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Array-Based Genomic Comparative Hybridization Analysis of Field Strains of Mycoplasma hyopneumoniae

ABSTRACT: Mycoplasma hyopneumoniae is the causative agent of porcine enzootic pneumonia and a major factor in the porcine respiratory disease complex. A clear understanding of the mechanisms of pathogenesis does not exist although it is clear that M. hyopneumoniae adheres to porcine ciliated epithelium by action of a protein called P97. Previous studies have shown variation in the gene encoding the P97cilium adhesin within different strains of M. hyopneumoniae, but the extent of genetic variation among field strains across the genome is not known. Since M. hyopneumoniae is a worldwide problem, it is reasonable to expect that a wide range of genetic variability may exist given all of the different breed and housing conditions. This variation may impact the overall virulence of a single strain. Using microarray technology, this study examined potential variation of fourteen field strains in comparison to strain 232 on which the array was based. Genomic DNA was obtained, amplified with TempliPhi™, and labeled indirectly with Alexa dyes. Post genomic hybridization, the arrays were scanned and data analyzed using a linear statistical model. Results indicate that genetic variation could be detected in all fourteen field strains but across different loci, suggesting that variation occurs throughout the genome. Fifty-nine percent of the variable loci were hypothetical genes. Twenty-two percent of the lipoprotein genes showed variation in at least one field strain. A permutation test identified a location in M. hyopneumoniae genome where spatial clustering of variability between the field strains and strain 232 exists. Keywords: CGH, Mycoplasma Hyopneumoniae

ORGANISM(S): Mesomycoplasma hyopneumoniae

PROVIDER: GSE8306 | GEO | 2008/01/01

SECONDARY ACCESSION(S): PRJNA101275

REPOSITORIES: GEO

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Project description:Mycoplasma hyopneumoniae is the causative agent of porcine enzootic pneumonia and a major factor in the porcine respiratory disease complex. A clear understanding of the mechanisms of pathogenesis does not exist although it is clear that M. hyopneumoniae adheres to porcine ciliated epithelium by action of a protein called P97. Previous studies have shown variation in the gene encoding the P97cilium adhesin within different strains of M. hyopneumoniae, but the extent of genetic variation among field strains across the genome is not known. Since M. hyopneumoniae is a worldwide problem, it is reasonable to expect that a wide range of genetic variability may exist given all of the different breed and housing conditions. This variation may impact the overall virulence of a single strain. Using microarray technology, this study examined potential variation of fourteen field strains in comparison to strain 232 on which the array was based. Genomic DNA was obtained, amplified with TempliPhiâ¢, and labeled indirectly with Alexa dyes. Post genomic hybridization, the arrays were scanned and data analyzed using a linear statistical model. Results indicate that genetic variation could be detected in all fourteen field strains but across different loci, suggesting that variation occurs throughout the genome. Fifty-nine percent of the variable loci were hypothetical genes. Twenty-two percent of the lipoprotein genes showed variation in at least one field strain. A permutation test identified a location in M. hyopneumoniae genome where spatial clustering of variability between the field strains and strain 232 exists. TempliPhiâ¢ amplified DNA samples from field isolates were compared to control strain 232 using a two-color experimental microarray design. Independent samples from one isolate labeled with one dye were paired with control samples labeled with the alternate dye; the samples were mixed and hybridized to the microarray. For nine of the fourteen isolates (95MP1501, 95MP1502, 95MP1503, 95MP1504, 95MP1508, 95MP1509, 97MP0001, 00MP1301, and 05MP2301), four independent field isolate DNA samples were paired with four independent DNA samples from control 232. In two of the four arrays, the control sample was labeled with Alexa 555 dye and compared to the field isolate sample labeled with Alexa 647 dye (Molecular Probes, Inc., Eugene, Ore.). The dye assignment to control and treated samples was reversed for the other two arrays (dye swap). The arrays were hybridized under identical conditions as described below. This procedure was repeated for isolates 95MP1510 for a total of four arrays, where the control sample was labeled with Alexa 647 dye in three of the arrays and Alexa 555 dye for the fourth array. For isolates 95MP1505, 95MP1506 and 95MP1507, a total of five arrays each, including two dye swaps, were done; and for isolate 00MP1502, a total of six arrays were done with the control labeled with Alexa 555 dye for four of the arrays and Alexa 647 for the other two arrays.

Project description:Kamminga2017 - Metabolic model of Mycoplasma hyopneumoniae growth This model is described in the article: Metabolic modeling of energy balances in Mycoplasma hyopneumoniae shows that pyruvate addition increases growth rate. Kamminga T, Slagman SJ, Bijlsma JJE, Martins Dos Santos VAP, Suarez-Diez M, Schaap PJ. Biotechnol. Bioeng. 2017 Jun; : Abstract: Mycoplasma hyopneumoniae is cultured on large-scale to produce antigen for inactivated whole-cell vaccines against respiratory disease in pigs. However, the fastidious nutrient requirements of this minimal bacterium and the low growth rate make it challenging to reach sufficient biomass yield for antigen production. In this study, we sequenced the genome of M. hyopneumoniae strain 11 and constructed a high quality constraint-based genome-scale metabolic model of 284 chemical reactions and 298 metabolites. We validated the model with time-series data of duplicate fermentation cultures to aim for an integrated model describing the dynamic profiles measured in fermentations. The model predicted that 84% of cellular energy in a standard M. hyopneumoniae cultivation was used for non-growth associated maintenance and only 16% of cellular energy was used for growth and growth associated maintenance. Following a cycle of model-driven experimentation in dedicated fermentation experiments, we were able to increase the fraction of cellular energy used for growth through pyruvate addition to the medium. This increase in turn led to an increase in growth rate and a 2.3 times increase in the total biomass concentration reached after 3-4 days of fermentation, enhancing the productivity of the overall process. The model presented provides a solid basis to understand and further improve M. hyopneumoniae fermentation processes. Biotechnol. Bioeng. 2017;9999: 1-9. © 2017 Wiley Periodicals, Inc. This model is hosted on BioModels Database and identified by: MODEL1704250001. To cite BioModels Database, please use: BioModels Database: An enhanced, curated and annotated resource for published quantitative kinetic models. To the extent possible under law, all copyright and related or neighbouring rights to this encoded model have been dedicated to the public domain worldwide. Please refer to CC0 Public Domain Dedication for more information.

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Array-Based Genomic Comparative Hybridization Analysis of Field Strains of Mycoplasma hyopneumoniae

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