Single-Dose Intranasal Immunisation with Novel Chimeric H1N1 Expressing the Receptor-Binding Domain of SARS-CoV-2 Induces Robust Mucosal Immunity, Tissue-Resident Memory T Cells, and Heterologous Protection in Mice.

Current COVID-19 vaccines can effectively reduce disease severity and hospitalisation; however, they are not considerably effective in preventing infection and transmission. In this context, mucosal vaccines are pertinent to prevent SARS-CoV-2 infection and spread. In this study, we generated a replication-competent recombinant chimeric influenza A virus (IAV) expressing the receptor-binding domain (RBD) of a SARS-CoV-2 prototype in the C-terminus of the neuraminidase (NA) of A/Puerto Rico/08/1934 H1N1 (PR8). The remaining seven segments from A/WSN/1933 H1N1 (WSN) were named PR8NARBD/WSN. We observed that the recombinant virus with the WSN backbone demonstrated improved expression of NA and RBD. A single intranasal dose of PR8NARBD/WSN(103PFU) in mice generated robust mucosal immunity, neutralising antibodies, cellular immunity, and tissue-resident memory T cells specific to SARS-CoV-2 and IAV. Importantly, immunisation with PR8NARBD/WSN viruses effectively protected mice against lethal challenges with H1N1, H3N2 IAV, and SARS-CoV-2 Beta variant and significantly reduced lung viral loads. Overall, our research demonstrates the promising potential of PR8NARBD/WSN as an attractive vaccine against emerging SARS-CoV-2 variants and influenza A virus infections.

Synergistic Immunity and Protection in Mice by Co-Immunization with DNA Vaccines Encoding the Spike Protein and Other Structural Proteins of SARS-CoV-2.

The emergence of new variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has generated recurring worldwide infection outbreaks. These highly mutated variants reduce the effectiveness of current coronavirus disease 2019 (COVID-19) vaccines, which are designed to target only the spike (S) protein of the original virus. Except for the S of SARS-CoV-2, the immunoprotective potential of other structural proteins (nucleocapsid, N; envelope, E; membrane, M) as vaccine target antigens is still unclear and worthy of investigation. In this study, synthetic DNA vaccines encoding four SARS-CoV-2 structural proteins (pS, pN, pE, and pM) were developed, and mice were immunized with three doses via intramuscular injection and electroporation. Notably, co-immunization with two DNA vaccines that expressed the S and N proteins induced higher neutralizing antibodies and was more effective in reducing the SARS-CoV-2 viral load than the S protein alone in mice. In addition, pS co-immunization with either pN or pE + pM induced a higher S protein-specific cellular immunity after three immunizations and caused milder histopathological changes than pS alone post-challenge. The role of the conserved structural proteins of SARS-CoV-2, including the N/E/M proteins, should be investigated further for their applications in vaccine design, such as mRNA vaccines.

Ant nest-like hierarchical porous imprinted resin-dispersive solid-phase extraction for selective extraction and determination of polychlorinated biphenyls in milk.

Polychlorinated biphenyls (PCBs) are persistent toxic, organic chemicals that tend to accumulate in the food chain. This study reports the rapid and selective extraction and determination of PCBs (PCB81, 153, 105, 126, and 157) in milk samples by a dispersive solid-phase extraction (DSPE) coupled with gas chromatography-tandem mass spectrometry (GC-MS/MS). An ionic liquid-molecularly imprinted porous resin (IL-MIPPR) as a DSPE adsorbent was synthesized from m-aminophenol, formaldehyde, and 2,2'-benzidinedisulfonic acid as the monomer, crosslinker, and virtual template, respectively. The IL-MIPPR had a fast mass transfer (1.0 min) and good selectivity (imprinting factors of 1.8-3.0). The IL-MIPPR - DSPE - GC-MS/MS method exhibited good linearity (R2 ≥ 0.9995), the limit of detections (LODs) < 0.6 pg/g, and the recoveries ranged from 82.8 % to 106 % with relative standard deviations ≤ 6.6 %. This method is thus better than previously reported methods in terms of the LOD, the adsorbent dosage, and the extraction time.

Green synthesis of phloroglucinol-urotropine porous polymer: Ingenious miniaturized solid phase extraction for efficient purification and determination of polycyclic aromatic hydrocarbons in lotus roots.

Polycyclic aromatic hydrocarbons (PAHs) posed a serious threat to food safety and human health due to long-term emission. In this work, a new method was established using phloroglucinol-urotropine porous polymer (PU-PP) in a pipette tip for solid phase extraction (PT-SPE) for the first time and used prior to determination of four PAHs (phenanthrene, anthracene, fluoranthene, and pyrene) in lotus roots. Synthesis of the PU-PP adsorbent was green compared with alternatives; urotropine was used as a cross-linker and ethanol-water as the solvent. PU-PP-based PT-SPE had the advantages of low solvent consumption, good purification, practicability, stability, and low-cost. The proposed pre-purification method offered low limits of detection (0.09-0.28 ng/g) and good recoveries (84.6-114.3 %, RSDs ≤ 5.6 %) for determination of the four PAHs, which were detected at trace concentrations in samples. This new method provides an alternative for monitoring trace pollutants in aquatic plant ingredients.

DNA Vaccines Expressing the Envelope and Membrane Proteins Provide Partial Protection Against SARS-CoV-2 in Mice.

The coronavirus disease 2019 (COVID-19) pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become a public health emergency of international concern, and an effective vaccine is urgently needed to control the pandemic. Envelope (E) and membrane (M) proteins are highly conserved structural proteins among SARS-CoV-2 and SARS-CoV and have been proposed as potential targets for the development of cross-protective vaccines. Here, synthetic DNA vaccines encoding SARS-CoV-2 E/M proteins (called p-SARS-CoV-2-E/M) were developed, and mice were immunised with three doses via intramuscular injection and electroporation. Significant cellular immune responses were elicited, whereas no robust humoral immunity was detected. In addition, novel H-2d-restricted T-cell epitopes were identified. Notably, although no drop in lung tissue virus titre was detected in DNA-vaccinated mice post-challenge with SARS-CoV-2, immunisation with either p-SARS-CoV-2-E or p-SARS-CoV-2-M provided minor protection and co-immunisation with p-SARS-CoV-2-E+M increased protection. Therefore, E/M proteins should be considered as vaccine candidates as they may be valuable in the optimisation of vaccination strategies against COVID-19.

Bioinformatic Prediction and Production of Four Recombinant Proteins from Different Developmental Stages of Trichinella spiralis and Testing of Their Diagnostic Sensitivity in Mice.

BACKGROUND: Trichinellosis is a serious food-borne parasitic zoonosis, thus finding high quality antigens is the key to serodiagnosis of trichinosis. This article reports the characterization and sensitivity of four recombinant proteins expressed by four genes (Wn10, Zh68, T668, and Wm5) from different developmental stages of Trichinella spiralis for the diagnosis of trichinellosis in mice. METHODS: This study was conducted in Jilin University and National Institute of Parasitic Diseases of Chinese Center for Disease Control and Prevention in 2017-2018. The structures and functions of the proteins encoded by four genes were predicted by bioinformatics analysis. The four genes were cloned and expressed, and the recombinant proteins were purified. Anti-Trichinella IgM and IgG antibodies in the sera of mice infected with T. spiralis from 1-45 d post-infection (dpi) were evaluated by ELISA. RESULTS: The optimal antigen epitopes of four proteins (P1, P2, P3, and P4) encoded by the four genes from T- and B-cells were predicted, and four purified recombinant proteins (r-P1, r-P2, r-P3, and r-P4) were successfully produced. For IgM, the antibody levels detected by the four recombinant antigens were approximately equal to the cut-off value. Anti-Trichinella IgG antibodies were first detected by r-P1 at 8 dpi, followed by r-P2, r-P3, and r-P4 at 10 dpi, 14 dpi, and 16 dpi, respectively, and the antibody levels remained high until 45 dpi. CONCLUSION: The recombinant antigens r-P1, r-P2, r-P3, and r-P4 could be antigens that react with antibodies, they showed high sensitivity in the detection of anti-Trichinella IgG antibodies in mice. Among these proteins, r-P1 may be a candidate antigen for the detection of anti-Trichinella IgG antibodies in the early infection phase and exhibited the best sensitivity among the antigens.

Kinetics Evaluation of IgM and IgG Levels in the Mice Infected with Trichinella spiralis Experimentally Using ES Antigens from Different Developmental Stages of the Parasite.

BACKGROUND: To assay the Trichinella-specific IgM and IgG antibody responses during the early stage of infection, serum was collected from mice infected with the muscle larvae (ML) of T. spiralis (ISS534) at different dpi (days post infection) up to 60 days. METHODS: The levels of IgM and IgG antibodies in serum were measured by ES antigens from different stage of T. spiralis using the ELISA method in Shanghai, China in 2017. RESULTS: The anti-Trichinella IgM and IgG could be detected by ES antigens from the adult three days worm (Ad3) as early as 5 dpi and 8 dpi, respectively. ES antigens from the mixture of adult six days worm & new born larvae (Ad6+NBL) was similar to Ad3. When antibodies were detected by these two antigens, the levels of IgM peaked at 14 dpi and then declined from 15 dpi to 60 dpi; the IgG peaked at 20 dpi, and gradually declined, however, higher detection levels were maintained until 60 dpi. CONCLUSION: Ad3 ES antigens showed more antigenicity than Ad6+NBL ES on titer detection of IgM and IgG antibodies, and the production of Ad3 ES is easier. In terms of early diagnosis, these two antigens are better than the ML ES antigens of T. spiralis, which antibodies could not be detected before 20dpi. Ad3 ES antigens might be good candidate for the early diagnosis of trichinellosis or the mixture of Ad3 and Ad6+NBL ES might be used.

Graphene/multi-walled carbon nanotubes as an adsorbent for pipette-tip solid-phase extraction for the determination of 17ß-estradiol in milk products.

The illegal addition and abuse of 17ß-estradiol (E2) in food has led to its accumulation in the body through the food chain, resulting in adverse effects on the human body through interference with the endocrine system. Because milk and dairy products are major components of the human diet, screening as well as quantitative analysis of E2 in these food types is important. In this work, a three-dimensional graphene/multi-walled carbon nanotube composite was synthesized and applied to a home-made pipette-tip solid-phase extraction cartridge and coupled with high-performance liquid chromatography-fluorescence detection to determine E2 in milk. The established method showed good linearity in the range of 5-250 ng mL-1 with correlation coefficient of 0.9998. The detection limit and quantitation limit were 0.7 and 2.3 ng mL-1, respectively. The repeatability and intermediate precision, expressed as the coefficient of variation were 3.7% and 6.0%, respectively. The accuracy of the method was investigated by recovery experiments with milk at three spiked levels (25, 100, and 250 ng mL-1), where the recoveries were in the 88.4-108.9% range with relative standard deviations ≤5.5%. The proposed method has the advantage of requiring less adsorbent (only 1.0 mg) and less organic reagent (1.0 mL of washing solvent, 1.2 mL of elution solvent) and exhibits excellent purification ability, with high sensitivity and accuracy, and was successfully applied for the determination of E2 in milk products.

High-resolution chalcogenide fiber bundles for longwave infrared imaging.

A flexible chalcogenide fiber bundle (FB) with a resolution as high as ~31 lp/mm has been fabricated for delivering thermal images of objects at room temperature. The FB is composed of ~200,000 single fibers with a Ge-As-Te-Se glass core 15 µm in diameter and a polyetherimide (PEI) cladding 16.8 µm in diameter. These Ge-As-Te-Se/PEI fibers show good transparency in the 3-12 µm spectral region. The fabricated FB presents a filling factor of ~72% and a crosstalk of ~1%. High-quality thermal images of a human hand were obtained through the FB, demonstrating good potential of the FB for longwave infrared imaging in the areas such as medicine, industry and defense.

High-resolution chalcogenide fiber bundles for infrared imaging.

An ordered chalcogenide fiber bundle with a high resolution for infrared imaging was fabricated using a stack-and-draw approach. The fiber bundle consisted of about 810,000 single fibers with an As2S3 glass core of 9 µm in diameter and a polyetherimide (PEI) polymer cladding of 10 µm in diameter. The As2S3/PEI fibers showed good transparency in the 1.5-6.5 µm spectral region. It presented a resolution of ∼45 lp/mm and a crosstalk of ∼2.5%. Fine thermal images of a hot soldering iron tip were delivered through the fiber bundle.

1.8-10 µm mid-infrared supercontinuum generated in a step-index chalcogenide fiber using low peak pump power.

By pumping an 11-cm-long step-index chalcogenide fiber with ∼330 fs pulses at 4.0 µm from an optical parametric amplifier, mid-infrared supercontinuum generation spanning from ∼1.8 to ∼10 µm within a dynamic range of ±15 dB has been demonstrated at a relatively low power threshold of ∼3000 W.