Alginate di-aldehyde-modified metal-organic framework nanocarriers as delivery platform and adjuvant in inactivated pseudorabies vaccination.

Pseudorabies virus (PRV) is a highly contagious viral disease, which leads to severe financial losses in the breeding industry worldwide. Presently, PRV is mainly controlled using live attenuated and inactivated vaccines. However, these vaccines have an innate tendency to lose their structural conformation upon exposure to environmental and chemical stressors and cannot provide full protection against the emerging prevalent PRV variants. In this work, first, we synthesized aminated ZIF-7/8 nanoparticles (NPs), and then chemical bond-coated alginate dialdehyde (ADA, a type of dioxide alginate saccharide) on their surface via Schiff base reaction to obtain ZIF-7/8-ADA NPs. The as-fabricated ZIF-7/8-ADA NPs exhibited high stability, monodispersity and a high loading ratio of antigen. Furthermore, the ZIF-7/8-ADA NPs showed good biocompatibility in vitro and in vivo. Using ZIF-7/8-ADA NPs as an adjuvant and inactivated PRV as a model antigen, we constructed a PR vaccine through a simple mixture. The immunity studies indicated that ZIF-7/8-ADA induced an enhancement in the Th1/Th2 immune response, which was superior to that of the commercial ISA201, alum adjuvant and ZIF-7/8. Due to the pH-sensitive release of the antigen in lysosomes, the as-prepared PR vaccine subsequently accelerated the antigen presentation and improved the immune responses in vitro and in vivo. The results of PRV challenge using mice as the model demonstrated that ZIF-7/8-ADA achieved the same preventive effect as the commercial ISA201 and was much better than the alum adjuvant, and thus can serve as a promising delivery system and adjuvant to enhance humoral and cellular responses against PRV infection.

Benzophenoxazine-based colorimetric and fluorescent probe for highly sensitive detection of amines and food freshness.

In this work, we reported a chromogenic and near infrared (NIR) region fluorogenic dual-channel probe NRB, which could visually detect gaseous amines with high sensitivity (eg. 50 and 17 ppt for methylamine (MeNH2) via naked eyes and fluorescence spectrometer respectively). It exhibited a wide fluorescent emission band extending to the NIR region with a peak at 615 nm when stimulated by the MeNH2 solution. The plausible sensing mechanism was proved by mass spectrometry, where the reaction process was based on a nucleophilic substitution between the probe and amines rather than the ester group hydrolysis. Furthermore, NRB was successfully applied to monitor the food freshness (seafood and meat food), because of its low cytotoxicity and excellent photophysical properties. It was worth mentioning that real time monitoring for food quality can be realized visually by using a 365 nm UV lamp. In addition, the probe was stable during the quality guarantee period for perishable packaged food. It was believed that the applied experiments have demonstrated the value of this probe in the practical applications for food safety.

Formulation and Evaluation of Lipidized Imiquimod as an Effective Adjuvant.

Adjuvants are essential for the induction of robust immune responses against vaccine antigens. Small-molecule TLR7 agonists hold high potential for this purpose. In this communication, imiquimod (IMQ) bearing a cholesterol lipid moiety derivative, IMQ-Chol, was designed and synthesized as a vaccine adjuvant, which could release parent IMQ molecules in aqueous conditions via amide bond hydrolysis. We performed a series of immunological evaluations by cooperating with the inactivated foot-and-mouth disease virus (FMDV). All of the results confirmed that IMQ-Chol could stimulate the body for a prolonged time to produce strong humoral and cellular immunity with a balanced Th1/Th2 immune response through a TLR7-related MAPK pathway. In addition, the results of the proof-of-concept vaccine indicated IMQ-Chol had a good effect on preventing and treating FMD in pigs.

A benzophenoxazine-based NIR fluorescent probe for the detection of hydrogen sulfide and imaging in living cells.

Hydrogen sulfide (H2S) plays a vital role in regulating many important physiological functions. However, H2S-containing industrial wastewater is inevitably pumped into the environment which seriously contaminates water supplies and foodstuffs. So it is crucial to develop a single fluorescent probe for H2S detection with high sensitivity and selectivity. Herein, a colorimetric and turn-on near infrared (NIR) fluorescent probe NRDNP based on benzophenoxazine was designed and synthesized by thiolysis of 2,4-dinitrophenyl (DNP) ether as the specific reaction site strategy to achieve highly specific H2S detection in living systems. The studies demonstrated that the probe NRDNP exhibited excellent sensing performance toward H2S with an about 80-fold NIR fluorescence enhancement, a rapid response within 10 min, excellent sensitivity with a detection limit of 19 nM and good selectivity. Furthermore, the NRDNP is an optical sensor which visually changes in terms of the fluorescence colour/intensity upon sensing gaseous H2S molecules. NRDNP has low cytotoxicity and has been successfully applied in the fluorescence imaging of H2S in living cells, suggesting that it would be an effective tool for H2S detection in living systems.

Biodegradable Imiquimod-Loaded Mesoporous Organosilica as a Nanocarrier and Adjuvant for Enhanced and Prolonged Immunity against Foot-and-Mouth Disease Virus in Mice.

Foot-and-mouth disease (FMD), a serious, fast-spreading, and virulent disease, has led to huge economic losses to people all over the world. Vaccines are the most effective way to control FMD. However, the weak immunogenicity of inactivated FMD virus (FMDV) requires the addition of adjuvants to enhance the immune effectiveness of the vaccines. Herein, we formulated and fabricated biodegradable dendritic mesoporous tetrasulfide-doped organosilica nanoparticles SOMSN with imiquimod complex (SOMSN-IMQ) and used it as a platform for FMD vaccine delivery and as an adjuvant. SOMSN-IMQ demonstrated excellent stability for 6 months when stored in PBS, while it could be completely degraded within 42 days in SBF at room temperature. Biosafety experiments such as cell toxicity, hemolysis, and histology indicated that the as-prepared SOMSN-IMQ showed nontoxicity and good biocompatibility. Furthermore, SOMSN-IMQ exhibited a maximum adsorption capacity of 1000 µg/mg for inactivated FMDV antigens. Our results showed that SOMSN-IMQ can be effectively engulfed by RAW264.7 cells in a dose-dependent manner. After immunization, SOMSN-IMQ@FMDV can elicit persistent higher antibody levels, higher IgG2a/IgG1 ratio, and cytokine expression, which indicated that SOMSN-IMQ@FMDV triggered superior humoral and cellular immune responses. Moreover, SOMSN-IMQ could provoke maturation and activation of dendritic cells in lymph nodes (LDCs) as well as the proliferation of lymphocytes in vivo. Thus, SOMSN-IMQ could promote effective and potent immunity and provide a promising adjuvant platform for FMDV vaccination with acceptable safety.

A pyrylium salt-based fluorescent probe for the highly sensitive detection of methylamine vapour.

The amine vapour produced by microorganisms is an important indicator of food spoilage. Amines are also ubiquitous in the chemical industry. The development of molecule-based ion sensors has been a pivotal issue that is currently receiving considerable attention. In this work, a new pyrylium salt-based fluorescent probe MTPY has been developed as a rapid, highly sensitive, and selective sensor for methylamine vapour. MTPY exhibits an obvious fluorescence response from yellow to cyan towards CH3NH2 vapour. The calibration curve of titration analysis shows a linear relationship of the fluorescence intensity at 514 nm versus the methylamine concentration in the range of 0.1-2 ppm. In addition to the linearity (R2 = 0.974) and short response time with a low detection limit (2.6 ppt, 8.4 × 10-8 M), the sensing mechanism was traced using mass spectrometry.

Effect of multiplicity of infection on the evasion of neutrophil killing by Streptococcus agalactiae isolated from clinical mastitis bovine.

Streptococcus agalactiae (S. agalactiae) causes intramammary infection in dairy cows. Increased neutrophils and a high bacterial load are important characteristics of bovine bacterial mastitis. We hypothesized that the multiplicity of infection (MOI) of S. agalactiae in bovine mastitis plays an important role in bacterial pathogenicity by modulating the neutrophil response to promote bacterial survival. Neutrophils from BALB/c mice were infected with the bovine mastitis isolate of S. agalactiae SAG-FX17 at various MOIs, and neutrophil responses were investigated. Infecting neutrophils with SAG-FX17 at an MOI of 1 induced reactive oxygen species (ROS) and neutrophil extracellular traps (NETs) formation. Bacteria at an MOI of 10 suppressed neutrophil responses, including ROS bursts, NET formation, and cell necrosis, which are conducive to bacterial multiplication within 30 min postinfection. In addition, neutrophils are destroyed by SAG-FX17 at an MOI of 100 or greater. This study identified the MOIs related to the ROS and NET suppression caused by SAG-FX17, and the findings suggested that interventions to decrease bacterial loads before the MOI of 10 could be necessary and effective to harness the power of innate immune response to eliminate pathogens.

Outer inflammatory protein of Helicobacter pylori impacts IL-8 expression, adherence, cell apoptosis and cell cycle of gastric cells independent of its copy number.

Outer inflammatory protein (OipA) is an important virulence factor of Helicobacter pylori (H. pylori), but the correlation between oipA copy number and its virulence remains unknown. The study was designed to investigate whether the duplicate oipA gene loci showed more virulent than one oipA gene in vitro. H. pylori strain CCS9803 (China Chongqing Strain 9803) that carries duplicate oipA loci was used to construct one or two oipA knockout mutant strain, which was further verified by qPCR and western blot. Gastric epithelial cells AGS and GES-1 were infected with wild-type (WT) or oipA mutants for 6 or 24 h. The expression levels of IL-8, bacterial adhesion, cell apoptosis and cell cycle were performed to analyze the function of oipA. The WT and oipA mutant strains induce significantly higher mRNA and protein levels of IL-8 than the uninfected group (P < 0.05), but only oipA2 mutants induced significantly decreased expression levels than the WT-infected group (P < 0.05). Adherence to gastric cells was significantly decreased by inactivated two oipA loci (P < 0.05). The WT strain caused a significant rising proportion of early apoptosis cell, which had dropped after duplicate oipA genes were both knockout (P < 0.05). WT and oipA1 mutants failed to affect cell cycle; however, the oipA2 mutants increased M phase and reduced S phase when compared to the uninfected group. In conclusion, our study demonstrated that oipA impacts IL-8 expression, adherence, cell apoptosis and cell cycle of gastric cells independent of its gene copy number.

In-vitro characterisation of a novel antimicrobial agent, TNP-2092, against Helicobacter pylori clinical isolates.

BACKGROUND AND OBJECTIVES: TNP-2092 is a novel dual-action lead compound consisting of rifamycin SV and 4H-4-oxo-quinolizine pharmacophores, with a broad spectrum of antibacterial activities. This compound is currently in the early stage of clinical development for Helicobacter pylori infection. The aim of the present study was to determine the antibacterial activity of TNP-2092 against H. pylori isolated from primary patients. METHODS: A total of 100 H. pylori clinical isolates from primary patients were selected. The minimum inhibitory concentrations (MICs) for clarithromycin, levofloxacin, rifampin and TNP-2092 were determined using an agar dilution method. A time-kill study was performed with different concentrations of TNP-2092 relevant to MIC against H. pylori ATCC strain 43504 for up to 24 hours. The time-kill study with drug concentrations of 0-4 × MIC was also used to determine the antibacterial activity of TNP-2092 against H. pylori under different pH conditions (pH 4-7). RESULTS: The primary resistance percentages to clarithromycin, levofloxacin, rifampin and TNP-2092 were 13, 18, 1 and 1%, respectively. TNP-2092 killing kinetics were both concentration and time dependent. The effectiveness of TNP-2092 against H. pylori was gradually reduced with a decrease in pH. CONCLUSIONS: TNP-2092 is highly active against H. pylori and against strains resistant to clarithromycin or levofloxacin. Its antibacterial activity is both concentration- and time-dependent .The antibacterial activity of TNP-2092 appears to be pH-dependent and is more active under neutral pH. TNP-2092 represents a promising new therapy for the treatment of H. pylori infection in primary patients.

Sequential versus concomitant therapy for treatment of Helicobacter pylori infection: an updated systematic review and meta-analysis.

BACKGROUND: Sequential and concomitant therapies are two innovative therapies for Helicobacter pylori (H. pylori) eradication. However, the comparative efficacy and safety of these treatments are controversial. Therefore, we aimed to conduct an updated systematic review and meta-analysis of studies that compared these two treatments. METHODS: A search of PubMed, Embase, the Cochrane Library, and Web of Science was carried out. Randomized controlled trials (RCTs) that compared sequential with concomitant therapies were selected for meta-analysis. RESULTS: Twenty RCTs were included in the analysis. The eradication rate of 10-day sequential therapy was superior to that of 5-day concomitant therapy (82.09 versus 77.79%, relative risk (RR) 1.052 (95% confidence interval (CI) 1.004-1.103), P = 0.035)), similar to that of 7-day concomitant therapy (82.40 versus 86.99%, RR 0.959 (95% CI 0.874-1.053), P = 0.382), and inferior to that of 10-day concomitant therapy (78.39 versus 83.32%, RR 0.945 (95% CI 0.907-0.984, P = 0.006); the occurrence of diarrhea was higher in 10-day concomitant therapy than that in 10-day sequential therapy. Compared with the eradication rate of sequential therapy, that of concomitant therapy was higher in metronidazole-resistant strains (RR 0.912 (95% CI 0.844-0.986, P = 0.020)) and strains resistant to metronidazole and clarithromycin (RR 0.542 (95% CI 0.308-0.956, P = 0.035)). CONCLUSION: The efficacy of concomitant therapy was duration dependent, and 10-day concomitant therapy was superior to 10-day sequential therapy. Compared to sequential therapy, concomitant therapy was more efficacious for metronidazole-resistant strains and metronidazole plus clarithromycin-resistant strains. However, diarrhea was more frequent with concomitant therapy than with sequential therapy.