Rapid and sensitive detection of SARS-CoV-2 virus in human saliva samples using glycan based nanozyme: a clinical study.

A highly sensitive colorimetric method (glycan-based nano(e)zyme) was developed for sensitive and rapid detection of the SARS-CoV-2 virus based on N-acetyl neuraminic acid (sialic acid)-functionalized gold nanoparticles (SA-Au NZs). A number of techniques were used to characterize the prepared nanomaterials including XRD, FT-IR, UV-vis, DLS, and TEM. DLS analysis indicates an average hydrodynamic size of 34 nm, whereas TEM analysis indicates an average particle size of 15.78 nm. This observation confirms that water interacts with nanoparticle surfaces, resulting in a large hydrodynamic diameter. The peroxidase-like activity of SA-Au NZs was examined with SARS-CoV-2 and influenza viruses (influenza A (H1N1), influenza A (H3N2), and influenza B). UV-visible spectroscopy was used to monitor and record the results, as well as naked eye detection (photographs). SA-Au NZs exhibit a change in color from light red to purple when SARS-CoV-2 is present, and they exhibit a redshift in their spectrum. N-acetyl neuraminic acid interacts with SARS-CoV-2 spike glycoprotein, confirming its ability to bind glycans. As a result, SA-Au NZs can detect COVID-19 with sensitivity and specificity of over 95% and 98%, respectively. This method was approved by testing saliva samples from 533 suspected individuals at Ghaem Hospital of Mashhad, Mashhad, Iran. Sensitivity and specificity were calculated by comparing the results with the definitive results. The positive results were accompanied by a color change from bright red to purple within five minutes. Statistical analysis was performed based on variables such as age, gender, smoking, diabetes, hypertension, and lung involvement. In clinical trials, it was demonstrated that this method can be used to diagnose SARS-CoV-2 in a variety of places, such as medical centers, hospitals, airports, universities, and schools.

Immunogenic evaluation of FMD virus immuno-dominant epitopes coupled with IL-2/FcIgG in BALB/c mice.

Previous studies on vaccine development against foot-and-mouth disease (FMD) virus reported that application of the inactivated vaccines for FMD virus is not completely effective. Novel vaccinations based on immune-dominant epitopes showed they induced immune responses. In addition, for better and safer immunization, access to of efficient adjuvants against FMD virus seems to be critical. In this study, we produced epitope recombinant vaccines from the VP1 protein of the FMD virus for serotype O of Iran that conjugated with Fc Immunoglobulin (FcIgG) and Interleukin-2 (IL-2). Multiple-epitope constructs included Polytope, Polytope-IL2-FcIgG, Polytope-IL2, Polytope-FcIgG that cloned and expressed in E. coli BL21 (DE3). To evaluate whether these epitope recombinant vaccines induce immune responses, BALB/c mice were injected with the epitope recombinant vaccines and their immune responses were compared with a negative control group. The humoral and cellular immune responses were measured by ELISA. The results showed there were significant differences between the negative control group and other immunized mice with recombinant epitope proteins (p < 0.05). The results of total IgG, IgG1, IgG2a levels and secretion of IFN-γ, IL-4 and IL-10 revealed that immune responses were enhanced when the epitope recombinant vaccine of FMD virus coupled with IL-2 and FcIgG. Observations indicated that the epitope recombinant plasmid of the VP1 protein co-expressed with IL-2 and FcIgG was effective in inducing an enhanced immune response. Therefore, IL-2 and FcIgG could be recommended as a potential adjuvant for epitope recombinant vaccine of the VP1 protein from FMD virus.

EPIYA Motif Genetic Characterization from Helicobacter pylori Isolates in Distinct Geographical Regions of Iran.

Background: This study aimed to determine the current EPIYA motifs of the cagA gene in Helicobacter pylori isolates from patients with gastric disorders, and evaluate the association between these patterns and the clinical outcome of H. pylori infection in different geographical regions of Iran. Materials and Methods: We examined 150 patients with gastrointestinal disorders from the central and eastern regions of Iran. The detection of H. pylori and screening of cagA was performed by polymerase chain reaction (PCR). The pattern of the motifs was determined by PCR followed by sequencing. Results: The overall prevalence of H. pylori was 66.3% in eastern (Mashad) and 50.6% in the central (Isfahan) part of Iran. The frequency of cagA-positive strains in Mashad and Isfahan were 63.4% and 56.7%, respectively. The pattern of EPIYA motif was as follows: 43 (79.6%) ABC, 7 (12.9%) AB, 4 (7.4%) ABCC, and one (1.9%) ABCCC. We also identified a novel EPIYA C sequence motif which showed association with gastric cancer (GC). The relationship between the frequency of specific EPIYA motifs and GC was statistically significant (P < 0.05). Conclusions: This is the first report for the determination of the cagA EPIYA motif of H. pylori in the Northeast and center of Iran. The prevalence of cagA positive H. pylori between the two regions was significant (P ≤ 0.05). All isolates of the H. pylori cagA were western type (ABC). The increase in the number of EPIYA-C repeats was associated with GC (P ≤ 0.01).

Epstein-barr virus/Helicobacter pylori coinfection and gastric cancer: the possible role of viral gene expression and shp1 methylation.

Background and Objectives: Among the various factors involved in the development of gastric cancer (GC), infectious agents are one of the most important causative inducers. This study aimed to investigate the possible role of EBV gene expression on SHP1 methylation in co-infection with Helicobacter pylori in patients with GC. Materials and Methods: Formalin-fixed paraffin-embedded samples were obtained from 150 patients with gastrointestinal disorders. The presence of the H. pylori and EBV genome were examined by PCR. The expression level of viral gene transcripts and methylation status of the SHP1 cellular gene was assessed by quantitative real-time PCR and methyl-specific PCR. Results: EBV and H. pylori coinfection were reported in 5.6% of patients. The mean DNA viral load was significant in patients coinfected with cagA-positive H. pylori (P= 0.02). The expression of BZLF1 and EBER was associated with GC. Also, the expression level of BZLF1in GC tissues was significantly higher in coinfection (P = 0.01). SHP1 methylation frequency was higher in the GC group than in the control group (P = 0.04). The correlation between the methylation rate and the H. pylori infection was highly significant (P<0.0001). The strongest positive correlation was observed in GC specimens between SHP1 methylation and H. pylori cagA-positive strains (p= 0.003). Conclusion: Our results suggested that cagA might involve in the elevation of EBV lytic gene expression and SHP1 methylation, and the development of gastric cancer. Understanding the mechanism of EBV H. pylori - cagA + coinfection, as well as host epigenetic changes, can play an important role in diagnosing and preventing gastric cancer.

Effect of IL-2 co-expressed or co-inoculated with immuno-dominant epitopes from VP1 protein of FMD virus on immune responses in BALB/c mice.

OBJECTIVES: The results of studies on vaccine development for foot-and-mouth disease (FMD) virus show that the use of inactivated vaccines for FMD virus is not completely effective. Novel vaccinations based on immuno-dominant epitopes have been shown to induce immune responses. Furthermore, for safety of immunization, access to efficient adjuvants against FMD virus seems to be critical. MATERIALS AND METHODS: In this study, we produced epitope recombinant vaccines from the VP1 protein of the FMD virus for serotype O of Iran. Constructs were included polytope (tandem-repeat multiple-epitope), polytope coupled with interleukin-2 (polytope-IL 2) as a molecular adjuvant and IL-2. Three expression vectors were constructed and expressed in Escherichia coli BL21 (DE3). To evaluate whether these recombinant vaccines induce immune responses, BALB/c mice were injected with the recombinant vaccines and their immune responses were compared with a negative control group. The humoral and cellular immune responses were measured by ELISA. RESULTS: The results showed that IL-2 co-expressed or co-inoculated with Polytope protein enhances the immune effect of multiple epitope recombinant vaccine against FMD virus. The results of total immunoglobulin G (IgG), IgG1, and IgG2a levels and secretion of interferon gamma (IFN-γ), IL-4 and IL-10 revealed that there were significant differences between negative control group and other injected mice with the recombinant vaccines (P<0.05). CONCLUSION: Observations indicated that the epitope recombinant plasmid of the VP1 protein co-expressed or co-inoculated with IL-2 was effective in inducing an enhanced immune response. Therefore, IL-2 can be recommended as a potential adjuvant for epitope recombinant vaccine of the VP1 protein from FMD virus.

Identification and phylogenetic analysis of contagious ecthyma virus from camels (Camelus dromedarius) in Iran.

Contagious ecthyma is a highly contagious disease affecting domestic and wild ruminants such as sheep, goats and camels. The identification and characterisation of a parapoxvirus (PPV) infecting camels is described here. The virus was detected in dromedary camels (Camelus dromedarius) from Kerman and Shiraz in Iran. PPV-specific amplification by polymerase chain reaction (PCR) further confirmed that the disease was associated with PPV infection. Phylogenetic analysis of ORF011 (B2L) gene sequences showed 99.79% and 82.13% similarity of the PPV identified in this study with the Jodhpur isolate and the bovine papular stomatitis virus (BPSV) isolates (CE41), respectively. Moreover, phylogenetic analysis of the ORF045 gene indicated that the Shiraz sample was in all probability closely related to VR634 and to F00.120R and PCPV776. In conclusion, the results suggest that camel PPV (CPPV) is a likely cause of contagious ecthyma in dromedary camels in Iran.

Exploring Hepatitis From the Perspective of Iranian Traditional Medicine: Using a Qualitative Approach.

Hepatitis is a global health problem, with significant adverse impacts on patients' quality of life. In this study, we aimed to review major resources of Iranian traditional medicine and determine whether the etiology and semiology of hepatitis, in particular chronic hepatitis, in traditional and conventional medicine might be aligned. Through such studies, we might be able to develop new approaches for clinical research to improve our current knowledge on the etiology and treatment of this condition. In this qualitative study, recently published studies, scientific databases, and reliable Iranian traditional medicine resources, including the Canon of Medicine, were searched. The integrative use of conventional and traditional medicine for diagnostic and therapeutic purposes could be evaluated to develop new modalities for dealing with this condition. An integrated approach is recommended in clinical research in order to find more efficient and safer treatment.

Design and Construction of Chimeric VP8-S2 Antigen for Bovine Rotavirus and Bovine Coronavirus.

PURPOSE: Bovine Rotavirus and Bovine Coronavirus are the most important causes of diarrhea in newborn calves and in some other species such as pigs and sheep. Rotavirus VP8 subunit is the major determinant of the viral infectivity and neutralization. Spike glycoprotein of coronavirus is responsible for induction of neutralizing antibody response. METHODS: In the present study, several prediction programs were used to predict B and T-cells epitopes, secondary and tertiary structures, antigenicity ability and enzymatic degradation sites. Finally, a chimeric antigen was designed using computational techniques. The chimeric VP8-S2 antigen was constructed. It was cloned and sub-cloned into pGH and pET32a(+) expression vector. The recombinant pET32a(+)-VP8-S2 vector was transferred into E.oli BL21CodonPlus (DE3) as expression host. The recombinant VP8-S2 protein was purified by Ni-NTA chromatography column. RESULTS: The results of colony PCR, enzyme digestion and sequencing showed that the VP8-S2 chimeric antigen has been successfully cloned and sub-cloned into pGH and pET32a(+).The results showed that E.coli was able to express VP8-S2 protein appropriately. This protein was expressed by induction of IPTG at concentration of 1mM and it was confirmed by Ni-NTA column, dot-blotting analysis and SDS-PAGE electrophoresis. CONCLUSION: The results of this study showed that E.coli can be used as an appropriate host to produce the recombinant VP8-S2 protein. This recombinant protein may be suitable to investigate to produce immunoglobulin, recombinant vaccine and diagnostic kit in future studies after it passes biological activity tests in vivo in animal model and or other suitable procedure.

Production of specific IgY antibody to the recombinant FanC protein produced in Escherichia coli.

OBJECTIVES: Enterotoxigenic Escherichia coli (ETEC) strains are one of the primary causes of diarrhea in newborn calves and in humans, pigs, and sheep. IgY technology has been identified as a promising alternative to generating a mass amount of specific antibody for use in immunotherapy and immunodiagnostics. The purpose of this study was to produce specific antibody by egg yolk antibody (IgY) to recombinant FanC protein from ETEC. MATERIALS AND METHODS: FanC (K99) gene was amplified from ETEC by specific primers and polymerase chain reaction. The gene was cloned and subcloned into pTZ57R/T and pET32a (+) vectors, respectively. Recombinant vector was transferred into E. coli BL21 CodonPlus (DE3). Protein expression was investigated by 1 mM IPTG induction. Hens were immunized by the purified recombinant FanC protein. The activity and specificity of the IgY antibody were detected by dot-blotting, Western blotting, and indirect ELISA. RESULTS: We obtained FanC specific IgYs by immunizing the hens with the recombinant FanC protein. The anti-FanC IgY showed binding specifically to the FanC protein of ETEC. CONCLUSION: The results emphasize that specific IgY against the recombinant FanC protein could be recommended as a candidate for passive immunization against ETEC infection in animals and humans.

Study of Lactic Acid Bacteria Community From Raw Milk of Iranian One Humped Camel and Evaluation of Their Probiotic Properties.

BACKGROUND: Camel milk is amongst valuable food sources in Iran. On the other hand, due to the presence of probiotic bacteria and bacteriocin producers in camel milk, probiotic bacteria can be isolated and identified from this food product. OBJECTIVES: The objectives of the present research were the isolation and molecular identification of lactic acid bacteria from camel milk and evaluation of their probiotic properties. MATERIALS AND METHODS: A total of ten samples of camel milk were collected from the Golestan province of Iran under aseptic conditions. Bacteria were isolated by culturing the samples on selective medium. Isolates were identified by amplification of the 16S rDNA and Internal Transcribed Spacer (ITS) region between the 16S and 23S rRNA genes by Polymerase Chain Reaction (PCR) and were then screened and grouped by the Amplified Ribosomal DNA Restriction Analysis (ARDRA) method. To evaluate probiotic properties, representative isolates of different ARDRA profiles were analyzed. The antimicrobial activity of Lactic Acid Bacteria (LAB) against Pediococcus pentosaceus, Escherichia coli and Bacillus cereus was examined by the agar diffusion assay. Acid and bile tolerance of isolates were evaluated. RESULTS: A total of 64 isolates were analyzed based on biochemical tests and morphological characteristics. The most frequently isolated LAB was Enterococci. Weissella, Leuconostoc, Lactobacilli and Pediococci were less frequently found. Based on restriction analysis of the ITS, the isolates were grouped into nine different ARDRA patterns that were identified by ribosomal DNA sequencing as P. pentosaceus, Enterococcus faecium strain Y-2, E. faecium strain JZ1-1, E. faecium strain E6, E. durans, E. lactis, Leuconostoc mesenteroides, Lactobacillus casei and Weissella cibaria. The results showed that antimicrobial activity of the tested isolates was remarkable and P. pentosaceus showed the most antibacterial activity. In addition, E. durans, E. lactis, L. casei and P. pentosaceus were selected as probiotic bacteria. CONCLUSIONS: This study revealed the presence of bacteriocin-producing bacteria and probiotic bacteria in camel milk from the Golestan province of Iran.