The date was July 20, 1969, at 20:17 UTC. Two men made a historic landing that would become arguably the greatest technological achievement of the second half of the 20th Century. On this day, commander Neil Armstrong and lunar module pilot Buzz Aldrin landed the Apollo 11 Lunar Module Eagle on the surface of the Moon. Six hours and 39 minutes after landing, Neil stepped out of the Eagle and thus became the first human to walk on Moon’s surface, and 19 minutes later, Buzz joined him. The stakes were then raised for a more intense space race that occurred in the decades that followed. Fast-forward half a century later, on the chilly morning of November 5, 2019, two men set out on a 160-kilometer trip, not by spaceflight but through forests and thick bushes, in search of microbes that can revolutionize the way oil spills and other organic pollutants are cleaned up. Thus, the stage was set for events that would eventually crystallize into this research paper. But why was such a search-and-discovery trip needed?
Around the globe, industrialization and increasing demand for energy have led to an unabating exploitation of natural resources, especially fossil fuels. This often results in anthropogenic contamination of aquatic and terrestrial ecosystems, which threatens the survival of species. While large-scale marine spills often make headlines, most petroleum spills occur on land, with significant health and ecological impacts. Consequently, the future of humanity hangs in the balance if our assault on “mother nature” does not receive commensurate attention. Can science help in reversing these effects?
The remediation of petroleum-contaminated sites is essential to mitigating human and ecological risks. Sadly, traditional methods of remediation are very expensive and environmentally unfriendly. For example, it was estimated that one such method called “dig-and-haul” (excavation and translocation of contaminated soils for offsite treatments) would require over US$1 million per hectare. No wonder many contaminated sites are left as they are, or their rehabilitation is postponed. In contrast, the United States Environmental Protection Agency (EPA) indicated that implementing bioremediation will result in cost savings of 50-80% over traditional methods. One eco-friendly bio-based remediation strategy is rhizoremediation – the degradation of pollutants in the rhizosphere (the area surrounding the plant roots) through microbial activity.
Rhizoremediation stands out as an integrated plant-microbe endeavor. During this process, the plant roots secrete exudates such as sugars and amino acids that stimulate the growth and activity of rhizospheric microbes. However, the slow metabolic activity of the indigenous microorganisms often leads to long remediation times, thereby limiting the effectiveness of this approach. To address this shortfall, there is a growing interest in the inoculation of pre-cultivated microbes able to enhance the growth of host plants and speed up biodegradation as they utilize the organic pollutants for carbon and energy source. In turn, the root exudates released by host plants provide a constant flow of nutrients to the associated bacteria, enabling continuous biodegradation of contaminants. This synergistic relationship has been described as the ecological driver of rhizoremediation. Although some studies have isolated microbial consortia for bioremediation, the difficulty in replicating most consortia creates an urgent need for single culturable strains suited to environmental biotechnological applications. Where can such microbes be found? This takes us to the trip described at the outset.
Scientists generally agree that microorganisms present in contaminated environments often manifest the environmental adaptability and genetic capability required for effective breakdown of contaminants. Hence, our search for novel strains of bacteria capable of plant growth promotion and hydrocarbon degradation took us to the crude oil tar pit in Wietze, Germany. Wietze is the site of the first commercial oil exploration in Germany dating back to 1859. It accounted for almost 80% of German oil production between 1900 and 1920. Although oil production ceased in Wietze some decades later, it continues to witness oil seepages. Therefore, it was the ideal place to mine for potential hydrocarbon-degrading microorganisms that can work in synergy with Medicago sativa plants. But why Medicago sativa?
Our earlier studies revealed that M. sativa, also known as alfalfa or lucerne can relatively withstand hydrocarbon toxicity under low concentrations of diesel fuel. Therefore, through genomic analyses and greenhouse-based experiments, we examined the synergistic interactions of Medicago sativa and Paraburkholderia tropica WTPI1 for enhanced rhizoremediation of diesel-contaminated soils. Whole genome analyses revealed that among the isolated single species, P. tropica WTPI1 has the highest potential for plant growth promotion and hydrocarbon degradation. Greenhouse-based experiments confirmed that the inoculation of M. sativa with P. tropica WTPI1 led to a 99% increase in plant biomass. Organic geochemical analysis revealed that 96% of all the distinctive diesel fuel hydrocarbons, including C10–C25 n-alkanes, branched alkanes, cycloalkanes and aromatic hydrocarbons were degraded in the M. sativa + P. tropica treatment within just 60 days. This is significantly greater than the 49% degradation observed under natural attenuation. Interestingly, biodegradation parameters (Pr/nC17, Ph/nC18, nor-Pr/nC16 and UCM/TPH) revealed that the removal of petroleum hydrocarbons in the different treatments resulted from biodegradation.
Metagenome analysis of residual soils revealed that the inoculated microbe (P. tropica) prospered in the soils. This is of great importance since a major drawback in the development of plant growth-promoting rhizobacteria has always been the failure of inoculated microorganisms to effectively thrive against indigenous microbes. It is important to mention here that owing to the complexity of soils and environmental variables, the viability of P. tropica WTPI1 may vary under different conditions. Nevertheless, our results will prove beneficial for biotechnological application of P. tropica WTPI1 for plant growth promotion and most importantly for environmental remediation of organic pollutants.