Chemokine receptors CCR2 and CX3CR1 regulate viral encephalitis-induced hippocampal damage but not seizures.

Viral encephalitis is a major risk factor for the development of seizures, epilepsy, and hippocampal damage with associated cognitive impairment, markedly reducing quality of life in survivors. The mechanisms underlying seizures and hippocampal neurodegeneration developing during and after viral encephalitis are only incompletely understood, hampering the development of preventive treatments. Recent findings suggest that brain invasion of blood-born monocytes may be critically involved in both seizures and brain damage in response to encephalitis, whereas the relative role of microglia, the brain's resident immune cells, in these processes is not clear. CCR2 and CX3CR1 are two chemokine receptors that regulate the responses of myeloid cells, such as monocytes and microglia, during inflammation. We used Ccr2-KO and Cx3cr1-KO mice to understand the role of these receptors in viral encephalitis-associated seizures and neurodegeneration, using the Theiler's virus model of encephalitis in C57BL/6 mice. Our results show that CCR2 as well as CX3CR1 plays a key role in the accumulation of myeloid cells in the CNS and activation of hippocampal myeloid cells upon infection. Furthermore, by using Cx3cr1-creER+/-tdTomatoSt/Wt reporter mice, we show that, with regard to CD45 and CD11b expression, some microglia become indistinguishable from monocytes during CNS infection. Interestingly, the lack of CCR2 or CX3CR1 receptors was associated with almost complete prevention of hippocampal damage but did not prevent seizure development after viral CNS infection. These data are compatible with the hypothesis that CNS inflammatory mechanism(s) other than the infiltrating myeloid cells trigger the development of seizures during viral encephalitis.

Impact of Astrocyte Depletion upon Inflammation and Demyelination in a Murine Animal Model of Multiple Sclerosis.

Astrocytes play a key role in demyelinating diseases, like multiple sclerosis (MS), although many of their functions remain unknown. The aim of this study was to investigate the impact of astrocyte depletion upon de- and remyelination, inflammation, axonal damage, and virus distribution in Theiler`s murine encephalomyelitis (TME). Groups of two to six glial fibrillary acidic protein (GFAP)-thymidine-kinase transgenic SJL mice and SJL wildtype mice were infected with TME virus (TMEV) or mock (vehicle only). Astrocyte depletion was induced by the intraperitoneal administration of ganciclovir during the early and late phase of TME. The animals were clinically investigated while using a scoring system and a rotarod performance test. Necropsies were performed at 46 and 77 days post infection. Cervical and thoracic spinal cord segments were investigated using hematoxylin and eosin (H&E), luxol fast blue-cresyl violet (LFB), immunohistochemistry targeting Amigo2, aquaporin 4, CD3, CD34, GFAP, ionized calcium-binding adapter molecule 1 (Iba1), myelin basic protein (MBP), non-phosphorylated neurofilaments (np-NF), periaxin, S100A10, TMEV, and immunoelectron microscopy. The astrocyte depleted mice showed a deterioration of clinical signs, a downregulation and disorganization of aquaporin 4 in perivascular astrocytes accompanied by vascular leakage. Furthermore, astrocyte depleted mice showed reduced inflammation and lower numbers of TMEV positive cells in the spinal cord. The present study indicates that astrocyte depletion in virus triggered CNS diseases contributes to a deterioration of clinical signs that are mediated by a dysfunction of perivascular astrocytes.

The influence of macrophage growth factors on Theiler's Murine Encephalomyelitis Virus (TMEV) infection and activation of macrophages.

Macrophages are common targets for infection and innate immune activation by many pathogenic viruses including the neurotropic Theiler's Murine Encephalomyelitis Virus (TMEV). As both infection and innate activation of macrophages are key determinants of viral pathogenesis especially in the central nervous system (CNS), an analysis of macrophage growth factors on these events was performed. C3H mouse bone-marrow cells were differentiated in culture using either recombinant macrophage colony stimulating factor (M-CSF) or granulocyte-macrophage colony-stimulating factor (GM-CSF), inoculated with TMEV (BeAn) and analyzed at various times thereafter. Cytokine RNA and protein analysis, virus titers, and flow cytometry were performed to characterize virological parameters under these culture conditions. GM-CSF-differentiated macrophages showed higher levels of TMEV viral RNA and proinflammatory molecules compared to infected M-CSF-differentiated cells. Thus, GM-CSF increases both TMEV infection and TMEV-induced activation of macrophages compared to that seen with M-CSF. Moreover, while infectious viral particles decreased from a peak at 12h to undetectable levels at 48h post infection, TMEV viral RNA remained higher in GM-CSF- compared to M-CSF-differentiated macrophages in concert with increased proinflammatory gene expression. Analysis of a possible basis for these differences determined that glycolytic rates contributed to heightened virus replication and proinflammatory cytokine secretion in GM-CSF compared to M-CSF-differentiated macrophages. In conclusion, we provide evidence implicating a role for GM-CSF in promoting virus replication and proinflammatory cytokine expression in macrophages, indicating that GM-CSF may be a key factor for TMEV infection and the induction of chronic TMEV-induced immunopathogenesis in the CNS.

Development of a reverse transcription recombinase polymerase amplification assay for rapid detection of Theiler's murine encephalomyelitis virus.

Theiler's murine encephalomyelitis virus (TMEV) is one of the most common viral pathogens that circulate widely in captive mouse colonies. A molecular biology detection method would be a useful tool to use in an integrated program to monitor and prevent TMEV infection and transmission. Thus, a reverse transcription recombinase polymerase amplification (RT-RPA) assay was developed to detect TMEV infection. The sensitivity of the RT-RPA assay approached 8 copies per reaction, which is equivalent to the sensitivity of RT-qPCR reactions. This assay did not detect RNA extracts from other murine pathogens included in this study or TMEV negative samples. Brain tissues and contaminated biological materials were used to assess the clinical performance of the RT-RPA. The detection results of RT-RPA and RT-qPCR were very similar, except that a contaminated biological material sample which was positive by RT-qPCR, with a CT value of 38, was negative by RT-RPA. In summary, the developed RT-RPA assay offers a rapid, sensitive and specific alternative method for monitoring of TMEV, especially in resource-limited conditions.

Saffold virus, an emerging human cardiovirus.

Saffold virus (SAFV) is an emerging human cardiovirus that has been shown to be ubiquitous. Initial studies of SAFV focused on respiratory and gastrointestinal infection; however, it has also recently been associated with diverse clinical symptoms including the endocrine, cardiovascular, and neurological systems. Given the systemic nature of SAFV, and its high prevalence, understanding its pathogenicity and clinical impact is of utmost importance. This comprehensive review highlights and discusses recent developments in epidemiology, human pathogenicity, animal, and molecular studies related to SAFV. It also provides detailed insights into the neuropathogenicity of SAFV. We argue that human studies have been confounded by coinfections and therefore require support from robust molecular and animal research. Thereby, we aim to provide foresight into further research to better understand this emerging virus.

The level of viral infection of antigen-presenting cells correlates with the level of development of Theiler's murine encephalomyelitis virus-induced demyelinating disease.

UNLABELLED: Intracerebral infection with Theiler's murine encephalomyelitis virus (TMEV) induces immune-mediated demyelinating disease in susceptible SJL/J mice but not in resistant C57BL/6 mice. Previous studies have indicated that the major histocompatibility complex (MHC) genes play the most prominent role in the development of TMEV-induced demyelinating disease. In this study, we used C57BL/6.S (B6.S) congenic mice, which carry H-2(s) MHC genes instead of H-2(b) MHC genes in conjunction with the C57BL/6 (B6) background genes. Our data show that virus-infected B6.S mice are free from disease and have significantly lower viral loads than susceptible SJL mice, particularly in the spinal cord. A strong protective Th1-type T helper response with virtually no pathogenic Th17 response was detected in B6.S mice, in contrast to the reduced Th1- and robust Th17-type responses in SJL mice. Notably, lower levels of viral infectivity in B6.S antigen-presenting cells (APCs) correlated with the disease resistance and T-cell-type response. In vitro studies using APCs from B6.S and SJL mice show that TLR2, -3, -4, and -7, but not TLR9, signaling can replace viral infection and augment the effect of viral infection in the differentiation of the pathogenic Th17 cell type. Taken together, these results strongly suggest that the viral replication levels in APCs critically affect the induction of protective versus pathogenic Th cell types via the signaling of pattern recognition receptors for innate immune responses. Our current findings further imply that the levels of viral infectivity/replication and TLR-mediated signaling play critical roles in the pathogenesis of chronic viral diseases. IMPORTANCE: This study indicates that innate immune cytokines produced in antigen-presenting cells stimulating the T cell immune responses during early viral infection play a critical role in determining the susceptibility of mice to the development of demyelinating disease. The level of innate immune cytokines reflects the level of initial viral infection in the antigen-presenting cells, and the level determines the development of T cell types, which are either protective or pathogenic. The level of initial viral infection to the cells is controlled by a gene or genes that are not associated with the major histocompatibility antigen complex genes. This finding has an important implication in controlling not only chronic viral infections but also infection-induced autoimmune-like diseases, which are closely associated with the pathogenic type of T cell responses.

Immunohistochemical insights into Saffold virus infection of the brain of juvenile AG129 mice.

BACKGROUND: Saffold Virus (SAFV) is a human cardiovirus that is suspected of causing infection of the central nervous system (CNS) in children. While recent animal studies have started to elucidate the pathogenesis of SAFV, very little is known about the mechanisms behind it. METHOD: In this study, we attempted to elucidate some of the mechanisms of the pathogenesis of SAFV in the brain of a juvenile mouse model by using immunohistochemical methods. RESULTS: We first showed that SAFV is able to infect both neuronal and glial cells in the brain of 2 week-old AG129 mice. We then showed that SAFV is able to induce apoptosis in both neuronal and glial cells in the brain. Lastly, we showed that SAFV infection does not show any signs of gross demyelination in the brain. CONCLUSION: Overall, our results provide important insights into the mechanisms of SAFV in the brain.

SAffold viruses in pediatric patients with diarrhea in Thailand.

Saffold virus (SAFV) is a newly discovered human virus which is classified into the genus Cardiovirus of the family Picornaviridae. A total of 608 fecal specimens collected during January 2012 to December 2013 from children with diarrhea in Chiang Mai, Thailand were investigated for SAFV by RT-nested PCR and sequence analysis. Of these, nine out of 608 (1.5%) were positive for SAFVs and four genotypes were identified, SAFV1, SAFV2, SAFV3, and SAFV4. SAFV mono-infection was found in five cases (CMH-S038-12, CMH-S071-12, CMH-S102-12, CMH-N029-12, and CMH-S048-13), while co-infection with other viruses causing diarrhea was observed in four cases (CMH-S021-12, CMH-S115-12, CMH-N048-13 and CMH-N103-13). This study provides more information about the genetic background of SAFV circulating in pediatric patients with diarrhea in Thailand.

[Molecular genetic characteristics of the virus isolated from patients with human acute encephalomyelitis and multiple sclerosis].

The study of the antigenic and molecular genetic structure of human acute encephalomyelitis virus (HAEV) showed a high similarity of the HAEV N gene with the homologous gene of the fixed rabies virus strain. The results of the nucleotide sequence analysis indicate that HAEV belongs to the lyssavirus genotype 1. The N gene sequence is the closest to those of the ERA-CB20-M and RV-97 strains of the rabies virus. The need for further research into the role of the human acute encephalomyelitis virus in human pathology stems from past surveys that revealed the presence of the VNAs against this virus in 6 per cent of the blood received from donors in the USA and in each third among the patients with multiple sclerosis in the former USSR.

Melanoma differentiation-associated gene 5 is critical for protection against Theiler's virus-induced demyelinating disease.

Infection of dendritic and glial cells with Theiler's murine encephalomyelitis virus (TMEV) induces various cytokines via Toll-like receptor- and melanoma differentiation-associated gene 5 (MDA5)-dependent pathways. However, the involvement and role of MDA5 in cytokine gene activation and the pathogenesis of TMEV-induced demyelinating disease are largely unknown. In this study, we demonstrate that MDA5 plays a critical role in the production of TMEV-induced alpha interferon (IFN-α) during early viral infection and in protection against the development of virus-induced demyelinating disease. Our results indicate that MDA5-deficient 129SvJ mice display significantly higher viral loads and apparent demyelinating lesions in the central nerve system (CNS) accompanied by clinical symptoms compared with wild-type 129SvJ mice. During acute viral infection, MDA5-deficient mice produced elevated levels of chemokines, consistent with increased cellular infiltration, but reduced levels of IFN-α, known to control T cell responses and cellular infiltration. Additional studies with isolated CNS glial cells from these mice suggest that cells from MDA5-deficient mice are severely compromised in the production of IFN-α upon viral infection, which results in increased cellular infiltration and viral loads in the CNS. Despite inadequate stimulation, the overall T cell responses to the viral determinants were significantly elevated in MDA5-deficient mice, reflecting the increased cellular infiltration. Therefore, the lack of MDA5-mediated IFN-α production may facilitate a massive viral load and elevated cellular infiltration in the CNS during early viral infection, leading to the pathogenesis of demyelinating disease.

Mapping loci influencing the persistence of Theiler's virus in the murine central nervous system.

Inbred strains of mice differ greatly in their susceptibility to the demyelinating disease caused by Theiler's Murine Encephalomyelitis Virus. In this murine disease, which is an animal model for the study of multiple sclerosis, demyelination depends on the persistent infection of the central nervous system. Previous studies identified a locus in the H-2D region of the major histocompatibility complex which controls susceptibility to the persistent infection, and also showed that other loci are involved. In order to identify these loci, we screened the genome of a set of backcross animals with a combination of polymorphic microsatellites and restriction enzymes sites. We now show that viral persistence is also controlled by a locus close to Ifg on chromosome 10 and possibly by a locus near Mbp on chromosome 18.

Identification of a novel neuropathogenic Theiler's murine encephalomyelitis virus.

Theiler's murine encephalitis viruses (TMEV) are divided into two subgroups based on their neurovirulence. Persistent strains resemble Theiler's original viruses (referred to as the TO subgroup), which largely induce a subclinical polioencephalomyelitis during the acute phase of the disease and can persist in the spinal cord of susceptible animals, inducing a chronic demyelinating disease. In contrast, members of the neurovirulent subgroup cause an acute encephalitis characterized by the rapid onset of paralysis and death within days following intracranial inoculation. We report herein the characterization of a novel neurovirulent strain of TMEV, identified using pyrosequencing technology and referred to as NIHE. Complete coverage of the NIHE viral genome was obtained, and it shares <90% nucleotide sequence identity to known TMEV strains irrespective of subgroup, with the greatest sequence variability being observed in genes encoding the leader and capsid proteins. The histopathological analysis of infected brain and spinal cord demonstrate inflammatory lesions and neuronal necrosis during acute infection with no evidence of viral persistence or chronic disease. Intriguingly, genetic analysis indicates the putative expression of the L protein, considered a hallmark of strains within the persistent subgroup. Thus, the identification and characterization of a novel neurovirulent TMEV strain sharing features previously associated with both subgroups will lead to a deeper understanding of the evolution of TMEV strains and new insights into the determinants of neurovirulence.

Interleukin-6, produced by resident cells of the central nervous system and infiltrating cells, contributes to the development of seizures following viral infection.

Cells that can participate in an innate immune response within the central nervous system (CNS) include infiltrating cells (polymorphonuclear leukocytes [PMNs], macrophages, and natural killer [NK] cells) and resident cells (microglia and sometimes astrocytes). The proinflammatory cytokine interleukin-6 (IL-6) is produced by all of these cells and has been implicated in the development of behavioral seizures in the Theiler's murine encephalomyelitis virus (TMEV)-induced seizure model. The assessment, via PCR arrays, of the mRNA expression levels of a large number of chemokines (ligands and receptors) in TMEV-infected and mock-infected C57BL/6 mice both with and without seizures did not clearly demonstrate the involvement of PMNs, monocytes/macrophages, or NK cells in the development of seizures, possibly due to overlapping function of the chemokines. Additionally, C57BL/6 mice unable to recruit or depleted of infiltrating PMNs and NK cells had seizure rates comparable to those of controls following TMEV infection, and therefore PMNs and NK cells do not significantly contribute to seizure development. In contrast, C57BL/6 mice treated with minocycline, which affects monocytes/macrophages, microglial cells, and PMNs, had significantly fewer seizures than controls following TMEV infection, indicating monocytes/macrophages and resident microglial cells are important in seizure development. Irradiated bone marrow chimeric mice that were either IL-6-deficient mice reconstituted with wild-type bone marrow cells or wild-type mice reconstituted with IL-6-deficient bone marrow cells developed significantly fewer behavioral seizures following TMEV infection. Therefore, both resident CNS cells and infiltrating cells are necessary for seizure development.

Isolation of saffold virus type 2 in green monkey kidney cells.

Saffold viruses (SAFV) have been discovered recently and they are classified into Theilovirus species in genus Cardiovirus in the Picornaviridae family. SAFV, especially those belonging to the genotype 2, have been difficult to propagate in laboratory cell lines. This study describes the successful isolation of an efficiently growing SAFV-2 strain directly from a stool specimen by standard virological methods. The availability of SAFV isolates that can be propagated to high titers is crucial to the future studies on pathogenesis and epidemiology of these novel human viruses.

Identification of cardioviruses related to Theiler's murine encephalomyelitis virus in human infections.

Cardioviruses comprise a genus of picornaviruses that cause severe illnesses in rodents, but little is known about the prevalence, diversity, or spectrum of disease of such agents among humans. A single cardiovirus isolate, Saffold virus, was cultured in 1981 in stool from an infant with fever. Here, we describe the identification of a group of human cardioviruses that have been cloned directly from patient specimens, the first of which was detected using a pan-viral microarray in respiratory secretions from a child with influenza-like illness. Phylogenetic analysis of the nearly complete viral genome (7961 bp) revealed that this virus belongs to the Theiler's murine encephalomyelitis virus (TMEV) subgroup of cardioviruses and is most closely related to Saffold virus. Subsequent screening by RT-PCR of 719 additional respiratory specimens [637 (89%) from patients with acute respiratory illness] and 400 cerebrospinal fluid specimens from patients with neurological disease (aseptic meningitis, encephalitis, and multiple sclerosis) revealed no evidence of cardiovirus infection. However, screening of 751 stool specimens from 498 individuals in a gastroenteritis cohort resulted in the detection of 6 additional cardioviruses (1.2%). Although all 8 human cardioviruses (including Saffold virus) clustered together by phylogenetic analysis, significant sequence diversity was observed in the VP1 gene (66.9%-100% pairwise amino acid identities). These findings suggest that there exists a diverse group of novel human Theiler's murine encephalomyelitis virus-like cardioviruses that hitherto have gone largely undetected, are found primarily in the gastrointestinal tract, can be shed asymptomatically, and have potential links to enteric and extraintestinal disease.

Management of an outbreak of rat theilovirus.

Rat theilovirus is a commonly reported infection in research rat colonies. The author's institution experienced an outbreak of rat theilovirus in a breeding colony of unique outbred rats. To manage this outbreak, the institution chose to use a 'test and cull' strategy because this approach is reported to be successful in mouse colonies infected with Theiler's murine encephalomyelitis virus, a related virus. Here the author describes the outbreak and subsequent management of rat theilovirus. The strategy successfully cleared the virus from the rat colony.

Restraint stress modulates virus specific adaptive immunity during acute Theiler's virus infection.

Multiple sclerosis (MS) is a devastating CNS disease of unknown origin. Multiple factors including genetic background, infection, and psychological stress affect the onset or progression of MS. Theiler's murine encephalomyelitis virus (TMEV) infection is an animal model of MS in which aberrant immunity leads to viral persistence and subsequently results in demyelination that resembles MS. Here, we examined how stress during acute TMEV infection altered virus-specific cell mediated responses. Using immunodominant viral peptides specific for either CD4(+) or CD8(+) T cells, we found that stress reduced IFN-gamma producing virus-specific CD4(+) and CD8(+) T cells in the spleen and CD8(+) T cells CNS. Cytokine production by cells isolated from the CNS or spleens following stimulation with virus or viral peptides, indicated that stress decreased both type 1 and type 2 responses. Glucocorticoids were implicated in the decreased T cell function as the effects of stress were partially reversed by concurrent RU486 administration but mimicked by dexamethasone. As T cells mediate viral clearance in this model, our data support the hypothesis that stress-induced immunosuppression may provide a mechanism for enhanced viral persistence within the CNS.

Limited remyelination in Theiler's murine encephalomyelitis due to insufficient oligodendroglial differentiation of nerve/glial antigen 2 (NG2)-positive putative oligodendroglial progenitor cells.

AIMS: Limited remyelination is a key feature of demyelinating Theiler's murine encephalomyelitis (TME). It is hypothesized that a dysregulation of differentiation of oligodendroglial progenitor cells (OPCs) represents the main cause of insufficient regeneration in this model of multiple sclerosis. METHODS: TME virus (TMEV)-infected SJL/J mice were evaluated by footprint analysis, light and electron microscopy, immunohistology, confocal immunofluorescence and RT-qPCR at multiple time points ranging from 1 h to 196 days post infection (dpi). RESULTS: Footprint analysis revealed a significantly decreased stride length at 147 and 196 dpi. Demyelination progressively increased from 14 towards 196 dpi. A mild amount of remyelination was detected at 147 and 196 dpi. Early onset axonal injury was detected from 14 dpi on. TMEV RNA was detectable throughout the observation period and markedly increased between 7 and 28 dpi. Intralesional nerve/glial antigen 2 (NG2)-positive OPCs were temporarily increased between 28 and 98 dpi. Similarly, a transient upregulation of NG2 and platelet-derived growth factor alpha-receptor mRNA was noticed. In contrast, intralesional 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNPase)-positive oligodendrocytes were decreased between 56 and 196 dpi. Although CNPase mRNA remained unchanged, myelin basic protein mRNA and especially its exon 2 containing splice variants were decreased. Glial fibrillary acidic protein (GFAP)-positive astrocytes and GFAP mRNA were increased in the late phase of TME. A mildly increased colocalization of both NG2/CNPase and NG2/GFAP was revealed at 196 dpi. CONCLUSIONS: Summarized, the present results indicated a dysregulation of OPC maturation as the main cause for the delayed and limited remyelination in TME. A shift of OPC differentiation from oligodendroglial towards astrocytic differentiation is postulated.

Differential transcription of matrix-metalloproteinase genes in primary mouse astrocytes and microglia infected with Theiler's murine encephalomyelitis virus.

The BeAn strain of Theiler's murine encephalomyelitis virus (TMEV) induces demyelinating disease in susceptible mice comparable to human multiple sclerosis. Recent in vivo studies showed that matrix metalloproteinases (MMPs) and their inhibitors (tissue inhibitors of MMPs, TIMPs) are associated with demyelination in Theiler's murine encephalomyelitis. The present study was performed to evaluate the in vitro MMP and TIMP expression in astrocytes and microglia following TMEV infection. Brain cell cultures from SJL/J mice were infected with the BeAn strain of TMEV and the expressions of 11 MMPs and 4 TIMPs were evaluated by reverse-transcription quantitative polymerase chain reaction (RT-qPCR) at different time points post infection (p.i.). In control astrocytes and microglia, a constitutive expression of MMP-2, -3, -9, -10, -12, -13, -14, -15, -24 and TIMP-2 to -4 was detected. In addition, TIMP-1 and MMP-11 was found in astrocytes only, and MMP-7 was absent in both cells cultures. RT-qPCR demonstrated high virus RNA copy numbers in astrocytes and a low amount in microglia. In accordance, TMEV antigen was detected in astrocytes, whereas it was below the limit of detection in microglia. MMP-3, -9, -10, -12, and -13 as well as TIMP-1 were the enzymes most prominently up-regulated in TMEV-infected astrocytes. In contrast, TMEV infection was associated with a down-regulation of MMPs and TIMPs in microglia. Conclusively, in addition to inflammatory infiltrates, TMEV-induced astrocytic MMPs might trigger a proteolysis cascade leading to an opening of the blood-brain barrier and demyelination in vivo.

Differential susceptibility of SD and CD rats to a novel rat theilovirus.

Antibodies to rat theilovirus (RTV) have been detected in rats for many years because of their serologic crossreactivity with strains of Theiler murine encephalomyelitis virus (TMEV) of mice. Little information exists regarding this pathogen, yet it is among the most common viruses detected in serologic surveys of rats used in research. In the study reported here, a novel isolate of RTV, designated RTV1, was cultured from the feces of infected rats. The RTV1 genome contained 8094 nucleotides and had approximately 95% identity with another rat theilovirus, NSG910, and 73% identity with TMEV strains. In addition, the genome size of RTV1 was similar to those of TMEV strains but larger than that reported for NSG910. Oral inoculation of Sprague-Dawley (SD) and CD male rats (n = 10 each group) with RTV1 revealed that SD rats were more susceptible than CD rats to RTV1 infection. At 14 d postinoculation, 100% of SD rats shed virus in the feces, and 70% were positive for RTV serum antibodies. By 56 d postinoculation 30% of SD rats continued to have detectable virus in the feces, and 90% had seroconverted. In contrast, in inoculated CD rats RTV was detected only in the feces at 14 d postinoculation, at which time 40% of CD rats were fecal positive. By 56 d postinoculation only 20% of CD rats had detectable RTV serum antibodies. Our data provide additional sequence information regarding a rat-specific Cardiovirus and indicate that SD rats are more susceptible than CD rats to RTV1 infection.