Yeonsu Oh

Comparison of the virulence of European and North American genotypes of porcine reproductive and respiratory syndrome virus in experimentally infected pigs

The objective of this study was to compare the virulence of Korean types 1 and 2 porcine reproductive and respiratory syndrome virus (PRRSV) isolated from weaned pigs with respiratory disease. Affected pigs were within the same herd and animals infected with type 2 virus had significantly higher mean rectal temperatures than those with type 1 virus between days 2 and 9 post-inoculation (P<0.05). Similarly, mean serum viral titres, expressed as tissue culture infective doses 50% (TCID50)/mL, as well as macroscopic and microscopic pulmonary lesion scores, were significantly higher at multiple time points in pigs infected with type 2 PRRSV compared to those infected with type 1 virus. Mean numbers of PRRSV-positive cells/unit area of lungs and lymph nodes were also significantly higher in type 2 PRRSV infected pigs. This study demonstrates that type 2 PRRSV is more virulent than type 1 PRRSV in this experimental setting as reflected by the pulmonary pathology induced, the extent of virus distribution, and oral shedding of the virus.

Comparative efficacy of commercial Mycoplasma hyopneumoniae and porcine circovirus 2 (PCV2) vaccines in pigs experimentally infected with M. hyopneumoniae and PCV2.

The efficacies of two commercial Mycoplasma hyopneumoniae bacterins and porcine circovirus 2 (PCV2) vaccines were compared in conventional pigs immunized at different ages based on humoral response, pathological observation, and growth performance from birth to finishing (175 days of age) using a M. hyopneumoniae and PCV2 co-infection challenge model. One-week-old pigs (n=110) were randomly assigned to five groups: three vaccinated and challenged (VC), and one each of non-vaccinated and challenged (NVC) and negative control. A significant difference was found in the number of genomic copies of M. hyopneumoniae in nasal swabs and PCV2 in serum samples, the average daily weight gain (gram/pig/day) between 63 and 133 dpi, gross and histopathological lung lesion scores, histopathological lymph node lesion scores, and the immunohistochemical analysis of PCV2 among the three VC groups. The single dose schedule for M. hyopneumoniae bacterins and PCV2 vaccines have the advantages of (i) improving daily weight gain (122.4%) and slaughter weight (120.5%), and (ii) reducing the incidence of clinical signs and lung and lymph node lesions.

Pathogenesis of Korean type 1 (European genotype) porcine reproductive and respiratory syndrome virus in experimentally infected pregnant gilts.

The aim of this study was to elucidate the pathogenesis of experimental infection with Korean type 1 porcine reproductive and respiratory syndrome virus (PRRSV) by defining the virus distribution, sites of viral replication, viraemia and gross and microscopical lesions in conventional pigs studied for 28 days after intranasal inoculation. Mean rectal temperature was significantly higher in infected pigs than in negative control pigs at 2 days post inoculation (dpi) (P=0.004), 3 dpi (P<0.001), 4 dpi (P=0.003) and 5 dpi (P=0.034). The log(10)TCID(50)/ml of type 1 PRRSV increased significantly at 0-1 dpi (P=0.024) and 5-7 dpi (P=0.029), but decreased at 10-14 dpi (P=0.026) and 14-21 dpi (P=0.012) in infected pigs. Infected pigs developed multifocal, tan-mottled areas of lung tissue with irregular and indistinct borders. Microscopical lesions, when present, were multifocal, mild to moderate, generally most extensive at 5-7 dpi (P=0.036), and were nearly resolved at 28 dpi. Type 1 PRRSV nucleic acid and antigen were detected exclusively within the cytoplasm of macrophages and type I and II pneumocytes. The score for PRRSV-positive cells increased at 3-7 dpi (P<0.05) and decreased at 10-14 dpi (P=0.034) in infected pigs. Thus, respiratory disease was reproduced in conventional pigs by infection with Korean type 1 PRRSV.

Effect of the Modified Live Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) Vaccine on European and North American PRRSV Shedding in Semen from Infected Boars

ABSTRACT The objective of the present study was to compare the effects of the modified live porcine reproductive and respiratory syndrome virus (PRRSV) vaccine (Ingelvac PRRS MLV; Boehringer Ingelheim Animal Health, St. Joseph, MO) on European and North American PRRSV shedding in the semen of experimentally infected boars. The boars were randomly divided into six groups. Vaccinated boars shed the North American PRRSV at the rate of 100.1 to 101.0 viral genome copies per ml and 3.63 to 101.1 50% tissue culture infective doses (TCID50)/ml, respectively, in semen, whereas nonvaccinated boars shed the North American PRRSV at the rate of 100.2 to 104.7 viral genome copies per ml and 1.14 to 103.07 TCID50/ml, respectively, in semen. Vaccinated boars shed the European PRRSV at the rate of 100.1 to 104.57 viral genome copies per ml and 1.66 to 103.10 TCID50/ml, respectively, in semen, whereas nonvaccinated boars shed the European PRRSV at the rate of 100.3 to 105.14 viral genome copies per ml and 1.69 to 103.17 TCID50/ml, respectively, in semen. The number of genomic copies of the European PRRSV in semen samples was not significantly different between vaccinated and nonvaccinated challenged European PRRSV boars. The present study demonstrated that boar vaccination using commercial modified live PRRSV vaccine was able to decrease subsequent shedding of North American PRRSV in semen after challenge but was unable to decrease shedding of European PRRSV in semen after challenge.

Evidence of shedding of porcine circovirus type 2 in milk from experimentally infected sows.

Detection of porcine circovirus type 2 (PCV2) was to evaluate the milk from experimentally infected sows using polymerase chain reaction (PCR) and virus isolation. Six pregnant sows were inoculated intranasally with PCV2 at 93 days of gestation, and milk samples were collected from all sows at 1, 3, 6, 9, 12, 15, 18, 21, 24, and 27 days of lactation. PCV2 was detected in milk as early as day 1 of lactation in all six sows. Thereafter, all infected sows remained positive by PCR for PCV2 in milk until 27 days of lactation. In addition, PCV2 itself was isolated from milk collected from a virus-infected sows. These results suggest that PCV2 may be shed in milk following infection of pregnant sows by the virus.

Efficacy of a reformulated inactivated chimeric PCV1-2 vaccine based on clinical, virological, pathological and immunological examination under field conditions.

Inactivated chimeric porcine circovirus (PCV) 1-2 vaccine was initially taken off the market due to concerns that the vaccine virus was not killed and thus further replicated and spread in the pig population. In August 2011, a reformulated inactivated chimeric PCV1-2 vaccine re-entered the market. The efficacy of the reformulated inactivated chimeric PCV1-2 vaccine was evaluated under field conditions for registration as recommended by the Republic of Koreas Animal, Plant & Fisheries Quarantine & Inspection Agency. Three farms were selected based on their history of postweaning multisystemic wasting syndrome (PMWS). On each farm, a total of 50 3-week-old pigs were randomly allocated to one of two treatment groups: (i) vaccinated at 3 weeks of age and (ii) non-vaccinated. Clinical examination indicated that vaccinated animals displayed an improved average daily weight gain (672.2g/day vs. 625g/day; difference of +47.3g/day; P<0.05) and a reduced time to market (177 days vs. 183 days; difference of -6 days; P<0.05). Virological examination indicated that vaccinated animals displayed a reduced PCV2 load in the blood and nasal swabs compared to non-vaccinated animals. Pathological examination indicated that vaccination of pigs against PCV2 effectively reduced the number of PMWS-associated microscopic lesions and the PCV2 load in lymphoid tissues compared to non-vaccinated animals in the 3 herds. Immunological examination indicated that vaccinated animals induced PCV2-specific neutralizing antibodies (NA) and interferon-γ-secreting cells (IFN-γ-SCs). A reduction in the PCV2 load in the blood coincided with the appearance of both PCV2-specific NA and IFN-γ-SCs in the vaccinated animals. The number of CD4(+) cells was decreased in non-vaccinated animals compared to vaccinated animals. The reformulated inactivated chimeric PCV1-2 vaccine seems to be very effective in controlling PCV2 infection based on clinical, virological, pathological, and immunological evaluations under field conditions.

Protective effect of the maternally derived porcine circovirus type 2 (PCV2)-specific cellular immune response in piglets by dam vaccination against PCV2 challenge.

The objective of the present study was to evaluate (i) the passive transfer of maternally derived functional porcine circovirus type 2 (PCV2)-specific lymphocytes of seronegative sows immunized with the PCV2 vaccine to newborn piglets and (ii) the functional role of the maternally derived PCV2-specific cellular immune response in protecting newborn piglets from challenge with PCV2. After ingesting colostrums, piglets from vaccinated sows (PT01 and PT02) have significantly higher numbers of PCV2-specific gamma interferon-secreting cells, an increased PCV2-specific delayed type hypersensitivity response, and a stronger proliferative response of peripheral blood mononuclear cells compared with piglets from non-vaccinated seronegative sows (PT03 and PT04). In the PCV2 challenge study, the number of serum genomic PCV2 copies was significantly less in piglets from vaccinated sows (PT02) compared with piglets from non-vaccinated sows (PT04) at 7-28 days post-inoculation (P<0.05 and P<0.001). The histopathological lesions and immunohistochemical scores were significantly lower in piglets of vaccinated sows compared with those of non-vaccinated sows. To our knowledge, this is the first report of transferring a maternally derived PCV2-specific cellular immune response from vaccinated dams to their offspring. Maternally derived adaptive cellular immune responses play a critical role in protecting newborn piglets challenged with PCV2 at 3 weeks of age.

Reduction of porcine circovirus type 2 (PCV2) viremia by a reformulated inactivated chimeric PCV1-2 vaccine-induced humoral and cellular immunity after experimental PCV2 challenge

BackgroundThe objective of the present study was to elucidate the humoral and cellular immune response mechanisms by which a reformulated inactivated chimeric PCV1-2 vaccine reduces the PCV2 viremia. Forty PCV2 seronegative 3-week-old pigs were randomly divided into the following four groups: vaccinated challenged (T01), vaccinated non-challenged (T02), non-vaccinated challenged (T03), and non-vaccinated non-challenged (T04) animals. The pigs in groups T01 and T02 were immunized with a reformulated inactivated chimeric PCV1-2 vaccine (Fostera™ PCV; Pfizer Animal Health) administered as a 2.0 ml dose at 21 days of age. At 35 days of age (0 days post-challenge), the pigs in groups T01 and T03 were inoculated intranasally with 2 ml each of PCV2b.ResultsA reduction of PCV2 viremia coincided with the appearance of both PCV2-specific neutralizing antibodies (NA) and interferon-γ-secreting cells (IFN-γ-SCs) in the vaccinated animals. However, the presence of anti-PCV2 IgG antibodies did not correlate with the reduction of PCV2 viremia. Lymphocyte subset analysis indicated that the numbers of CD3+ and CD4+ cells increased in vaccinated animals but the numbers of CD4+ cells decreased transiently in non-vaccinated animals. The observation of a delayed type hypersensitivity response in only the vaccinated animals also supports a CD4+ cell-associated protective cellular immune response induced by the reformulated inactivated chimeric PCV1-2 vaccine.ConclusionsThe induction of PCV2-specific NA and IFN-γ-SCs, and CD4+ cells by the reformulated inactivated chimeric PCV1-2 vaccine is the important protective immune response leading to reduction of the PCV2 viremia and control of the PCV2 infection. To our knowledge this is the first demonstration of protective humoral and cellular immunity induced by the reformulated inactivated chimeric PCV1-2 vaccine and its effect on reduction of PCV2 viremia by vaccination.

Comparison of sow and/or piglet vaccination of 3 commercial porcine circovirus type 2 (PCV2) single-dose vaccines on pigs under experimental PCV2 challenge.

The objective of the present study was to compare the effect of sow and/or piglet vaccination regimen by three commercial porcine circovirus type 2 (PCV2) vaccines on pigs experimentally challenged with PCV2 at 84 days of age based on immunological, virological, and pathological evaluation. One hundred and nineteen piglets born to vaccinated or non-vaccinated sows were divided into 17 groups. A portion of the pigs with or without passively acquired immunity was vaccinated at 21 or 49 days of age. Regardless of the PCV2 vaccine, the combination of sow and pig (49 days of age) vaccinations significantly (P<0.05) reduced PCV2 viremia, induced higher log2 transformed neutralizing antibody titers, and resulted in higher proportion of CD4(+)CD8(+)IFN-γ(+) lymphocyte subsets than the sow vaccination alone, the pig (21 or 49 days of age) vaccination alone, and the combination of sow and pig (21 days of age) vaccinations at various days post challenge. The results showed a significant negative correlation between maternally derived antibodies at the day of vaccination and the increment of antibody titers to PCV2 at 28 days post vaccination in the combination of sow and pig (21 days of age) vaccinations but not the combination of sow and pig (49 days of age) vaccinations. The combination of sow and pig (49 days of age) vaccinations could be more effective for controlling PCV2 infection if PCV2 the infection occurs during the growing-finishing period in herds. Optimal vaccination strategies must balance the advantage of delayed vaccination with the need to induce immunity prior to exposure to pathogens under field conditions.

Formation of liposome by microfluidic flow focusing and its application in gene delivery

We report the formation of liposomes in a simple procedure using a microfluidic hydrodynamic flow focusing method for the application in gene delivery. We fabricated microfluidic device using soft lithography and polydimethylsiloxane (PDMS) molding technique. Lipid-containing stream was surrounded by aqueous stream and liposomes were formed at the lipid-water interface. Size distribution of liposomes and zeta potential of liposome dispersion were investigated under various flow rate ratio (FRR) and processing temperature. Size distributions of liposomes were measured by dynamic light scattering (DLS), and zeta potential was measured to quantify the colloidal stability. Prepared liposomes were used as a vehicle for gene delivery, and the successful expression of delivered gene was observed by fluorescent microscope.