Field efficacy of a novel porcine reproductive and respiratory syndrome modified‐live virus vaccine with an emphasis on growth performance

Abstract

Background

This field evaluation was designed to evaluate the efficacy of a new porcine reproductive and respiratory syndrome virus‐2 (PRRSV‐2) modified live virus vaccine at three independent pig farms.

Methods

Three farms were selected for this study based on their respiratory disease status caused by PRRSV‐2 infection in post‐weaning and growing pigs. Each farm housed a total of 40, 18‐day‐old pigs that were randomly allocated to one of two treatment groups. Pigs were administered a 1.0 mL dose of the bivalent vaccine intramuscularly at 21 days of age in accordance with the manufacturer's recommendations, whereas unvaccinated pigs were administered a single dose of phosphate buffered saline at the same age.

Results

Vaccinated groups were measured and calculated significantly (p < 0.05) higher in body weight and average daily weight gain on all three farms compared with unvaccinated groups. Vaccinated groups elicited PRRS antibodies and PRRSV‐2‐specific interferon‐γ secreting cells, which reduced the amount of PRRSV‐2 genomic copies in the blood and reduced macroscopic and microscopic lung lesions severity when compared with unvaccinated groups.

Conclusions

The field evaluation data demonstrated that a new PRRSV‐2 modified live virus vaccine was efficacious in swine herds suffering from respiratory diseases caused by PRRSV‐2 infection.

INTRODUCTION

Porcine reproductive and respiratory syndrome (PRRS) continues to threaten the global swine industry resulting in enormous economic losses. PRRS virus (PRRSV), the etiological agent of PRRS, causes reproductive failures in sows and respiratory distress in pigs of all ages (Zimmerman et al., 2019). PRRSV is an enveloped single stranded positive-sense RNA virus with a genome of approximately 15 kb in length that belongs to the family Arteriviridae within the genus Betaarterivirus (Snijder & Meulenberg, 1998; Walker et al., 2020). PRRSV is divided taxonomically into two species: PRRSV-1 (European type) as the species Betaarterivirus suid 1 and PRRSV-2 (North American type) as the species Betaarterivirus suid 2 (Walker et al., 2020). The first identified emergence of a PRRSV-2, in Korea, occurred through isolation from a growing pig with respiratory problems in 1994 (Kweon et al., 1994). Eleven years later (2005), PRRSV-1 was isolated (Nam et al., 2009).

To date, a perfect strategy does not exist in the prevention and control of PRRS (Chae, 2021). Despite this limitation, the impact that PRRSV has on the swine population is still best minimized through vaccination (Chae, 2021; Charerntantanakul, 2012). Several additional Modified live  vaccines (MLV) have been licensed in Korean market (Oh et al., 2019; Park, Seo, Han et al., 2014), since the first was introduced to the swine market more than two decades ago (Jeong et al., 2016). According to the Korean Animal Health Products Association (), 85% of PRRSV MLV vaccinated herds have received PRRSV-2 MLV, whereas only 15% are vaccinated with PRRSV-1 MLV. Swine farm infections are traditionally minimized through animal vaccination with PRRSV-2 MLV. A brand new PRRSV-2 MLV was recently launched into the market, although the full economic benefits have not yet been assessed. The objective of this study was to evaluate the efficacy of a new PRRSV-2 MLV with an emphasis on growth performance in farms suffering from PRRS.

MATERIALS AND METHODS

Farm history

The clinical field trial was conducted on three farms designated as follows: farm A (380-sow), B (100-sow) and C (420-sow). Each farm represented a unique site, and all were categorized as an all-in-all-out production system. Each farm tested seropositive for porcine circovirus type 2 (PCV2), but clinical signs of PCV2-associated disease were absent during observation. Pigs were vaccinated with a bivalent vaccine containing PCV2 and Mycoplasma hyopneumoniae at 21 days of age. PRRSV MLV had not been administered for at least 1 year in both sow and growing pigs. The three farms were selected based on respiratory disease status caused by PRRSV-2 infection in post-weaning and growing pigs. They were PRRS unstable as confirmed in a herd prescreening with PRRSV serology and by quantitative polymerase chain reaction (Holtkamp et al., 2011). Serum and lung tissue samples were collected from three growing pigs exhibiting respiratory symptoms characteristic of PRRSV at each farm and analysed. The serum samples from all three pigs were collected for virus isolation. The Lung samples were fixed in 10% neutral buffered formalin for histopathology and bacterial isolation. Histopathological lung lesions were then characterized by typical bronchointerstitial pneumonia. Trueperella pyogenes was isolated from the lung of one pig from farm A. Pasteurella multocida was isolated from lung of two pigs from farm B. Glaesserella parasuis was isolated from lung of two pigs from farm C. At the time of the study, there were no reports of reproductive failures in sows from the three selected farms. Clinical respiratory signs first appeared at approximately 7–10 weeks of age and reached peak mortality (approximately 2%–4% = farm A, 3%–4% = farm B and 4%–6% = farm C) between 10 and 15 weeks of age. Five post-weaned pigs from all farms were submitted to the Department of Veterinary Pathology for the isolation of PRRSV-2 at 8 weeks of age.

Field trial design

The guidelines of the Republic of Korea's Animal and Plant Quarantine Agency require that 20 piglets be selected and assigned to each group. Study design considerations included randomization, personnel blinding and animals were both weight-matched and sex-matched under a controlled clinical field trial format. To minimize sow variation, 83-day-old pigs were randomly selected from eight sows. Pigs were assigned to each of two uniform groups (20 pigs per group; 10 = male and 10 = female) using the random number generator function (Excel, Microsoft Corporation) (Table 1).

TABLE 1 Average daily weight gain (ADWG), body weight and mortality rate between vaccinated (Vac) and unvaccinated (UnVac) groups on three farms.

Age (day)	Farm A	Farm B	Farm C
VacA	UnVacA	VacB	UnVacB	VacC	UnVacC
ADWG (gram/pig/day)
21–70	381 ± 19	376 ± 21	391 ± 13	384 ± 11	394 ± 16	385 ± 15
70–112	733 ± 33*	646 ± 41	743 ± 52*	665 ± 38	745 ± 46*	667 ± 40
112–175	789 ± 51	779 ± 26	816 ± 47	799 ± 36	822 ± 48	799 ± 40
21–175	643 ± 16*	616 ± 10	660 ± 13*	630 ± 11	664 ± 15*	632 ± 11
Body weight (kg)
21	5.2 ± 0.1	5.3 ± 0.3	5.4 ± 0.2	5.4 ± 0.3	5.3 ± 0.3	5.4 ± 0.3
70	23.9 ± 1.0	23.7 ± 1.0	24.5 ± 0.6	24.2 ± 0.5	24.7 ± 0.7	24.2 ± 0.6
112	54.6 ± 1.1*	50.8 ± 1.2	55.7 ± 1.8*	52.1 ± 1.5	55.9 ± 1.9*	52.1 ± 1.7
175	104.2 ± 2.5*	100.2 ± 1.6	107.0 ± 2.1*	102.2 ± 1.8	107.5 ± 2.4*	102.7 ± 1.7
Mortality rate	1/20	2/20	0/20	2/20	1/20	3/20

At 0 days post-vaccination (dpv, 21 days of age), pigs in the VacA, VacB and VacC groups received a 1.0 mL dose of PRRSV-2 MLV (IMMUNIS PRRS-NA, WOOGENE B&G CO. LTD., ), a cell passaged attenuated strain containing 1 × 10⁵ TCID₅₀/mL derived from PRRSV-2 strain KNA090690 belonging to lineage 1 (GenBank no. ON936845 for ORF5 and ON936846 for ORF7). Each farm received a different serial of the vaccine as follows: farm A = Lot No: PRRS-A F001, farm B = Lot No: PRRS-A F002 and farm C = Lot No: PRRS-A F003. Pigs in the UnVacA, UnVacB and UnVacC groups were injected intramuscularly in the right side of the neck at 21 (24 days of age) with 1.0 mL of phosphate buffered saline (PBS, 0.01 M, pH 7.4). Pigs were weaned at 24 days in all three farms.

Upon vaccination, pigs from each of two groups were housed by treatment, with a minimum of two pens per treatment and 10 pigs per pen in the same building. Pens were randomly assigned to litters/treatments with an empty pen between each occupied pen to minimize the shedding of the vaccine virus to the individual pigs in the control group. All animals were housed within the same building in similar conditions, receiving the same feed and subjected to the same management practices. Five pigs from each group were randomly selected and euthanized for necropsy at 112 days of age. The rest of pigs from each group were euthanized for necropsy at 175 days of age. Pigs were sedated by an intravenous injection of sodium pentobarbital and then euthanized by electrocution as previously described (Beaver et al., 2001). Lung, liver, tonsil, kidney, spleen, small and large intestine and superficial inguinal lymph node tissues were collected from each pig at the time of necropsy. Tissues were fixed for 24 h in 10% neutral buffered formalin, routinely processed, and embedded in paraffin. The protocol for this field study was approved by the Institutional Animal Care and Use Committee.

Sampling collection

Blood samples were collected at 0 (21 days old), 28 (49 days old), 49 (70 days old), 91 (112 days old) and 154 (175 days old) dpv.

Clinical observations

The pigs were monitored daily for abnormal clinical signs and scored weekly using scores ranging from 0 (normal) to 6 (severe dyspnea and abdominal breathing) (Halbur et al., 1995). Observers were blinded to vaccination and type of vaccine status. Mortality rate was calculated as the number of pigs that died divided by the number of pigs initially assigned to that group within batch. Pigs that died or were culled throughout the study were necropsied. Evaluation of injection site reaction including palpation was performed 24 h post-vaccination.

Diagnostic test for mortality

Pigs that died were subjected to gross pathological examination within 24 h at local veterinary practitioners. All major organs, such as brain, lung, superficial inguinal lymph node, small and large intestine, liver, kidney and tonsils, were collected from each pig submitted to the diagnostic laboratory. Polymerase chain reaction assays were used in order to detect specific nucleic acids for PCV2, PRRSV, swine influenza virus and M. hyopneumoniae (Cai et al., 2007; Chung et al., 2002; Kim & Chae, 2004; Lee et al., 2008). All other bacterial isolation and identifications were carried out by using routine methods.

Average daily weight gain

The live weight of each pig was weighed at 21 (0 dpv), 70 (49 dpv), 112 (91 dpv) and 175 (154 dpv) days of age. The average daily weight gain (ADWG; grams/pig/day) was analysed over three time periods: (i) between 21 and 70 days old, (ii) between 70 and 112 days old, (ii) between 112 and 175 days old and (iv) between 21 and 175 days old. ADWG during the different production stages was calculated as the difference between the starting and final weight divided by the duration of the stage. Data for dead or removed pigs were included in the calculation.

Quantification of PRRSV cDNA

RNA was extracted from serum and tissue samples using a commercial kit (QIAamp Viral RNA Mini Kit, Qiagen). Genomic cDNA copy numbers were quantified using real-time PCR (Wasilk et al., 2004). PRRSV-2 forward and reverse primers were 5′-TGGCCAGTCAGTCAATCAAC-3′ and 5′-AATCGATTGCAAGCAGAGGGAA-3′, respectively (Wasilk et al., 2004). Real-time PCR for the quantification of genomic cDNA from the vaccine viruses was performed as previously described (Oh et al., 2019; Park et al., 2014a; Wasilk et al., 2004). The PCR amplification was set in a 20 µL volume consisted of 10 µL Maxima SYBR Green/ROX qPCR Master Mix (Thermo Scientific), 6 µL RNase-free DW, 1 µL of each primer of which the concentration is 10 pmole/µL and 2 µL of transcript cDNA. The reaction was carried out in thermal status of 95°C for 10 min, followed by 40 cycles of 95°C for 30 s, 60°C for 30 s and 72°C for 30 s.

To construct the standard curve for each assay, the plasmid DNA was built with the cDNA of the vaccine virus (PRRSV-2 strain KNA090690) following modified methods described previously (Chung et al., 2005). Briefly, PCR products of the target primers for real-time PCR were purified with Wizard PCR Preps DNA Purification kit (Promega). Resulting products were TA-cloned into the pCR II vector (Invitrogen), and transfection was performed on DH5α competent Escherichia coli (Enzynomics). The recombinant plasmid was purified using a plasmid Miniprep kit (Qiagen), and the concentration was quantified using OD₂₆₀ with a BioTek Epoch 2 microplate spectrophotometer (Agilent). In each run of real-time PCR, standard curve was generated with decimally diluted plasmid DNA of 10⁰–10⁹ copies/µL. The melting curves of all reactions were analysed for verification of the amplification results, and the detection limit of this assay was 10 copies/µL.

Serology

The serum samples were tested using the commercially available PRRSV (IDEXX PRRS X3 Ab test, IDEXX Laboratories Inc.). Samples were considered positive for PRRSV antibodies if the sample-to-positive (S/P) ratio was ≥0.4 in accordance with the manufacturer's instructions for each kit.

The serum samples were tested using serum virus neutralization (SVN) test against vaccine virus (PRRSV-2 strain KNA090690) with modified methods of previous researches (Yoon et al, 1994; Zuckermann et al., 2007). Serum samples were heated for 45 min at 56°C for inactivation before tests. Each serum was binarily diluted in Dulbecco's Modified Eagle's Medium (Sigma) with 10% FCS (Sigma), 20 mM l-glutamine (Cell-grow) and antibiotic–antimycotic mixture (Sigma). A volume of 100 µL each diluted serum was added to an equal volume of the vaccine virus (PRRSV-2 strain KNA090690) in the concentration of 1 × 10³ TCID₅₀/mL. The mixtures were incubated in 37°C for 1 h and were incorporated into confluent MARC-145 cell in a 96 well microplate. The MARC-145 cells in this microplate were suspended with 200 µL of the RPMI medium with 10% FCS. For 7 days, the cells were in the 37°C and 5% CO₂ incubator and cytopathic effect within each well of the microplate was recorded daily. Back titration of the inoculum was carried out to verify the titre of inoculated virus. The threshold of the SVN test for the serum samples was the titre of 2.0 times dilution.

Enzyme-linked immunospot assay

Enzyme-linked immunospot (ELISPOT) assay was conducted to measure the numbers of PRRSV-specific interferon-γ secreting cells (IFN-γ-SC) (Park, Seo, Han et al., 2014). The numbers of PRRSV-specific IFN-γ-SC were determined in peripheral blood mononuclear cells (PBMC) by ELISPOT (MABTECH) according to the manufacturer's instructions. Porcine PBMCs were isolated from 4 to 5 mL of pig blood samples collected in lithium–heparin as anticoagulant by density gradient in Histopaque-1077 solution (Sigma) according to the manufacturer's instructions (Ferrari et al., 2013). In brief, 5 × 10⁵ PBMC was plated in 96-well microplate precoated with swine specific IFN-γ antibody (10 µg/mL, MABTECH) and stimulated by addition of vaccine virus (PRRSV-2 strain KNA090690) solution in RPMI 1640 medium for 20 h at 37°C in a 5% humidified CO₂ atmosphere. The linear response was tested between 0.01 and 0.1 multiplicity of infection. Phytohemagglutinin (10 µg/mL, Roche Diagnostics GmbH) and culture medium were used as positive and negative control, respectively. The IFN-γ positive spots on the membranes were imaged, analysed, and counted using an automated ELISPOT Reader (AID ELISPOT Reader, AID GmbH). The results were expressed as the numbers of IFN-γ-SC per million PBMC. ELISPOT assay was done in duplicate.

Pathology

Lungs were evaluated with a morphometric analysis of the macroscopic lung lesion score by assessing each individual lung lobe. Each lobe received a percentile score that ranges from 0% to 100% of the affected area. The entire lung was scored in the same manner (Halbur et al., 1995).

Microscopic interstitial pneumonia was scored on a scale of ‘0’ to ‘4’. The score of ‘0’ was considered normal, followed by ‘1’ which indicated mild interstitial pneumonia. A score of ‘2’ was assigned as moderate multifocal interstitial pneumonia; ‘3’ was designated moderate diffuse interstitial pneumonia; and a score of ‘4’ indicated severe interstitial pneumonia (Halbur et al., 1995).

Statistical analysis

Statistical analyses were performed with IBM SPSS Statistics for Windows version 29.0 (IBM Corp.). Real-time PCR and neutralizing antibody data were recalculated to log₁₀ and log₂ values, respectively, prior to statistical analysis. The collected data were assessed by the Shapiro–Wilk test for a normal distribution. Then, Student's t test was performed to determine if there were statistically significant differences between different groups at each time point. In case of the normality assumption was not met, Mann–Whitney test was performed to compare the differences between different groups. The value of p < 0.05 was considered significant and reported in p-values.

RESULTS

PRRSV isolation

PRRSV-2 was isolated from pig serum samples from all three farms before the animals received their first vaccination. All PRRSV-2 field isolates were categorized as lineage 5, despite the geographic difference between each farm. The vaccine strain (KNA090690, GenBank no. ON936845) and field PRRSV-2 viruses shared an 85.7% (farm A), 85.9% (farm B) and 86.2% (farm C) identity of their ORF5 amino acid sequences, respectively.

Clinical signs

The mean clinical respiratory scores were significantly (p < 0.05) lower in vaccinated groups than in unvaccinated groups from 28 to 119 dpv in farm A (Figure 1a). By 28 dpv, several pigs from the VacA group displayed mild transient dyspnea and tachypnea at rest. Additional pigs from the same group displayed these same clinical signs from 42 to 84 dpv after stress from handling. By 28 dpv, several pigs from the UnVacA group displayed mild dyspnea and/or tachypnea when at rest. Additional pigs from the same groups displayed moderate dyspnea and tachypnea after stress by handling from 49 to 84 dpv.

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Vaccination significantly lowered the mean clinical respiratory scores (p < 0.05) lower in farm B compared to the unvaccinated groups from 21 to 98 dpv (Figure 1b). Pigs from the VacB group had mild dyspnea and/or tachypnea at rest following a period of stress between 21 and 98 dpv. Within this period, moderate respiratory disease characterized by moderate dyspnea and tachypnea was documented from 35 to 56 dpv. By 63 to 98 dpv, clinical scores decreased anf were documented as mild dyspnea and/or tachypnea when at rest.

The mean clinical respiratory scores were significantly (p < 0.05) lower in farm C vaccinated groups than in unvaccinated groups from 14 to 84 dpv (Figure 1c). VacC pigs displayed mild dyspnea and/or tachypnea when stress following a period of 14 to 98 dpv. Within this timeframe, clinical signs were considered mild between 14 and 35 dpv as they occurred only at rest and heighted to a moderate level from 42 to 84 dpv.

Mortality

Pig mortality rate is summarized in Table 1. PRRSV-2 was not detected as a cause of death for any vaccinated pigs. Study mortality of unvaccinated pigs was primarily related to co-infections of PRRSV-2 and other bacterial pathogens on all three farms. Vaccinated and unvaccinated death were categorized further as follows: Streptococcus suis was isolated from the brain of a VacA groups pig died, whereas T. pyogenes was isolated from the lungs of a VacC group that died. PRRSV-2, G. parasuis and P. multocida were isolated from lungs of two UnVacA group pigs that died. PRRSV-2, T. pyogenes and P. multocida were isolated from lungs of two UnVacB group pigs that died. PRRSV-2 and G. parasuis were isolated from lungs of three UnVacC group pigs that died.

Growth performance

A significant difference in average body weight (±standard deviation) between vaccinated and unvaccinated groups in all three farms was not present at the start of the field trial. Within each of the three farms, vaccinated groups were significantly (p < 0.05) higher in body weight compared to unvaccinated groups at 112 (91 dpv) and 175 (154 dpv) days of age. A significantly (p < 0.05) higher ADWG was measured in vaccinated groups between 70 and 112 days of age and between 21 and 175 days of age, when compared with unvaccinated groups (Table 1).

Quantitation of PRRSV cDNA in blood and lung

PRRSV genomic copies of cDNA were not detected from serum samples of any pigs from the three farms in either the designated vaccinate and non-vaccinate groups at 0 dvp (at the time of vaccination). The number of PRRSV cDNA genomic copies measured from vaccinated group serum samples was significantly (p < 0.05) lower on 49, 91 and 154 dpv compared to those from the unvaccinated groups in farm A (Figure 2a), B (Figure 2b) and C (Figure 2c). The prevalence of PRRSV positive pigs in vaccinated and unvaccinated groups from each of the farms is summarized in Table 2.

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TABLE 2 Prevalence of porcine reproductive and respiratory syndrome virus (PRRSV) viraemia and pathology between vaccinated (Vac) and unvaccinated (UnVac) groups on three farms.

Dpv	Farm A	Farm B	Farm C
	VacA	UnVacA	VacB	UnVacB	VacC	UnVacC
PRRSV viraemia
0	0/20	0/20	0/20	0/20	0/20	0/20
28	11/20	10/20	7/20	12/20	7/20	10/20
49	13/20	17/20	10/20	15/20	11/20	14/20
91	13/19	16/18	12/20	15/19	12/19	14/18
154	10/14	11/13	10/15	11/13	9/14	9/12
Lung genomic copy of PRRSV
91	1.7 ± 1.6*	3.7 ± 0.5	2.1 ± 1.3*	3.5 ± 0.9	1.7 ± 1.6	2.8 ± 1.6
154	1.1 ± 0.9*	2.5 ± 1.2	0.8 ± 0.8*	2.3 ± 1.2	1.1 ± 1.0*	2.1 ± 1.5
Lung lesion score
Macroscopic
91	22 ± 4.9*	46 ± 5.7	22 ± 3.4*	41 ± 11.7	21 ± 6.2	39 ± 0.9
154	21 ± 6.2*	33 ± 4.9	20 ± 8.1*	32 ± 7.6	19 ± 2.4*	32 ± 10.2
Microscopic
91	1.7 ± 0.6*	2.8 ± 0.3	1.6 ± 0.4*	2.8 ± 0.5	1.6 ± 0.4*	2.6 ± 0.6
154	1.6 ± 0.3*	2.5 ± 0.6	1.6 ± 0.2*	2.1 ± 0.5	1.4 ± 0.3*	2.2 ± 0.3

The PRRSV cDNA genomic copies measured from vaccinated group lungs were significantly (p < 0.05) lower on 91 and 154 dpv compared to those from the unvaccinated groups in farms A and B. The PRRSV cDNA genomic copies measured from vaccinated groups lungs were significantly (p < 0.05) lower on 154 dpv compared to those from the unvaccinated groups in farm C (Table 2).

Immune responses

The presence of PRRSV antibodies was tested by Enzyme-linked immunosorbent assay (ELISA) and was detected in 7 pigs (VacA), 4 pigs (UnVacA), 1 pig (VacB), 4 pigs (UnVacB), 2 pigs (VacC) and 2 pigs (UnVacC) at 0 dvp (at the time of vaccination). The mean PRRSV antibody S/P ratio and the mean numbers of PRRSV-specific IFN-γ-SC were significantly (p < 0.05) higher in the vaccinated groups on 28, 49, 91 and 154 dpv compared to those from the unvaccinated groups in farm A (Figure 3a), B (Figure 3b) and C (Figure 3c).

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The mean number of neutralizing antibodies that were produced against the against vaccine virus was low (either 1:4 or 1:8), with no statistical differences between the vaccinated and unvaccinated groups in farm A (Figure 4a), B (Figure 4b) and C (Figure 4c), with the number 0 pigs that produced these titres as follows: farm A vaccinated pigs at 0 (0/20 pigs), 28 (11/20 pigs), 49 (11/20 pigs), 91 (13/19 pigs) and 154 (12/14 pigs) dpv; farm A unvaccinated pigs 0 (1/20 pigs), 28 (6/20 pigs), 49 (10/20 pigs), 91 (11/18 pigs) and 154 (6/13 pigs) dpv; farm B vaccinated pigs at 0 (6/20 pig), 28 (11/20 pigs), 49 (15/20 pigs), 91 (17/20 pigs) and 154 (12/20 pigs) dpv; farm B unvaccinated pigs at 0 (0/ 20 pigs), 28 (8/20 pigs), 49 (10/20 pigs), 91 (12/19 pigs) and 154 (7/13 pigs) dpv; farm C vaccinated pigs at 0 (0/20 pigs), 28 (11/20 pigs), 49 (15/20 pigs), 91 (16/19 pigs) and 154 (11/14 pigs) dpv; and farm C unvaccinated pigs at 0 (1/20 pigs), 28 (9/20 pigs), 49 (8/20 pigs), 91 (10/18 pigs) and 154 (6/ 12 pigs) dpv.

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The mean numbers of PRRSV-specific IFN-γ-SC were significantly (p < 0.05) higher in the vaccinated groups on 28, 49, 91 and 154 dpv compared to those from unvaccinated groups in farm A (Figure 5a), B (Figure 5b) and C (Figure 5c).

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Pathology

Macroscopic and microscopic lung lesion severity at 91 and 154 dpv scored significantly (p < 0.05) lower in the vaccinated groups compared with those in the unvaccinated groups in three farms (Table 2). Vaccinated pigs showed mild interstitial pneumonia (Figure 6a), whereas unvaccinated pigs showed moderate interstitial pneumonia (Figure 6b).

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DISCUSSION

Clinical field trials are useful in evaluating the growth performance of PRRS MLV. ADWG has been improved in pigs through use of MLV vaccination as previously evaluated under field conditions (Park, Seo, Kang et al., 2014). The amount of economic loss reduction in the field was calculated as ADWG between the unvaccinated and MLV vaccinated pigs and resulted in approximately 650 g/days. Pigs in commercial farms settings are continuously exposed and re-exposed to field PRRSV strains circulating within the farms and to co-infections with bacterial pathogens such as P. multocida, T. pyogenes and G. parasuis. Therefore, the effect of growth performance obtained from this field trial provides useful data that can be directly referenced to a farmer.

Levels of PRRSV blood load play a key role in understanding the development of respiratory diseases and how they are correlated with the severity of interstitial pneumonia (Han et al., 2013; Lager et al., 2014). It is therefore considered an important index in assessing the efficacy of PRRSV vaccines in their ability to reduce PRRS blood viral load (Wei et al., 2019; Roca et al, 2012). The PRRS blood viral load was significantly reduced in vaccinated groups, which led to reduced interstitial pneumonia severity compared to the unvaccinated groups. The key elements of immune response involvement in PRRSV-2 viraemia reduction are not completely elucidated as PRRSV viraemia is often resolved before neutralizing antibodies are even able to be detected in infected pigs (Mateu & Diaz, 2008; Nelson et al., 1994). The same principal was evaluated in another study with vaccinated pigs (Mengeling et al., 2003). These observations all lead to the potential for cell-mediated immunity playing a role in this field study (Loving et al., 2015; Park et al., 2014b). In the preset field study, vaccination reduced PRRS blood viral load as indicated by the low titres (1:4 or 1:8) of neutralizing antibodies, supporting that cell-mediated immunity can confer protection (Vu et al., 2017). Vaccination with this new PRRSV-2 MLV elicited cellular immune responses as measured by PRRSV specific IFN-SC. This leads to reduced levels of PRRS blood viral load and thus a reduction in the degree of interstitial pneumonia.

In general, vaccination unfavourably resulted in lower S/P ratios and IFN-γ responses compared to those of unvaccinated groups. These field trials challenged these findings where the opposite was concluded. Many of the pigs were exposed to natural infection later in these field trials resulting on stronger immune responses, as indicated by the higher S/P rations and IFN-γ responses from the vaccinated groups compared to the unvaccinated groups as previous studies (Madaponga et al., 2020; Zuckermann et al., 2007).

There is some existing concern with MLV and their reversion to virulence. For PRRSV, this has only been reported for a limited number of cases and with one particular vaccine only (Nielsen et al., 2001). The safety and stability of the PRRSV-2 KNA090690 master seed virus demonstrated that reversion to virulence did not occur through a 6 virus back passage as required prior to MLV license approval (data not shown). An ADWG difference between vaccinated and unvaccinated groups was not calculated in the present field trials within the first 7 weeks post-vaccination. Safety was established during the 7 weeks following vaccination, as it did not negatively influence the health status of the pigs.

Predictions of protective efficacy have been associated with antigenic, but not genetic, similarities between the vaccine and field wild-type strain (Lager & Mengeling, 1999; Murtaugh & Genzow, 2011; Prieto et al., 2008). In the present study, the vaccine strain and the field strain are genetically distant. Nevertheless, this PRRSV-2 MLV provided good protection against field wild-type strains and improved the growth performance. This information from this trial is of clinical significance, as many swine practitioners still request genetic analysis of field wild-type viruses as a criterion before they select a proper PRRS MLV that is genetically similar to the strain circulating in their specific farms. These field data provide scientific evidence that the degree of antigenic similarities but not genetic similarities predicts the cross-protective ability of a PRRS MLV.

AUTHOR CONTRIBUTIONS

Sehyeong Ham and Hyunjoon Lee: Conceptualization; data curation; investigation; methodology. Jeongmin Suh: Data curation; resources; visualization. Chonghan Kim: Methodology; software. Woo Ju Kwon: Formal analysis. Gyeong-Seo Park: Formal analysis. Chanhee Chae: Conceptualization; project administration; supervision; writing – review and editing.

CONFLICT OF INTEREST STATEMENT

The authors declare no conflicts of interest with respect to their authorship.

FUNDING INFORMATION

Ministry of Agriculture, Food and Rural Affairs (MAFRA), Grant number: 821032-03; BK 21 FOUR Future Veterinary Medicine Leading Education and Research Center, Grant number: A0449-20200100

ETHICS STATEMENT

All of the methods were previously approved by the Seoul National University Institutional Animal Care and Use and Ethics Committee. Sample collection was carried out according to the animal welfare code of Korea.

DATA AVAILABILITY STATEMENT

The data that support the findings of this study are available from the corresponding author upon reasonable request.

PEER REVIEW

The peer review history for this article is available at .

References

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