Browsing by Person "Dietz, Stefanie"

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Microbiological and proteome analysis to gain insights into the pathogenesis of the highly adapted not-cultivated hemotrophic Mycoplasma suis
(2016) Dietz, Stefanie; Hölzle, Ludwig
The aims of this work were to establish a chronic pig infection model and to clarify unrecognized transmission pathways of M. suis. In addition, proteomics-based investigations of M. suis should be performed to improve the knowledge on the host pathogen interactions and host adaptation in IAP. Based on the succeeded and frequently applied splenectomized M. suis pig model (acute infection model) it was possible to establish a novel non-splenectomized M. suis pig model. This infection model enables the experimental investigation of the chronic M. suis infection. To this end, the piglets were infected intramuscularly with a highly virulent M. suis strain. Infected animals exhibited clinical signs (e.g. Morbus maculosus) including the typical cyclic course of chronic IAP. In the next step, potential transmission pathways of M. suis were analyzed during acute and chronic M. suis infection based on these two pig infection models. Feces, urine, air and dust as well as nasal, vaginal and saliva excretions were collected during the course of infection and examined for M. suis DNA by Rt-PCR. For the first time it was possible to detect M. suis in urine with and without erythrocytes as well as in nasal, vaginal and saliva excretions during acute and chronic infections. These results indicate blood-independent M. suis transmission via vaginal discharge, nasal excretions, saliva, and urine. The non-culturability limited the improvement of proteomic-based investigations of M. suis-related host-pathogen interactions. Therefore, we used modern and sophisticated proteome analysis to solve this problem. Blood samples from experimentally infected pigs at different time points of infection were investigated. For this, novel enrichment methods for M. suis proteins (especially membrane proteins) were established. These methods enabled an improved resolution of the protein expression profile of M. suis and thereby deeper insights into the pathogenesis of this microorganism. Despite of the missing cultivation system it was possible to identify more than 50% of the predicted M. suis proteins during acute infection. This identification ratio is similar to the one found in cultivable bacteria. Furthermore, the results of the proteome analysis indicate that nutrients such as glucose, hexose-6-phosphate, spermidine, putrescine, phosphate, amino acids, magnesium, potassium, sodium and iron are taken up by M. suis from the host leading to the high degree of host adaptation. Therefore, gained information on expressed M. suis proteins involved in transport are helpful in the establishment of an in vitro cultivation system in future. Particularly the supplementation of individual nutrients can play key functions in the media to support growth. Besides the M. suis proteome the acquired dataset firstly enables also the quantitative identification of Sus scrofa proteins differentially expressed during M. suis infection. This information can be used to unravel infection-relevant processes in the host in further studies.