ABSTRACT
Bovine respiratory disease (BRD) is the costliest complex disease affecting the cattle industry worldwide, with significant economic losses. BRD pathogenesis involves several interactions between microorganisms, such as bacteria and viruses, and management factors. The present study aimed to characterize the nasal virome from 43 pooled nasal swab samples collected from Egyptian nonvaccinated cow-calf operations with acute BRD from January to February 2020 using metagenomic sequencing. Bovine herpesvirus-1 (BHV-1), first detection of bovine herpesvirus-5 (BHV-5), and first detection of bovine parvovirus-3 (BPV-3) were the most commonly identified in Egyptian cattle. Moreover, phylogenetic analysis of glycoprotein B revealed that the BHV-1 isolate is closely related to the Cooper reference strain (genotype 1.1), whereas the BHV-5 isolate is closely related to the reference virus GenBank NP_954920.1. In addition, the whole-genome sequence of BPV-3 showed 93.02% nucleotide identity with the reference virus GenBank AF406967.1. In this study, several DNA viruses, such as BHV-1 and first detection BHV-5, and BPV-3, were detected and may have an association with the BRD in Egyptian cattle. Therefore, further research, including investigating more samples from different locations to determine the prevalence of detected viruses and their contributions to BRD in cattle in Egypt, is needed.
Subject(s)
Cattle Diseases , Herpesvirus 1, Bovine , Herpesvirus 5, Bovine , Respiratory Tract Diseases , Viruses , Animals , Cattle , Cattle Diseases/epidemiology , Female , Herpesvirus 1, Bovine/genetics , Phylogeny , Virome , Viruses/geneticsABSTRACT
Objetivou-se descrever a ocorrência do Bovine alphaherpesvirus 5 (BoHV5) como causa de meningoencefalite não supurativa em bovinos do estado de Pernambuco, Brasil. Para tanto, 32 amostras de sistema nervoso embebidas em parafina foram obtidas de animais acometidos por doenças neurológicas atendidos na Clínica de Bovinos de Garanhuns da Universidade Federal Rural de Pernambuco (CBG-UFRPE), entre 2012 e 2016. As amostras foram analisadas quanto à presença do gene da glicoproteína C do BoHV5 por reação em cadeia da polimerase (PCR). Dois animais (6,25%) tiveram resultado positivo à PCR, e sua análise de sequenciamento indicou 100% de similaridade para o BoHV5. Os resultados histopatológicos desses dois animais revelaram lesões multifocais de meningoencefalite não supurativa associada à polioencefalomalácia, presença de corpúsculos de inclusão basofílicos, infiltração de células de Gitter e presença de manguitos perivasculares. A PCR se mostra uma importante ferramenta para diferenciação das infecções por BoHV5 de outras enfermidades neurológicas de bovinos, especialmente a raiva.(AU)
Subject(s)
Animals , Cattle , Herpesvirus 5, Bovine/isolation & purification , Meningoencephalitis/veterinary , Paraffin , Central Nervous System , Polymerase Chain Reaction/veterinary , Central Nervous System Viral Diseases/epidemiologyABSTRACT
The Bovine herpes virus type 5 glycoprotein D (gD) is essential for viral penetration into host permissive cells. The Herpes virus gD glycoprotein has been used for bovine immunization, being efficient in reduction of viral replication, shedding and clinical signs, however sterilizing immunity is still not achieved. Recombinant subunit vaccines are, in general, poorly immunogenic requiring additional adjuvant components. Interleukin 17A (IL17A) is a pro-inflammatory cytokine produced by T helper 17 cells that mediate mucosal immunity. IL17 production during vaccine-induced immunity is a requirement for mucosal protection to several agents. In this study, we investigated the potential of a recombinant IL17A to act as an adjuvant for a recombinant BoHV-5 glycoprotein D vaccine in cattle. Three cattle groups were divided as: group 1) rgD5 + alumen + rIL-17A; 2) rgD5 + alumen; and 3) PBS + alumen. The cattle (3 per group) received two doses of their respective vaccines at an interval of 21 days. The group that received rIL17 in its vaccine formulation at the 7th day after the prime immunization had significant higher levels of specific rgD-IgG than the alumen group. Addition of rIL17 also led to a significant fold increase in specific anti-rgD IgG and neutralizing antibodies to the virus, respectively, when compared with the alumen group. Cells stimulated with rIL17A responded with IL17 transcription, as well IL2, IL4, IL10, IL15, Bcl6 and CXCR5. Our findings suggest that the rIL17A has adjuvant potential for use in vaccines against BoHV-5 as well as potentially other pathogens of cattle.
Subject(s)
Antibodies, Viral/immunology , Cattle Diseases/prevention & control , Encephalitis, Viral/veterinary , Herpesviridae Infections/veterinary , Herpesvirus 5, Bovine/immunology , Herpesvirus Vaccines/immunology , Meningoencephalitis/veterinary , Adjuvants, Immunologic , Animals , Antibodies, Neutralizing/immunology , Cattle , Encephalitis, Viral/prevention & control , Herpesviridae Infections/prevention & control , Herpesvirus 5, Bovine/genetics , Immunization/veterinary , Interleukin-17/genetics , Interleukin-17/immunology , Meningoencephalitis/prevention & control , Vaccines, Synthetic , Viral Envelope Proteins/genetics , Viral Envelope Proteins/immunologyABSTRACT
Bacterial spores of the genus Bacillus are being evaluated as adjuvant molecules capable of improving the immune response to vaccines. In this study, we investigate whether subcutaneously administered spores of B. toyonensis BCT-7112T could enhance a vaccine immune response in mice. Three groups of mice were subcutaneously vaccinated on day 0 and received a booster on day 21 of the experiment, with the following vaccine formulations: 40 µg of recombinant glycoprotein D (rgD) from bovine herpesvirus type 5 (BoHV-5) adsorbed in 10% aluminum hydroxide (alum) without B. toyonensis spores (group 1) and B. toyonensis (1 × 106 viable spores) + 40 µg of rgD adsorbed in 10% alum (group 2); and B. toyonensis (1 × 106 viable spores) without rgD (group 3). Group 2 showed significantly higher titers (P < 0.05) of total specific serum IgG, IgG2a, and neutralizing antibodies, when compared with the groups 1 and 3. A significantly higher (P < 0.05) transcription level of cytokines IL-4, IL-12, and IFN-γ was observed in splenocytes from mice that received the B. toyonensis spores in the vaccine formulation. In addition, stimulation of the macrophage-like cell line RAW264.7 with spores of B. toyonensis markedly enhanced the cell proliferation and mRNA transcription levels of IL-4, and IL-12 cytokines in these cells. Our findings indicated that the subcutaneous administration of B. toyonensis BCT-7112T spores enhanced the humoral and cellular immune response against BoHV-5 in mice.
Subject(s)
Adjuvants, Immunologic , Bacillus , Herpesviridae Infections/prevention & control , Viral Vaccines/immunology , Animals , Bacillus/immunology , Disease Models, Animal , Herpesvirus 5, Bovine , Interleukin-12 , Interleukin-4 , Mice , Oligopeptides , Spores, Bacterial/immunologyABSTRACT
Neurotrophic factors have been implicated in the control of neuronal survival and plasticity in different brain diseases. Meningoencephalitis caused by bovine alpha-herpesvirus 5 (BoHV-5) infection is a frequent neurological disease of young cattle, being the involvement of apoptosis in the development of neuropathological changes frequently discussed in the literature. It's well known that Toll-like receptors (TLRs) can activate neuroinflammatory response and consequently lead to neuronal loss. However, there are no studies evaluating the expression of neurotrophic factors and their association with brain pathology and TLRs during the infection by BoHV-5. The current study aimed to analyze brain levels of neurotrophic factors along with neuropathological changes during acute infection by BoHV-5 in wild-type (WT) and TLR3/7/9 (TLR3/7/9-/-) deficiency mice. The infection was induced by intracranial inoculation of 1 × 104 TCID50 of BoHV-5. Infected animals presented similar degrees of clinical signs and neuropathological changes. Both infected groups had meningoencephalitis and neuronal damage in CA regions from hippocampus. BoHV-5 infection promoted the proliferation of Iba-1 positive cells throughout the neuropil, mainly located in the frontal cortex. Moreover, significant lower levels of brain-derived neurotrophic factor (BDNF) were detected in both BoHV-5 infected WT and TLR3/7/9 deficient mice, compared with non-infected animals. Our study showed that BDNF down regulation was associated with brain inflammation, reactive microgliosis and neuronal loss after bovine alpha-herpesvirus 5 infection in mice. Moreover, we demonstrated that combined TLR3/7/9 deficiency does not alter those parameters.
Subject(s)
Brain-Derived Neurotrophic Factor/metabolism , Cattle Diseases/metabolism , Herpesviridae Infections/veterinary , Herpesvirus 5, Bovine , Toll-Like Receptors/deficiency , Animals , Brain-Derived Neurotrophic Factor/genetics , Cattle , Cattle Diseases/virology , Down-Regulation , Herpesviridae Infections/metabolism , Mice , Toll-Like Receptor 3/deficiency , Toll-Like Receptor 7/deficiency , Toll-Like Receptor 9/deficiencyABSTRACT
Spores of the genus Bacillus are molecules capable of increasing the vaccine adjuvanticity. Bovine herpesvirus type 5 (BoHV-5) is responsible for meningoencephalitis that causes important economic losses in cattle. BoHV-5 glycoprotein D (gD) is a target of vaccine antigen and plays an important role in host cell penetration. The present study aimed to evaluate the adjuvanticity of Bacillus toyonensis (B.t) spores, live and heat-killed, associated with a vaccine formulated with aluminum hydroxide (alum) and the recombinant BoHV-5 glycoprotein D (rgD) in an experimental murine model. Six experimental groups of mice were subcutaneously vaccinated on day 0 and received a booster on day 21 of the experiment, with the following vaccine formulations: rgD (40 µg) + live spores (2 × 109 CFU); rgD + killed spores; rgD + live spores + alum (2.0 mg); rgD + killed spores + alum; rgD + alum, and rgD + PBS. Mice from rgD + live spores group showed an increase in rgD IgG titers from the 21st day until the end of the experiment. The groups of live and killed spores, associated to alum, had similar levels of IgG titers with no significant difference between each other; however, by the 14th and 28th day until the end of the experiment, presented higher IgG titers in comparison to the rgD + alum group. Moreover, increased serum levels of IgG1, IgG2a, and IgG2b were detected in mice that received spores in the vaccine formulation. The spores associated with alum groups showed neutralizing BoHV-5 antibodies and high mRNA transcription of the cytokines IFN-γ (66-fold), IL-17 (14-fold), and IL-12 (2.8-fold). In conclusion, our data demonstrated that the B. toyonensis spores, live or killed, associated with alum increased the adjuvanticity for BoHV-5 rgD in mice, suggesting the use of B. toyonensis spores as a promising component for vaccine formulations.
Subject(s)
Bacillus , Herpesvirus 1, Bovine , Herpesvirus 5, Bovine , Adjuvants, Immunologic , Alum Compounds , Animals , Antibodies, Viral , Cattle , Immunity , Mice , Spores , Vaccines, SubunitABSTRACT
Rabies and herpetic encephalitis are the main viral infections in bovines with neurological symptoms. Bovine rabies has a high prevalence in Central and South America, while bovine encephalitis associated with herpesvirus is especially important in South America. Viral isolation is the classical way to confirm herpesvirus infection, but molecular evidence of the presence of the virus in affected animals is gaining importance in the diagnosis of the disease in the laboratory. This study investigated the presence of herpesvirus type 1 and 5 (BoHV-1 and BoHV-5) in 182 encephalon of rabies-suspected cattle in Rio Grande do Sul state (RS), Brazil using multiplex real-time polymerase chain reaction (mRT-PCR). The rabies virus was investigated by direct fluorescent antibody assay and intracerebral suckling mouse inoculation. The genomes of BoHV-1 and BoHV-5 were detected in 17% of samples. BoHV-5 and BoHV-1 were detected in 100% and 19% of BoHV positive samples, respectively, indicating the circulation of the pathogens in cattle herds in RS. The high Ct values and the absence of isolation suggest viral latency. Coinfection of herpesvirus and the rabies virus was detected in 28% of samples, although no significant association between pathogens was observed. Rabies was detected in 57.7% of suspected samples, confirming the importance of the disease in the state. Concerning the method by which samples were conserved, no significant difference was observed between the number of positive results in frozen and refrigerated samples.
Subject(s)
Cattle Diseases/virology , Herpesviridae Infections/veterinary , Herpesvirus 1, Bovine/genetics , Herpesvirus 5, Bovine/genetics , Rabies/veterinary , Animals , Brain/virology , Brazil/epidemiology , Cattle , Cattle Diseases/epidemiology , Cryopreservation/veterinary , DNA, Viral/isolation & purification , Herpesviridae Infections/epidemiology , Herpesviridae Infections/virology , Mice , Multiplex Polymerase Chain Reaction/veterinary , Rabies/epidemiology , Refrigeration/veterinaryABSTRACT
Helminths have developed complex mechanisms to suppress the host immune response. These mechanisms may impair the host vaccine response. This study aimed to evaluate the effect of Toxocara spp. infection on the vaccine immune response to bovine herpesvirus type 5 (BoHV-5). First, 30 heifers received two doses of an experimental BoHV-5 vaccine. At 42nd days after the primo vaccination the vaccine efficacy was evaluated, and the presence of anti-Toxocara antibodies. Second, 20 Balb/c mice were divided into two groups, one infected with T. canis and the other without infection. After infection, both groups received two doses of vaccine. The vaccine immune response was assessed by BoHV-5 serum neutralization and splenic cytokines transcription by qPCR. All heifers positive for Toxocara spp. (40%) showed BoHV-5 SN titer ≤1:32, whereas heifers negative for Toxocara spp. (60%) had BoHV-5 SN titer ≥1: 128. Infected T. canis mice showed BoHV-5 SN titer ≤1:2, whereas mice not infected with T. canis BoHV-5 SN titer ≥1:8. Splenocytes from control mice stimulated with BoHV-5 had a significant (p < .05) mRNA transcription for the cytokines IL-12, IL-17, and IL-23, whereas the same cytokines were down-regulated in T. canis infected mice. These results suggest that Toxocara spp. infection may impair BoHV-5 immunization and should be considered for efficient herd immunization.
Subject(s)
Cattle Diseases/prevention & control , Herpesviridae Infections/veterinary , Herpesvirus 5, Bovine , Toxocara canis , Toxocariasis/pathology , Animals , Cattle , Cattle Diseases/immunology , Female , Herpesviridae Infections/prevention & control , Mice , Spleen , Toxocariasis/immunology , Vaccination/veterinaryABSTRACT
Bovine herpesvirus (BoHV) types 1 and 5 are two closely related alpha-herpesviruses of cattle with neuroinvasive potential. BoHV-5 causes non-suppurative meningoencephalitis in calve whereas encephalitis caused by BoHV-1 has been occasionally reported. As an initial step to understand the biology of both BoHV types in neural cells, undifferentiated SH-SY5Y human neuroblastoma cells were infected with BoHV-1 strains Cooper and Los Angeles (LA), BoHV-5 strain 97/613 and A663, a BoHV-5/BoHV-1 natural recombinant. Cytopathic effect (CPE) in these cells was evident earlier for BoHV-5 strain 97/613 and CPE progression was slower for BoHV-1, particularly for Cooper strain. Virus antigen was detected as early as 8 h post-infection (hpi) for all strains, with the exception of BoHV-1 Cooper for which antigen expression was detectable by 24 hpi. All strains released detectable infectious virus in the extracellular medium by 8 hpi, confirming that undifferentiated SH-SY5Y cells are fully permissive to BoHV infection. Significantly different extracellular virus titers among the different strains were detected by 24 hpi, with BoHV-5 97/613 reaching the maximal virus production. The lowest extracellular titer was recorded for BoHV-1 Cooper at all the evaluated time-points. BoHV-1 Cooper, BoHV-1 LA and BoHV-5 97/613 had a steady increase in intracellular virus production. The evaluation of lysis plaques formation revealed that BoHV-5 A663 produced the largest plaques followed by BoHV-5 97/613. Both BoHV-1 strains produced smaller plaques when compared with BoHV-5. Despite a slower replicative cycle, strain A663 is more efficient in cell to cell dissemination. Thus, it is evident that BoHV-5 strains have growth advantages in undifferentiated neural cells compared with BoHV-1. This in vitro model might be useful to analyze the neuropathogenic potential of bovine alphaherpesviruses.
Subject(s)
Herpesvirus 1, Bovine/physiology , Herpesvirus 5, Bovine/physiology , Neurons/virology , Virus Replication , Animals , Antigens, Viral/genetics , Cattle , Cattle Diseases/virology , Cell Line , Herpesvirus 1, Bovine/genetics , Herpesvirus 5, Bovine/genetics , Humans , Neuroblastoma , Viral LoadABSTRACT
Bovine herpesvirus 1 (BoHV1) and 5 (BoHV5) are genetically and antigenically related alphaherpesviruses. Infection with one virus induces protective immunity against the other. However, disease associated with BoHV1 and BoHV5 varies significantly; whereas BoHV1 infection is usually associated with rhinotracheitis and abortion, BoHV5 causes encephalitis in cattle. BoHV5 outbreaks are sporadic and mainly restricted to the South American countries. We report BoHV5 infection for the first time from aborted cattle in India. Based on the characteristic cytopathic effects in MDBK cells, amplification of the viral genome by PCR, differential PCR for BoHV1/BoHV5, nucleotide sequencing and restriction endonuclease patterns, identity of the virus was confirmed as BoHV5 subtype A. Serum samples from the aborted cattle strongly neutralized both BoHV1 and BoHV5 suggesting an active viral infection in the herd. Upon UL27, UL44 and UL54 gene-based sequence and phylogenetic analysis, the isolated virus clustered with BoHV5 strains and showed highest similarity with the Brazilian BoHV5 strains.
Subject(s)
Herpesvirus 5, Bovine/genetics , Herpesvirus 5, Bovine/isolation & purification , Herpesvirus 5, Bovine/metabolism , Alphaherpesvirinae/genetics , Animals , Antibodies, Viral/blood , Cattle , Cattle Diseases/epidemiology , Disease Outbreaks/veterinary , Genome, Viral/genetics , Herpesviridae Infections/virology , Herpesvirus 1, Bovine/genetics , Herpesvirus 1, Bovine/metabolism , India , PhylogenyABSTRACT
Bovine herpesvirus 5 (BoHV-5) is a pathogen of cattle responsible for fatal meningoencephalitis. Like alpha herpesvirus subfamily members, BoHV-5 also encodes microRNA in lytic infections of epithelial cells. BoHV-5-miR-B10 was the most abundant miRNA detected in a high-throughput sequencing study. Here, we evaluated the kinetics of miR-B10 expression after BoHV-5 productive infection by stem-loop real-time quantitative PCR. miR-B10 candidate target sites in the virus were predicted, and BoHV-5 UL39 was confirmed as a target gene by dual-luciferase assay with the design of an miR-B10 tough decoy (TuD). The UL39 gene encoding ribonucleotide reductase (RR) large subunit plays an important role in the early stage of BoHV-5 lytic infection. As BoHV-5-miR-B10 is located in internal and terminal repeat regions, we generated a TuD gene-integrated BoHV-5 strain, which effectively down-regulated miR-B10-3p. Strikingly, the suppression of miR-B10-3p significantly improved BoHV-5 replication. Taking these findings together, our study established an efficient method to deliver and express TuD RNA for viral miRNA suppression, and demonstrated that virus-encoded miRNA suppresses viral-genome biogenesis with a feedback mode, which might serve as a brake for viral replication. Herpesviruses infect humans and a variety of animals. Almost all herpesviruses can encode miRNAs, but the functions of these miRNAs remain to be elucidated. Most herpesvirus-encoded miRNA harbours dual copies, which is difficult to be deleted by current genetic modulation. Here, we developed an efficient method to deliver and express TuD RNA to efficiently suppress viral miRNA with multiple copies. Using this method, we demonstrated for the first time that viral miRNA feedback regulates viral replication by suppressing the expression of RR.
Subject(s)
Cattle Diseases/virology , Encephalitis, Viral/virology , Feedback, Physiological , Herpesviridae Infections/virology , Herpesvirus 5, Bovine/genetics , Meningoencephalitis/virology , MicroRNAs/metabolism , Virus Replication/genetics , Animals , Cattle , Gene Expression Regulation, Viral , HEK293 Cells , Humans , MicroRNAs/genetics , RNA, Viral/genetics , Real-Time Polymerase Chain ReactionABSTRACT
Bovine herpesvirus types 1 (BoHV-1) and 5 (BoHV-5) are two closely related alphaherpesviruses. BoHV-1 causes several syndromes in cattle, including respiratory disease and sporadic cases of encephalitis, whereas BoHV-5 is responsible for meningoencephalitis in calves. Although both viruses are neurotropic, they differ in their neuropathogenic potential. This review summarizes the findings on the specific mechanisms and pathways known to modulate the pathogenesis of BoHV-1 and BoHV-5, particularly in relation to respiratory and neurological syndromes, which characterize BoHV-1 and BoHV-5 infections, respectively.
Subject(s)
Cattle Diseases/virology , Herpesviridae Infections/veterinary , Herpesviridae Infections/virology , Herpesvirus 1, Bovine/pathogenicity , Herpesvirus 5, Bovine/pathogenicity , Host-Pathogen Interactions/physiology , Animals , Cattle , HumansABSTRACT
BACKGROUND: Bovine herpesvirus type 5 is an important agent of meningoencephalitis in cattle and has been identified in outbreaks of bovine neurological disease in several Brazilian states. In recent years, oxoquinoline derivatives have become an important focus in antiviral drug research. METHODS: The cytotoxicity and anti BoHV-5RJ42/01 activity of a set of synthetic 4-oxoquinoline derivatives 4a-k were assayed on Madin-Darby Bovine Kidney cell and antiviral activity by plaque reduction assay. RESULTS: The most promising substance (4h) exhibited CC50 and EC50 values of 1,239 µM ±5.5 and 6.0 µM ±1.5, respectively, with an SI =206. Two other compounds 4j (CC50 = 35 µM ±2 and EC50 = 24 µM ±7.0) and 4k (CC50= 55 µM ±2 and EC50 = 24 µM ±5.1) presented similar inhibitory profile and selectivity indexes of 1.4 and 2.9, respectively. The results of the time-of-addition studies revealed expressive reduction of virus production (≥80%) in different stages of virus replication cycle except for compound 4h that slightly inhibited virus yield in the first 2 h post infection, but it showed expressive virus inhibition after this time. CONCLUSIONS: All three compounds slightly interact with the virus on the virucidal assay and they are not able to block virus attachment and penetration. Antiviral effect of oxoquinoline 4h was more prominent than acyclovir which leads us to suggest compound 4h as a promising molecule for further anti-BoHV-5 drug design.
Subject(s)
4-Quinolones/pharmacology , Antiviral Agents/pharmacology , Herpesvirus 5, Bovine/drug effects , Animals , Dogs , Madin Darby Canine Kidney Cells/drug effects , Viral Plaque Assay , Virus Inactivation/drug effects , Virus Replication/drug effectsABSTRACT
Bovine alphaherpesvirus types 1 (BoHV-1) and 5 (BoHV-5) are closely related alphaherpesviruses. BoHV-5 causes non-suppurative meningoencephalitis in calves. BoHV-1 is associated with several syndromes and, occasionally, can cause encephalitis. Although both viruses are neurotropic and they share similar biological properties, it is unknown why these alphaherpesviruses differ in their ability to cause neurological disease. Neural tissue samples were collected from BoHV-1- and BoHV-5-intranasally inoculated calves during acute infection, latency and reactivation and the levels of cyclins mRNA expression were analyzed by qRT-PCR. Striking differences in the levels of cyclins mRNA were particularly detected in trigeminal ganglion (TG). The expression levels of cyclins in TG during BoHV-5 latency suggest that these viruses utilize different strategies to persist in the host. It is apparent that a relationship between virus loads and cyclin mRNA levels can be established only during acute infection and other factors might be involved in the regulation of cell cycle components during BoHV latency and reactivation. Bovine alphaherpesviruses neuropathogenicity might be influenced by the differential control of cell cycle components by these herpesviruses. This is the first report on BoHV-5 modulation of cyclins expression in neural tissues from its natural host.
Subject(s)
Cattle Diseases/pathology , Cyclins/biosynthesis , Encephalitis, Viral/veterinary , Herpesviridae Infections/veterinary , Herpesvirus 1, Bovine/growth & development , Herpesvirus 5, Bovine/growth & development , Meningoencephalitis/veterinary , RNA, Messenger/biosynthesis , Animals , Brain/pathology , Cattle , Cyclins/genetics , Encephalitis, Viral/pathology , Gene Expression Profiling , Herpesviridae Infections/pathology , Meningoencephalitis/virology , RNA, Messenger/genetics , Real-Time Polymerase Chain ReactionABSTRACT
Bovine herpesvirus type 1 and type 5 (BoHV-1 and BoHV-5) are two alphaherpesviruses that affect cattle with two different syndromes. While BoHV-1 mainly produces respiratory symptoms, BoHV-5 is highly neuropathogenic and responsible for meningoencephalitis in young cattle. The latency-related (LR) gene, which is not conserved between these two herpesviruses, is the only viral gene abundantly expressed in latently infected neurons. The antiapoptotic action of this gene has been demonstrated during acute infection and reactivation from latency and seems to be mainly mediated by a LR protein (ORF-2) which is truncated in amino acid 51 in the case of BoHV-5. In this work, we show that the BoHV-5 LR gene is less efficient at cell survival and apoptosis inhibition in transient as well as in established neuronal cell lines compared to its BoHV-1 homolog. We hypothesize that the BoHV-5 LR gene may have novel functions that are lacking in the BoHV-1 LR gene and that these differences may contribute to its enhanced neuropathogenesis.
Subject(s)
Herpesviridae Infections/veterinary , Herpesvirus 1, Bovine/genetics , Herpesvirus 5, Bovine/genetics , Infectious Bovine Rhinotracheitis/metabolism , Meningoencephalitis/veterinary , Viral Proteins/genetics , Virus Latency/genetics , Animals , Apoptosis/genetics , Cattle , Cell Line , Gene Expression , Herpesviridae Infections/pathology , Herpesviridae Infections/virology , Herpesvirus 1, Bovine/growth & development , Herpesvirus 1, Bovine/metabolism , Herpesvirus 5, Bovine/growth & development , Herpesvirus 5, Bovine/metabolism , Host-Pathogen Interactions/genetics , Infectious Bovine Rhinotracheitis/pathology , Infectious Bovine Rhinotracheitis/virology , Meningoencephalitis/pathology , Meningoencephalitis/virology , Neurons/metabolism , Neurons/virology , Protein Isoforms/genetics , Protein Isoforms/metabolism , Viral Proteins/metabolism , Virus ActivationABSTRACT
Bovine alphaherpesviruses 1 and 5 (BoHV-1/5) are main pathogens of respiratory, reproductive and neurological diseases in cattle. The aim of this study was to investigate the frequency of neutralizing antibodies against BoHV-1/5 in serum samples and to detect viral DNA in semen of bulls from beef cattle farms located in RS. A total of 372 serum and semen sample from bulls were collected in eighteen farms. Serum samples were submitted to virus neutralization (VN) assay, while semen samples were used to detect BoHV-1 and BoHV-5 DNA by PCR. VN results showed that BoHV-1/5 antibodies were detected in bulls of 66.7% (12/18) of the farms, 295 (79.5%) BoHV positive bulls, 287 for BoHV-1 and 234 for BoHV-; at 43 vaccinated bulls 72.1% (31/43) showing serology negative. BoHV-1/5 DNA was detected in the semen of three bulls; one of the them presenting BoHV-1, one out three presenting BoHV-5 and one BoHV-1/5.co-infection All BoHV DNA positive samples came from animals presenting posthitis and other genital lesions at sampling. Results showed a high seroprevalence of BoHV-1/5 antibodies in bulls as well as strong evidence that these viruses are actively circulating in the cattle farms. A remarkable finding is that in the presence of clinically evident lesions in the genital tract, both BoHV-1 and 5 may found in semen.(AU)
Os alfa-herpesvírus bovinos 1 e 5 (BoHV-1/5) são importantes patógenos de doença respiratória, reprodutiva e neurológica em bovinos. O objetivo deste estudo foi investigar a frequência de detecção de anticorpos neutralizantes contra BoHV-1/5 em amostra de soro e detectar DNA viral em sêmen de touros do rebanho bovino localizado nas fazendas de gado de corte do RS. Um total de 371 amostras de soro e sêmen foi coletado de touros em 18 fazendas, 325 das quais são provenientes de touros não vacinados e 43 de vacinados. Amostras de soro foram submetidas à técnica de vírus-neutralização (VN), enquanto as amostras de sêmen foram submetidas à extração de DNA e posterior PCR (polymerase chain reaction) para detecção de BoHV-1 e 5. Os resultados da VN demostraram que anticorpos contra BoHV-1/5 foram detectados nos touros não vacinados em 66,7% (12/18) das fazendas, 295 (79,5%) touros mostraram-se positivos para BoHV, 287 para BoHV-1 e 234 para BoHV-5; e para 43 touros vacinados, observou-se que 72,1% (31/43) foram negativos na sorologia DNA de BoHV-1/5, detectado no sêmen de três touros: um deles apresentava BoHV-1, outro BoHV-5 e em um foi detectada coinfecção por BoHV-1/5. Todas as amostras positivas para o DNA viral eram provenientes de animais que apresentavam lesões de postite e outras lesões genitais. Esses resultados demonstram que há uma alta soroprevalência de BoHV-1/5 em touros, bem como uma forte evidência de que esses vírus estão circulando ativamente no rebanho bovino dessas fazendas. Um achado interessante foi a detecção de BoHV-1 e 5 em touros com lesões na região do trato genital.(AU)
Subject(s)
Animals , Cattle , Wounds and Injuries , Cattle/genetics , Herpesvirus 1, Bovine/genetics , Herpesvirus 5, BovineABSTRACT
This article has been retracted: please see Elsevier Policy on Article Withdrawal (https://www.elsevier.com/about/our-business/policies/article-withdrawal). This article has been retracted at the request of the Editors-in-Chief and Authors. Fig 1A is a duplicate of a figure that has already been published in da Silva SEL et al. Archives of Virology 2018;163:1043-1049; 10.1007/s00705-018-3704-2. These two papers report studies performed with cells from two different animal species (bovine cells for the Veterinary Microbiology paper and chicken cells for the Archives of Virology paper). The reuse of the same figure in the Veterinary Microbiology paper to describe cells that were supposed to be from a different species is thus inappropriate and also puts into question the reliability of the other results presented in this paper. In addition, the Editors-in-Chief have remaining concerns about the strong similarities of other data presented in the two papers. Even if these concerns were addressed, the re-use of any data has to be clearly indicated and appropriately cited. As such this article represents a misuse of the scientific publishing system. The scientific community takes a very strong view on this matter and apologies are offered to readers of the journal that this was not detected during the submission process.
Subject(s)
Herpesvirus 5, Bovine , Macrophages/virology , Mitochondria/pathology , Virus Replication/physiology , Animals , Cattle , Cell Proliferation , Cell Survival , Cells, Cultured , Macrophages/metabolism , Membrane Potential, Mitochondrial , Nitric OxideABSTRACT
Bovine herpesvirus 5 is an alphaherpesvirus that causes nonsuppurative meningoencephalitis in cattle. This disease occurs naturally in either outbreaks or isolated cases, and exhibits low morbidity and high lethality. Although previous studies elucidated crucial aspects involved in the pathogenesis of the disease, there is a paucity of information regarding the molecular events contributing to infection and replication of BoHV-5. The objective of the present study was to determine the in vitro gene expression pattern of BoHV-5 (e.g., alpha, beta, and gamma genes) and host cells genes (GAPDH and 18S) over time utilizing different quantities of inoculated virus. Three BoHV-5 genes (bICP0, UL9, US4) and one structural bovine cell gene had their expression accessed by real-time PCR. While the expression of BoHV-5 genes increased during the course of infection, GAPDH gene expression decreased in the host cells, evidencing the effect of viral infection on the expression of bovine cell genes. The 18S ribosomal RNA (rRNA) gene was constitutively expressed throughout BoHV-5 infection. Our data clearly demonstrates that GAPDH gene should not be used as a reference gene in studies of BoHV-5 infection because it was influenced by viral infection. However, 18S rRNA was constitutively expressed and, therefore, is recommended for normalization of BoHV-5 infection studies in bovine cells. The expression of viral genes transcripts was not altered by increasing number of viral particles added to the culture. All viral genes included here demonstrated the same expression pattern over time and there was no difference in the expression of viral genes among the various time points. Our data show important differences comparing to classical studies regarding herpesvirus alpha, beta, and gamma genes expression. More research is necessary to improve our understanding about the BoHV-5 biology during infection. Studies employing next-generation sequencing (i.e., RNA-seq), using both in vitro and in vivo models, would be the next logical step to grasp the virus and host cell's transcriptome changes over the course of infection.(AU)
Herpesvirus bovino 5 é um alfaherpesvírus causador de meningoencefalite não supurativa em bovinos. Esta doença possui ocorrência natural em surtos ou casos isolados, associadas a baixa morbidade e alta letalidade. Embora estudos anteriores tenham elucidado aspectos relacionados a patogenia da doença, há uma lacuna de conhecimento relacionado aos eventos moleculares que contribuem para a infecção e replicação do BoHV-5. O objetivo do presente estudo foi determinar a expressão gênica in vitro de genes virais (i.e., alfa, beta e gama) e das células hospedeiras (GAPDH e 18S) durante a infecção considerando diferentes momentos de infecção e quantidade de vírus utilizado. Três genes do BoHV-5 (bICP0, UL9, US4), um gene estrutural (GAPDH) e um gene constitutivo (18S) da célula bovina tiveram suas expressões avaliadas por PCR quantitativa (qPCR). Enquanto os genes virais tiveram sua expressão aumentada ao longo do tempo de infecção, o gene hospedeiro teve sua expressão diminuída, demonstrando a ação do vírus na expressão gênica de células bovinas in vitro. O gene constitutivo 18S teve sua expressão mantida durante todos os momentos do experimento. Nossos resultados claramente demonstraram que o GAPDH não deve ser usado como gene de referência em estudos com infecção por BoHV-5 pois é influenciado pela infecção viral. Entretanto, o 18S rRNA foi constitutivamente expresso e pode ser recomendado para normalização em células bovinas infectadas pelo vírus. A expressão de mRNA viral não foi alterada pela quantidade de vírus usada. Todos os genes virais demonstraram o mesmo padrão de expressão ao longo do tempo de infecção. Nossos resultados trazem importantes diferenças comparando aos estudos clássicos que avaliaram a expressão de genes alfa, beta e gama. Mais estudos são necessários para aumentar o conhecimento da biologia molecular do BoHV-5. Estudo utilizando sequenciamento de última geração (i.e., RNA-seq), usando modelos in vitro e in vivo, aparentam ser o próximo passo lógico para acessar as alterações do transcriptoma do hospedeiro e viral ao longo do curso da infecção.(AU)
Subject(s)
Gene Expression , Polymerase Chain Reaction/veterinary , Herpesvirus 5, Bovine/classification , Molecular BiologyABSTRACT
Bovine herpesvirus 5 (BoHV-5) is responsible for outbreaks of meningoencephalitis that cause important economic losses in young cattle. BoHV-5 glycoprotein D (gD5) is essential for attachment and penetration into permissive cells and targeting of host immune systems, inducing strong humoral and cellular immune responses. The aim of this study was to evaluate the vaccinal immune response of vaccines formulated with the recombinant BoHV-5 gD (rgD5) in bovines. For the experiment, 72 heifers were randomly allotted into 6 different groups with 12 animals each. Group 1: vaccine formulated using inactivated BoHV-5 (iBoHV-5) adjuvanted with ISA50V2; Group 2: iBoHV-5 associated with 100⯵g of rgD5 adjuvanted with ISA50V2; Group 3: 100⯵g of rgD5 adjuvanted with ISA50V2; Group 4: 100⯵g of rgD5 adjuvanted with Al(OH)3; Group 5: commercial vaccine; and Group 6: control group. Two doses were administered in a 26-day interval and the third after 357â¯days from primo vaccination. Cattle vaccinated with the vaccines formulated with iBoHV-5 plus rgD5 showed a significant (pâ¯<â¯0.01) five-fold increase in total immunoglobulin G (IgG) for BoHV-5, BoHV-1, and rgD5 as compared with the commercial and control groups. Also, a significant (pâ¯<â¯0.05) increase in IgG1 and IgG2a levels was induced in serum for rgD5. In addition, these same vaccines showed significant (pâ¯<â¯0.01) four-fold higher titers of BoHV-1 and -5 neutralizing antibodies. The results demonstrated that the rgD5 conserved important epitopes that were able to stimulate bovine humoral immunity response capable of viral neutralization of BoHV-1 and -5, suggesting it as a promising vaccine antigen to be used in vaccine for BoHV-1 and -5 endemic areas.
Subject(s)
Cattle Diseases/prevention & control , Herpesviridae Infections/veterinary , Viral Envelope Proteins/immunology , Viral Vaccines/immunology , Animals , Antibodies, Neutralizing/blood , Antibodies, Viral/blood , Antigens, Viral/genetics , Antigens, Viral/immunology , Cattle , Cattle Diseases/immunology , Female , Herpesviridae Infections/immunology , Herpesviridae Infections/prevention & control , Herpesvirus 1, Bovine/immunology , Herpesvirus 5, Bovine/immunology , Immunization Schedule , Immunoglobulin G/blood , Recombinant Proteins/genetics , Recombinant Proteins/immunology , Vaccines, Synthetic/administration & dosage , Vaccines, Synthetic/genetics , Vaccines, Synthetic/immunology , Viral Envelope Proteins/genetics , Viral Vaccines/administration & dosage , Viral Vaccines/geneticsABSTRACT
Bovine herpesvirus 1 and 5 (BoHV-1 and -5) are antigenically and genetically related and can establish latent infection. We aimed to analyze the applicability of the milk sample to detect latently BoHV-infected cattle. BoHV-1 non-vaccinated clinically healthy cows from five dairy cattle herds (herd 1, n=24; herd 2, n=39; herd 3, n=39; herd 4, n=36; herd 5, n=70) were studied. We confirmed the presence of BoHV-1, and for the first time, BoHV-5 in the milk of naturally infected dairy cattle.