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Differences in virus-induced cell morphology and in virus maturation between MVA and other strains (WR, Ankara, and NYCBH) of vaccinia virus in infected human cells.

ABSTRACT: Live recombinants based on attenuated modified vaccinia virus Ankara (MVA) are potential vaccine candidates against a broad spectrum of diseases and tumors. To better understand the efficacy of MVA as a human vaccine, we analyzed by confocal and electron microscopy approaches MVA-induced morphological changes and morphogenetic stages during infection of human HeLa cells in comparison to other strains of vaccinia virus (VV): the wild-type Western Reserve (WR), Ankara, and the New York City Board of Health (NYCBH) strains. Confocal microscopy studies revealed that MVA infection alters the cytoskeleton producing elongated cells (bipolar), which do not form the characteristic actin tails. Few virions are detected in the projections connecting neighboring cells. In contrast, cells infected with the WR, Ankara, and NYCBH strains exhibit a stellated (multipolar) or rounded morphology with actin tails. A detailed transmission electron microscopy analysis of HeLa cells infected with MVA showed important differences in fine ultrastructure and amounts of the viral intermediates compared to cells infected with the other VV strains. In HeLa cells infected with MVA, the most abundant viral forms are intracellular immature virus, with few intermediates reaching the intracellular mature virus (IMV) form, at various stages of maturation, which exhibit a more rounded shape than IMVs from cells infected with the other VV strains. The "IMVs" from MVA-infected cells have an abnormal internal structure ("atypical" viruses) with potential alterations in the core-envelope interactions and are unable to significantly acquire the additional double envelope to render intracellular envelope virus. The presence of potential cell-associated envelope virus is very scarce. Our findings revealed that MVA in human cells promotes characteristic morphological changes to the cells and is able to reach the IMV stage, but these virions were not structurally normal and the subsequent steps in the morphogenetic pathway are blocked.

SUBMITTER: Gallego-Gomez JC

PROVIDER: S-EPMC228399 | biostudies-other | 2003 Oct

REPOSITORIES: biostudies-other

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Project description:Viral vectors are increasingly used as delivery means to induce a specific immunity in humans and animals. However, they also impact the immune system, and it depends on the given context whether this is beneficial or not. The attenuated vaccinia virus strain modified vaccinia virus Ankara (MVA) has been used as a viral vector in clinical studies intended to treat and prevent cancer and infectious diseases. The adjuvant property of MVA is thought to be due to its capability to stimulate innate immunity. Here, we confirmed that MVA induces interleukin-8 (IL-8), and this chemokine was upregulated significantly more in monocytes and HLA-DRbright dendritic cells (DCs) of HIV-infected patients on combined antiretroviral therapy (ART) than in cells of healthy persons. The effect of MVA on cell surface receptors is mostly unknown. Using mass cytometry profiling, we investigated the expression of 17 cell surface receptors in leukocytes after ex vivo infection of human whole-blood samples with MVA. We found that MVA downregulates most of the characteristic cell surface markers in particular types of leukocytes. In contrast, C-X-C motif chemokine receptor 4 (CXCR4) was significantly upregulated in each leukocyte type of healthy persons. Additionally, we detected a relative higher cell surface expression of the HIV-1 co-receptors C-C motif chemokine receptor 5 (CCR5) and CXCR4 in leukocytes of HIV-ART patients than in healthy persons. Importantly, we showed that MVA infection significantly downregulated CCR5 in CD4+ T cells, CD8+ T cells, B cells, and three different DC populations. CD86, a costimulatory molecule for T cells, was significantly upregulated in HLA-DRbright DCs after MVA infection of whole blood from HIV-ART patients. However, MVA was unable to downregulate cell surface expression of CD11b and CD32 in monocytes and neutrophils of HIV-ART patients to the same extent as in monocytes and neutrophils of healthy persons. In summary, MVA modulates the expression of many different kinds of cell surface receptors in leukocytes, which can vary in cells originating from persons previously infected with other pathogens.

Project description:Although vaccinia virus (VACV) was once used as a vaccine to eradicate smallpox on a worldwide scale, the biological origins of VACV are uncertain, as are the historical relationships between the different strains once used as smallpox vaccines. Here, we sequenced additional VACV strains that either represent relatively pristine examples of old vaccines (e.g., Dryvax, Lister, and Tashkent) or have been subjected to additional laboratory passage (e.g., IHD-W and WR). These genome sequences were compared with those previously reported for other VACVs as well as other orthopoxviruses. These extant VACVs do not always cluster in simple phylogenetic trees that are aligned with the known historical relationships between these strains. Rather, the pattern of deletions suggests that all existing strains likely come from a complex stock of viruses that has been passaged, distributed, and randomly sampled over time, thus obscuring simple historical or geographic links. We examined surviving nonclonal vaccine stocks, like Dryvax, which continue to harbor larger and now rare variants, including one that we have designated "clone DPP25." DPP25 encodes genes not found in most VACV strains, including an ankyrin-F-box protein, a homolog of the variola virus (Bangladesh) B18R gene which we show can be deleted without affecting virulence in mice. We propose a simple common mechanism by which recombination of a larger and hypothetical DPP25-like ancestral strain, combined with selection for retention of critically important genes near the terminal inverted repeat boundaries (vaccinia virus growth factor gene and an interferon alpha/beta receptor homolog), could produce all known VACV variants.Smallpox was eradicated by using a combination of intensive disease surveillance and vaccination using vaccinia virus (VACV). Interestingly, little is known about the historical relationships between different strains of VACV and how these viruses may have evolved from a common ancestral strain. To understand these relationships, additional strains were sequenced and compared to existing strains of VACV as well as other orthopoxviruses by using whole-genome sequence alignments. Extant strains of VACV did not always cluster in simple phylogenetic trees based on known historical relationships between these strains. Based on these findings, it is possible that all existing strains of VACV are derived from a single complex stock of viruses that has been passaged, distributed, and sampled over time.

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Differences in virus-induced cell morphology and in virus maturation between MVA and other strains (WR, Ankara, and NYCBH) of vaccinia virus in infected human cells.

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