Genetic analysis of human parainfluenza type 2 virus in Riyadh, Saudi Arabia.

The extensive mass gathering of pilgrims from all over the world, as well as the constant flow of foreign workers via country entry crossings, raises the likelihood of respiratory virus outbreaks spreading and evolving in Saudi Arabia. Here, we report the sequence and phylogenetic analysis of the human parainfluenza type-2 (HPIV-2) in nasopharyngeal aspirates (NPAs) collected from Riyadh, Saudi Arabia, from 2020/21 to 2021/22 seasons. RNA was extracted from the clinical samples and subjected to RT-PCR analysis for the detection of IAV and IBV. The full-length HN gene of HPIV-2 was amplified and sequenced. Multiple sequence alignments (both nucleotides and deduced amino acids) were aligned using Clustal W, MegAlign program of Lasergene software, and MEGA 7.0. HPIV-2 was found in (4; 2% of 200) NPAs. Sequence and phylogenetic analysis results showed that indicated a genotype shifting from G3 to G4a with 83% sequence homology 62-M786 from Japan, which was prominent throughout the winter seasons of 2008/09. Multiple amino acid sequence alignment revealed 25 sites of possible difference between G3 genotypes and G4a. A total of twenty- two of these locations were shared by the other G4a genotypes, whereas three positions, 67 V, 175 S, and 377Q, were exclusively shared by G3. Only eight conserved N-glycosylation sites were found at amino acids 6(NLS), 286(NTT), 335(NIT), 388(NNS), 498(NES), 504(NPT), 517(NTT), and 539(NGT) in four Riyadh isolates. Our findings also revealed that the G4a genotype of HPIV-2 predominated in our samples population during the winter seasons of 2020/21 and 2021/22. Further research with a larger sample size covering numerous regions of Saudi Arabia throughout different epidemic seasons is needed to achieve an improved knowledge of HPIV-2 circulation.

Phytoconstituents of Artemisia Annua as potential inhibitors of SARS CoV2 main protease: an in silico study.

BACKGROUND: In November 2019, the world faced a pandemic called SARS-CoV-2, which became a major threat to humans and continues to be. To overcome this, many plants were explored to find a cure. METHODS: Therefore, this research was planned to screen out the active constituents from Artemisia annua that can work against the viral main protease Mpro as this non-structural protein is responsible for the cleavage of replicating enzymes of the virus. Twenty-five biocompounds belonging to different classes namely alpha-pinene, beta-pinene, carvone, myrtenol, quinic acid, caffeic acid, quercetin, rutin, apigenin, chrysoplenetin, arteannunin b, artemisinin, scopoletin, scoparone, artemisinic acid, deoxyartemisnin, artemetin, casticin, sitogluside, beta-sitosterol, dihydroartemisinin, scopolin, artemether, artemotil, artesunate were selected. Virtual screening of these ligands was carried out against drug target Mpro by CB dock. RESULTS: Quercetin, rutin, casticin, chrysoplenetin, apigenin, artemetin, artesunate, sopolin and sito-gluside were found as hit compounds. Further, ADMET screening was conducted which represented Chrysoplenetin as a lead compound. Azithromycin was used as a standard drug. The interactions were studied by PyMol and visualized in LigPlot. Furthermore, the RMSD graph shows fluctuations at various points at the start of simulation in Top1 (Azithromycin) complex system due to structural changes in the helix-coil-helix and beta-turn-beta changes at specific points resulting in increased RMSD with a time frame of 50 ns. But this change remains stable after the extension of simulation time intervals till 100 ns. On other side, the Top2 complex system remains highly stable throughout the time scale. No such structural dynamics were observed bu the ligand attached to the active site residues binds strongly. CONCLUSION: This study facilitates researchers to develop and discover more effective and specific therapeutic agents against SARS-CoV-2 and other viral infections. Finally, chrysoplenetin was identified as a more potent drug candidate to act against the viral main protease, which in the future can be helpful.

Global and local evolutionary dynamics of Dengue virus serotypes 1, 3, and 4.

Evolutionary studies on Dengue virus (DENV) in endemic regions are necessary since naturally occurring mutations may lead to genotypic variations or shifts in serotypes, which may lead to future outbreaks. Our study comprehends the evolutionary dynamics of DENV, using phylogenetic, molecular clock, skyline plots, network, selection pressure, and entropy analyses based on partial CprM gene sequences. We have collected 250 samples, 161 in 2017 and 89 in 2018. Details for the 2017 samples were published in our previous article and that of 2018 are presented in this study. Further evolutionary analysis was carried out using 800 sequences, which incorporate the study and global sequences from GenBank: DENV-1 (n = 240), DENV-3 (n = 374), and DENV-4 (n = 186), identified during 1944-2020, 1956-2020, and 1956-2021, respectively. Genotypes V, III, and I were identified as the predominant genotypes of the DENV-1, DENV-3, and DENV-4 serotypes, respectively. The rate of nucleotide substitution was found highest in DENV-3 (7.90 × 10-4 s/s/y), followed by DENV-4 (6.23 × 10-4 s/s/y) and DENV-1 (5.99 × 10-4 s/s/y). The Bayesian skyline plots of the Indian strains revealed dissimilar patterns amongst the population size of the three serotypes. Network analyses showed the presence of different clusters within the prevalent genotypes. The data presented in this study will assist in supplementing the measures for vaccine development against DENV.

Mutation in the CX3C Motif of G Protein Disrupts Its Interaction with Heparan Sulfate: A Calorimetric, Spectroscopic, and Molecular Docking Study.

Respiratory syncytial virus (RSV) is the leading cause of lower respiratory tract infection in children and infants. To date, there is no effective vaccine available against RSV. Heparan sulfate is a type of glycosaminoglycan that aids in the attachment of the RSV to the host cell membrane via the G protein. In the present study, the effect of amino acid substitution on the structure and stability of the ectodomain G protein was studied. Further, it was investigated whether mutation (K117A) in the CX3C motif of G protein alters the binding with heparan sulfate. The point mutation significantly affects the conformational stability of the G protein. The mutant protein showed a low binding affinity with heparan sulfate as compared to the wild-type G protein, as determined by fluorescence quenching, isothermal titration calorimetry (ITC), and molecular docking studies. The low binding affinity and decreased stability suggested that this mutation may play an important role in prevention of attachment of virion to the host cell receptors. Collectively, this investigation suggests that mutation in the CX3C motif of G protein may likely improve the efficacy and safety of the RSV vaccine.

Induction of Immune Responses and Immune Evasion by Human Bocavirus.

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.

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

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.

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

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 (P = 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; P = 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.

Structural Refolding and Thermal Stability of Myoglobin in the Presence of Mixture of Crowders: Importance of Various Interactions for Protein Stabilization in Crowded Conditions.

The intracellular environment is overcrowded with a range of molecules (small and large), all of which influence protein conformation. As a result, understanding how proteins fold and stay functional in such crowded conditions is essential. Several in vitro experiments have looked into the effects of macromolecular crowding on different proteins. However, there are hardly any reports regarding small molecular crowders used alone and in mixtures to observe their effects on the structure and stability of the proteins, which mimics of the cellular conditions. Here we investigate the effect of different mixtures of crowders, ethylene glycol (EG) and its polymer polyethylene glycol (PEG 400 Da) on the structural and thermal stability of myoglobin (Mb). Our results show that monomer (EG) has no significant effect on the structure of Mb, while the polymer disrupts its structure and decreases its stability. Conversely, the additive effect of crowders showed structural refolding of the protein to some extent. Moreover, the calorimetric binding studies of the protein showed very weak interactions with the mixture of crowders. Usually, we can assume that soft interactions induce structural perturbations while exclusion volume effects stabilize the protein structure; therefore, we hypothesize that under in vivo crowded conditions, both phenomena occur and maintain the stability and function of proteins.

Structural Characterization of Ectodomain G Protein of Respiratory Syncytial Virus and Its Interaction with Heparan Sulfate: Multi-Spectroscopic and In Silico Studies Elucidating Host-Pathogen Interactions.

The global burden of disease caused by a respiratory syncytial virus (RSV) is becoming more widely recognized in young children and adults. Heparan sulfate helps in attaching the virion through G protein with the host cell membrane. In this study, we examined the structural changes of ectodomain G protein (edG) in a wide pH range. The absorbance results revealed that protein maintains its tertiary structure at physiological and highly acidic and alkaline pH. However, visible aggregation of protein was observed in mild acidic pH. The intrinsic fluorescence study shows no significant change in the λmax except at pH 12.0. The ANS fluorescence of edG at pH 2.0 and 3.0 forms an acid-induced molten globule-like state. The denaturation transition curve monitored by fluorescence spectroscopy revealed that urea and GdmCl induced denaturation native (N) ↔ denatured (D) state follows a two-state process. The fluorescence quenching, molecular docking, and 50 ns simulation measurements suggested that heparan sulfate showed excellent binding affinity to edG. Our binding study provides a preliminary insight into the interaction of edG to the host cell membrane via heparan sulfate. This binding can be inhibited using experimental approaches at the molecular level leading to the prevention of effective host-pathogen interaction.

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

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.

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

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.

Human Cytokinome Analysis for Interferon Response.

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-ß 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-ß 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.

Epidemiology of 11 respiratory RNA viruses in a cohort of hospitalized children in 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.

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

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.

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

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.

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

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.

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

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

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.

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

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.

Human bocavirus in Saudi Arabia: Molecular epidemiology and Co-infections among children with acute respiratory tract infections during 2014-2016.

Respiratory tract infections due to a variety of viruses continue to threaten the human population worldwide, particularly in developing countries. Among the responsible viruses, Human Bocavirus (HBoV), a novel discovered virus, causes respiratory tract and gastroenteritis disorders in young children. In Saudi Arabia, data regarding virus molecular epidemiology and evolution and its implication in respiratory tract infection are scarce. In the current study, genetic diversity and circulation pattern of HBoV-1 among hospitalized children due to acute respiratory tract infection (ARTI) during two consecutive years were charted. We found that 3.44% (2014/2015) and 11.25% (2015/2016) of children hospitalized due to ARTI were infected by HBoV-1. We have shown that HBoV was detected year-round without a marked seasonal peak. HBoV-1 also was co-detected with one or multiple other respiratory viruses. The multisequence analysis showed high sequence identity (∼99%) (few point mutation sites) between strains of each genotype and high sequence variation (∼79%) between HBoV-1 and the other 3 genotypes. Phylogenetic analysis showed the clustering of the study's isolates in the HBoV-1 subclade. Our data reveal that genetically conserved HBoV-1 was circulating among admitted children during the course of the study. Further epidemiological and molecular characterization of multiple HBoV-1 strains for different years and from all regions of Saudi Arabia are required to understand and monitor the virus evolution.