Chimeric Hepatitis B core virus-like particles harboring SARS-CoV2 epitope elicit a humoral immune response in mice.

BACKGROUND: Virus-like particles are an interesting vector platform for vaccine development. Particularly, Hepatitis B virus core antigen has been used as a promising VLP platform. It is highly expressed in different recombinant expression systems, such as E. coli, and self-assembled in vitro. It effectively improves the immunogenicity of foreign antigenic epitopes on its surface. Various foreign antigens from bacteria, viruses, and protozoa can be genetically inserted into such nanoparticles. The effective immunogenicity due to VLP vaccines has been reported. However, no research has been performed on the SARS-CoV2 vaccine within this unique platform through genetic engineering. Considering the high yield of target proteins, low cost of production, and feasibility of scaling up, E. coli is an outstanding expression platform to develop such vaccines. Therefore, in this investigation, we planned to study and develop a unique HBc VLP-based vaccine against SARS-Cov2 utilizing the E. coli expression system due to its importance. RESULTS: Insertion of the selected epitope was done into the major immunodominant region (MIR) of truncated (149 residues) hepatitis B core capsid protein. The chimeric protein was constructed in PET28a+ and expressed through the bacterial E. coli BL21 expression system. However, the protein was expressed in inclusion body forms and extracted following urea denaturation from the insoluble phase. Following the extraction, the vaccine protein was purified using Ni2 + iminodiacetic acid (IDA) affinity chromatography. SDS-PAGE and western blotting were used to confirm the protein expression. Regarding the denaturation step, the unavoidable refolding process was carried out, so that the chimeric VLP reassembled in native conformation. Based on the transmission electron microscopy (TEM) analysis, the HBC VLP was successfully assembled. Confirming the assembled chimeric VLP, we explored the immunogenic effectivity of the vaccine through mice immunization with two-dose vaccination with and without adjuvant. The utilization of adjuvant was suggested to assess the effect of adjuvant on improving the immune elicitation of chimeric VLP-based vaccine. Immunization analysis based on anti-spike specific IgG antibody showed a significant increase in antibody production in harvested serum from immunized mice with HBc-VLP harboring antigenic epitope compared to HBc-VLP- and PBS-injected mice. CONCLUSIONS: The results approved the successful production and the effectiveness of the vaccine in terms of humoral IgG antibody production. Therefore, this platform can be considered a promising strategy for developing safe and reasonable vaccines; however, more complementary immunological evaluations are needed.

Antifungal activity of protective cultures against the yogurt drink-specific spoilage yeasts.

Fungal agents emerged as post-pasteurization contamination are responsible for the spoilage in yogurt drink. In this work, the antifungal effects of some lactic acid bacteria (LAB) on the spoilage yeasts isolated from yogurt drink (Doogh) were evaluated. First, the microbial growth in the yogurt drink samples during the storage time was investigated, and the isolated microorganisms were identified using biochemical methods and sequencing of the specific amplicons. Yeasts (3-7 log CFU ml-1 ) were found to be the most abundant microorganisms (specific spoilage organisms) in several samples. Using the amplification technique of rDNA by ITS1 and ITS4 primers, the dominant yeasts were identified as Pichia kudriavzevii, Kluyveromyces marxianus, and Candida parapsilosis. Then, the antimicrobial activity of 37 strains of LAB against the isolated yeasts was studied using broth microdilution. Eventually, the strains of Lacticplantibacillus plantarum (245, 24, P6, and P7), Lactiplantibacillus pentosus (20), and Levilactobacillus brevis (30) exhibited significant antifungal activity. In the most effective impacts, lag times of C. parapsilosis, K. marxianus, and P. kudriavzevii were increased by almost 12-19 h, 12-19 h, and 2-6 h, respectively, while the area under the growth curve for these yeasts was reduced to lower than 40%, near 16%, and approximately 67%, in the order given. Overall, these bacteria showed high potential as the substituents for chemical preservatives in yogurt drinks. PRACTICAL APPLICATION: Spoilage yeasts were isolated from yogurt drink and identified by molecular method. Isolated yeasts belonged to Pichia, Kluyveromyces, and Candida genera. Inhibitory effects of 37 strains were evaluated against the spoilage yeasts. Cell-free supernatant was used against the isolated fungi in microdilution method. Several LAB strains showed a significant antimicrobial activity.

Mining transcriptomic data to identify Saccharomyces cerevisiae signatures related to improved and repressed ethanol production under fermentation.

Saccharomyces cerevisiae is known for its outstanding ability to produce ethanol in industry. Underlying the dynamics of gene expression in S. cerevisiae in response to fermentation could provide informative results, required for the establishment of any ethanol production improvement program. Thus, representing a new approach, this study was conducted to identify the discriminative genes between improved and repressed ethanol production as well as clarifying the molecular responses to this process through mining the transcriptomic data. The significant differential expression probe sets were extracted from available microarray datasets related to yeast fermentation performance. To identify the most effective probe sets contributing to discriminate ethanol content, 11 machine learning algorithms from RapidMiner were employed. Further analysis including pathway enrichment and regulatory analysis were performed on discriminative probe sets. Besides, the decision tree models were constructed, the performance of each model was evaluated and the roots were identified. Based on the results, 171 probe sets were identified by at least 5 attribute weighting algorithms (AWAs) and 17 roots were recognized with 100% performance Some of the top ranked presets were found to be involved in carbohydrate metabolism, oxidative phosphorylation, and ethanol fermentation. Principal component analysis (PCA) and heatmap clustering validated the top-ranked selective probe sets. In addition, the top-ranked genes were validated based on GSE78759 and GSE5185 dataset. From all discriminative probe sets, OLI1 and CYC3 were identified as the roots with the best performance, demonstrated by the most weighting algorithms and linked to top two significant enriched pathways including porphyrin biosynthesis and oxidative phosphorylation. ADH5 and PDA1 were also recognized as differential top-ranked genes that contribute to ethanol production. According to the regulatory clustering analysis, Tup1 has a significant effect on the top-ranked target genes CYC3 and ADH5 genes. This study provides a basic understanding of the S. cerevisiae cell molecular mechanism and responses to two different medium conditions (Mg2+ and Cu2+) during the fermentation process.

Integrating expression data and genomic sequences to investigate transcriptional regulation in barley in response to abiotic stress.

Abiotic stress responses are regulated critically at the transcriptional level. Clarifying the intricate mechanisms that regulate gene expression in response to abiotic stress is crucial and challenging. For this purpose, the factors that regulate gene expression and their binding sites in DNA should be determined. By using bioinformatics tools, the differentially expressed probe sets were studied. A meta-analysis of transcriptomic responses to several abiotic stresses in barley was performed. Motif enrichments revealed that AP2/ERF (APETALA2/Ethylene-Responsive Factor) has the most frequent binding sites. We found that the bHLH transcription factor family has the highest number of transcription factor members. Moreover, network construction revealed that AP2 has the highest number of connections with other genes, which indicates its critical role in abiotic stress responses. The present research further predicted 49 miRNAs belonging to 23 miRNA families. This study identified the probable conserved and enriched motifs, which might have a role in the regulation of differentially expressed genes under abiotic stresses. In addition to shedding light on gene expression regulation, a toolbox of available promoters for genetic engineering of crop plants under such abiotic stresses was developed.

A study on the regulatory network with promoter analysis for Arabidopsis DREB-genes.

Dehydration response element binding factors (DREBs) are one of the principal plant transcription factor subfamilies that regulate the expression of many abiotic stress-inducible genes. This sub-family belongs to AP2 transcription factor family and plays a considerable role in improving abiotic stresses tolerance in plants. Therefore, it is of interest to identify critical cis-acting elements involved in abiotic stress responses. In this study, we survey promoter cis-elements for ATDREBs genes (Arabidopsis thaliana DREBs). Regulatory networks based on ATDREB candidate genes were also generated to find other genes that are functionally similar to DREBs. The study was conducted on all 20 Arabidopsis thaliana non redundant DREB genes stored in RefSeq database. Promoter analysis and regulatory network prediction was accomplished by use of Plant CARE program and GeneMANIA web tool, respectively. The results indicated that among all genes, DREB1A, DREB1C, DREB2C, DREB2G and DEAR3 have the most type of diverse motifs involved in abiotic stress responses. It is implied that co-operation of abscisic acid, ethylene, salicylic acid and methyl jasmonate signaling is crucial for the regulation of the expression of drought and cold responses through DREB transcription factors. Gene network analysis showed different co-expressed but functionally similar genes that had physical and functional interactions with candidate DREB genes.