Mycoplasmal Pneumonia Of Swine Research Articles

Hijacking of Host Plasminogen by Mesomycoplasma (Mycoplasma) hyopneumoniae via GAPDH: an Important Virulence Mechanism To Promote Adhesion and Extracellular Matrix Degradation.

Mesomycoplasma hyopneumoniae is the etiological agent of mycoplasmal pneumonia of swine (MPS), which causes substantial economic losses to the world's swine industry. Moonlighting proteins are increasingly being shown to play a role in the pathogenic process of M. hyopneumoniae. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH), a key enzyme in glycolysis, displayed a higher abundance in a highly virulent strain of M. hyopneumoniae than in an attenuated strain, suggesting that it may have a role in virulence. The mechanism by which GAPDH exerts its function was explored. Flow cytometry and colony blot analysis showed that GAPDH was partly displayed on the surface of M. hyopneumoniae. Recombinant GAPDH (rGAPDH) was able to bind PK15 cells, while the adherence of a mycoplasma strain to PK15 was significantly blocked by anti-rGAPDH antibody pretreatment. In addition, rGAPDH could interact with plasminogen. The rGAPDH-bound plasminogen was demonstrated to be activated to plasmin, as proven by using a chromogenic substrate, and to further degrade the extracellular matrix (ECM). The critical site for GAPDH binding to plasminogen was K336, as demonstrated by amino acid mutation. The affinity of plasminogen for the rGAPDH C-terminal mutant (K336A) was significantly decreased according to surface plasmon resonance analysis. Collectively, our data suggested that GAPDH might be an important virulence factor that facilitates the dissemination of M. hyopneumoniae by hijacking host plasminogen to degrade the tissue ECM barrier. IMPORTANCE Mesomycoplasma hyopneumoniae is a specific pathogen of pigs that is the etiological agent of mycoplasmal pneumonia of swine (MPS), which is responsible for substantial economic losses to the swine industry worldwide. The pathogenicity mechanism and possible particular virulence determinants of M. hyopneumoniae are not yet completely elucidated. Our data suggest that GAPDH might be an important virulence factor in M. hyopneumoniae that facilitates the dissemination of M. hyopneumoniae by hijacking host plasminogen to degrade the extracellular matrix (ECM) barrier. These findings will provide theoretical support and new ideas for the research and development of live-attenuated or subunit vaccines against M. hyopneumoniae.

Genome‐wide detection of changes in allelic frequency in Landrace pigs selected for resistance to mycoplasma pneumonia of swine

Closed-pig line breeding could change the genetic structure at a genome-wide scale because of the selection in a pig breeding population. We investigated the changes in population structure among generations at a genome-wide scale and the selected loci across the genome by comparing the observed and expected allele frequency changes in mycoplasma pneumonia of swine (MPS)-selected pigs. Eight hundred and seventy-four Landrace pigs, selected for MPS resistance without reducing average daily gain over five generations, had 37,299 single nucleotide polymorphisms (SNPs) and were used for genomic analyses. Regarding population structure, individuals in the first generation were the most widely distributed and then converged into a specific group, as they were selected over five generations. For allele frequency changes, 96 and 14 SNPs had higher allele frequency changes than the 99.9% and 99.99% thresholds of the expected changes, respectively. These SNPs were evenly spread across the genome, and a few of these selected regions overlapped with previously detected quantitative trait loci for MPS and immune-related traits. Our results indicated that the considerable changes in allele frequency were identified in many regions across the genome by closed-pig line breeding based on estimated breeding value.

Development of a Multi-Epitope Vaccine for Mycoplasma hyopneumoniae and Evaluation of Its Immune Responses in Mice and Piglets.

Mycoplasma hyopneumoniae (Mhp), the primary pathogen causing Mycoplasma pneumonia of swine (MPS), brings massive economic losses worldwide. Genomic variability and post-translational protein modification can enhance the immune evasion of Mhp, which makes MPS prone to recurrent outbreaks on farms, even with vaccination or other treatments. The reverse vaccinology pipeline has been developed as an attractive potential method for vaccine development due to its high efficiency and applicability. In this study, a multi-epitope vaccine for Mhp was developed, and its immune responses were evaluated in mice and piglets. Genomic core proteins of Mhp were retrieved through pan-genome analysis, and four immunodominant antigens were screened by host homologous protein removal, membrane protein screening, and virulence factor identification. One immunodominant antigen, AAV27984.1 (membrane nuclease), was expressed by E. coli and named rMhp597. For epitope prioritization, 35 B-cell-derived epitopes were identified from the four immunodominant antigens, and 10 MHC-I and 6 MHC-II binding epitopes were further identified. The MHC-I/II binding epitopes were merged and combined to produce recombinant proteins MhpMEV and MhpMEVC6His, which were used for animal immunization and structural analysis, respectively. Immunization of mice and piglets demonstrated that MhpMEV could induce humoral and cellular immune responses. The mouse serum antibodies could detect all 11 synthetic epitopes, and the piglet antiserum suppressed the nuclease activity of rMhp597. Moreover, piglet serum antibodies could also detect cultured Mhp strain 168. In summary, this study provides immunoassay results for a multi-epitope vaccine derived from the reverse vaccinology pipeline, and offers an alternative vaccine for MPS.

Genome-wide association studies for production, respiratory disease, and immune-related traits in Landrace pigs

Identification of a quantitative trait locus (QTL) related to a chronic respiratory disease such as Mycoplasmal pneumonia of swine (MPS) and immune-related traits is important for the genetic improvement of disease resistance in pigs. The objective of this study was to detect a novel QTL for a total of 22 production, respiratory disease, and immune-related traits in Landrace pigs. A total of 874 Landrace purebred pigs, which were selected based on MPS resistance, were genotyped using the Illumina PorcineSNP60 BeadChip. We performed single nucleotide polymorphism (SNP)-based and haplotype-based genome-wide association studies (GWAS) to detect a novel QTL and to evaluate the possibility of a pleiotropic QTL for these traits. SNP-based GWAS detected a total of six significant regions in backfat thickness, ratio of granular leucocytes to lymphatic cells, plasma concentration of cortisol at different ages, and complement alternative pathway activity in serum. The significant region detected by haplotype-based GWAS was overlapped across the region detected by SNP-based GWAS. Most of these detected QTL regions were novel regions with some candidate genes located in them. With regard to a pleiotropic QTL among traits, only three of these detected QTL regions overlapped among traits, and many detected regions independently affected the traits.

Effect of Pregnane X Receptor on CYP3A29 Expression in Porcine Alveolar Macrophages during Mycoplasma hyopneumoniae Infection.

Simple SummaryIn the currently intense production process, infection of swine with Mycoplasma pneumonia is common in pig farms around the world, and reduction in the feeding efficiency and the growth rate of sick pigs causes considerable economic losses to the pig-rearing industry. Our study aimed to determine the molecular mechanism by which Mycoplasma hyopneumoniae induces inflammation in pigs. Our study showed that Mycoplasma hyopneumoniae can regulate the expression of CYP3A29 by upregulating PXR during the inflammatory response induced in porcine alveolar macrophages. These findings may provide useful information for breeding pigs that are resistant to disease.Mycoplasma hyopneumoniae (M. hyopneumoniae, Mhp) is the causative agent of mycoplasma pneumonia of swine (MPS). M. hyopneumoniae infection causes inflammation in pigs and leads to considerable economic losses in the pig industry. Pregnane X receptor (PXR) is a pluripotent gene regulatory protein that plays an important role in regulating cytochrome P-450 (CYP) in pigs in the context of inflammatory responses, drug metabolism, homeostasis, etc. We previously reported that cytochrome P450 3A29 (CYP3A29) expression was significantly upregulated in pigs infected with M. hyopneumoniae compared with healthy control pigs. This experiment mainly focused on identifying the role of PXR in the regulation of CYP3A29 and inflammatory factors after M. hyopneumoniae infection by establishing pig alveolar macrophage (PAM) cells in which PXR was overexpressed or silenced. Our results showed that the overexpression of PXR could significantly improve the protein and the mRNA expression levels of CYP3A29 with and without M. hyopneumoniae infection in PAM cells. After the expression of PXR was inhibited, protein and mRNA expression levels of CYP3A29 were significantly reduced with and without M. hyopneumoniae infection in PAM cells. Moreover, PXR can regulate the mRNA expression levels of IL-6 and IL-8 during M. hyopneumoniae infection of PAM cells. In conclusion, these results suggest that PXR positively regulates CYP3A29 expression during the inflammatory response caused by M. hyopneumoniae infection.

Immune responses induced by a combined vaccination with a recombinant chimera of Mycoplasma hyopneumoniae antigens and capsid virus-like particles of porcine circovirus type 2

BackgroundMycoplasma hyopneumoniae (Mhp) and porcine circovirus type 2 (PCV2) are two important pathogens causing Mycoplasma pneumonia of swine (MPS) and porcine circovirus diseases and porcine circovirus-associated diseases (PCVDs/PCVADs), respectively, and resulted in considerable economic loss to the swine industry worldwide. Currently, vaccination is one of the main measures to control these two diseases; however, there are few combination vaccines that can prevent these two diseases. To determine the effect of combination immunization, we developed capsid-derived (Cap) virus-like particles (VLPs) of PCV2 and a new recombinant chimera composed of the P97R1, P46, and P42 antigens of Mhp. Then we investigated the immune responses induced by the immunization with this combination vaccine in mice and piglets.ResultsThe high level antibodies against three protein antigens (P97R1, P46, and P42 of Mhp) were produced after immunization, up to or higher than 1:400,000; the antibody levels in Pro group continuously increased throughout the 42 days for all the antigens tested. The lymphocyte proliferative response in PCV2 group was stronger than that in PBS, VP, Mhp CV in mice. The antibody levels for Cap remained stable and reached the peak at 35 DAI. The IFN-γ and IL-4 in sera were significantly enhanced in the Pro group than that in the negative control-VP group on Day 14 and 28 post-the first immunization in piglets.ConclusionsAbove all, the combination immunization could induce humoral and cellular immune responses against all four antigens in mice and piglets. Therefore, our approach is a simple and effective vaccination strategy to protect pigs against MPS and PCVD/PCVAD.

IL-12p40 gene expression in lung and hilar lymph nodes of MPS-resistant pigs.

Mycoplasma pneumonia of swine (MPS) is caused by Mycoplasma hyopneumoniae (M.hp) and is a common chronic respiratory disease of pigs. Recently, a genetically selected variant of the Landrace pig (Miyagino L2) has a lower incidence of pulmonary MPS lesions. We investigated the pathological and immunological characteristics of MPS resistance in these pigs (n=24) by comparing with the normal landrace pig (control: n=24). The pathological MPS lung lesion score in MPS-selected landrace pigs was significantly lower than in the control. The gene expression of interleukin (IL)-12p40, which acts as a chemoattractant and a component of the bioactive cytokines IL-12 and IL-23, was significantly higher at the hilar lymph nodes, lung, and spleen in MPS-selected landrace pigs than in control landrace pigs, and these were negatively correlated with the macroscopic MPS lung lesion score. In summary, we demonstrate that resistance against MPS in Miyagino L2 pigs is associated with IL-12p40 up-regulation, in comparison with normal landrace pigs without the MPS vaccine. In addition, a comparative study of macroscopic MPS lung lesions and IL-12p40 gene expression in lung and hilar lymph nodes may lead to beneficial selection traits for the genetic selection for MPS resistance in pigs.

Mycoplasmal pneumonia of swine: Update and importance of the disease

The paper provides an overview of updated knowledge on this common disease with a strong impact on pig production efficiency. Mycoplasma hyopneumoniae (Mhp), the etiologic agent of the condition, is the smallest self-replicating bacteria, distinguished from most other bacteria by the lack of a cell wall. Colonization of the respiratory cilia is a unique feature of Mhp, which results in a persistent presence of Mhp in the airways and creates a population of infected asymptomatic animals that continually expose other pigs to infection. The economic consequences of a long-term persistence of infection within the herd include a chronic reduction in feed efficiency and increased opportunity for co-infection with other respiratory pathogens, leading to PRDC (porcine respiratory disease complex) syndrome. Chronic Mhp infection in the form of a respiratory disease with high morbidity and low mortality has been estimated to affect up to 70% of pigs worldwide. Diagnostic tools include clinical diagnosis, examination of gross lesions and histopathology. Mycoplasma hyopneumoniaespecific laboratory tests are critical to identify asymptomatic carriers or to detect antibodies as a sign of endemic infection. Tools that demonstrate the presence of Mhp rely on visualizing the organism (culture, FIA) or on detection of nucleic acid (PCR) or antigen specific for that pathogen (ELISA). Control of Mycoplasma hyopneumoniae has been attempted by several widely practiced methods. These include sow vaccination programmes, early vaccination of suckling piglets, use of antibiotics at peak transmission times, introduction of piglet flow in the all-in/all-out system and a segregated system of pig production. Several Mhp elimination protocols have been developed and can be implemented to eradicate this pathogen from the herd. One of them is a partial depopulation procedure, which consists in depopulation of pigs below 9 months of age. The other animals are subjected to medication.

Development of a Combined Genetic Engineering Vaccine for Porcine Circovirus Type 2 and Mycoplasma Hyopneumoniae by a Baculovirus Expression System.

Mycoplasma hyopneumoniae (Mhp) and porcine circovirus type 2 (PCV2) are the main pathogens for mycoplasmal pneumonia of swine (MPS) and post-weaning multisystemic wasting syndrome (PMWS), respectively. Infection by these pathogens often happens together and causes great economic losses. In this study, a kind of recombinant baculovirus that can display P97R1P46P42 chimeric protein of Mhp and the capsid (Cap) protein of PCV2 was developed, and the protein location was identified. Another recombinant baculovirus was constructed without tag proteins (EGFP, mCherry) and was used to evaluate the immune effect in experiments with BALB/c mice and domestic piglets. Antigen proteins P97R1P46P42 and Cap were expressed successfully; both were anchored on the plasma membrane of cells and the viral envelope. It should be emphasized that in piglet immunization, the recombinant baculovirus vaccine achieved similar immunological effects as the mixed commercial vaccine. Both the piglet and mouse experiments showed that the recombinant baculovirus was able to induce humoral and cellular responses effectively. The results of this study indicate that this recombinant baculovirus is a potential candidate for the further development of more effective combined genetic engineering vaccines against MPS and PMWS. This experiment also provides ideas for vaccine development for other concomitant diseases using the baculovirus expression system.

Association of swine leukocyte antigen class II haplotypes and immune-related traits in a swine line selected for resistance to mycoplasmal pneumonia.

By selective breeding for five generations, a Landrace line has been recently established to improve resistance to mycoplasmal pneumonia of swine (MPS), daily gain (DG), back fat thickness (BF), and plasma cortisol concentrations (COR). To clarify the involvement of swine leukocyte antigen (SLA) polymorphisms in the selection process, we investigated possible associations of 11 SLA-class II haplotypes with selected traits or immune parameters. Pigs with the low-resolution SLA haplotype Lr-0.23 or Lr-0.13, which increased in frequency with the passage of generations, had less severe pathological lesions of MPS, increased leukocyte phagocytic activity, and higher white blood cell counts. In contrast, Lr-0.12 and Lr-0.2, which decreased in subsequent generations, were weakly associated with more severe pathological lesions of MPS. Therefore, in the studied Landrace line, the Lr-0.23 and Lr-0.13 haplotypes are potentially useful genetic markers for selecting and breeding animals with less severe pathological lesions of MPS.

Mycoplasma pneumonia of swine (MPS) resistance and immune characteristics of pig lines generated by crossing an MPS pulmonary lesion selected Landrace line and a highly immune capacity selected Large White line.

To understand the influence of crossbreeding on Mycoplasma pneumonia of swine (MPS) resistance and immune characteristics, two crossbred lines were characterized. One crossbred line, LaWa, was generated by crossing the MPS pulmonary lesion selected Landrace line (La) and the highly immune-selected Large White line (Wa). The second crossbred line, LaWb, was generated by crossing the La line and the nonselected Large White line (Wb). The crossbred LbWb line (nonselected Landrace line × nonselected Large White line) and the La line were used as controls. The LaWa and LaWb lines had an intermediate level of MPS lung lesions between La and LbWb lines, although the difference was not statistically significant. After stimulation with sheep red blood cells (SRBCs), the LaWb and LaWa lines showed immune characteristics similar to that of the La line; the number of monocytes in peripheral blood increased, while B cells, T cells, secretion of SRBC-specific immunoglobulin G, and interleukin (IL)-13 decreased. Additionally, the number of natural killer (NK) cells and the expression of IL-4 and IL-17 were significantly higher in the LaWb and LaWa lines, respectively. These data suggested that crossbreeding of La and Wa lines resulted in the inheritance of some of the selected immune responses.

Correlated responses of respiratory disease and immune capacity traits of Landrace pigs selected for Mycoplasmal pneumonia of swine (MPS) lesion

Five generations of Landrace pigs selected for average daily gain, backfat thickness, Mycoplasmal pneumonia of swine (MPS) lesion score, and plasma cortisol levels, was executed to decrease the MPS lesion score. Genetic parameters and correlated genetic responses for respiratory disease and peripheral blood immune traits were estimated in 1395 Landrace pigs. We estimated the negative genetic correlation of MPS lesion score with phagocytic activity (PA) at 7 weeks of age (-0.67). The breeding values of PA at 7 weeks of age and 105 kg body weight and the correlated selection response of the ratio of granular leukocytes to lymphocytes at 105 kg body weight were significantly increased, and sheep red blood cell-specific antibody production (AP) was significantly decreased in a selection-dependent manner. Increasing of natural immunological indicators (e.g. PA) and decreasing of humoral immunological indicator (e.g. AP) were observed due to genetically decreasing MPS lesion score.

Correlated response of peripheral blood cytokines with selection for reduced mycoplasma pneumonia of swine lesions in Landrace pigs.

Mycoplasma pneumonia of swine (MPS) is responsible for significant economic losses in the swine industry. We selected Landrace pigs for reduced MPS pulmonary lesions over five generations, and measured concentrations of the following cytokines: interleukin (IL)-10, IL-13, IL-17, tumor necrosis factor (TNF)-α and interferon (IFN)-γ to estimate their correlation with MPS lesions. Sheep red blood cells (SRBC) were injected twice intramuscularly at 70 and 95 kg body weight. Blood serum samples were collected after 1 week of secondary SRBC inoculation and cytokine concentrations were analyzed by ELISA. Genetic parameters and breeding values were estimated. The heritability estimates of IL-10, IL-13, IL-17, TNF-α and IFN-γ were 0.20 ± 0.06, 0.12 ± 0.06, 0.27 ± 0.07, 0.20 ± 0.10 and 0.05 ± 0.03, respectively. Genetic correlations of IL-17 and TNF-α with pulmonary MPS lesions were high (-0.86 ± 0.13 and 0.69 ± 0.29, respectively) and those of IFN-γ and IL-13 with MPS lesions were moderately negative (-0.45). Through selection, the breeding values of IL-17 and IFN-γ increased substantially and those of TNF-α decreased. These results suggest that innate and cellular immunity are more important for the suppression of pulmonary lesions in MPS than humoral-mediated immunity, such as antibody response.

Immunogenic properties and mycoplasmal pneumonia of swine (MPS) lung lesions in Large White pigs selected for higher peripheral blood immune capacity.

Immunogenic properties and mycoplasmal pneumonia of swine (MPS) lung lesions were compared between the immunity-selected Large White line and the non-selected Large White line. The selected Large White line showed a higher level of pulmonary MPS lesions compared with the non-selected Large White line. Subsequent to vaccination, the percentage of natural killer cells and T cells (CD3(+) CD4(+) CD8(-) and CD3(+) CD4(-) CD8(+) T cells) were significantly increased in the non-selected line but remained unchanged in the immunity-selected Large White line. Secretion of Mycoplasma hyopneumoniae vaccine-specific immunoblogulin G and phagocyte activity in peripheral blood were significantly higher in the immunity-selected Large White line than in the non-selected line. Expression of interleukin (IL)-4 and IL-6 messenger RNA in hilar lymph nodes was significantly lower in the immunity-selected Large White line than in the non-selected line. However, expression of IL-10 in all immune tissues was significantly higher in the immunity-selected Large White line. These results suggest that the selection for high immunity was not effective in increasing resistance to MPS lung lesions.

Immunogenic properties of Landrace pigs selected for resistance to mycoplasma pneumonia of swine.

Mycoplasma pneumonia of swine (MPS) lung lesions and immunogenic properties were compared between a Landrace line that was genetically selected for reduced incidence of pulmonary MPS lesions, and a non‐selected Landrace line. The MPS‐selected Landrace line showed significantly lower degrees of pulmonary MPS lesions compared with the non‐selected Landrace line. When changes in immunity before and after vaccination were compared, the percentage of B cells in the peripheral blood of the MPS‐selected Landrace line was significantly lower than that of the non‐selected line. Furthermore, the concentration of growth hormone and the mitogen activity of peripheral blood mononuclear cells in the MPS‐selected Landrace line showed significantly (P < 0.05) lower increases after vaccination than the non‐selected line. Conversely, the concentration of peripheral blood interferon (IFN)‐γ and salivary immunoglobulin A (IgA) after Mycoplasma hyopneumoniae vaccination was significantly higher in the MPS‐selected Landrace line than in the non‐selected line. Gene expression of toll‐like receptor (TLR)2 and TLR4 was significantly higher in the MPS‐selected Landrace line in immune tissues, with the exception of the hilar lymph nodes. The present results suggest that peripheral blood IFN‐γ, salivary IgA TLR2, and TLR4 are important immunological factors influencing the development of MPS lesions.

Genetic selection for resistance to mycoplasmal pneumonia of swine (MPS) in the Landrace line influences the expression of soluble factors in blood after MPS vaccine sensitization

We recently developed a Landrace line that is resistant to mycoplasmal pneumonia of swine (MPS) infection by genetic selection for five generations, and we reported that the immunophenotype of this line is different from that of the non-selected line in terms of changes in peripheral blood leukocyte population after MPS vaccination. This study followed up previous findings demonstrating changes in soluble factors in blood, namely, hormones, Mycoplasma hyopneumoniae-specific immunoglobulin G (IgG), and cytokines. These two lines were injected with MPS vaccine on days -7 and 0 after blood sampling on those days, and blood samples were collected on days -14, -7, 0, 2, 7 and 14. We found changes in the levels of many hormones and cytokines in both lines. However, we found that only growth hormone (GH) and interferon (IFN)-γ levels were statistically different between these two lines. GH concentration was reduced (day 0) and IFN-γ concentration was increased (day 14) in the MPS-selected line compared with the non-selected line, despite unchanged IFN-γ messenger RNA expression in blood cells. Although detailed mechanisms underlying these phenotypes remain unsolved, these traits would be useful to improve MPS resistance in pig production and provide an insight into MPS infection.

Immunological characterization of peripheral blood leukocytes using vaccine for mycoplasmal pneumonia of swine (MPS) in swine line selected for resistance toMPS

This study was conducted to evaluate immunological changes in peripheral blood leukocytes in pigs that were genetically selected for their improved resistance to mycoplasmal pneumonia of swine (MPS), using MPS vaccine as an antigen. Twelve castrated MPS-selected Landrace pigs were compared with the same number of pigs from a nonselected line by using a time-course analysis at the hematological level. After the second sensitization with MPS vaccine, the percentages of B cells, CD4(+) T cells, and natural killer (NK) cells in total leukocytes were lower in the selected line than in the nonselected line, whereas the percentage of granulocytes in total leukocytes increased in the MPS-selected line. We also assessed the proliferative ability of peripheral blood mononuclear cells (PBMCs) stimulated with Mycoplasma hyopneumoniae, lipopolysaccharide or concanavalin A, and found that although the proliferative ability of the PBMC was not different between the two lines at a steady state, the nonselected line showed a significantly higher proliferative ability after sensitization with MPS vaccine than the selected line regardless of antigens used. These results thus indicate that the selection of pigs on the basis of MPS resistance changes their immunophenotype, and would give us beneficial information for the prevention of MPS infection.

A genome‐wide scan for quantitative trait loci affecting respiratory disease and immune capacity in Landrace pigs

Respiratory disease is the most important health concern for the swine industry. Genetic improvement for disease resistance is challenging because of the difficulty in obtaining good phenotypes related with disease resistance; however, identification of genes or markers associated with disease resistance can help in the genetic improvement of pig health. The purpose of our study was to investigate whether quantitative trait loci (QTL) associated with disease resistance were segregated in a purebred population of Landrace pigs that had been selected for meat production traits and mycoplasmal pneumonia of swine (MPS) scores over five generations. We analysed 1395 pigs from the base to the fifth generation of this population. Two respiratory disease traits [MPS scores and atrophic rhinitis (AR) scores] and 11 immune-capacity traits were measured in 630-1332 animals at 7 weeks of age and when the animal's body weight reached 105 kg. Each of the pigs, except sires in the base population, was genotyped using 109 microsatellite markers, and then, QTL analysis of the full-sib family population with a multi-generational pedigree structure was performed. Variance component analysis was used to detect QTL associated with MPS or AR scores, and the logarithm of odds (LOD) score and genotypic heritability of the QTL were estimated. Five significant (LOD > 2.51) and 18 suggestive (LOD > 1.35) QTL for respiratory disease traits and immune-capacity traits were detected. The significant QTL for Log-MPS score, located on S. scrofa chromosome 2, could explain 87% of the genetic variance of this score in this analysis. This is the first report of QTL associated with respiratory disease lesions.

Immunophenotype Characterization for Swine Selected Line, Which is Resistant for the Mycoplasma Pneumonia

Mycoplasma Pneumonia of swine (MPS) decreases the daily growth of pigs, and, co-infection with a virus sometimes causes severe pneumonia. Genetic selection of pigs resistant to the pulmonary MPS lesion might solve the economic loss due to MPS in animal production. Here, we examined the immunophenotype of Landrace line (Miyagino L2), genetically selected to reduce the incidence of pulmonary MPS lesion for 5 generations in Miyagi Prefecture Animal Industry Experiment Station. Although this line is expected to be resistant to the pulmonary MPS lesion, the biological characteristics of its immune function are not clear. We investigated details of the immunorelated phenotype of Miyagino L2 at the hematological and molecular biological level, including cytokine expression, and compared the results with that of non-genetically selected Landrace. Miyagino L2 showed decreased antigen-specific IgG and IgM production and increased CD8-positive T-cell population, and high levels of cortisol concentration, suggesting that the MPS-resistant phenotype is associated these immunological differences. Additionally, T-cell CD4 expression was highly correlated with the MPS expected breeding value. Although the detailed mechanisms underlying this high correlation remain unknown, our result suggested that the genetic selection of the expression level of CD4 might be useful to improve MPS resistance in pig production.

Immune Responses to the Attenuated Mycoplasma hyopneumoniae 168 Strain Vaccine by Intrapulmonic Immunization in Piglets

To investigate the immune responses to the attenuated Mycoplasma hyopneumoniae 168 strain vaccine, 8-15 d old piglets were immunized with M. hyopneumoniae 168 strain vaccine by intrapulmonic route. And the specific IgG antibody in serum, lymphoproliferation, IFNγ, and specific secretory IgA (SIgA) antibody in bronchoalveolar lavage fluid were detected on 30 and 60 d post-immunization (DPI), respectively. On 60 DPI, all the pigs except for those in health control group were challenged with a field M. hyopneumoniae strain JS. Necropsy was performed on 30 d post-challenge (DPC). The results showed that IFNγ and specific SIgA were stimulated on surface of respiratory tract after immunization. And peripheral blood mononuclear cells could also be proliferated about 1.81 and 2.12 fold on 30 and 60 DPI when stimulated by M. hyopneumoniae protein in vitro. However, no serum IgG antibody against M. hyopneumoniae was detected during the whole immune phage. After challenge, vaccinated pigs were observed with only very slight histological lesion in individual lobes. None of vaccinated pigs showed any clinical signs. While the unvaccinated pigs from challenge control group showed varying degrees of clinical sign and severe macroscopical lesion of mycoplasmal pneumonia of swine (MPS). The result suggested that the attenuated M. hyopneumoniae 168 strain vaccine inoculated by intrapulmonic route could activate the systemic cellular immunity, the local mucosal immunity and IFNγ secretion in respiratory tract to against M. hyopneumoniae infection in piglets.