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1.
Rev Argent Microbiol ; 56(2): 125-133, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-38143232

RESUMEN

Bovine respiratory syncytial virus (BRSV) affects both beef and dairy cattle, reaching morbidity and mortality rates of 60-80% and 20%, respectively. The aim of this study was to obtain a recombinant MVA expressing the BRSV F protein (MVA-F) as a vaccine against BRSV and to evaluate the immune response induced by MVA-F after systemic immunization in homologous and heterologous vaccination (MVA-F alone or combined with a subunit vaccine), and after intranasal immunization of mice. MVA-F administered by intraperitoneal route in a homologous scheme elicited levels of neutralizing antibodies similar to those obtained with inactivated BRSV as well as better levels of IFN-γ secretion. In addition, nasal administration of MVA-F elicited local and systemic immunity with a Th1 profile. This study suggests that MVA-F is a good candidate for further evaluations combining intranasal and intramuscular routes, in order to induce local and systemic immune responses, to improve the vaccine efficacy against BRSV infection.


Asunto(s)
Administración Intranasal , Ratones Endogámicos BALB C , Virus Sincitial Respiratorio Bovino , Animales , Virus Sincitial Respiratorio Bovino/inmunología , Ratones , Femenino , Bovinos , Proteínas Virales de Fusión/inmunología , Proteínas Virales de Fusión/genética , Proteínas Virales de Fusión/administración & dosificación , Vacunas contra Virus Sincitial Respiratorio/inmunología , Vacunas contra Virus Sincitial Respiratorio/administración & dosificación , Vectores Genéticos , Infecciones por Virus Sincitial Respiratorio/prevención & control , Infecciones por Virus Sincitial Respiratorio/inmunología , Infecciones por Virus Sincitial Respiratorio/veterinaria , Virus Vaccinia/inmunología , Virus Vaccinia/genética , Anticuerpos Antivirales/sangre , Inmunidad Mucosa , Anticuerpos Neutralizantes/sangre , Anticuerpos Neutralizantes/inmunología , Inmunización/métodos , Vacunas de Subunidad/inmunología , Vacunas de Subunidad/administración & dosificación
2.
Appl Microbiol Biotechnol ; 102(23): 10139-10146, 2018 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-30238142

RESUMEN

The baculovirus Autographa californica multiple nuclear polyhedrosis virus (AcMNPV) infects lepidopteran invertebrates as natural hosts, although it also has been used as display vector for vaccine development. In this work, we evaluated the effectiveness of repetitive doses of AcMNPV-based vectors on the cytotoxic immune response specific to the capsid-displayed heterologous antigen ovalbumin (OVA). Our results demonstrate that baculovirus vectors induce a boosting effect in the cytotoxic immune response to OVA, making possible to recover the levels obtained in the primary response. Moreover, mice preimmunized with wild-type baculovirus showed a complete lack of antigen-specific CD8 cytotoxic T lymphocytes (CTLs) that may be related to the presence of antibodies directed to baculoviral surface proteins, particularly to GP64. However, baculovirus was able to induce the innate immune response in spite of a previous response against this vector, although some quantitative differences reflect a distinct activation of the immune cells in prime and boost. This is the first report in which the novel capsid display strategy is evaluated in prime-boost schemes to improve efficient CTL responses.


Asunto(s)
Proteínas de la Cápside/inmunología , Cápside/inmunología , Nucleopoliedrovirus/inmunología , Vacunación , Animales , Antígenos Virales/inmunología , Linfocitos T CD8-positivos/inmunología , Línea Celular , Citocinas/sangre , Femenino , Interacciones Microbiota-Huesped/inmunología , Inmunidad Innata , Ratones , Ratones Endogámicos C57BL , Ovalbúmina/inmunología , Spodoptera/inmunología , Spodoptera/virología , Linfocitos T Citotóxicos/inmunología
3.
Vaccine ; 2024 Jun 15.
Artículo en Inglés | MEDLINE | ID: mdl-38880691

RESUMEN

Vaccination is the best strategy to control Paratuberculosis (PTB), which is a significant disease in cattle and sheep. Previously we showed the humoral and cellular immune response induced by a novel vaccine candidate against PTB based on the Argentinian Mycobacterium avium subspecies paratuberculosis (Map) 6611 strain. To improve 6611 immunogenicity and efficacy, we evaluated this vaccine candidate in mice with two different adjuvants and a heterologous boost with a recombinant modified vaccinia Ankara virus (MVA) expressing the antigen 85A (MVA85A). We observed that boosting with MVA85A did not improve total IgG or specific isotypes in serum induced by one or two doses of 6611 formulated with incomplete Freund's adjuvant (IFA). However, when 6611 was formulated with ISA201 adjuvant, MVA85A boost enhanced the production of IFNγ, Th1/Th17 cytokines (IL-2, TNF, IL-17A) and IL-6, IL-4 and IL-10. Also, this group showed the highest levels of IgG2b and IgG3 isotypes, both important for better protection against Map infection in the murine model. Finally, the heterologous scheme elicited the highest levels of protection after Map challenge (lowest CFU count and liver lesion score). In conclusion, our results encourage further evaluation of 6611 strain + ISA201 prime and MVA85A boost in bovines.

4.
Mol Immunol ; 139: 115-122, 2021 11.
Artículo en Inglés | MEDLINE | ID: mdl-34481269

RESUMEN

Modified vaccinia Ankara virus (MVA) is extensively used as a vaccine vector. We have previously observed that MVAΔ008, an MVA lacking the gene that codes for interleukin-18 binding protein, significantly increases CD8+ and CD4+ T-cell responses to vaccinia virus (VACV) epitopes and recombinant HIV antigens. However, the efficacy of this vector against pathogens or tumor cells remains unclear. Thus, the aim of this study was to evaluate the cellular immune response and the protection induced by recombinant MVAs encoding the model antigen ovalbumin (OVA). We used the MO5 melanoma tumor model (OVA-expressing tumor) as an approach for evaluating the vector-induced efficacy. Our results show that MVAΔ008-OVA (optimized vector) induced higher in vivo specific cytotoxicity and ex vivo T-cell IFN-γ responses against OVA than the conventional MVA vector. Importantly, the recombinant vectors were capable of controlling MO5 tumor growth. Indeed, the administration of MVAΔ008-OVA or MVA-OVA in prophylactic and therapeutic schemes provided total protection and longer survival of mice, respectively. Overall, our results demonstrate the improved immunogenicity and the protective capacity of MVAΔ008 against a heterologous model antigen. These findings suggest that MVAΔ008 constitutes an excellent candidate for vaccine development against pathogens or cancer therapy.


Asunto(s)
Melanoma Experimental/inmunología , Ovalbúmina/inmunología , Vacunas Sintéticas/inmunología , Vacunas Virales/inmunología , Animales , Vectores Genéticos , Ratones , Vacunas de ADN
5.
Front Cell Infect Microbiol ; 10: 581812, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-33072631

RESUMEN

Mycobacterium tuberculosis (Mtb) infection is one of the leading causes of death worldwide. The Modified Vaccinia Ankara (MVA) vaccine vector expressing the mycobacterial antigen 85A (MVA85A) was demonstrated to be safe, although it did not improve BCG efficacy, denoting the need to search for improved tuberculosis vaccines. In this work, we investigated the effect of IL-12 DNA -as an adjuvant- on an Ag85A DNA prime/MVA85A boost vaccination regimen. We evaluated the immune response profile elicited in mice and the protection conferred against intratracheal Mtb H37Rv challenge. We observed that the immunization scheme including DNA-A85A+DNA-IL-12/MVA85A induced a strong IFN-γ production to Ag85A in vitro, with a significant expansion of IFN-γ+CD4+ and IFN-γ+CD8+ anti-Ag85A lymphocytes. Furthermore, we also detected a significant increase in the proportion of specific CD8+CD107+ T cells against Ag85A. Additionally, inclusion of IL-12 DNA in the DNA-A85A/MVA85A vaccine scheme induced a marked augment in anti-Ag85A IgG levels. Interestingly, after 30 days of infection with Mtb H37Rv, DNA-A85A+DNA-IL-12/MVA85A vaccinated mice displayed a significant reduction in lung bacterial burden. Together, our findings suggest that IL-12 DNA might be useful as a molecular adjuvant in an Ag85A DNA/MVA prime-boost vaccine against Mtb infection.


Asunto(s)
Mycobacterium tuberculosis , Vacunas contra la Tuberculosis , Tuberculosis , Vacunas de ADN , Aciltransferasas/genética , Animales , Antígenos Bacterianos/genética , Vacuna BCG , ADN , Inmunización Secundaria , Interleucina-12/genética , Ratones , Mycobacterium tuberculosis/genética , Tuberculosis/prevención & control , Vacunas de ADN/genética
7.
Methods Mol Biol ; 1581: 15-28, 2017.
Artículo en Inglés | MEDLINE | ID: mdl-28374241

RESUMEN

Canarypox viruses (CNPV) are excellent candidates to develop recombinant vector vaccines due to both their capability to induce protective immune responses and their incompetence to replicate in mammalian cells (safety profile). In addition, CNPV and the derived recombinants can be manipulated under biosafety level 1 conditions. There is no commercially available system to obtain recombinant CNPV; however, the methodology and tools required to develop recombinant vaccinia virus (VV), prototype of the Poxviridae family, can be easily adapted. This chapter provides protocols for the generation, plaque isolation, molecular characterization, amplification and purification of recombinant CNPV.


Asunto(s)
Virus de la Viruela de los Canarios/crecimiento & desarrollo , Fibroblastos/virología , Vacunas Sintéticas/inmunología , Animales , Virus de la Viruela de los Canarios/genética , Virus de la Viruela de los Canarios/inmunología , Línea Celular , Pollos , Fibroblastos/inmunología , Vacunas Virales/inmunología
8.
Viral Immunol ; 30(1): 70-76, 2017.
Artículo en Inglés | MEDLINE | ID: mdl-27809679

RESUMEN

In this study, we evaluated the immunogenicity and efficacy of mucosal delivery of a recombinant modified vaccinia Ankara virus (MVA) expressing the secreted version of bovine herpesvirus type 1 (BoHV-1) glycoprotein D (MVA-gDs) without addition of adjuvant in two animal models. First, we demonstrated the capability of MVA-gDs of inducing both local and systemic anti-gD humoral immune response after intranasal immunization of mice. Then, we confirmed that two doses of MVA-gDs administered intranasally to rabbits induced systemic anti-gD antibodies and conferred protection against BoHV-1 challenge. Our results show the potential of using MVA as a vector for the rational design of veterinary vaccines capable of inducing specific and protective immune responses both at local and systemic level.


Asunto(s)
Portadores de Fármacos , Infecciones por Herpesviridae/prevención & control , Herpesvirus Bovino 1/inmunología , Vacunas contra Herpesvirus/inmunología , Virus Vaccinia/genética , Proteínas Virales/inmunología , Administración Intranasal , Animales , Anticuerpos Antivirales/sangre , Modelos Animales de Enfermedad , Femenino , Infecciones por Herpesviridae/inmunología , Vacunas contra Herpesvirus/administración & dosificación , Vacunas contra Herpesvirus/genética , Ratones Endogámicos BALB C , Conejos , Vacunas Sintéticas/administración & dosificación , Vacunas Sintéticas/genética , Vacunas Sintéticas/inmunología , Proteínas Virales/genética
9.
Viruses ; 8(5)2016 05 23.
Artículo en Inglés | MEDLINE | ID: mdl-27223301

RESUMEN

MVA is an attenuated vector that still retains immunomodulatory genes. We have previously reported its optimization after deleting the C12L gene, coding for the IL-18 binding-protein. Here, we analyzed the immunogenicity of MVA vectors harboring the simultaneous deletion of A44L, related to steroid synthesis and A46R, a TLR-signaling inhibitor (MVAΔA44L-A46R); or also including a deletion of C12L (MVAΔC12L/ΔA44L-A46R). The absence of biological activities of the deleted genes in the MVA vectors was demonstrated. Adaptive T-cell responses against VACV epitopes, evaluated in spleen and draining lymph-nodes of C57Bl/6 mice at acute/memory phases, were of higher magnitude in those animals that received deleted MVAs compared to MVAwt. MVAΔC12L/ΔA44L-A46R generated cellular specific memory responses of higher quality characterized by bifunctionality (CD107a/b⁺/IFN-γ⁺) and proliferation capacity. Deletion of selected genes from MVA generated innate immune responses with higher levels of determining cytokines related to T-cell response generation, such as IL-12, IFN-γ, as well as IL-1ß and IFN-ß. This study describes for the first time that simultaneous deletion of the A44L, A46R and C12L genes from MVA improved its immunogenicity by enhancing the host adaptive and innate immune responses, suggesting that this approach comprises an appropriate strategy to increase the MVA vaccine potential.


Asunto(s)
Inmunidad Innata , Eliminación de Secuencia , Linfocitos T/inmunología , Virus Vaccinia/genética , Virus Vaccinia/inmunología , Proteínas Virales/genética , Animales , Antígenos Virales/inmunología , Citocinas/metabolismo , Epítopos/inmunología , Ganglios Linfáticos/inmunología , Ratones Endogámicos C57BL , Bazo/inmunología
10.
Vaccine ; 32(36): 4625-32, 2014 Aug 06.
Artículo en Inglés | MEDLINE | ID: mdl-24968152

RESUMEN

In this study, a recombinant modified vaccinia virus Ankara vector expressing a chimeric multi-antigen was obtained and evaluated as a candidate vaccine in homologous and heterologous prime-boost immunizations with a recombinant protein cocktail. The chimeric multi-antigen comprises immunodominant B and T cell regions of three Babesia bovis proteins. Humoral and cellular immune responses were evaluated in mice to compare the immunogenicity induced by different immunization schemes. The best vaccination scheme was achieved with a prime of protein cocktail and a boost with the recombinant virus. This scheme induced high level of specific IgG antibodies and secreted IFN and a high degree of activation of IFNγ(+) CD4(+) and CD8(+) specific T cells. This is the first report in which a novel vaccine candidate was constructed based on a rationally designed multi-antigen and evaluated in a prime-boost regime, optimizing the immune response necessary for protection against bovine babesiosis.


Asunto(s)
Babesia bovis/inmunología , Babesiosis/prevención & control , Inmunización Secundaria , Vacunas Antiprotozoos/inmunología , Proteínas Recombinantes/inmunología , Animales , Babesia bovis/genética , Babesiosis/inmunología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Células Cultivadas , Embrión de Pollo , Cricetinae , Fibroblastos/inmunología , Vectores Genéticos/inmunología , Inmunidad Celular , Inmunoglobulina G/sangre , Interferón gamma/sangre , Masculino , Ratones , Ratones Endogámicos BALB C , Porcinos , Virus Vaccinia/genética
11.
J Vet Sci ; 13(2): 199-201, 2012 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-22705743

RESUMEN

A recombinant modified vaccinia Ankara (MVA) virus expressing mature viral protein 2 (VP2) of the infectious bursal disease virus (IBDV) was constructed to develop MVA-based vaccines for poultry. We demonstrated that this recombinant virus was able to induce a specific immune response by observing the production of anti-IBDV-seroneutralizing antibodies in specific pathogen-free chickens. Besides, as the epitopes of VP2 responsible to induce IBDV-neutralizing antibodies are discontinuous, our results suggest that VP2 protein expressed from MVA-VP2 maintained the correct conformational structure. To our knowledge, this is the first report on the usefulness of MVA-based vectors for developing recombinant vaccines for poultry.


Asunto(s)
Infecciones por Birnaviridae/veterinaria , Virus de la Enfermedad Infecciosa de la Bolsa/inmunología , Enfermedades de las Aves de Corral/prevención & control , Virus Vaccinia/genética , Proteínas Estructurales Virales/inmunología , Vacunas Virales/inmunología , Animales , Anticuerpos Antivirales , Infecciones por Birnaviridae/prevención & control , Células Cultivadas , Embrión de Pollo , Pollos , Fibroblastos/metabolismo , Enfermedades de las Aves de Corral/virología , Organismos Libres de Patógenos Específicos , Virus Vaccinia/inmunología , Virus Vaccinia/metabolismo , Proteínas Estructurales Virales/genética , Proteínas Estructurales Virales/metabolismo
12.
PLoS One ; 7(2): e32220, 2012.
Artículo en Inglés | MEDLINE | ID: mdl-22384183

RESUMEN

BACKGROUND: Modified Vaccinia Ankara (MVA) is an attenuated strain of Vaccinia virus (VACV) currently employed in many clinical trials against HIV/AIDS and other diseases. MVA still retains genes involved in host immune response evasion, enabling its optimization by removing some of them. The aim of this study was to evaluate cellular immune responses (CIR) induced by an IL-18 binding protein gene (C12L) deleted vector (MVAΔC12L). METHODOLOGY/PRINCIPAL FINDINGS: BALB/c and C57BL/6 mice were immunized with different doses of MVAΔC12L or MVA wild type (MVAwt), then CIR to VACV epitopes in immunogenic proteins were evaluated in spleen and draining lymph nodes at acute and memory phases (7 and 40 days post-immunization respectively). Compared with parental MVAwt, MVAΔC12L immunization induced a significant increase of two to three-fold in CD8(+) and CD4(+) T-cell responses to different VACV epitopes, with increased percentage of anti-VACV cytotoxic CD8(+) T-cells (CD107a/b(+)) during the acute phase of the response. Importantly, the immunogenicity enhancement was also observed after MVAΔC12L inoculation with different viral doses and by distinct routes (systemic and mucosal). Potentiation of MVA's CIR was also observed during the memory phase, in correlation with a higher protection against an intranasal challenge with VACV WR. Of note, we could also show a significant increase in the CIR against HIV antigens such as Env, Gag, Pol and Nef from different subtypes expressed from two recombinants of MVAΔC12L during heterologous DNA prime/MVA boost vaccination regimens. CONCLUSIONS/SIGNIFICANCE: This study demonstrates the relevance of IL-18 bp contribution in the immune response evasion during MVA infection. Our findings clearly show that the deletion of the viral IL-18 bp gene is an effective approach to increase MVA vaccine efficacy, as immunogenicity improvements were observed against vector antigens and more importantly to HIV antigens.


Asunto(s)
Vacunas contra el SIDA/uso terapéutico , Péptidos y Proteínas de Señalización Intercelular/química , Virus Vaccinia/metabolismo , Vacunas contra el SIDA/química , Animales , Linfocitos T CD8-positivos/metabolismo , Pollos , Femenino , Eliminación de Gen , Vectores Genéticos , Memoria Inmunológica , Péptidos y Proteínas de Señalización Intercelular/genética , Interferón gamma/metabolismo , Interleucina-18 , Interleucina-2/metabolismo , Ganglios Linfáticos/patología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Bazo/metabolismo
13.
Vet Microbiol ; 153(3-4): 361-6, 2011 Dec 15.
Artículo en Inglés | MEDLINE | ID: mdl-21665386

RESUMEN

The US3 protein is a unique protein kinase only present in the Alphaherpesvirinae subfamily of the herpesviruses. Studies performed with several alphaherpesviruses demonstrated that the US3 protein is involved in cytoskeleton modifications during viral infection and displays anti-apoptotic activity. However, the US3 protein of BoHV-5 has not been studied up to now. As reported for other alphaherpesviruses, our results showed that BoHV-5 US3 confers resistance against apoptosis and induces cytoskeletal reorganization leading to cell rounding, actin stress fiber breakdown and cell projections that interconnect cells. The expression of a kinase-dead version of BoHV-5 US3 showed that the anti-apoptotic activity and the induction of cell projections are kinase-dependent whereas kinase activity is not absolutely required for actin stress fiber breakdown. Besides, the kinase-dead version of US3, but not the wild type protein, was found excluded from the nucleus. These results constitute the first report on the BoHV-5 US3 functions, and highlight that there are functional differences and similarities among US3 proteins of different alphaherpesviruses.


Asunto(s)
Actinas/metabolismo , Apoptosis/fisiología , Citoesqueleto/metabolismo , Herpesvirus Bovino 5/enzimología , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Virales/metabolismo , Animales , Bovinos , Núcleo Celular/enzimología , Chlorocebus aethiops , Citoplasma/enzimología , Microtúbulos/metabolismo , Datos de Secuencia Molecular , Mutación , Proteínas Serina-Treonina Quinasas/genética , Células Vero , Proteínas Virales/genética
14.
Viral Immunol ; 24(4): 331-9, 2011 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-21830904

RESUMEN

Bovine herpesvirus-1 (BoHV-1) infection is distributed worldwide and the development of new tools to fight against this pathogen has become extremely important. In this work a recombinant modified vaccinia virus Ankara (MVA) vector expressing the secreted version of glycoprotein D, MVA-gDs, was obtained and evaluated as a candidate vaccine. First, the correct expression, antigenicity, and N-glycosylation of glycoprotein D were confirmed by molecular techniques. Then MVA-gDs was used as parenteral immunogen in BALB/C mice in which a specific anti-gD humoral immune response was induced and maintained for 7 mo. Two doses of MVA-gDs supplemented with cholera toxin delivered by intranasal immunization induced IgA anti-gD humoral immune responses in nasal and bronchopulmonary washes, as well as IgG anti-gD antibodies in serum samples. In order to evaluate the protection conferred by MVA-gDs immunization, a rabbit BoHV-1 challenge assay was performed. A shorter viral excretion period and a reduction in the number of animals shedding BoHV-1 was observed in the group immunized with recombinant MVA-gDs. In conclusion our data encourage further studies to evaluate MVA-gDs, alone or combined with other immunogens, as a candidate vaccine for BoHV-1.


Asunto(s)
Portadores de Fármacos , Vacunas contra Herpesvirus/inmunología , Virus Vaccinia/genética , Proteínas Virales/inmunología , Adyuvantes Inmunológicos/administración & dosificación , Adyuvantes Inmunológicos/genética , Administración Intranasal , Animales , Anticuerpos Antivirales/análisis , Anticuerpos Antivirales/sangre , Líquido del Lavado Bronquioalveolar/inmunología , Toxina del Cólera/administración & dosificación , Toxina del Cólera/genética , Modelos Animales de Enfermedad , Femenino , Vectores Genéticos , Infecciones por Herpesviridae/inmunología , Infecciones por Herpesviridae/prevención & control , Vacunas contra Herpesvirus/administración & dosificación , Vacunas contra Herpesvirus/genética , Inmunoglobulina A/análisis , Inmunoglobulina G/sangre , Masculino , Ratones , Ratones Endogámicos BALB C , Mucosa Nasal/inmunología , Enfermedades de los Roedores/inmunología , Enfermedades de los Roedores/prevención & control , Factores de Tiempo , Vacunas Sintéticas/administración & dosificación , Vacunas Sintéticas/genética , Vacunas Sintéticas/inmunología , Proteínas Virales/genética , Esparcimiento de Virus
15.
J Virol Methods ; 161(1): 75-83, 2009 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-19501619

RESUMEN

Bovine herpesviruses 1 (BoHV-1) and 5 (BoHV-5) are closely related alphaherpesviruses infecting cattle. In countries where both viruses circulate, co-infection of cattle is likely. It was shown that recombination occurs at a high frequency in cattle infected dually with two BoHV-1 mutants. In addition, interspecific recombinants are generated in cell culture co-infected with BoHV-1 and BoHV-5. Even if the process of interspecific recombination appears inefficient relative to intraspecific recombination, BoHV-1 and BoHV-5 may give rise to interspecific recombinants in co-infected cattle. Since molecular tools for differentiating BoHV-1 from BoHV-5 are limited and do not allow to localize recombination events between these closely related virus species, 13 PCR sequencing assays were developed to discriminate between BoHV-1 and BoHV-5 at regular intervals throughout the entire respective viral DNA genomes. These assays were used to determine the genetic background of two interspecific BoHV-1/-5 recombinants generated previously. The two crossover points where recombination events occurred between the parental strains were determined. This study provides a detailed analysis of two interspecific recombinant viruses generated in vitro from closely related alphaherpesviruses infecting the same natural host. It demonstrates that recombination can occur within very short fragments of sequence homology. This finding raises questions about the mechanisms involved in the strands exchange and resolution step of the homologous recombination used by herpesviruses. This method will allow monitoring generation of recombinants between closely related herpesvirus species both in vitro and in vivo.


Asunto(s)
ADN Viral/genética , Herpesvirus Bovino 1/crecimiento & desarrollo , Herpesvirus Bovino 1/genética , Herpesvirus Bovino 5/crecimiento & desarrollo , Herpesvirus Bovino 5/genética , Reacción en Cadena de la Polimerasa/métodos , Recombinación Genética , Animales , Secuencia de Bases , Bovinos , Línea Celular , Herpesvirus Bovino 1/clasificación , Herpesvirus Bovino 5/clasificación , Datos de Secuencia Molecular , Alineación de Secuencia , Análisis de Secuencia de ADN
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