The effect of human interferon alpha on replication of different bovine viral diarrhea virus strains.

Bovine viral diarrhea virus (BVDV) exists in two main biotypes: cytopathic (cp) and noncytopathic (ncp). Although some studies were done on the effect of interferon alpha (IFN-α) on BVDV, the effect of exogenous IFN against BVDV biotypes remains unclear. In the present study, we evaluated the comparative effect of exogenous human IFN-α (HuIFN-α) on different BVDV biotypes and genotypes. The results showed that exogenous HuIFN-α greatly inhibited the growth of different BVDV biotypes and genotypes. However, HuINF-α has a significant inhibitory effect on cp biotype compared to ncp one without significant variation between different genotypes. The effect of HuIFN-α on BVDV reached the maximum level at early stages of infection (0-20 h post infection) and increased in a dose-dependent manner (10-500 U/ml). Quantitative real-time RT-PCR was used to evaluate the effect of exogenous HuIFN-α on RNA synthesis of both BVDV biotypes. HuIFN-α reduced RNA production of cp by 4 logs compared to only 2 logs for ncp strains. Additionally, the antiviral effect of IFN-α against both BVDV biotypes seems to be independent of the RNA-dependent protein kinase (PKR) activation as assayed by direct analysis of in vivo phosphorylation of eIF2-α and by 2-aminopurine (2-AP) treatment. Collectively, these results indicated that the exogenous HuIFN-α treatment has an inhibitory effect not only on cp BVDV biotype but also on the ncp BVDV. The antiviral effect of exogenous HuIFN-α was biotype, time, dose but not genotype dependent. PKR has no role in the inhibitory effect suggesting that other IFN-antiviral pathways were involved. Keywords: BVDV biotypes; HuIFN-α; RNA synthesis; PKR-independent.

Porcine reproductive and respiratory syndrome virus infection of gnotobiotic pigs: sites of virus replication and co-localization with MAC-387 staining at 21 days post-infection.

The organ distribution of PRRSV-infected cells in gnotobiotic piglets at 21 days after infection with PRRSV isolate VR-2332 was examined by in situ hybridization. Cells that expressed PRRSV RNA were identified in all tissues examined, including organs not usually characterized as sites of PRRSV infection. PRRSV-infected cells frequently appeared in clusters and were not always associated with microscopic lesions. The expression of PRRSV RNA co-localized with a macrophage monoclonal antibody, MAC-387, in lymph nodes. Some, but not all infected cells stained with MAC-387. The wide distribution of PRRSV-infected cells and co-localization with MAC-387 staining is consistent with the macrophage-tropism of PRRSV and is similar to observations made during persistent infection with other arteriviruses.

The localization of porcine reproductive and respiratory syndrome virus nucleocapsid protein to the nucleolus of infected cells and identification of a potential nucleolar localization signal sequence.

The nucleocapsid (N) protein of porcine reproductive and respiratory syndrome virus (PRRSV) possesses two regions in the N-terminal half of the protein that are enriched in basic amino acids. Presumably, these basic regions are important for packaging the RNA genome within the nucleocapsid of the virus. The PSORT computer program identified the same regions as nuclear localization signal (NLS) sequence motifs. N protein localization to the nucleus of infected MARC-145 and porcine pulmonary macrophages was observed following staining with SDOW-17 and SR-30 anti-N monoclonal antibodies. Furthermore, the co-localization of SR-30 antibody with human ANA-N autoimmune serum identified the nucleolus as the primary site for N protein localization within the nucleus. The localization of the N protein in the absence of infection was studied by following fluorescence in MARC-145 cells transfected with a plasmid, which expressed the nucleocapsid protein fused to an enhanced green fluorescent protein (N-EGFP). Similar to infected cells, N-EGFP localized to the cytoplasm and the nucleolus. Results following the transfection of cells with pEGFP fused to truncated portions of the N gene identified a region containing the second basic stretch of amino acids as the nucleolar localization signal (NoLS) sequence. Another outcome following transfection was the rapid disappearance of cells that expressed high levels of N-EGFP. However, cell death did not correlate with localization of N-EGFP to the nucleolus.

Serological and genotypic characterization of group A rotavirus reassortants from diarrheic calves born to dams vaccinated against rotavirus.

Two strains of bovine rotavirus (BRV), designated strain Nebraska Scottsbluff-1 (NS-1) and NS-2, were isolated from 2 neighboring cow-calf beef cattle ranches where dams had been vaccinated with a commercial vaccine containing group A BRV strain Neonatal Calf Diarrhea Virus (NCDV)-Lincoln (P1:G6). Northern blot hybridizations using whole genomic RNA probes indicated that strains NS-1 and NS-2 had identical group A RNA electrophoretic patterns and were homologous at all gene segments. Strain NS-1 was compared with reference group A BRV strains using serological and genotypic methods. In vitro virus neutralization assays indicated that strain NS-1 was neutralized by a G6-specific neutralizing monoclonal antibody (mAb) and guinea pig hyperimmune serum (GPHS) raised against BRV strain B641 (P5:G6), but not by G10-specific neutralizing mAb or GPHS raised against BRV strain B223 (P11:G10). Nucleic acid hybridization experiments using whole-genomic RNA probes revealed that gene segment 4 of strain NS-1 differed from BRV strains NCDV-Lincoln and B223, but hybridized with strain B641. Conversely, gene segment 5 of strain NS-1 hybridized with BRV strain B223, but not with BRV strains NCDV-Lincoln and B641. A G-specific cDNA probe produced by reverse transcription polymerase chain reaction (RT-PCR) amplification of strain NS-1 hybridized specifically only with G6 strains NCDV-Lincoln and B641, but not with G10 strain B223. Co-electrophoresis experiments using strains NS-1, B641, and B223 further confirmed these results, suggesting that strain NS-1 was a naturally-occurring reassortant BRV between strains B641 and B223. Taken together these results indicated that a naturally-occurring group A BRV reassortant with a P gene different from the vaccine virus was responsible for the diarrheal syndrome observed on both ranches. Results from this study also indicate the existence of at least 2 different gene segments 5 among group A BRV infecting cattle.

Serum immune responses to the proteins of porcine reproductive and respiratory syndrome (PRRS) virus.

The antibody responses of pigs to porcine reproductive and respiratory syndrome virus (isolate VR-2332) were evaluated by indirect immunofluorescence, virus neutralization, and immunoblotting. All pigs in each group were positive by indirect immunofluorescence 14-21 days postexposure (DPE), and antibodies to specific viral proteins (15, 19 or 26 kD) were initially demonstrated by immunoblotting at 7-21 days DPE. Neutralizing antibodies were detected in only 2 pigs that were inoculated intranasally and given additional parenteral injections with adjuvant. These antibodies appeared much later, 51-70 DPE, than did antibodies detected by indirect immunofluorescence. The titer of the neutralizing antibodies increased until 127 DPE, after which the titers decreased, and 1 animal became seronegative for neutralizing antibody by 262 DPE.

Detection and duration of porcine reproductive and respiratory syndrome virus in semen, serum, peripheral blood mononuclear cells, and tissues from Yorkshire, Hampshire, and Landrace boars.

Because transmission of porcine reproductive and respiratory syndrome virus (PRRSV) can occur through boar semen, it is important to identify persistently infected boars. However, even for boars given the same PRRSV strain and dose, variability in the duration of viral shedding in semen has been observed, suggesting that host factors are involved in PRRSV persistence. To determine whether there are host genetic factors, particularly litter and breed differences related to the persistence of PRRSV, 3 litters from 3 purebred swine breeds were used for this study. It was also determined whether PRRSV could be detected for a longer period of time in serum, semen, or peripheral blood mononuclear cells (PBMC) and if PRRSV could still be detected in tissues after these antemortem specimens were PRRSV negative for a minimum of 2-3 weeks. Three Hampshire, 3 Yorkshire, and 2 Landrace PRRSV-naive boars were obtained and inoculated intranasally with a wild-type PRRSV isolate (SD-23983). All boars within each breed were from the same litter, and litters were within 9 days of age. Serum and PBMC were collected twice weekly from each boar and analyzed for the presence of PRRSV by virus isolation and the polymerase chain reaction (PCR). Serum was also used to obtain virus neutralization titers and enzyme-linked immunosorbent assay S/P values. Semen was collected twice weekly from 7 of 8 boars and analyzed by PCR. After all specimens were PRRSV negative for a minimum of 2-3 weeks, each boar was euthanized, and 21 tissues plus saliva, serum, feces, and urine were collected. All postmortem specimens were evaluated by virus isolation. Specimens that were PRRSV negative by virus isolation were then evaluated by PCR. The mean number of days (+/-SD) for the duration of PRRSV shedding in semen was 51+/-26.9 days, 7.5+/-4.9 days, and 28.3+/-17.5 days for Landrace, Yorkshire, and Hampshire boars, respectively. Because of small sample sizes and large SDs, the differences in duration of PRRSV shedding in semen between breeds were not considered significant. However, the trend suggested that Yorkshire boars were more resistant to PRRSV shedding in semen than were Landrace boars, requiring further investigation using a larger numbers of boars. PRRSV was detected for a longer period in semen than in serum or PBMC in 4 of 7 boars. Viremia could be detected for a longer period in serum than in PBMC in 6 of 8 boars. After a minimum of 2-3 weeks of PRRSV-negative serum, semen, and PBMC, PRRSV could still be detected in the tonsil of 3 of 8 boars by virus isolation, indicating that boars still harbor PRRSV within the tonsil even though antemortem specimens are PRRSV negative.

Relative prevalence of typical and atypical strains among rotaviruses from diarrheic pigs in conventional swine herds.

Polyacrylamide gel electrophoresis was conducted on genomic RNA extracted from rotaviruses detected in diarrheic pigs from conventional swine herds. Ninety samples contained sufficient virus for RNA band visualization and genome classification. Genome profiles were characteristic of typical group A rotaviruses in 67.8% of the 90 samples, of group B rotaviruses in 10.0%, and of group C rotaviruses in 11.1%. In 11.1% of the samples, the presence of more than 11 bands suggested concurrent infection with more than 1 strain of rotavirus. In infections among nursing pigs, 76.4% were group A rotaviruses, 7.4% were group B, 7.4% were group C, and 8.8% were coinfections. In infections among weaned pigs, 40.9% were group A, 18.2% were group B, 22.7% were group C, and 18.2% were coinfections. Coelectrophoresis with prototype OSU and Gottfried strains revealed a great diversity in electropherotype among field strains of rotavirus.

Experimental porcine reproductive and respiratory syndrome virus infection in one-, four-, and 10-week-old pigs.

One-, 4-, and 10-week-old pigs were exposed to porcine reproductive and respiratory syndrome virus (PRRSV) to determine the effect of age on clinical signs, hematologic alterations, the onset and duration of viremia, routes of virus shedding, antibody production, and microscopic lesions produced by PRRSV isolate ATCC VR-2332. The response to PRRSV infection was similar among age groups. Fever, usually prolonged, and a marked dyspnea with cutaneous erythema when restrained for sample collection were the most consistent clinical signs. Prolonged periocular edema was unique to the 1-week-old pigs. The white blood cell count was decreased on day 4 postexposure (PE) due to decreases in neutrophils and lymphocytes. The virus was isolated from buffy coats at day 1 PE and was isolated from serum, buffy coat, or plasma at each sample collection period through the end of the trial (day 28 PE). Virus was most consistently isolated from lung, lymph node, spleen, and tonsil on day 7 PE and exclusively from lymph node, spleen, and tonsil on day 28 PE. Virus was infrequently isolated from urine and fecal and nasal swabs. Consistent microscopic changes in all age groups included interstitial pneumonia and lymph node hypertrophy and hyperplasia on days 7 and 28 PE, lymph node necrosis on day 7 PE, and subacute mononuclear myocarditis on day 28 PE. Findings presented here indicate that interstitial pneumonia, lymphoid necrosis, and mononuclear myocarditis are characteristic lesions of PRRSV isolate ATCC VR-2332 infection in 1-, 4-, and 10-week-old pigs.

Characterization of swine infertility and respiratory syndrome (SIRS) virus (isolate ATCC VR-2332).

The characterization of an isolate of swine infertility and respiratory syndrome (SIRS) virus (ATCC VR-2332) is reported. A commercial cell line (CL2621) was used for the propagation of the virus for all assays. Laboratory studies indicate that this isolate is a fastidious, nonhemagglutinating, enveloped RNA virus. Cesium chloride-purified virions visualized by electron microscopy were spherical particles with an average diameter of 62 nm (range: 48-83 nm) and a 25-30 nm core surrounded by an envelope. Virus replication was restricted to the cytoplasm, as demonstrated by immunofluorescence. The virus did not react serologically with antisera to several common porcine viruses or with antisera to known viruses in the alphavirus, rubivirus, pestivirus, and ungrouped lactic dehydrogenase virus genera of the Togaviridae. However, convalescent sow sera and rabbit hyperimmune sera neutralized the SIRS virus at titers of 1:256 and 1:512, respectively. The virus was stable at 4 and -70 C, but was labile at 37 and 56 C. The properties of this isolate of SIRS virus resemble those of the family Togaviridae but do not match the described genera.

Isolation of swine infertility and respiratory syndrome virus (isolate ATCC VR-2332) in North America and experimental reproduction of the disease in gnotobiotic pigs.

A recent epizootic of swine infertility and respiratory syndrome (SIRS) in a Minnesota swine herd was investigated. Examination of a sow, neonatal piglets, and stillborn fetuses obtained during the epizootic from the affected herd revealed interstitial pneumonitis, lymphomononuclear encephalitis, and lymphomononuclear myocarditis in the piglets and focal vasculitis in the brain of the sow. Fetuses did not have microscopic lesions. No cause for the infertility and respiratory syndrome was determined. Therefore, attempts were made to experimentally reproduce the disease. Eleven 3-day-old gnotobiotic piglets exposed intranasally to tissue homogenates of piglets from the epizootic became inappetent and febrile by 2-4 days postexposure and had interstitial pneumonitis and encephalitis similar to that seen in the field outbreak. After 2 blind passages in gnotobiotic piglets, tissue homogenates were cultured on continuous cell line CL2621, and a cytopathic virus (ATCC VR-2332), provisionally named SIRS virus, was isolated. Gnotobiotic piglets exposed intranasally to the SIRS virus developed clinical signs and microscopic lesions that were the same as those in piglets exposed to the tissue homogenates, and the virus was reisolated from their lungs. This is the first isolate of SIRS virus in the United States that fulfills Koch's postulates in producing the respiratory form of the disease in gnotobiotic piglets and the first report of isolation and propagation of the virus on a continuous cell line (CL2621). The virus is designated as American Type Culture Collection VR-2332.

Persistence of porcine reproductive and respiratory syndrome virus in serum and semen of adult boars.

Four seronegative adult boars were intranasally inoculated with porcine reproductive and respiratory syndrome virus (PRRSV) isolate VR-2332. Serum and semen were collected 2-3 times weekly for over 100 days postinoculation (DPI). Serum samples were assayed for PRRSV by virus isolation (VI) and a polymerase chain reaction (PCR) and screened for antibodies to PRRSV using the indirect fluorescent antibody (IFA) and virus neutralization (VN) tests. Semen was assayed for PRRSV RNA by PCR. Virus and viral RNA was detected in the serum of all boars within 1 DPI by Vi and/or PCR. However, VI results indicated that viremia was transient and occurred from 1 to 9 DPI. Viral RNA was detected in serum from 1 to 31 DPI. In the acute stage of the infection, PRRSV RNA was detected in serum by PCR prior to the presence of viral RNA in semen. The PRRSV RNA was detected in semen as early as 3 DPI and persisted for 25 DPI in 2 of the boars and 56 and 92 DPI in the remaining 2 boars. Detection of PRRSV RNA in semen occurred 2-8 and 28-35 days prior to the detection of antibodies by IFA and VN, respectively. PRRSV was isolated from the bulbourethral gland of the boar that shed viral RNA in semen for 92 DPI. These results suggest that PRRSV RNA can be detected by PCR in boar serum and semen, and may persist for variable periods of time. Viremia and the serologic status of the boar are not adequate indicators of when PRRSV or PRRSV RNA is being shed in the semen. Preliminary findings also indicated that neither shipping stress nor reinoculation with homologous PRRSV resulted in viremia or viral RNA shedding in semen.

Antigenic comparison of Canadian and US isolates of porcine reproductive and respiratory syndrome virus using monoclonal antibodies to the nucleocapsid protein.

Fifteen Canadian field isolates of porcine reproductive and respiratory syndrome (PRRS) virus from Quebec and Ontario were compared with 5 US PRRS virus (PRRSV) isolates and with the European Lelystad isolate using monoclonal antibodies (MAbs) SDOW17, EP147, and VO17 directed to the 15-kDa nucleocapsid protein of PRRSV. All Canadian and US isolates tested by indirect immunofluorescence were recognized by the 3 MAbs, and individual titers of MAbs towards Canadian and US PRRSV isolates were similar as well. In contrast, the Lelystad virus isolate reacted only with the SDOW17 MAb and showed no reactivity with either EP147 or VO17. The reactivity pattern with these MAbs suggests that the Canadian isolates of PRRSV tested are antigenically similar to US isolates of PRRSV, and that these North American isolates share highly conserved epitopes on the 15-kDa nucleocapsid protein that clearly differentiate them from the European Lelystad virus isolate.

Comparative virulence of two porcine group-A rotavirus isolates in gnotobiotic pigs.

The virulence of 2 porcine group-A rotavirus isolates was compared. Forty hysterotomy-derived 3-day-old gnotobiotic pigs were inoculated orally with 2 ml of intestinal homogenate containing either the Ohio State University (OSU) or the South Dakota State University (SDSU) strain of porcine rotavirus or were inoculated with medium only. Clinical signs of disease, body weight, distribution of viral antigen, fecal excretion of virus, and histologic lesions (observed by light and scanning electron microscopy) were determined. Morphometric measurements of villi and crypts were made. In pigs inoculated with OSU or SDSU strains, diarrhea began at postinoculation hours (PIH) 19 to 48 and PIH 24 to 54, respectively. None of the virus-infected pigs died as a consequence of infection and all had similar clinical signs of disease, body weight changes, and virus-shedding patterns, regardless of the strain of rotavirus with which they were infected. Microscopic findings in the small intestine of virus-infected pigs were similar, except that the SDSU strain caused more severe villus atrophy and villus fusion in the duodenum at PIH 72 and 168 than was associated with the OSU strain. Viral antigen in the small intestine of pigs infected with either virus was observed by use of immunofluorescence at PIH 24 and 72, but was seldom seen at PIH 168.

Effects of a modified-live virus vaccine against porcine reproductive and respiratory syndrome in boars.

OBJECTIVES: To determine whether vaccine virus is found in serum and semen of vaccinated boars, whether vaccination prevents subsequent shedding of wild-type virus after challenge exposure, and whether semen and blood variables are altered after vaccination or challenge exposure with wild-type virus, or both. DESIGN: Throughout the 50-day postvaccination period, serum and semen from exposed boars were evaluated for the presence of porcine reproductive and respiratory syndrome virus (PRRSV). All boars were then challenge-exposed with PRRSV isolate VR-2332 and evaluated for an additional 27 days. Semen quality variables, serostatus, and blood variables were monitored. ANIMALS: 7 PRRSV-seronegative adult boars. PROCEDURE: Semen was collected 3 times weekly and evaluated by use of a nested reverse-transcriptase polymerase chain reaction for detection of PRRSV RNA. Serum was obtained weekly and evaluated by nested reverse-transcriptase polymerase chain reaction, virus isolation, and PRRSV ELISA. Semen quality variables were evaluated 3 times weekly, and CBC was performed weekly. RESULTS: Vaccine virus was shed in the semen of all vaccinated boars, but shedding was of shorter duration in 4 of 5 vaccinated boars than that generally observed after exposure to wild-type virus. After challenge exposure, shedding of wild-type virus in semen was shortened or eliminated in 4 of 5 vaccinated boars. Percentage of forward movement and normal spermatozoal morphology and motility were significantly reduced in vaccinated boars after challenge exposure. CONCLUSIONS: Vaccine virus was shed in semen of vaccinated boars, but vaccination generally reduced or eliminated shedding of wild-type PRRSV after challenge exposure. Semen quality appeared to be less than optimal, particularly after vaccination and subsequent challenge exposure with wild-type virus. CLINICAL RELEVANCE: Extra-label use of the PRRSV vaccine in boars remains controversial because some boars may still shed wild-type virus in semen after challenge exposure at postvaccination day 50. Semen quality also appeared to be altered after vaccination and subsequent challenge exposure.

Comparison of a commercial enzyme-linked immunosorbent assay with electron microscopy, fluorescent antibody, and virus isolation for the detection of bovine and porcine rotavirus.

The purpose in this study was to compare the sensitivity of a commercial enzyme-linked immunosorbent assay (ELISA) with electron microscopy (EM), fluorescent antibody (FA), and virus isolation (VI) for the detection of bovine and porcine rotavirus (RV). Seventy-three bovine and 116 porcine accessions were evaluated by 1 or all 4 diagnostic tests, where suitable specimens were available. For the bovine samples, agreement was 33% between FA and EM, 33% between FA and ELISA, and 92% between EM and ELISA. For the porcine samples, agreement was 79% between EM and FA, 72% between EM and ELISA, and 82% between ELISA and FA. Virus was isolated from 68% and 41% of the bovine and porcine fecal samples, respectively. Commercial ELISA was as sensitive as EM, but was more sensitive than FA or VI for the detection of RV in bovine feces. Electron microscopy was more sensitive than FA, ELISA, or VI for detection of RV in porcine feces. The ELISA was an advantageous alternative to the conventional methods of EM, FA, and VI for the diagnosis of RV in calf feces, but not for porcine feces.

Production and characterization of VP4/VP7 reassortant swine rotaviruses derived from Gottfried and OSU parental strains.

The ability of viral glycoproteins (VP) VP4/VP7 reassortant swine rotaviruses (RV) to induce cross-neutralizing antibody against parental serotypes was investigated in guinea pigs. Using selective culture conditions, we produced 10 reassortant viruses that contained gene segment 4 of the OSU RV strain and gene segment 9 of the Gottfried RV strain. These reassortant RV grew to high titer in cell culture and were neutralized by monospecific antisera against both parental RV strains. The reassortant RV were chemically inactivated with binary ethylenimine, adjuvanted with aluminum hydroxide, and used to produce antisera in guinea pigs. The hyperimmune antisera had high neutralization titer against both parent RV strains. These results indicate that several of the reassortant RV may be capable of inducing neutralizing antibodies to VP4 and VP7 and may have future use as bivalent vaccine strains.

Combined rotavirus and K99 Escherichia coli infection in gnotobiotic pigs.

Fifty nine 3-day-old gnotobiotic pigs were randomly assigned to 4 experimental groups: 14 pigs were orally inoculated with rotavirus (RV), 14 were orally inoculated with enterotoxigenic Escherichia coli (ETEC), 18 were orally inoculated with both agents, and 13 were controls. Pigs inoculated with RV plus ETEC were given the RV inoculum at 3 days of age and then, 24 hours later, were given the ETEC inoculum. Three pigs inoculated only with RV, 3 pigs inoculated only with ETEC, 4 pigs inoculated with RV plus ETEC, and 3 pigs in the control group were euthanatized at 5 and 7 days of age. Two pigs in each of the 4 experimental groups also were euthanatized at 9 days of age. Intestinal segments from 6 sites in the small intestine were examined by virologic, bacteriologic, and histologic procedures. For 10 days after inoculation, the remaining pigs in each group were observed clinically to monitor severity and duration of diarrhea, mortality, and shedding of RV or ETEC. Pigs inoculated with the combined RV plus ETEC inoculum developed more severe diarrhea, compared with pigs inoculated with the single agents; all dually inoculated pigs died between 3 and 6 days after inoculation. There was no mortality in pigs inoculated with either RV or ETEC. Lesions were restricted to the small intestine in pigs inoculated with RV plus ETEC and in pigs inoculated with RV or ETEC. There was no difference in the severity of the villus atrophy between the dually inoculated pigs and pigs inoculated only with RV.(ABSTRACT TRUNCATED AT 250 WORDS)

Experimental reproduction of swine infertility and respiratory syndrome in pregnant sows.

The purpose of this study was to experimentally reproduce swine infertility and respiratory syndrome (SIRS). Six multiparous sows were intranasally inoculated at 93 days of gestation with lung homogenates from clinically affected pigs, and 3 additional sows were similarly inoculated with a virus isolated in cell culture from the lung homogenate (SIRS virus, isolate ATCC VR-2332). Inoculated sows developed transient anorexia, farrowed up to 7 days prematurely, and delivered a mean of 5.8 live pigs and 6.0 dead fetuses/litter. Clinical signs of disease were not observed in 3 sham-inoculated control sows that delivered a mean of 12.7 live pigs and 0.3 stillborn fetuses/litter. The SIRS virus was isolated from 50 of 76 live-born and stillborn fetuses from the 9 infected litters. Virus was not isolated from 26 autolyzed fetuses or 15 control pigs. Six of 9 inoculated sows developed neutralizing antibodies to SIRS virus. The reproductive effects found in these experiments were identical to those found in field cases. On the basis of our findings, virus isolate ATCC VR-2332 causes the reproductive failure associated with SIRS.