Interactions between sewage sludge and the survival of pathogenic bacteria in soil

Abstract

Sewage sludge is a potentially valuable resource that can enhance both the structure and fertility of soil. However, it can also harbour enteric pathogens which pose a significant socio-economic risk to society. Therefore it is important to understand the factors that govern the persistence of such pathogens in soil, when co-introduced with sewage sludge, in order to mitigate risk and to further avail of such a valuable resource. This research aimed to clarify how microbial activity and the presence of sewage sludge would influence the persistence of co-introduced enteric pathogens in soil. It was theorised that the addition of sewage sludge to soil would cause the formation of organic matter (OM) and nutrient-rich niches. Such niches, in turn, would encourage the enhanced activity of the local soil microbial community, instigating greater competition for local resources, i.e. a hot spot of microbial activity that would lead to a decline in the introduced enteric pathogens. It was also hypothesised that the interface between the soil and sewage sludge may influence such interactions, as the physicochemical characteristics could affect the extent of exposure and subsequent interactions between enteric pathogens and the soil microbial community. These theories were investigated using four different perspectives that linked closely with each other. In initial studies, two cohorts of microcosms consisting of different proportions of sewage sludge to soil were inoculated with either E. coli or S. Dublin and destructively sampled over a 42 day period. E. coli prevailed at greater numbers when inoculated directly into soil and sewage sludge, whilst it declined to the greatest extent within mixed microcosms containing 25% sludge. All treatments containing S. Dublin appeared to decline at a similar rate, which was more linear than the decline observed within treatments inoculated with E. coli. From these findings, it can be concluded that there are no direct relationships between the proportion of sludge to soil and its affect on pathogen survival. A subsequent experiment implemented a similar treatment strategy, whilst using indigenous sewage sludge E. coli. The use of this microbe provided data which was more suited to the original premise of this work, as under such scenarios it would be indigenous sewage sludge E. coli that would be of concern. Therefore, microcosms consisting of different proportions of sewage sludge, containing indigenous E. coli, were destructively sampled over a 56 day period. The indigenous sewage sludge E. coli exhibited a more consistent linear decline after the first week. However, the indigenous E. coli were again not significantly affected by different proportions of sewage sludge to soil. It was theorised that this lack of variation in response to varying proportions of sewage sludge to soil may have been associated with a lack of available substrate within the system, or some form of partitioning effect between soil and sewage sludge matrices, which prevented the microbial communities from interacting. To further develop these concepts, the effect of two contrasting substrate amendments and their location (either sewage sludge, soil or within both matrices) was also investigated in relation to the persistence of sewage sludge-derived E. coli. Microcosms consisting of both pure samples and mixtures of sewage sludge or soil were inoculated with sewage sludge-derived E. coli and destructively sampled over a 42 day period. Respired CO2 and microbial carbon were also quantified. The addition of a simple substrate, glucose, instigated a peak in microbial respiration and accelerated the decline of sewage sludge-derived E. coli and also marginally increased the microbial biomass. This is similar to the original concept proposing that a hot spot of microbial activity could instigate pathogen die-off. In contrast, amendment with a more complex substrate, yeast extract, had little effect on the decline of sewage sludge-derived E. coli. Nor did respiration increase immediately after amendment. There was also no observable partitioning effects between soil or sewage sludge with either amendment. This suggests that a lack of available substrate could influence microbial dynamics and thus the decline of E. coli. To further explore this phenomenon the repeated addition of glucose and its effect on the survival of sewage sludge-derived E. coli was investigated. It aimed to highlight the impact of sustained competition for resources on persistence, whilst mimicking the recurrent input of carbon that occurs in plant/soil systems. Microcosms consisting of both pure and mixtures of sewage sludge or soil were inoculated with sewage sludge-derived E. coli and destructively sampled over a period of 105 days. Respired CO2 and microbial carbon were again analysed. It was found that the repeated addition of glucose did not cause a significant decline in the survival of sewage sludge-derived E. coli. Notably, some small increases in E. coli numbers were observed after the second and third amendments of glucose. Overall, these findings suggest that hot spots of activity can instigate a decline in enteric pathogens, though such interactions are dependent upon the availability and quantity of nutrients and organic carbon within the matrices. These findings could aid in developing the use of amendments in sewage sludge that would minimise the survival of enteric pathogens in soil. They also provide a framework which pinpoints the factors that should be considered when investigating the persistence of enteric pathogens in the soil environment. Such amendments and knowledge pertaining to the key factors in the survival of enteric pathogens could further decrease the social and economic risk which the use of sewage sludge poses when used in agricultural systems.