Characterisation of non-degraded oligosaccharides in enzymatically hydrolysed and fermented, dilute ammonia-pretreated corn stover for ethanol production.

BACKGROUND: Corn stover is lignocellulosic biomass that has potential to be used as raw material for bioethanol production. In the current research, dilute ammonia pretreatment was used to improve the accessibility of corn stover carbohydrates to subsequently added hydrolytic enzymes. Some carbohydrates, however, were still present after enzymatic hydrolysis and fermentation. Hence, this research was aimed to characterise the recalcitrant carbohydrates, especially the oligosaccharides that remained after hydrolysis and fermentation of dilute ammonia-pretreated corn stover (DACS). RESULTS: About 35% (w/w) of DACS carbohydrates remained after enzymatic hydrolysis and fermentation of the released monosaccharides. One-third of these recalcitrant carbohydrates were water soluble and composed of diverse oligosaccharides. By using UHPLC-MS n , more than 50 oligosaccharides were detected. Glucurono-xylooligosaccharides (UAXOS) with a degree of polymerisation (DP) less than 5 were the most abundant oligosaccharides. The (4-O-methyl) glucuronosyl substituent was mostly attached onto the terminal xylosyl residue. It was shown that the glucuronosyl substituent in some UAXOS was modified into a hexenuronosyl, a glucuronamide or a hexenuronamide residue due to the dilute ammonia pretreatment. Another group of abundant oligosaccharides comprised various xyloglucan oligosaccharides (XGOS), with a DP 5 annotated as XXG as the most pronounced. In addition, disaccharides annotated as xylosyl-glucose with different ß linkages as well as larger carbohydrates were present in the fermentation slurry. CONCLUSIONS: Around one-third of the 35% (w/w) recalcitrant DACS carbohydrates remained as water-soluble saccharides. In this study, more than 50 recalcitrant oligosaccharides were detected, which mostly composed of xylosyl and/or glucosyl residues. The most pronounced oligosaccharides were UAXOS and XGOS. Hence, α-glucuronidase and α-xylosidase were suggested to be added to the enzyme mixture to degrade these oligosaccharides further, and hence the fermentation yield is potentially increased.

Assessment of hydration biomarkers including salivary osmolality during passive and active dehydration.

BACKGROUND/OBJECTIVES: Hydration state can be assessed via body mass change (BMΔ), serum and urine osmolality (Sosm, Uosm), urine-specific gravity (Usg) and urine volume (Uvol). As no hydration index has been shown to be valid in all circumstances, value exists in exploring novel biomarkers such as salivary osmolality (Vosm). Utilizing acute BMΔ as the reference standard, this research examined the efficacy of Sosm, Vosm, Uosm, Uvol and Usg, during passive (PAS) and active (ACT) heat exposure. SUBJECTS/METHODS: Twenty-three healthy men (age, 22±3 years; mass, 77.3±12.8 kg; height, 179.9±8.8cm; body fat, 10.6±4.5%) completed two randomized 5-h dehydration trials (36±1 °C). During PAS, subjects sat quietly, and during ACT, participants cycled at 68±6% maximal heart rate. Investigators measured all biomarkers at each 1% BMΔ. RESULTS: Average mass loss during PAS was 1.4±0.3%, and 4.1±0.7% during ACT. Significant between-treatment differences at -1% BMΔ were observed for Sosm (PAS, 296±4; ACT, 301±4 mOsm/kg) and Uosm (PAS, 895±207; ACT, 661±192 mOsm/kg). During PAS, only Uosm, Uvol and Usg increased significantly (-1 and -2% BMΔ versus baseline). During ACT, Vosm most effectively diagnosed dehydration 2% (sensitivity=86%; specificity=91%), followed by Sosm (sensitivity=83%; specificity=83%). Reference change values were validated for Sosm, Usg and BMΔ. CONCLUSIONS: The efficacy of indices to detect dehydration 2% differed across treatments. At rest (PAS), only urinary indices increased in concert with body water loss. During exercise (ACT), Sosm and Vosm exhibited the highest sensitivity and specificity. Sosm, Usg and BMΔ exhibited validity in serial measurements. These findings indicate hydration biomarkers should be selected by considering daily activities.

Lung tumor development and spontaneous regression in lambs coinfected with Jaagsiekte sheep retrovirus and ovine lentivirus.

Ovine pulmonary adenocarcinoma (OPA) is a naturally occurring and experimentally inducible lung cancer of sheep caused by Jaagsiekte sheep retrovirus (JSRV). The first aim of this study was to monitor the development of OPA with minimally invasive, real-time observations of animals experimentally infected with JSRV as well as ovine lentivirus (maedi-visna virus). Worldwide, simultaneous infection of sheep with these 2 retroviruses is a common occurrence, naturally and experimentally; consequently, the lung tumor homogenates used as inocula contained both viruses. Following inoculation, computed tomography was used to detect tumor nodules early, before the onset of clinical signs, and to monitor tumor advancement. However, not only was OPA disease progression observed, but the apparent spontaneous regression of OPA was witnessed. In fact, regression was more common than progression following JSRV inoculation of neonatal lambs. Immune responses were detected, particularly involving CD3(+) T cells and the production of antibodies against JSRV that may mediate the spontaneous regression of JSRV-induced OPA. The second aim of this study was to determine whether OPA tumors harbor genetic alterations similar to those found in human lung adenocarcinoma. No mutations were found in the tyrosine kinase domain of the epidermal growth factor receptor, KRAS codons 12 and 13, or the DNA-binding domain of p53 in tumor DNA from naturally occurring and experimentally-induced OPA cases. Overall, the genetic profile combined with the disease development data provides further important characterization of OPA and describes, for the first time, spontaneous regression of OPA tumors in experimentally infected sheep.

A sero-survey of rinderpest in nomadic pastoral systems in central and southern Somalia from 2002 to 2003, using a spatially integrated random sampling approach.

A cross-sectional sero-survey, using a two-stage cluster sampling design, was conducted between 2002 and 2003 in ten administrative regions of central and southern Somalia, to estimate the seroprevalence and geographic distribution of rinderpest (RP) in the study area, as well as to identify potential risk factors for the observed seroprevalence distribution. The study was also used to test the feasibility of the spatially integrated investigation technique in nomadic and semi-nomadic pastoral systems. In the absence of a systematic list of livestock holdings, the primary sampling units were selected by generating random map coordinates. A total of 9,216 serum samples were collected from cattle aged 12 to 36 months at 562 sampling sites. Two apparent clusters of RP seroprevalence were detected. Four potential risk factors associated with the observed seroprevalence were identified: the mobility of cattle herds, the cattle population density, the proximity of cattle herds to cattle trade routes and cattle herd size. Risk maps were then generated to assist in designing more targeted surveillance strategies. The observed seroprevalence in these areas declined over time. In subsequent years, similar seroprevalence studies in neighbouring areas of Kenya and Ethiopia also showed a very low seroprevalence of RP or the absence of antibodies against RP. The progressive decline in RP antibody prevalence is consistent with virus extinction. Verification of freedom from RP infection in the Somali ecosystem is currently in progress.

Regulation of surfactant protein and defensin mRNA expression in cultured ovine type II pneumocytes by all-trans retinoic acid and VEGF.

Beta-defensins and surfactant proteins are components of the pulmonary innate immune system. Their gene expression is regulated by development, hormones, growth and immunoregulatory factors. It was our hypothesis that growth and differentiation factors such as all-trans retinoic acid (RA) and vascular endothelial growth factor (VEGF) may affect expression of selected innate immune genes by respiratory epithelial cells. Ovine JS7 cells (alveolar type II pneumocytes) were incubated in serum-free Dulbecco's modified Eagle's medium (DMEM) complete media that contained: no treatment (negative control), RA (500 nM), or VEGF (100 ng/ml) for 6, 12 or 24 h incubation. Total RNA was isolated, cDNA synthesized, and relative mRNA levels of surfactant protein A (SP-A) and SP-D, and sheep beta-defensin-1 (SBD-1) were determined by real-time reverse transcriptase-polymerase chain reaction (RT-PCR). Cells had significantly increased expression of SP-D mRNA at 6 h and 24 h, decreased expression of SP-A mRNA at 12 h, and unchanged levels of SBD-1 mRNA after the treatment with RA compared with their respective negative controls. VEGF did not alter the expression of the three innate immune genes. These findings suggest that SP-A and SP-D have different transcription regulation pathways, and that expression of SBD-1 is not inducible by RA similar to its human homolog HBD-1. The lack of changes induced by VEGF treatment suggests that VEGF does not have a direct effect on epithelial cells, but may affect gene expression indirectly.

Ovine lentivirus-associated leucomyelitis in naturally infected North American sheep.

Leucomyelitis was the predominant feature in four North American adult sheep (cases 1-4) with ovine lentivirus (OvLV) infection. All four animals were OvLV-seropositive and a syncytogenic virus consistent with OvLV was isolated from the brain of case 3 and the lungs of case 4. Clinically, the sheep had dyspnoea and neurologic signs of varying severity. Changes in the central nervous system included asymmetrical meningoleucomyelitis with white matter degeneration in all four sheep and scattered foci of leucoencephalitis in periventricular, subependymal and other white matter areas of the brain of the three animals (cases 1, 2 and 4) for which the brain was examined. In the lungs of two sheep (cases 3 and 4), there was lymphoid interstitial pneumonia with marked lymphoid hyperplasia. The viral capsid antigen (p25) was detected by immunohistochemistry (IHC) in sections of lung, brain and spinal cord of the four sheep and OvLV RNA was detected by in-situ hybridization (ISH) in lung and spinal cord samples. The results confirm the usefulness of the IHC and ISH for differential diagnosis of visna.

Colostral transmission of maedi visna virus: sites of viral entry in lambs born from experimentally infected ewes.

Maedi visna virus (MVV) vertical transmission in sheep via infected colostrums is a very important route of infection in lambs. To verify colostral transmission and to study early viral entry in lambs, colostrum samples, and small intestine and mesenteric lymph nodes of lambs born from experimentally infected ewes were examined by histopathology, immunohistochemistry (IHC) and in situ hybridisation (ISH) studies. In particular, newborn lambs were naturally fed maternal colostrum and humanely killed at 10, 24, 48, 72, 96 h and 7 and 10 days after birth; two caesarian-derived lambs served as uninfected controls. No lesions suggestive of MVV infection were found, but marked immunoreactions for MVV capsid antigen (CA, p28) were detected in lambs fed maternal colostrum and in macrophages cultured from colostrum. IHC results in lambs suggest an initial viral absorption by intestinal epithelial cells at the tip of the villi, passage to mononuclear cells in the lamina propria and involvement of ileum Peyers' patches and mesenteric lymph nodes, with different staining patterns depending on infection times. ISH on intestinal sections of the 72 h lamb revealed the presence of proviral DNA in epithelial cells at the tip of the villi, suggesting a role for these cells in early MVV replication. The results contribute to knowledge about the pathogenesis of ovine lentivirus infection suggesting that the small intestine and mesenteric nodes are the sites of entry and propagation of MVV in lambs fed colostrums from infected ewes.

Natural history of JSRV in sheep.

Ovine pulmonary adenocarcinoma (OPA) is a contagious lung tumour of sheep and, rarely, goats that arises from two types of secretory epithelial cell that retain their luxury function of surfactant synthesis and secretion. It is classified as a low-grade adenocarcinoma and is viewed as a good model for epithelial neoplasia because of its morphological resemblance to the human lung tumour, bronchioloalveolar adenocarcinoma. OPA is present in most of the sheep rearing areas of the globe and, in affected flocks, tumours are present in a high proportion of sheep. OPA is associated with the ovine retrovirus, jaagsiekte sheep retrovirus (JSRV), and is transmissible only with inocula that contain JSRV. All sheep contain JSRV-related endogenous viruses, but JSRV is an exogenous virus that is associated exclusively with OPA. JSRV is detected consistently in the lung fluid, tumour and lymphoid tissues of sheep affected by both natural and experimental OPA or unaffected in-contact flockmates and never in sheep from unaffected flocks with no history of the tumour. JSRV replicates principally in the epithelial tumour cells, but also establishes a disseminated infection of several lymphoid cell types, including peripheral blood leukocytes (PBLs). Longitudinal studies in flocks with endemic OPA have revealed JSRV in PBLs before the onset of clinical OPA and even in the absence of discernible lung tumour. The prevalence of JSRV infection is 40%-80%, although only 30% of sheep appear to develop OPA lesions. A unique feature of OPA is the absence of a specific humoral immune response to JSRV, despite the highly productive infection in the lungs and the disseminated lymphoid infection. This feature is associated with reduced responsiveness to some mitogens, although the phenotypic profile of the peripheral blood remains unaltered. The reduced response is an early and sustained event during infection and may indicate that the failure of infected sheep to produce specific antibodies to JSRV is a direct consequence of infection.

Endogenous retroviruses related to jaagsiekte sheep retrovirus.

Ovine betaretroviruses consist of exogenous viruses [jaagsiekte sheep retrovirus (JSRV) and enzootic nasal tumor virus, (ENTV)] associated with neoplastic diseases of the respiratory tract and 15-20 endogenous viruses (enJSRV) stably integrated in the ovine and caprine genome. Phylogenetic analysis of this group of retroviruses suggests that the enJSRV can be considered as 'modern' endogenous retroviruses with active, exogenous counterparts. Sequence analysis of JSRV, ENTV and enJSRV suggests that enJSRV do not directly contribute to the pathogenesis of ovine pulmonary adenocarcinoma (OPA) or enzootic nasal tumor through large-scale recombination events, but small-scale recombination or complementation of gene function cannot be excluded; experiments involving enJSRV-free sheep, which have not been found, would be needed to investigate this possibility. Evidence of expression of enJSRV structural proteins in tissues of the reproductive tract and lung implies that they do not have a primary role in disease. However, experimental exploitation of exogenous/endogenous retrovirus sequence differences by producing chimeras has been useful in establishing the determinants of JSRV Env-induced transformation. Even if enJSRV do not have a direct role in OPA, their expression during ontogeny or in neonatal life may impact the likelihood of exogenous JSRV infection and disease outcome via the induction of immunological tolerance. Aside from any role in disease, enJSRV loci may serve as useful genetic markers in the sheep and their frequent expression in the reproductive tract of the ewe may portend an important physiologic role in sheep.

Transformation and oncogenesis by jaagsiekte sheep retrovirus.

Jaagsiekte sheep retrovirus (JSRV) is an exogenous retrovirus of sheep that induces a contagious lung cancer, ovine pulmonary adenocarcinoma (OPA). JSRV is a potent carcinogen in the experimental setting, inducing end-stage tumors at around 6 weeks of age when newborn lambs are inoculated intratracheally. Despite this rapid oncogenesis, inspection of the JSRV genome sequence does not reveal any obvious viral oncogenes. In this review, recent advances in studies of JSRV oncogenic transformation are described. Molecular cloning of an infectious and oncogenic JSRV provirus was instrumental in the studies. DNA transfection of JSRV proviral DNA into mouse NIH3T3 cells results in morphological transformation, indicating that the JSRV genome carries an oncogene. Further experiments identified the JSRV envelope protein as the transforming gene, and a PI3 kinase docking site in the cytoplasmic tail of the transmembrane (TM) protein was shown to be necessary for transformation. Avian DF-1 cells infected with an avian retroviral vector (RCAS) expressing the JSRV envelope protein also undergo tumorigenic transformation. Possible mechanisms of transformation are discussed, and a cooperating role for insertional activation of proto-oncogenes in tumorigenesis is also considered. The transforming potential of the JSRV envelope protein may be necessary for JSRV infection and replication in vivo.

Alveolar type II cells expressing jaagsiekte sheep retrovirus capsid protein and surfactant proteins are the predominant neoplastic cell type in ovine pulmonary adenocarcinoma.

Ovine pulmonary adenocarcinoma is caused by jaagsiekte sheep retrovirus. To gain insight into the histogenesis and viral pathogenesis of this neoplasm, the tumor cell phenotypes and differentiation state were correlated with the distribution of jaagsiekte sheep retrovirus capsid protein in neoplastic and normal cells of the lung in nine naturally occurring and 12 experimentally induced cases of ovine pulmonary adenocarcinoma. Overall, 82% of tumor cells had ultrastructural features consistent with alveolar type II cells, 7% of tumor cells had features of Clara cells, and 11% of tumor cells were insufficiently differentiated to classify. The proportion of the neoplastic cell phenotypes varied within tumors, and no tumor consisted of a morphologically uniform cell population. To further characterize the neoplastic cell population, sections of tumors were immunostained with antibodies to surfactant protein A, surfactant protein C, and Clara cell 10-kd protein. Overall, surfactant proteins A and C were expressed in 70% and 80% of tumor cells, respectively, whereas Clara cell 10-kd protein was expressed in 17% of tumor cells. Jaagsiekte sheep retrovirus capsid protein was detected in 71% of tumor cells and in macrophages (5/21 tumors examined) and in nonneoplastic alveolar and bronchiolar cells (6/14 tumors). Expression of this viral protein in neoplastic cells, classified morphologically and by immunophenotyping primarily as of the alveolar type II lineage, implies an important role for specific virus-cell interactions in the pathogenesis of ovine pulmonary adenocarcinoma.

Retroviral antibody binding of the MHC class II molecule: a biochemical influence on CD4 T cell differentiation in HIV infection?

Retroviral antibody capable of binding to the major histocompatibility complex (MHC) Class II molecule has been documented in human immunodeficiency virus-1 (HIV-1)-infected patients. Interactions between the MHC Class II receptor and the T-cell receptor (TCR) are central to the immune response. Importantly, retroviral antibody possesses a much higher binding affinity for the MHC Class II receptor, when compared to the TCR. Experiments have manipulated a number of factors related to antigen-presenting cell (APC) interaction with differentiating T-cells. These studies have observed the effects of lowering antigen dose and reducing ligand density on precursor Th (T helper) cell differentiation. Studies have also examined the effect of downregulated MHC Class II receptors and co-stimulatory molecules on APC-Th cell interaction. In addition, the sequestration of antigens away from the Class II processing pathway has been studied. These investigations reveal a general trend that can determine whether a naive CD4 T-cell becomes a Th1 or Th2-like cell. If the experimental manipulation weakens the APC-Th cell interaction, a weak ligating TCR signal results. Consequently, a weak ligating TCR signal can influence precursor Th cells to become Th2-like cells. Retroviral antibody binding of MHC Class II receptors may mimic a number of experimental conditions responsible for creating a weak ligating TCR signal.

Jaagsiekte sheep retrovirus proviral clone JSRV(JS7), derived from the JS7 lung tumor cell line, induces ovine pulmonary carcinoma and is integrated into the surfactant protein A gene.

Ovine pulmonary carcinoma (OPC) is a contagious neoplasm of alveolar epithelial type II (ATII) or Clara cells caused by a type D/B chimeric retrovirus, jaagsiekte sheep retrovirus (JSRV). Here we report the isolation, sequencing, pathogenicity, and integration site of a JSRV provirus isolated from a sheep lung tumor cell line (JS7). The sequence of the virus was 93 to 99% identical to other JSRV isolates and contained all of the expected open reading frames. To produce virions and test its infectivity, the JS7 provirus (JSRV(JS7)) was cloned into a plasmid containing a cytomegalovirus promoter and transfected into 293T cells. After intratracheal inoculation with virions from concentrated supernatant fluid, JSRV-associated OPC lesions were found in one of four lambs, confirming that JSRV(JS7) is pathogenic. In JS7-cell DNA, the viral genome was inserted in the protein-coding region for the surfactant protein A (SP-A) gene, which is highly expressed in ATII cells, in an orientation opposite to the direction of transcription of the SP-A gene. No significant transcription was detected from either the viral or the SP-A gene promoter in the JS7 cell line at passage level 170. The oncogenic significance of the JSRV proviral insertion involving the SP-A locus in the JS7 tumor cell line is unknown.

Ovine lentivirus is aetiologically associated with chronic respiratory disease of sheep on the Laikipia Plateau in Kenya.

A study was undertaken to investigate the occurrence of ovine lentivirus (OvLV) infection in sheep with chronic respiratory disease on the Laikipia Plateau, Kenya. All seven Merino crossbred sheep with chronic dyspnoea and emaciation examined for gross and microscopic lesions had lymphoid interstitial pneumonia (LIP), and one also had pulmonary abscesses. Two of the sheep with LIP also had lesions of ovine pulmonary carcinoma (OPC, jaagsiekte). Using in situ hybridization, OvLV DNA localized to a high proportion of pulmonary macrophages in lungs with lesions of LIP. Lung tissue samples from six of these sheep were positive for a syncytium-inducing virus in cultures of lamb testis cells. Thin-section electron microscopy of infected cells showed virions with morphogenesis typical of lentiviruses. In a western blotting assay, monoclonal antibodies to the OvLV capsid (CA, p27) and matrix (MA, p15) proteins of a North American OvLV isolate reacted with similar-sized bands of the virus, and serum from six of the sheep were reactive with CA from the Kenyan viral isolate. Using an OvLV agar gel immunodiffusion (AGID) test, all seven sheep were positive for serum antiviral antibody, as were 29% of 63 clinically normal sheep from Laikipia District. However, when sera from the healthy sheep were tested in a western blot assay, only 52% had IgG reactive to the OvLV CA, indicating a high rate of false negative reactions with the AGID test. Serum samples from 87 Red Maasai or Dorper crossbred sheep from two farms in other parts of Kenya were OvLV seronegative by both the AGID test and the western blot assay. These results document the first identification of OvLV as a cause of chronic respiratory disease in sheep in Kenya and show a high rate of infection in sheep flocks, with a high prevalence of chronic respiratory disease.

Epizootic malignant catarrhal fever in three bison herds: differences from cattle and association with ovine herpesvirus-2.

Three bison herds in Colorado experienced high mortality from malignant catarrhal fever (MCF). In comparison with cattle, the bison had a more rapidly progressive disease, fewer clinical signs, and milder inflammatory histologic lesions. There was consistent association with ovine herpesvirus-2 (OHV-2). Contact with sheep was not consistent. Of 17 animals in herd A, 15 died of acute MCF; 1 was slaughtered while healthy; and 1 developed clinical signs of MCF, was treated with corticosteroids and antibiotics, and died of fungal abomasitis and rhinitis after 5 months. In herds B and C, approximately 300 of 900 and 18 of 20 died of MCF following brief clinical disease. The nearest sheep were 1 mile away from herd A, but direct contact with sheep could be documented in herds B and C. Complete gross and histologic examinations were conducted on 34 animals, including all animals in herd A, and MCF was diagnosed in 31. In addition, field necropsies were performed on all dead animals in herd B and most in herd C and MCF was diagnosed on the basis of the gross lesions in most animals. Clinical signs of each animal in herd A were recorded. Illness was brief, usually 8-48 hours. Clinical signs were subtle; separation from the herd was often observed. In all 3 herds, hemorrhagic cystitis and multifocal ulceration of the alimentary tract were consistently found at necropsy. Mild lymphocytic vasculitis was present in multiple organs. Ovine herpesvirus-2 was found by polymerase chain reaction (PCR) in 71 of 105 formalin-fixed tissue specimens from 29 of 31 animals with MCF. In herd A, blood samples from 13 animals were collected at 5 time points and tested by PCR for the presence of OHV-2 viral sequences in peripheral blood leukocytes. Nine bison with a positive PCR test and 4 with negative results prior to clinical illness died of MCF.

Malignant catarrhal fever: polymerase chain reaction survey for ovine herpesvirus 2 and other persistent herpesvirus and retrovirus infections of dairy cattle and bison.

Using a polymerase chain reaction (PCR) test for sequences of ovine herpesvirus 2 (OHV2), this virus was shown to be significantly associated with sheep-associated malignant catarrhal fever (SA-MCF) in terminal cases of disease in 34 cattle and 53 bison. Ovine herpesvirus 2 was not detected in cattle (38) and bison (10) that succumbed to other diseases. Other persistent herpesviruses, retroviruses, and pestivirus, some of which have been previously isolated from cases of SA-MCF, were not associated with the disease. These included bovine herpesvirus 4 (BHV4), bovine lymphotrophic herpesvirus (BLHV), bovine syncytial virus (BSV, also known as bovine spumavirus), bovine immunodeficiency virus (BIV), and bovine viral diarrhea virus (BVDV). A PCR survey for OHV2 in DNA from individual cow's peripheral blood lymphocytes in 4 dairies showed that the 1 dairy that was in close contact to sheep had a prevalence of OHV2 of 21.3%, whereas the 3 other dairies had no OHV2. Prevalence of the other herpesviruses and retroviruses in the dairy cows was variable, ranging from 2% to 51% for BHV4, 52% to 78.7% for BLHV, and 10% to 34% for BSV. Bovine lymphotrophic herpesvirus and BSV were also found in a few (1-4 of 21 tested) cases of terminal SA-MCF, but BIV and BVDV were not found in either the dairy cows sampled, or in the cases of SA-MCE No significant correlation was found between the presence of any 2 viruses (OHV2, BHV4, BLHV, BSV) in the dairy cows or terminal cases of SA-MCE

Retrovirus vectors bearing jaagsiekte sheep retrovirus Env transduce human cells by using a new receptor localized to chromosome 3p21.3.

Jaagsiekte sheep retrovirus (JSRV) is a type D retrovirus associated with a contagious lung tumor of sheep, ovine pulmonary carcinoma. Other than sheep, JSRV is known to infect goats, but there is no evidence of human infection. Until now it has not been possible to study the host range for JSRV because of the inability to grow this virus in culture. Here we show that the JSRV envelope protein (Env) can be used to pseudotype Moloney murine leukemia virus (MoMLV)-based retrovirus vectors and that such vectors can transduce human cells in culture. We constructed hybrid retrovirus packaging cells that express the JSRV Env and the MoMLV Gag-Pol proteins and can produce JSRV-pseudotype vectors at titers of up to 10(6) alkaline phosphatase-positive focus-forming units/ml. Using this high-titer virus, we have studied the host range for JSRV, which includes sheep, human, monkey, bovine, dog, and rabbit cells but not mouse, rat, or hamster cells. Considering the inability of the JSRV-pseudotype vector to transduce hamster cells, we used the hamster cell line-based Stanford G3 panel of whole human genome radiation hybrids to phenotypically map the JSRV receptor (JVR) gene within the p21.3 region of human chromosome 3. JVR is likely a new retrovirus receptor, as none of the previously identified retrovirus receptors localizes to the same position. Several chemokine receptors that have been shown to serve as coreceptors for lentivirus infection are clustered in the same region of chromosome 3; however, careful examination shows that the JSRV receptor does not colocalize with any of these genes.

Evolutionary stable strategy: a test for theories of retroviral pathology which are based upon the concept of molecular mimicry.

The genetic makeup of animal and plant populations is determined by established principles and concepts. Ecology and evolution provide a basic theoretical framework for understanding how genetic changes occur in populations. Whether these rules can be applied to host retroviral populations is unknown. Individuals infected with the human immunodeficiency virus (HIV) contain within their bodies a viral population. This population is known as a viral quasispecies. Located in the transmembrane protein of HIV-1 is the viral sequence Gly-Thr-Asp-Arg-Val. Previous immunological studies have shown that viral antibody is produced in response to this five-amino-acid sequence. Antibody to this viral sequence also crossreacts and binds to a related peptide sequence found on certain immune cells. This related sequence, Gly-Thr-Glu-Arg-Val, is found on immune cells bearing a structure known as the major histocompatibility complex (MHC). The viral transmembrane sequence, Gly-Thr-Asp-Arg-Val, can be substituted with alanine residues utilizing site-directed mutagenesis. This creates a viral clone devoid of the genetic similarity with the MHC. Chimpanzees progressing to AIDS contain both sequences of interest. Suppression of the chimpanzee quasispecies utilizing anti-retroviral drugs is proposed. This action serves to suppress the presence of the viruses containing the sequence Gly-Thr-Asp-Arg-Val. When viral load has been reduced significantly, a drug resistant, alanine altered clone is to be introduced in large numbers. The concept of evolutionary stable strategy predicts that a viable HIV clone with alanine residues can genetically dominate the viral population. Immune system recognition of the alanine sequence is likely to result in renewed antibody production. Antibodies to the alanine containing viral sequence should not recognize or bind to the MHC. Immunological parameters can then be measured to determine the physiological impact of eliminating a sequence responsible for molecular mimicry between virus and host.

Comparison of a maedi-visna virus CA-TM fusion protein ELISA with other assays for detecting sheep infected with North American ovine lentivirus strains.

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.