limit of detection (LOD) reached 4.0 nM. Moreover, the whole detection process could be completed within 5 min, and it also achieved excellent performance in real samples detection with recoveries in the range of 86.22-109.89%. The results indicate that target-induced AuNPs colorimetric sensing coupled with aptamers for the direct detection of kana is simple, rapid and high-sensitivity, has the promising potential applications in the fields of food safety and environmental monitoring.

Comparative Immunoreactivity Analyses of Hantaan Virus Glycoprotein-Derived MHC-I Epitopes in Vaccination.

MHC-I antigen processes and presentation trigger host-specific anti-viral cellular responses during infection, in which epitope-recognizing cytotoxic T lymphocytes eliminate infected cells and contribute to viral clearance through a cytolytic killing effect. In this study, Hantaan virus (HTNV) GP-derived 9-mer dominant epitopes were obtained with high affinity to major HLA-I and H-2 superfamilies. Further immunogenicity and conservation analyses selected 11 promising candidates, and molecule docking (MD) was then simulated with the corresponding MHC-I alleles. Two-way hierarchical clustering revealed the interactions between GP peptides and MHC-I haplotypes. Briefly, epitope hotspots sharing good affinity to a wide spectrum of MHC-I molecules highlighted the biomedical practice for vaccination, and haplotype clusters represented the similarities among individuals during T-cell response establishment. Cross-validation proved the patterns observed through both MD simulation and public data integration. Lastly, 148 HTNV variants yielded six types of major amino acid residue replacements involving four in nine hotspots, which minimally influenced the general potential of MHC-I superfamily presentation. Altogether, our work comprehensively evaluates the pan-MHC-I immunoreactivity of HTNV GP through a state-of-the-art workflow in light of comparative immunology, acknowledges present discoveries, and offers guidance for ongoing HTNV vaccine pursuit.