RESUMO
Despite SRLV infection being endemic in Mexico, there is little information regarding which genotypes are present. We compared serotyping and PCR-sequencing results from sheep and goats infected with SRLV. We separated plasma and peripheral blood leukocytes (PBL) from 1940 blood samples from sheep and goats from 12 states across Mexico. To detect SRLV infection, we tested plasma samples using two commercial ELISA kits (VMRD and Eradikit SRLV Screening). Then, we serotyped the infecting virus (A/ B) using Eradikit SRLV Genotyping. PBL DNA was used to detect the proviral genome via PCR. Positive amplicons were sequenced to identify viral genotypes using a phylogenetic analysis. Also, we analysed for residues differences in the sequences of a capsid epitope between genotypes. The serological results indicated a higher detection of seropositive animals using the VMRD ELISA compared to Eradikit, with 21 % and 15.3 % more in sheep and goats respectively. Only 25.7 % of the ELISA serotyping results matched those from PCR-sequencing. PCR-sequencing was able to identify genotype A, B and coinfections in animals classified as indeterminate by the ELISA test. This lack of sensitivity may be related to the lack of epitopes from the matrix and transmembrane peptides used by ELISA screening. Sequences analysis revealed that SRLVs found in sheep cluster with genetic subtypes A2 and B1, while those in goats cluster with subtypes A1 and B1. Serotyping did not prove to be an adequate method for predicting the viral genotype (A and / or B) in infections caused by SRLV.
Assuntos
Anticorpos Antivirais/sangue , Doenças das Cabras/virologia , Infecções por Lentivirus/veterinária , Lentivirus/imunologia , Doenças dos Ovinos/virologia , Animais , Ensaio de Imunoadsorção Enzimática/veterinária , Genótipo , Cabras , Lentivirus/genética , Lentivirus/isolamento & purificação , Infecções por Lentivirus/virologia , Filogenia , Reação em Cadeia da Polimerase/veterinária , Ruminantes , Sensibilidade e Especificidade , Sorotipagem/veterinária , OvinosRESUMO
Caprine arthritis encephalitis (CAE) is a chronic disease caused by a retrovirus from the Lentivirus genus. No effective vaccines or treatments exist, and therefore genetic selection for CAE resistance might be a feasible alternative. To our best knowledge, no other studies have investigated the genetic architecture of CAE resistance in dairy goats. In this context, this study was designed to estimate genetic parameters for CAE infection in Alpine and Saanen goats using a Bayesian threshold model. A total of 542 adult goats (and >3-generation pedigree), which were group-housed in a population with high CAE prevalence, were tested based on a serological infection assessment test (negative = 1 or positive = 2) and used for this study. Genetic parameters were estimated using the BLUPF90 family programs. There was considerable genetic variability for CAE resistance, and pedigree-based heritability was significantly different from zero (0.026 < heritability < 0.128). Our findings indicate that the prevalence of CAE in goat herds can be reduced or eliminated through direct genetic selection for CAE resistance in addition to proper management strategies.
Assuntos
Vírus da Artrite-Encefalite Caprina , Predisposição Genética para Doença , Doenças das Cabras/virologia , Infecções por Lentivirus/veterinária , Animais , Teorema de Bayes , Doenças das Cabras/epidemiologia , Cabras , Infecções por Lentivirus/genética , Infecções por Lentivirus/virologiaRESUMO
Small ruminant lentiviruses (SRLV) have high genetic variability which results in different viral strains around the world. This create a challenge to design sensible primers for molecular diagnosis in different regions. This work proposes a protocol of duplex nested-PCR for the precise diagnosis of SRLV. The technique was designed and tested with the control strains CAEV Co and MVV 1514. Then, field strains were submitted to the same protocol of duplex nested-PCR. Blood samples of sheep and goats were tested with AGID and nested PCR with specific primers for pol, gag and LTR. The AGID results showed low detection capacity of positive animals, while the nested PCR demonstrated a greater capacity of virus detection. Results demonstrated that LTR-PCR was more efficient in detecting positive sheep samples, whereas gag-PCR allowed a good detection of samples of positive goats and positive sheep. In addition, pol-PCR was more efficient with goat samples than for sheep. Duplex nested PCR performed with standard virus samples and field strains demonstrated that the technique is more efficient for the detection of multiple pro-viral DNA sequences. This study demonstrated a successful duplex nested PCR assay allowing a more accurate diagnosis of SRLV.
Assuntos
Doenças das Cabras/virologia , Infecções por Lentivirus/veterinária , Reação em Cadeia da Polimerase/métodos , Doenças dos Ovinos/virologia , Animais , Primers do DNA/genética , DNA Viral/genética , Doenças das Cabras/diagnóstico , Cabras , Infecções por Lentivirus/diagnóstico , Infecções por Lentivirus/virologia , Ovinos , Doenças dos Ovinos/diagnósticoRESUMO
The caprine arthrite encephalite (CAE) is a disease that affects especially dairy goat. The virus shows compartmentalization features, that allows it to hide at certain times during the course of the disease, making it difficult to control. The present study was conducted to identify the major seminal plasma protein profile of goats infected by CAE and its associations with seroconversion using Western blotting. Two groups containing five males each, were used in this experiment. The first group was composed by seropositive animals and the control by seronegative confirmed by Western blotting and PCR. The semen was collected through artificial vagina and after that, two-dimensional electrophoresis and MALDI-TOF MS were used. Seventy-five spots were identified in the goat seminal plasma gels, equivalent to 13 different proteins with more expression. The similar proteins found in both groups and related to reproduction were spermadhesin Z13-like, bodhesin and bodhesin-2, Lipocalin, protein PDC-109-like, and albumin. In infected goats, proteases such as arisulfatase A have been identified, whose function probably is related to metabolism control of sulfatides, involved to virus control. The other ones were bifunctional ATP-dependent dihydroxyacetone kinase/FAD-AMP lyase, cathepsin F isoform X1, disintegrin and metalloproteinase domain-containing protein 2-like isoform X1, clusterin, carbonic anhydrase 2, electron transfer flavoprotein subunit beta, and epididymal secretory glutathione peroxidase. The results of this study show the reaction of the innate immune system against chronic infection of goats by CAE.
Assuntos
Vírus da Artrite-Encefalite Caprina/isolamento & purificação , Doenças das Cabras/diagnóstico , Infecções por Lentivirus/veterinária , Proteínas de Plasma Seminal/análise , Animais , Western Blotting/veterinária , Eletroforese em Gel Bidimensional/veterinária , Doenças das Cabras/virologia , Cabras/genética , Infecções por Lentivirus/diagnóstico , Infecções por Lentivirus/virologia , Masculino , Reação em Cadeia da Polimerase/veterinária , Sêmen/química , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/veterináriaRESUMO
Abstract Small ruminant lentiviruses (SRLV) have high genetic variability which results in different viral strains around the world. This create a challenge to design sensible primers for molecular diagnosis in different regions. This work proposes a protocol of duplex nested-PCR for the precise diagnosis of SRLV. The technique was designed and tested with the control strains CAEV Co and MVV 1514. Then, field strains were submitted to the same protocol of duplex nested-PCR. Blood samples of sheep and goats were tested with AGID and nested PCR with specific primers for pol, gag and LTR. The AGID results showed low detection capacity of positive animals, while the nested PCR demonstrated a greater capacity of virus detection. Results demonstrated that LTR-PCR was more efficient in detecting positive sheep samples, whereas gag-PCR allowed a good detection of samples of positive goats and positive sheep. In addition, pol-PCR was more efficient with goat samples than for sheep. Duplex nested PCR performed with standard virus samples and field strains demonstrated that the technique is more efficient for the detection of multiple pro-viral DNA sequences. This study demonstrated a successful duplex nested PCR assay allowing a more accurate diagnosis of SRLV.
Assuntos
Animais , Doenças dos Ovinos/virologia , Doenças das Cabras/virologia , Reação em Cadeia da Polimerase/métodos , Infecções por Lentivirus/veterinária , Doenças dos Ovinos/diagnóstico , DNA Viral/genética , Cabras , Ovinos , Doenças das Cabras/diagnóstico , Infecções por Lentivirus/diagnóstico , Infecções por Lentivirus/virologia , Primers do DNA/genéticaRESUMO
The transmission frequency of small ruminant lentiviruses (SRLVs) through the placenta is controversial and may be associated with breed susceptibility. In Mexico, SRLV infections in sheep have been poorly studied. This work explores the presence of antibodies and proviral DNA in Mexican Pelibuey sheep. Enzyme-linked immunosorbent assays (ELISAs; three commercial kits and two on the basis of synthetic peptides) and polymerase chain reaction (PCR; amplifying the long terminal repeat and gag segments) were performed to diagnose SRLV infection in 25 adult Pelibuey ewes with an average age of 2.5 years and 32 fetuses with gestational ages ranging from 40 to 90 days without clinical signs of SRLV. Two of the three commercial ELISAs and the synthetic peptide-based ones were positive for SRLV antibody detection in 28% and 24% of the ewes, respectively, whereas none of the fetuses were positive by any of the ELISAs. By PCR, 31% of the ewes and, interestingly, two fetuses were positive. Characteristic SRLV lesions were not found in the fetal and/or ewe tissues, including those with positive PCR results. These findings demonstrate the susceptibility of Pelibuey sheep to SRLV infection and the low transmission frequency through the placenta.
Assuntos
Infecções por Lentivirus/veterinária , Lentivirus Ovinos-Caprinos/isolamento & purificação , Doenças dos Ovinos/virologia , Animais , Ensaio de Imunoadsorção Enzimática/veterinária , Feminino , Infecções por Lentivirus/epidemiologia , Infecções por Lentivirus/virologia , Lentivirus Ovinos-Caprinos/classificação , México/epidemiologia , Ovinos , Doenças dos Ovinos/epidemiologiaRESUMO
This paper reports a first-time study performed in El Salvador on the presence or absence of antibodies to three important animal diseases in small ruminants. The work was conducted in the west and central departments of the country, selecting 42 and 43 cantons with an existing sheep and goat population, respectively. Serum samples were collected from 396 sheep and 335 goats and tested for seropositivity to Brucella (B.) spp. The specimens from goats were also tested for antibodies to caprine arthritis-encephalitis (CAE) virus. Four (1 %) sheep and none of the goats were seropositive by Rose Bengal test. All animals were negative by indirect ELISA (iELISA) for B. abortus. All animals were negative by iELISA for CAE. A total of 383 sheep and 330 goats underwent the single intradermal cervical tuberculin (SICT) test for tuberculosis. Seventy (18 %) sheep and 43 (13 %) goats reacted to the SICT test. Those reactors were subjected to the single intradermal comparative cervical tuberculin (SICCT) test, and one (0.3 %) goat was deemed to be a positive reactor. No mycobacteria were diagnosed in concluding analyses, and further studies are considered necessary to determine the prevalence of the investigated diseases. Additionally, it is recommended that small ruminants should be included in the national eradication program on bovine brucellosis and tuberculosis to prevent potential reservoirs.
Assuntos
Brucelose/veterinária , Doenças das Cabras/epidemiologia , Infecções por Lentivirus/veterinária , Doenças dos Ovinos/epidemiologia , Tuberculose/veterinária , Animais , Anticorpos Antibacterianos/análise , Vírus da Artrite-Encefalite Caprina/isolamento & purificação , Brucella/isolamento & purificação , Brucelose/epidemiologia , Brucelose/microbiologia , El Salvador/epidemiologia , Doenças das Cabras/microbiologia , Doenças das Cabras/virologia , Cabras , Infecções por Lentivirus/epidemiologia , Infecções por Lentivirus/virologia , Mycobacterium/isolamento & purificação , Prevalência , Estudos Soroepidemiológicos , Ovinos , Doenças dos Ovinos/microbiologia , Doenças dos Ovinos/virologia , Carneiro Doméstico , Tuberculose/epidemiologia , Tuberculose/microbiologiaRESUMO
This study was conducted in order to evaluate the transmission of caprine lentivirus to sheep using different experimental groups. The first one (colostrum group) was formed by nine lambs receiving colostrum from goats positive for small ruminant lentiviruses (SRLV). The second group (milk group) was established by nine lambs that received milk of these goats. Third was a control group, consisting of lambs that suckled colostrum and milk of negative mothers. Another experimental group (contact group) was formed by eight adult sheep, confined with two naturally infected goats. The groups were monitored by immunoblotting (IB), enzyme-linked immunosorbent assay (ELISA), agar gel immunodiffusion (AGID) and nested polymerase chain reaction (nPCR). All lambs that suckled colostrum and milk of infected goats and six sheep of the contact group had positive results in the nPCR, although seroconversion was detected only in three of the exposed animals, with no clinical lentiviruses manifestation, in 720 days of observation. There was a close relationship between viral sequences obtained from infected animals and the prototype CAEV-Cork. Thus, it was concluded that SRLV can be transmitted from goats to sheep, however, the degree of adaptation of the virus strain to the host species probably interferes with the infection persistence and seroconversion rate.
Assuntos
Vírus da Artrite-Encefalite Caprina/patogenicidade , Colostro/virologia , Doenças das Cabras/transmissão , Infecções por Lentivirus/transmissão , Doenças dos Ovinos/transmissão , Vírus Visna-Maedi/patogenicidade , Animais , Anticorpos Antivirais/sangue , Doenças das Cabras/virologia , Cabras/virologia , Interações Hospedeiro-Patógeno/fisiologia , Infecções por Lentivirus/virologia , Ruminantes/virologia , Soroconversão/fisiologia , Ovinos/virologia , Doenças dos Ovinos/virologiaRESUMO
This study was conducted in order to evaluate the transmission of caprine lentivirus to sheep using different experimental groups. The first one (colostrum group) was formed by nine lambs receiving colostrum from goats positive for small ruminant lentiviruses (SRLV). The second group (milk group) was established by nine lambs that received milk of these goats. Third was a control group, consisting of lambs that suckled colostrum and milk of negative mothers. Another experimental group (contact group) was formed by eight adult sheep, confined with two naturally infected goats. The groups were monitored by immunoblotting (IB), enzyme-linked immunosorbent assay (ELISA), agar gel immunodiffusion (AGID) and nested polymerase chain reaction (nPCR). All lambs that suckled colostrum and milk of infected goats and six sheep of the contact group had positive results in the nPCR, although seroconversion was detected only in three of the exposed animals, with no clinical lentiviruses manifestation, in 720 days of observation. There was a close relationship between viral sequences obtained from infected animals and the prototype CAEV-Cork. Thus, it was concluded that SRLV can be transmitted from goats to sheep, however, the degree of adaptation of the virus strain to the host species probably interferes with the infection persistence and seroconversion rate.
.Assuntos
Animais , Vírus da Artrite-Encefalite Caprina/patogenicidade , Colostro/virologia , Doenças das Cabras/transmissão , Infecções por Lentivirus/transmissão , Doenças dos Ovinos/transmissão , Vírus Visna-Maedi/patogenicidade , Anticorpos Antivirais/sangue , Doenças das Cabras/virologia , Cabras/virologia , Interações Hospedeiro-Patógeno/fisiologia , Infecções por Lentivirus/virologia , Ruminantes/virologia , Soroconversão/fisiologia , Doenças dos Ovinos/virologia , Ovinos/virologiaRESUMO
This study was conducted in order to evaluate the transmission of caprine lentivirus to sheep using different experimental groups. The first one (colostrum group) was formed by nine lambs receiving colostrum from goats positive for small ruminant lentiviruses (SRLV). The second group (milk group) was established by nine lambs that received milk of these goats. Third was a control group, consisting of lambs that suckled colostrum and milk of negative mothers. Another experimental group (contact group) was formed by eight adult sheep, confined with two naturally infected goats. The groups were monitored by immunoblotting (IB), enzyme-linked immunosorbent assay (ELISA), agar gel immunodiffusion (AGID) and nested polymerase chain reaction (nPCR). All lambs that suckled colostrum and milk of infected goats and six sheep of the contact group had positive results in the nPCR, although seroconversion was detected only in three of the exposed animals, with no clinical lentiviruses manifestation, in 720 days of observation. There was a close relationship between viral sequences obtained from infected animals and the prototype CAEV-Cork. Thus, it was concluded that SRLV can be transmitted from goats to sheep, however, the degree of adaptation of the virus strain to the host species probably interferes with the infection persistence and seroconversion rate..(AU)