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Infections by small ruminant lentiviruses (SRLVs) affect goats and sheep causing chronic multisystemic diseases that generate great economic losses. The caprine lentivirus (CLV) and the ovine lentivirus (OLV) present tropism for cells of the monocyte/macrophage lineage, which are directly associated with the main route of transmission through the ingestion of milk and colostrum from infected animals. In this manner, controlling this route is of paramount importance. Currently, researches have investigated the use of chemical additives in milk that can preserve colostrum or milk and inactivate microbiological agents. Among the compounds, sodium dodecyl sulfate (SDS) has been shown to be satisfactory in the chemical inactivation of HIV and CLV in milk, and also as a biocide in goat colostrum. [Emprego do dodecil sulfato de sódio como inativador viral e suas perspectivas no controle de lentivírus de pequenos ruminantes]. Resumo: As lentiviroses de pequenos ruminantes (LVPRs) são infecções que afetam caprinos e ovinos, causando doenças multissistêmicas crônicas, ocasionando grandes perdas econômicas. Os agentes causadores, lentivírus caprino (LVC) e o lentivírus ovino (LVO), apresentam tropismo por células da linhagem monocítico-fagocitária, as quais estão diretamente associadas à principal via de transmissão, por meio da ingestão de leite e colostro provindos de animais infectados. Desse modo, o controle por esta via é de suma importância. Atualmente, pesquisas vêm sendo desenvolvidas para o uso de aditivos químicos no leite, que possam conservar o colostro ou leite, e inativar agentes microbiológicos presentes. Dentre estes, o dodecil sulfato de sódio (SDS) vem apresentando resultados satisfatórios na inativação química do HIV e LVC em leite, e ainda como biocida em colostro caprino. Made available in DSpace on 2019-11-18T18:06:07Z (GMT). No. of bitstreams: 1 cnpc2019Sodium.pdf: 498207 bytes, checksum: 0395f8468452fc5d662d9c754b53f271 (MD5) Previous issue date: 2019 [ALICE ANDRIOLI PINHEIRO]

This study was conducted to evaluate caprine arthritis encephalitis virus (CAEV) transmission among sheep using 15 lambs that were distributed in 2 experimental groups. The exposed group consisted of 10 lambs that remained with their mothers, who were experimentally infected with CAEV. The non-exposed group was characterized as the control group and was comprised of 5 lambs that remained with their CAEV-negative mothers. Blood samples were collected monthly from birth until 1 year of life. To evaluate the transmission, an agar gel immunodiffusion test (AGID), enzyme immunoassay (ELISA), immunoblotting (IB), and nested polymerase chain reaction (nPCR) techniques were used. The non-exposed group was negative in all of the tests throughout the whole experiment. In the exposed group, 2 individuals had positive nPCR results. Positive nPCR samples were sequenced for comparison with the original goat strains and were shown to be similar to the CAEV-Cork strain. Seroconversion was not detected, and clinical manifestations were not observed. Thus, after 1 year of observation, it was verified that CAEV transmission among sheep is possible; however, with discreet frequency. This was an initial study, and other experiments are needed to analyze the adaptive capacity of the CAEV to remain in an infected sheep flock and cause the disease. [Transmissão do vírus da artrite encefalite caprina entre ovinos]. Resumo: O estudo foi conduzido para avaliar a transmissão do vírus da artrite encefalite caprina (CAEV) entre ovinos, utilizando 15 cordeiros, distribuídos em dois grupos experimentais. O grupo exposto foi constituído por 10 cordeiros, mantidos com suas mães, que foram infectadas, experimentalmente, com CAEV. O grupo não exposto caracterizou-se como grupo controle e foi formado por cinco cordeiros, mantidos com suas matrizes, negativas para CAEV. Foram colhidas amostras de sangue mensalmente, do periodo que compreende o nascimento até um ano de vida. Para avaliar a transmissão, foram utilizadas as técnicas de imunodifusão em gel de agarose (IDGA), ensaio imunoenzimático (ELISA), immunoblotting (IB) e reação em cadeia da polimerase do tipo nested (nPCR). O grupo não exposto se manteve negativo aos testes durante todo o experimento. Já no grupo exposto, dois indivíduos apresentaram resultados positivos na nPCR. As amostras positivas na nPCR foram sequenciadas para serem comparadas com as cepas originais de caprinos, comprovando se tratar de lentivírus semelhante à cepa CAEV-Cork. A soroconversão não foi detectada e a manifestação clínica não foi observada. Sendo assim, após um ano de observação, verificou-se que a transmissão do CAEV entre ovinos é possível, entretanto, com discreta frequência. Este foi um estudo inicial, e outros experimentos são necessários para analisar a capacidade adaptativa do CAEV de permanecer em rebanho ovino infectado e, com isso, causar doença. Made available in DSpace on 2018-09-21T00:46:25Z (GMT). No. of bitstreams: 1 CNPC2018Transmission.pdf: 1669152 bytes, checksum: 57ddeb25c242586e1b38691f197f4dae (MD5) Previous issue date: 2018-09-20

The objective of this study was to demonstrate iatrogenic transmission of small ruminant lentivirus (SRLV) from goats to sheep and horizontal transmission between sheep. The study was conducted on a farm with separate goat and sheep rearing, and animals were monitored for lentivirus occurrence by clinical examination and testing by immunoblotting (IB), agar gel immunodiffusion (AGID), and nested polymerase chain reaction (nPCR). Positive results had not been observed in the sheep flock until this study. Conversely, virus positive dairy goats were known. For this reason, the farm performed the caprine arthritis-encephalitis (CAE) control program. The study was designed with a sheep group that presented positive animals for SRLV by nPCR. It was verified that three newborn animals in this group were rejected by their mothers and consequently received milk from the goat herd. These three animals remained with another 20 sheep of the same age, totaling 23 animals. After one year, during monitoring, 11 of the 23 animals in the group presented positive results in the nPCR and three demonstrated seroconversion by IB. Of the animals that had received goat milk, two had positive results in the nPCR and IB. The 11 animals positive in the nPCR were followed and it was verified that five animals did not present further positive results in the nPCR, nor seroconversion; two continued presenting positive results in the nPCR but were negative in the IB and AGID and four were positive in the nPCR, IB, and AGID. Thus, it was possible to demonstrate iatrogenic interspecific infection and the occurrence of horizontal caprine lentivirus transmission among sheep. [Infecção de ovinos pelo lentivírus caprino]. Resumo: O objetivo foi demonstrar a transmissão iatrogênica de lentivírus de pequenos ruminantes (LVPR) de caprinos para ovinos e a transmissão horizontal entre ovinos. O estudo foi conduzido em propriedade com criação não consorciada de caprinos e ovinos, monitorada para ocorrência de lentiviroses, a partir de acompanhamento clínico e testes de immunoblotting (IB), imunodifusão em gel de agarose (IDGA) e reação em cadeia da polimerase do tipo nested (nPCR). Resultados positivos no rebanho ovino não haviam sido observados até então. Por outro lado, sabia-se que o rebanho caprino leiteiro era positivo. Por este motivo, a propriedade realizava programa de controle da artrite-encefalite caprina (CAE). O estudo foi delineado em um lote de ovinos que apresentou animais positivos para LVPR na nPCR. Verificou-se que três animais neonatos deste lote foram rejeitados por suas mães e por isso receberam leite do rebanho caprino. Estes três animais permaneceram com outros 20 ovinos de mesma faixa etária, totalizando 23 animais. Após um ano, durante a realização do monitoramento, dos 23 animais do lote, 11 apresentaram resultados positivos na nPCR e três demonstraram soroconversão por IB. Dos animais que haviam recebido leite do rebanho caprino, dois tiveram resultados positivos na nPCR e no IB. Os 11 animais positivos na nPCR foram acompanhados, verificando-se que cinco animais não apresentaram mais resultados positivos na nPCR, nem soroconverteram; dois continuaram apresentando resultados positivos na nPCR, mas negativos no IB e IDGA e quatro apresentaram-se positivos na nPCR, no IB e no IDGA. Dessa forma, foi possível demonstrar a infecção interespecífica de forma iatrogênica e a ocorrência de transmissão horizontal do lentivírus caprino entre ovinos. Made available in DSpace on 2018-10-06T00:41:36Z (GMT). No. of bitstreams: 1 CNPC2018Sheep.pdf: 169811 bytes, checksum: 20856e1b8518b49b8c3f85f3c43af5bc (MD5) Previous issue date: 2018-10-05

Ovine lentivirus (OvLV) is a macrophage-tropic lentivirus found in many countries that causes interstitial pneumonia, mastitis, arthritis and cachexia in sheep. There is no preventive vaccine and no cure, but breed differences suggest marker-assisted selective breeding might improve odds of infection and control of OvLV post-infection. Although variants in TMEM154 have consistent association with odds of infection, no variant in any gene has been associated with host control of OvLV post-infection in multiple animal sets. Proviral concentration is a live-animal diagnostic measure of OvLV control post-infection related to severity of OvLV-induced lesions. A recent genome-wide association study identified a region including four zinc finger genes associated with proviral concentration in one Rambouillet flock. To refine this region, we tested additional variants and identified a small insertion/deletion variant near ZNF389 that showed consistent association with proviral concentration in three animal sets (P 87%, including yearlings) and mean proviral concentration (>950 copies/μg), possibly due to needle sharing. The best estimate of proviral concentration by genotype, obtained from all 1310 OvLV-positive animals tested, showed insertion homozygotes had less than half the proviral concentration of other genotypes (P

Maedi-visna virus (MVV) is an ovine lentivirus that is widespread in many countries worldwide. Both clinical and subclinical MVV infections cause substantial economic losses. MVV infection in live sheep is usually diagnosed serologically, with antibody-positive sheep being regarded as infected. There have been few reports of maedi-visna in China, with no detailed epidemic analysis of MVV infection in ovine herds. In order to investigate the seroprevalence and spatial distribution of maedi-visna among ovine flocks in China, a total of 672 serum samples were collected from different ovine flocks in 12 regions (provinces, autonomous regions or municipalities) of China in 2011, and serum antibody levels were determined using a commercial ELISA Kit. This study represents the first investigation of the seroepidemiology of maedi-visna in China, indicating a circulation of MMV among sheep.

Ovine lentivirus (OvLV) also known as maedi-visna virus, infects and replicates primarily in macrophages. This investigation examined the role of nitric oxide in the replication of OvLV in cultured macrophages. Peripheral blood mononuclear cells were collected from OvLV-free sheep and cultured in Teflon coated flasks at a high concentration of lamb serum. The cells were subsequently infected with OvLV strain 85/34. OvLV replication was assessed under different experimental treatments by comparison of reverse transcriptase (RT) activity in culture supernatant. Cultures that were treated with exogenous nitric oxide via S-nitroso-acetylpenicillamine did not have altered levels of RT activity compared to cultures treated with the inactive control compound, acetylpenicillamine. However, blockage of nitric oxide production by treatment with aminoguanidine, a competitive inhibitor of inducible nitric oxide synthase (iNOS), led to a significant rise in RT activity. This rise in RT activity was partially reversed in aminoguanidine treated cultures by L-arginine, the normal substrate for iNOS. Finally, the number of viral antigen producing cells was also quantified after aminoguanidine treatment and found to be significantly higher than untreated cultures. Collectively, these results indicate that nitric oxide is a negative regulator of OvLV replication in macrophages.

Small ruminant lentivirus (SRLV), also called ovine progressive pneumonia virus or maedi-visna, is present in 24% of US sheep. Like human immunodeficiency virus, SRLV is a macrophage-tropic lentivirus that causes lifelong infection. The production impacts from SRLV are due to a range of disease symptoms, including pneumonia, arthritis, mastitis, body condition wasting and encephalitis. There is no cure and no effective vaccine for preventing SRLV infection. However, breed differences in prevalence and proviral concentration indicate a genetic basis for susceptibility to SRLV. Animals with high blood proviral concentration show increased tissue lesion severity, so proviral concentration represents a live animal test for control post-infection in terms of proviral replication and disease severity. Recently, it was found that sheep with two copies of TMEM154 haplotype 1 (encoding lysine at position 35) had lower odds of SRLV infection. In this study, we examined the relationship between SRLV control post-infection and variants in two genes, TMEM154 and CCR5, in four flocks containing 1403 SRLV-positive sheep. We found two copies of TMEM154 haplotype 1 were associated with lower SRLV proviral concentration in one flock (P < 0.02). This identified the same favorable diplotype for SRLV control post-infection as for odds of infection. However, frequencies of haplotypes 2 and 3 were too low in the other three flocks to test. The CCR5 promoter deletion did not have consistent association with SRLV proviral concentration. Future work in flocks with more balanced allele frequencies is needed to confirm or refute TMEM154 association with control of SRLV post-infection.

A maedi-visna virus CA-TM fusion protein ELISA (MVV ELISA) was evaluated for the detection of antibody in sheep infected with North American ovine lentivirus (OvLV). The results of the MVV ELISA were compared with other assays for OvLV antibody and with viral infection in an intensively studied group of 38 sheep with a high prevalence of OvLV infection and disease. The sensitivity, specificity, and concordance of assays for OvLV antibody (MVV ELISA, indirect ELISA, Western blot, and AGID), virus (virus isolation, PCR, antigen ELISA), and OvLV-induced disease in each animal were compared with OvLV infection status as defined by a positive result in two or more of the assays. Five sheep met the criteria for absence of OvLV infection. The sensitivity of the MVV ELISA in detecting OvLV infected sheep was 64%, whereas the sensitivity of the other three tests for antibody ranged from 85 to 94%. All the antibody assays were 100% specific in this group of animals. Of the assays for virus, the PCR test had the highest sensitivity and the best concordance with OvLV infection, but it also had the lowest specificity of any of the virus or antibody assays. Among the antibody tests, the concordance of the MVV ELISA compared most favorably with the AGID test for detecting OvLV-infected sheep. Analysis of serum samples from 28 lambs experimentally-infected with one of three North American strains of OvLV suggested that there were no significant strain differences detectable by antibody assay. Twenty virus-inoculated lambs were positive by both the MVV ELISA and the AGID test, five lambs were MVV ELISA negative and AGID test positive, and three lambs were MVV ELISA positive and AGID test negative. No pre-inoculation samples were positive by either assay. In a longitudinal study involving seven lambs, antibodies to OvLV were detected by AGID 3–5 weeks post-inoculation, but were not detected by MVV ELISA until 5–10 weeks post-inoculation. Among 128 naturally and experimentally-infected sheep that were seropositive in the AGID test, the overall sensitivity of the MVV ELISA was higher in the naturally infected sheep (84%) than in the experimentally infected sheep (69%). The data indicated that the MVV ELISA represents a less sensitive, but specific alternative for the detection of OvLV antibodies.

We used a previously described sensitive and specific ovine lentivirus (OLV) recombinant transmembrane (rTM) protein enzyme-linked immunosorbent assay (ELISA) to detect anti-OLV antibodies and define OLV infection in breeding ewes from nine US Meat Animal Research Center (MARC) flocks. We estimated the production impacts of dam rTM ELISA seropositivity on ewe and lamb productivity in the birth-to-weaning interval using production data from 1466 breeding ewes (of which 1242 actually lambed) and their 2452 lambs born in spring 1992 using several multiple linear and logistic regression models. By adjusting for lamb weaning age, gender, type of birth and rearing, birth difficulty, dam age, and flock, the component of ewe or lamb productivity related to ewe OLV infection alone was isolated. The rTM ELISA-negative ewes produced significantly more total weight of weaned lamb per ewe-lambing (3.84 kg) and per ewe ram-exposed (4.95 kg) compared to their OLV-positive flockmates. Negative ewes also weaned 0.11 more lambs per ewe-lambing and 0.09 more lambs per ewe ram-exposed, gave birth to 0.13 more lambs per ewe ram-exposed, and were more likely to lamb after breeding (odds ratio (OR) = 1.9) compared to equivalent OLV-positive ewes. Lambs reared by OLV-negative ewes weighed 0.15 kg more at birth, gained 8 g more per day through weaning, and weighed 0.59 kg more at 56-day weaning. Preweaning mortality was lower (OR = 0.8) among lambs born to OLV-negative compared to OLV-positive ewes, although this difference was not significant. Our results suggest that subclinical OLV infection has important detrimental effects on sheep production which occur in cumulative fashion from breeding through weaning and that OLV control efforts may be financially justified in some sheep flocks.

Although ovine lentiviruses have been described in the United States since the early part of this century, North American strains of sheep lentiviruses remain relatively uncharacterized at the molecular level. The LTR of a North American ovine lentivirus, OLV-CU1, was found to be closely related at the molecular and functional levels to visna virus, the Icelandic ovine lentivirus. Sequence analysis of the LTR revealed high identity to other ovine and caprine lentiviruses in key regulatory elements of the upstream promoter region (-25 to -115). However, the R region of the LTR was much less homologous. Transcriptional control of OLV-CU1 in transient transcriptional assays required a conserved putative AP-4 region and possibly an AP-1 like element in the upstream promoter region for moderate to high levels of transcription, much like visna virus. In contrast to visna virus, the down-stream region beyond the transcriptional start site was required for virus-specific transactivation.

Recombinant major core protein p25 (rP25) and transmembrane protein gp40 (rTM) of the ovine lentivirus (OLV) were used as immobilized antigens in enzyme-linked immunosorbent assays (ELISAs) for the detection of antibodies against caprine arthritis-encephalitis virus (CAEV). The sensitivity and specificity of these assays were compared with an agar gel immunodiffusion (AGID) test and a whole-virus ELISA. The results showed that the rTM ELISA was more effective than rP25 ELISA or AGID test in identifying CAEV antibodies in goat populations. The rTM ELISA had similar sensitivity and specificity as the whole-virus ELISA, with an overall concordance of 87.5%. When data for rP25 and rTM were combined, the overall test agreement between whole-virus ELISA and combined recombinant ELISAs increased to 89.3%. The high quantity and purity characteristic of recombinant proteins should make them suitable as routine diagnostic antigens for CAEV and OLV serology.

The effect of the indurative lymphocytic mastitis caused by infection with maedi-visna virus was quantified by comparing the pre-weaning growth of lambs from infected and uninfected ewes under the same conditions. A total of 73 infected, but clinically healthy, ewes and 75 ewes from a maedi-visna virus-free source were purchased to form a new flock; they were all three years old. The ewes were mated and the flock was managed as a normal field flock. Serum samples were taken at regular intervals and tested for antibodies to maedi-visna virus. The lambs were weighed at birth and at 14, 30, 50 and, finally, 80 days old, when they were weaned. The ewes were slaughtered, their udders were examined histologically, and the lesions were assessed by counting the typical lymphocytic follicles. Sixty-six per cent of the ewes that were seropositive at slaughter appeared to have follicles. A statistically significant association was found between the number of follicles in the udder and the reduction in the growth rate of the lambs. Lambs from ewes with the mean number of follicles weighed 1.7 kg less at weaning.

The antiviral effects of recombinant ovine interferon-τ (roIFN-τ) were studied in 26 lambs inoculated with ovine lentivirus (OvLV) or mock-infected. Six of the OvLV-infected lambs and three of the mock-infected lambs were treated with 106 antiviral units (AVU) per kg roIFN-τ daily for 30 days starting at day 0 post-inoculation (p.i.) and twice a week thereafter (early treatment). Six of the OvLV-infected lambs and three of the mock-infected lambs were treated with 106 AVU/kg roIFN-τ daily for 30 days starting at day 150 p.i. and twice a week thereafter (late treatment). Six OvLV-infected and two mock-infected lambs were treated either early or late with placebo. Cell-associated viraemia was quantified by an end-point dilution method. The weekly antibody response against OvLV proteins was studied by ELISA. All experimental animals were killed at 27 weeks p.i. and histological sections of lung were scored for the degree of lymphoid interstitial pneumonia (LIP). A 90% reduction in OvLV titres was detected at 4 weeks post-treatment in lambs that received early roIFN-τ treatment (PPP in vivo and reduces the likelihood of development of lentivirus-induced LIP when infected lambs are treated during the initial phases of OvLV infection.

Maedi Visna Virus (MVV) is a prototypic lentivirus that causes infection only in cells of macrophage lineage, unlike the primate lentiviruses which infect both CD4+ T lymphocytes and macrophages. In primates, the earliest viral invasion is associated with the ability of the virus to infect and activate T cells which convey virus to the brain. Infected monocytes in blood rarely cause CNS infection in absence of activation of CD4+ T cells. In the face of lack of infection or activation of T cells by MVV in sheep, the question arises, how does MVV gain access to the brain to cause the classical lesions of visna? In previous studies on experimental induction of visna, sheep were inoculated with virus directly in the brain. In this study, we asked whether neuroinvasion by MVV would occur if sheep were inoculated with virus in a non-neural site. Nine sheep were inoculated intratracheally and all developed systemic infection when examined 3 weeks later. At this time, five were injected intramuscularly with brain white matter homogenized in Freund's complete adjuvant to induce EAE. None of the four animals inoculated with virus alone developed CNS infection despite typical lentiviral infection in lungs, lymphoid tissues and blood-borne mononuclear cells. In contrast, all five of the sheep injected with brain homogenate developed infection in the brain. Virus was produced by macrophages associated with the EAE lesions. This study illustrated that both activated T cells specific for antigen in the CNS and infected macrophages are essential for lentivirus neuropathogenesis.

In order to localize the immunodominant regions, 12 ovine lentivirus (OLV) gag-coding gene fragments were cloned and expressed in Escherichia coli and when tested in a Western blot (WB) assay against a panel of sera collected from US and Italian OLV-infected sheep. The most immunoreactive regions were mapped to the amino-terminal of p25 and carboxyl-terminal of p14. In addition, we found that the reactivity pattern between US and Italian sheep was very similar, suggesting the antigenic domain between US and Italian isolates in the gag gene structures could be conserved. Given the broad immunoreactivity of the amino-terminal of p25, this region could serve as an ideal diagnostic antigen for the serological identification of OLV-infected sheep.

This chapter covers economic importance, epidemiology and possibilities for the control of maedi-visna and related ovine lentivirus infections, following an historical introduction in conjunction with an outline of their geographic distribution.

Summary Enzyme-linked immunosorbent assay (ELISA) and agar gel immunodiffusion (AGID) are the most widely used serological tests for Maedi—Visna diagnostics. The purpose of the present study was to develop an indirect whole virus ELISA and an immunodiffusion test and compare their sensitivity. A total of 747 ovine serum specimens were analysed for antibodies against this ovine lentivirus. The number of positive results in the ELISA was 430 (57.56%). In the AGID test, a positive result was found in 380 samples (50.87%). In the group of discordant results 78 (10.4%) samples tested positive by the ELISA and negative by the AGID test and 28 sera (3.7%) were found to be positive by the AGID test and negative by the ELISA. The data in this report show the ELISA to be more sensitive than the AGID test, but accurate serological diagnostics should be based on a combination of the two tests.