Effect of vaccination with a modified-live porcine reproductive and respiratory syndrome virus vaccine on dynamics of homologous viral infection in pigs

Jean Paul Cano Swine Disease Eradication Center, College of Veterinary Medicine, University of Minnesota, Saint Paul, MN 55108.

Search for other papers by Jean Paul Cano in
Current site
Google Scholar
PubMed
Close
 DVM
,
Scott A. Dee Swine Disease Eradication Center, College of Veterinary Medicine, University of Minnesota, Saint Paul, MN 55108.

Search for other papers by Scott A. Dee in
Current site
Google Scholar
PubMed
Close
 DVM, PhD
,
Michael P. Murtaugh Department of Veterinary and Biomedical Sciences, College of Veterinary Medicine, University of Minnesota, Saint Paul, MN 55108.

Search for other papers by Michael P. Murtaugh in
Current site
Google Scholar
PubMed
Close
 PhD
,
Carlos A. Trincado Swine Disease Eradication Center, College of Veterinary Medicine, University of Minnesota, Saint Paul, MN 55108.

Search for other papers by Carlos A. Trincado in
Current site
Google Scholar
PubMed
Close
 DVM, MS
, and
Carlos B. Pijoan Swine Disease Eradication Center, College of Veterinary Medicine, University of Minnesota, Saint Paul, MN 55108.

Search for other papers by Carlos B. Pijoan in
Current site
Google Scholar
PubMed
Close
 DVM, PhD†

Abstract

Objective—To determine effects of vaccination protocols with modified-live porcine reproductive and respiratory syndrome virus (PRRSV) vaccine on persistence and transmission of virus in pigs infected with a homologous isolate and determine clinical and virologic responses following heterologous viral challenge.

Animals—Four hundred forty 6- to 8-week-old PRRSV-naïve pigs.

Procedures—Pigs were allocated into 5 groups. Groups A to D were inoculated with wild-type PRRSV VR2332. Group A (positive control pigs) received PRRSV only. Groups B, C, and D received modified-live PRRSV vaccine (1, 2, or 3 doses). Group E served as a negative control group. To evaluate viral transmission, sentinel pigs were introduced into each group at intervals from 37 to 67, 67 to 97, and 97 to 127 days postinoculation (DPI). To evaluate persistence, pigs were euthanized at 37, 67, 97, or 127 DPI. To assess clinical and virologic response after challenge, selected pigs from each group were inoculated at 98 DPI with a heterologous isolate (PRRSV MN-184).

Results—Mass vaccination significantly reduced the number of persistently infected pigs at 127 DPI. Vaccination did not eliminate wild-type PRRSV; administration of 2 or 3 doses of modified-live virus vaccine reduced viral shedding after 97 DPI. Previous exposure to wild-type and vaccine virus reduced clinical signs and enhanced growth following heterologous challenge but did not prevent infection.

Conclusions and Clinical Relevance—Findings suggest that therapeutic vaccination may help to reduce economic losses of PRRSV caused by infection; further studies to define the role of modified-live virus vaccines in control-eradication programs are needed.

Abstract

Objective—To determine effects of vaccination protocols with modified-live porcine reproductive and respiratory syndrome virus (PRRSV) vaccine on persistence and transmission of virus in pigs infected with a homologous isolate and determine clinical and virologic responses following heterologous viral challenge.

Animals—Four hundred forty 6- to 8-week-old PRRSV-naïve pigs.

Procedures—Pigs were allocated into 5 groups. Groups A to D were inoculated with wild-type PRRSV VR2332. Group A (positive control pigs) received PRRSV only. Groups B, C, and D received modified-live PRRSV vaccine (1, 2, or 3 doses). Group E served as a negative control group. To evaluate viral transmission, sentinel pigs were introduced into each group at intervals from 37 to 67, 67 to 97, and 97 to 127 days postinoculation (DPI). To evaluate persistence, pigs were euthanized at 37, 67, 97, or 127 DPI. To assess clinical and virologic response after challenge, selected pigs from each group were inoculated at 98 DPI with a heterologous isolate (PRRSV MN-184).

Results—Mass vaccination significantly reduced the number of persistently infected pigs at 127 DPI. Vaccination did not eliminate wild-type PRRSV; administration of 2 or 3 doses of modified-live virus vaccine reduced viral shedding after 97 DPI. Previous exposure to wild-type and vaccine virus reduced clinical signs and enhanced growth following heterologous challenge but did not prevent infection.

Conclusions and Clinical Relevance—Findings suggest that therapeutic vaccination may help to reduce economic losses of PRRSV caused by infection; further studies to define the role of modified-live virus vaccines in control-eradication programs are needed.

Porcine reproductive and respiratory syndrome has been estimated to cost the US swine industry 560 million dollars in losses each year.1 Results of the same study1 indicate that 88% of the total cost of PRRSV infections in the United States is attributable to increased mortality rates and decreased growth performance in postweaning pigs, whereas the impact of the disease on breeding herds represents only 12% of the total cost. Since the disease was first reported in 1989,2 anorexia, lethargy, hyperemia of the skin, dyspnea, increase in mortality rates, and reduction in ADG have been the clinical signs consistently described following PRRSV infection in nursery, grower, or finisher pigs.2–4 The PRRSV replicates in pulmonary alveolar macrophages,5 facilitating bacterial coinfections and resulting in cases of streptococcal meningitis, septicemic salmonellosis, Hemophilus parasuis infection, and bacterial bronchopneumonia.6

Porcine reproductive and respiratory syndrome virus is an arterivirus7 that initially replicates in macrophages, establishing nonclinical persistent infections.8 Porcine reproductive and respiratory syndrome virus RNA has been detected in the lymphoid tissues of pigs up to 251 DPI.9 It continuously replicates at a low level10 and can be transmitted to susceptible animals following direct contact with pigs inoculated up to 86 days after infection.11 A known risk factor for transmission is the presence of subpopulations of PRRSV-naïve and PRRSV-infected swine, coexisting within endemically infected herds,12 a problem that is frequently exacerbated by introduction of PRRSV-naïve replacement gilts.13 Techniques such as herd closure,14,15 acclimation of gilts to their new environment,16–18 and mass exposure19–22 have been proposed to eliminate such subpopulations and reduce the risk of PRRSV shedding; however, results have been inconsistent among farms.

Another method to maximize population immunity to PRRSV is vaccination. The induction of both humoral and cell-mediated immune responses has been described following the application of modified-live PRRSV vaccines in pigs.23–25 Although it has been reported that vaccination with MLV provides incomplete heterologous protection against PRRSV infection,26 multiple experiments23,24,27,28 have revealed substantial reduction in lesions and clinical signs in vaccinated pigs following homologous and heterologous PRRSV challenge. By contrast, research on killed PRRSV vaccines has generated contrasting results regarding stimulation of production of neutralizing antibody and cell-mediated immune responses in pigs29 or the absence of these responses following use of killed-virus vaccines.25,30 In commercial conditions, the strategic combination of mass vaccination by use of modified-live PRRSV products with herd closure and unidirectional pig flow has been a successful approach to control and, in some instances, to eliminate PRRSV from swine herds.31–34 Although these field reports provide important information regarding the practical application of PRRSV therapeutic vaccination, the need for specific variables to scientifically validate the technique such as the use of positive and negative control groups; exact knowledge of infection time; and control of other factors such as feeding, genetics, health status, and management motivated us to perform a large-scale experiment controlling for those variables and including the use of valuable diagnostic techniques to understand the effect on infection dynamics after mass vaccination.

The study reported here tested the hypothesis that the use of a modified-live PRRSVvaccine would significantly reduce the persistence and transmission of PRRSV in a population of pigs infected with the homologous isolate. The specific aims of the study were to determine whether 3 protocols of vaccination against PRRSV reduced the proportion of persistently infected pigs in the population, to evaluate the effect of 3 protocols of vaccination on PRRSV transmission to susceptible pigs, and to determine the clinical and virologic response of vaccinated pigs following challenge with a highly virulent heterologous PRRSV isolate.

Materials and Methods

Swine and housing—Three hundred thirty-two 6- to 8-week-old pigs (principals) were obtained from a herd known to be free of PRRSV on the basis of 10 years of diagnostic testing. After arrival, pigs were confirmed to be uninfected by use of a commercial ELISAa and RT-PCR assay.b Pigs were individually identified with ear tags and randomly assigned to 5 groups (A to E) that were housed in separate rooms at the research farm of the University of Minnesota Swine Disease Eradication Center in west central Minnesota. Groups were designated as follows: group A, infected with wild-type PRRSV only (positive control pigs); group B, infected with PRRSV and administered 1 dose of MLV vaccine; group C, infected with PRRSV and administered 2 doses of MLV vaccine; group D, infected with PRRSV and administered 3 doses of MLV vaccine; and group E, sham inoculated only (negative control pigs; Table 1). Groups A, B, C, and D contained 80 pigs each, and group E contained 12 pigs. All pigs were vaccinated on arrival against H parasuis, Erysipelothrix rhusiopathiae,c and Lawsonia intracellularis.d In conjunction with the 332 principal pigs, 108 PRRSV-naïve age-matched sentinel pigs were introduced at designated periods following the initiation of the study. All protocols and procedures of pig management and care were reviewed and approved by the University of Minnesota Institutional Animal Care and Use Committee. Personnel practiced PRRSV-specific biosecurity protocols35 across all study groups throughout the experiment.

Table 1—

Experimental design for a study of effect of a modified-live PRRSV vaccine on the dynamics of homologous viral infection in pig populations.

GroupDPIPeriods of sentinel contact (DPI)
0737679712737-6767-9797-127
AInoculation with PRRSVNAEuthanize-sampleEuthanize-sampleEuthanize-sampleEuthanize-sample all remaining pigs1st sentinel group*2nd sentinel group3rd sentinel group
BInoculation with PRRSVMLV administrationEuthanize-sampleEuthanize-sampleEuthanize-sampleEuthanize-sample all remaining pigs1st sentinel group2nd sentinel group3rd sentinel group
CInoculation with PRRSVMLV administrationMLV administrationEuthanize-sampleEuthanize-sampleEuthanize-sample all remaining pigsNA1st sentinel group2nd sentinel group
DInoculation with PRRSVMLV administrationMLV administrationMLV administrationEuthanize-sampleEuthanize-sample all remaining pigsNANA1st sentinel group
ESham inoculationNAEuthanize-sampleEuthanize-sampleEuthanize-sampleEuthanize-sample all remaining pigs1st sentinel group2nd sentinel group3rd sentinel group

Groups of sentinel pigs were in contact with originally inoculated pigs for 30 days and then removed and euthanized, and samples were collected.

Group A = Pigs infected with wild-type virus infection only (positive control pigs; n = 80). Group B = Pigs infected with wild-type virus infection and administered 1 dose of MLV (n = 80). Group C = Pigs infected with wild-type virus infection and administered 2 doses of MLV (n = 80). Group D = Pigs infected with wild-type virus infection and administered 3 doses of MLV (n = 80). Group E = Negative control pigs (n = 12). NA = Not applicable. Euthanize-sample = Euthanize and collect tissues from 10 originally inoculated pigs to assess persistence of infection.

Infection and vaccination—On day 0, all 320 pigs in groups A, B, C, and D were intranasally inoculated with 2 mL of cell culture fluid containing 104 TCID50/mL of PRRSV VR2332 (cell culture passage 3). At 7 DPI, pigs in groups B, C, and D were vaccinated IM with 2 mL of a homologous modified-live PRRSV vaccine.e Groups C and D were then revaccinated 30 days later (37 DPI), and a final dose was administered to group D at 67 DPI. Pigs in group E were sham inoculated intranasally with 2 mL of sterile saline (0.9% NaCl) solution (Table 1). Modifiedlive PRRSV vaccinee and PRRSV VR2332 GenBank accession numbers are AF066183 and PRU87392, respectively. To monitor the protocols of infection and vaccination, 12 pigs from groups A to D and 6 pigs in group E were randomly selected and serum samples were collected in sterile vacuum tubesf via jugular venipuncture at 0, 7, 14, 37, 67, 97, and 127 DPI.

Assessment of transmission—To determine whether different protocols of vaccination affected virus shedding, groups of 10 individually tagged PRRSV-naïve pigs were introduced 30 days after the last vaccination into each treatment group (Table 1). All sentinel pigs were removed and euthanized, and samples were collected 30 days after introduction. Transmission was defined as either detection of PRRSV nucleic acid in serum or tissues by use of RT-PCR procedures in 1 or more sentinel pigs or detection of anti-PRRSV antibodies as determined by positive results of ELISA that was subsequently confirmed by positive results of indirect fluorescent antibody test36 in 1 or more sentinel pigs.

Assessment of persistence—To evaluate whether different protocols of vaccination affected the proportion of PRRSV persistently infected pigs through time, 10 pigs/group were randomly selected and euthanized, and samples were collected 30 days following the last vaccination (Table 1). The study was terminated, and all remaining pigs were euthanized and sampled at 127 DPI. Serum and tissue samples were tested by use of RT-PCR assay for PRRSV nucleic acid. Tonsil, sternal lymph nodes, and superficial inguinal lymph nodes were collected in the processing plant, stored in separate plastic bags, and transported on ice to the laboratory for testing.

Clinical and virologic response to heterologous challenge—At 97 DPI, 10 randomly selected pigs from every group and 2 additional negative control pigs were transported to the isolation facilities of the College of Veterinary Medicine at the University of Minnesota. Each group was housed in 2 rooms (5 pigs/room) at a density of 1.2 m2/pig. Each isolation room had an independent ventilation system and a slurry pit to prevent cross-contamination of pathogens between rooms. All pigs in groups A, B, C, and D and 10 pigs from group E were intranasally inoculated with 2 mL of cell culture fluid containing PRRSV MN-184 isolate (104 TCID50/mL). Challenge control PRRSV-naïve pigs were labeled as E+. The group of 2 additional negative control pigs was sham inoculated intranasally with 2 mL of sterile saline solution and labeled as E–. The challenge virus, PRRSV MN-184 (GenBank accession No. AY656992), a highly virulent isolate from a farm in southern Minnesota affected with severe reproductive disease and high sow mortality rate in 2001,37 was used at cell culture passage 2. The percentage divergence in the ORF 5 nucleotide sequence between MN-184 and VR2332 or the modified-live PRRSV was 15.5%.37

Blood was collected from all 52 pigs at 0, 7, 14, and 21 DPI to evaluate viremia and antibody response. At 21 DPI, all pigs were euthanized and samples of tonsil, sternal lymph nodes, and superficial inguinal lymph nodes were collected in separate plastic bags. To evaluate the clinical response following heterologous challenge, rectal temperature, appetite, and mortality rate were measured at 0, 2, 4, 7, 10, 14, 17, and 21 DPI. Pigs were also weighed at 0 and 21 DPI, and ADG (g) per group was calculated. Rectal temperature (°C) was measured by the same person every sampling day between 8 and 9 AM. Every morning, the same operator measured the volume of feed still remaining from the previous day in each room and the percentage of reduction in feed intake was estimated (0% to 100%). Study personnel were unaware of group assignments.

Diagnostic testing—The PRRSV antibody response was evaluated by use of a commercial ELISA test.a The presence of PRRSV nucleic acid in serum and tissues was determined by use of RT-PCR assay.b Tissue samples collected at euthanasia were pooled by individual animal and 1 g was placed in 15 mL of lysis bufferg in a sterile plastic tube.h After homogenization with the appropriate equipment,i samples were centrifuged (700 X g for 15 minutes) to remove disrupted cells and debris. Total RNA was extracted and purified with a commercial kitg according to the manufacturer's protocol from 200 μL of serum or 50 μL of the middle layer of homogenized tissue supernatant. The RNA was eluted in 50 μL of water, dried in a vacuum centrifuge,j and rehydrated in 5 μL of water. Every sample was assayed in duplicate by use of 2 μL of the rehydrated sample in a 20-μL RT-PCR assay with primers and probe directed to the ORF 7 region of the North American PRRSV.38 All reactions were conducted in a real-time PCR instrument.k

Extracted RNA from selected tissue samples with positive PCR assay results was submitted to the University of Missouri, Columbia Veterinary Diagnostic Laboratory to sequence the PRRSV ORF 5 region by use of a procedure modified from a published protocol.39 Amino acid sequence comparisons were used to differentiate wild-type from vaccine viruses.

Tissue RNA from pigs in the heterologous challenge phase of the study was tested by use of an isolate-specific PCR assay targeting a nucleotide sequence on the ORF 5 region of PRRSV MN-184. Total RNA was extracted as described. The PCR included the RT-PCR kit,b forward primer (5a–TAACTTAACGATATGTGAGCTGAATG-GCAC–3a), reverse primer (5a–ACACAGTGATCAG-GCCGACC–3a), and probe (6FAM-CTGGCTGAACAA TCATTTTAGTTGGGCAGTGGAGACTTTCGTTATC-TAMRA). A standard curve was developed for the quantitative RT-PCR procedure by preparing 10-fold dilutions of PRRSV MN-184 stock starting at 105 TCID50/mL. Results were reported in number of RNA copies (ie, RNAc) per gram of tissue.

Statistical analysis—The proportions of PRRSV persistently infected pigs were compared among groups within the same DPI by use of the Fisher exact test. The number of PRRSV RNA copies per gram of tissue was log transformed to stabilize the variance prior to analysis. Rectal temperature (°C), ELISA S:P ratio mean, ADG (g) and log10 RNAc per gram of tissue were compared among groups by use of 1-way ANOVA. All analyses were performed with standard software.l A value of P <0.05 was considered significant.

Results

Viremia and antibody response in monitor pigs—Porcine reproductive and respiratory syndrome virus nucleic acid was detected in serum at 7 DPI in 85% of monitor pigs from all inoculated groups (A to D). The proportion of viremic pigs decreased over time up to 67 DPI when all serum samples from monitor pigs in all groups yielded negative results of PCR assays. At 14 DPI, 90% of monitor pigs in all inoculated groups had positive results of ELISA (S: P ratio > 0.4). No significant differences in S:P ratio means were observed among inoculated groups during the first 97 DPI independently of the vaccination protocol; however, at 127 DPI, pigs in groups A and B had significantly (P = 0.008) higher S:P ratio mean values than pigs in groups C and D (Table 2). Negative control pigs (group E) had negative results of ELISAb and PCRa assay throughout the study.

Table 2—

Mean ± SE PRRSV ELISA S:P ratios in pigs on various days after inoculation with PRRSV.

GroupDPI
0714376797127
A0.03 ± 0.01a0.07 ± 0.03a2.00 ± 0.34a2.47 ± 0.28a1.60 ± 0.23a1.11 ± 0.2a1.08 ± 0.16a
B0.02 ± 0.01a0.04 ± 0.01a1.33 ± 0.28a1.91 ± 0.24a1.64 ± 0.2a1.28 ± 0.18a1.04 ± 0.15a
C0.02 ± 0.01a0.06 ± 0.02a1.28 ± 0.18a1.59 ± 0.15a1.19 ± 0.14a0.79 ± 0.1a0.50 ± 0.09b
D0.04 ± 0.01a0.04 ± 0.01a1.29 ± 0.24a1.92 ± 0.23a1.46 ± 0.18a0.90 ± 0.12a0.62 ± 0.08b
E0.01 ± 0a0.02 ± 0a0.02 ± 0.01b0.19 ± 0.01b0.04 ± 0.01b0.02 ± 0.01b-0.02 ± 0.03c

Within each column, values with different superscripts are significantly (P < 0.05) different.

See Table 1 for remainder of key.

Transmission to sentinel pigs—Transmission of PRRSV was detected at 37 to 67 DPI and 67 to 97 DPI in all groups in which sentinel pigs were introduced (Table 3). Between 97 and 127 DPI, sentinel pigs introduced into groups A (positive control pigs) and B became infected as determined by results of PCR assay or ELISA, whereas sentinel pigs in groups C and D remained uninfected.

Table 3—

Proportions of persistently infected pigs and transmission of PRSSV to sentinel pigs.

GroupDPI
376797127
PersistencePersistenceTransmissionPersistenceTransmissionPersistenceTransmission
A9/109/9Yes8/10Yes16/42aYes
B9/109/10Yes5/10aYes5/39bYes
CNT8/10NT5/10aYes4/46bNo
DNTNTNT4/10aNT8/57bNo
E0/60/6No0/6bNo0/5cNo

Persistence = Proportion of persistently infected pigs per number of necropsied pigs at each DPI. Transmission = Determination of infection of the sentinel group (Yes or No). NT = Not tested.

See Tables 1 and 2 for remainder of key.

Proportion of persistently infected pigs—The modified-live PRRSV vaccine did not reduce the proportion of persistently infected pigs at 37, 67, or 97 DPI (Table 3). However, at 127 DPI, the proportion of infected pigs was significantly reduced in vaccinated groups.

Twenty tissue samples that yielded positive results of PCR assay at 127 DPI from groups B, C, and D were submitted for nucleic acid sequencing of the ORF 5 region. Seven samples yielded sufficient product for sequencing. In every sample, the sequence corresponded more closely to that of VR2332 wild-type virus than to that of MLV vaccine.

Clinical and virologic response to MN-184 challenge—Porcine reproductive and respiratory syndrome virus RNA was not detected in serum of previously infected pigs (groups A, B, C, and D) at 7, 14, or 21 days after MN-184 inoculation. Only the challenge controls (group E+) had a detectable viremia: 10 of 10 pigs at 7 DPI, 7 of 10 at 14 DPI, and 1 of 10 at 21 DPI. The mean S:P ratios among previously infected groups (A to D) before MN-184 inoculation were not significantly (P = 0.321) different. Seven days after MN-184 administration, all challenge controls (E+) yielded negative ELISA results. In contrast, previously exposed pigs had increased ELISA S:P ratios by day 7, without significant (P = 0.934) difference across these groups. One week later, every pig in group E+ yielded ELISA positive results and the S:P ratio means were not significantly (P = 0.864) different among all inoculated groups (A to E+). Similar results were obtained 21 days after MN-184 inoculation (P = 0.965). Negative control pigs (E–) had negative results of ELISA and PCR assays during this phase of the study.

Isolate MN-184 was detected in tissue samples of 70% to 100% of the pigs from all challenged groups (A to E+) at 21 DPI. The proportion of infected pigs per group was not significantly affected by previous PRRSV exposure status. No significant (P = 0.366) difference among groups was detected in the mean number of PRRSV RNA copies per gram of tissue (Table 4).

Table 4—

Virologic and clinical variables in pigs after challenge inoculation with PRRSV MN-184.

GroupViremiaSeroconversion*Mean ± SE log10 RNAc/gDeathsin appetiteSE ADG (g)Hyperthermia
A7 DPI3.26 ± 0.3b510 ± 68a2 DPI
B7 DPI3.81 ± 0.1b603 ± 69a2 DPI
C7 DPI3.67 ± 0.2b717 ± 85a2 DPI
D7 DPI.62 ± 0.2b574 ± 110a2 DPI
E++14 DPI3.8 ± 0.3b1 pig died 14 DPI+ (4-16 DPI)208 ± 89b2, 4, 7, 10, and 14 DPI
E-0a810 ± 54a

Significant (P < 0.05) increase in group mean ELISA S:P ratio.

Group mean rectal temperature (°C) was significantly (P < 0.05) different from that of the negative control group on that DPI.

RNAc = RNA copies. + = Detected. — = Not detected.

Within a column, values with different superscripts are significantly (P < 0.05) different.

See Table 1 for remainder of key.

Overall, pigs from groups that had been previously exposed to PRRSV had less severe clinical signs than newly infected pigs, 1 of which died 14 days after MN-184 challenge. It had fever (41.3°C), dyspnea, cyanosis of extremities, lethargy, and anorexia before death. Noncollapsed lungs and enlarged lymph nodes were the only remarkable lesions observed during necropsy. Interstitial pneumonia was detected via histologic examination, and PRRSV RNA was detected in lymph nodes and serum by use of RT-PCR assay. Although appetite was not affected in negative control pigs (E–) or groups A to D, a 5% to 15% mean reduction in feed intake was recorded in group E+ from 4 to 16 days after MN-184 inoculation. Pigs in group E+ also had significantly (P = 0.002) lower ADG than pigs in groups A, B, C, and D. Mean rectal temperatures in all groups were not significantly different before MN-184 inoculation (P = 0.071). Inoculated groups (A to E+) had significantly (P = 0.036) higher rectal temperatures than negative control pigs at 2 DPI; however, between 4 and 14 DPI, pigs in group E+ had significantly higher rectal temperature than previously infected (A to D) or negative control pigs (Table 4).

Discussion

The purpose of the study reported here was to understand the dynamics of endemic PRRSV infection in pig populations following the application of a modified-live PRRSV vaccine as an initial means to determine whether vaccination could be a potential tool in control or eradication programs or both. After inoculation of pigs with PRRSV VR2332, the infection and seroconversion pattern was consistent with previous observations.37,40 Various protocols of repeated vaccination with MLV did not affect the ELISA s:p ratio dynamic from 14 to 97 DPI; however, at 127 DPI, the 2 groups that received multiple vaccinations (C and D) had significantly lower S:P ratios than the positive control (A) and 1-dose group (B). These results are in accordance with previous reports41,42 that the commercial ELISA test detected an antibody response to initial infection but not to single or repeated homologous inoculations. In the present study, repeated vaccination reduced the variation in ELISA results, as reflected in the SE of the S:P ratio mean.

Another observation was that repeated vaccination (2 or 3 doses) reduced the duration of PRRSV transmission to sentinels, perhaps because vaccinated groups had a significantly lower proportion of persistently infected pigs at 127 DPI. Despite the significant effect of the vaccination on viral persistence, the originally inoculated wild-type virus was not eliminated from the pigs in 127 days.

Although the reduction in persistence and duration of transmission by vaccination of infected populations may offer relevant information to improve PRRSV-control strategies such as herd closure and acclimation of gilts, results of the present study need to be interpreted with caution for a number of reasons. A limitation of the study was that the wild-type virus in the population was homologous to the commercially available MLV vaccine, which is not common. Because this experiment was part of a long-term study, the first logical step was to analyze the use of mass vaccination under homologous conditions to determine whether vaccination provided any benefit under optimum conditions. Detection of actively shedding pigs at 127 DPI was unexpected, and therefore, a longer trial may have provided more information. However, given the duration of shedding reported in the literature11 and the standard acclimation period used by the industry,16–18 it was expected that any evidence of vaccination effect on transmission or persistence would have been observed within 127 days. It is possible that performing mass vaccination on a population with low infection prevalence would reveal a larger benefit of mass vaccination than in the present experiment. Most of the prior reports of success of vaccination in controlling PRRSV resulted from a study that combined this strategy with herd closure.31–34 Constant introduction of sentinels in the study, which was necessary to assess transmission over time, may have allowed for reinfection of principal pigs. However, reinfection of pigs with a homologous PRRSV has not been reported.

The final phase of the study attempted to recreate the common situation of external PRRSV reintroduction to endemically infected swine herds to gain knowledge about the clinical and virologic responses against a highly virulent and heterologous challenge. After MN-184 challenge, only the challenge controls had viremia, whereas previously exposed pigs generated a rapid serologic response without evidence of viremia. The MN-184 virus was found in the tissues of nearly all pigs from all groups regardless of the previous PRRSV infection status. A low level of transient viremia may have occurred before the first sampling at 7 DPI because an antibody response and tissue infection were observed. Consistent with partial protection against infection, fewer clinical signs and enhanced growth performance were observed in all groups that were previously exposed to PRRSV.

Under the conditions of this study, the use of a modified-live PRRSV vaccine reduced the proportion of persistently infected pigs and the duration of shedding of wild-type virus in a population of pigs infected with a homologous PRRSV isolate. Previous exposure to PRRSV significantly improved the clinical response against a highly virulent heterologous challenge, but vaccination did not eliminate the wild-type virus from lymphoid tissues or prevent heterologous infection. These results offer a detailed and controlled evaluation of the benefits and risks of the use of PRRSV mass vaccination under controlled field conditions. Although some field experiences have been reported, to the authors' knowledge, large-scale controlled experiments evaluating the effect of therapeutic vaccination on persistence, transmission, and clinical response against a heterologous PRRSV challenge have not been published. Although previous reports31–34 provide valuable information about the potential applications of therapeutic vaccination by interpreting field results, the present experiment included positive and negative control groups; intense diagnosis monitoring via modern diagnosis tools; and strict control of a number of factors such as infection time, feeding, genetics, health status, and management to analyze the results with scientific criteria. Further studies are needed to assess the effect of mass vaccination on PRRSV-infected populations with heterologous isolates under field conditions to further define the role of MLV vaccines in control-eradication programs.

ABBREVIATIONS

PRRSV

Porcine reproductive and respiratory syndrome virus

ADG

Average daily gain

DPI

Days postinoculation

MLV

Modified-live virus

RT

Reverse transcriptase

ORF

Open reading frame

S:P ratio

Sample-to-positive ratio

a.

Herd Chek PRRS Antibody 2XR Test Kit, IDEXX Laboratories, Westbrook, Me.

b.

Taqman RT-PCR kit, Perkin-Elmer Applied Biosystems, Foster City, Calif.

c.

Ingelvac HPE-1, Boehringer Ingelheim Vetmedica Inc, St Joseph, Mo.

d.

Enterisol Ileitis, Boehringer Ingelheim Vetmedica Inc, St Joseph, Mo.

e.

Ingelvac PRRS MLV, Boehringer Ingelheim Vetmedica Inc, St Joseph, Mo.

f.

Becton-Dickinson Vacutainer, Franklin Lakes, NJ.

g.

Nucleospin II kit, BD Biosciences, Palo Alto, Calif.

h.

Falcon tube, Becton-Dickinson, Franklin Park, NJ.

i.

Polytron PT 3100, Kinematica AG, Lucerne, Switzerland.

j.

Savant, Speedvac, GMI Inc, Ramsey, Minn.

k.

ABI 7700, Perkin-Elmer Applied Biosystems, Foster City, Calif.

l.

Statistix 8 Trial Version, Analytical Software, Tallahassee, Fla.

References

  • 1

    Neumann EJ, Kliebenstein JB, Johnson CD, et al. Assessment of the economic impact of porcine reproductive and respiratory syndrome on swine production in the United States. J Am Vet Med Assoc 2005;227:385392.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 2

    Keffaber KK. Reproductive failure of unknown etiology. Am Assoc Swine Pract Newsl 1989;1:110.

  • 3

    Loula T. Mystery pig disease. Agri-Practice 1991;12:2334.

  • 4

    Moore C. Clinical presentation of the mystery swine disease in the growing pig, in Proceedings. Mystery Swine Dis Comm Meet 1990;4149.

    • Search Google Scholar
    • Export Citation
  • 5

    Molitor TW, Xiao J, Choi CS. PRRS virus infection of macrophages: regulation by maturation and activation state, in Proceedings. 25th Annu Meet Am Assoc Swine Pract 1996;563569.

    • Search Google Scholar
    • Export Citation
  • 6

    Zimmerman J, Benfield DA, Murtaugh MP, et al. Porcine reproductive and respiratory syndrome virus (porcine arterivirus). In: Straw BE, Zimmerman J, D'Allaire S, et al, eds.Diseases of swine. 9th ed. Ames, Iowa: Blackwell Publishing, 2006;387418.

    • Search Google Scholar
    • Export Citation
  • 7

    Cavanagh D. Nidovirales: a new order comprising Coronaviridae and Arteviridae. Arch Virol 1997;142:629633.

  • 8

    Plagemann P, Moenning V. Lactate dehydrogenase-elevating virus, equine arteritis virus, and simian hemorrhagic fever virus: a new group of positive strand RNA viruses. Adv Virus Res 1992;41:99192.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 9

    Wills RW, Doster AR, Galeota JA, et al. Duration of infection and proportion of pigs persistently infected with porcine reproductive and respiratory syndrome virus. J Clin Microbiol 2003;41:5862.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 10

    Allende R, Laegreid WW, Kutish GF, et al. Porcine reproductive and respiratory syndrome virus: description in individual pigs upon experimental infection. J Virol 2000;74:1083410837.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 11

    Bierk MD, Dee SA, Rossow KD, et al. Transmission of porcine reproductive and respiratory syndrome virus from persistently infected sows to contact controls. Can J Vet Res 2001;65:261266.

    • Search Google Scholar
    • Export Citation
  • 12

    Dee SA, Joo HS, Henry S, et al. Detecting subpopulations after PRRS virus infection in large breeding herds using multiple serologic tests. J Swine Health Prod 1995;4:181184.

    • Search Google Scholar
    • Export Citation
  • 13

    Dee SA, Joo HS. Recurrent reproductive failure associated with porcine reproductive and respiratory syndrome in a swine herd. J Am Vet Med Assoc 1994;205:10171018.

    • Search Google Scholar
    • Export Citation
  • 14

    Dee SA, Joo HS, Pijoan C. Controlling the spread of PRRS virus in the breeding herd through management of the gilt pool. J Swine Health Prod 1995;3:6469.

    • Search Google Scholar
    • Export Citation
  • 15

    Torremorell M, Henry S, Christianson WT. Eradication using herd closure. In:Zimmerman J, Yoon KJ, ed.The PRRS compendium. 2nd ed.Des Moines: National Pork Board, 2003;157161.

    • Search Google Scholar
    • Export Citation
  • 16

    Dee SA, Philips RE. Use of polymerase chain reaction (PCR) to detect vertical transmission of porcine reproductive and respiratory syndrome virus (PRRSV) in piglets from gilt litters. J Swine Health Prod 1997;7 (5):237239.

    • Search Google Scholar
    • Export Citation
  • 17

    Batista L, Pijoan C, Torremorel M. Experimental injection of gilts with porcine reproductive and respiratory syndrome virus (PRRSV) during acclimatization. J Swine Health Prod 2002;10:147150.

    • Search Google Scholar
    • Export Citation
  • 18

    Fano E, Olea L, Pijoan C. Eradication of porcine reproductive and respiratory syndrome virus by serum inoculation of naive gilts. Can J Vet Res 2005;69:7174.

    • Search Google Scholar
    • Export Citation
  • 19

    Desrosiers R, Boutin M. An attempt to eradicate porcine reproductive and respiratory syndrome virus (PRRSV) after an outbreak in a breeding herd: eradication strategy and persistence of antibody titers in sows. J Swine Health Prod 2002;10:2325.

    • Search Google Scholar
    • Export Citation
  • 20

    Batista L, Pijoan C, Baidoo S. Eradication of porcine reproductive and respiratory syndrome virus (PRRSV) by serum inoculation with the homologous PRRSV strain, in Proceedings. 18th Int Pig Vet Soc Congr 2004;815.

    • Search Google Scholar
    • Export Citation
  • 21

    Wagner M. PRRS planned exposure: a summary of results, in Proceedings. 35th Annu Meet Am Assoc Swine Vet 2005;329336.

  • 22

    FitzSimmons MA. Principles of dealing with PRRS, in Proceedings. 35th Annu Meet Am Assoc Swine Vet 2005;319321.

  • 23

    Charerntanakul W, Platt R, Johnson W, et al. Immune responses and protection by vaccine and various vaccine adjuvant candidates to virulent porcine reproductive and respiratory syndrome virus. Vet Immunol Immunopathol 2006;109:99115.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 24

    Foss DL, Zillox MJ, Meier WA, et al. Adjuvant danger signals increase the immune response to porcine reproductive and respiratory syndrome virus. Viral Immunol 2002;15:557566.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 25

    Meier WA, Galeota J, Osorio FA, et al. Gradual development of the interferon-G response of swine to porcine reproductive and respiratory syndrome virus infection or vaccination. Virology 2003;309:1831.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 26

    Labarque G, Van Gucht S, Van Reeth K, et al. Respiratory tract protection upon challenge of pigs vaccinated with attenuated porcine reproductive and respiratory syndrome virus vaccines. Vet Microbiol 2003;95:187197.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 27

    Mengeling WL, Lager KM, Vorwald AC, et al. Comparative safety and efficacy of attenuated single-strain and multi-strain vaccines for porcine reproductive and respiratory syndrome. Vet Microbiol 2003;93:2538.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 28

    Opriessnig T, Pallarés FJ, Nilubol D, et al. Genomic homology of ORF 5 gene sequence between modified live vaccine virus and porcine reproductive and respiratory syndrome virus challenge isolates is not predictive of vaccine efficacy. J Swine Health Prod 2005;13:246253.

    • Search Google Scholar
    • Export Citation
  • 29

    Nilubol D, Platt KB, Halbur PG, et al. The effect of a killed porcine reproductive and respiratory syndrome virus (PRRSV) vaccine treatment on virus shedding in previously PRRSV infected pigs. Vet Microbiol 2004;102:1118.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 30

    Plana-Durán J, Bastons M, Urniza A, et al. Efficacy of an inactivated vaccine for prevention of reproductive failure induced by porcine reproductive and respiratory syndrome virus. Vet Microbiol 1997;55:361370.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 31

    Dee SA, Phillips R. Using vaccination and unidirectional pig flow to control PRRSV transmission. J Swine Health Prod 1998;6:2125.

  • 32

    Gillispie TG, Carroll AL. Methods of control and elimination of porcine reproductive and respiratory syndrome virus using modified live vaccine in a two-site production system. J Swine Health Prod 2003;11:291295.

    • Search Google Scholar
    • Export Citation
  • 33

    Ridremont B, Lebret A. Validation of a mass vaccination protocol with a PRRS modified live vaccine to stabilize French breeding herds, in Proceedings. 19th Int Pig Vet Soc Congr 2006;2: 43.

    • Search Google Scholar
    • Export Citation
  • 34

    Rodriguez E, Escuder M, Riera P, et al. A two-year study of the evolution of PRRS infection dynamics in a herd vaccinated with a modified live European-type strain, in Proceedings. 19th Int Pig Vet Soc Congr 2006;2: 51.

    • Search Google Scholar
    • Export Citation
  • 35

    Otake S, Dee SA, Rossow KD, et al. Transmission of porcine reproductive and respiratory syndrome virus by fomites (boots and coveralls). J Swine Health Prod 2002;10:5965.

    • Search Google Scholar
    • Export Citation
  • 36

    Yoon IJ, Joo HS, Christianson WT, et al. An indirect fluorescent antibody test for the detection of antibody of swine infertility and respiratory syndrome virus in swine sera. J Vet Diagn Invest 1992;4:144147.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 37

    Johnson W, Roof M, Vaughn E, et al. Pathogenic and humoral immune responses to porcine reproductive and respiratory syndrome virus (PRRSV) are related to viral load in acute infection. Vet Immunol Immunopathol 2004;102:233247.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 38

    Schurrer JA, Dee SA, Moon RD, et al. Retention of ingested porcine reproductive and respiratory syndrome virus in houseflies. Am J Vet Res 2005;66:15171525.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 39

    Pardo ID, Johnson GC, Kleiboeker SB. Phylogenetic characterization of canine distemper viruses detected in naturally infected dogs in North America. J Clin Microbiol 2005;43:50095017.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 40

    Opriessnig T, Halbur PG, Yoon KJ, et al. Comparison of molecular and biological characteristics of a modified live porcine reproductive and respiratory syndrome virus (PRRSV) vaccine (Ingelvac PRRS MLV), the parent strain of the vaccine (ATCC VR2332), ATCC VR2385, and two recent field isolates of PRRSV. J Virol 2002;76:1183711844.

    • Search Google Scholar
    • Export Citation
  • 41

    McCaw M, Murtaugh M, Laster S, et al. PRRSV rORF specific antibody responses following repeated homologous wild-type virus challenges, in Proceedings. 18th Int Pig Vet Soc Congr 2004;1: 36.

    • Search Google Scholar
    • Export Citation
  • 42

    Foss DL, Zilliox MJ, Meier W, et al. Adjuvant danger signals increase the immune response to porcine reproductive and respiratory syndrome virus. Viral Immunol 2002;15:557566.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 1

    Neumann EJ, Kliebenstein JB, Johnson CD, et al. Assessment of the economic impact of porcine reproductive and respiratory syndrome on swine production in the United States. J Am Vet Med Assoc 2005;227:385392.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 2

    Keffaber KK. Reproductive failure of unknown etiology. Am Assoc Swine Pract Newsl 1989;1:110.

  • 3

    Loula T. Mystery pig disease. Agri-Practice 1991;12:2334.

  • 4

    Moore C. Clinical presentation of the mystery swine disease in the growing pig, in Proceedings. Mystery Swine Dis Comm Meet 1990;4149.

    • Search Google Scholar
    • Export Citation
  • 5

    Molitor TW, Xiao J, Choi CS. PRRS virus infection of macrophages: regulation by maturation and activation state, in Proceedings. 25th Annu Meet Am Assoc Swine Pract 1996;563569.

    • Search Google Scholar
    • Export Citation
  • 6

    Zimmerman J, Benfield DA, Murtaugh MP, et al. Porcine reproductive and respiratory syndrome virus (porcine arterivirus). In: Straw BE, Zimmerman J, D'Allaire S, et al, eds.Diseases of swine. 9th ed. Ames, Iowa: Blackwell Publishing, 2006;387418.

    • Search Google Scholar
    • Export Citation
  • 7

    Cavanagh D. Nidovirales: a new order comprising Coronaviridae and Arteviridae. Arch Virol 1997;142:629633.

  • 8

    Plagemann P, Moenning V. Lactate dehydrogenase-elevating virus, equine arteritis virus, and simian hemorrhagic fever virus: a new group of positive strand RNA viruses. Adv Virus Res 1992;41:99192.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 9

    Wills RW, Doster AR, Galeota JA, et al. Duration of infection and proportion of pigs persistently infected with porcine reproductive and respiratory syndrome virus. J Clin Microbiol 2003;41:5862.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 10

    Allende R, Laegreid WW, Kutish GF, et al. Porcine reproductive and respiratory syndrome virus: description in individual pigs upon experimental infection. J Virol 2000;74:1083410837.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 11

    Bierk MD, Dee SA, Rossow KD, et al. Transmission of porcine reproductive and respiratory syndrome virus from persistently infected sows to contact controls. Can J Vet Res 2001;65:261266.

    • Search Google Scholar
    • Export Citation
  • 12

    Dee SA, Joo HS, Henry S, et al. Detecting subpopulations after PRRS virus infection in large breeding herds using multiple serologic tests. J Swine Health Prod 1995;4:181184.

    • Search Google Scholar
    • Export Citation
  • 13

    Dee SA, Joo HS. Recurrent reproductive failure associated with porcine reproductive and respiratory syndrome in a swine herd. J Am Vet Med Assoc 1994;205:10171018.

    • Search Google Scholar
    • Export Citation
  • 14

    Dee SA, Joo HS, Pijoan C. Controlling the spread of PRRS virus in the breeding herd through management of the gilt pool. J Swine Health Prod 1995;3:6469.

    • Search Google Scholar
    • Export Citation
  • 15

    Torremorell M, Henry S, Christianson WT. Eradication using herd closure. In:Zimmerman J, Yoon KJ, ed.The PRRS compendium. 2nd ed.Des Moines: National Pork Board, 2003;157161.

    • Search Google Scholar
    • Export Citation
  • 16

    Dee SA, Philips RE. Use of polymerase chain reaction (PCR) to detect vertical transmission of porcine reproductive and respiratory syndrome virus (PRRSV) in piglets from gilt litters. J Swine Health Prod 1997;7 (5):237239.

    • Search Google Scholar
    • Export Citation
  • 17

    Batista L, Pijoan C, Torremorel M. Experimental injection of gilts with porcine reproductive and respiratory syndrome virus (PRRSV) during acclimatization. J Swine Health Prod 2002;10:147150.

    • Search Google Scholar
    • Export Citation
  • 18

    Fano E, Olea L, Pijoan C. Eradication of porcine reproductive and respiratory syndrome virus by serum inoculation of naive gilts. Can J Vet Res 2005;69:7174.

    • Search Google Scholar
    • Export Citation
  • 19

    Desrosiers R, Boutin M. An attempt to eradicate porcine reproductive and respiratory syndrome virus (PRRSV) after an outbreak in a breeding herd: eradication strategy and persistence of antibody titers in sows. J Swine Health Prod 2002;10:2325.

    • Search Google Scholar
    • Export Citation
  • 20

    Batista L, Pijoan C, Baidoo S. Eradication of porcine reproductive and respiratory syndrome virus (PRRSV) by serum inoculation with the homologous PRRSV strain, in Proceedings. 18th Int Pig Vet Soc Congr 2004;815.

    • Search Google Scholar
    • Export Citation
  • 21

    Wagner M. PRRS planned exposure: a summary of results, in Proceedings. 35th Annu Meet Am Assoc Swine Vet 2005;329336.

  • 22

    FitzSimmons MA. Principles of dealing with PRRS, in Proceedings. 35th Annu Meet Am Assoc Swine Vet 2005;319321.

  • 23

    Charerntanakul W, Platt R, Johnson W, et al. Immune responses and protection by vaccine and various vaccine adjuvant candidates to virulent porcine reproductive and respiratory syndrome virus. Vet Immunol Immunopathol 2006;109:99115.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 24

    Foss DL, Zillox MJ, Meier WA, et al. Adjuvant danger signals increase the immune response to porcine reproductive and respiratory syndrome virus. Viral Immunol 2002;15:557566.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 25

    Meier WA, Galeota J, Osorio FA, et al. Gradual development of the interferon-G response of swine to porcine reproductive and respiratory syndrome virus infection or vaccination. Virology 2003;309:1831.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 26

    Labarque G, Van Gucht S, Van Reeth K, et al. Respiratory tract protection upon challenge of pigs vaccinated with attenuated porcine reproductive and respiratory syndrome virus vaccines. Vet Microbiol 2003;95:187197.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 27

    Mengeling WL, Lager KM, Vorwald AC, et al. Comparative safety and efficacy of attenuated single-strain and multi-strain vaccines for porcine reproductive and respiratory syndrome. Vet Microbiol 2003;93:2538.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 28

    Opriessnig T, Pallarés FJ, Nilubol D, et al. Genomic homology of ORF 5 gene sequence between modified live vaccine virus and porcine reproductive and respiratory syndrome virus challenge isolates is not predictive of vaccine efficacy. J Swine Health Prod 2005;13:246253.

    • Search Google Scholar
    • Export Citation
  • 29

    Nilubol D, Platt KB, Halbur PG, et al. The effect of a killed porcine reproductive and respiratory syndrome virus (PRRSV) vaccine treatment on virus shedding in previously PRRSV infected pigs. Vet Microbiol 2004;102:1118.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 30

    Plana-Durán J, Bastons M, Urniza A, et al. Efficacy of an inactivated vaccine for prevention of reproductive failure induced by porcine reproductive and respiratory syndrome virus. Vet Microbiol 1997;55:361370.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 31

    Dee SA, Phillips R. Using vaccination and unidirectional pig flow to control PRRSV transmission. J Swine Health Prod 1998;6:2125.

  • 32

    Gillispie TG, Carroll AL. Methods of control and elimination of porcine reproductive and respiratory syndrome virus using modified live vaccine in a two-site production system. J Swine Health Prod 2003;11:291295.

    • Search Google Scholar
    • Export Citation
  • 33

    Ridremont B, Lebret A. Validation of a mass vaccination protocol with a PRRS modified live vaccine to stabilize French breeding herds, in Proceedings. 19th Int Pig Vet Soc Congr 2006;2: 43.

    • Search Google Scholar
    • Export Citation
  • 34

    Rodriguez E, Escuder M, Riera P, et al. A two-year study of the evolution of PRRS infection dynamics in a herd vaccinated with a modified live European-type strain, in Proceedings. 19th Int Pig Vet Soc Congr 2006;2: 51.

    • Search Google Scholar
    • Export Citation
  • 35

    Otake S, Dee SA, Rossow KD, et al. Transmission of porcine reproductive and respiratory syndrome virus by fomites (boots and coveralls). J Swine Health Prod 2002;10:5965.

    • Search Google Scholar
    • Export Citation
  • 36

    Yoon IJ, Joo HS, Christianson WT, et al. An indirect fluorescent antibody test for the detection of antibody of swine infertility and respiratory syndrome virus in swine sera. J Vet Diagn Invest 1992;4:144147.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 37

    Johnson W, Roof M, Vaughn E, et al. Pathogenic and humoral immune responses to porcine reproductive and respiratory syndrome virus (PRRSV) are related to viral load in acute infection. Vet Immunol Immunopathol 2004;102:233247.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 38

    Schurrer JA, Dee SA, Moon RD, et al. Retention of ingested porcine reproductive and respiratory syndrome virus in houseflies. Am J Vet Res 2005;66:15171525.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 39

    Pardo ID, Johnson GC, Kleiboeker SB. Phylogenetic characterization of canine distemper viruses detected in naturally infected dogs in North America. J Clin Microbiol 2005;43:50095017.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 40

    Opriessnig T, Halbur PG, Yoon KJ, et al. Comparison of molecular and biological characteristics of a modified live porcine reproductive and respiratory syndrome virus (PRRSV) vaccine (Ingelvac PRRS MLV), the parent strain of the vaccine (ATCC VR2332), ATCC VR2385, and two recent field isolates of PRRSV. J Virol 2002;76:1183711844.

    • Search Google Scholar
    • Export Citation
  • 41

    McCaw M, Murtaugh M, Laster S, et al. PRRSV rORF specific antibody responses following repeated homologous wild-type virus challenges, in Proceedings. 18th Int Pig Vet Soc Congr 2004;1: 36.

    • Search Google Scholar
    • Export Citation
  • 42

    Foss DL, Zilliox MJ, Meier W, et al. Adjuvant danger signals increase the immune response to porcine reproductive and respiratory syndrome virus. Viral Immunol 2002;15:557566.

    • Crossref
    • Search Google Scholar
    • Export Citation

Advertisement