Leishmaniasis is a zoonotic and vector-borne infectious disease that is caused by the genus Leishmania belonging to the trypanosomatid family. The protozoan parasite has a digenetic life cycle involving a mammalian host and an insect vector. Leishmaniasisis is a worldwide public health problem falling under the neglected tropical disease category, with over 90 endemic countries, and approximately 1 million new cases and 20,000 deaths annually. Leishmania infection can progress toward the development of species–specific pathologic disorders, ranging in severity from self-healing cutaneous lesions to disseminating muco-cutaneous and fatal visceral manifestations. The severity and the outcome of leishmaniasis is determined by the parasite’s antigenic epitope characteristics, the vector physiology, and most importantly, the immune response and immune status of the host. This review examines the nature of host–pathogen interaction in leishmaniasis, innate and adaptive immune responses, and various strategies that have been employed for vaccine development.

C4b Binding Protein (C4BP) is a major fluid phase inhibitor of the classical and lectin pathways of the complement system. Complement inhibition is achieved by binding to and restricting the role of activated complement component C4b. C4BP functions as a co-factor for factor I in proteolytic inactivation of both soluble and cell surface-bound C4b, thus restricting the formation of the C3-convertase, C4b2a. C4BP also accelerates the natural decay/dissociation of the C3 convertase. This makes C4BP a prime target for exploitation by pathogens to escape complement attack, as seen in Streptococcus pyogenes or Flavivirus. Here, we examined whether C4BP can act on its own in a complement independent manner, against pathogens. C4BP bound H1N1 and H3N2 subtypes of Influenza A Virus (IAV) most likely via multiple sites in Complement Control Protein (CCP) 1-2, 4-5, and 7-8 domains of its α-chain. In addition, C4BP CCP1-2 bound H3N2 better than H1N1. C4BP bound three IAV envelope proteins: Haemagglutinin (~70 kDa), Neuraminidase (~55 kDa), and Matrix protein 1 (~25kDa). C4BP suppressed H1N1 subtype infection into the lung epithelial cell line, A549, while it promoted infection by H3N2 subtype. C4BP restricted viral entry for H1N1 but had the opposite effect on H3N2, as evident from experiments using pseudo-typed viral particles. C4BP downregulated mRNA levels of pro-inflammatory IFN-α, IL-12, and NFκB in the case of H1N1, while it promoted a pro-inflammatory immune response by upregulating IFN- α, TNF-α, RANTES, and IL-6 in the case of H3N2. We conclude that C4BP differentially modulates the efficacy of IAV entry, and hence, replication in a target cell in a strain-dependent manner, and acts as an entry inhibitor for H1N1. Thus, CCP containing complement proteins such as factor H and C4BP may have additional defense roles against IAV that do not rely on the regulation of complement activation.

BACKGROUND: Data on human papillomavirus (HPV) prevalence and survival rates among HPV-infected women are scarce in Saudi Arabia. OBJECTIVE: Assess the prevalence of HPV genotypes in cervical biopsy specimens and its effect on survival over a 10-year timeframe. DESIGN: Retrospective, cross-sectional. SETTINGS: Saudi referral hospital. PATIENTS AND METHODS: Cervical biopsy specimens were collected from women aged 23-95 years old who underwent HPV detection, HPV genotyping, p16 INK4a expression measurement using immunohistochemistry. Kaplan-Meier plots were constructed to analyze overall survival rates. MAIN OUTCOME MEASURES: Survival rate of HPV-positive cervical cancer patients. SAMPLE SIZE: 315 cervical biopsy specimens. RESULTS: HPV was detected in 96 patients (30.4%): 37.3% had cervical cancer; 14.2% cervical intraepithelial neoplasia (CIN) III, 4.1% CIN II, and 17.0% CIN I. A significant association was found between HPV presence and cervical cancer (χ 2 =56.78; P <.001). The expression of p16 INK4a was a significant predictor of survival: women who had p16 INK4a overexpression had poorer survival rates (multivariate Cox regression, hazard ratio, 3.2; 95% CI, 1.1–8.8). In addition, multivariate models with HPV status and cervical cancer diagnosis showed that HPV status was a significant predictor of survival: HPV-positive women had better survival rates than HPV-negative women. CONCLUSION: These findings suggest that implementing cervical and HPV screening programs may decrease cervical cancer rates and improve survival rates of women in Saudi Arabia. LIMITATION: Single center and small sample size. CONFLICT OF INTEREST: None.

An innate immune response is essential to mobilize protective immunity upon the infection of respiratory epithelial cells with influenza A virus (IAV). The response is classified as early (nonspecific effectors), local systematic (effector cells recruitment) and late (antigen to lymphoid organ transport, naive B and T cells recognition, effector cells clonal expansion and differentiation). Virus particles are detected by the host cells as non-self by various sensors that are present on the cell surface, endosomes and cytosol. These sensors are collectively termed as pattern recognition receptors (PRRs). The PRRs distinguish unique molecular signatures known as pathogen-associated molecular pattern, which are present either on the cell surface or within intracellular compartments. PRRs have been classified into five major groups: C-Type Lectin Receptor (CLR), Toll-like receptor (TLR), Nod-like receptor (NLR), Retinoic acid-inducible gene-I-like receptor (RLR), which play a role in innate immunity to IAV infection, and the pyrin and hematopoietic interferon-inducible nuclear (PYHIN) domain protein. Here, we discuss the role of PRRs in cellular infectivity of IAV and highlight the recent progress.

Leishmania species cause a range of moderate to severe diseases affecting cutaneous to visceral regions, thus having a great impact on morbidity and mortality. Leishmania is initially dealt with by human innate immune system immediately after its entry into the body, primarily by the complement system, one of the most potent humoral immune mechanisms that link innate and adaptive immunity. Complement can be activated via all its three pathways on the surface of Leishmania to varying degrees; however, it is considered that the classical pathway is the first to respond due to a lag phase preceding the alternative pathway activation. The present study illustrated that the recognition molecule, C1q bound to soluble leishmania antigens (SLA) derived from L. major via its domains and activates the classical pathway. This was confirmed in complement consumption assay where SLA was able to induce only ~22% haemolysis of sensitized cells. Moreover, this study showed that recombinant individual head regions of C1q chains bind differentially to SLA while interaction studies with substitution mutants revealed that C1q-SLA binding relied on charge-charge interaction. When macrophage-like cell line, THP-1, was challenged with SLA in the presence of human C1q, it downregulated a number of cytokine and chemokine response considerably; the effects were mostly suppressive of Th1 immune response, suggesting that the requirements for potent adjuvants in SLA-mediated vaccine strategies.

Human Papillomavirus (HPV) is the causative agent in the majority of anal, head and neck, oral, oropharyngeal, penile, vaginal, vulvar, and cervical cancers. Cervical cancer is the fourth most common cancer among women worldwide. Of all diagnosed human malignant neoplasms, approximately 4.5% are attributable to HPV, including cervical, anal cancers, vaginal, vulvar, penile, and oropharyngeal cancers. Over 182 HPV types have been identified and sequenced to date however, only certain types of HPV are more frequent in malignant lesions and considered to be a major risk factor in the development of some cancers. Because most HPV infections are transient, and an individual's immunocompetent may clear the infection, HPV infection has received little attention from clinicians, the general public, or policy makers. This lack of attention may underpin a deadly and increasing problem because each newly acquired infection has the potential to persist and become an incurable, lifelong affliction. In addition, no successful treatment of HPV infection currently exists despite the great strides toward understanding the mechanisms underlying HPV pathogenesis. Moreover, ample research has proven that the use of prophylactic vaccines, such as Gardasil and Cervarix, have led to documented progress in decreasing the burden of HPV infection, however not all countries introduced a government-funded National HPV Vaccination Program to protect young men and women. This chapter summarizes the HPV infection, detection and prevention. We also shed light on non-cervical HPV-related cancers, which is rapidly increasing in more developed countries toward cervical cancer.

Cadmium telluride quantum dots (CdTe-QDs) are acquiring great interest in terms of their applications in biomedical sciences. Despite earlier sporadic studies on possible oncogenic roles and anticancer properties of CdTe-QDs, there is limited information regarding the oncogenic potential of CdTe-QDs in cancer progression. Here, we investigated the oncogenic effects of CdTe-QDs on the gene expression profiles of Chang cancer cells. Chang cancer cells were treated with 2 different doses of CdTe-QDs (10 and 25 μg/ml) at different time intervals (6, 12, and 24 h). Functional annotations helped identify the gene expression profile in terms of its biological process, canonical pathways, and gene interaction networks activated. It was found that the gene expression profiles varied in a time and dose-dependent manner. Validation of transcriptional changes of several genes through quantitative PCR showed that several genes upregulated by CdTe-QD exposure were somewhat linked with oncogenesis. CdTe-QD-triggered functional pathways that appear to associate with gene expression, cell proliferation, migration, adhesion, cell-cycle progression, signal transduction, and metabolism. Overall, CdTe-QD exposure led to changes in the gene expression profiles of the Chang cancer cells, highlighting that this nanoparticle can further drive oncogenesis and cancer progression, a finding that indicates the merit of immediate in vivo investigation.

Human papillomavirus (HPV) is a causative agent of cervical and other cancers. Sexually transmitted Infections (STIs) may play a crucial role in HPV persistence, leading to serious complications, including cervical cancer. This study investigated the association of HPV/STI co-infection in cervical samples with cervical dysplasia among women in Saudi Arabia. HPV-positive cervical samples (n = 142) were obtained from previous studies and newly collected samples (n = 209) were obtained from women aged 19-83 years. For HPV detection and genotyping, PCR and Genoflow HPV assay kits were used. STIs were detected using a Genoflow STD array kit. Of 351 samples, 94 (27%) were positive for STIs. Among HPV-positive samples, 36 (25%) were positive for STIs; the most common pathogens were Ureaplasma urealyticum/Ureaplasma parvu (13%) and Mycoplasma hominis (6%). A global significant correlation was detected between HPV and STIs with progression of abnormal cervical cytology (chi(2) = 176, P < 0.0001). Associations between cervical cytology diagnosis and HPV status, STI types (opportunistic and pathogenic), and the presence of Ureaplasma spp., and Mycoplasma hominis were significant (P < 0.05). Our results suggest that additional study in a larger population is warranted to determine the association between HPV/STI co-infection and cervical neoplasia in Saudi women. (C) 2020 The Author(s). Published by Elsevier B.V. on behalf of King Saud University.

The emergence of a novel coronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-COV-2), from Wuhan, China, in December 2019 has challenged many countries. The current pandemic caused by this coronavirus has already negatively affected millions of people and the economies of countries worldwide. However, the challenges faced by Saudi Arabia during the Middle East respiratory syndrome coronavirus (MERS-CoV) epidemic that began in 2012 led to marked improvements in the government's response to the current pandemic. Saudi Arabia is one of largest countries in the Middle East and is home to the holiest Muslim sites. Since the global risk of the virus was declared by the World Health Organization (WHO), Saudi Arabia has taken substantial public health measures to control the spread of the infection. This review reports on the transmission of SARS-COV-2 in Saudi Arabia and the proactive responses taken by the government, comparing the Saudi government's actions and their effects with those of other countries. Although Saudi Arabia is currently experiencing the peak of the pandemic, their early precautionary responses have shortened the period of individual/family isolation, reduced the number of confirmed infections and infection-related fatality rates, and decreased the economic burden of the people and the country compared with other countries in the Middle East and elsewhere.

On 31 December 2019 the Wuhan Health Commission reported a cluster of atypical pneumonia cases that was linked to a wet market in the city of Wuhan, China. The first patients began experiencing symptoms of illness in mid-December 2019. Clinical isolates were found to contain a novel coronavirus with similarity to bat coronaviruses. As of 28 January 2020, there are in excess of 4,500 laboratory-confirmed cases, with > 100 known deaths. As with the SARS-CoV, infections in children appear to be rare. Travel-related cases have been confirmed in multiple countries and regions outside mainland China including Germany, France, Thailand, Japan, South Korea, Vietnam, Canada, and the United States, as well as Hong Kong and Taiwan. Domestically in China, the virus has also been noted in several cities and provinces with cases in all but one provinence. While zoonotic transmission appears to be the original source of infections, the most alarming development is that human-to-human transmission is now prevelant. Of particular concern is that many healthcare workers have been infected in the current epidemic. There are several critical clinical questions that need to be resolved, including how efficient is human-to-human transmission? What is the animal reservoir? Is there an intermediate animal reservoir? Do the vaccines generated to the SARS-CoV or MERS-CoV or their proteins offer protection against 2019-nCoV? We offer a research perspective on the next steps for the generation of vaccines. We also present data on the use of in silico docking in gaining insight into 2019-nCoV Spike-receptor binding to aid in therapeutic development. Diagnostic PCR protocols can be found at https://www.who.int/health-topics/coronavirus/laboratory-diagnostics-for-novel-coronavirus.

Background: Human papillomavirus (HPV), one of the most common sexually transmitted viral infections worldwide, is the leading cause of cervical cancer. In Middle East and North Africa (MENA) Region HPV data is at scarce, and most of the countries haven't implemented any vaccination programs. This present meta-analysis and systematic review aimed to describe human papillomavirus (HPV) epidemiology by clinical subgroups in the (MENA) region. Methods: Studies assessing HPV prevalence rates were systematically reviewed, and the selected articles were reported following the PRISMA guideline. Random-effects meta-analyses and meta-regression were used to estimate HPV pooled mean prevalence rates and their association with other factors. Results: For the cervical cancer population in the MENA region, the pooled HPV prevalence rate was 81% (95% CI, 70%-90%). HPV detected in cervical cancer samples was most prevalent in the Maghreb countries (88%; 95% CI, 78%-96%) and least prevalent in Iran (73%; 95% CI, 62%-83%).For the subgroup with abnormal-cervical cytology in the MENA region, the pooled HPV prevalence rate was 54% (95% CI, 41%-67%), with the highest prevalence reported in Northeast Africa (94%; 95% CI, 91%-96%), and the lowest prevalence in the Levant region (31%; 95 CI, 16%-49%). In the general population subgroup in the MENA region, the pooled HPV prevalence rate was 16% (95% CI, 14%-17%), HPV was most prevalent in the Northeast Africa region (21%; 95 CI, 7%-40%) and least prevalent in the Levant region (7%; 95 CI, 2%-14%). Conclusion: The present meta-analysis comprehensively described the current HPV prevalence rates in the MENA region and found that the rates have continued to increase with time, especially in African regions. Designing personalized awareness and vaccination programs that respect the various cultural and religious values remains the main challenge in prevention of cervical cancer in the MENA region. (C) 2020 The Author(s). Published by Elsevier Ltd on behalf of King Saud Bin Abdulaziz University for Health Sciences.

Background and Objectives. Malaria infection, caused by Plasmodium falciparum, is the most lethal and frequently culminates in severe clinical complications. Interleukin-22 (IL-22) has been implicated in several diseases including malaria. The objective of this study was to investigate the role of IL-22 gene polymorphisms in P. falciparum infection. Material and Methods. Ten single-nucleotide polymorphisms (SNPs), rs976748, rs1179246, rs2046068, rs1182844, rs2227508, rs2227513, rs2227478, rs2227481, rs2227491, and rs2227483, of IL-22 gene were genotyped through PCR-based assays of 250 P. falciparum-infected patients and 200 healthy controls. In addition, a luciferase reporter assay was done to assess the role of the rs2227513 SNP in IL-22 gene promoter activity. Results. We found that the rs2227481 TT genotype (odds ratio 0.254, confidence interval = 0.097-0.663, P=0.002) and the T allele is associated with protection against P. falciparum malaria as well as the rs2227483 AT genotype (odds ratio 0.375, confidence interval = 0.187-0.754, P=0.004). The haplotype A-T-T of rs1179246, rs1182844, and rs976748 was statistically more frequent in the control group (frequency 41%, P=0.034) as well as the haplotype A-G of rs2046068 and rs2227491 (frequency 49.4%, P=0.041). The variant rs2227513 G allele had a statistically higher activity (P<0.0001) with the luciferase reporter assay. Conclusion. The study suggests that IL-22 polymorphisms in rs2227481 and rs2227483 could contribute to protection against P. falciparum malaria. Also, the G allele of rs2227513, located in the promoter region of IL-22 gene, could be essential for higher expression levels of IL-22 cytokine.

Abstract The present study aimed to screen for and genotype HPV among Saudi women; develop sensitive quantitative polymerase chain reaction (qPCR) assays to detect the viral load for the two most common HPV types, namely 16 and 18; and assess whether HPV viral load could be used as a marker for cervical abnormality and disease progression. This study examined 733 specimens (both formalin-fixed, paraffin-embedded specimens and PAP smear samples) from women who underwent cervical screening. The specimens and samples were processed for DNA extraction and then tested for HPV DNA. Approximately 165 specimens (18%) were positive for HPV. Those specimens were genotyped using a reverse line blotting hybridization assay. The results indicated that the most common HPV types detected were a single infection with HPV 16 (51%) or with HPV 18 (28%) followed by infections with multiple HPV types (~7%). A qPCR TaqMan assay developed in-house was used to determine the viral load for HPV genotypes 16 (n=80) and 18 (n=45). Viral loads for both HPV types were significantly associated with cervical cytology grade (P < 0.05). The odds ratio (OR) for the HPV 16 viral load was high for specimens with cervical cancer (OR, 18.8; 95% CI, 4.3–82.9) or for those with high-grade squamous intraepithelial lesions (OR, 14.7; 95% Cl, 2.43–88.49). For the HPV 18 viral load, the OR was significant only for specimens with cervical cancer (OR, 11.1; 95% Cl, 2.2–54.9). Logistic regression models for HPV 16 and for HPV 18 viral load levels were significant, with higher viral load associated with cervical abnormalities. These findings indicate that viral load is a predictor significantly associated with cytology abnormality in women who are positive for high-risk HPVs and suggest integrating a viral load test into current clinical screening practices for HPV-positive women. Citation Format: Dalia Obeid, Shihana Almatrrouk, Hadeel Khayat, Turfah Al-Muammer, Asma Tullbah, Ismail Albadawi, Mohammed Alahdal, Fatimah Alhamlan. Human papillomavirus type 16 and 18 viral loads as predictors associated with abnormal cervical cytology among women in Saudi Arabia [abstract]. In: Proceedings of the AACR Special Conference on the Microbiome, Viruses, and Cancer; 2020 Feb 21-24; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2020;80(8 Suppl):Abstract nr A17.

INTRODUCTION:Human papillomavirus (HPV) infection is typically critical in the oncogenesis of cervical cancer. However, available HPV detection kits differ in their ability and sensitivity to detect various types of HPV, and this variability has led to inconsistencies in the reporting of the geographic prevalence of HPV types, especially in developing countries. Here, we compared results of the recently developed GenoFlow HPV array test, which detects 33 HPV genotypes, to those of the well-established reverse line blot (RLB) assay, which detects 23 HPV types.METHODOLOGY:In total, 608 cervical specimens with cytology results ranging from normal to cancer were collected using an endocervical brush from women attending outpatient clinics in Riyadh, Saudi Arabia.RESULTS:Sixty-nine specimens (11%) were positive for HPV. HPV genotype detection using the GenoFlow test had a sensitivity of 62% and a specificity of 100%. Overall agreement between the two HPV genotyping methods was 97%, with a concordance rate of 95%. Among the GenoFlow test results, 2% indicated additional HPV types that were not detected in the RLB assay, whereas the GenoFlow test missed 0.3% of the HPV types that were detected by the RLB; however, both tests were in agreement in detecting all major HPV types.CONCLUSION:The GenoFlow test was reliable, with results comparable to the RLB test. However, because the GenoFlow test is less labor-intensive and takes less total time (3 hours), it is a promising, affordable alternative to the RLB for HPV diagnosis and screening programs.

Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb) infection, remains a major global health emergency. It is estimated that one third of global population are affected, predominantly with latent granuloma form of the disease. Mtb co-evolved with humans, for its obligatory intra-macrophage phagosome habitat and slow replication, balanced against unique mycobacterial innate immunity, which appears to be highly complex. TB is transmitted via cough aerosol Mtb inhalation. Bovine TB attenuated Bacillus Calmette Guerin (BCG) live vaccine has been in practice for protection of young children from severe disseminated Mtb infection, but not sufficiently for their lungs, as obtained by trials in TB endemic community. To augment BCG vaccine-driven innate and adaptive immunity for neonates and better protection against adult pulmonary TB, a number of BCG pre-vaccination based, subset vaccine candidates have been tested via animal preclinical, followed by safe clinical trials. BCG also enhances innate macrophage trained immunity and memory, through primordial intracellular Toll-like receptors (TLRs) 7 and 9, which recognise distinct mycobacterial molecular pattern signature. This signature is transmitted by TLR signalling via nuclear factor-κB, for activating innate immune transcription and expression of gene profiling in a mycobacterial signature-specific manner. These are epigenetically imprinted in reprogramming of distinct chromatin areas for innate immune memory, to be recalled following lung reinfection. Unique TB innate immunity and its trained memory are considered independent from adaptive immune B and T cells. On the other hand, adaptive immunity is crucial in Mtb containment in granulomatous latency, supported by innate immune cell infiltration. In nearly 5–10 % of susceptible people, latent TB may be activated due to immune evasion by Mtb from intracellular phagosome within macrophage, perpetrating TB. However, BCG and new recombinant BCG vaccines have the capacity, as indicated in pre- and clinical trials, to overcome such Mtb evasion. Various strategies include pro-inflammatory-bactericidal type 1 polarisation (M1) phenotype of the infected macrophage, involving thrombospondin-TLR pathway. Saprophytic M. smegmatis-based recombinant vaccines are also promising candidates against TB. BCG vaccination of neonates/infants in TB endemic countries also reduced their pneumonia caused by various microbes independent of TB immunity. Here, we discuss host immune response against Mtb, its immune evasion strategies, and the important role innate immunity plays in the development of protection against TB.

The Kingdom of Saudi Arabia is one of the G20 nations and its global influence is arguably undisputed. The country hosts millions of expatriate residents whose mobility has been affected by the COVID-19 pandemic.

The detection of HPV viral DNA is regularly conducted with cervical screening. However, using a molecular marker such as the viral load may serve as a predictor associated with disease detection and progression. The present study aimed to screen for and genotype HPV among women in Saudi Arabia, develop and validate sensitive quantitative polymerase chain reaction (qPCR) assays to detect viral load for the two most common HPV types, namely 16 and 18, and assess whether HPV viral load could be used as a marker for cervical abnormality and disease progression. This study examined 733 specimens (both formalin-fixed paraffin embedded specimens and PAP smear samples) from women who underwent cervical screening. The specimens and samples were processed for DNA extraction and then tested for HPV DNA using nested PCR. Approximately 165 specimens (18%) were positive for HPV. Those specimens were genotyped using a reverse line blotting hybridization assay. The results indicated that the most common HPV types detected were a single infection with HPV 16 (51%) or with HPV 18 (28%) followed by infections with multiple HPV types (~7%). A qPCR TaqMan assay developed and validated in-house was used to determine viral load for HPV genotypes 16 (n = 80) and 18 (n = 45). Viral loads for both HPV types were significantly associated with cervical cytology grade (P < 0.05). The odds ratio (OR) for the HPV 16 viral load was high for specimens with cervical cancer (OR, 18.8; 95% CI, 4.3–82.9) or for those with high-grade squamous intraepithelial lesions (OR, 14.7; 95% Cl, 2.43–88.49). For the HPV 18 viral load, the OR was significant only for specimens with cervical cancer (OR, 11.1; 95% Cl, 2.2–54.9). Logistic regression models for HPV 16 and for HPV 18 viral load levels were significant, with higher viral load associated with cervical abnormalities. These findings indicate that viral load is a predictor significantly associated with cytology abnormality in women who are positive for high-risk HPVs and suggest that integrating a viral load test into current clinical screening practices for HPV-positive women is warranted in Saudi Arabia.

Familial hypercholesterolemia (FH) is an autosomal dominant disease most often caused by mutations in the low-density lipoprotein receptor (LDLR) gene, which consists of 18 exons spanning 45 kb and codes for a precursor protein of 860 amino acids. Mutations in the LDLR gene lead to a reduced hepatic clearance of LDL as well as a high risk of coronary artery disease (CAD) and sudden cardiac death (SCD). Recently, LDLR transgenes have generated interest as potential therapeutic agents. However, LDLR packaging using a lentiviral vector (LVV) system pseudotyped with a vesicular stomatitis virus (VSV)-G envelope is not efficient. In this study, we modified the LVV system to improve transduction efficiency and investigated the LDLR regions responsible for transduction inhibition. Transduction efficiency of 293T cells with a 5'-LDLReGFP-3' fusion construct was only 1.55% compared to 42.32% for the eGFP construct. Moreover, co-expression of LDLR affected eGFP packaging. To determine the specific region of the LDLR protein responsible for packaging inhibition, we designed constructs with mutations or sequential deletions at the 3' and 5' ends of LDLR cDNA. All constructs except one without the ligand-binding domain (LBD) (pWoLBD-eGFP) resulted in low transduction efficiency, despite successful packaging of viral RNA in the VSV envelope, as confirmed through RT-PCR. When we evaluated a direct interaction between LDLR and the VSV envelope glycoprotein using MD simulation and protein-protein interactions, we uncovered Val119, Thr120, Thr67, and Thr118 as exposed residues in the LDLR receptor that interact with the VSV protein. Together, our results suggest that the LBD of LDLR interacts with the VSV-G protein during viral packaging, which significantly reduces transduction efficiency.

Despite the implementation of various vaccination programs, hepatitis B virus (HBV) poses a considerable health problem in Saudi Arabia. Insight on HBV evolutionary history in the region is limited. We performed a comprehensive epidemiological and phylogenetic reconstruction based on a large cohort of HBV infected patients. Three hundred and nineteen HBV-infected patients with different clinical manifestations, including inactive and active chronic carriers and patients with cirrhosis and hepatocellular carcinoma (HCC), were enrolled in this study. The full-length large S gene was amplified and sequenced. Phylogenetic analysis was performed to determine the genotype and subgenotypes of the isolates. Phylogenetic tree analysis revealed that genotype D is the most dominant genotype among patients. Moreover, this analysis identified two strains with genotype E isolated from active carriers. Detailed phylogenetic analyses confirmed the presence of four HBV D subgenotypes, D1 (93%, n = 296), D2 (0.02%, n = 5), D3 (0.003%, n = 1), and D4 (0.003%, n = 1). In addition, six genotype D strains were not assigned to any existing HBV D subgenotype. The large S gene of eight strains showed signatures of genotype recombination between the genotypes D and A and between D and E. Several strains harbored medically important point mutations at the protein level. Along with the dominance of the HBV genotype D, isolation of the E genotype and several recombinant strains from patients with Saudi Arabian origin is an essential result for decisions involving therapeutic measures for patients. Development of vaccines and detection of diagnostic escape mutations at antigenic epitopes on the HBsAg will be valuable to public health authorities. Furthermore, the diversity at the nucleotide and amino acid levels and different proportions of dN/dS at the PreS1, PreS2, and HBsAg reveal the selective pressure trend from inactive status towards advanced liver diseases.

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