Publications by authors named "Naif Khalaf Alharbi"

18 Publications

  • Page 1 of 1

Nationwide Seroprevalence of SARS-CoV-2 in Saudi Arabia.

J Infect Public Health 2021 Jul 24;14(7):832-838. Epub 2021 Apr 24.

King Abdullah International Medical Research Center, Riyadh, Saudi Arabia; King Saud Bin Abdulaziz University for Health Sciences, Riyadh, Saudi Arabia.

Background: Estimated seroprevalence of Coronavirus Infectious Disease 2019 (COVID-19), caused by the Severe Acute Respiratory Syndrome-Coronavirus-2 (SARS-CoV-2) is a critical evidence for a better evaluation of the virus spread and monitoring the progress of COVID-19 pandemic in a population. In the Kingdom of Saudi Arabia (KSA), SARS-CoV-2 seroprevalence has been reported in specific regions, but an extensive nationwide study has not been reported. Here, we report a nationwide study to determine the prevalence of SARS-CoV-2 in the population of KSA during the pandemic, using serum samples from healthy blood donors, non-COVID patients and healthcare workers (HCWs) in six different regions of the kingdom, with addition samples from COVID-19 patients.

Methods: A total of 11,703 serum samples were collected from different regions of the KSA including; 5395 samples from residual healthy blood donors (D); 5877 samples from non-COVID patients collected through residual sera at clinical biochemistry labs from non-COVID patients (P); and 400 samples from consented HCWs. To determine the seroprevalence of SARS-CoV-2, all serum samples, in addition to positive control sera from RT-PCR confirmed COVID-19 patients, were subjected to in-house ELISA with a sample pooling strategy, which was further validated by testing individual samples that make up some of the pools, with a statistical estimation method to report seroprevalence estimates.

Results: Overall (combining D and P groups) seroprevalence estimate was around 11% in Saudi Arabia; and was 5.1% (Riyadh), 1.5% (Jazan), 18.4% (Qassim), 20.8% (Hail), 14.7% (ER; Alahsa), and 18.8% in Makkah. Makkah samples were only D group and had a rate of 24.4% and 12.8% in the cities of Makkah and Jeddah, respectively. The seroprevalence in Saudi Arabia across the sampled areas would be 12 times the reported COVID-19 infection rate. Among HCWs, 7.5% (4.95-10.16 CI 95%) had reactive antibodies to SARS-CoV-2 without reporting any previously confirmed infection. This was higher in HCWs with hypertension. The study also presents the demographics and prevalence of co-morbidities in HCWs and subset of non-COVID-19 population.

Interpretation: Our study estimates the overall national serological prevalence of COVID-19 in Saudi Arabia to be 11%, with an apparent disparity between regions. This indicates the prevalence of asymptomatic or mild unreported COVID-19 cases.
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http://dx.doi.org/10.1016/j.jiph.2021.04.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8188888PMC
July 2021

Viruses Causing Aseptic Meningitis: A Tertiary Medical Center Experience With a Multiplex PCR Assay.

Front Neurol 2020 7;11:602267. Epub 2020 Dec 7.

College of Medicine, King Saud bin Abdulaziz University for Health Sciences, Riyadh, Saudi Arabia.

Central nervous system (CNS) infection is associated with high rates of morbidity and mortality, and despite advancements in molecular testing, aseptic meningitis remains challenging to diagnose. Aseptic meningitis cases are often underreported worldwide, which impacts the quality of patient care. Therefore, we aimed to assess the results of BioFire® FilmArray® meningitis/encephalitis (ME) PCR panel, clinical characteristics, and etiologies of aseptic meningitis patients. From January 2018 to January 2020, all pediatric and adult patients in a large tertiary medical center who underwent lumbar puncture and cerebrospinal fluid (CSF) testing by a ME multiplex PCR panel and who fit the aseptic meningitis definition were retrospectively reviewed. Data were reviewed from 1,607 patients; 240 met the inclusion criteria (54.6% males; 68.8% <4 years of age). The rate of detected viral causes of aseptic meningitis was 40.4%; therefore, 59.6% of the patients remained with unidentified etiology. Among the identified viral meningitis, enterovirus and human herpesvirus 6 (HHV-6) were the most common (25 and 7.9%, respectively). The median length of hospital stay was 6 days, and it was longer in patients with unidentifiable aseptic meningitis ( < 0.0001). Aseptic meningitis is common among suspected meningitis patients, but most cases remained of unknown etiology. The most common identified viruses were enterovirus followed by HHV-6, and there is predominance in males and the pediatric age group. These results highlight that further research is needed to identify other etiologies and possible additional viral pathogens for aseptic meningitis in the current diagnostic methods.
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http://dx.doi.org/10.3389/fneur.2020.602267DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7793969PMC
December 2020

Seroprevalence of SARS-CoV-2 (COVID-19) among healthcare workers in Saudi Arabia: comparing case and control hospitals.

Diagn Microbiol Infect Dis 2021 Mar 20;99(3):115273. Epub 2020 Nov 20.

Public Health Lab, Saudi Center for Disease Prevention and Control, Riyadh, Saudi Arabia; Department of Pathology, School of Medicine, King Saud University, Riyadh, Saudi Arabia.

Healthcare workers (HCWs) stand at the frontline for fighting coronavirus disease 2019 (COVID-19) pandemic. This puts them at higher risk of acquiring the infection than other individuals in the community. Defining immunity status among health care workers is therefore of interest since it helps to mitigate the exposure risk. This study was conducted between May 20 and 30, 2020. Eighty-five hospitals across Kingdom of Saudi Arabia were divided into 2 groups: COVID-19 referral hospitals are those to which RT-PCR-confirmed COVID-19 patients were admitted or referred for management (Case-hospitals). COVID-19 nonaffected hospitals where no COVID-19 patients had been admitted or managed and no HCW outbreak (Control hospitals). Next, seroprevalence of severe acute respiratory syndrome coronavirus 2 among HCWs was evaluated; there were 12,621 HCWs from the 85 hospitals. There were 61 case-hospitals with 9379 (74.3%) observations, and 24 control-hospitals with 3242 (25.7%) observations. The overall positivity rate by the immunoassay was 299 (2.36%) with a significant difference between the case-hospital (2.9%) and the control-group (0.8%) (P value <0.001). There was a wide variation in the positivity rate between regions and/or cities in Saudi Arabia, ranging from 0% to 6.31%. Of the serology positive samples, 100 samples were further tested using the SAS2pp neutralization assay; 92 (92%) samples showed neutralization activity. The seropositivity rate in Kingdom of Saudi Arabia is low and varies across different regions with higher positivity in case-hospitals than control-hospitals. The lack of neutralizing antibodies (NAb) in 8% of the tested samples could mean that assay is a more sensitive assay or that neutralization assay has a lower detection limits; or possibly that some samples had cross-reaction to spike protein of other coronaviruses in the assay, but these were not specific to neutralize severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).
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http://dx.doi.org/10.1016/j.diagmicrobio.2020.115273DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7677039PMC
March 2021

High Rate of Circulating MERS-CoV in Dromedary Camels at Slaughterhouses in Riyadh, 2019.

Viruses 2020 10 27;12(11). Epub 2020 Oct 27.

Vaccine Development Unit, Department of Infectious Disease Research, King Abdullah International Medical Research Center, Riyadh 11564, Saudi Arabia.

MERS-CoV is a zoonotic virus that has emerged in humans in 2012 and caused severe respiratory illness with a mortality rate of 34.4%. Since its appearance, MERS-CoV has been reported in 27 countries and most of these cases were in Saudi Arabia. So far, dromedaries are considered to be the intermediate host and the only known source of human infection. This study was designed to determine the seroprevalence and the infection rate of MERS-CoV in slaughtered food-camels in Riyadh, Saudi Arabia. A total of 171 nasal swabs along with 161 serum samples were collected during the winter; from January to April 2019. Nasal swabs were examined by Rapid test and RT-PCR to detect MERS-CoV RNA, while serum samples were tested primarily using S1-based ELISA Kit to detect MERS-CoV (IgG) antibodies and subsequently by MERS pseudotyped viral particles (MERSpp) neutralization assay for confirmation. Genetic diversity of the positive isolates was determined based on the amplification and sequencing of the spike gene. Our results showed high prevalence (38.6%) of MERS-CoV infection in slaughtered camels and high seropositivity (70.8%) during the time of the study. These data indicate previous and ongoing MERS-CoV infection in camels. Phylogenic analysis revealed relatively low genetic variability among our isolated samples. When these isolates were aligned against published spike sequences of MERS-CoV, deposited in global databases, there was sequence similarity of 94%. High seroprevalence and high genetic stability of MERS-CoV in camels indicating that camels pose a public health threat. The widespread MERS-CoV infections in camels might lead to a risk of future zoonotic transmission into people with direct contact with these infected camels. This study confirms re-infections in camels, highlighting a challenge for vaccine development when it comes to protective immunity.
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http://dx.doi.org/10.3390/v12111215DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7692456PMC
October 2020

Transcriptomics-Based Characterization of the Toxicity of ZnO Nanoparticles Against Chronic Myeloid Leukemia Cells.

Int J Nanomedicine 2020 13;15:7901-7921. Epub 2020 Oct 13.

Brighton and Sussex Medical School, University of Sussex, Brighton, UK.

Introduction: Zinc oxide nanoparticles (ZnO NPs) have recently attracted attention as potential anti-cancer agents. To the best of our knowledge, the toxicity of ZnO NPs against human chronic myeloid leukemia cells (K562 cell line) has not been studied using transcriptomics approach.

Objective: The goals of this study were to evaluate the capability of ZnO NPs to induce apoptosis in human chronic myeloid leukemia cells (K562 cells) and to investigate the putative mechanisms of action.

Methods: We used viability assay and flowcytometry coupled with Annexin V-FITC and propidium iodide to investigate the toxicity of ZnO NPs on K562 cells and normal peripheral blood mononuclear cells. Next we utilized a DNA microarray-based transcriptomics approach to characterize the ZnO NPs-induced changes in the transcriptome of K562 cells.

Results: ZnO NPs exerted a selective toxicity (mainly by apoptosis) on the leukemic cells (≤0.005) and altered their transcriptome; 429 differentially expressed genes (DEGs) with fold change (FC)≥4 and ≤0.008 with corrected ≤0.05 were identified in K562 cells post treatment with ZnO NPs. The over-expressed genes were implicated in "response to zinc", "response to toxic substance" and "negative regulation of growth" (corrected ≤0.05). In contrast, the repressed genes positively regulated "cell proliferation", "cell migration", "cell adhesion", "receptor signaling pathway via JAK-STAT" and "phosphatidylinositol 3-kinase signaling" (corrected ≤0.05). Lowering the FC to ≥1.5 with ≤0.05 and corrected ≤0.1 showed that ZnO NPs over-expressed the anti-oxidant defense system, drove K562 cells to undergo mitochondrial-dependent apoptosis, and targeted NF-κB pathway.

Conclusion: Taken together, our findings support the earlier studies that reported anti-cancer activity of ZnO NPs and revealed possible molecular mechanisms employed by ZnO NPs to induce apoptosis in K562 cells.
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http://dx.doi.org/10.2147/IJN.S261636DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7568638PMC
November 2020

Challenge infection model for MERS-CoV based on naturally infected camels.

Virol J 2020 06 17;17(1):77. Epub 2020 Jun 17.

Ministry of Environment, Water and Agriculture (MEWA), Riyadh, Saudi Arabia.

Background: Middle East Respiratory Syndrome coronavirus (MERS-CoV) is an emerging virus that infects humans and camels with no approved antiviral therapy or vaccine. Some vaccines are in development for camels as a one-health intervention where vaccinating camels is proposed to reduce human viral exposure. This intervention will require an understanding of the prior exposure of camels to the virus and appropriate vaccine efficacy studies in camels.

Methods: We conducted a cross sectional seroprevalence study in young dromedary camels to determine the rate of MERS-CoV seropositivity in young camels. Next, we utilised naturally infected camels as a natural challenge model that can be used by co-housing these camels with healthy naive camels in a ratio of 1 to 2. This model is aimed to support studies on natural virus transmission as well as evaluating drug and vaccine efficacy.

Results: We found that 90% of the screened camels have pre-existing antibodies for MERS-CoV. In addition, the challenge model resulted in MERS-CoV transmission within 48 h with infections that continued for 14 days post challenge.

Conclusions: Our finding suggests that the majority of young dromedary camels in Saudi Arabia are seropositive and that naturally infected camels can serve as a challenge model to assess transmission, therapeutics, and vaccine efficacy.
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http://dx.doi.org/10.1186/s12985-020-01347-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7298446PMC
June 2020

Preparedness and response to COVID-19 in Saudi Arabia: Building on MERS experience.

J Infect Public Health 2020 Jun 11;13(6):834-838. Epub 2020 May 11.

Department of Medical Microbiology and Parasitology, Faculty of Medicine, King Abdulaziz University, Jeddah, Saudi Arabia; Vaccines and Immunnotherapy Unit, King Fahd Medical Research Center, King Abdulaziz University, Jeddah, Saudi Arabia. Electronic address:

Nearly four months have passed since the emergence of the severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2), which caused the rapidly spreading Coronavirus Disease 2019 (COVID-19) pandemic. To date, there have been more than 2.3 million confirmed cases and more than 160,000 deaths globally caused by COVID-19. Chinese health authorities, where the virus emerged, have taken prompt strict public health measures to control and prevent the spread of the outbreak. In Saudi Arabia, unprecedented precautionary strict measures were applied to prevent virus entry to the country or to mitigate its impact when it arrives. Here, we review the response of Saudi Arabia to COVID-19 pandemic and how did the experience learned from the Middle East respiratory syndrome coronavirus (MERS-CoV) epidemic since 2012 has helped the country to be better prepared for the current COVID-19 pandemic. We also discuss the country readiness, improvement in research and development, and the unprecedented rapid precautionary measures that have been taken by the Saudi government thus far.
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http://dx.doi.org/10.1016/j.jiph.2020.04.016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7211706PMC
June 2020

Seroprevalence of MERS-CoV in healthy adults in western Saudi Arabia, 2011-2016.

J Infect Public Health 2020 May 28;13(5):697-703. Epub 2020 Jan 28.

Vaccines and Immunotherapy Unit, King Fahd Medical Research Center, King Abdulaziz University, Jeddah, Saudi Arabia; Department of Medical Microbiology and Parasitology, Faculty of Medicine, King Abdulaziz University, Jeddah, Saudi Arabia. Electronic address:

Background: The Middle East respiratory syndrome coronavirus (MERS-CoV) is a newly recognized zoonotic coronavirus. Current evidence confirms the role of dromedaries in primary human infections but does not explain the sporadic community cases. However, asymptomatic or subclinical cases could represent a possible source of infection in the community.

Methods: Archived human sera (7461) collected between 2011 and 2016 from healthy adult blood donors from 50 different nationalities in the western part of Saudi Arabia were obtained for MERS-CoV seroprevalence investigation. Samples were tested for MERS-CoV S1-specific antibodies (Abs) by ELISA and confirmed by testing for neutralizing Abs (nAbs) using both pseudotyped and live virus neutralization assays.

Results: Out of 7461 samples, 174 sera from individuals with 18 different nationalities were ELISA positive (2.3%, 95% CI 2.0-2.7). Presence of nAbs was confirmed in 17 samples (0.23%, 95% CI 0.1-0.4) of which one sample exhibited positivity in both neutralization assays. Confirmed seropositivity was identified in young (15-44 years) men and women from Saudi Arabia, Egypt, Yemen, Pakistan, Palestine, Sudan, and India without significant preference.

Conclusions: An increasing trend of MERS-CoV seroprevalence was observed in the general population in western Saudi Arabia, suggesting that asymptomatic or mild infections might exist and act as an unrecognized source of infection. Seropositivity of individuals from different nationalities underscores the potential MERS exportation outside of the Arabian Peninsula. Thus, enhanced and continuous surveillance is highly warranted.
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http://dx.doi.org/10.1016/j.jiph.2020.01.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7104088PMC
May 2020

Generation of MERS-CoV Pseudotyped Viral Particles for the Evaluation of Neutralizing Antibodies in Mammalian Sera.

Methods Mol Biol 2020 ;2099:117-126

Department of Medical Microbiology and Parasitology, Faculty of Medicine, King Abdulaziz University, Jeddah, Saudi Arabia.

Pseudotyped viral particle production has been used extensively and broadly for many viruses to evaluate levels of neutralizing antibodies, viral entry inhibitors and vaccine immunogenicity. This assay is extremely safe and useful alternative to live virus-based assay without the need for high containment facilities. In this chapter, we describe the generation of MERS-CoV pseudotyped viral particles (MERSpp) expressing full-length spike protein using second-generation lentiviral packaging system. This platform is optimized to generate high titer of MERSpp and to test sera from different mammalian species.
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http://dx.doi.org/10.1007/978-1-0716-0211-9_10DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7123069PMC
September 2020

Humoral Immunogenicity and Efficacy of a Single Dose of ChAdOx1 MERS Vaccine Candidate in Dromedary Camels.

Sci Rep 2019 11 8;9(1):16292. Epub 2019 Nov 8.

Department of Infectious Disease Research, King Abdullah International Medical Research Center, Riyadh, Saudi Arabia.

MERS-CoV seronegative and seropositive camels received a single intramuscular dose of ChAdOx1 MERS, a replication-deficient adenoviral vectored vaccine expressing MERS-CoV spike protein, with further groups receiving control vaccinations. Infectious camels with active naturally acquired MERS-CoV infection, were co-housed with the vaccinated camels at a ratio of 1:2 (infected:vaccinated); nasal discharge and virus titres were monitored for 14 days. Overall, the vaccination reduced virus shedding and nasal discharge (p = 0.0059 and p = 0.0274, respectively). Antibody responses in seropositive camels were enhancedby the vaccine; these camels had a higher average age than seronegative. Older seronegative camels responded more strongly to vaccination than younger animals; and neutralising antibodies were detected in nasal swabs. Further work is required to optimise vaccine regimens for younger seronegative camels.
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http://dx.doi.org/10.1038/s41598-019-52730-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6841732PMC
November 2019

A Highly Immunogenic, Protective, and Safe Adenovirus-Based Vaccine Expressing Middle East Respiratory Syndrome Coronavirus S1-CD40L Fusion Protein in a Transgenic Human Dipeptidyl Peptidase 4 Mouse Model.

J Infect Dis 2019 10;220(10):1558-1567

Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston.

Background: Infection control measures have played a major role in limiting human/camel-to-human transmission of Middle East respiratory syndrome coronavirus (MERS-CoV); however, development of effective and safe human or camel vaccines is warranted.

Methods: We extended and optimized our previous recombinant adenovirus 5 (rAd5)-based vaccine platform characterized by in vivo amplified and CD40-mediated specific responses to generate MERS-CoV S1 subunit-based vaccine. We generated rAd5 constructs expressing CD40-targeted S1 fusion protein (rAd5-S1/F/CD40L), untargeted S1 (rAd5-S1), and Green Fluorescent Protein (rAd5-GFP), and evaluated their efficacy and safety in human dipeptidyl peptidase 4 transgenic (hDPP4 Tg+) mice.

Results: Immunization of hDPP4 Tg+ mice with a single dose of rAd5-S1/F/CD40L elicited as robust and significant specific immunoglobulin G and neutralizing antibodies as those induced with 2 doses of rAd5-S1. After MERS-CoV challenge, both vaccines conferred complete protection against morbidity and mortality, as evidenced by significantly undetectable/reduced pulmonary viral loads compared to the control group. However, rAd5-S1- but not rAd5-S1/F/CD40L-immunized mice exhibited marked pulmonary perivascular hemorrhage post-MERS-CoV challenge despite the observed protection.

Conclusions: Incorporation of CD40L into rAd5-based MERS-CoV S1 vaccine targeting molecule and molecular adjuvants not only enhances immunogenicity and efficacy but also prevents inadvertent pulmonary pathology after viral challenge, thereby offering a promising strategy to enhance safety and potency of vaccines.
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http://dx.doi.org/10.1093/infdis/jiz137DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7107499PMC
October 2019

Poxviral promoters for improving the immunogenicity of MVA delivered vaccines.

Hum Vaccin Immunother 2019 6;15(1):203-209. Epub 2018 Sep 6.

a Infectious Disease Research Department , King Abdullah International Medical Research Center (KAIMRC) , Riyadh , Saudi Arabia.

Modified vaccinia virus Ankara (MVA) is a replication-deficient poxvirus, attenuated in chick embryo fibroblast primary cells. It has been utilised as a viral vector to develop many vaccines against cancer and infectious diseases such as malaria, HIV/AIDS, influenza, and tuberculosis, MERS-CoV, and Ebola virus infection. There is accumulating data from many preclinical and clinical studies that highlights the excellent safety and immunogenicity of MVA. However, due to the complex nature of many pathogens and their pathogenicity, MVA vectored vaccine candidates need to be optimised to improve their immunogenicity. One of the main approaches to improve MVA immunogenicity focuses on optimising poxviral promoters that drive recombinant vaccine antigens, encoded within recombinant MVA vector genome. A number of promoters were described or optimised to improve the development of MVA based vaccines such as p7.5, pF11, and mH5 promoters. This review focuses on poxviral promoters, their optimisation, genetic stability, and clinical use.
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http://dx.doi.org/10.1080/21645515.2018.1513439DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6363155PMC
February 2020

ChAdOx1 and MVA based vaccine candidates against MERS-CoV elicit neutralising antibodies and cellular immune responses in mice.

Vaccine 2017 06 1;35(30):3780-3788. Epub 2017 Jun 1.

The Jenner Institute, University of Oxford, Oxford OX3 7DQ, UK.

The Middle East respiratory syndrome coronavirus (MERS-CoV) has infected more than 1900 humans, since 2012. The syndrome ranges from asymptomatic and mild cases to severe pneumonia and death. The virus is believed to be circulating in dromedary camels without notable symptoms since the 1980s. Therefore, dromedary camels are considered the only animal source of infection. Neither antiviral drugs nor vaccines are approved for veterinary or medical use despite active research on this area. Here, we developed four vaccine candidates against MERS-CoV based on ChAdOx1 and MVA viral vectors, two candidates per vector. All vaccines contained the full-length spike gene of MERS-CoV; ChAdOx1 MERS vaccines were produced with or without the leader sequence of the human tissue plasminogen activator gene (tPA) where MVA MERS vaccines were produced with tPA, but either the mH5 or F11 promoter driving expression of the spike gene. All vaccine candidates were evaluated in a mouse model in prime only or prime-boost regimens. ChAdOx1 MERS with tPA induced higher neutralising antibodies than ChAdOx1 MERS without tPA. A single dose of ChAdOx1 MERS with tPA elicited cellular immune responses as well as neutralising antibodies that were boosted to a significantly higher level by MVA MERS. The humoral immunogenicity of a single dose of ChAdOx1 MERS with tPA was equivalent to two doses of MVA MERS (also with tPA). MVA MERS with mH5 or F11 promoter induced similar antibody levels; however, F11 promoter enhanced the cellular immunogenicity of MVA MERS to significantly higher magnitudes. In conclusion, our study showed that MERS-CoV vaccine candidates could be optimized by utilising different viral vectors, various genetic designs of the vectors, or different regimens to increase immunogenicity. ChAdOx1 and MVA vectored vaccines have been safely evaluated in camels and humans and these MERS vaccine candidates should now be tested in camels and in clinical trials.
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http://dx.doi.org/10.1016/j.vaccine.2017.05.032DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5516308PMC
June 2017

Vaccines against Middle East respiratory syndrome coronavirus for humans and camels.

Rev Med Virol 2017 03 27;27(2). Epub 2016 Oct 27.

Infectious Diseases Research Department, King Abdullah International Medical Research Center (KAIMRC), Riyadh, Saudi Arabia.

Middle East respiratory syndrome coronavirus (MERS-CoV) is caused by a novel betacoronavirus that was isolated in late 2012 in Saudi Arabia. The viral infections have been reported in more than 1700 humans, ranging from asymptomatic or mild cases to severe pneumonia with a mortality rate of 40%. It is well documented now that dromedary camels contract the infection and shed the virus without notable symptoms, and such animals had been infected by at least the early 1980s. The mechanism of camel to human transmission is still not clear, but several primary cases have been associated with camel contact. There is no approved antiviral drug or vaccine against MERS-CoV despite the active research in this area. Vaccine candidates have been developed using various platforms and regimens and have been tested in several animal models. Here, this article reviews the published studies on MERS-CoV vaccines with more focus on vaccines tested in large animals, including camels. It is foreseeable that the 1-health approach could be the best way of tackling the MERS-CoV endemic in the Arabian Peninsula, by using the mass vaccination of camels in the affected areas to block camel to human transmission. Camel vaccines can be developed in a faster time with fewer regulations and lower costs and could clear this virus from the Arabian Peninsula if accompanied by efficient public health measures.
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http://dx.doi.org/10.1002/rmv.1917DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7169231PMC
March 2017

Enhancing cellular immunogenicity of MVA-vectored vaccines by utilizing the F11L endogenous promoter.

Vaccine 2016 Jan 24;34(1):49-55. Epub 2015 Nov 24.

The Jenner Institute, University of Oxford, Oxford OX3 7DQ, UK.

Modified vaccinia virus Ankara (MVA)-vectored vaccines against malaria, influenza, tuberculosis and recently Ebola virus are in clinical development. Although this vector is safe and immunogenic in humans, efforts remain on-going to enhance immunogenicity through various approaches such as using stronger promoters to boost transgene expression. We previously reported that endogenous MVA promoters such as pB8 and pF11 increased transgene expression and immunogenicity, as compared to the conventional p7.5 promoter. Here, we show that both promoters also rivalled the mH5 promoter in enhancing MVA immunogenicity. We investigated the mechanisms behind this improved immunogenicity and show that it was a result of strong early transgene expression in vivo, rather than in vitro as would normally be assessed. Moreover, keeping the TK gene intact resulted in a modest improvement in immunogenicity. Utilizing pB8 or pF11 as ectopic promoters at the TK locus instead of their natural loci also increased transgene expression and immunogenicity. In addition to a reporter antigen, the pF11 promoter was tested with the expression of two vaccine antigens for which cellular immunogenicity was significantly increased as compared to the p7.5 promoter. Our data support the use of the pF11 and pB8 promoters for improved immunogenicity in future MVA-vectored candidate vaccines.
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http://dx.doi.org/10.1016/j.vaccine.2015.11.028DOI Listing
January 2016

Deletion of Fifteen Open Reading Frames from Modified Vaccinia Virus Ankara Fails to Improve Immunogenicity.

PLoS One 2015 8;10(6):e0128626. Epub 2015 Jun 8.

The Jenner Institute, University of Oxford, Oxford, OX3 7DQ, United Kingdom.

Modified vaccinia virus Ankara (MVA) is a highly attenuated strain of vaccinia virus, which has been used as a recombinant vaccine vector in many vaccine development programmes. The loss of many immunosuppressive and host-range genes resulted in a safe and immunogenic vaccine vector. However it still retains some immunomodulatory genes that may reduce MVA immunogenicity. Earlier reports demonstrated that the deletion of the A41L, B15R, C6L, or C12L open reading frames (ORFs) enhanced cellular immune responses in recombinant MVA (rMVA) by up to 2-fold. However, previously, we showed that deletion of the C12L, A44L, A46R, B7R, or B15R ORFs from rMVA, using MVA-BAC recombineering technology, did not enhance rMVA immunogenicity at either peak or memory cellular immune responses. Here, we extend our previous study to examine the effect of deleting clusters of genes on rMVA cellular immunogenicity. Two clusters of fifteen genes were deleted in one rMVA mutant that encodes either the 85A antigen of Mycobacterium tuberculosis or an immunodominant H2-Kd-restricted murine malaria epitope (pb9). The deletion mutants were tested in prime only or prime and boost vaccination regimens. The responses showed no improved peak or memory CD8+ T cell frequencies. Our results suggest that the reported small increases in MVA deletion mutants could not be replicated with different antigens, or epitopes. Therefore, the gene deletion strategy may not be taken as a generic approach for improving the immunogenicity of MVA-based vaccines, and should be carefully assessed for every individual recombinant antigen.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0128626PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4459983PMC
February 2016

Investigation of IRES Insertion into the Genome of Recombinant MVA as a Translation Enhancer in the Context of Transcript Decapping.

PLoS One 2015 26;10(5):e0127978. Epub 2015 May 26.

The Jenner Institute, University of Oxford, Oxford, OX3 7DQ, United Kingdom.

Recombinant modified vaccinia virus Ankara (MVA) has been used to deliver vaccine candidate antigens against infectious diseases and cancer. MVA is a potent viral vector for inducing high magnitudes of antigen-specific CD8+ T cells; however the cellular immune responses to a recombinant antigen in MVA could be further enhanced by increasing transgene expression. Previous reports showed the importance of utilizing an early poxviral promoter for increasing transgene expression and therefore enhancing cellular immune responses. However, the vaccinia D10 decapping enzyme is reported to target and decap vaccinia virus early transcripts - a mechanism that could limit the usefulness of early promoters in MVA viral vectors if this enzyme shows the same activity in this closely related virus. Therefore, we attempted to increase transgene expression in recombinant MVA by inserting the encephalomyocarditis virus (EMCV) internal ribosome entry site (IRES) upstream of a transgene sequence that is controlled by the B8R early promoter, and assessed D10 enzyme decapping activity in MVA. The aim of the IRES element was to initiate translation of the transgene transcript (after the removal of the cap structure by the D10 decapping protein) in a cap-independent manner. Here, we report that overexpression of the D10 decapping protein, in trans, in MVA reduced growth and transgene expression; however, the IRES element was not able to compensate for the negative effect of the D10 decapping protein. Recombinant MVA with EMCV IRES induced levels of both gene expression and transcription that were similar to the control recombinant MVA, encoding the same transgene but without the IRES element. Both viruses were tested in BALB/c mice and induced similar magnitudes of epitope-specific CD8+ T cells. This work indicates that the MVA version of the D10 decapping enzyme, overexpressed using a plasmid, is functional, but its negative effect on transgene expression by recombinant MVA cannot be overcome by the use of the EMCV IRES inserted upstream of the transgene initiation codon.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0127978PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4444188PMC
April 2016

Expression and cellular immunogenicity of a transgenic antigen driven by endogenous poxviral early promoters at their authentic loci in MVA.

PLoS One 2012 27;7(6):e40167. Epub 2012 Jun 27.

The Jenner Institute, University of Oxford, Oxford, United Kingdom.

CD8(+) T cell responses to vaccinia virus are directed almost exclusively against early gene products. The attenuated strain modified vaccinia virus Ankara (MVA) is under evaluation in clinical trials of new vaccines designed to elicit cellular immune responses against pathogens including Plasmodium spp., M. tuberculosis and HIV-1. All of these recombinant MVAs (rMVA) utilize the well-established method of linking the gene of interest to a cloned poxviral promoter prior to insertion into the viral genome at a suitable locus by homologous recombination in infected cells. Using BAC recombineering, we show that potent early promoters that drive expression of non-functional or non-essential MVA open reading frames (ORFs) can be harnessed for immunogenic expression of recombinant antigen. Precise replacement of the MVA orthologs of C11R, F11L, A44L and B8R with a model antigen positioned to use the same translation initiation codon allowed early transgene expression similar to or slightly greater than that achieved by the commonly-used p7.5 or short synthetic promoters. The frequency of antigen-specific CD8(+) T cells induced in mice by single shot or adenovirus-prime, rMVA-boost vaccination were similarly equal or marginally enhanced using endogenous promoters at their authentic genomic loci compared to the traditional constructs. The enhancement in immunogenicity observed using the C11R or F11L promoters compared with p7.5 was similar to that obtained with the mH5 promoter compared with p7.5. Furthermore, the growth rates of the viruses were unimpaired and the insertions were genetically stable. Insertion of a transgenic ORF in place of a viral ORF by BAC recombineering can thus provide not only a potent promoter, but also, concomitantly, a suitable insertion site, potentially facilitating development of MVA vaccines expressing multiple recombinant antigens.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0040167PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3384612PMC
January 2013
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