Factors influencing the fate of <em>Salmonella</em> in the plant and soil environment

Abstract

Fresh fruits and vegetables are increasingly recognized as part of a healthy diet. However, disease outbreaks caused by Salmonella linked to fresh produce indicate that raw fruits and vegetables could be a vector for human pathogens. Contamination of fresh produce with Salmonella can occur along the whole production chain and one possible source is contaminated soil. Soil, rhizosphere and phyllosphere are heterogenous habitats where conditions can change quickly and survival of human pathogens in this environment is influenced by various biotic and abiotic factors. So far, the ecology of Salmonella in the environment is largely unknown and this thesis aimed at extending the knowledge of factors influencing the persistence of Salmonella in the soil-plant environment.
In this thesis, microcosm and greenhouse experiments were conducted to analyze factors influencing the fate of Salmonella under defined conditions. Lettuce (Lactuca sativa cv. Tizian) was used as a model plant and Salmonella enterica serovar Typhimurium strain LT2 as a model human pathogen. S. Typhimurium LT2 was inoculated into soil and its fate was monitored using cultivation-dependent and -independent methods.
Among the factors investigated in this thesis, the influence of preadaptation, meaning the conditions which the bacteria encounter before their introduction into the analyzed environment, on the survival of S. Typhimurium LT2 was investigated in soil microcosms. It was found that numbers of cultivable S. Typhimurium LT2 cells were significantly higher in the treatment with preadaptation than in the respective treatmment without preadaptation, showing that preadaptation can prolong survival of this strain in soil. These results indicate that the medium used to pre-grow Salmonella before inoculation into soil influenced the ability to survive in soil in a strain-specific manner. Therefore, conditions which bacteria encounter before their introduction into the environment might have a major influence on their survival and should be considered also with respect to comparability between studies.

Furthermore, in this thesis it was analyzed whether survival of S. Typhimurium LT2 inoculated into soil and colonization of lettuce plants by this strain is fostered by addition of sewage sludge or the plant-parasitic nematodes Meloidogyne hapla or Pratylenchus crenatus to soil. Unexpectedly, higher counts of cultivable S. Typhimurium LT2 were observed in untreated soil compared to soil with sludge. There were no differences in numbers of cultivable Salmonella in samples with nematode inoculation compared to control samples. No S. Typhimurium LT2 was detected in any of the leaf samples of lettuce plants grown in soil that was inoculated with S. Typhimurium LT2 after 21 and 49 days; neither in the samples where sludge or nematodes were added to the soil nor in the respective controls, indicating that the presence of sewage sludge or nematodes in soil did not foster colonization of lettuce plants by S. Typhimurium LT2. This suggests that the presence of Salmonella in soil even in relatively high concentrations and after addition of sewage sludge or plant-parasitic nematodes does not necessarily lead to a contamination of the aboveground part of plants by internalization through the roots.
The application of sewage sludge to soil was also analyzed with respect to the dynamics of the bacterial soil community as well as its associated mobile genetic elements (MGEs) and antibiotic resistance genes (ARGs) in a microcosm experiment over the course of four months. Comparison of the microbial diversity and relative abundances of ARGs in soil samples with sewage sludge compared to control soil revealed increased relative abundance of plasmids, integrons, transposons and resistance genes that in some cases were still detectable after four months. Soil bacterial communities showed reduced richness and diversity compared to control soil but differences decreasing with time. Resistance plasmids were captured directly from sewage sludge in Pseudomonas putida. The majority of the plasmids isolated belonged to the broad host range IncP-1 plasmids.
Altogether, this thesis extends the knowledge of factors influencing the fate of Salmonella in the soil - plant environment demonstrating that the fate of Salmonella in the environment of plants is very complex with many factors influencing its survival.

Upon author request this thesis is available as printed version only.