Small Ruminant Lentivirus Infection Research Articles

Influence of true within-herd prevalence of small ruminant lentivirus infection in goats on agreement between serological immunoenzymatic tests

The study was conducted to evaluate influence of the true within-herd prevalence of small ruminant lentivirus (SRLV) infection on agreement beyond chance between three different types of commercial serological ELISAs. Blood samples were collected from 865 goats from 12 dairy goat herds. Serum samples were tested using three commercial ELISA kits: whole-virus indirect ELISA (wELISA), indirect ELISA based on recombined TM and CA antigens (TM/CA-ELISA), and competitive-inhibition ELISA based on SU antigen (SU-ELISA). Herds were classed into three prevalence strata of high (>50%), moderate (10–50%) and low (<10%) true within-herd prevalence of SRLV infection. The latter was estimated on the basis of results of wELISA adjusted by its sensitivity and specificity. Agreement beyond chance between the three ELISAs was assessed at two levels. First, the general agreement was determined using two coefficients corrected for chance-agreement: Cohen’s kappa and Gwet’s AC1. Then, agreement between tests was evaluated using Gwet’s AC1 separately in the three prevalence strata and compared between them by computing 95% confidence intervals for differences with a Bonferroni correction for multiple comparisons. The general agreement between the three tests was very good: wELISA and TM/CA-ELISA – Cohen’s kappa of 81.8% (CI 95%: 77.9% to 85.7%), Gwet’s AC1 of 82.7% (CI 95%: 79.0% to 86.4%); wELISA and SU-ELISA – Cohen’s kappa of 83.2% (CI 95%: 79.4% to 86.9%), Gwet’s AC1 of 83.9% (CI 95%: 80.4% to 87.5%); TM/CA-ELISA and SU-ELISA – Cohen’s kappa of 86.0% (CI 95%: 82.6% to 89.5%), Gwet’s AC1 of 86.9% (CI 95%: 83.6% to 90.1%). However, agreement between ELISAs was significantly related to the within-herd true prevalence – it was significantly lower (although still high) when within-herd true prevalence was moderate (Gwet’s AC1 between 67.2% and 78.7%), whereas remained very high, when true within-herd prevalence was either >50% (Gwet’s AC1 between 91.9% and 98.8%) or <10% (Gwet’s AC1 between 94.7% and 98.4%). Concluding, the three different commercial ELISAs for SRLV infection in goats available on the market yield highly consistent results. However, their agreement is affected by the true within-herd prevalence in a tested population, and the worse (although still high) agreement should be expected, when the percentage of infected goats is moderate.

Small Ruminant Lentiviruses in Sheep: Pathology and Tropism of 2 Strains Using the Bone Marrow Route.

The objective of this work was to comparatively study the tissue tropism and the associated pathology of 2 autochthonous small ruminant lentivirus (SRLV) field strains using an experimental infection in sheep through the bone marrow. Fifteen male, SRLV-free lambs of the Rasa Aragonesa breed were inoculated with strain 697 (nervous tissue origin, animals A1-A6), with strain 496 (articular origin, animals B1-B6), or with uninfected culture medium (C1-C3). Clinical, serologic, and polymerase chain reaction (PCR) evaluations were performed periodically. Two lambs from each infected group and a control animal were euthanized at 134, 273, and 319 days postinfection. Tissues were analyzed by gross and histopathologic evaluation; immunohistochemistry for CD3, CD4, CD8, CD68, and FoxP3 cell markers; lung morphometric evaluation; and tissue proviral quantification by PCR. All infected animals became positive either by enzyme-linked immunosorbent assay and/or PCR, with group B lambs showing the highest serologic values and more consistently positive PCR reactions. Group A lambs showed representative lung lesions but only mild histopathologic changes in the central nervous system (CNS) or in carpal joints. Contrarily, group B lambs demonstrated intense carpal arthritis and interstitial pneumonia but an absence of lesions in the CNS. Proviral copies in tissues were detected only in group B lambs. Experimental infection with these SRLV strains indicates that strain 496 is more virulent than strain 697 and more prone to induce arthritis, whereas strain 697 is more likely to reproduce encephalitis in Rasa Aragonesa lambs. Host factors as well as viral factors are responsible for the final clinicopathologic picture during SRLV infections.

Goat umbilical cord cells are permissive to small ruminant lentivirus infection in vitro.

Small ruminant lentiviruses isolated from peripheral blood leukocytes and target organs can be propagated in vitro in fibroblasts derived from goat synovial membrane cells. These cells are obtained from tissues collected from embryos or fetuses and are necessary for the establishment of the fibroblast primary culture. A new alternative type of host cells, derived from goat umbilical cord, was isolated and characterized phenotypically with its main purpose being to obtain cell monolayers that could be used for the diagnosis and isolation of small ruminant lentiviruses in cell culture. To accomplish this goal, cells were isolated from umbilical cords; characterized phenotypically by flow cytometry analysis; differentiate into osteogenic, chondrogenic and adipogenic lineage; and submitted to viral challenge. The proliferation of goat umbilical cord cells was fast and cell monolayers formed after 15 days. These cells exhibited morphology, immunophenotype, growth characteristics, and lineage differentiation potential similar to mesenchymal stem cells of other origins. The goat umbilical cord derived cells stained positive for vimentin and CD90, but negative for cytokeratin, CD34 and CD105 markers. Syncytia and cell lysis were observed in cell monolayers infected by CAEV-Cork and MVV-K1514, showing that the cells are permissive to small ruminant lentivirus infection in vitro. These data demonstrate the proliferative competence of cells derived from goat umbilical cords and provide a sound basis for future research to standardize this cell lineage.

Impaired Expression of Cytokines as a Result of Viral Infections with an Emphasis on Small Ruminant Lentivirus Infection in Goats.

Knowing about the genes involved in immunity, and being able to identify the factors influencing their expressions, helps in gaining awareness of the immune processes. The qPCR method is a useful gene expression analysis tool, but studies on immune system genes are still limited, especially on the caprine immune system. Caprine arthritis encephalitis, a disease caused by small ruminant lentivirus (SRLV), causes economic losses in goat breeding, and there is no therapy against SRLV. The results of studies on vaccines against other viruses are promising. Moreover, the Marker-Assisted Selection strategy against SRLV is possible, as has been shown in sheep breeding. However, there are still many gaps in our knowledge on the caprine immune response to infection. All types of cytokines play pivotal roles in immunity, and SRLV infection influences the expression of many cytokines in different types of cells. This information encouraged the authors to examine the results of studies conducted on SRLV and other viral infections, with an emphasis on the expression of cytokine genes. This review attempts to summarize the results of studies on the expression of cytokines in the context of the SRLV infection.

The presence of small ruminant lentiviruses in Mexican Pelibuey sheep

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.

Risk factors associated with small ruminant lentivirus infection in eastern Poland sheep flocks

An analysis of the risk factors for ovine lentivirus infection was performed in sheep flocks located throughout the central-eastern region of Poland. Here, we report the infection details for 98 flocks with a total of 6470 ewes, 15 sheep breeds. The identification of infected animals and a review of the epidemiological status of each flock were based on an evaluation of serological tests performed on collected blood serum samples. Blood for examination was obtained from 2925 ewes of the 98 flocks under observation. Specific antibodies for Maedi Visna Virus (MVV) were detected via ELISA. Data illustrating the conditions at each sheep farm were obtained through questionnaires completed by farmers, as well as observations, measurements, and breeding records that were available. These observations were used to assess risk factors contributing to small ruminant lentivirus (SRLV) infection in sheep flocks. It was found that both sheep flock size and the type of management system had a significant effect on the increased risk of lentiviral infection. In addition, we demonstrate that there is a significant (p<0.0001) relationship between the occurrence of mastitis (OR 2.01, CI: 1.55-2.61) and diarrhea (OR 4.22, CI: 3.30-5.39) with SRLV infection in the observed sheep. Additionally, the infection rate of the animals translated directly to an impaired physical condition. Notably, the risk of infection could potentially be reduced if sheep producers are further acquainted with SRLV detection and invoke a control program based on diagnostic tests. Moreover, marketing approval should be granted for solely SRLV-seronegative animals.

Post-entry blockade of small ruminant lentiviruses by wild ruminants.

Small ruminant lentivirus (SRLV) infection causes losses in the small ruminant industry due to reduced animal production and increased replacement rates. Infection of wild ruminants in close contact with infected domestic animals has been proposed to play a role in SRLV epidemiology, but studies are limited and mostly involve hybrids between wild and domestic animals. In this study, SRLV seropositive red deer, roe deer and mouflon were detected through modified ELISA tests, but virus was not successfully amplified using a set of different PCRs. Apparent restriction of SRLV infection in cervids was not related to the presence of neutralizing antibodies. In vitro cultured skin fibroblastic cells from red deer and fallow deer were permissive to the SRLV entry and integration, but produced low quantities of virus. SRLV got rapidly adapted in vitro to blood-derived macrophages and skin fibroblastic cells from red deer but not from fallow deer. Thus, although direct detection of virus was not successfully achieved in vivo, these findings show the potential susceptibility of wild ruminants to SRLV infection in the case of red deer and, on the other hand, an in vivo SRLV restriction in fallow deer. Altogether these results may highlight the importance of surveilling and controlling SRLV infection in domestic as well as in wild ruminants sharing pasture areas, and may provide new natural tools to control SRLV spread in sheep and goats.

Small ruminant lentivirus infections and diseases

Small ruminant lentiviruses include viruses with diverse genotypes that frequently cross the species barrier between sheep and goats and that display a great genetic variability. These characteristics stress the need to consider the whole host range and to perform local surveillance of the viruses to opt for optimum diagnostic tests, in order to establish control programes. In the absence of effective vaccines, a comprehensive knowledge of the epidemiology of these infections is of major importance to limit their spread. This article intends to cover these aspects and to summarise information related to characteristics of the viruses, pathogenesis of the infection and description of the various syndromes produced, as well as the diagnostic tools available, the mechanisms involved in transmission of the pathogens and, finally, the control strategies that have been designed until now, with remarks on the drawbacks and the advantages of each one. We conclude that there are many variables influencing the expected cost and benefits of control programs that must be evaluated, in order to put into practice measures that might lead to control of these infections.

Diagnosing infection with small ruminant lentiviruses of genotypes A and B by combining synthetic peptides in ELISA

The major challenges in diagnosing small ruminant lentivirus (SRLV) infection include early detection and genotyping of strains of epidemiological interest. A longitudinal study was carried out in Rasa Aragonesa sheep experimentally infected with viral strains of genotypes A or B from Spanish neurological and arthritic SRLV outbreaks, respectively. Sera were tested with two commercial ELISAs, three based on specific peptides and a novel combined peptide ELISA. Three different PCR assays were used to further assess infection status.The kinetics of anti-viral antibody responses were variable, with early diagnosis dependent on the type of ELISA used. Peptide epitopes of SRLV genotypes A and B combined in the same ELISA well enhanced the overall detection rate, whereas single peptides were useful for genotyping the infecting strain (A vs. B). The results of the study suggest that a combined peptide ELISA can be used for serological diagnosis of SRLV infection, with single peptide ELISAs useful for subsequent serotyping.

Genetic subgroup of small ruminant lentiviruses that infects sheep homozygous for TMEM154 frameshift deletion mutation A4Δ53.

Small ruminant lentivirus (SRLV) infections of sheep are influenced by genetics on both the host and pathogen sides. Genetic variation in the ovine transmembrane 154 (TMEM154) gene associates with infection susceptibility, and distinct SRLV genetic subgroups infect sheep in association with their TMEM154 diplotypes. In this study, a novel SRLV subgroup was identified that naturally infected sheep with various TMEM154 diplotypes, including those homozygous for a rare frameshift mutation (A4 delta53), which is predicted to abolish TMEM154 protein function. Thus, these SRLVs may infect sheep that lack functional TMEM154, and may not be restricted by TMEM154 diplotypes in establishing infections.Electronic supplementary materialThe online version of this article (doi:10.1186/s13567-015-0162-7) contains supplementary material, which is available to authorized users.

Seropositive bucks and within-herd prevalence of small ruminant lentivirus infection.

Caprine arthritis-encephalitis is an economically important disease of goats. It is evident that horizontal transmission through respiratory secretions and milk plays an important part in the disease spread whereas the role of sexual transmission remains questionable. The cross-sectional study was carried out to investigate the relationship between presence of small ruminant lentivirus (SRL V)-seropositive bucks and seroprevalence of SRL V infection in does in herds. The analysis included 76 goat herds seropositive for SRL V infection. A sample of adult female goats from each herd was selected in a simple random fashion. All males present in a herd were also enrolled in the study. The animals were screened with commercial serological immunoenzymatic tests. Standardized questionnaires were used to gather knowledge of 3 hypothesized herd-level confounding factors: number of years for which a herd had existed until testing, goat replacement from other herds in Poland and use of machine milking. Three-level hierarchical linear regression model was developed to evaluate the relationship (α = 0.05). Median (interquartile range) within-herd seroprevalence of SRL V was 60.1% (35.7% to 87.9%) and 35.8% (10.1% to 49.6%) in herds where seropositive males were present and absent, respectively. Controlling for possible confounders presence of SRL V-seropositive bucks proved to be an independent factor linked to the higher within-herd seroprevalence of SRL V (p = 0.001). The study indicates that seropositive bucks may facilitate the spread of SRL V infection in goat herds and therefore their presence should be considered as a risk factor.

Influence of small ruminant lentivirus infection on cheese yield in goats

Three-year cohort study was carried out to investigate the influence of small ruminant lentivirus (SRLV) infection on cheese yield in goats. For this purpose records of milk yield, milk composition and cheese yield were collected in a dairy goat herd. Cheese yield was recorded as the amount of fresh cheese obtained from 1 kg milk. All goats were serologically tested for SRLV infection twice a year. The analysis included 247 records in total (71 for seropositive and 176 from seronegative individuals) and was carried out with the use of the four-level hierarchical linear model (α = 0·05). SRLV infection proved to be a statistically significant independent factor reducing cheese yield (P = 0·013)--when other covariates were held constant cheese yield was reduced by 4·6 g per each 1 kg milk in an infected goat compared with an uninfected goat. Other statistically significant covariates positively associated with cheese yield were protein contents, fat contents and the 3rd stage of lactation (P < 0·001 for all).

Comparison of primer pairs: Greater degeneracy improves small ruminant lentivirus (SRLVs) detection by seminested PCR

The identification of small ruminant lentivirus (SRLV) infected animals is crucial to obtain success through eradication, control and monitoring programs worldwide. Several authors have demonstrated that serology alone is not sufficient to detect SRLV infection. Polymerase chain reaction (PCR) is suggested as the choice test to identify seronegative infected animals. The aim of this study was to evaluate and compare degenerate primer pairs in order to detect a great number of virus variants by a seminested PCR. Based on already described primers and the alignment of gag nucleotide sequences from databases, degenerate primers were designed. Increasing the degree of degeneracy has improved the detection of the provirus in ovine and caprine genomic DNA samples. The number of positive PCR samples was twice higher when using the more degenerate primers in the established experimental conditions. Thus, we propose the use of this seminested PCR as a diagnostic tool, in combination with serology, especially in the situations where the circulating virus variants are unknown.

Diagnostic performance of ID screen MVV-CAEV Indirect Screening ELISA in identifying small ruminant lentiviruses-infected goats.

Diagnostic performance of ID Screen MVV-CAEV Indirect Screening ELISA in identifying goats infected with small ruminant lentiviruses (SRLV) was evaluated. In total 299 serum samples from the collection of the Laboratory of Veterinary Epidemiology and Economics--109 truly positive and 190 truly negative--were used. To be enrolled in the study a serum sample had to come from at least 2 year-old goat which had reacted identically in two serological surveys preceding sample collection and was kept in a herd of stable serological status confirmed at least twice during preceding 5 years. Moreover, in seropositive herds at least 20% of goats had to be serologically positive at the moment when the serum sample was collected for the study. The test proved to have high accuracy. Area under curve was 98.8% (95% CI: 97.5%, 100%). Diagnostic performance of the test was almost identical (Youlden's index of 90%, sensitivity > 90% and specificity > 95%) within a fairly wide range of cut-off values--between 20% and 60%. At manufacturer's cut-off of 50% sensitivity and specificity were 91.7% (95% CI: 85.0%, 95.6%) and 98.9% (95% CI: 96.2%, 99.7%), respectively. For this cut-off positive likelihood ratio was 87 (95% CI: 22, 346) and negative likelihood ratio was 0.08 (95% CI: 0.04, 0.16). In conclusion, the results of this study indicate that ID Screen MVV-CAEV Indirect Screening ELISA is a highly accurate diagnostic test for SRLV infection.

Mutations in Ovis aries TMEM154 are associated with lower small ruminant lentivirus proviral concentration in one sheep flock.

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.

Investigation of risk factors associated with infections caused by small ruminant lentiviruses

Abstract An epidemiological study was conducted to identify risk factors related to small ruminant lentivirus (SRLV) infection in the central region of Spain. Between October 1998 and October 2000, a total of 194 sheep from 10 flocks and 163 goats from three flocks were tested for SRLV antibodies, resulting in 65.5% and 8.0% of seroprevalence, respectively. The relationship between differences in prevalence of SRLV, geographical location of the flock, and possible factors related to the flock that could enhance transmission were studied. Results of multivariable analysis showed an association between SRLV infection and geographical location of the flock and the rearing system. In addition, the differences in the productivity between infected and non-infected animals were explored. The productivity parameters were measured in 62 sheep and 28 goats. All productivity parameters studied (milk production, number of milking days, and lambing rate) appeared to be reduced in the SRLV-seropositive group in both goats and sheep. Even though, these differences were not statistically significant, it seems that animals infected are less productive than these non-infected. Statistical analyses comparing infected and non-infected sheep showed no statistical relationship between SRLV infection and milk quality.

Development, validation and evaluation of added diagnostic value of a q(RT)-PCR for the detection of genotype A strains of small ruminant lentiviruses

Small ruminant lentiviruses (SRLV) infect sheep and goats. Diagnosis of SRLV infection mostly relies on serological testing but more recently, also PCR is regarded as a useful complementary tool in SRLV diagnosis. The goal of this study was to develop and validate a quantitative PCR capable to detect a broad range of SRLV strains from genotype A, including strains circulating in Belgium.The developed q(RT)-PCR targets a region of the gag gene and showed to be highly sensitive and specific with a limit of detection of 6 DNA and 40 RNA copies/reaction respectively. SRLV sequences could be detected in lung samples and leukocytes pellets. The q(RT)-PCR identified SRLV positive animals in Belgian sheep flocks, but also SRLV isolates and samples from Scotland, The Netherlands, Spain, Portugal, UK, Iceland, Finland and USA were found positive. Samples known to contain ‘CAEV like’ SRLV from France and Spain were not identified as positive. Combined serological and PCR analysis of a limited number (n=35) of Belgian sheep underlined the usefulness of the described PCR as a complementary diagnostic tool since 3 seronegative animals were found positive by the PCR.In conclusion, the validated q(RT)-PCR shows excellent analytical characteristics and is capable to detect SRLV strains belonging to genotype A from various countries.

Retroviral Infections in Sheep and Goats: Small Ruminant Lentiviruses and Host Interaction

Small ruminant lentiviruses (SRLV) are members of the Retrovirus family comprising the closely related Visna/Maedi Virus (VMV) and the Caprine Arthritis-Encephalitis Virus (CAEV), which infect sheep and goats. Both infect cells of the monocyte/macrophage lineage and cause lifelong infections. Infection by VMV and CAEV can lead to Visna/Maedi (VM) and Caprine Arthritis-Encephalitis (CAE) respectively, slow progressive inflammatory diseases primarily affecting the lungs, nervous system, joints and mammary glands. VM and CAE are distributed worldwide and develop over a period of months or years, always leading to the death of the host, with the consequent economic and welfare implications. Currently, the control of VM and CAE relies on the control of transmission and culling of infected animals. However, there is evidence that host genetics play an important role in determining Susceptibility/Resistance to SRLV infection and disease progression, but little work has been performed in small ruminants. More research is necessary to understand the host-SRLV interaction.

Immunization against Small Ruminant Lentiviruses

Multisystemic disease caused by Small Ruminant Lentiviruses (SRLV) in sheep and goats leads to production losses, to the detriment of animal health and welfare. This, together with the lack of treatments, has triggered interest in exploring different strategies of immunization to control the widely spread SRLV infection and, also, to provide a useful model for HIV vaccines. These strategies involve inactivated whole virus, subunit vaccines, DNA encoding viral proteins in the presence or absence of plasmids encoding immunological adjuvants and naturally or artificially attenuated viruses. In this review, we revisit, comprehensively, the immunization strategies against SRLV and analyze this double edged tool individually, as it may contribute to either controlling or enhancing virus replication and/or disease.

Toll Like Receptor 9 (TLR9) Polymorphism G520R in Sheep Is Associated with Seropositivity for Small Ruminant Lentivirus

Infectious diseases of sheep are of major economic importance causing direct and indirect losses. Among the major sheep infectious agents are Small Ruminant Lentivirus, Chlamydophila abortus and Mycobacterium avium subsp. paratuberculosis infections, mainly due to their worldwide distribution and economic impact that they cause. Based on the differential susceptibility to infectious diseases between and within breeds and on the recent findings regarding the putative involvement of TLR9 in disease susceptibility, the aim of this study was to evaluate the levels of nucleotide variation of TLR9 and its mediator MyD88 in three sheep flocks originated from different breeds and assess their possible association with seropositivity/seronegativity for different infectious agents. The analysis indicated that the change of G to R at codon 520 of TLR9 polypeptide shows a significant association with Small Ruminant Lentivirus seropositivity. This amino-acid substitution, which can result in polarity change, might influence structure and function of LRR17, interfering with ligand binding and thus could be used in studies investigating susceptibility/resistance to Small Ruminant Lentivirus infections in sheep.