Physicochemical and structural characterization, epitope mapping and vaccine potential investigation of a new protein containing Tetratrico Peptide Repeats of Acinetobacter baumannii: An in-silico and in-vivo approach.

Acinetobacter baumannii is an opportunistic multidrug-resistant pathogen that causes a significant mortality rate. The proteins containing Tetratrico Peptide Repeats (TPRs) are involved in the pathogenicity and virulence of bacteria and have different roles such as transfer of bacterial virulence factors to host cells, binding to the host cells and inhibition of phagolysosomal maturation. So, in this study, physicochemical properties of a new protein containing TPRs in A. baumannii which was named PcTPRs1 by this study were characterized and its 3D structure was predicted by in-silico tools. The protein B and T cell epitopes were mapped and its vaccine potential was in-silico and in-vivo investigated. Domain analysis indicated that the protein contains the Flp pilus assembly protein TadD domain which has three TPRs. The helix is dominant in the protein structure, and this protein is an outer membrane antigen which, is extremely conserved among A. baumannii strains; thus, has good properties to be applied as a recombinant vaccine. The best-predicted and refined model was applied in ligand-binding sites and conformational epitopes prediction. Based on epitope mapping results, several epitopes were characterized which could stimulate both immune systems. BLAST results showed the introduced epitopes are completely conserved among A. baumannii strains. The in-vivo analysis indicates that a 101 amino acid fragment of the protein which contains the best selected epitope, can produce a good protectivity against A. baumannii as well as the whole TPR protein and thus could be investigated as an effective subunit and potential vaccines.

Protective Efficacy of the OprF/OprI/PcrV Recombinant Chimeric Protein Against Pseudomonas aeruginosa in the Burned BALB/c Mouse Model.

BACKGROUND: Pseudomonas aeruginosa infection is the major cause of death in burn patients. Thus, in this study, a chimeric vaccine harboring the OprF185-350-OprI22-83-PcrV was designed and expressed in Escherichia coli. The immunogenicity of the recombinant chimer, OprI, OprF, and PcrV was studied in a burned mouse model. METHODOLOGY: Recombinant proteins including the proposed chimer, OprF, OprI, and PcrV were expressed in the E.coli. Mice were immunized with the purified recombinant proteins, and the antibody titre was estimated in the sera obtained from immunized mice. Immunized and control mice were challenged with 2, 5, and 10xLD50 of the P. aeruginosa strains (PAO1, PAK, and R5), and microbial counts were measured in the skin, liver, spleen, and kidney of the studied mice. RESULTS: Results showed that the antibody titre (total IgG) was significantly increased by injection of 10 µg of chimeric protein in the experimental groups compared to the control groups. The antibody survival titre was high until 235 days after administration of the second booster. The survival rate of the mice infected with 10xLD50 was significantly increased and the number of bacteria was reduced, especially in the internal organs (kidney, spleen, and liver) compared to the mice immunized with any of the OprF, OprI, and PcrV proteins alone. CONCLUSION: The findings of our study revealed that the chimeric protein is a promising vaccine candidate for control of the P. aeruginosa infection.

In-Silico Analysis and Protective Efficacy of the PcrV Recombinant Vaccine against Pseudomonas Aeruginosa in the Burned and PA-Infected BALB/c Mouse Model.

BACKGROUND: Pseudomonas aeruginosa is considered as the most severe cause of infections in burn patients and pneumonia infections. OBJECTIVE: To study the protective effects of recombinant protein vaccine harboring the PcrV of P. aeruginosa in the mouse model of burn and respiratory infections. METHODS: Recombinant protein vaccine harboring the PcrV was expressed in the E. coli BL-21 strain. Mice were immunized with the purified recombinant protein, and the antibody titer was measured in the sera obtained from the immunized mice. Immunized and control mice were challenged by active and passive immunization. The microbial counts in the lung, skin, liver, spleen, and kidney were compared with the control mice. RESULTS: Bioinformatics analysis indicated that the PcrV protein was conserved in 1552 clinical and environmental isolates. Also, the isoelectric point (pI), molecular weight, and Grand Average of Hydropathy (GRAVY) score were analyzed. Mice were injected with recombinant protein, and serum from immunized mice reacted strongly with recombinant antigen at a dilution of 1:64000. The survival rate of mice infected with 5xLD50 of the P. aeruginosa increased significantly up to 75% in the standard strains (PAO1 and PAK), and the number of bacteria, especially in the internal organs (kidney, spleen, and liver) significantly reduced compared to the mice immunized with placebo. CONCLUSION: Our results demonstrated that the PcrV protein could be an effective candidate vaccine for the generation of antibody response against P. aeruginosa infection.

Chemical Composition and Antifungal Activity of Cuminum cyminum L. Essential Oil From Alborz Mountain Against Aspergillus species.

BACKGROUND: Aflatoxin B1 (AFB1) is a highly toxic and hepatocarcinogenic metabolite produced by Aspergillus species. Some natural products are known to kill fungi and destroy toxins and toxin-producing agents. OBJECTIVES: The purpose of this study is to provide experimental data on the antifungal activity of cumin oils and their components that could be considered suitable for application in foods and drugs. MATERIALS AND METHODS: The essential oil (EO) of Cuminum cyminum L. collected from Alborz Mountain, Iran, was obtained by hydro-distillation. The oil was analyzed by gas chromatography (GC) and chromatography/mass spectrophotometry (GC/MS). The antifungal activity of the oil was studied with regard to the inhibition of the growth of Aspergillus flavus PICC-AF39 , Aspergillus flavus PICC-AF24, Aspergillus parasiticus NRRL-2999 and Aspergillus niger. The minimal inhibitory (MIC) and minimal fungicidal (MFC) concentrations of the oil were determined. RESULTS: α-Pinene (29.2%), limonene (21.7%), 1,8-cineole (18.1%), linalool (10.5%), linalyl acetate (4.8%), and α-terpineole (3.17%) were the major components of the essential oil from C. cyminum L., and the oil showed a strong inhibitory effect on fungal growth. CONCLUSIONS: Essential oils could be safely used as preservatives in pharmaceuticals as well as health and food products to protect them against toxigenic fungal infections.

Antimycotoxigenic characteristics of Rosmarinus officinalis and Trachyspermum copticum L. essential oils.

Aflatoxin B1 (AFB1) is a highly toxic and carcinogenic metabolite produced by Aspergillus species on food and agricultural commodities. Natural products may regulate the cellular effects of aflatoxins and evidence suggests that aromatic organic compounds of spices can control the production of aflatoxins. With a view to controlling aflatoxin production, the essential oils from Rosmarinus officinalis and Trachyspermum copticum L. were obtained by hydrodistillation. Antifungal activities of the oils were studied with special reference to the inhibition of Aspergillus parasiticus growth and aflatoxin production. Minimal inhibitory (MIC) and minimal fungicidal (MFC) concentrations of the oils were determined. T. copticum L. oil showed a stronger inhibitory effect than R. officinalis on the growth of A. parasiticus. Aflatoxin production was inhibited at 450 ppm of both oils with that of R. officinalis being stronger inhibitor. The oils were analyzed by GC and GC/MS. The major components of R. officinalis and T. copticum L. oils were Piperitone (23.65%), alpha-pinene (14.94%), Limonene (14.89%), 1,8-Cineole (7.43%) and Thymol (37.2%), P-Cymene (32.3%), gamma-Terpinene (27.3%) respectively. It is concluded that the essential oils could be safely used as preservative materials on some kinds of foods to protect them from toxigenic fungal infections.