Publications by authors named "Fahad N Almajhdi"

32 Publications

Sequence and phylogentic analysis of MERS-CoV in Saudi Arabia, 2012-2019.

Virol J 2021 04 30;18(1):90. Epub 2021 Apr 30.

Department of Botany and Microbiology, College of Science, King Saud University, Riyadh, 11451, Saudi Arabia.

Background: The Middle East Respiratory Syndrome-related Coronavirus (MERS-CoV) continues to exist in the Middle East sporadically. Thorough investigations of the evolution of human coronaviruses (HCoVs) are urgently required. In the current study, we studied amplified fragments of ORF1a/b, Spike (S) gene, ORF3/4a, and ORF4b of four human MERS-CoV strains for tracking the evolution of MERS-CoV over time.

Methods: RNA isolated from nasopharyngeal aspirate, sputum, and tracheal swabs/aspirates from hospitalized patients with suspected MERS-CoV infection were analyzed for amplification of nine variable genomic fragments. Sequence comparisons were done using different bioinformatics tools available.

Results: Several mutations were identified in ORF1a/b, ORF3/4a and ORF4b, with the highest mutation rates in the S gene. Five codons; 4 in ORF1a and 1 in the S gene, were found to be under selective pressure. Characteristic amino acid changes, potentially hosted and year specific were defined across the S protein and in the receptor-binding domain Phylogenetic analysis using S gene sequence revealed clustering of MERS-CoV strains into three main clades, A, B and C with subdivision of with clade B into B1 to B4.

Conclusions: In conclusion, MERS-CoV appears to continuously evolve. It is recommended that the molecular and pathobiological characteristics of future MERS-CoV strains should be analyzed on regular basis to prevent potential future outbreaks at early phases.
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http://dx.doi.org/10.1186/s12985-021-01563-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8085657PMC
April 2021

Induction of Immune Responses and Immune Evasion by Human Bocavirus.

Int Arch Allergy Immunol 2021 Apr 19:1-8. Epub 2021 Apr 19.

Virology Research Group, Department of Botany and Microbiology, College of Science, King Saud University, Riyadh, Saudi Arabia.

Respiratory tract infections are the primary cause of morbidity and mortality globally. Human bocavirus 1 (HBoV1), a member of the Parvoviridae family causes a wide spectrum of respiratory diseases in children, and gastroenteritis in adults. The mechanisms of latency, persistence, and reinfection of Bocavirus are poorly understood at present due to the lack of permissive cell lines and efficient animal models. Moreover, the dual infections of HBoV and other respiratory viruses further complicate the study of the pathogenicity of Bocaviruses. The data on immunological consequences of Bocavirus infection are sparse. However, the existing data have highlighted the role of CD4 T cells in Bocavirus infection. High titres of HBoV-specific antibodies have been detected in different populations suggesting its ubiquitous prevalence. Interestingly, the mechanism employed by Bocavirus to evade the immune system mostly targets type I IFN pathways and cause pyroptotic cell death of host cells. This review summarizes the immune responses evoked in response to Bocavirus infection, escape mechanism employed by the virus, and the vaccination strategies, including antisense technology to combat Bocavirus infections.
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http://dx.doi.org/10.1159/000514688DOI Listing
April 2021

Influence of RFC1 c.80A>G Polymorphism on Methotrexate-Mediated Toxicity and Therapeutic Efficacy in Rheumatoid Arthritis: A Meta-analysis.

Ann Pharmacother 2021 Mar 22:10600280211002053. Epub 2021 Mar 22.

Research Chair for Biomedical Application of Nanomaterials, Biochemistry Department, College of Science, King Saud University, Riyadh, Saudi Arabia.

Background: Methotrexate (MTX) is an antirheumatic drug, transported by reduced folate carrier-1 (RFC1). The most common RFC1 gene variant, c.80 A>G (rs1051266) is ambiguously linked to adverse effects of MTX therapy in some rheumatoid arthritis (RA) patients.

Objective: The purpose of meta-analysis was to summarize all major published studies on c.80 A>G SNP to clarify this ambiguity in MTX therapy.

Methods: A total of 18 studies representing 3592 RA patients comprising 699 men and 2893 women were included. Both fixed and random effect models were applied to study the data.

Results: The RFC1 80A-allele showed null association with MTX-mediated toxicity in both fixed (odds ratio [OR] = 0.91; 95% CI = 0.80-1.03) and random effects (OR = 0.89; 95% CI: 0.71-1.11) models. Because heterogeneity was observed in this association ( = 0.0006), data were segregated based on use of folate therapy. In 7 studies (n = 1191) where folate was used along with MTX, RFC1 AA patients showed reduced risk for MTX-mediated toxicity (OR = 0.67; 95% CI: 0.50-0.89; = 0.0006). The RFC1 80A-allele was found to increase the efficacy of MTX therapy by 1.53-fold (95% CI: 1.24-1.88), whereas the 80AA-genotype increased the efficacy by 1.85-fold (95% CI: 1.41-2.42). No publication bias was observed in these associations.

Conclusion And Relevance: RFC1 c.80 A>G is an important pharmacogenetic determinant of MTX therapy in RA. The RFC1 80A-allele robustly increased therapeutic efficacy and safety when folate was used along with MTX. Findings are relevant to decision-making in the clinical use of MTX as a treatment for RA patients harboring the RFC1 gene variant.
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http://dx.doi.org/10.1177/10600280211002053DOI Listing
March 2021

Double-edged role of natural killer cells during RSV infection.

Int Rev Immunol 2020 29;39(5):233-244. Epub 2020 May 29.

Virology Research Group, Department of Botany and Microbiology, College of Science, King Saud University, Riyadh, Saudi Arabia.

Natural killer cells play a vital role in the rejection of tumors and pathogen-infected cells. NK cells are indispensable in the early immune response against viral infections by directly targeting infected cells. Furthermore, NK cells influence adaptive immunity by driving virus-specific T-cell responses. Respiratory syncytial virus, a highly contagious virus that causes bronchiolitis, is the main reason for mortality in infants and elderly patients. RSV infection triggers both innate and adaptive immune responses. However, immunity against RSV is ephemeral due to the impaired development of immunological memory. The role of NK cells during RSV infection remains ambiguous. NK cells play a dual role in RSV infection; initially, their role is a protective one as they utilize their intrinsic cytotoxicity, followed by a detrimental one that induces lung injury due to the inhibition of antibody responses and the secretion of pro-inflammatory factors. Noteworthy, IFN-γ released from NK cells play a critical role in promoting a shift to adaptive responses and inhibiting antibody responses in neonates. Indeed, NK cells have a pro-inflammatory and inhibitory role rather than a cytotoxic one that contributes to the severity of the disease. Therapeutic options, including DNA-protein-based vaccines, synthetic peptides, and attenuated strains, are presently under tests. However, there is a need for effective strategies to augment NK cell activity and circumvent the pro-inflammatory activity to benefit the host. In this review, we focused on the role played by NK cells in the immune response and its outcome on the immunopathogenesis of RSV disease.
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http://dx.doi.org/10.1080/08830185.2020.1770748DOI Listing
May 2020

Inhibition of hepatitis C virus genotype 4 replication using siRNA targeted to the viral core region and the CD81 cellular receptor.

Cell Stress Chaperones 2020 03 14;25(2):345-355. Epub 2020 Feb 14.

College of Medicine Research Center, King Saud University, Riyadh, Saudi Arabia.

Hepatitis C virus (HCV) is one of the most important causative agents of hepatitis worldwide. The current study aimed to evaluate the silencing effect of the small interference RNA (siRNA) molecules designed against the core region of HCV genotype 4 (HCV-4) and the CD81 gene, which is the cellular receptor for HCV in the human hepatocytes. RT-PCR was used to measure the changes in both the viral HCV core and the cellular CD81 genes induced by the specific siRNA molecules. Additionally, the fluctuations in either the viral or the cellular proteins of the target regions were tested by flow cytometry and immunofluorescence. The results showed the effectiveness of the used siRNA molecules against the target genes in either RNA or protein levels. The effect of 100 nM of siCD81 and 40 nM of siCore was more evident at 24 and 48 h post-transfection. The combination of the two siRNA molecules resulted in an extra inhibitory effect of the HCV core at both the RNA (85.6%) and protein (98.5%) levels. The current study suggested that targeting of the CD81 cellular receptor and/or the viral HCV core region by the small interference molecules might be a suitable choice in the suppression of HCV-4 replication. This might assist the development of new antiviral medications and provides a new alternative strategy for the targeting and treatment of HCV genotype 4.
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http://dx.doi.org/10.1007/s12192-020-01077-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7058739PMC
March 2020

The emergence of subgenotype ON-1 of Human orthopneumovirus type A in Riyadh, Saudi Arabia: A new episode of the virus epidemiological dynamic.

J Med Virol 2020 08 9;92(8):1133-1140. Epub 2019 Dec 9.

Department of Botany and Microbiology, College of Science, King Saud University, Riyadh, Saudi Arabia.

Lower respiratory tract infections caused by Human orthopneumovirus are still a threat to the pediatric population worldwide. To date, the molecular epidemiology of the virus in Saudi Arabia has not been adequately charted. In this study, a total of 205 nasopharyngeal aspirate samples were collected from hospitalized children with lower respiratory tract symptoms during the winter seasons of 2014/15 and 2015/16. Human orthopneumovirus was detected in 89 (43.4%) samples, of which 56 (27.3%) were positive for type A and 33 (16.1%) were positive for type B viruses. The fragment that spans the two hypervariable regions (HVR1 and HVR2) of the G gene of Human orthopneumovirus A was amplified and sequenced. Sequence and phylogenetic analyses have revealed a genotype shift from NA1 to ON-1, which was prevalent during the winter seasons of 2007/08 and 2008/09. Based on the intergenotypic p-distance values, ON-1 was reclassified as a subgenotype of the most predominant genotype GA2. Three conserved N-glycosylation sites were observed in the HVR2 of Saudi ON-1 strains. The presence of a 23 amino acid duplicated region in ON-1 strains resulted in a higher number of O-glycosylation sites as compared to other genotypes. The data presented in this report outlined the replacement of NA1 and NA2 subgenotypes in Saudi Arabia with ON-1 within 7 to 8 years. The continuous evolution of Human orthopneumovirus through point mutations and nucleotide duplication may explain its ability to cause recurrent infections.
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http://dx.doi.org/10.1002/jmv.25643DOI Listing
August 2020

Epidemiology of respiratory viruses in Saudi Arabia: toward a complete picture.

Arch Virol 2019 Aug 28;164(8):1981-1996. Epub 2019 May 28.

Department of Botany and Microbiology, College of Science, King Saud University, P.O. Box 2455QA6, Riyadh, 11451, Saudi Arabia.

Acute lower respiratory tract infection is a major health problem that affects more than 15% of the total population of Saudi Arabia each year. Epidemiological studies conducted over the last three decades have indicated that viruses are responsible for the majority of these infections. The epidemiology of respiratory viruses in Saudi Arabia is proposed to be affected mainly by the presence and mobility of large numbers of foreign workers and the gathering of millions of Muslims in Mecca during the Hajj and Umrah seasons. Knowledge concerning the epidemiology, circulation pattern, and evolutionary kinetics of respiratory viruses in Saudi Arabia are scant, with the available literature being inconsistent. This review summarizes the available data on the epidemiology and evolution of respiratory viruses. The demographic features associated with Middle East respiratory syndrome-related coronavirus infections are specifically analyzed for a better understanding of the epidemiology of this virus. The data support the view that continuous entry and exit of pilgrims and foreign workers with different ethnicities and socioeconomic backgrounds in Saudi Arabia is the most likely vehicle for global dissemination of respiratory viruses and for the emergence of new viruses (or virus variants) capable of greater dissemination.
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http://dx.doi.org/10.1007/s00705-019-04300-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7087236PMC
August 2019

Novel recombinant DNA vaccine candidates for human respiratory syncytial virus: Preclinical evaluation of immunogenicity and protection efficiency.

Hum Vaccin Immunother 2017 07 8;13(7):1586-1597. Epub 2017 Mar 8.

a Department of Botany and Microbiology , College of Science, King Saud University , Riyadh , Saudi Arabia.

The development of safe and potent vaccines for human respiratory syncytial virus (HRSV) is still a challenge for researchers worldwide. DNA-based immunization is currently a promising approach that has been used to generate human vaccines for different age groups. In this study, novel HRSV DNA vaccine candidates were generated and preclinically tested in BALB/c mice. Three different versions of the codon-optimized HRSV fusion (F) gene were individually cloned into the pPOE vector. The new recombinant vectors either express full-length (pPOE-F), secretory (pPOE-TF), or M2 linked (pPOE-FM2) forms of the F protein. Distinctive expression of the F protein was identified in HEp-2 cells transfected with the different recombinant vectors using ELISA and immunofluorescence. Mice immunization verified the potential for recombinant vectors to elicit significant levels of neutralizing antibodies and CD8 T-cell lymphocytes. pPOE-TF showed higher levels of gene expression in cell culture and better induction of the humoral and cellular immune responses. Following virus challenge, mice that had been immunized with the recombinant vectors were able to control virus replication and displayed lower inflammation compared with mice immunized with empty pPOE vector or formalin-inactivated HRSV vaccine. Moreover, pulmonary cytokine profiles of mice immunized with the 3 recombinant vectors were similar to those of the mock infected group. In conclusion, recombinant pPOE vectors are promising HRSV vaccine candidates in terms of their safety, immunogenicity and protective efficiency. These data encourage further evaluation in phase I clinical trials.
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http://dx.doi.org/10.1080/21645515.2017.1295190DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5512761PMC
July 2017

Surface gene variants of hepatitis B Virus in Saudi Patients.

Saudi J Gastroenterol 2016 Mar-Apr;22(2):133-8

Department of Botany and Microbiology, College of Science, King Saud University, Riyadh, Saudi Arabia.

Background/aims: Hepatitis B virus (HBV) continues to be one of the most important viral pathogens in humans. Surface (S) protein is the major HBV antigen that mediates virus attachment and entry and determines the virus subtype. Mutations in S gene, particularly in the "a" determinant, can influence virus detection by ELISA and may generate escape mutants. Since no records have documented the S gene mutations in HBV strains circulating in Saudi Arabia, the current study was designed to study sequence variation of S gene in strains circulating in Saudi Arabia and its correlation with clinical and risk factors.

Patients And Methods: A total of 123 HBV-infected patients were recruited for this study. Clinical and biochemical parameters, serological markers, and viral load were determined in all patients. The entire S gene sequence of samples with viral load exceeding 2000 IU/mL was retrieved and exploited in sequence and phylogenetic analysis.

Results: A total of 48 mutations (21 unique) were recorded in viral strains in Saudi Arabia, among which 24 (11 unique) changed their respective amino acids. Two amino acid changes were recorded in "a" determinant, including F130L and S135F with no evidence of the vaccine escape mutant G145R in any of the samples. No specific relationship was recognized between the mutation/amino acid change record of HBsAg in strains in Saudi Arabia and clinical or laboratory data. Phylogenetic analysis categorized HBV viral strains in Saudi Arabia as members of subgenotypes D1 and D3.

Conclusion: The present report is the first that describes mutation analysis of HBsAg in strains in Saudi Arabia on both nucleotide and amino acid levels. Different substitutions, particularly in major hydrophilic region, may have a potential influence on disease diagnosis, vaccination strategy, and antiviral chemotherapy.
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http://dx.doi.org/10.4103/1319-3767.167186DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4817297PMC
January 2017

Human Respiratory Syncytial Virus: Role of Innate Immunity in Clearance and Disease Progression.

Viral Immunol 2016 Jan-Feb;29(1):11-26. Epub 2015 Dec 17.

Department of Botany and Microbiology, King Saud University , Riyadh, Saudi Arabia .

Human respiratory syncytial virus (HRSV) infections have worldwide records. The virus is responsible for bronchiolitis, pneumonia, and asthma in humans of different age groups. Premature infants, young children, and immunocompromised individuals are prone to severe HRSV infection that may lead to death. Based on worldwide estimations, millions of cases were reported in both developed and developing countries. In fact, HRSV symptoms develop mainly as a result of host immune response. Due to inability to establish long lasting adaptive immunity, HRSV infection is recurrent and hence impairs vaccine development. Once HRSV attached to the airway epithelia, interaction with the host innate immune components starts. HRSV interaction with pulmonary innate defenses is crucial in determining the disease outcome. Infection of alveolar epithelial cells triggers a cascade of events that lead to recruitment and activation of leukocyte populations. HRSV clearance is mediated by a number of innate leukocytes, including macrophages, natural killer cells, eosinophils, dendritic cells, and neutrophils. Regulation of these cells is mediated by cytokines, chemokines, and other immune mediators. Although the innate immune system helps to clear HRSV infection, it participates in disease progression such as bronchiolitis and asthma. Resolving the mechanisms by which HRSV induces pathogenesis, different possible interactions between the virus and immune components, and immune cells interplay are essential for developing new effective vaccines. Therefore, the current review focuses on how the pulmonary innate defenses mediate HRSV clearance and to what extent they participate in disease progression. In addition, immune responses associated with HRSV vaccines will be discussed.
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http://dx.doi.org/10.1089/vim.2015.0098DOI Listing
October 2016

Characterization of H5N1 influenza A virus that caused the first highly pathogenic avian influenza outbreak in Saudi Arabia.

J Infect Dev Ctries 2015 Nov 30;9(11):1210-9. Epub 2015 Nov 30.

Research Center, King Faisal Specialist Hospital and Research Center, Riyadh, Saudi Arabia.

Introduction: Saudi Arabia (SA) experienced a highly pathogenic avian influenza (HPAI) H5N1 outbreak in domesticated birds in 2007.

Methodology: Forty-three hemagglutinin (HA) and 41 neuraminidase (NA) genes of HPAI H5N1 viruses were sequenced and phylogenetic analyses of completely sequenced genes were performed to compare with other viral HA and NA gene sequences available in the public databases.

Results: Molecular characterization of the H5N1 viruses revealed two genetically distinct clades, 2.2.2 and 2.3.1, of H5N1 viruses circulating in the area. Amino acid sequence analysis of the HA gene indicated that the virus from 2.2.2 contained the sequence SPQGERRRK-R/G at the cleavage site, while the virus from 2.3.1 contained the sequence SPQRERRRK-R/G. Additionally, a few mutations with amino acid substitutions such as M226I at N-link glycosylation site were identified in two of these isolates. Amino acid sequence of the NA gene showed a 20-amino-acid deletion in the NA stalk region, required for enhanced virulence of influenza viruses and its adaptation from wild birds to domestic chickens. As close contact between humans and birds is unavoidable, there is a need for a thorough understanding of the virus epidemiology, factors affecting the spread of the virus, and molecular characterization such as phylogeny and substitution rates of H5N1 viruses circulating in the region.

Conclusion: Two genetically distinct clades were found to be circulating in the country, which could likely result in recombination and emergence of more virulent viral strains. These findings could be helpful for the authorities devising control measures against these viruses.
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http://dx.doi.org/10.3855/jidc.6546DOI Listing
November 2015

Epidemiology of 11 respiratory RNA viruses in a cohort of hospitalized children in Riyadh, Saudi Arabia.

J Med Virol 2016 Jun 1;88(6):1086-91. Epub 2015 Dec 1.

Department of Botany and Microbiology, College of Science, King Saud University, Riyadh, Saudi Arabia.

Respiratory tract infections are a principal cause of illness and mortality in children worldwide and mostly caused by viruses. In this study, the epidemiology of 11 respiratory RNA viruses was investigated in a cohort of hospitalized children at a tertiary referral center in Riyadh from February 2008 to March 2009 using conventional and real-time monoplex RT-PCR assays. Among 174 nasopharyngeal aspirates, respiratory syncytial virus (RSV) was detected in 39 samples (22.41%), influenza A virus in 34 (19.54%), metapneumovirus (MPV) in 19 (10.92%), coronaviruses in 14 (8.05%), and parainfluenza viruses (PIVs) in 11 (6.32%). RSV, PIVs and coronaviruses were most prevalent in infants less than 6 months old, whereas MPV and influenza A virus were more prominent in children aged 7-24 and 25-60 months, respectively. The majority of the viruses were identified during winter with two peaks observed in March 2008 and January 2009. The presented data warrants further investigation to understand the epidemiology of respiratory viruses in Saudi Arabia on spatial and temporal basis.
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http://dx.doi.org/10.1002/jmv.24435DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7167021PMC
June 2016

Human Cytokinome Analysis for Interferon Response.

J Virol 2015 Jul 29;89(14):7108-19. Epub 2015 Apr 29.

Molecular BioMedicine Program and Department of Microbiology, King Faisal Specialist Hospital and Research Centre, Riyadh, Saudi Arabia

Unlabelled: Cytokines are a group of small secreted proteins that mediate a diverse range of immune and nonimmune responses to inflammatory and microbial stimuli. Only a few of these cytokines mount an antiviral response, including type I, II, and III interferons (IFNs). During viral infections and under inflammatory conditions, a number of cytokines and chemokines are coproduced with IFN; however, no systematic study exists on the interactions of the cytokine repertoire with the IFN response. Here, we performed the largest cytokine and chemokine screen (the human cytokinome, with >240 members) to investigate their modulation of type I and type II IFN responses in a cell line model. We evaluated the cytokine activities in both IFN-stimulated response element (ISRE) and IFN-γ activation sequence (GAS) reporter systems. Several cytokine clusters that augment either or both ISRE- and GAS-mediated responses to IFNs were derived from the screen. We identified novel modulators of IFN response-betacellulin (BTC), interleukin 11 (IL-11), and IL-17F-that caused time-dependent induction of the IFN response. The ability to induce endogenous IFN-β and IFN-stimulated genes varies among these cytokines and was largely dependent on Stat1, as assessed by Stat1 mutant fibroblasts. Certain cytokines appear to augment the IFN-β response through the NF-κB pathway. The novel IFN-like cytokines augmented the antiviral activity of IFN-α against several RNA viruses, including encephalomyocarditis virus, vesicular stomatitis virus, and influenza virus, in susceptible cell lines. Overall, the study represents a large-scale analysis of cytokines for enhancing the IFN response and identified cytokines capable of enhancing Stat1, IFN-induced gene expression, and antiviral activities.

Importance: Innate immunity to viruses is an early defense system to ward off viruses. One mediator is interferon (IFN), which activates a cascade of biochemical events that aim to control the virus life cycle. In our work, we examined more than 200 cytokines, soluble mediators produced within the body as a result of infection, for the ability to enhance IFN action. We identified enhanced interactions with specific IFNs and cytokines. We also revealed that betacellulin, IL-17, and IL-11 cytokines have the novel property of enhancing the antiviral action of IFN against several viruses. These results demonstrate that the human genome codes for previously unknown proteins with unrelated functions that can augment the innate immunity to viruses. Knowing these interactions not only helps our understanding of immunity to viruses and emerging diseases, but can also lead to devising possible new therapeutics by enhancing the mediator of antiviral action itself, IFN.
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http://dx.doi.org/10.1128/JVI.03729-14DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4473565PMC
July 2015

Hemagglutinin-neuraminidase gene sequence-based reclassification of human parainfluenza virus 3 variants.

Authors:
Fahad N Almajhdi

Intervirology 2015 13;58(1):35-40. Epub 2015 Jan 13.

Department of Botany and Microbiology, College of Science, King Saud University, Riyadh, Saudi Arabia.

The most comprehensive phylogenetic classification of human parainfluenza virus 3 (HPIV-3) was recently developed [PLoS One 2012;7:e43893]. This classification included three distinct clusters (A, B and C) with subdivision of cluster C into four subclusters (C1-4). In the present report, the classification of HPIV-3 was refined by inclusion of 27 overlooked beside newly characterized Saudi variants. The new phylogram was developed and included the same clusters described before, in which cluster A remained unchanged and cluster B contained more recent isolates. The organization of cluster C was altered through inclusion of a new subcluster (C5), subdivision of C1 into two lineages C1a and C1b and subdivision of C3 into three lineages C3a, C3b and C3c. The majority of Saudi variants were classified as members of subcluster C1b, whereas only one variant was placed in each of subclusters C2 and C5. This study illustrates an up-to-date phylogenetic classification of HPIV-3 variants.
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http://dx.doi.org/10.1159/000369208DOI Listing
August 2015

Design of a highly effective therapeutic HPV16 E6/E7-specific DNA vaccine: optimization by different ways of sequence rearrangements (shuffling).

PLoS One 2014 25;9(11):e113461. Epub 2014 Nov 25.

Department of Chemistry and Biotechnology, Aachen University of Applied Sciences, Jülich, Germany.

Persistent infection with the high-risk Human Papillomavirus type 16 (HPV 16) is the causative event for the development of cervical cancer and other malignant tumors of the anogenital tract and of the head and neck. Despite many attempts to develop therapeutic vaccines no candidate has entered late clinical trials. An interesting approach is a DNA based vaccine encompassing the nucleotide sequence of the E6 and E7 viral oncoproteins. Because both proteins are consistently expressed in HPV infected cells they represent excellent targets for immune therapy. Here we report the development of 8 DNA vaccine candidates consisting of differently rearranged HPV-16 E6 and E7 sequences within one molecule providing all naturally occurring epitopes but supposedly lacking transforming activity. The HPV sequences were fused to the J-domain and the SV40 enhancer in order to increase immune responses. We demonstrate that one out of the 8 vaccine candidates induces very strong cellular E6- and E7- specific cellular immune responses in mice and, as shown in regression experiments, efficiently controls growth of HPV 16 positive syngeneic tumors. This data demonstrates the potential of this vaccine candidate to control persistent HPV 16 infection that may lead to malignant disease. It also suggests that different sequence rearrangements influence the immunogenecity by an as yet unknown mechanism.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0113461PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4244082PMC
August 2015

Human papillomavirus (HPV) type 16 E7 protein bodies cause tumour regression in mice.

BMC Cancer 2014 May 24;14:367. Epub 2014 May 24.

Department of Molecular and Cell Biology, University of Cape Town, Private Bag X3, Cape Town, Rondebosch 7700, South Africa.

Background: Human papillomaviruses (HPV) are the causative agents of cervical cancer in women, which results in over 250 000 deaths per year. Presently there are two prophylactic vaccines on the market, protecting against the two most common high-risk HPV types 16 and 18. These vaccines remain very expensive and are not generally affordable in developing countries where they are needed most. Additionally, there remains a need to treat women that are already infected with HPV, and who have high-grade lesions or cervical cancer.

Methods: In this paper, we characterize the immunogenicity of a therapeutic vaccine that targets the E7 protein of the most prevalent high-risk HPV - type 16 - the gene which has previously been shown to be effective in DNA vaccine trials in mice. The synthetic shuffled HPV-16 E7 (16E7SH) has lost its transforming properties but retains all naturally-occurring CTL epitopes. This was genetically fused to Zera®, a self-assembly domain of the maize γ-zein able to induce the accumulation of recombinant proteins into protein bodies (PBs), within the endoplasmic reticulum in a number of expression systems.

Results: High-level expression of the HPV 16E7SH protein fused to Zera® in plants was achieved, and the protein bodies could be easily and cost-effectively purified. Immune responses comparable to the 16E7SH DNA vaccine were demonstrated in the murine model, with the protein vaccine successfully inducing a specific humoral as well as cell mediated immune response, and mediating tumour regression.

Conclusions: The fusion of 16E7SH to the Zera® peptide was found to enhance the immune responses, presumably by means of a more efficient antigen presentation via the protein bodies. Interestingly, simply mixing the free PBs and 16E7SH also enhanced immune responses, indicating an adjuvant activity for the Zera® PBs.
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http://dx.doi.org/10.1186/1471-2407-14-367DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4041048PMC
May 2014

Single-walled carbon nanotubes induce cytotoxicity and DNA damage via reactive oxygen species in human hepatocarcinoma cells.

In Vitro Cell Dev Biol Anim 2014 Sep 2;50(8):714-22. Epub 2014 May 2.

Department of Zoology, College of Science, King Saud University, Box 2454, 11451, Riyadh, Saudi Arabia.

Carbon nanotubes (CNTs) are gradually used in various areas including drug delivery, nanomedicine, biosensors, and electronics. The current study aimed to explore the DNA damage and cytotoxicity due to single-walled carbon nanotubes (SWCNTs) on human hepatocarcinoma cells (HepG2). Cellular proliferative assay showed the SWCNTs to exhibit a significant cell death in a dose- and time-dependent manner. However, SWCNTs induced significant intracellular reactive oxygen species (ROS) production and elevated lipid peroxidation, catalase, and superoxide dismutase in the HepG2 cells. SWCNTs also induced significant decrease in GSH and increase caspase-3 activity in HepG2 cells. DNA fragmentation analysis using the alkaline single-cell gel electrophoresis showed that the SWCNTs cause genotoxicity in a dose- and time-dependent manner. Therefore, the study points towards the capability of the SWCNTs to induce oxidative stress resulting cytotoxicity and genomic instability. This study warrants more careful assessment of SWCNTs before their industrial applications.
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http://dx.doi.org/10.1007/s11626-014-9760-3DOI Listing
September 2014

In-vitro anticancer and antimicrobial activities of PLGA/silver nanofiber composites prepared by electrospinning.

J Mater Sci Mater Med 2014 Apr 31;25(4):1045-53. Epub 2013 Dec 31.

Department of Botany and Microbiology, College of Science, King Saud University, Riyadh, Saudi Arabia.

In the present work, a series of 0, 1 and 7 wt% silver nano-particles (Ag NPs) incorporated poly lactic-co-glycolic acid (PLGA) nano-fibers were synthesized by the electrospinning process. The PLGA/Ag nano-fibers sheets were characterized using SEM, TEM and DSC analyses. The three synthesized PLGA/silver nano-fiber composites were screened for anticancer activity against liver cancer cell line using MTT and LDH assays. The anticancer activity of PLGA nano-fibers showed a remarkable improvement due to increasing the concentration of the Ag NPs. In addition to the given result, PLGA nano-fibers did not show any cytotoxic effect. However, PLGA nano-fibers that contain 1 % nano silver showed anticancer activity of 8.8 %, through increasing the concentration of the nano silver to 7 % onto PLGA nano-fibers, the anticancer activity was enhanced to a 67.6 %. Furthermore, the antibacterial activities of these three nano-fibers, against the five bacteria strains namely; E.coli o157:H7 ATCC 51659, Staphylococcus aureus ATCC 13565, Bacillus cereus EMCC 1080, Listeria monocytogenes EMCC 1875 and Salmonella typhimurium ATCC25566 using the disc diffusion method, were evaluated. Sample with an enhanced inhibitory effect was PLGA/Ag NPs (7 %) which inhibited all strains (inhibition zone diameter 10 mm); PLGA/Ag NPs (1 %) sample inhibited only one strain (B. cereus) with zone diameter 8 mm. The PLGA nano-fiber sample has not shown any antimicrobial activity. Based on the anticancer as well as the antimicrobial results in this study, it can be postulated that: PLGA nanofibers containing 7 % nano silver are suitable as anticancer- and antibiotic-drug delivery systems, as they will increase the anticancer as well as the antibiotic drug potency without cytotoxicity effect on the normal cells. These findings also suggest that Ag NPs, of the size (5-10 nm) evaluated in the present study, are appropriate for therapeutic application from a safety standpoint.
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http://dx.doi.org/10.1007/s10856-013-5131-yDOI Listing
April 2014

Group B strains of human respiratory syncytial virus in Saudi Arabia: molecular and phylogenetic analysis.

Virus Genes 2014 Apr 27;48(2):252-9. Epub 2013 Dec 27.

Department of Botany and Microbiology, College of Science, King Saud University, P.O. Box 2455, Riyadh, 11451, Saudi Arabia,

The genetic variability and circulation pattern of human respiratory syncytial virus group B (HRSV-B) strains, identified in Riyadh during the winters of 2008 and 2009, were evaluated by partial sequencing of the attachment (G) protein gene. The second hypervariable region (HVR-2) of G gene was amplified by RT-PCR, sequenced and compared to representatives of different HRSV-B genotypes. Sequence and phylogenetic analysis revealed that all Saudi strains belonged to the genotype BA, which is characterized by 60-nucleotide duplication at HVR-2. Only strains of 2008 were clustered with subgroup BA-IV, while those isolated at 2009 were clustered among the most recent subgroups (particularly BA-X and CB-B). Amino acid sequence analysis demonstrated 18 amino acid substitutions in Saudi HRSV-B strains; among which five are specific for individual strains. Furthermore, two potential N-glycosylation sites at residues 230 and 296 were identified for all Saudi strains, and an additional site at amino acid 273 was found only in Riyadh 28/2008 strain. O-glycosylation was predicted in 42-43 sites, where the majority (no = 38) are highly conserved among Saudi strains. The average ratio between non-synonymous and synonymous mutations (ω) implied stabilizing selection pressure on G protein, with evidences of positive selection on certain Saudi strains. This report provides preliminary data on the circulation pattern and molecular characteristics of HRSV-B strains circulating in Saudi Arabia.
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http://dx.doi.org/10.1007/s11262-013-1030-zDOI Listing
April 2014

Hemagglutinin and neuraminidase genes of influenza B viruses circulating in Riyadh, Saudi Arabia during 2010-2011: evolution and sequence analysis.

J Med Virol 2014 Jun 22;86(6):1003-16. Epub 2013 Oct 22.

Center of Excellence in Biotechnology Research, King Saud University, Riyadh, Saudi Arabia.

Influenza viruses are known as continuing threats to human public health every year worldwide. Evolutionary dynamics of influenza B viruses in humans are in a unique progression having two lineages; B/Yam and B/Vic-like viruses, which are circulating simultaneously worldwide. There is a considerable lack of data on influenza B viruses circulating in Saudi Arabia. During the winter-spring season of 2010-2011, 80 nasopharyngeal aspirates were collected from hospitalized patients with flu-like symptoms in Riyadh. Screening of samples by one-step RT-PCR identified three (3.8%) influenza B viruses. Sequencing of hemagglutinin (HA) and neuraminidase (NA) genes was performed to analyze influenza B viruses circulating in Riyadh as compared to the globally circulating strains. Several common and six unique amino acid substitutions were observed for both HA and NA genes of influenza B Saudi strains. Three unique substitutions (T182A, D196N, and K254R) were identified in HA gene of the B/Yam-like Riyadh strains. In NA gene, a unique common substitution (D53G) was found in all Riyadh strains, while two unique substitutions (L38P, G233R) were recognized only in B/Vic-like Riyadh strains. Riyadh strains were also found to contain N-glycosylation site in HA gene of both B/Vic and B/Yam lineages at positions 197-199 (NET) and 196-198 (NNK/DNK), respectively. The significance of these mutations on the antigenicity of both lineages is discussed herein. The unique changes observed in HA and NA genes of influenza B Riyadh strains support strongly the need for continuous surveillance and monitoring of new evolving strains that might pose threat to the Saudi community.
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http://dx.doi.org/10.1002/jmv.23819DOI Listing
June 2014

Report on influenza A and B viruses: their coinfection in a Saudi leukemia patient.

Biomed Res Int 2013 2;2013:290609. Epub 2013 Sep 2.

Department of Botany and Microbiology, College of Science, King Saud University, P.O. Box 2455, Riyadh 11451, Saudi Arabia ; Center of Excellence in Biotechnology Research, King Saud University, P.O. Box 2460, Riyadh 11451, Saudi Arabia.

Purpose: Influenza A and B viruses are the leading cause of respiratory infections in children worldwide, particularly in developing countries. There is a lack of data on coinfection of influenza A and B viruses circulating in Saudi Arabia. In this study, we aimed to identify the circulation of influenza viruses that contribute to respiratory tract infections in Saudi children.

Methods: We collected 80 nasopharyngeal aspirates (NPAs) from hospitalized children with acute respiratory illness (ARI) at Riyadh during the period extended from October 2010 till April 2011. Samples were tested for the common respiratory viruses including influenza viruses by RT-PCR.

Results: Overall, 6 samples were found positive for influenza A and/or B viruses. Among these positive clinical samples, only one collected sample from a female one-year-old immunocompromised child with leukemia showed a coinfection with influenza A and B viruses. In present study coinfection was confirmed by inoculation of the clinical specimen in specific pathogenfree embryonating chicken eggs and identification of the virus isolates by hemagglutination and one-step RT-PCR.

Conclusion: This study opens the scene for studying the role of influenza virus's coinfection in disease severity and virus evolution. Further studies are required to better understand the clinical importance of viral coinfection.
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http://dx.doi.org/10.1155/2013/290609DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3775437PMC
April 2014

Effect of gamma radiation and accelerated aging on the mechanical and thermal behavior of HDPE/HA nano-composites for bone tissue regeneration.

Biomed Eng Online 2013 Sep 24;12:95. Epub 2013 Sep 24.

Biomedical Engineering Department, Helwan University, Faculty of Engineering, Helwan, Egypt.

Background: The replacement of hard tissues demands biocompatible and sometimes bioactive materials with properties similar to those of bone. Nano-composites made of biocompatible polymers and bioactive inorganic nano particles such as HDPE/HA have attracted attention as permanent bone substitutes due to their excellent mechanical properties and biocompatibility.

Method: The HDPE/HA nano-composite is prepared using melt blending at different HA loading ratios. For evaluation of the degradation by radiation, gamma rays of 35 kGy, and 70 kGy were used to irradiate the samples at room temperature in vacuum. The effects of accelerated ageing after gamma irradiation on morphological, mechanical and thermal properties of HDPE/HA nano-composites were measured.

Results: In Vitro test results showed that the HDPE and all HDPE/HA nano-composites do not exhibit any cytotoxicity to WISH cell line. The results also indicated that the tensile properties of HDPE/HA nano-composite increased with increasing the HA content except fracture strain decreased. The dynamic mechanical analysis (DMA) results showed that the storage and loss moduli increased with increasing the HA ratio and the testing frequency. Finally, it is remarked that all properties of HDPE/HA is dependent on the irradiation dose and accelerated aging.

Conclusion: Based on the experimental results, it is found that the addition of 10%, 20% and 30% HA increases the HDPE stiffness by 23%, 44 and 59% respectively. At the same time, the G' increased from 2.25E11 MPa for neat HDPE to 4.7E11 MPa when 30% HA was added to the polymer matrix. Also, significant improvements in these properties have been observed due to irradiation. Finally, the overall properties of HDPE and its nano-composite properties significantly decreased due to aging and should be taken into consideration in the design of bone substitutes. It is attributed that the developed HDPE/HA nano-composites could be a good alternative material for bone tissue regeneration due to their acceptable properties.
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http://dx.doi.org/10.1186/1475-925X-12-95DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3850061PMC
September 2013

Genetic diversity in the G protein gene of group A human respiratory syncytial viruses circulating in Riyadh, Saudi Arabia.

Arch Virol 2014 Jan 25;159(1):73-81. Epub 2013 Jul 25.

Department of Botany and Microbiology, College of Science, King Saud University, P.O. Box 2455, Riyadh, 11451, Saudi Arabia,

Human respiratory syncytial virus (HRSV) is a frequent cause of hospitalization and mortality in children worldwide. The molecular epidemiology and circulation pattern of HRSV in Saudi Arabia is mostly uncharted. In the current study, the genetic variability and phylogenetic relationships of HRSV type A strains circulating in Riyadh Province were explored. Nasopharyngeal aspirates were collected from hospitalized children with acute respiratory symptoms during the winter-spring seasons of 2007/08 and 2008/09. Among 175 samples analyzed, 39 (22.3 %) were positive for HRSV by one-step RT-PCR (59 % type A and 41 % type B). Propagation of positive samples in HEp-2 cells permitted the recovery of the first Saudi HRSV isolates. Genetic variability among Saudi HRSV-A strains was evaluated by sequence analysis of the complete attachment (G) protein gene. The nucleotide sequence was compared to representatives of the previously identified HRSV-A genotypes. Sequence and phylogenetic analysis showed that the strains examined in this study were very closely related at both the nucleotide and amino acid level, and all of them are clustered in the GA2 genotype (and mostly belonged to the NA-1 subtype). A total of 23 mutation sites, 14 of which resulted in an amino acid change, were recorded only in Saudi strains. This is the first report on genetic diversity of HRSV-A strains in Saudi Arabia. Further analysis of strains on a geographical and temporal basis is needed to fully understand HRSV-A circulation patterns in Saudi Arabia.
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http://dx.doi.org/10.1007/s00705-013-1792-6DOI Listing
January 2014

Differential expression of transforming growth factor-β1 and HBx enhances hepatitis B virus replication and augments host immune cytokines and chemokines.

Ann Hepatol 2013 May-Jun;12(3):408-15

Department of Botany and Microbiology College of Science, King Saud University, Riyadh, Saudi Arabia.

Background/aims: This study investigated how HBV replication and host immune response are effected by reduced expression of TGF-β1 and HBx.

Material And Methods: Short interfering RNA (siRNA) knockdown technology has been used to examine the role of TGF-β1 in hepatitis B virus replication. The siTGF-β1 has been transfected along with 1.3mer HBV x-null to investigate the knockdown effect of TGF-β1 on HBV replication and host immune factors.

Results: In this study, we found that diminished expression of TGF-β1 and increased expression of HBx enhances HBV replication several folds. The differential expression of TGF-β1 and HBx also stimulated transcriptional viral replicative intermediate (pgRNA) and secretion of core and 'e' antigen at translational level. Consequently, several cytokines such as IL-2, IL-8 and chemokine monocyte- chemoattractant protein (MCP-1) were increased significantly in response to stimulation of HBV replication. In contrast, TNF-α and RANTES mRNA expression increased insignificantly in response to enhanced HBV replication.

Conclusions: We concluded that reduced expression of TGF-β1 together with HBx expression stimulate HBV replication and immune response, although the underlying mechanism of stimulation most likely differs.
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October 2013

Single nucleotide polymorphisms in CXCR1 gene and its association with hepatitis B infected patients in Saudi Arabia.

Ann Hepatol 2013 Mar-Apr;12(2):220-7

Department of Botany and Microbiology, College of Science, King Saud University, Riyadh, Saudi Arabia.

Background/aim: This study aims to investigate whether the SNPs of CXCR1 gene, could predict the likelihood of viral persistence and/or disease progression.

Material And Methods: We investigated the association of two different SNPs (rs2234671, and rs142978743) in 598 normal healthy controls and 662 HBV patients from a Saudi ethnic population. The HBV patients were categorized into inactive carriers (n = 428), active carriers (n = 162), cirrhosis (n = 54) and Cirrhosis-HCC (n = 18) sub-groups. Genetic variants in CXCR1 were determined by polymerase chain reaction (PCR)-based DNA direct sequencing.

Results: The frequency of the risk allele 'C' for the SNP, rs2234671 was found to be insignificant when the patient group was compared to the uninfected control group, however, a significant distribution of the allele 'C' of rs2234671 was observed among active HBV carriers + cirrhosis + cirrhosis - HCC vs. inactive HBV carriers with an OR = 1.631 (95% C.I. 1.016-2.616) and p = 0.032. However, no significant association was observed for rs142978743 when the distribution of risk allele was analyzed among the different patient groups (i.e. inactive carriers, active carriers, cirrhosis and HCC). Furthermore, the most common haplotype, Haplo-1 (AG), was found to have an insignificant frequency distribution between HBV cases and controls, while the same haplotype was found to be significantly distributed when active carriers + cirrhosis + cirrhosis - HCC patients were compared to inactive HBV carriers with a frequency of 0.938 and p = 0.0315. Haplo-2 (AC) was also found to be significantly associated with a frequency of 0.058 and p = 0.0163.

Conclusion: The CXCR1 polymorphism, rs2234671 was found to be associated with chronic HBV infection and may play a role in disease activity.
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October 2013

Human parainfluenza virus type 2 hemagglutinin-neuramindase gene: sequence and phylogenetic analysis of the Saudi strain Riyadh 105/2009.

Virol J 2012 Dec 22;9:316. Epub 2012 Dec 22.

Department of Botany and Microbiology, College of Science, King Saud University, P,O, Box, 2455, Riyadh, 11451, Kingdom of Saudi Arabia.

Background: Although human parainfluenza type 2 (HPIV-2) virus is an important respiratory pathogen, a little is known about strains circulating in Saudi Arabia.

Findings: Among 180 nasopharyngeal aspirates collected from suspected cases in Riyadh, only one sample (0.56%) was confirmed HPIV-2 positive by nested RT-PCR. The sample that was designated Riyadh 105/2009 was used for sequencing and phylogenetic analysis of the most variable virus gene; the haemagglutinin-neuramindase (HN). Comparison of HN gene of Riyadh 105/2009 strain and the relevant sequences available in GenBank revealed a strong relationship with Oklahoma-94-2009 strain. Phylogenetic analysis indicated four different clusters of HPIV-2 strains (G1-4). Twenty-three amino acid substitutions were recorded for Riyadh 105/2009, from which four are unique. The majority of substitutions (n=18) had changed their amino acids characteristics. By analyzing the effect of the recorded substitutions on the protein function using SIFT program, only two located at positions 360 and 571 were predicted to be deleterious.

Conclusions: The presented changes of Riyadh 105/2009 strain may possess potential effect on the protein structure and/or function level. This is the first report that describes partial characterization of Saudi HPIV-2 strain.
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http://dx.doi.org/10.1186/1743-422X-9-316DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3547708PMC
December 2012

Toll-like receptor 3 polymorphism and its association with hepatitis B virus infection in Saudi Arabian patients.

J Med Virol 2012 Sep;84(9):1353-9

Department of Infection and Immunity, Research Center, King Faisal Specialist Hospital & Research Center, Riyadh, Saudi Arabia.

Hepatitis B virus (HBV) is the major causative agent of chronic liver complications including cirrhosis and hepatocellular carcinoma (HCC). Individuals infected with HBV show a wide spectrum of disease manifestations ranging from asymptomatic carriers to HCC. TLR3 is part of the innate immune system that recognizes double-stranded RNA (dsRNA) and provides early immune response to exogenous antigens. The genetic polymorphisms such as single nucleotide polymorphisms (SNPs) in the TLR3 could be considered as factors for the susceptibility to viral pathogens including HBV. Due to lack of knowledge on the role of TLR3 polymorphisms in HBV infection, this study investigated the distribution of nine SNPs in the TLR3 gene and its association with Saudi Arabian patients infected with HBV. A total of 707 patients and 600 uninfected controls were examined for different parameters including the nine SNPs (rs5743311, rs5743312, rs1879026, rs5743313, rs5743314, rs5743315, rs111611328, rs78726532 and a newly identified SNP located at position 184322913 of chr4). The association analysis confirmed that only one SNP, rs1879026 (G/T), showed a significant difference (P = 0.0480; OR = 0.809, 95% CI = 0.655-0.999) in the distribution between HBV carriers and uninfected controls. While, the rest of the SNPs showed no significant association with regards to HBV infection or in the progression to cirrhosis of the liver and HCC. Furthermore, haplotype analysis revealed that one haplotype GCGA (rs1879026, rs5743313, rs5743314, and rs5743315, respectively), was associated significantly with HBV infection in this population. These findings indicate that genetic variations in the TLR3 gene could affect the outcome of HBV infection among Saudis.
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http://dx.doi.org/10.1002/jmv.23271DOI Listing
September 2012

Molecular characterization and phylogenetic analysis of human parainfluenza virus type 3 isolated from Saudi Arabia.

J Med Virol 2012 Aug;84(8):1304-11

Department of Botany and Microbiology, College of Science, King Saud University, Riyadh, Saudi Arabia.

Human parainfluenza virus 3 (HPIV-3) is a leading cause of respiratory disease in children worldwide. Previous sequence analyses of the entire virus genome, among different HPIV-3 strains, demonstrated that HN is the most variable gene. There is a dearth of data on HPIV-3 strains circulating in Saudi Arabia. In this report, HPIV-3 was screened in nasopharyngeal aspirates collected from hospitalized children with acute respiratory disease during two successive seasons (2007/08 and 2008/09) using nested RT-PCR. Out of 73 samples collected during 2007/08, seven (9.59%) were positive; while 3 out of 107 samples collected during 2008/09 (2.8%) were positive. Virus isolation in cell culture was successful using HEp2, but not Vero cells. The identity of the isolated viruses was confirmed using immunofluorescence and neutralization assays. To elucidate the genetic characteristics and phylogeny of Saudi HPIV-3 strains, the complete HN gene sequence of two selected Saudi strains was analyzed in comparison to 20 strains isolated by others from different countries worldwide. Both strains showed the highest degree of sequence homology with Indian strains, followed by Chinese and most Japanese strains. Phylogenetic analysis confirmed that these strains fell into a distinct Asian lineage. This study is the first in Saudi Arabia to recover HPIV-3 isolates of confirmed identity, and to generate sequence data that may help in understanding virus diversity and evolution.
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http://dx.doi.org/10.1002/jmv.23326DOI Listing
August 2012

Green fluorescent protein reporter system with transcriptional sequence heterogeneity for monitoring the interferon response.

J Virol 2011 Sep 13;85(18):9268-75. Epub 2011 Jul 13.

P3354, MBC-03, Riyadh 11211, Saudi Arabia.

The interferon (IFN) response is initiated by a variety of triggers, including viruses and foreign RNA, and involves several receptors and intracellular mediators. Although there are common cis-acting consensus sequences in the promoters of many genes stimulated during the IFN response, they exhibit core and context heterogeneity that may lead to differential transcriptional activity. We have developed and validated a live cell-based enhanced green fluorescent protein (EGFP) reporter system employing more than a hundred constructs containing multiple viruses and IFN response elements derived from a variety of promoters involved in immunity to viruses. Common and distinct response patterns were observed due to promoter heterogeneity in response to different stimuli, including IFN-α, TLR3-agonist double-stranded RNA, and several viruses. This information should serve as a resource in selecting specific reporters for sensing nonself ligands.
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http://dx.doi.org/10.1128/JVI.00772-11DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3165742PMC
September 2011

Immunological and molecular epidemiological characteristics of acute and fulminant viral hepatitis A.

Virol J 2011 May 23;8:254. Epub 2011 May 23.

PCR Hepatitis Laboratory, Department of Medicine, Maulana Azad Medical College, New Delhi, India.

Background: Hepatitis A virus is an infection of liver; it is hyperendemic in vast areas of the world including India. In most cases it causes an acute self limited illness but rarely fulminant. There is growing concern about change in pattern from asymptomatic childhood infection to an increased incidence of symptomatic disease in the adult population.

Objective: In-depth analysis of immunological, viral quantification and genotype of acute and fulminant hepatitis A virus.

Methods: Serum samples obtained from 1009 cases of suspected acute viral hepatitis was employed for different biochemical and serological examination. RNA was extracted from blood serum, reverse transcribed into cDNA and amplified using nested PCR for viral quantification, sequencing and genotyping. Immunological cell count from freshly collected whole blood was carried out by fluorescence activated cell sorter.

Results: Fulminant hepatitis A was mostly detected with other hepatic viruses. CD8+ T cells count increases in fulminant hepatitis to a significantly high level (P = 0.005) compared to normal healthy control. The immunological helper/suppressor (CD4+/CD8+) ratio of fulminant hepatitis was significantly lower compared to acute cases. The serologically positive patients were confirmed by RT-PCR and total of 72 (69.2%) were quantified and sequenced. The average quantitative viral load of fulminant cases was significantly higher (P < 0.05). There was similar genotypic distribution in both acute and fulminant category, with predominance of genotype IIIA (70%) compared to IA (30%).

Conclusions: Immunological factors in combination with viral load defines the severity of the fulminant hepatitis A. Phylogenetic analysis of acute and fulminant hepatitis A confirmed genotypes IIIA as predominant against IA with no preference of disease severity.
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http://dx.doi.org/10.1186/1743-422X-8-254DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3117845PMC
May 2011