Maria Fort

Porcine circovirus type 2 (PCV2) vaccination of conventional pigs prevents viremia against PCV2 isolates of different genotypes and geographic origins.

The efficacy of recently developed porcine circovirus type 2 (PCV2) vaccines has not been tested yet against PCV2 isolates of the two proposed genotypes. In the present work, the efficacy of a subunit vaccine containing PCV2 capsid protein was evaluated by using a challenge model with four different PCV2 isolates of different genotype and geographic origin. The vaccine prevented the development of viremia in all cases as well as significantly decreased nasal and faecal shedding of the virus. Also, the vaccine elicited PCV2-specific neutralizing antibodies to PCV2 even in the presence of maternally derived immunity.

Development of cell-mediated immunity to porcine circovirus type 2 (PCV2) in caesarean-derived, colostrum-deprived piglets.

Abstract The interaction between porcine circovirus type 2 (PCV2) and the pig immune system has been suggested to be a determinant event for the pathogenesis of postweaning multisystemic wasting syndrome (PMWS). To gain insight into the host immune mechanisms developed upon PCV2 infection, early innate and adaptive immune responses were examined in 1-week-old, caesarean-derived, colostrum-deprived piglets using a subclinical infection model of PCV2 in combination with lipopolysaccharide (LPS) as a potential immunostimulation factor. The use of LPS did not show any significant effect on the course of PCV2 infection, nor did in the evolution of the immunological parameters evaluated. Ex vivo responses were detected as early as 1 day post-infection (PI) and consisted of an elevation of the plasmatic levels of interleukin (IL)-8 in PCV2-inoculated pigs followed by an increase on plasmatic IFN-α at day 5 PI. Regarding IL-10, only one PCV2-inoculated pig was positive (day 7 PI); this pig was the only one in which viremia persisted until the end of the study. In vitro cytokine determination showed that, regardless of the treatment administrated to the pigs, an IL-10 release was observed when peripheral blood mononuclear cells (PBMC) cultures were stimulated with PCV2. Seroconvertion to PCV2 measured by an immunoperoxidase monolayer assay (IPMA) occurred between 7 and 14 days PI, whereas neutralizing antibodies (NA) did not appear until day 29 PI. PCV2 DNA was first detected in serum at day 7 PI, reaching the peak of viremia between days 14 and 21 PI, followed by a drop in viral load that was found coincident with the appearance of PCV2-specific IFN-γ-secreting cells (PCV2-IFN-γ-SC) and NA. Results from the present work suggest that viral clearance might be mediated by the development of PCV2-IFN-γ-SC in contribution to the PCV2-specific NA.

Granuloma Encapsulation Is a Key Factor for Containing Tuberculosis Infection in Minipigs

A transthoracic infection involving a low dose of Mycobacterium tuberculosis has been used to establish a new model of infection in minipigs. The 20-week monitoring period showed a marked Th1 response and poor humoral response for the whole infection. A detailed histopathological analysis was performed after slicing the formalin-fixed whole lungs of each animal. All lesions were recorded and classified according to their microscopic aspect, their relationship with the intralobular connective network and their degree of maturity in order to obtain a dissemination ratio (DR) between recent and old lesions. CFU counts and evolution of the DR with time showed that the proposed model correlated with a contained infection, decreasing from week 9 onwards. These findings suggest that the infection induces an initial Th1 response, which is followed by local fibrosis and encapsulation of the granulomas, thereby decreasing the onset of new lesions. Two therapeutic strategies were applied in order to understand how they could influence the model. Thus, chemotherapy with isoniazid alone helped to decrease the total number of lesions, despite the increase in DR after week 9, with similar kinetics to those of the control group, whereas addition of a therapeutic M. tuberculosis fragment-based vaccine after chemotherapy increased the Th1 and humoral responses, as well as the number of lesions, but decreased the DR. By providing a local pulmonary structure similar to that in humans, the mini-pig model highlights new aspects that could be key to a better understanding tuberculosis infection control in humans.

Immune responses and vaccine-induced immunity against Porcine circovirus type 2.

Porcine circovirus type 2 (PCV2) is essential but not sufficient for postweaning multisystemic wasting syndrome (PMWS) occurrence in pigs. The outcome of PCV2 infection depends on the specific immune responses that are developing during the infection. Diseased pigs are immunosupressed and unable to mount effective immune responses to clear the virus from circulation. In the final stage, PMWS-affected pigs suffer from extensive lymphoid lesions and altered cytokine expression patterns in peripheral blood mononuclear cells (PBMCs) and lymphoid organs. PCV2 infection can also be asymptomatic, demonstrating that not every infection will guarantee the occurrence of severe immunopathological disturbances. Asymptomatic animals have higher virus specific and neutralising antibody titres than PMWS-affected animals. Recent results have pointed out that the mechanisms by which PCV2 can affect the immune responses involve the induction of IL-10, virus accumulation into and modulation of plasmacytoid dendritic cells and the role of viral DNA in regulation of immune cell functions. Fourteen years after the first description of PMWS in Canada, efficient commercial vaccines against PCV2 are available. The vaccine success is based on activated humoral and cellular immune responses against PCV2. This review focuses on the recent research on immunological aspects during PCV2 infections and summarizes what is currently known about the vaccine-induced immunity.

Immunity conferred by an experimental vaccine based on the recombinant PCV2 Cap protein expressed in Trichoplusia ni-larvae.

Porcine circovirus type 2 (PCV2) vaccination has been recently included as a measure to control postweaning multisystemic wasting syndrome (PMWS) in the field. Aiming to obtain a more affordable vaccine to be extensively implemented in the field, a highly efficient non-fermentative expression platform based on Trichoplusia ni (T. ni) larvae was used to produce a baculovirus-derived recombinant PCV2 Cap protein (rCap) for vaccine purposes. Vaccination of pigs with rCap induced solid protection against PCV2 experimental infection, inhibiting both the viremia and the viral shedding very efficiently. The protection afforded by the rCap vaccine strongly correlated with the induction of specific humoral immune responses, even in the presence of PCV2-specific maternal immunity, although cellular responses also seemed to play a partial role. In summary, we have shown that rCap expressed in T. ni larvae could be a cost-effective PCV2 vaccine candidate to be tested under field conditions.

Porcine circovirus type 2 (PCV2) Cap and Rep proteins are involved in the development of cell-mediated immunity upon PCV2 infection.

The aim of the present study was to investigate the role of the capside (Cap) and replicase (Rep) proteins of Porcine circovirus type 2 (PCV2) as well as the whole PCV2 (both PCV2a and PCV2b genotypes) in the induction of cell-mediated immunity upon infection. At 6 weeks of age, six pigs were intranasally inoculated with the Stoon 1010 (Stoon) isolate (PCV2a) and seven with the Sp-7-10-54-13 (Sp) isolate (PCV2b). None of the pigs developed clinical disease but the Sp group had significantly higher proportion of pigs with PCV2-associated lesions and PCV2 load in tissues compared to the Stoon group. In both groups, development of IFN-gamma secreting cells (SC) in response to the whole PCV2 and Cap protein was detected by means of an ELISPOT from day 7 post-inoculation (PI) to the end of the study (21 days PI). Significant responses against Rep protein were only detected in Sp-inoculated pigs. No differences in ELISPOT results were seen when either PCV2a or PCV2b was used in vitro to recall peripheral blood mononuclear cells (PBMC) in any group. Stimulation of PBMC with the whole virus but not with Cap or Rep protein induced IL-10-SC in all pigs regardless of their PCV2 infection status, indicating an innate origin of this response. The results from this study demonstrate that PCV2-infected pigs developed cell-mediated immunity to Cap and Rep proteins and that, in the course of a sub-clinical infection, development and strength of such responses are possibly related to the levels of PCV2 replication.

Lack of transmission of porcine circovirus type 2 to weanling pigs by feeding them spray-dried porcine plasma.

An experiment was conducted to determine whether spray-dried porcine plasma containing 2·47 × 105 dna copies of porcine circovirus type 2 (pcv-2) could infect weanling pigs when fed to them. Five specific pathogen-free (spf) weanling pigs were fed ad libitum for 45 days a control diet and six pigs were fed a test diet containing 8 kg sdpp per 100 kg feed. The two groups were housed in separate biosecurity level-3 rooms. None of the pigs in either group developed any clinical signs or became pcv-2 viraemic or seroconverted.

Evaluation of cell-mediated immune responses against porcine circovirus type 2 (PCV2) Cap and Rep proteins after vaccination with a commercial PCV2 sub-unit vaccine.

This study investigated the development of cellular immunity to Porcine circovirus type 2 (PCV2) Cap and Rep proteins in pigs vaccinated with a commercial PCV2 genotype a (PCV2a) based sub-unit vaccine, before and after a heterologous challenge with a PCV2b isolate. At three weeks of age, 20 pigs were inoculated intramuscularly with either the vaccine product (V group, n=9) or phosphate buffered saline solution (PBS) (NV group, n=11). Three weeks after vaccination, pigs were challenged intranasally with PCV2b (V-C and NV-C groups) or PBS (V-NC and NV-NC groups). None of the pigs developed clinical signs during the whole experiment, but all NV-C and 3/5 V-C pigs developed viraemia. Vaccination induced the development IFN-γ-secreting cells in response to the Cap protein of PCV2, which appeared three weeks post-vaccination and increased after challenge. By that time, no significant differences were detected on PCV2 antibody titres between vaccinated and non-vaccinated pigs, although there were significant differences on day 7 post-challenge. PCV2-inoculation induced a cellular response against the Rep protein. Such response was significantly reduced or even absent in PCV2-inoculated pigs that were previously vaccinated (V-C group), presumably as a result of a lower PCV2 replication in vaccinated animals compared to non-vaccinated ones.

Simultaneous porcine circovirus type 2 and Mycoplasma hyopneumoniae co-inoculation does not potentiate disease in conventional pigs.

The aim of this study was to assess the effect of simultaneous experimental inoculation of porcine circovirus type 2 (PCV2; intranasal delivery) and Mycoplasma hyopneumoniae (Mhyo; transtracheal delivery) into conventional, seropositive 6-week-old piglets. Thirty-six male piglets were assigned randomly to four groups: control (n=6), PCV2 (n=6), Mhyo (n=12) and PCV2+Mhyo (n=12). Blood samples and faecal and nasal swabs were collected at 0, 7, 14 and 21 days post inoculation (dpi). No significant clinical signs attributable to PCV2 infection were observed during the experiment. Coughing was recorded in three pigs from the Mhyo group and six from the PCV2+Mhyo group. No significant differences in mean body weight and rectal temperature were observed between the groups. Mild microscopical lesions similar to those reported for post-weaning multisystemic wasting syndrome were observed in two PCV2 pigs and in one PCV2+Mhyo animal. Mhyo-compatible lung lesions were observed in 21/24 pigs inoculated with Mhyo (10 from the Mhyo group and 11 from the PCV2+Mhyo group). PCV2 was detected by in-situ hybridization in 3/12 PCV2 and in 4/12 PCV2+Mhyo animals. No significant differences in PCV2 load (serum and nasal and faecal swabs), duration of viraemia or antibody titre were detected between PCV2-inoculated groups. No significant differences in Mhyo load in nasal swabs, percentage of Mhyo-seropositive pigs and mean lung score was detected between Mhyo-inoculated groups. Under the conditions of the present study, concurrent inoculation of PCV2 and Mhyo did not result in potentiation of clinical signs and lesions attributed to either infection.

Pigs naturally exposed to porcine circovirus type 2 (PCV2) generate antibody responses capable to neutralise PCV2 isolates of different genotypes and geographic origins.

Porcine circovirus type 2 (PCV2) is the essential infectious agent for PCV2-systemic disease (PCV2-SD, formerly known as postweaning multisystemic wasting syndrome) and other pathological conditions. Recent studies indicated antigenic variability amongst different PCV2 isolates and suggested that single amino acid changes within the capsid protein determine differences in the level of neutralization by specific monoclonal antibodies. The objective of the present study was to examine the cross-reactivity of PCV2 antibodies induced in the context of a natural infection against different PCV2 isolates belonging to genotypes PCV2a and PCV2b. Sera taken from several farms from animals of varying health status (PCV2-SD and age-matched healthy pigs and a set of slaughter-aged animals) were assayed for neutralizing activity against four PCV2 isolates from both predominant genotypes (PCV2a and PCV2b) and of differing geographic origins (Europe and North-America). Results showed that most of studied pigs (79 out of 82) contained neutralizing antibodies (NA) able to neutralize all four studied viral strains. Overall, pigs had significantly higher NA titres against PCV2a than against PCV2b (P < 0.001). Accordingly, studied serums were able to better neutralize Burgos390L4 and Stoon-1010 strains (PCV2a) than L-33-Sp-10-54 and MO/S-06 strains (PCV2b) (P < 0.001). No differences between capabilities of seroneutralization of viruses from different geographic origin were observed. Present data suggests that sequence differences between PCV2 isolates translate to functional antigenic differences in viral neutralization in vivo.