Human Herpesvirus 6 Infection and Risk of Chronic Fatigue Syndrome: A Systematic Review and Meta-Analysis.

INTRODUCTION: Chronic fatigue syndrome (CFS) is a neurological disease that is accompanied by excessive fatigue or tiredness. There are several reports confirming the association between human herpesvirus 6 (HHV-6) infection and CFS illness. This systematic review and meta-analysis was performed to integrate the information of published studies with regard to this association until May 2021. METHODS: The literature search was based on keywords including "chronic fatigue syndrome and HHV 6," "chronic fatigue syndrome and HHV-6," "chronic fatigue syndrome and HHV6," "chronic fatigue syndrome and Herpes virus 6," and "chronic fatigue syndrome and Herpesvirus6" in MEDLINE (PubMed), Web of Science, and EMBASE. RESULTS: The literature search identified 17 studies to be included in the systematic review and 11 studies in meta-analysis. The symmetry funnel plot and Egger's test (p value = 0.2) identified no publication bias among studies. Moreover, the low level of I2 revealed homogeneity across studies. DISCUSSION: In conclusion, the association between the HHV-6 infection and CFS incidence was substantiated. However, the results of this study also suggest that further comprehensive studies are needed to solidify the association between HHV-6 and CFS. Future studies should consider additional factors that may have affected the significance of such a correlation.

Identifying novel biomarkers of the pediatric influenza infection by weighted co-expression network analysis.

BACKGROUND: Despite the high yearly prevalence of Influenza, the pathogenesis mechanism and involved genes have not been fully known. Finding the patterns and mapping the complex interactions between different genes help us to find the possible biomarkers and treatment targets. METHODS: Herein, weighted gene co-expression network analysis (WGCNA) was employed to construct a co-expression network among genes identified by microarray analysis of the pediatric influenza-infected samples. RESULTS: Three of the 38 modules were found as the most related modules to influenza infection. At a functional level, we found that the genes in these modules regulate the immune responses, protein targeting, and defense to virus. Moreover, the analysis of differentially expressed genes disclosed 719 DEGs between the normal and infected subjects. The comprehensive investigation of genes in the module involved in immune system and viral defense (yellow module) revealed that SP110, HERC5, SAMD9L, RTP4, C19orf66, HELZ2, EPSTI1, and PHF11 which were also identified as DEGs (except C19orf66) have the potential to be as the biomarkers and also drug targeting for the treatment of pediatric influenza. CONCLUSIONS: The WGCN analysis revealed co-expressed genes which were involved in the innate immune system and defense to virus. The differentially expressed genes in the identified modules can be considered for designing drug targets. Moreover, modules can help to find pathogenesis routes in the future.

Reconnaissance of the candidate genes involved in the pathogenesis of human immunodeficiency virus and targeted by antiretroviral therapy.

The expression levels of many genes change after treatment of human immunodeficiency virus (HIV)-infected subjects by antiretroviral drugs. High-throughput analysis of tremendous datasets led to the discovery of genes that are implicated in the treatment pathways. In this study, we performed a gene-enrichment analysis after determining the differentially expressed genes (DEGs) between untreated HIV-positive and HIV-negative subjects and also between treated HIV-positive subjects with antiretroviral therapy (ART; who receiving nucleoside reverse transcriptase inhibitor-based ART) and untreated HIV-positive cases in the peripheral blood mononuclear cells (PBMCs), adipose, and muscle tissues. In sum, the genes that activate inflammatory, immune response, proliferation, metabolism, and viral involvement pathways have different expression patterns in the untreated HIV-positive subjects and treated HIV-positive cases. Moreover, the expression levels of the genes including ACLY, ALDH18A1, HADHA, and YARS in the PBMCs tissue and HBEGF, PKN3, DEGS2, and EDN3 in the fat tissue were found to be different in the HIV-infected patients, which can be considered as new biomarkers for HIV infection.

Genetic and Phylogenetic Characterization of the M Gene of Influenza A Virus Isolated from Iranian Patients.

BACKGROUND: A few studies have been done on the molecular analysis of Iranian influenza A isolates M gene. METHODS: In 2014, nasal swabs collected from outpatients with clinical symptoms in the hospital clinics of Tehran, Iran were subjected for influenza detection and subtyping using Real-Time RT-PCR. Sequence and phylogenetic analysis performed on four randomly selected isolates from each subtype (H1N1 and H3N2) using neighbor-joining method. RESULTS: Phylogenetic dendrograms drawn based on M nucleotide sequence of H1N1 isolates showed close relatedness with Omanian isolates while the most isolates of H3N2 have clustered with Kuwait isolates and isolates from outside of geographical location. Amino acid sequence analysis showed S31N substitution in all isolates rendering the virus resistant to adamantanes. CONCLUSION: This study determined the sequence identity and phylogenetic relatedness of M gene sequence got from Iranian influenza A isolates to elucidate the modality of relationship of this gene in comparison with its counterparts from other regions.

A Feasibility Study to Evaluate Bacillus subtilis as a Host for Producing Recombinant Human Parathyroid Hormone.

BACKGROUND: Biosynthetic teriparatide (1-34) (TPD) is a N-terminally truncated version of human parathyroid hormone (hPTH). The recombinant form of this polypeptide has been expressed in Escherichia coli (E. coli) and approved as the first anabolic treatment of osteoporosis in the EU and the USA. Feasibility of expression and secretion of a tag- fused form of TPD into Bacillus subtilis (B. subtilis) was examined due to several advantages of B. subtilis over E. coli in production of recombinant proteins with pharmacological activities. METHODS: A codon optimized gene containing TPD open reading frame carrying enterokinase site in its upstream was fully synthesized. According to our cloning scheme, this synthetic polynucleotide was used as a template for PCR amplification using engineered primers in such a way that a polyhistidin tag was added in frame to the upstream of the amplicon as well as two restriction sites at its ends. The resulted amplicon, a cassette containing His-tag, enterokinase site and TPD, from 5' to 3', was cloned into pTZ57R/T vector and subjected to sequencing.The cassette was then subcloned into pHT43 shuttle vector and transformed into B. subtilis. Expression of target protein was analyzed by SDS-PAGE and western blotting upon induction by IPTG. RESULTS: The accuracy of construction of pHT43-TPD was confirmed by sequencing and restriction map analyses. SDS-PAGE and western blotting results showed that the recombinant fusion form of hPTH was successfully expressed and secreted into cytoplasm and extracellular medium. CONCLUSION: TPD may be successfully expressed and secreted in B. subtilis; however, optimization of expression conditions is required for enhancing target protein yield.

Characterization of the neuraminidase genes from human influenza A viruses circulating in Iran from 2010 to 2015.

BACKGROUND: Characterization of influenza viruses is critical for detection of new emerging variants. Herein, we analyzed the genetic diversity and drug susceptibility of the neuraminidase gene (NAs) expressed by influenza A/H1N1pdm09 and A/H3N2 viruses circulating in Iran from 2010 to 2015. METHODS: We genetically analyzed the NAs of 38 influenza A/H1N1pdm09 and 35 A/H3N2 isolates. RESULTS: The Iranian A/H1N1pdm09 viruses belonged to seven genogroups/subgenogroups, with the dominant groups being genogroups 6B and 6C. The A/H3N2 isolates fell into six gneogroups/subgenogroups, with the dominant genogroups being 3C and 3C.2a. The most common mutations detected among the A/H1N1pdm09 viruses included N44S, V106I, N200S, and N248D. All H1N1pdm09 viruses were genetically susceptible to the NAIs. However, one A/H1N1pdm09 virus from the 2013-2014 season possessed an NA-S247N mutation, which reduces the susceptibility to oseltamivir. In case of H3N2, none of the analyzed Iranian strains carried a substitution that might affect its susceptibility to NAIs. CONCLUSION: The ongoing evolution of influenza viruses and the detect of influenza viruses with reduced susceptibility to NAIs warrants continuous monitoring of the circulating strains.

Prevalence of human influenza virus in Iran: Evidence from a systematic review and meta-analysis.

This systematic review and meta-analysis was conducted to consolidate the information on the prevalence of the human influenza virus, including H1N1 and H3N2 as well as B-type influenza across Iran from 2000 to December 2016. The literature search was based on keywords including "influenza and Iran", "human influenza", "prevalence", "relative frequency", "incidence", and "drug" in MEDLINE (PubMed), Web of Science, Scopus, ScienceDirect, the Iranian Research Institute for Information Science and Technology (IranDoc), the Regional Information Centre for Science & Technology (RICeST), and the Scientific Information Database (SID). The literature search revealed 25 prevalence and seven drug resistance studies. In order to investigate the publication bias among studies, funnel plots and Egger's test were used. Additionally, the statistical tests of I2, Chi2, and Tau2 were computed, and the results were visualized with forest plots. A high level of I2 and Chi2 were obtained among studies, which are representative of the high variation and remarkable heterogeneity between studies. This may be because of various methodologies applied in the studies such as study design, age groups, and different populations. The prevalence of influenza H1N1, H3N2, and B in Iran have not yet been well evaluated. The heterogeneity among studies reveals that more attention should be paid to considering various factors, including gender, population size, and underlying conditions.

Molecular characterization and phylogenetic analysis of human influenza A viruses isolated in Iran during the 2014-2015 season.

Influenza A viruses are an important cause of severe infectious diseases in humans and are characterized by their fast evolution rate. Global monitoring of these viruses is critical to detect newly emerging variants during annual epidemics. Here, we sought to genetically characterize influenza A/H1N1pdm09 and A/H3N2 viruses collected in Iran during the 2014-2015 influenza season. A total of 200 nasopharyngeal swabs were collected from patients with influenza-like illnesses. Swabs were screened for influenza A and B using real-time PCR. Furthermore, positive specimens with high virus load underwent virus isolation and genetic characterization of their hemagglutinin (HA) and M genes. Of the 200 specimens, 80 were influenza A-positive, including 44 A/H1N1pdm09 and 36 A/H3N2, while 18 were influenza B-positive. Phylogenetic analysis of the HA genes of the A/H1N1pdm09 viruses revealed the circulation of clade 6C, characterized by amino acid substitutions D97N, V234I and K283E. Analysis of the A/H3N2 viruses showed a genetic drift from the vaccine strain A/Texas/50/2012 with 5 mutations (T128A, R142G, N145S, P198S and S219F) belonging to the antigenic sites A, B, and D of the HA protein. The A/H3N2 viruses belonged to phylogenetic clades 3C.2 and 3C.3. The M gene trees of the Iranian A/H1N1pdm09 and A/H3N2 mirrored the clustering patterns of their corresponding HA trees. Our results reveal co-circulation of several influenza A virus strains in Iran during the 2014-2015 influenza season.

Molecular Analysis of Ganciclovir-Resistant Cytomegalovirus in Renal Transplant Recipients with High Viral Load.

BACKGROUND: Gancyclovir-resistant (GanR) cytomegalovirus (CMV) remains an issue, especially in solid organ transplant (SOT) recipients. Some mutations in UL54 and UL97 confer this resistance. Long-lasting high-dose drug exposure, high viral load, together with lack of sufficient compliance with treatment may account for these mutations. The aim of this study was to detect UL97 and UL54 putative mutations conferring ganciclovir-resistance in renal organ transplant recipients with high CMV load. METHODS: In this cross-sectional study, 58 serum samples were collected from renal transplant recipients who had referred to three hospitals in Tehran from January 2014 to June 2015. Specific criteria such as CMV syndrome, presence of CMV in blood and organ dysfunction were considered. Then, they were tested for viral load in their early fourth month of intravenous ganciclovir treatment. Fifty cases revealing more than 200 copies/mL were analyzed for mutations. Two fragments of UL54 and Ul97 genes were amplified and sequenced bidirectionally. Sequence alignment and statistical analysis were performed by Mutation Surveyor software and t-test respectively. RESULTS: A significant difference was observed in viral load between seronegative and seropositive recipients (P = 0.036). The most frequent mutation was related to D605E in UL97 gene with the rate of 25%. Regardless of viral load, neither putative mutation nor simultaneous mutation was detected in either UL97 and UL54 regions. CONCLUSION: In spite of high viral load and persistence of symptoms, our population study did not reveal putative mutations. Hence, the direct relationship between the presence of high quantity of CMV and the occurrence of putative mutation cannot be considered. Non-putative gancyclovir resistant mutations and prolonged drug exposure may have a role in these manifestations.

Construction and eukaryotic expression of recombinant large hepatitis delta antigen.

BACKGROUND: Hepatitis delta virus (HDV) is a subviral human pathogen that exploits host RNA editing activity to produce two essential forms of the sole viral protein, hepatitis delta antigen (HDAg). Editing at the amber/W site of HDV antigenomic RNA leads to the production of the large form (L-HDAg), which is required for RNA packaging. METHODS: In this study, PCR-based site-directed mutagenesis by the overlap extension method was used to create the point mutation converting the small-HDAg (S-HDAg) stop codon to a tryptophan codon through three stages. RESULTS: Sequencing confirmed the desirable mutation and integrity of the L-HDAg open reading frame. The amplicon was ligated into pcDNA3.1 and transfected to Huh7 and HEK 293 cell lines. Western blot analysis using enhanced chemiluminescence confirmed L-HDAg expression. The recombinant L-HDAg localized within the nuclei of cells as determined by immunofluorescence and confocal microscopy. CONCLUSION: Because L-HDAg requires extensive post-translational modifications, the recombinant protein expressed in a mammalian system might be fully functional and applicable as a tool in HDV molecular studies, as well as in future vaccine research.

Design and characterization of an aequorin-based bacterial biosensor for detection of toluene and related compounds.

An aequorin-based Escherichia coli strain JM109 biosensor was constructed and characterized for its potential to detect toluene and related compounds in aqueous solutions. The biosensor was constructed based on a PGL2 plasmid carrying the lower pathway promoter (Pu) of the xyl operon of Pseudomonas putida mt-2, which was incorporated with transcriptional activator xylR and fused to aequorin cDNA named pGL2-aequorin. Binding of xylR protein to a subset of toluene-like compounds activates transcription at the Pu promoter, thus expression of aequorin is controlled by xylR and Pu. In this work we have compared the effect of Shine-Dalgarno (SD) and T2 rrnß terminator sequence in the expression of aequorin. According to the sensitivity of aequorin and increase in the signal-to-noise ratio, this reporter enzyme has reasonable sensitivity compared with other reporter systems. The results indicate higher expression of aequorin in the presence of SD and T2 rrnß. The activity of aequorin in recombinant whole-cell biosensor was linear from 1 to 500 µm of toluene. The bioluminescence response was specific for toluene-like molecules, so this biosensor cells would be able to detect toluene derivative contamination in environmental samples, accurately.

Enhancement of protective humoral immune responses against Herpes simplex virus-2 in DNA-immunized guinea-pigs using protein boosting.

Genital Herpes is a common sexually transmitted disease that is caused mostly by Herpes simplex virus type 2 (HSV-2). Its prevalence has increased in developing countries in spite of the availability of valuable antiviral drug therapy. Considering the importance of HSV-2 infections, effective vaccines remain the most likely hope for controlling the spread of HSV diseases. In the present study, the complete HSV-2 glycoprotein D gene was isolated and cloned into different plasmid vectors to construct a DNA vaccine and prepare recombinant subunit vaccines using a baculovirus expression system. The vaccines were tested alone or in combination to evaluate their ability to induce protective immunity in guinea-pigs against genital HSV infections. Immunization elicited humoral responses as measured by neutralization tests and enzyme-linked immunosorbent assay, and immunized animals had less severe genital skin disease as well as reduced replication of the challenging virus in the genital tract during experimental infection. Our results further demonstrate that DNA priming-protein boosting induced a neutralizing antibody titer higher than that obtained with DNA-DNA vaccination. The massive increase of antibody titer following DNA priming-protein boosting might be attributed to a recall of B cell memory.

In vitro replication of hepatitis D virus using a new construct containing a cDNA dimer of HDV genome.

BACKGROUND: There is no cell line susceptible to hepatitis D virus (HDV) infection and capable of stable replication of its genome. Different genetic-based approaches have been introduced to initiate HDV replication events so far. METHODS: In order to construct a replicative model for HDV made from a unique genome sequence, two monomeric units of HDV full-length cDNA were joined together through a four-step cloning scheme. The resulting vector (pcDNA3.1-D2) containing two tandem repeats of HDV cDNA under CMV promoter control was then used in transfection experiments into COS7 and HuH7 cell lines. RESULTS: HDV replication markers including expression of hepatitis delta antigen (HDAg), the only HDV-specific antigen, and synthesis of antigenomic RNA were shown in both transfected cell lines, indicating initiation of HDV genome replication. CONCLUSIONS: Our results suggested that pcDNA3.1-D2, a vector containing a cDNA dimer of the HDV genome, originated from a unique full-length HDV molecule that is capable of replicating in cultured cells. This vector can be used conveniently for transfection experiments to study HDV molecular biology.

Molecular phylogenetic analysis of Iranian HDV complete genome.

Hepatitis D (delta) virus (HDV) is a subviral pathogen agent and a satellite of Hepatitis B virus. Three distinct genotypes are described for HDV; genotype I is distributed worldwide but other genotypes appear to be more restricted geographically. In the present study, the entire nucleotide sequence of an HDV isolate from an Iranian patient (IR-1) was obtained using twelve pairs of primers to amplify six overlapping fragments covering the whole HDV genome by RT-nested PCR. Phylogenetic and pairwise alignments were done on this new isolate to determine IR-1 position among other isolates. Our results indicate that IR-1 contains 1676 nucleotides encoding 214 a.a. of the hepatitis delta antigen (HDAg). This new isolate belongs to genotype I with most sequence similarity to an Italian HDV isolate (92.6%). At amino acid level, predicted HDAg sequence of IR-1 revealed the most homology with those of Italian and Lebanese isolates. Data analysis confirmed genetic variability and heterogeneity of the HDV species isolated from different geographical areas.