Publications by authors named "Jessica A Plante"

40 Publications

The N501Y spike substitution enhances SARS-CoV-2 infection and transmission.

Nature 2021 Nov 24. Epub 2021 Nov 24.

Institute for Human Infections and Immunity, University of Texas Medical Branch, Galveston, TX, USA.

Beginning in the summer of 2020, a variant of SARS-CoV-2, the cause of the COVID-19 pandemic, emerged in the United Kingdom. This B.1.1.7 variant, also known as Alpha, increased rapidly in prevalence, attributed to an increase in infection and/or transmission efficiency. The Alpha variant has 19 nonsynonymous mutations across its viral genome, including 8 substitutions or deletions in the spike protein, which interacts with cellular receptors to mediate infection and tropism. Here, using a reverse genetics approach, we show that, of the 8 individual spike protein substitutions, only N501Y exhibited consistent fitness gains for replication in the upper airway in the hamster model as well as primary human airway epithelial cells. The N501Y substitution recapitulated the phenotype of enhanced viral transmission seen with the combined 8 Alpha spike mutations, suggesting it is a major determinant of increased transmission of this variant. Mechanistically, the N501Y substitution improved the affinity of the viral spike protein for cellular receptors. As suggested by its convergent evolution in Brazil, South Africa, and elsewhere, our results indicate that N501Y substitution is a major adaptive spike mutation of major concern.
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http://dx.doi.org/10.1038/s41586-021-04245-0DOI Listing
November 2021

Lineage Divergence and Vector-Specific Adaptation Have Driven Chikungunya Virus onto Multiple Adaptive Landscapes.

mBio 2021 Nov 9:e0273821. Epub 2021 Nov 9.

Institute for Human Infections and Immunology, University of Texas Medical Branch, Galveston, Texas, USA.

Previous studies have shown that the adaptation of Indian Ocean lineage (IOL) chikungunya virus (CHIKV) strains for Aedes albopictus transmission was mediated by an E1-A226V substitution, followed by either a single substitution in E2 or synergistic substitutions in the E2 and E3 envelope glycoproteins. Here, we examined whether Asian lineage strains, including those that descended from the 2014 Caribbean introduction, are likely to acquire these A. albopictus-adaptive E2 substitutions. Because Asian lineage strains cannot adapt through the E1-A226V substitution due to an epistatic constraint, we first determined that the beneficial effect of these E2 mutations in IOL strains is independent of E1-A226V. We then introduced each of these E2 adaptive mutations into the Asian lineage backbone to determine if they improve infectivity for . Surprisingly, our results indicated that in the Asian lineage backbone, these E2 mutations significantly decreased CHIKV fitness in . Furthermore, we tested the effects of these mutations in Aedes aegypti and observed different results from those in , suggesting that mosquito species-specific factors that interact with the envelope proteins are involved in vector infection efficiency. Overall, our results indicate that the divergence between Asian lineage and IOL CHIKVs has led them onto different adaptive landscapes with differing potentials to expand their vector host range. Since its introduction into the Caribbean in October 2013, CHIKV has rapidly spread to almost the entire neotropical region. However, its potential to further spread globally, including into more temperate climates, depends in part on its ability to be transmitted efficiently by Aedes albopictus, which can survive colder winters than A. aegypti. We examined in an Asian lineage backbone -adaptive mutations that arose from 2005 to 2009 in Indian Ocean lineage (IOL) strains. Our results predict that the Asian CHIKV lineage now in the Americas will not readily adapt for enhanced transmission via the same mechanisms or adaptive mutations used previously by IOL strains. The vector species- and CHIKV lineage-specific effects caused by adaptive CHIKV envelope glycoprotein substitutions may elucidate our understanding of the mechanisms of mosquito infection and spread.
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http://dx.doi.org/10.1128/mBio.02738-21DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8576524PMC
November 2021

Mouse-adapted SARS-CoV-2 protects animals from lethal SARS-CoV challenge.

PLoS Biol 2021 11 4;19(11):e3001284. Epub 2021 Nov 4.

Departments of Microbiology and Immunology, University of Texas Medical Branch, Galveston, Texas, United States of America.

The emergence of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has resulted in a pandemic causing significant damage to public health and the economy. Efforts to understand the mechanisms of Coronavirus Disease 2019 (COVID-19) have been hampered by the lack of robust mouse models. To overcome this barrier, we used a reverse genetic system to generate a mouse-adapted strain of SARS-CoV-2. Incorporating key mutations found in SARS-CoV-2 variants, this model recapitulates critical elements of human infection including viral replication in the lung, immune cell infiltration, and significant in vivo disease. Importantly, mouse adaptation of SARS-CoV-2 does not impair replication in human airway cells and maintains antigenicity similar to human SARS-CoV-2 strains. Coupled with the incorporation of mutations found in variants of concern, CMA3p20 offers several advantages over other mouse-adapted SARS-CoV-2 strains. Using this model, we demonstrate that SARS-CoV-2-infected mice are protected from lethal challenge with the original Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV), suggesting immunity from heterologous Coronavirus (CoV) strains. Together, the results highlight the use of this mouse model for further study of SARS-CoV-2 infection and disease.
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http://dx.doi.org/10.1371/journal.pbio.3001284DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8594810PMC
November 2021

Nucleocapsid mutations in SARS-CoV-2 augment replication and pathogenesis.

bioRxiv 2021 Oct 15. Epub 2021 Oct 15.

While SARS-CoV-2 continues to adapt for human infection and transmission, genetic variation outside of the spike gene remains largely unexplored. This study investigates a highly variable region at residues 203-205 in SARS-CoV-2 nucleocapsid protein. Recreating the alpha variant mutation in an early pandemic (WA-1) background, we found that the R203K/G204R mutation is sufficient to enhance replication, fitness, and pathogenesis of SARS-CoV-2. Importantly, the R203K/G204R mutation increases nucleocapsid phosphorylation, providing a molecular basis for these phenotypes. Notably, an analogous alanine substitution mutant also increases SARS-CoV-2 fitness and phosphorylation, suggesting that infection is enhanced through ablation of the ancestral 'RG' motif. Overall, these results demonstrate that variant mutations outside spike are also key components in SARS-CoV-2's continued adaptation to human infection.

One-sentence Summary: A mutation in the nucleocapsid gene of the SARS-CoV-2 alpha variant is found to enhance replication, fitness, and pathogenesis.
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http://dx.doi.org/10.1101/2021.10.14.464390DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8528077PMC
October 2021

Tiled-ClickSeq for targeted sequencing of complete coronavirus genomes with simultaneous capture of RNA recombination and minority variants.

Elife 2021 09 28;10. Epub 2021 Sep 28.

Department of Biochemistry and Molecular Biology, The University of Texas Medical Branch, Galveston, United States.

High-throughput genomics of SARS-CoV-2 is essential to characterize virus evolution and to identify adaptations that affect pathogenicity or transmission. While single-nucleotide variations (SNVs) are commonly considered as driving virus adaption, RNA recombination events that delete or insert nucleic acid sequences are also critical. Whole genome targeting sequencing of SARS-CoV-2 is typically achieved using pairs of primers to generate cDNA amplicons suitable for next-generation sequencing (NGS). However, paired-primer approaches impose constraints on where primers can be designed, how many amplicons are synthesized and requires multiple PCR reactions with non-overlapping primer pools. This imparts sensitivity to underlying SNVs and fails to resolve RNA recombination junctions that are not flanked by primer pairs. To address these limitations, we have designed an approach called , which uses hundreds of tiled-primers spaced evenly along the virus genome in a single reverse-transcription reaction. The other end of the cDNA amplicon is generated by azido-nucleotides that stochastically terminate cDNA synthesis, removing the need for a paired-primer. A sequencing adaptor containing a Unique Molecular Identifier (UMI) is appended to the cDNA fragment using click-chemistry and a PCR reaction generates a final NGS library. Tiled-ClickSeq provides complete genome coverage, including the 5'UTR, at high depth and specificity to the virus on both Illumina and Nanopore NGS platforms. Here, we analyze multiple SARS-CoV-2 isolates and clinical samples to simultaneously characterize minority variants, sub-genomic mRNAs (sgmRNAs), structural variants (SVs) and D-RNAs. Tiled-ClickSeq therefore provides a convenient and robust platform for SARS-CoV-2 genomics that captures the full range of RNA species in a single, simple assay.
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http://dx.doi.org/10.7554/eLife.68479DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8478411PMC
September 2021

Characterization of a Dengue Virus Serotype 1 Isolated from a Patient in Ciudad Juarez, Mexico.

Pathogens 2021 Jul 10;10(7). Epub 2021 Jul 10.

Department of Biological Sciences and Border Biomedical Research Center, University of Texas at El Paso, El Paso, TX 79968, USA.

Dengue (DEN) is the most important human arboviral disease worldwide. Sporadic outbreaks of DEN have been reported since 1980 in urban communities located along the border in southeast Texas and northern Mexico. Other than the Rio Grande Valley region of TX, autochthonous transmission of DENV has not been reported from any other US border communities. As part of a surveillance program for arthropod-borne viruses in Ciudad Juarez, Mexico, during November 2015, a blood sample was obtained from a female patient who experienced an undifferentiated fever and arthralgia. The plasma of the sample was tested for virus in Vero-76 and C6/36 cells. DENV serotype 1 (DENV-1) was isolated in the C6/36 cells, and nucleotide sequencing of the envelope gene and full genome grouped the DENV-1 isolate in the Central America clade. The patient had not traveled outside of Ciudad Juarez, Mexico, thus suggesting DENV-1 infection was acquired in this community.
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http://dx.doi.org/10.3390/pathogens10070872DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8308707PMC
July 2021

Mouse Adapted SARS-CoV-2 protects animals from lethal SARS-CoV challenge.

bioRxiv 2021 May 4. Epub 2021 May 4.

Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA.

The emergence of SARS-CoV-2 has resulted in a worldwide pandemic causing significant damage to public health and the economy. Efforts to understand the mechanisms of COVID-19 disease have been hampered by the lack of robust mouse models. To overcome this barrier, we utilized a reverse genetic system to generate a mouse-adapted strain of SARS-CoV-2. Incorporating key mutations found in SARSCoV-2 variants, this model recapitulates critical elements of human infection including viral replication in the lung, immune cell infiltration, and significant disease. Importantly, mouse-adaptation of SARS-CoV-2 does not impair replication in human airway cells and maintains antigenicity similar to human SARS-CoV-2 strains. Utilizing this model, we demonstrate that SARS-CoV-2 infected mice are protected from lethal challenge with the original SARS-CoV, suggesting immunity from heterologous CoV strains. Together, the results highlight the utility of this mouse model for further study of SARS-CoV-2 infection and disease.
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http://dx.doi.org/10.1101/2021.05.03.442357DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8109199PMC
May 2021

Adjuvanting a subunit COVID-19 vaccine to induce protective immunity.

Nature 2021 06 19;594(7862):253-258. Epub 2021 Apr 19.

Tulane National Primate Research Center, Covington, LA, USA.

The development of a portfolio of COVID-19 vaccines to vaccinate the global population remains an urgent public health imperative. Here we demonstrate the capacity of a subunit vaccine, comprising the SARS-CoV-2 spike protein receptor-binding domain displayed on an I53-50 protein nanoparticle scaffold (hereafter designated RBD-NP), to stimulate robust and durable neutralizing-antibody responses and protection against SARS-CoV-2 in rhesus macaques. We evaluated five adjuvants including Essai O/W 1849101, a squalene-in-water emulsion; AS03, an α-tocopherol-containing oil-in-water emulsion; AS37, a Toll-like receptor 7 (TLR7) agonist adsorbed to alum; CpG1018-alum, a TLR9 agonist formulated in alum; and alum. RBD-NP immunization with AS03, CpG1018-alum, AS37 or alum induced substantial neutralizing-antibody and CD4 T cell responses, and conferred protection against SARS-CoV-2 infection in the pharynges, nares and bronchoalveolar lavage. The neutralizing-antibody response to live virus was maintained up to 180 days after vaccination with RBD-NP in AS03 (RBD-NP-AS03), and correlated with protection from infection. RBD-NP immunization cross-neutralized the B.1.1.7 SARS-CoV-2 variant efficiently but showed a reduced response against the B.1.351 variant. RBD-NP-AS03 produced a 4.5-fold reduction in neutralization of B.1.351 whereas the group immunized with RBD-NP-AS37 produced a 16-fold reduction in neutralization of B.1.351, suggesting differences in the breadth of the neutralizing-antibody response induced by these adjuvants. Furthermore, RBD-NP-AS03 was as immunogenic as a prefusion-stabilized spike immunogen (HexaPro) with AS03 adjuvant. These data highlight the efficacy of the adjuvanted RBD-NP vaccine in promoting protective immunity against SARS-CoV-2 and have led to phase I/II clinical trials of this vaccine (NCT04742738 and NCT04750343).
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http://dx.doi.org/10.1038/s41586-021-03530-2DOI Listing
June 2021

The variant gambit: COVID-19's next move.

Cell Host Microbe 2021 04 1;29(4):508-515. Epub 2021 Mar 1.

Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, USA; World Reference Center for Emerging Viruses and Arboviruses, University of Texas Medical Branch, Galveston, TX, USA; Institute for Human Infections and Immunity, University of Texas Medical Branch, Galveston, TX, USA. Electronic address:

More than a year after its emergence, COVID-19, the disease caused by SARS-CoV-2, continues to plague the world and dominate our daily lives. Even with the development of effective vaccines, this coronavirus pandemic continues to cause a fervor with the identification of major new variants hailing from the United Kingdom, South Africa, Brazil, and California. Coupled with worries over a distinct mink strain that has caused human infections and potential for further mutations, SARS-CoV-2 variants bring concerns for increased spread and escape from both vaccine and natural infection immunity. Here, we outline factors driving SARS-CoV-2 variant evolution, explore the potential impact of specific mutations, examine the risk of further mutations, and consider the experimental studies needed to understand the threat these variants pose. In this review, Plante et al. examine SARS-CoV-2 variants including B.1.1.7 (UK), B.1.351 (RSA), P.1 (Brazil), and B.1.429 (California). They focus on what factors contribute to variant emergence, mutations in and outside the spike protein, and studies needed to understand the impact of variants on infection, transmission, and vaccine efficacy.
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http://dx.doi.org/10.1016/j.chom.2021.02.020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7919536PMC
April 2021

Tiled-ClickSeq for targeted sequencing of complete coronavirus genomes with simultaneous capture of RNA recombination and minority variants.

bioRxiv 2021 Mar 11. Epub 2021 Mar 11.

Department of Biochemistry and Molecular Biology, The University of Texas Medical Branch, Galveston, TX, USA.

High-throughput genomics of SARS-CoV-2 is essential to characterize virus evolution and to identify adaptations that affect pathogenicity or transmission. While single-nucleotide variations (SNVs) are commonly considered as driving virus adaption, RNA recombination events that delete or insert nucleic acid sequences are also critical. Whole genome targeting sequencing of SARS-CoV-2 is typically achieved using pairs of primers to generate cDNA amplicons suitable for Next-Generation Sequencing (NGS). However, paired-primer approaches impose constraints on where primers can be designed, how many amplicons are synthesized and requires multiple PCR reactions with non-overlapping primer pools. This imparts sensitivity to underlying SNVs and fails to resolve RNA recombination junctions that are not flanked by primer pairs. To address these limitations, we have designed an approach called ' '. Tiled-ClickSeq uses hundreds of tiled-primers spaced evenly along the virus genome in a single reverse-transcription reaction. The other end of the cDNA amplicon is generated by azido-nucleotides that stochastically terminate cDNA synthesis, obviating the need for a paired-primer. A sequencing adaptor containing a Unique Molecular Identifier (UMI) is appended using click-chemistry and a PCR reaction using Illumina adaptors generates a final NGS library. Tiled-ClickSeq provides complete genome coverage, including the 5'UTR, at high depth and specificity to virus on both Illumina and Nanopore NGS platforms. Here, we analyze multiple SARS-CoV-2 isolates and simultaneously characterize minority variants, sub-genomic mRNAs (sgmRNAs), structural variants (SVs) and D-RNAs. Tiled-ClickSeq therefore provides a convenient and robust platform for SARS-CoV-2 genomics that captures the full range of RNA species in a single, simple assay.
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http://dx.doi.org/10.1101/2021.03.10.434828DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7987005PMC
March 2021

The N501Y spike substitution enhances SARS-CoV-2 transmission.

bioRxiv 2021 Mar 9. Epub 2021 Mar 9.

Beginning in the summer of 2020, a variant of SARS-CoV-2, the cause of the COVID-19 pandemic, emerged in the United Kingdom (UK). This B.1.1.7 variant increased rapidly in prevalence among sequenced strains, attributed to an increase in infection and/or transmission efficiency. The UK variant has 19 nonsynonymous mutations across its viral genome including 8 substitutions or deletions in the spike protein, which interacts with cellular receptors to mediate infection and tropism. Here, using a reverse genetics approach, we show that, of the 8 individual spike protein substitutions, only N501Y exhibited consistent fitness gains for replication in the upper airway in the hamster model as well as primary human airway epithelial cells. The N501Y substitution recapitulated the phenotype of enhanced viral transmission seen with the combined 8 UK spike mutations, suggesting it is a major determinant responsible for increased transmission of this variant. Mechanistically, the N501Y substitution improved the affinity of the viral spike protein for cellular receptors. As suggested by its convergent evolution in Brazil and South Africa, our results indicate that N501Y substitution is a major adaptive spike mutation of major concern.
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http://dx.doi.org/10.1101/2021.03.08.434499DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7986995PMC
March 2021

Outbreak of coronavirus disease 2019 (COVID-19) among operating room staff of a tertiary referral center: An epidemiologic and environmental investigation.

Infect Control Hosp Epidemiol 2021 Mar 19:1-7. Epub 2021 Mar 19.

Department of Infection Control and Healthcare Epidemiology, University of Texas Medical Branch, Galveston, Texas.

Objective: Investigate an outbreak of coronavirus disease 2019 (COVID-19) among operating room staff utilizing contact tracing, mass testing for severe acute respiratory coronavirus virus 2 (SARS-CoV-2), and environmental sampling.

Design: Outbreak investigation.

Setting: University-affiliated tertiary-care referral center.

Patients: Operating room staff with positive SARS-CoV-2 molecular testing.

Methods: Epidemiologic and environmental investigations were conducted including contact tracing, environmental surveys, and sampling and review of the operating room schedule for staff-to-staff, staff-to-patient, and patient-to-staff SARS-CoV-2 transmission.

Results: In total, 24 healthcare personnel (HCP) tested positive for SARS-CoV-2, including nurses (29%), surgical technologists (25%), and surgical residents (16%). Moreover, 19 HCP (79%) reported having used a communal area, most commonly break rooms (75%). Overall, 20 HCP (83%) reported symptomatic disease. In total, 72 environmental samples were collected from communal areas for SARS-CoV-2 genomic testing; none was positive. Furthermore, 236 surgical cases were reviewed for transmission: 213 (90%) had negative preoperative SARS-CoV-2 testing, 21 (9%) had a positive test on or before the date of surgery, and 2 (<1%) did not have a preoperative test performed. In addition, 40 patients underwent postoperative testing (mean, 13 days to postoperative testing), and 2 returned positive results. Neither of these 2 cases was linked to our outbreak.

Conclusions: Complacency in infection control practices among staff during peak community transmission of SARS-CoV-2 is believed to have driven staff-to-staff transmission. Prompt identification of the outbreak led to rapid interventions, ultimately allowing for uninterrupted surgical service.
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http://dx.doi.org/10.1017/ice.2021.116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8042383PMC
March 2021

Antiviral activity of oleandrin and a defined extract of Nerium oleander against SARS-CoV-2.

Biomed Pharmacother 2021 Jun 3;138:111457. Epub 2021 Mar 3.

Department of Experimental Therapeutics, University of Texas MD Anderson Cancer Center, Houston, TX 77030, USA; Phoenix Biotechnology, Inc., San Antonio, TX 78217, USA. Electronic address:

With continued expansion of the coronavirus disease (COVID-19) pandemic, caused by severe acute respiratory syndrome 2 (SARS-CoV-2), both antiviral drugs as well as effective vaccines are desperately needed to treat patients at high risk of life-threatening disease. Here, we present in vitro evidence for significant inhibition of SARS-CoV-2 by oleandrin and a defined extract of N. oleander (designated as PBI-06150). Using Vero cells, we found that prophylactic (pre-infection) oleandrin (as either the pure compound or as the active principal ingredient in PBI-06150) administration at concentrations as low as 0.05 µg/ml exhibited potent antiviral activity against SARS-CoV-2, with an 800-fold reduction in virus production, and a 0.1 µg/ml concentration resulted in a greater than 3000-fold reduction in infectious virus production. The half maximal effective concentration (EC) values were 11.98 ng/ml when virus output was measured at 24 h post-infection, and 7.07 ng/ml measured at 48 h post-infection. Therapeutic (post-infection) treatment up to 24 h after SARS-CoV-2 infection of Vero cells also reduced viral titers, with 0.1 µg/ml and 0.05 µg/ml concentrations causing greater than 100-fold reduction as measured at 48 h, and the 0.05 µg/ml concentration resulting in a 78-fold reduction. Concentrations of oleandrin up to 10 µg/ml were well tolerated in Vero cells. We also present in vivo evidence of the safety and efficacy of defined N. oleander extract (PBI-06150), which was administered to golden Syrian hamsters in a preparation containing as high as 130 µg/ml of oleandrin. In comparison to administration of control vehicle, PBI-06150 provided a statistically significant reduction of the viral titer in the nasal turbinates (nasal conchae). The potent prophylactic and therapeutic antiviral activities demonstrated here, together with initial evidence of its safety and efficacy in a relevant hamster model of COVID-19, support the further development of oleandrin and/or defined extracts containing this molecule for the treatment of SARS-CoV-2 and associated COVID-19 disease and potentially also for reduction of virus spread by persons diagnosed early after infection.
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http://dx.doi.org/10.1016/j.biopha.2021.111457DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7927596PMC
June 2021

A trans-complementation system for SARS-CoV-2 recapitulates authentic viral replication without virulence.

Cell 2021 04 23;184(8):2229-2238.e13. Epub 2021 Feb 23.

Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX, USA; Institute for Human Infections and Immunity, University of Texas Medical Branch, Galveston, TX, USA; Sealy Institute for Vaccine Sciences, University of Texas Medical Branch, Galveston, TX, USA; Sealy Center for Structural Biology & Molecular Biophysics, University of Texas Medical Branch, Galveston, TX, USA. Electronic address:

The biosafety level 3 (BSL-3) requirement to culture severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a bottleneck for research. Here, we report a trans-complementation system that produces single-round infectious SARS-CoV-2 that recapitulates authentic viral replication. We demonstrate that the single-round infectious SARS-CoV-2 can be used at BSL-2 laboratories for high-throughput neutralization and antiviral testing. The trans-complementation system consists of two components: a genomic viral RNA containing ORF3 and envelope gene deletions, as well as mutated transcriptional regulator sequences, and a producer cell line expressing the two deleted genes. Trans-complementation of the two components generates virions that can infect naive cells for only one round but does not produce wild-type SARS-CoV-2. Hamsters and K18-hACE2 transgenic mice inoculated with the complementation-derived virions exhibited no detectable disease, even after intracranial inoculation with the highest possible dose. Thus, the trans-complementation platform can be safely used at BSL-2 laboratories for research and countermeasure development.
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http://dx.doi.org/10.1016/j.cell.2021.02.044DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7901297PMC
April 2021

Adjuvanting a subunit SARS-CoV-2 nanoparticle vaccine to induce protective immunity in non-human primates.

bioRxiv 2021 Feb 11. Epub 2021 Feb 11.

The development of a portfolio of SARS-CoV-2 vaccines to vaccinate the global population remains an urgent public health imperative. Here, we demonstrate the capacity of a subunit vaccine under clinical development, comprising the SARS-CoV-2 Spike protein receptor-binding domain displayed on a two-component protein nanoparticle (RBD-NP), to stimulate robust and durable neutralizing antibody (nAb) responses and protection against SARS-CoV-2 in non-human primates. We evaluated five different adjuvants combined with RBD-NP including Essai O/W 1849101, a squalene-in-water emulsion; AS03, an alpha-tocopherol-containing squalene-based oil-in-water emulsion used in pandemic influenza vaccines; AS37, a TLR-7 agonist adsorbed to Alum; CpG 1018-Alum (CpG-Alum), a TLR-9 agonist formulated in Alum; or Alum, the most widely used adjuvant. All five adjuvants induced substantial nAb and CD4 T cell responses after two consecutive immunizations. Durable nAb responses were evaluated for RBD-NP/AS03 immunization and the live-virus nAb response was durably maintained up to 154 days post-vaccination. AS03, CpG-Alum, AS37 and Alum groups conferred significant protection against SARS-CoV-2 infection in the pharynges, nares and in the bronchoalveolar lavage. The nAb titers were highly correlated with protection against infection. Furthermore, RBD-NP when used in conjunction with AS03 was as potent as the prefusion stabilized Spike immunogen, HexaPro. Taken together, these data highlight the efficacy of the RBD-NP formulated with clinically relevant adjuvants in promoting robust immunity against SARS-CoV-2 in non-human primates.
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http://dx.doi.org/10.1101/2021.02.10.430696DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7885918PMC
February 2021

A -complementation system for SARS-CoV-2.

bioRxiv 2021 Jan 19. Epub 2021 Jan 19.

The biosafety level-3 (BSL-3) requirement to culture severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a bottleneck for research and countermeasure development. Here we report a -complementation system that produces single-round infectious SARS-CoV-2 that recapitulates authentic viral replication. We demonstrate that the single-round infectious SARS-CoV-2 can be used at BSL-2 laboratories for high-throughput neutralization and antiviral testing. The -complementation system consists of two components: a genomic viral RNA containing a deletion of ORF3 and envelope gene, and a producer cell line expressing the two deleted genes. complementation of the two components generates virions that can infect naive cells for only one round, but does not produce wild-type SARS-CoV-2. Hamsters and K18-hACE2 transgenic mice inoculated with the complementation-derived virions exhibited no detectable disease, even after intracranial inoculation with the highest possible dose. The results suggest that the -complementation platform can be safely used at BSL-2 laboratories for research and countermeasure development.
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http://dx.doi.org/10.1101/2021.01.16.426970DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7836106PMC
January 2021

Loss of furin cleavage site attenuates SARS-CoV-2 pathogenesis.

Nature 2021 03 25;591(7849):293-299. Epub 2021 Jan 25.

Department of Pathology and Immunology, Washington University School of Medicine, St Louis, MO, USA.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-a new coronavirus that has led to a worldwide pandemic-has a furin cleavage site (PRRAR) in its spike protein that is absent in other group-2B coronaviruses. To explore whether the furin cleavage site contributes to infection and pathogenesis in this virus, we generated a mutant SARS-CoV-2 that lacks the furin cleavage site (ΔPRRA). Here we report that replicates of ΔPRRA SARS-CoV-2 had faster kinetics, improved fitness in Vero E6 cells and reduced spike protein processing, as compared to parental SARS-CoV-2. However, the ΔPRRA mutant had reduced replication in a human respiratory cell line and was attenuated in both hamster and K18-hACE2 transgenic mouse models of SARS-CoV-2 pathogenesis. Despite reduced disease, the ΔPRRA mutant conferred protection against rechallenge with the parental SARS-CoV-2. Importantly, the neutralization values of sera from patients with coronavirus disease 2019 (COVID-19) and monoclonal antibodies against the receptor-binding domain of SARS-CoV-2 were lower against the ΔPRRA mutant than against parental SARS-CoV-2, probably owing to an increased ratio of particles to plaque-forming units in infections with the former. Together, our results demonstrate a critical role for the furin cleavage site in infection with SARS-CoV-2 and highlight the importance of this site for evaluating the neutralization activities of antibodies.
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http://dx.doi.org/10.1038/s41586-021-03237-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8175039PMC
March 2021

Acute Respiratory Distress in Aged, SARS-CoV-2-Infected African Green Monkeys but Not Rhesus Macaques.

Am J Pathol 2021 02 7;191(2):274-282. Epub 2020 Nov 7.

Tulane National Primate Research Center, Covington, Louisiana; Department of Microbiology and Immunology, Tulane University School of Medicine, New Orleans, Louisiana.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) induces a wide range of disease severity, ranging from asymptomatic infection to a life-threating illness, particularly in the elderly population and individuals with comorbid conditions. Among individuals with serious coronavirus 2019 (COVID-19) disease, acute respiratory distress syndrome (ARDS) is a common and often fatal presentation. Animal models of SARS-CoV-2 infection that manifest severe disease are needed to investigate the pathogenesis of COVID-19-induced ARDS and evaluate therapeutic strategies. We report two cases of ARDS in two aged African green monkeys (AGMs) infected with SARS-CoV-2 that had pathological lesions and disease similar to severe COVID-19 in humans. We also report a comparatively mild COVID-19 phenotype characterized by minor clinical, radiographic, and histopathologic changes in the two surviving, aged AGMs and four rhesus macaques (RMs) infected with SARS-CoV-2. Notable increases in circulating cytokines were observed in three of four infected, aged AGMs but not in infected RMs. All the AGMs had increased levels of plasma IL-6 compared with baseline, a predictive marker and presumptive therapeutic target in humans infected with SARS-CoV-2. Together, our results indicate that both RMs and AGMs are capable of modeling SARS-CoV-2 infection and suggest that aged AGMs may be useful for modeling severe disease manifestations, including ARDS.
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http://dx.doi.org/10.1016/j.ajpath.2020.10.016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7648506PMC
February 2021

Spike mutation D614G alters SARS-CoV-2 fitness.

Nature 2021 04 26;592(7852):116-121. Epub 2020 Oct 26.

Institute for Human Infections and Immunity, University of Texas Medical Branch, Galveston, TX, USA.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein substitution D614G became dominant during the coronavirus disease 2019 (COVID-19) pandemic. However, the effect of this variant on viral spread and vaccine efficacy remains to be defined. Here we engineered the spike D614G substitution in the USA-WA1/2020 SARS-CoV-2 strain, and found that it enhances viral replication in human lung epithelial cells and primary human airway tissues by increasing the infectivity and stability of virions. Hamsters infected with SARS-CoV-2 expressing spike(D614G) (G614 virus) produced higher infectious titres in nasal washes and the trachea, but not in the lungs, supporting clinical evidence showing that the mutation enhances viral loads in the upper respiratory tract of COVID-19 patients and may increase transmission. Sera from hamsters infected with D614 virus exhibit modestly higher neutralization titres against G614 virus than against D614 virus, suggesting that the mutation is unlikely to reduce the ability of vaccines in clinical trials to protect against COVID-19, and that therapeutic antibodies should be tested against the circulating G614 virus. Together with clinical findings, our work underscores the importance of this variant in viral spread and its implications for vaccine efficacy and antibody therapy.
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http://dx.doi.org/10.1038/s41586-020-2895-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8158177PMC
April 2021

Spike mutation D614G alters SARS-CoV-2 fitness and neutralization susceptibility.

bioRxiv 2020 Sep 2. Epub 2020 Sep 2.

Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston TX, USA.

A spike protein mutation D614G became dominant in SARS-CoV-2 during the COVID-19 pandemic. However, the mutational impact on viral spread and vaccine efficacy remains to be defined. Here we engineer the D614G mutation in the SARS-CoV-2 USA-WA1/2020 strain and characterize its effect on viral replication, pathogenesis, and antibody neutralization. The D614G mutation significantly enhances SARS-CoV-2 replication on human lung epithelial cells and primary human airway tissues, through an improved infectivity of virions with the spike receptor-binding domain in an "up" conformation for binding to ACE2 receptor. Hamsters infected with D614 or G614 variants developed similar levels of weight loss. However, the G614 virus produced higher infectious titers in the nasal washes and trachea, but not lungs, than the D614 virus. The hamster results confirm clinical evidence that the D614G mutation enhances viral loads in the upper respiratory tract of COVID-19 patients and may increases transmission. For antibody neutralization, sera from D614 virus-infected hamsters consistently exhibit higher neutralization titers against G614 virus than those against D614 virus, indicating that (i) the mutation may not reduce the ability of vaccines in clinical trials to protect against COVID-19 and (ii) therapeutic antibodies should be tested against the circulating G614 virus before clinical development.

Importance: Understanding the evolution of SARS-CoV-2 during the COVID-19 pandemic is essential for disease control and prevention. A spike protein mutation D614G emerged and became dominant soon after the pandemic started. By engineering the D614G mutation into an authentic wild-type SARS-CoV-2 strain, we demonstrate the importance of this mutation to (i) enhanced viral replication on human lung epithelial cells and primary human airway tissues, (ii) improved viral fitness in the upper airway of infected hamsters, and (iii) increased susceptibility to neutralization. Together with clinical findings, our work underscores the importance of this mutation in viral spread, vaccine efficacy, and antibody therapy.
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http://dx.doi.org/10.1101/2020.09.01.278689DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7480025PMC
September 2020

Furin Cleavage Site Is Key to SARS-CoV-2 Pathogenesis.

bioRxiv 2020 Aug 26. Epub 2020 Aug 26.

SARS-CoV-2 has resulted in a global pandemic and shutdown economies around the world. Sequence analysis indicates that the novel coronavirus (CoV) has an insertion of a furin cleavage site (PRRAR) in its spike protein. Absent in other group 2B CoVs, the insertion may be a key factor in the replication and virulence of SARS-CoV-2. To explore this question, we generated a SARS-CoV-2 mutant lacking the furin cleavage site (ΔPRRA) in the spike protein. This mutant virus replicated with faster kinetics and improved fitness in Vero E6 cells. The mutant virus also had reduced spike protein processing as compared to wild-type SARS-CoV-2. In contrast, the ΔPRRA had reduced replication in Calu3 cells, a human respiratory cell line, and had attenuated disease in a hamster pathogenesis model. Despite the reduced disease, the ΔPRRA mutant offered robust protection from SARS-CoV-2 rechallenge. Importantly, plaque reduction neutralization tests (PRNT ) with COVID-19 patient sera and monoclonal antibodies against the receptor-binding domain found a shift, with the mutant virus resulting in consistently reduced PRNT titers. Together, these results demonstrate a critical role for the furin cleavage site insertion in SARS-CoV-2 replication and pathogenesis. In addition, these findings illustrate the importance of this insertion in evaluating neutralization and other downstream SARS-CoV-2 assays.

Importance: As COVID-19 has impacted the world, understanding how SARS-CoV-2 replicates and causes virulence offers potential pathways to disrupt its disease. By removing the furin cleavage site, we demonstrate the importance of this insertion to SARS-CoV-2 replication and pathogenesis. In addition, the findings with Vero cells indicate the likelihood of cell culture adaptations in virus stocks that can influence reagent generation and interpretation of a wide range of data including neutralization and drug efficacy. Overall, our work highlights the importance of this key motif in SARS-CoV-2 infection and pathogenesis.

Article Summary: A deletion of the furin cleavage site in SARS-CoV-2 amplifies replication in Vero cells, but attenuates replication in respiratory cells and pathogenesis in vivo. Loss of the furin site also reduces susceptibility to neutralization .
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http://dx.doi.org/10.1101/2020.08.26.268854DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7457603PMC
August 2020

High Seroprevalence of Dengue Virus Infection in Sudan: Systematic Review and Meta-Analysis.

Trop Med Infect Dis 2020 Jul 18;5(3). Epub 2020 Jul 18.

World Reference Center for Emerging Viruses and Arboviruses, University of Texas Medical Branch, Galveston, TX 77555, USA.

The goal of this study was to systematically review the published data on dengue virus (DENV) seroprevalence in Sudan and to estimate disease burden through meta-analysis. We searched, reviewed, and extracted online available reports on DENV in Sudan. Among 168 identified records, 19 were selected. Dengue infections were documented in 11/18 states. The overall seroprevalence of DENV in Sudan was estimated to be 27%, while the prevalence of dengue IgM was 22% and IgG was 38%. The prevalence of dengue estimated from community and hospital-based cross-sectional studies were 26% and 30% respectively. Additionally, one cohort study and a single PCR-based study reported a prevalence of 1% and 4%, respectively. Regional analysis revealed that the variation in seroprevalence in East, North, West, and Central Sudan was 23%, 24%, 36% and 43%, respectively. Interestingly, we found that DENV is circulating countrywide with a significant spatiotemporal variation in the disease seroprevalence. Furthermore, publications on dengue prevalence are temporally and geographically fragmented, perhaps due to limited resources. However, this gap in data and knowledge highlights the urgent need for a country-wide surveillance system and continued study of dengue burden in Sudan to accurately estimate the disease prevalence and determine the associated risk factors.
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http://dx.doi.org/10.3390/tropicalmed5030120DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7559303PMC
July 2020

Prophylactic and Therapeutic Inhibition of In Vitro SARS-CoV-2 Replication by Oleandrin.

bioRxiv 2020 Jul 15. Epub 2020 Jul 15.

With continued expansion of the COVID-19 pandemic, antiviral drugs are desperately needed to treat patients at high risk of life-threatening disease and even to limit spread if administered early during infection. Typically, the fastest route to identifying and licensing a safe and effective antiviral drug is to test those already shown safe in early clinical trials for other infections or diseases. Here, we tested oleandrin, derived from the plant and shown previously to have inhibitory activity against several viruses. Using Vero cells, we found that prophylactic oleandrin administration at concentrations down to 0.05 μg/ml exhibited potent antiviral activity against SARS-CoV-2, with an 800-fold reduction in virus production, and a 0.1 μg/ml dose resulted in a greater than 3,000-fold reduction in infectious virus production. The EC values were 11.98ng/ml when virus output was measured at 24 hours post-infection, and 7.07ng/ml measured at 48 hours post-infection. Therapeutic (post-infection) treatment up to 24 hours after infection of Vero cells also reduced viral titers, with the 0.1 μg/ml dose causing greater than 100-fold reductions as measured at 48 hours, and the 0.05 μg/ml dose resulting in a 78-fold reduction. The potent prophylactic and therapeutic antiviral activities demonstrated here strongly support the further development of oleandrin to reduce the severity of COVID-19 and potentially also to reduce spread by persons diagnosed early after infection.

Importance: COVID-19, a pandemic disease caused by infection with SARS-CoV-2, has swept around the world to cause millions of infections and hundreds-of-thousands of deaths due to the lack of vaccines and effective therapeutics. We tested oleandrin, derived from the plant and shown previously to reduce the replication of several viruses, against SARS-CoV-2 infection of Vero cells. When administered both before and after virus infection, nanogram doses of oleandrin significantly inhibited replication by up to 3,000-fold, indicating the potential to prevent disease and virus spread in persons recently exposed to SARS-CoV-2, as well as to prevent severe disease in persons at high risk. These results indicate that oleandrin should be tested in animal models and in humans exposed to infection to determine its medical usefulness in controlling the pandemic.
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http://dx.doi.org/10.1101/2020.07.15.203489DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7373128PMC
July 2020

Isolation and characterization of SARS-CoV-2 from the first US COVID-19 patient.

bioRxiv 2020 Mar 7. Epub 2020 Mar 7.

Department of Microbiology and Immunology, Institute for Human Infection and Immunity, University of Texas Medical Branch, Galveston TX, USA.

The etiologic agent of the outbreak of pneumonia in Wuhan China was identified as severe acute respiratory syndrome associated coronavirus 2 (SARS-CoV-2) in January, 2020. The first US patient was diagnosed by the State of Washington and the US Centers for Disease Control and Prevention on January 20, 2020. We isolated virus from nasopharyngeal and oropharyngeal specimens, and characterized the viral sequence, replication properties, and cell culture tropism. We found that the virus replicates to high titer in Vero-CCL81 cells and Vero E6 cells in the absence of trypsin. We also deposited the virus into two virus repositories, making it broadly available to the public health and research communities. We hope that open access to this important reagent will expedite development of medical countermeasures.
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http://dx.doi.org/10.1101/2020.03.02.972935DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7239045PMC
March 2020

Severe Acute Respiratory Syndrome Coronavirus 2 from Patient with Coronavirus Disease, United States.

Emerg Infect Dis 2020 06 17;26(6):1266-1273. Epub 2020 Jun 17.

The etiologic agent of an outbreak of pneumonia in Wuhan, China, was identified as severe acute respiratory syndrome coronavirus 2 in January 2020. A patient in the United States was given a diagnosis of infection with this virus by the state of Washington and the US Centers for Disease Control and Prevention on January 20, 2020. We isolated virus from nasopharyngeal and oropharyngeal specimens from this patient and characterized the viral sequence, replication properties, and cell culture tropism. We found that the virus replicates to high titer in Vero-CCL81 cells and Vero E6 cells in the absence of trypsin. We also deposited the virus into 2 virus repositories, making it broadly available to the public health and research communities. We hope that open access to this reagent will expedite development of medical countermeasures.
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http://dx.doi.org/10.3201/eid2606.200516DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7258473PMC
June 2020

Peptidoglycan-Associated Cyclic Lipopeptide Disrupts Viral Infectivity.

J Virol 2019 11 29;93(22). Epub 2019 Oct 29.

Department of Microbiology and Immunology, University of Texas Medical Branch at Galveston, Galveston, Texas, USA

Enteric viruses exploit bacterial components, including lipopolysaccharides (LPS) and peptidoglycan (PG), to facilitate infection in humans. Because of their origin in the bat enteric system, we wondered if severe acute respiratory syndrome coronavirus (SARS-CoV) or Middle East respiratory syndrome CoV (MERS-CoV) also use bacterial components to modulate infectivity. To test this question, we incubated CoVs with LPS and PG and evaluated infectivity, finding no change following LPS treatment. However, PG from reduced infection >10,000-fold, while PG from other bacterial species failed to recapitulate this. Treatment with an alcohol solvent transferred inhibitory activity to the wash, and mass spectrometry revealed surfactin, a cyclic lipopeptide antibiotic, as the inhibitory compound. This antibiotic had robust dose- and temperature-dependent inhibition of CoV infectivity. Mechanistic studies indicated that surfactin disrupts CoV virion integrity, and surfactin treatment of the virus inoculum ablated infection Finally, similar cyclic lipopeptides had no effect on CoV infectivity, and the inhibitory effect of surfactin extended broadly to enveloped viruses, including influenza, Ebola, Zika, Nipah, chikungunya, Una, Mayaro, Dugbe, and Crimean-Congo hemorrhagic fever viruses. Overall, our results indicate that peptidoglycan-associated surfactin has broad viricidal activity and suggest that bacteria by-products may negatively modulate virus infection. In this article, we consider a role for bacteria in shaping coronavirus infection. Taking cues from studies of enteric viruses, we initially investigated how bacterial surface components might improve CoV infection. Instead, we found that peptidoglycan-associated surfactin is a potent viricidal compound that disrupts virion integrity with broad activity against enveloped viruses. Our results indicate that interactions with commensal bacterial may improve or disrupt viral infections, highlighting the importance of understanding these microbial interactions and their implications for viral pathogenesis and treatment.
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http://dx.doi.org/10.1128/JVI.01282-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6819921PMC
November 2019

Immunogenicity and Efficacy of a Measles Virus-Vectored Chikungunya Vaccine in Nonhuman Primates.

J Infect Dis 2019 07;220(5):735-742

Institute for Human Infections and Immunity, University of Texas Medical Branch, Galveston.

Background: Chikungunya virus (CHIKV) infection can result in chikungunya fever (CHIKF), a self-limited acute febrile illness that can progress to chronic arthralgic sequelae in a large percentage of patients. A new measles virus-vectored vaccine was developed to prevent CHIKF, and we tested it for immunogenicity and efficacy in a nonhuman primate model.

Methods: Nine cynomolgus macaques were immunized and boosted with the measles virus-vectored chikungunya vaccine or sham-vaccinated. Sera were taken at multiple times during the vaccination phase to assess antibody responses against CHIKV. Macaques were challenged with a dose of CHIKV previously shown to cause fever and viremia, and core body temperature, viremia, and blood cell and chemistry panels were monitored.

Results: The vaccine was well tolerated in all macaques, and all seroconverted (high neutralizing antibody [PRNT80 titers, 40-640] and enzyme-linked immunosorbent assay titers) after the boost. Furthermore, the vaccinated primates were protected against viremia, fever, elevated white blood cell counts, and CHIKF-associated cytokine changes after challenge with the virulent La Reunión CHIKV strain.

Conclusions: These results further document the immunogenicity and efficacy of a measles-vectored chikungunya vaccine that shows promise in Phase I-II clinical trials. These findings are critical to human health because no vaccine to combat CHIKF is yet licensed.
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http://dx.doi.org/10.1093/infdis/jiz202DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6667792PMC
July 2019

ZIKV Demonstrates Minimal Pathologic Effects and Mosquito Infectivity in Viremic Cynomolgus Macaques.

Viruses 2018 11 21;10(11). Epub 2018 Nov 21.

Department of Pathology, University of Texas Medical Branch, Galveston, TX 77555, USA.

To evaluate the effects of ZIKV infection on non-human primates (NHPs), as well as to investigate whether these NHPs develop sufficient viremia to infect the major urban vector mosquito, , four cynomolgus macaques () were subcutaneously infected with 5.0 log focus-forming units (FFU) of DNA clone-derived ZIKV strain FSS13025 (Asian lineage, Cambodia, 2010). Following infection, the animals were sampled (blood, urine, tears, and saliva), underwent daily health monitoring, and were exposed to at specified time points. All four animals developed viremia, which peaked 3⁻4 days post-infection at a maximum value of 6.9 log genome copies/mL. No virus was detected in urine, tears, or saliva. Infection by ZIKV caused minimal overt disease: serum biochemistry and CBC values largely fell within the normal ranges, and cytokine elevations were minimal. Strikingly, the minimally colonized population of exposed to viremic animals demonstrated a maximum infection rate of 26% during peak viremia, with two of the four macaques failing to infect a single mosquito at any time point. These data indicate that cynomolgus macaques may be an effective model for ZIKV infection of humans and highlights the relative refractoriness of for ZIKV infection at the levels of viremia observed.
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http://dx.doi.org/10.3390/v10110661DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6267344PMC
November 2018
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