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Infant rats are susceptible to persistent rat virus (RV) infection, but risk of persistent infection after prenatal exposure to virus is unclear. We examined this aspect of RV infection in the progeny of dams inoculated with virus during or prior to pregnancy. Sprague-Dawley (SD) dams were infected during pregnancy (gestation day 9) by oronasal inoculation with 10(5) TCID50 of the UMass strain of RV. SD rats were infected prior to pregnancy by oronasal inoculation of two-day-old females with 10(2) TCID50 of RV-UMass, which induced persistent infection. They were mated to non-immune males after reaching sexual maturity. Rats were assessed for RV infection by virus isolation, in situ hybridization, contact transmission, or serologic testing. The progeny of dams inoculated with virus during gestation had high prevalence of infection through postpartum week 9 (9 of 12 rats were virus positive at week 3, and 7 of 10 were virus positive at week 9). Additionally, 2 of 10 rats were virus positive at least through postpartum week 15. The progeny from persistently infected, seropositive dams had no evidence of infection and did not transmit infection to contact sentinels. However, 12 dams were virus positive at necropsy and 9 had transmitted infection to their breeding partners. These results indicate that prenatal infection in non-immune dams can lead to RV persistence in their progeny. By contrast, the progeny of persistently infected dams are protected from infection, presumably by maternal antibody, although their dams can transmit infection to their breeding partners.

Rat virus (RV) infection can cause disease or disrupt responses that rely on cell proliferation. Therefore, persistent infection has the potential to amplify RV interference with research. As a step toward determining underlying mechanisms of persistence, we compared acute and persistent RV infections in infant euthymic and athymic rats inoculated oronasally with the University of Massachusetts strain of RV. Rats were assessed by virus isolation, in situ hybridization, and serology. Selected tissues also were analyzed by Southern blotting or immunohistochemistry. Virus was widely disseminated during acute infection in rats of both phenotypes, whereas vascular smooth muscle cells (SMC) were the primary targets during persistent infection. The prevalence of virus-positive cells remained moderate to high in athymic rats through 8 weeks but decreased in euthymic rats by 2 weeks, coincident with seroconversion and perivascular infiltration of mononuclear cells. Virus-positive pneumocytes and renal tubular epithelial cells also were detected through 8 weeks, implying that kidney and lung excrete virus during persistent infection. Viral mRNA was detected in SMC of both phenotypes through 8 weeks, indicating that persistent infection includes virus replication. However, only half of the SMC containing viral mRNA at 4 weeks stained for proliferating cell nuclear antigen, a protein expressed in cycling cells. The results demonstrate that vasculotropism is a significant feature of persistent infection, that virus replication continues during persistent infection, and that host immunity reduces, but does not eliminate, infection.

Background: The recently identified autonomous mouse parvovirus designated mouse parvovirus-1 (MPV-1) persists in adult BALB/c mice for at least 9 weeks, infects lymphoid tissues, interferes with the ability of cloned T cells to proliferate, and exhibits immunomodulatory properties. As a consequence of these findings, the present studies were undertaken to characterize further the inmunomodulatory effects of MPV-1 on T cell-mediated immune responses in vivo and in vitro., Methods: To evaluate the effect of MPV-1 infection on CD8+ T cell-mediated responses, BALB/c-H2dm2 mice were infected after transplantation of allogeneic BALB/c skin., Results: MPV-1 potentiated the rejection of allogeneic skin grafts. This potentiation was not a result of virus infecting the cellular or vascular component of the graft as determined by in situ hybridization, but was mediated by T cells. However, the proliferative capacity of alloantigen-reactive lymphocytes from graft-sensitized infected mice was diminished. MPV-1 also induced the rejection of syngeneic skin grafts, and T cells from these infected graft-sensitized mice lysed syngeneic P815 target cells., Conclusions: These results suggest that MPV-1 infection of skin-grafted mice may disrupt normal mechanisms of peripheral tolerance and provide a unique model to study virus-induced autoimmunity.

A newly recognized parvovirus of laboratory rats, designated rat parvovirus type 1a (RPV-1a), was found to be antigenically distinct. It was cloned, sequenced, and compared with the University of Massachusetts strain of rat virus (RV-UMass) and other autonomous parvoviruses. RPV-1a VP1 identity with these viruses never exceeded 69%, thus explaining its antigenic divergence. In addition, RPV-1a had reduced amino acid identity in NS coding regions (82%), reflecting phylogenetic divergence from other rodent parvoviruses. RPV-1a infection in rats had a predilection for endothelium and lymphoid tissues as previously reported for RV. Infectious RPV-1a was isolated 3 weeks after inoculation of infant rats, suggesting that it, like RV, may result in persistent infection. In contrast to RV, RPV-1a was enterotropic, a characteristic previously associated with parvovirus infections of mice rather than rats. RPV-1a also differed from RV in that infection was nonpathogenic for infant rats under conditions where RV infection causes high morbidity and mortality. Thus, RPV-1a is the prototype virus of an antigenically, genetically, and biologically distinct rodent parvovirus serogroup.

Parvoviruses are prevalent and disruptive infectious agents of laboratory rats. Risks to rat-based research from infection are increased by the persistence of virus in immune rats and by prenatal transmission of infection. The mechanisms leading to viral persistence and prenatal infection are poorly understood and have been difficult to study for lack of reliable and humane induction methods. We report here protocols for inducing persistent and prenatal infection without causing clinical disease using the UMass strain of rat virus (RV), a common rat parvovirus. Infant rats inoculated by the oronasal route at 6 days of age had greater than 90% prevalence of persistent infection. RV-UMass also induced intrauterine infection in pregnant rats inoculated by the oronasal route. Inoculation of dams at gestation day 9 frequently caused severe disease in the fetuses whereas inoculation at gestation day 12 caused primarily asymptomatic fetal infection that persisted post partum RV-UMass infection facilitates study of parvoviralhost interactions that are relevant to laboratory rats and which also may improve understanding of persistent and prenatal human parvovirus infection.

Parvoviruses are among the most common infectious agents of laboratory rodents and major impediments to rodent-based research. The original prototypic rodent parvoviruses-minute virus of mice, rat virus, and H-1 virus-have recently been joined by biologically and antigenically distinct parvoviruses in mice, rats, and hamsters. Recognition of the increased diversity of rodent parvoviruses presents new challenges for determining the impact of parvovirus infection on research and for detecting, preventing, and eliminating infection. This review summarizes current knowledge about rodent parvoviruses and parvovirus infections, highlighting recent research on newly isolated virus strains.

Inoculation of the UMass strain of rat virus (RV-UMass) into adult immunocompetent rats results in a prolonged subclinical infection that is resolved in 4 to 8 wk. Co-labeling studies, using in situ hybridization (ISH) and immunohistochemistry (IHC), confirmed that RV-UMass was lymphocytotropic and capable of infecting CD4+ and CD8+ T cells as well as B cells. ISH studies also revealed that virus replication was restricted in unstimulated cells but was productive in concanavalin A-stimulated lymphocytes. A corollary of productive infection of lymphocytes was the suppression of lymphocyte functions. Although RV-UMass did not appear to induce phenotypic changes during the course of infection, cells from infected rats had diminished proliferation and cytolytic responses. Both peripheral and mesenteric lymph node cells exhibited only partial recovery of their proliferative and cytolytic capacities one month after infection. Furthermore, RV-UMass-infected tissue culture maintained alloreactive CD4+ T cells in vitro, and a nonlethal infection of this T cell line inhibited Ag- and IL-2-induced proliferation. Because parvoviruses are widespread among laboratory rodents, these findings emphasize the importance of identifying and excluding parvovirus infection in rodents and in cultures of rat T lymphocytes.

Infection of young adult BALB/cByJ mice with mouse parvovirus-1, a newly recognized, lymphocytotropic, nonpathogenic parvovirus, was examined by in situ hybridization. Virus appeared to enter through the small intestine and was disseminated to the liver and lymphoid tissues. Strand-specific probes detected virion DNA in a consistently larger number of cells than replicative forms of viral DNA and/or viral mRNA. The number of signal-positive cells in the intestinal mucosa, lymph nodes, spleen, and thymus increased through day 10 after oral inoculation but decreased after seroconversion. Positive cells were still detected, however, in peripheral lymphoid tissues of mice examined at 9 weeks postinoculation. The results underscore the need to assess potential effects of persistent mouse parvovirus-1 infection on immune function in mice.

In contrast to euthymic juvenile rats, which develop acute, self-limiting infection with rat virus (RV), RV infection of juvenile athymic rats was persistent for up to 12 weeks as demonstrated by recovery of infective virus, transmission to cagemates, and detection of viral DNA in the lungs. Administration of RV antiserum at the time of virus inoculation prevented persistent infection in five of six rats. Among rats given RV antiserum 1 week after virus, the interval at which euthymic rats begin to seroconvert, RV was not detected 1 week later but was recovered from four of six rats 3 weeks later. Results of these studies confirm that T-cell deficiency facilitates persistent RV infection and indicate that antibody provides significant protection from persistent infection only if it is present at the time of virus inoculation. The results support the concept that factors which prevent persistent infection in euthymic rats act early after virus inoculation and may include cellular immunity.

The environmental stability and transmission of a field isolate of rat virus was tested under conditions resembling those that may be encountered during the housing of laboratory rats. The rat virus kept in physiologic salt solutions at room temperature remained infective for at least 5 weeks. Similar virus preparations remained infective after drying on a plastic surface for 3 to 5 weeks, depending on initial virus concentration. Varying the protein concentration in the medium had no significant effect on stability. Bedding from cages housing infected litters induced seroconversions in sentinel rats for at least 5 weeks after storage of rat virus at room temperature. Infection was transmitted between rats housed in open cages in a Trexler isolator but not between rats housed in microisolator cages connected by tunnels partitioned by wire screens with a mesh size of 1.67 mm. The results indicate that rat virus can remain infective after prolonged exposure to an ambient environment and suggest that infection is more readily transmitted by animal-to-animal contact or by fomites than by aerosolization of exhaled virus.

In situ hybridization and virus titration were used to characterize early stages of rat virus (RV) infection of rat pups after oronasal inoculation. Results suggest that virus enters through the lung and that early viremia leads rapidly to pantropic infection. Cells derived from all three germ layers were infected with RV, but those of endodermal and mesodermal origin were the predominant targets. Infection of vascular endothelium was widespread and was associated with hemorrhage and infarction in the brain. Convalescence from acute infection was accompanied by mononuclear cell infiltrates at sites containing RV DNA. Viral DNA was also detected in endothelium, fibroblasts and smooth muscle myofibers four weeks after inoculation. Further examination of these cells as potential sites of persistent infection is warranted.

A virus antigenically related to, but distinct from, minute virus of mice was assessed for infectivity in neonatal and weanling random-bred mice and was equally infectious for both age groups. The virus, designated a mouse "orphan" parvovirus (OPV), was also localized in tissues of experimentally infected random-bred, inbred, and immunodeficient mice by in situ hybridization. Hybridization signal was seen in exocrine and endocrine pancreas, abdominal lymph nodes, mesentery, intestine, and sporadically in other tissues of Sencar, C3H, and DBA mice inoculated as infants. In adult BALB/c severe combined immunodeficient (scid) mice, signal was seen in lung, liver, spleen, lymph nodes, and intestine but not in pancreas. Transmission of OPV by Sencar mice inoculated as infants was intermittent, whereas transmission by Sencar mice inoculated as weanlings was consistent during the first 2 weeks both by direct contact and by exposure to soiled bedding. The longest duration of transmission was 6 weeks among mice inoculated as infants. The results implicate a role for urinary, fecal, and perhaps respiratory excretion of virus, depending on host genotype and route of virus exposure. They also suggest that evaluation of pancreatic and immune function during acute infection is warranted.

Neonates of various inbred strains of mice expressed three susceptibility phenotypes in response to infection with the lymphocyte-specific variant of minute virus of mice (MVMi). MVMi caused asymptomatic infections in C57BL/6 (B6) mice, lethal infections with intestinal hemorrhage in DBA/2 mice, and lethal infections with renal papillary hemorrhage in BALB/c, SWR, SJL, CBA, and C3H (H) mice. Sequential virus titration, histology, in situ hybridization with a full-length MVMi genomic probe, and immunohistochemistry for viral capsid antigen were used to compare the pathogenesis of MVMi infection in B6 and H mice. Peak infectious virus titers in heart, lung, liver, spleen, kidney and intestine did not differ between strains but brains of B6 mice, unlike H mice, were refractory to infection. Lesions in H mice consisted of renal papillary infarcts and accelerated involution of hepatic erythropoietic foci. No lesions were seen in B6 mice. In situ hybridization and immunohistochemistry indicated that three cell types were primary targets of MVMi; endothelium, lymphocytes, and hepatic erythropoietic precursors. Renal papillary infarcts in H mice were associated with virus replication in endothelial nuclei of the vasa recta. In contrast to the parity of infectious virus titers between strains, fewer cells in target organs of B6 mice were labeled with the MVMi probe then were labeled in H mice and fewer cells expressed viral capsid antigen. These results indicate (a) that the allotropic variants of minute virus of mice may be useful tools to dissect molecular mechanisms of parvovirus virulence, (b) that the virulence of MVMi for neonatal mice does not reside in its lymphotropism, and (c) that genetic susceptibility to lethal MVMi infection may result from overproduction of noninfectious virus products.

The duration of infection with rat virus (RV), an autonomous rodent parvovirus, was examined at multiple intervals over 6 months in rats inoculated by the oronasal route at 2 days of age or 4 weeks of age and individually housed after weaning to prevent cross-infection. Infectious virus was recovered by explant culture from 32 of 80 rats inoculated as pups and was detected as late as 6 months after inoculation. Rats inoculated as juveniles developed acute infection, but virus was not detected beyond 7 weeks after inoculation. Tissues from rats in both age groups were surveyed for RV DNA by Southern blotting using a double-stranded DNA probe made from a 1700 bp cloned fragment of RV spanning map units 0.19-0.52. Band patterns representative of acute infection (juvenile rats) were consistent with the replicating form of RV DNA, whereas patterns representative of persistent infection (rats inoculated as pups) were suggestive of defective or non-productive viral replication.

Two day-old athymic (rnu/rnu) and euthymic (rnu/+) rat pups nursing immune or non-immune dams were inoculated oronasally with the Yale strain of rat virus (RV-Y). All athymic and euthymic pups (57/57) from immune dams remained clinically normal, whereas 51 of 66 athymic and euthymic pups from non-immune dams died within 30 days. Infectious RV was detected by explant culture in 12 of 15 surviving pups of both genotypes from non-immune dams 30 days after inoculation, but in none of the 57 surviving pups from immune dams. RV-Y DNA was detected by Southern blotting in kidneys of surviving athymic pups from non-immune dams but was not detected in pups from immune dams. Euthymic pups from immune dams appeared not to produce endogenous antibody to RV after virus challenge, whereas euthymic pups from non-immune dams produced high-titered RV immune serum. Pups of both genotypes given immune serum prior to or with RV were fully protected from disease and persistent infection, whereas pups given immune serum 24 hours after RV were partially protected. These studies show that RV antibody offers significant protection against lethal and persistent RV infection.

Euthymic (SD or outbred rnu/+) and athymic (rnu/rnu) rats were inoculated oronasally or intraperitoneally with the RV-Y strain of rat virus when they were 2 days or 4 weeks old. Clinical signs of infection in athymic infants were similar to those in euthymic infants, but significantly more athymic infants died. Some infants developed anemia and thrombocytopenia. After inoculation of infants. RV-Y was detected in surviving euthymic rats for 7 weeks and in surviving athymic rats for at least 10 weeks. After oronasal inoculation of 4 week-old rats no clinical illness was observed. RV-Y persisted less than 6 weeks in juvenile euthymic rats but at least 12 weeks in athymic juvenile rats. Intraperitoneal inoculation of juveniles resulted in infection for at least 6 weeks. The antibody response of athymic rats to RV-Y was significantly reduced compared to that of euthymic rats. These studies indicate that T cell deficiency increases the severity and duration of RV infection and imply that T cells are required for the full expression of resistance to RV infection. They also suggest that RV-Y induced anemia could serve as a model for human parvovirus-induced anemia.

The pathogenesis of rat virus (RV) infection was studied in random-bred Sprague-Dawley rats after oronasal inoculation of a recent RV isolate designated RV-Yale (RV-Y). RV-Y was pathogenic for rats inoculated as infants (2 days) whereas rats inoculated as juveniles (4 weeks) had asymptomatic infection and no lesions. Rats inoculated as infants developed pantropic infection accompanied by hepatic necrosis, granuloprival cerebellar hypoplasia and hemorrhagic encephalopathy. Virological and serological studies showed that virus could persist in inoculated rats for at least 35 days and for at least 28 days after seroconversion was first detected. Immunohistochemical results indicated that RV-Y infects tissues conducive to virus excretion including kidney and lung. RV-Y also was found in genital tissues of some rats. Athymic juvenile rats inoculated intraperitoneally with RV-Y had a poor humoral immune response and harbored infectious virus for at least 3 weeks, whereas infection in euthymic control rats was detected for 1 week. These studies indicate that RV-Y can persists in the presence of humoral immunity and suggest that transmission of infection could occur for a substantial period after seroconversion. They also suggest that immunodeficient rats have increased susceptibility to persistent infection.

The duration of transmission of rat virus (RV) infection was determined using Sprague-Dawley rats inoculated oronasally as juveniles (4 weeks old) or as infants (2 days old). Contact transmission from rats inoculated as juveniles was detected for 3 weeks, whereas transmission from rats inoculated as infants occurred for 10 weeks. Transmission continued for at least 7 weeks after seroconversion occurred in rats inoculated as infants. Two of three rats that had ceased to transmit infection harbored infectious virus as detected by explantation of kidney. Intrauterine transmission occurred only after pregnant dams were inoculated with large doses of virus and was more efficient when virus was inoculated intravenously than by the oronasal route. Enzyme immunoassay antibody titers to RV in offspring of previously infected dams decreased steadily during the first 13 weeks of life and 27 of 29 offspring tested by immunofluorescence assay at 12 or 13 weeks of age were seronegative. These results indicate that RV was transmitted by rats inoculated as infants for long periods after seroconversion occurred. They also suggest that the offspring of previously-infected dams were not infected. In utero transmission of RV-Y is unlikely to occur after oronasal inoculation unless rats are exposed to large doses of virus.

Evidence of recent infection with human parvovirus B19 (HPV) was found in two patients with early rheumatoid arthritis (RA) and in four patients with acute inflammatory arthritis (IA). Both of the patients with RA but only one of the four patients with IA carried RA associated haplotypes. No evidence of persistent infection with HPV was found, but evidence of past infection with HPV was significantly more common in patients with RA than in controls. The results confirm the arthritogenic potential of HPV and are consistent with the hypothesis that rheumatoid arthritis may develop in a genetically predisposed patient after an arthritogenic insult such as an HPV infection.