Effects of urbanization and air pollution on Platanus x hispanica bacterial communities and cultivation of strains with potential applications in phytoremediation

Abstract

Plant-associated bacterial communities are valuable to their plant hosts for assistance in maintaining the plant’s health and growth. Roadside and urban trees are exposed to various sources of stress such as air pollution and infrastructure constraints that challenge their health and longevity. Years of experience has determined which tree species are better suited for urban environments. One such tree is Platanus x hispanica, also known as Platanus x acerifolia or more commonly as a Plane tree. However, little is known about how the bacterial communities associated with P. x hispanica respond to increasing urbanization and air pollution as well as how such changes may influence its health in the long-term. Additionally, bacteria isolated from soil and plant tissues can be exploited for diverse biotechnological applications such as phytoremediation of other polluted sites. This thesis aims to improve our understanding of the effect urban environments are having on the bacterial communities associated with P. x hispanica and to investigate isolated bacteria for potential uses in promoting and protecting plant health. In the first section, we focused on bacteria associated with the leaves of P. x hispanica, otherwise known as the phyllosphere. To understand the impact urban air pollution has on the total epiphytic bacterial communities we conducted a biennial study on trees along gradients of urban intensity and the nearby traffic intensity in chapter 4. There were large differences in taxonomical diversity and composition between the two sampling years, but we determined that the pattern of dissimilarity between the sites was repeated along the gradient of urban intensity and some taxa were consistently more prevalent in either the rural or the urban samples. In addition, although the community composition of rural leaf epiphytes was more stable over time, the urban communities consistently had more human-associated bacteria and xenobiotic degradation pathways. The nearby traffic intensity did not appear to exert any significant influence on the epiphytes. We then cultivated diesel-vapor tolerant bacteria from the leaves to isolate strains with potential traits for plant growth promotion and biodegradation of hydrocarbons in chapter 5. Exposure to diesel vapor had less of an effect on the number of cultivable bacteria from the urban leaves than the rural. 60-70 colonies were isolated from each tree for further study, of which Bacillus was the most commonly identified genus. The highest percentage of bacterial strains with positive in vitro results for diesel degradation, tolerance to polycyclic aromatic hydrocarbons, and plant growth promotion (PGP) traits all came from the Inner-city leaves. Laccase-like multicopper oxidase (LMCO) genes were identified in nine strains and phenol oxidase activity in three strains, however only one strain had positive results for both. Two strains were tested for phenanthrene (PHE) degradation and although neither showed an appreciable decrease of PHE compared to the control flask after 12 days, both strains were able to increase the solubility of PHE in vitro, potentially making the compound more bioavailable for other microbes to degrade in future studies. The second section then concentrated on the below-ground bacterial communities of the same P. x hispanica trees. In chapter 7, we investigated the effects of urbanization and traffic intensity on the total bacterial communities of the bulk soil, rhizosphere soil and root endophytes. The diversity, taxonomic composition and predicted functional composition of endophytic bacteria in the roots from each location showed no significant differences. However, urban intensity did seem to have a negative effect on the alpha diversity and the abundance of Proteobacteria in the rhizosphere and bulk soil communities. The taxonomic and inferred functional ortholog composition was unique to the soil communities at each location. Differential abundance of the predicted metabolic pathways revealed the rural rhizosphere was abundant in PAH degradation, indicating that non-urban locations should not necessarily be overlooked when screening bacteria strains for PAH degraders. Alternatively, the urban samples were enriched in antimicrobial biosynthesis pathways, suggesting that city parks and street trees may have a greater need to recruit bacterial communities capable of aiding their defense against plant pathogens. Chapter 8 explores phenotypic and genotypic traits of bacteria cultured from the bulk soil, rhizosphere soil, and roots. The number of colony forming units was depressed in the rhizosphere soil of the Intersection and this could be the result of nonculturable bacteria or non-bacterial microorganisms playing a greater role at this location. In total, more than 350 bacteria were isolated and tested in vitro for PGP traits, of which 36 strains were selected. Pseudomonas and Stenotrophomonas were the most abundant genera in the collection. Seven strains showed moderate tolerance to the trace elements, copper, nickel, and zinc. 16 strains had LMCO genes and three strains had both the gene and the phenol oxidase activity. Sequencing revealed all of the LMCO amplicons to be most similar to either the CopA copper resistance protein family or the SufI conserved protein domain, both of which have laccase-like oxidase activity. Again, two strains were tested for phenanthrene (PHE) degradation and although neither showed an appreciable decrease of PHE compared to the control flask after 12 days, both strains were able to increase the solubility of PHE in vitro. In conclusion, we present new insights into the bacterial communities associated with Platanus x hispanica trees (Chapter 4 and 7). Urbanization and the nearby traffic volume appeared to influence community diversity, structure, and function differently depending on the sampling compartment (i.e. leaves, bulk soil, rhizosphere, roots, etc.). We also succeeded in isolating bacterial strains with potential applications as biotechnological tools for remediating polluted environments and protecting plant health (Chapter 5 and 8).