Detection, Isolation And Comparison Of Different Strains Of Mycoplasma Hyorhinis And The Primary Study Of An Experimental Infection Model

Despite the absence of a cell wall and a small genome, the genera mycoplasma has been able to cause chronic infections in different hosts. High infection rates of Mycoplasma hyorhinis in swine may be attributed to the fact that the environment of pigs has changed to a greater extent due to the evolution of pork production systems towards specialization and intensive confinement. Therefore, transmission of M. hyorhinis by nose-to-nose contacts from infected excreting pig or by airborne transmission inside the barns or neighboring farms is common. Although M. hyorhinis is part of normal flora of the upper respiratory tract of pigs it can also be pathogenic. It is the etiological agent of arthritis, pleuritis, pericarditis and peritonitis and implicated with enzootic pneumonia in sucking pigs. Most infections are subclinical and manifestation of clinical signs may vary although co-infection with cohorts of other pathogens is also common. Strain virulence variations, weather changes, concomitant pathogens and the host immune system are some of the mechanisms that dictate its systemic nature.At the genomic level, great heterogeneity has been reported among mycoplasma isolates from different herds. It is possible to find different strains in the same animal in a farm, which could differ in virulence or cause interferences and cross reactions. The M. hyorhinis genome varies in some components concerned with metabolism and evasion of the host’s immune system, which might contribute to its growth aggressiveness.The current study has integrated different aspects that are ultimately geared to controlling M. hyorhinis in herds.(1) It addresses the detection of the main porcine pathogens; M. hyorhinis and M. hyopneumoniae. Nasal swabs clinical samples of 7-35 days pigs were collected from different herds in Jiangsu Province, China and their prevalence was determined after the detection of the pathogen’s DNA by nested PCR (nPCR) assay. Our findings suggest that the prevalence of M. hyorhinis is about 70.9% while that of M. hyopneumoniae is about 13.5%.7.5% of the nasal swabs samples were positive of both M. hyopneumoniae and M. hyorhinis.(2) Different strains of M. hyorhinis were isolated after detection of the pathogen in the lung samples and exudates of infected pigs with typical pneumonic lung lesions collected from different slaughterhouses. The isolates were identified as M. hyorhinis by microbiological culture and nPCR and then subjected to antimicrobial profiling against 12 antimicrobial agents that belong to different classes and are commonly used in the swine industry for the control of mycoplasmosis. (3) The field isolates were subsequently used to conduct immunogenicity testing in mice models to determine which one of them elicits the highest immune response. Mice were inoculated with antigens prepared from different strains of M. hyorhinis, then IgG antibody titers over a specific period of time was quantified by ELISA by using a recombinant detecting protein of VlpA-G as the coating antigen. The level of IgG elicited by the strain 100928 was more hence the isolate is highly immunogenic, followed by strains 11022402 and 11081111 respectively.(4) A challenge model was designed where pathogenicity studies were conducted by intratracheal challenge with a combination of a pure culture of different strains of M. hyorhinis in Specific Pathogen Free (SPF) pigs in order to determine and understand the mechanism of infection as well as form the basis of research on the pathogenesis of M. hyorhinis. The pigs were observed for clinical signs and presence of pathological lesions specific for M. hyorhinis infection. In an effort to induce pneumonic lesions, an inflammatory lung tissue was also used in combination with a pure culture of M. hyorhinis. This produced microscopic lesions as revealed by Hematoxylin and Eosin (H&E) staining. However, co-infection of M. hyorhinis and M. hyopneumoniae was noted in this study. Subsequently, screening the most virulent strain was conducted where chicken embryos experimentally inoculated with a standardized dose of a pure culture of M. hyorhinis. Mortality of the chicken embryos was observed and the most virulent strain was screened out for the purpose of future studies of inducing M. hyorhinis pathogenesis in swine. The most virulent strain was observed to be 11051207 followed by 100928 and 11081111.Although further investigation is warranted, this study has successfully achieved its objectives by detecting, isolating and characterizing different isolates of M. hyorhinis by assessing their virulence through pathogenicity studies in pigs and chicken embryos as well as by testing the immunogenicity of the isolates in mice models.