Browsing by Person "Detert, Katharina Margarete"

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    Uptake of pathogenic Escherichia coli into crop plants
    (2023) Detert, Katharina Margarete; Schmidt, Herbert
    Contaminations of fresh produce by enterohemorrhagic Escherichia coli (EHEC) have been an increasing source of human disease outbreaks in the past few decades. In particular, the large disease outbreak in 2011 in Germany has left many questions about the survival of EHEC in soil and the colonization of plants unanswered. Agricultural soil as a reservoir for EHEC strains represents an important contamination source for crop plants. The use of cattle manure for soil fertilization or contaminated irrigation water can result in the introduction of pathogens in fields. To prevent the transfer of pathogens into the food chain, the German fertilizer ordinance disallows the use of organic fertilizer 12 weeks before harvesting the crop plants. However, it is expected that EHEC survives for longer and therefore continue to pose a risk for crop plant contamination. In this study, the overall survival ability of the pathogenic E. coli O104:H4 strain C227/11Φcu, which represents a stx2a-negative derivative of the 2011 outbreak strain C227/11, in agricultural soil was investigated. Thereby, different environmental conditions, soil types and genetic factors of the bacterial strain were identified as influencing factors. Furthermore, the ability of E. coli O104:H4 strain C227/11Φcu to colonize lamb’s lettuce via the root system was investigated to demonstrate that contamination of soil can result in crop plant contamination. In the first publication, the survival of E. coli O104:H4 strain C227/11Φcu in soil microenvironments containing either diluvial sand or alluvial loam was investigated. Two different temperatures (4°C and 22°C) were used and the samples were incubated for more than 12 weeks. The study aimed to analyze whether cattle manure addition prolongs EHEC survival in the soil microenvironments. In the last step, the survival studies were performed using ΔrpoS and ΔfliC deletion mutant strains of C227/11Φcu. The results demonstrated that E. coli O104:H4 strain C227/11Φcu survived for at least 12 weeks in the soil microenvironment model. The survival rate was influenced by the soil type and the temperature. In more detail, the incubation at lower temperature prolonged the survival rate and pathogens were detected up to 20 weeks after inoculation. The application of contaminated cattle manure increased the survival ability at 22°C. Sigma factor RpoS was recognized as an important factor for soil survival. The rpoS deletion mutants showed significant reduction of the survival period while FliC did not influence the overall survival ability in these experiments. To investigate the influence of further genetic factors of the bacterial strain in more detail and thereby characterize all transcription activities, transcriptome analysis was performed in the second publication. Since the strain showed the best survival in alluvial loam at 4°C, this condition was used for the next experiments. To decrease the amount of competing soil microbiota, the soil samples were autoclaved prior inoculation. After inoculation and after 1 and 4 weeks, samples were taken for RNA isolation. The differential expression analysis was performed using the sample from time point 0 as control. The analysis revealed that stress response genes and genes of the primary metabolism were upregulated after 1 and 4 weeks. In addition, genes and gene sets for the uptake of various carbohydrates or amino acids were upregulated, indicating adaption to an environment with low nutrient availability. Moreover, the results of the second publication demonstrated that persistence of C227/11Φcu in soil is related with a complex interface of metabolic networks. The third paper focused on the colonization of lamb’s lettuce by E. coli O104:H4 strain C227/11Φcu via the root system. Surface-sterilized seeds were cultivated on Murashige-Skoog agar or in autoclaved agricultural soil and migration into the edible portions of the plants was analyzed after 2, 4 and 8 weeks. The results of this publication demonstrated that migration into the edible parts occurred when the surrounding agar or soil was contaminated. This highlighted the threat of plant contamination with pathogenic E. coli on the field as a result of soil contamination.