Update on Epidemiology and Circulating Genotypes of Rotavirus in Iranian Children With Severe Diarrhea: 1986-2015

Rotaviruses are the most common cause of severe diarrhea in children under 5 years of age worldwide with a higher prevalence in developing countries. In accordance with the World Health Organization (WHO) recommendations for the global use of rotavirus vaccines, it is important to review trends of rotavirus epidemiology, distribution and diversity of rotavirus strains in the pre-vaccine period. In Iran, the average rotavirus positivity rate is 40.04% in all patients under 5 years of age hospitalized for acute gastroenteritis (AGE). Studies have shown a substantial increase in the rotavirus detection rate over time from 1986 to 2013. Moreover, there has been continued predominance of G (G1) and P (P[8]) genotypes, although the peak prevalence of G1 appeared to decline in 2007-2011 compared to 2001-2006. The data presented in this review, which suggests a change in the pattern of rotavirus genotypes in the Iranian population, further highlights the important role of continuous monitoring of rotavirus genotypes before starting any national rotavirus vaccination program

Recombinant λ-phage nanobioparticles for tumor therapy in mice models

Lambda phages have considerable potential as gene delivery vehicles due to their genetic tractability, low cost, safety and physical characteristics in comparison to other nanocarriers and gene porters. Little is known concerning lambda phage-mediated gene transfer and expression in mammalian hosts. We therefore performed experiments to evaluate lambda-ZAP bacteriophage-mediated gene transfer and expression in vitro. For this purpose, we constructed recombinant λ-phage nanobioparticles containing a mammalian expression cassette encoding enhanced green fluorescent protein (EGFP) and E7 gene of human papillomavirus type 16 (λ-HPV-16 E7) using Lambda ZAP- CMV XR vector. Four cell lines (COS-7, CHO, TC-1 and HEK-239) were transduced with the nanobioparticles. We also characterized the therapeutic anti-tumor effects of the recombinant λ-HPV-16 E7 phage in C57BL/6 tumor mice model as a cancer vaccine. Obtained results showed that delivery and expression of these genes in fibroblastic cells (COS-7 and CHO) are more efficient than epithelial cells (TC-1 and HEK-239) using these nanobioparticles. Despite the same phage M.O.I entry, the internalizing titers of COS-7 and CHO cells were more than TC-1 and HEK-293 cells, respectively. Mice vaccinated with λ-HPV-16 E7 are able to generate potent therapeutic antitumor effects against challenge with E7- expressing tumor cell line, TC-1 compared to group treated with the wild phage. The results demonstrated that the recombinant λ-phages, due to their capabilities in transducing mammalian cells, can also be considered in design and construction of novel and safe phage-based nanomedicines

Expression of Leishmania major LmSTI1 in Yeast Pichia Pastoris

Background: Leishmania major LmSTI1 is a conserved protein among different species of leishmania, and expressed in both amastigote and promastigote forms of L. major life cycle. It has previously been expressed in bacterial systems.Materials and Methods: To express LmSTI1 in the methylotrophic yeast         Pichia pastoris (P. pastoris), the shuttle vector pPICZA containing gene lmsti1 was constructed under the control of the AOX1 promoter. The recombinant vector was electro-transformed into P. pastoris, and induced by 0.5% methanol in the buffered medium. The expression of the LmSTI1 protein was visualized in the total soluble protein of P. pastoris by 12% SDS-PAGE, and further confirmed by Western blotting with L.major-infected mouse sera and HRP-conjugated goat anti-mouse IgG as the first and secondary antibodies, respectively.Results: The expression level was 0.2% of total soluble proteins.Conclusion: It might be possible to use this formulation as a whole yeast candidate vaccine against cutaneous leishmanization

Identification of a Functional, CRM-1-Dependent Nuclear Export Signal in Hepatitis C Virus Core Protein

Hepatitis C virus (HCV) infection is a major cause of chronic liver disease worldwide. HCV core protein is involved in nucleocapsid formation, but it also interacts with multiple cytoplasmic and nuclear molecules and plays a crucial role in the development of liver disease and hepatocarcinogenesis. The core protein is found mostly in the cytoplasm during HCV infection, but also in the nucleus in patients with hepatocarcinoma and in core-transgenic mice. HCV core contains nuclear localization signals (NLS), but no nuclear export signal (NES) has yet been identified

PCR-based isolation, cloning and expression of streptokinase gene from a group C streptococcus equisimilis in E.Coli

Ph.D. - Doctoral Progra

Designing, Constructing and Immunogenic Evaluation of Polytope DNA Constructs by the Application of Hepatitis C Virus Immunodominant Epitopes in BALB/c Mouse

Objective: Polytope DNA vaccines, capable of focusing the cytotoxic T lymphocyte (CTL)response on critical epitopes, represent a promising approach in HCV immunotherapy. Nevertheless,due to controversial rules governing epitope processing and the low level expression/immunogenicity of recombinant polytope peptides, designing and primary expression/immunogenicity analysis of these vaccine types should be the first consideration prior tocostly transgenic animal studies.Materials and Methods: Four HLA-A2 and H-2d restricted CTL epitopes were selected anddesigned in three appropriate sequential tandems based on epitope and proteasomal cleavagepredictions. The related nucleotide sequences were synthesized using SOEing PCRmethod and cloned into a pcDNA3.1 vector, either alone or fused to the small hepatitis B surfaceantigen (HBsAg-S) gene. Following the preparation of polyclonal anti-sera, expression/secretion of polytopes was evaluated in Cos-7 cells by using immunofluorescence, Westernblot,dot blot, ELISA and RT-PCR techniques. The immunogenicity of the plasmids was alsoassessed through the delayed-type hypersensitivity (DTH) assay in BALB/c mice.Results: Due to in silico designs and optimizations, the polytope products of constructedplasmids were efficiently detected in vitro through common techniques and HBsAg-S-basedparticles were shown to be secreted into the culture media (up to 30%). Moreover, all plasmidswere able to efficiently induce a positive DTH response while HBsAg-S fusion constructsindicated a significant immunopotential effect towards the incorporated mouse epitopes.Conclusion: Designed polytope constructs of this study are efficiently expressed and processed.They have the required initial potency for further immunogenicity analysis in transgenicmice

The β‐domain of streptokinase affects several functionalities, including specific/proteolytic activity kinetics

Streptokinase (SK) is a plasminogen activator which converts inactive plasminogen (Pg) to active plasmin (Pm), which cleaves fibrin clots. SK secreted by groups A, C, and G Streptococcus (SKA/SKC/SKG) is composed of three domains: SKα, SKβ and SKγ. Previous domain‐swapping studies between SK1/SK2b‐cluster variants revealed that SKβ plays a major role in the activation of human Pg. Here, we carried out domain‐swapping between skcg‐SK/SK2‐cluster variants to determine the involvement of SKβ in several SK functionalities, including specific/proteolytic activity kinetics, fibrinogen‐bound Pg activation and α2‐antiplasmin resistance. Our results indicate that SKβ has a minor to determining role in these diverse functionalities for skcg‐SK and SK2b variants, which might potentially be accompanied by few critical residues acting as hot spots. Our findings enhance our understanding of the roles of SKβ and hot spots in different functional characteristics of SK clusters and may aid in the engineering of fibrin‐specific variants of SK for breaking down blood clots with potentially higher efficacy and safety

A Dual-Type L2 11-88 Peptide from HPV Types 16/18 Formulated in Montanide ISA 720 Induced Strong and Balanced Th1/Th2 Immune Responses, Associated with High Titers of Broad Spectrum Cross-Reactive Antibodies in Vaccinated Mice

E. coli-derived concatenated, multitype L2-conserved epitopes of human papillomavirus (HPV) L2 protein might represent a less expensive and pan-type vaccine alternative (compared to type-specific HPV L1 virus-like particles), if stable protein expression and strong immunogenicity features could be met. Herein, three dual-type- (DT-) HPV L2 fusion peptides comprising the three head-to-tail tandem repeats (multimers) of either HPV 16 epitope “17-36” or “69-81” or one copy (monomer) of 11-88 fused to the same residues of HPV 18 were constructed and expressed in E. coli. SDS-PAGE and Western blot analyses indicated the proper expression and stability of the E. coli-derived DT peptides. Mice immunized by formulation of the purified DT peptides and Freund’s adjuvant (CFA/IFA) raised neutralizing antibodies (NAbs; the highest for DT: 11-88 peptide) which showed proper cross-reactivity to HPV types: 18, 16, 31, and 45 and efficiently neutralized HPV 18/16 pseudoviruses in vitro. Immunization studies in mice by formulation of the DT: 11-88 × 1 peptide with various adjuvants (alum, MF59, and Montanides ISA 720 and 50) indicated that Montanide adjuvants elicited the highest cross-reactive titers of NAbs and similar levels of IgG1 and IgG2a (switching towards balanced Th1/Th2 responses). The results implied development of low-cost E. coli-derived DT: 11-88 peptide formulated in human compatible ISA 720 adjuvant as a HPV vaccine

Adenovirus vector-based vaccines as forefront approaches in fighting the battle against flaviviruses

Flaviviruses are arthropod-borne viruses (arboviruses) that have been recently considered among the significant public health problems in defined geographical regions. In this line, there have been vaccines approved for some flaviviruses including dengue virus (DENV), Japanese encephalitis virus (JEV), yellow fever virus (YFV), and tick-borne encephalitis virus (TBEV), although the efficiency of such vaccines thought to be questionable. Surprisingly, there are no effective vaccine for many other hazardous flaviviruses, including West Nile and Zika viruses. Furthermore, in spite of approved vaccines for some flaviviruses, for example DENV, alternative prophylactic vaccines seem to be still needed for the protection of a broader population, and it originates from the unsatisfying safety, and the efficacy of vaccines that have been introduced. Thus, adenovirus vector-based vaccine candidates are suggested to be effective, safe, and reliable. Interestingly, recent widespread use of adenovirus vector-based vaccines for the COVID-19 pandemic have highlighted the importance and feasibility of their widespread application. In this review, the applicability of adenovirus vector-based vaccines, as promising approaches to harness the diseases caused by Flaviviruses, is discussed