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1.
Science ; 385(6704): 105-112, 2024 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-38963841

RESUMO

Introns containing homing endonucleases are widespread in nature and have long been assumed to be selfish elements that provide no benefit to the host organism. These genetic elements are common in viruses, but whether they confer a selective advantage is unclear. In this work, we studied intron-encoded homing endonuclease gp210 in bacteriophage ΦPA3 and found that it contributes to viral competition by interfering with the replication of a coinfecting phage, ΦKZ. We show that gp210 targets a specific sequence in ΦKZ, which prevents the assembly of progeny viruses. This work demonstrates how a homing endonuclease can be deployed in interference competition among viruses and provide a relative fitness advantage. Given the ubiquity of homing endonucleases, this selective advantage likely has widespread evolutionary implications in diverse plasmid and viral competition as well as virus-host interactions.


Assuntos
Endonucleases , Íntrons , Fagos de Pseudomonas , Pseudomonas aeruginosa , Interferência Viral , Proteínas Virais , Endonucleases/metabolismo , Endonucleases/genética , Interferência Viral/genética , Proteínas Virais/genética , Proteínas Virais/metabolismo , Montagem de Vírus , Replicação Viral , Fagos de Pseudomonas/enzimologia , Fagos de Pseudomonas/genética , Pseudomonas aeruginosa/virologia
2.
Viruses ; 16(2)2024 01 30.
Artigo em Inglês | MEDLINE | ID: mdl-38399986

RESUMO

The Saint Louis encephalitis virus (SLEV) is an encephalitogenic arbovirus (Flaviviridae family) that has a wide geographical distribution in the western hemisphere, especially in the Americas. The negevirus Brejeira (BREV) was isolated for the first time in Brazil in 2005. This study aimed to verify the existence of a possible interfering effect of BREV on the course of SLEV infection and vice versa. We used clone C6/36 cells. Three combinations of MOIs were used (SLEV 0.1 × BREV 1; SLEV 1 × BREV 0.1; SLEV 1 × BREV 1) in the kinetics of up to 7 days and then the techniques of indirect immunofluorescence (IFA), a plaque assay on Vero cells, and RT-PCR were performed. Our results showed that the cytopathic effect (CPE) caused by BREV was more pronounced than the CPE caused by SLEV. Results of IFA, the plaque assay, and RT-PCR showed the suppression of SLEV replication in the co-infection condition in all the MOI combinations used. The SLEV suppression was dose-dependent. Therefore, the ISV Brejeira can suppress SLEV replication in Aedes albopictus cells, but SLEV does not negatively interfere with BREV replication.


Assuntos
Aedes , Vírus da Encefalite de St. Louis , Interferência Viral , Animais , Chlorocebus aethiops , Vírus da Encefalite de St. Louis/genética , Células Vero , Insetos
3.
J Infect Dis ; 229(Supplement_1): S25-S33, 2024 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-37249267

RESUMO

BACKGROUND: Previous studies reported inconsistent findings regarding the association between respiratory syncytial virus (RSV) subgroup distribution and timing of RSV season. We aimed to further understand the association by conducting a global-level systematic analysis. METHODS: We compiled published data on RSV seasonality through a systematic literature review, and unpublished data shared by international collaborators. Using annual cumulative proportion (ACP) of RSV-positive cases, we defined RSV season onset and offset as ACP reaching 10% and 90%, respectively. Linear regression models accounting for meteorological factors were constructed to analyze the association of proportion of RSV-A with the corresponding RSV season onset and offset. RESULTS: We included 36 study sites from 20 countries, providing data for 179 study-years in 1995-2019. Globally, RSV subgroup distribution was not significantly associated with RSV season onset or offset globally, except for RSV season offset in the tropics in 1 model, possibly by chance. Models that included RSV subgroup distribution and meteorological factors explained only 2%-4% of the variations in timing of RSV season. CONCLUSIONS: Year-on-year variations in RSV season onset and offset are not well explained by RSV subgroup distribution or meteorological factors. Factors including population susceptibility, mobility, and viral interference should be examined in future studies.


Assuntos
Vírus Sincicial Respiratório Humano , Humanos , Modelos Lineares , Estações do Ano , Interferência Viral
4.
Viruses ; 14(12)2022 11 26.
Artigo em Inglês | MEDLINE | ID: mdl-36560647

RESUMO

In nature, viral coinfection is as widespread as viral infection alone. Viral coinfections often cause altered viral pathogenicity, disrupted host defense, and mixed-up clinical symptoms, all of which result in more difficult diagnosis and treatment of a disease. There are three major virus-virus interactions in coinfection cases: viral interference, viral synergy, and viral noninterference. We analyzed virus-virus interactions in both aspects of viruses and hosts and elucidated their possible mechanisms. Finally, we summarized the protocol of viral coinfection studies and key points in the process of virus separation and purification.


Assuntos
Coinfecção , Viroses , Vírus , Humanos , Vírus/genética , Interferência Viral
6.
Cells ; 11(6)2022 03 08.
Artigo em Inglês | MEDLINE | ID: mdl-35326378

RESUMO

BACKGROUND: Hepatitis C virus (HCV) constitutes a global health problem, while hepatitis E virus (HEV) is the major cause of acute viral hepatitis globally. HCV/HEV co-infections have been poorly characterized, as they are hampered by the lack of robust HEV cell culture systems. This study developed experimental models to study HCV/HEV co-infections and investigate viral interference in cells and humanized mice. METHODS: We used state-of-the art human hepatocytes tissue culture models to assess HEV and HCV replication in co- or super-transfection settings. Findings were confirmed by co- and super-infection experiments in human hepatocytes and in vivo in human liver chimeric mice. RESULTS: HEV was inhibited by concurrent HCV replication in human hepatocytes. This exclusion phenotype was linked to the protease activity of HCV. These findings were corroborated by the fact that in HEV on HCV super-infected mice, HEV viral loads were reduced in individual mice. Similarly, HCV on HEV super-infected mice showed reduced HCV viral loads. CONCLUSION: Direct interference of both viruses with HCV NS3/4A as the determinant was observed. In vivo, we detected reduced replication of both viruses after super-infection in individual mice. These findings provide new insights into the pathogenesis of HCV-HEV co-infections and should contribute to its clinical management in the future.


Assuntos
Coinfecção , Hepatite C , Vírus da Hepatite E , Animais , Hepacivirus/genética , Vírus da Hepatite E/genética , Camundongos , Interferência Viral , Replicação Viral
7.
Viruses ; 14(2)2022 01 24.
Artigo em Inglês | MEDLINE | ID: mdl-35215819

RESUMO

Both influenza A virus (IAV) and influenza D virus (IDV) are enzootic in pigs. IAV causes approximately 100% morbidity with low mortality, whereas IDV leads to only mild respiratory diseases in pigs. In this study, we performed a series of coinfection experiments in vitro and in vivo to understand how IAV and IDV interact and cause pathogenesis during coinfection. The results showed that IAV inhibited IDV replication when infecting swine tracheal epithelial cells (STECs) with IAV 24 or 48 h prior to IDV inoculation and that IDV suppressed IAV replication when IDV preceded IAV inoculation by 48 h. Virus interference was not identified during simultaneous IAV/IDV infections or with 6 h between the two viral infections, regardless of their order. The interference pattern at 24 and 48 h correlated with proinflammatory responses induced by the first infection, which, for IDV, was slower than for IAV by about 24 h. The viruses did not interfere with each other if both infected the cells before proinflammatory responses were induced. Coinfection in pigs further demonstrated that IAV interfered with both viral shedding and virus replication of IDV, especially in the upper respiratory tract. Clinically, coinfection of IDV and IAV did not show significant enhancement of disease pathogenesis, compared with the pigs infected with IAV alone. In summary, this study suggests that interference during coinfection of IAV and IDV is primarily due to the proinflammatory response; therefore, it is dependent on the time between infections and the order of infection. This study facilitates our understanding of virus epidemiology and pathogenesis associated with IAV and IDV coinfection.


Assuntos
Coinfecção/virologia , Vírus da Influenza A/fisiologia , Infecções por Orthomyxoviridae/veterinária , Doenças dos Suínos/virologia , Thogotovirus/fisiologia , Interferência Viral , Animais , Coinfecção/imunologia , Vírus da Influenza A/genética , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/virologia , Suínos , Doenças dos Suínos/imunologia , Thogotovirus/genética , Fatores de Tempo , Replicação Viral
8.
Viruses ; 14(2)2022 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-35215988

RESUMO

The types of interactions between severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and other respiratory viruses are not well-characterized due to the low number of co-infection cases described since the onset of the pandemic. We have evaluated the interactions between SARS-CoV-2 (D614G mutant) and influenza A(H1N1)pdm09 or respiratory syncytial virus (RSV) in the nasal human airway epithelium (HAE) infected simultaneously or sequentially (24 h apart) with virus combinations. The replication kinetics of each virus were determined by RT-qPCR at different post-infection times. Our results showed that during simultaneous infection, SARS-CoV-2 interferes with RSV-A2 but not with A(H1N1)pdm09 replication. The prior infection of nasal HAE with SARS-CoV-2 reduces the replication kinetics of both respiratory viruses. SARS-CoV-2 replication is decreased by a prior infection with A(H1N1)pdm09 but not with RSV-A2. The pretreatment of nasal HAE with BX795, a TANK-binding kinase 1 inhibitor, partially alleviates the reduced replication of SARS-CoV-2 or influenza A(H1N1)pdm09 during sequential infection with both virus combinations. Thus, a prior infection of nasal HAE with SARS-CoV-2 interferes with the replication kinetics of A(H1N1)pdm09 and RSV-A2, whereas only A(H1N1)pdm09 reduces the subsequent infection with SARS-CoV-2. The mechanism involved in the viral interference between SARS-CoV-2 and A(H1N1)pdm09 is mediated by the production of interferon.


Assuntos
Células Epiteliais/virologia , Vírus da Influenza A Subtipo H1N1/fisiologia , Nasofaringe/citologia , Vírus Sincicial Respiratório Humano/fisiologia , SARS-CoV-2/fisiologia , Interferência Viral , Replicação Viral , Coinfecção , Humanos , Interações Microbianas , Nasofaringe/virologia
9.
J Immunol ; 208(3): 720-731, 2022 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-35022269

RESUMO

Respiratory infections are a leading cause of morbidity and mortality. The presence of multiple heterologous virus infections is routinely observed in a subset of individuals screened for the presence of respiratory viruses. However, the impact overlapping infections has on disease severity and the host immune response is not well understood. Respiratory syncytial virus (RSV) and influenza A virus (IAV) are two of the most common respiratory infections observed in hospitalized patients, particularly in the very young and aged populations. In this study, we examined how the order in which BALB/c mice were infected with both RSV and IAV impacts disease severity. RSV infection prior to an IAV infection was associated with decreased weight loss and increased survival as compared with IAV infection alone. In contrast, IAV infection prior to an RSV infection was associated with similar morbidity and mortality as compared with an IAV infection alone. Our results suggest that the order in which viral infections are acquired plays a critical role in the outcome of disease severity and the host immune response.


Assuntos
Vírus da Influenza A/imunologia , Infecções por Orthomyxoviridae/imunologia , Infecções por Vírus Respiratório Sincicial/imunologia , Vírus Sinciciais Respiratórios/imunologia , Interferência Viral/fisiologia , Animais , Anticorpos Antivirais/imunologia , Linfócitos T CD8-Positivos/imunologia , Coinfecção/imunologia , Coinfecção/virologia , Citocinas/imunologia , Feminino , Interferon Tipo I/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Infecções por Orthomyxoviridae/prevenção & controle
10.
Emerg Infect Dis ; 28(2): 273-281, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-35075991

RESUMO

Multiple respiratory viruses can concurrently or sequentially infect the respiratory tract and lead to virus‒virus interactions. Infection by a first virus could enhance or reduce infection and replication of a second virus, resulting in positive (additive or synergistic) or negative (antagonistic) interaction. The concept of viral interference has been demonstrated at the cellular, host, and population levels. The mechanisms involved in viral interference have been evaluated in differentiated airway epithelial cells and in animal models susceptible to the respiratory viruses of interest. A likely mechanism is the interferon response that could confer a temporary nonspecific immunity to the host. During the coronavirus disease pandemic, nonpharmacologic interventions have prevented the circulation of most respiratory viruses. Once the sanitary restrictions are lifted, circulation of seasonal respiratory viruses is expected to resume and will offer the opportunity to study their interactions, notably with severe acute respiratory syndrome coronavirus 2.


Assuntos
COVID-19 , Infecções Respiratórias , Vírus , Animais , Humanos , Pandemias , Infecções Respiratórias/epidemiologia , SARS-CoV-2 , Interferência Viral
11.
J Infect Dis ; 225(1): 42-49, 2022 01 05.
Artigo em Inglês | MEDLINE | ID: mdl-34120189

RESUMO

BACKGROUND: Whether acute respiratory illnesses (ARIs), often associated with virus detection, are associated with lower risk for subsequent ARI remains unclear. We assessed the association between symptomatic ARI and subsequent ARI in young children. METHODS: In a prospective cohort of Peruvian children <3 years, we examined the impact of index ARI on subsequent ARI risk. Index ARI were matched with ≤3 asymptomatic observations and followed over 28 days. We compared risk of subsequent ARI between groups using conditional logistic regression adjusting for several covariates, accounting for repeat observations from individual children. RESULTS: Among 983 index ARI, 339 (34%) had an ARI event during follow-up, compared with 876/2826 (31%) matched asymptomatic observations. We found no significant association of index ARI and subsequent ARI risk during follow-up overall (adjusted odds ratio [aOR], 1.10; 95% confidence interval [CI], .98-1.23) or when limited to index ARI with respiratory viruses detected (aOR, 1.03; 95% CI, .86-1.24). Similarly, when the outcome was limited to ARI in which viruses were detected, no significant association was seen (aOR, 1.05; 95% CI, .87-1.27). CONCLUSIONS: ARIs were not associated with short-term protection against subsequent ARI in these children. Additional longitudinal studies are needed to understand drivers of recurrent ARI in young children.


Assuntos
Sistema Respiratório/virologia , Infecções Respiratórias/epidemiologia , Infecções Respiratórias/virologia , Viroses/diagnóstico , Viroses/virologia , Vírus/isolamento & purificação , Doença Aguda , Criança , Pré-Escolar , Estudos de Coortes , Feminino , Humanos , Lactente , Masculino , Peru/epidemiologia , Estudos Prospectivos , Interferência Viral
12.
Viruses ; 13(12)2021 12 14.
Artigo em Inglês | MEDLINE | ID: mdl-34960776

RESUMO

The global spread of invasive mosquito species increases arbovirus infections. In addition to the invasive species Aedes albopictus and Aedes japonicus, Aedes koreicus has spread within Central Europe. Extensive information on its vector competence is missing. Ae. koreicus from Germany were investigated for their vector competence for chikungunya virus (CHIKV), Zika virus (ZIKV) and West Nile virus (WNV). Experiments were performed under different climate conditions (27 ± 5 °C; 24 ± 5 °C) for fourteen days. Ae. koreicus had the potential to transmit CHIKV and ZIKV but not WNV. Transmission was exclusively observed at the higher temperature, and transmission efficiency was rather low, at 4.6% (CHIKV) or 4.7% (ZIKV). Using a whole virome analysis, a novel mosquito-associated virus, designated Wiesbaden virus (WBDV), was identified in Ae. koreicus. Linking the WBDV infection status of single specimens to their transmission capability for the arboviruses revealed no influence on ZIKV transmission. In contrast, a coinfection of WBDV and CHIKV likely has a boost effect on CHIKV transmission. Due to its current distribution, the risk of arbovirus transmission by Ae. koreicus in Europe is rather low but might gain importance, especially in regions with higher temperatures. The impact of WBDV on arbovirus transmission should be analyzed in more detail.


Assuntos
Aedes/virologia , Infecções por Arbovirus/transmissão , Mosquitos Vetores/virologia , Interferência Viral , Animais , Febre de Chikungunya/transmissão , Infecção por Zika virus/transmissão
13.
Virology ; 562: 50-62, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34256244

RESUMO

We describe the isolation and characterization of a novel insect-specific flavivirus (ISFV), tentatively named Aripo virus (ARPV), that was isolated from Psorophora albipes mosquitoes collected in Trinidad. The ARPV genome was determined and phylogenetic analyses showed that it is a dual host associated ISFV, and clusters with the main mosquito-borne flaviviruses. ARPV antigen was significantly cross-reactive with Japanese encephalitis virus serogroup antisera, with significant cross-reactivity to Ilheus and West Nile virus (WNV). Results suggest that ARPV replication is limited to mosquitoes, as it did not replicate in the sandfly, culicoides or vertebrate cell lines tested. We also demonstrated that ARPV is endocytosed into vertebrate cells and is highly immunomodulatory, producing a robust innate immune response despite its inability to replicate in vertebrate systems. We show that prior infection or coinfection with ARPV limits WNV-induced disease in mouse models, likely the result of a robust ARPV-induced type I interferon response.


Assuntos
Flavivirus/imunologia , Imunomodulação , Vírus de Insetos/imunologia , Vertebrados/imunologia , Animais , Antígenos Virais/imunologia , Reações Cruzadas , Culicidae/virologia , Modelos Animais de Doenças , Flavivirus/genética , Flavivirus/isolamento & purificação , Flavivirus/patogenicidade , Genoma Viral/genética , Especificidade de Hospedeiro , Imunidade Inata , Vírus de Insetos/genética , Vírus de Insetos/isolamento & purificação , Vírus de Insetos/patogenicidade , Macrófagos/imunologia , Camundongos , Filogenia , Vertebrados/virologia , Interferência Viral , Replicação Viral , Febre do Nilo Ocidental/imunologia , Vírus do Nilo Ocidental/imunologia , Vírus do Nilo Ocidental/patogenicidade
14.
Front Immunol ; 12: 674216, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34177916

RESUMO

Virus interference is a phenomenon in which two viruses interact within a host, affecting the outcome of infection of at least one of such viruses. The effect of this event was first observed in the XVIII century and it was first recorded even before virology was recognized as a distinct science from microbiology. Studies on virus interference were mostly done in the decades between 1930 and 1960 in viruses infecting bacteria and different vertebrates. The systems included in vivo experiments and later, more refined assays were done using tissue and cell cultures. Many viruses involved in interference are pathogenic to humans or to economically important animals. Thus the phenomenon may be relevant to medicine and to animal production due to the possibility to use it as alternative to chemical therapies against virus infections to reduce the severity of disease/mortality caused by a superinfecting virus. Virus interference is defined as the host resistance to a superinfection caused by a pathogenic virus causing obvious signs of disease and/or mortality due to the action of an interfering virus abrogating the replication of the former virus. Different degrees of inhibition of the superinfecting virus can occur. Due to the emergence of novel pathogenic viruses in recent years, virus interference has recently been revisited using different pathogens and hosts, including commercially important farmed aquatic species. Here, some highly pathogenic viruses affecting farmed crustaceans can be affected by interference with other viruses. This review presents data on the history of virus interference in hosts including bacteria and animals, with emphasis on the known cases of virus interference in crustacean hosts. Life Science Identifiers (LSIDs) Escherichia coli [(Migula 1895) Castellani & Chalmers 1919] Aedes albopictus (Skuse 1894) Liocarcinus depurator (Linnaeus 1758): urn:lsid:marinespecies.org:taxname:107387 Penaeus duorarum (Burkenroad 1939): urn:lsid:marinespecies.org:taxname:158334 Carcinus maenas (Linnaeus 1758): urn:lsid:marinespecies.org:taxname:107381 Macrobrachium rosenbergii (De Man 1879): urn:lsid:marinespecies.org:taxname:220137 Penaeus vannamei (Boone 1931): urn:lsid:zoobank.org:pub:C30A0A50-E309-4E24-851D-01CF94D97F23 Penaeus monodon (Fabricius 1798): urn:lsid:zoobank.org:act:3DD50D8B-01C2-48A7-B80D-9D9DD2E6F7AD Penaeus stylirostris (Stimpson 1874): urn:lsid:marinespecies.org:taxname:584982.


Assuntos
Crustáceos/virologia , Interferência Viral , Viroses/virologia , Animais
15.
Viruses ; 13(5)2021 05 02.
Artigo em Inglês | MEDLINE | ID: mdl-34063220

RESUMO

Viral recombination is a key mechanism in the evolution and diversity of noroviruses. In vivo, synchronous single-cell coinfection by multiple viruses, the ultimate prerequisite to viral recombination, is likely to be a rare event and delayed secondary infections are a more probable occurrence. Here, we determine the effect of a temporal separation of in vitro infections with the two homologous murine norovirus strains MNV-1 WU20 and CW1 on the composition of nascent viral populations. WU20 and CW1 were either synchronously inoculated onto murine macrophage cell monolayers (coinfection) or asynchronously applied (superinfection with varying titres of CW1 at half-hour to 24-h delays). Then, 24 h after initial co-or superinfection, quantification of genomic copy numbers and discriminative screening of plaque picked infectious progeny viruses demonstrated a time-dependent predominance of primary infecting WU20 in the majority of viral progenies. Our results indicate that a time interval from one to two hours onwards between two consecutive norovirus infections allows for the establishment of a barrier that reduces or prevents superinfection.


Assuntos
Infecções por Caliciviridae/virologia , Norovirus/fisiologia , Doenças dos Roedores/virologia , Interferência Viral , Animais , Genoma Viral , Camundongos , Células RAW 264.7 , RNA Viral , Recombinação Genética , Superinfecção
16.
Virulence ; 12(1): 1111-1121, 2021 12.
Artigo em Inglês | MEDLINE | ID: mdl-34034617

RESUMO

Coronaviruses and influenza viruses are circulating in humans and animals all over the world. Co-infection with these two viruses may aggravate clinical signs. However, the molecular mechanisms of co-infections by these two viruses are incompletely understood. In this study, we applied air-liquid interface (ALI) cultures of well-differentiated porcine tracheal epithelial cells (PTECs) to analyze the co-infection by a swine influenza virus (SIV, H3N2 subtype) and porcine respiratory coronavirus (PRCoV) at different time intervals. Our results revealed that in short-term intervals, prior infection by influenza virus caused complete inhibition of coronavirus infection, while in long-term intervals, some coronavirus replication was detectable. The influenza virus infection resulted in (i) an upregulation of porcine aminopeptidase N, the cellular receptor for PRCoV and (ii) in the induction of an innate immune response which was responsible for the inhibition of PRCoV replication. By contrast, prior infection by coronavirus only caused a slight inhibition of influenza virus replication. Taken together, the timing and the order of virus infection are important determinants in co-infections. This study is the first to show the impact of SIV and PRCoV co- and super-infection on the cellular level. Our results have implications also for human viruses, including potential co-infections by SARS-CoV-2 and seasonal influenza viruses.


Assuntos
Células Epiteliais/virologia , Vírus da Influenza A Subtipo H3N2/fisiologia , Coronavirus Respiratório Porcino/fisiologia , Interferência Viral , Animais , Antígenos CD13/metabolismo , Células Cultivadas , Coinfecção/virologia , Infecções por Coronavirus/virologia , Células Epiteliais/imunologia , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Imunidade Inata , Infecções por Orthomyxoviridae/virologia , Suínos , Traqueia/citologia , Replicação Viral
18.
J Virol ; 95(15): e0080221, 2021 07 12.
Artigo em Inglês | MEDLINE | ID: mdl-34011551

RESUMO

Gene drives are genetic systems designed to efficiently spread a modification through a population. They have been designed almost exclusively in eukaryotic species, especially in insects. We recently developed a CRISPR-based gene drive system in herpesviruses that relies on similar mechanisms and could efficiently spread into a population of wild-type viruses. A common consequence of gene drives in insects is the appearance and selection of drive-resistant sequences that are no longer recognized by CRISPR-Cas9. In this study, we analyzed in cell culture experiments the evolution of resistance in a viral gene drive against human cytomegalovirus. We report that after an initial invasion of the wild-type population, a drive-resistant population is positively selected over time and outcompetes gene drive viruses. However, we show that targeting evolutionarily conserved sequences ensures that drive-resistant viruses acquire long-lasting mutations and are durably attenuated. As a consequence, and even though engineered viruses do not stably persist in the viral population, remaining viruses have a replication defect, leading to a long-term reduction of viral levels. This marks an important step toward developing effective gene drives in herpesviruses, especially for therapeutic applications. IMPORTANCE The use of defective viruses that interfere with the replication of their infectious parent after coinfecting the same cells-a therapeutic strategy known as viral interference-has recently generated a lot of interest. The CRISPR-based system that we recently reported for herpesviruses represents a novel interfering strategy that causes the conversion of wild-type viruses into new recombinant viruses and drives the native viral population to extinction. In this study, we analyzed how targeted viruses evolved resistance against the technology. Through numerical simulations and cell culture experiments with human cytomegalovirus, we showed that after the initial propagation, a resistant viral population is positively selected and outcompetes engineered viruses over time. We show, however, that targeting evolutionarily conserved sequences ensures that resistant viruses are mutated and attenuated, which leads to a long-term reduction of viral levels. This marks an important step toward the development of novel therapeutic strategies against herpesviruses.


Assuntos
Sistemas CRISPR-Cas/genética , Sequência Conservada/genética , Citomegalovirus/genética , Tecnologia de Impulso Genético/métodos , Interferência Viral/genética , Sequência de Aminoácidos , Sequência de Bases , Linhagem Celular , Citomegalovirus/crescimento & desenvolvimento , Infecções por Citomegalovirus/prevenção & controle , Infecções por Citomegalovirus/terapia , Vírus Defeituosos/genética , Farmacorresistência Viral/genética , Genes Virais/genética , Humanos , Alinhamento de Sequência , Proteínas Virais/genética
19.
J Virol ; 95(13): e0048621, 2021 06 10.
Artigo em Inglês | MEDLINE | ID: mdl-33853961

RESUMO

Wild-type adeno-associated virus (AAV) can only replicate in the presence of helper factors, which can be provided by coinfecting helper viruses such as adenoviruses and herpesviruses. The AAV genome consists of a linear, single-stranded DNA (ssDNA), which is converted into different molecular structures within the host cell. Using high-throughput sequencing, we found that herpes simplex virus 1 (HSV-1) coinfection leads to a shift in the type of AAV genome end recombination. In particular, open-end inverted terminal repeat (ITR) recombination was enhanced, whereas open-closed ITR recombination was reduced in the presence of HSV-1. We demonstrate that the HSV-1 protein ICP8 plays an essential role in HSV-1-mediated interference with AAV genome end recombination, indicating that the previously described ICP8-driven mechanism of HSV-1 genome recombination may be underlying the observed changes. We also provide evidence that additional factors, such as products of true late genes, are involved. Although HSV-1 coinfection significantly changed the type of AAV genome end recombination, no significant change in the amount of circular AAV genomes was identified. IMPORTANCE Adeno-associated virus (AAV)-mediated gene therapy represents one of the most promising approaches for the treatment of genetic diseases. Currently, various GMP-compatible production methods can be applied to manufacture clinical-grade vector, including methods that employ helper factors derived from herpes simplex virus 1 (HSV-1). Yet, to date, we do not fully understand how HSV-1 interacts with AAV. We observed that HSV-1 modulates AAV genome ends similarly to the genome recombination events observed during HSV-1 replication and postulate that further improvements of the HSV-1 production platform may enhance packaging of the recombinant AAV particles.


Assuntos
Dependovirus/crescimento & desenvolvimento , Dependovirus/genética , Genoma Viral/genética , Vírus Auxiliares/genética , Herpesvirus Humano 1/genética , Recombinação Genética/genética , Animais , Linhagem Celular , Chlorocebus aethiops , Coinfecção/patologia , Células HEK293 , Células HeLa , Herpes Simples/patologia , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Infecções por Parvoviridae/patologia , Sequências Repetidas Terminais/genética , Células Vero , Interferência Viral/genética , Replicação Viral/genética
20.
Viruses ; 13(3)2021 03 02.
Artigo em Inglês | MEDLINE | ID: mdl-33801276

RESUMO

The SAM and HD domain-containing protein 1 (SAMHD1) is a dNTP triphosphohydrolase that plays a crucial role for a variety of different cellular functions. Besides balancing intracellular dNTP concentrations, facilitating DNA damage repair, and dampening excessive immune responses, SAMHD1 has been shown to act as a major restriction factor against various virus species. In addition to its well-described activity against retroviruses such as HIV-1, SAMHD1 has been identified to reduce the infectivity of different DNA viruses such as the herpesviruses CMV and EBV, the poxvirus VACV, or the hepadnavirus HBV. While some viruses are efficiently restricted by SAMHD1, others have developed evasion mechanisms that antagonize the antiviral activity of SAMHD1. Within this review, we summarize the different cellular functions of SAMHD1 and highlight the countermeasures viruses have evolved to neutralize the restriction factor SAMHD1.


Assuntos
Infecções por Vírus de DNA/imunologia , Interações Hospedeiro-Patógeno/imunologia , Infecções por Retroviridae/imunologia , Proteína 1 com Domínio SAM e Domínio HD/imunologia , Vírus de DNA/imunologia , Humanos , Retroviridae/imunologia , Interferência Viral
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