Endophyte-enhanced phytoremediation of DDE-contaminated using Cucurbita pepo: A field trial.

Although the use of the pesticide 2,2-bis(p-chlorophenyl)-1,1,1-trichloroethane (DDT) was banned from the mid-1970s, its most abundant and recalcitrant degradation product, 2,2-bis(p-chlorophenyl)-1,1-dichloro-ethylene (DDE), is still present in terrestrial and aquatic ecosystems worldwide. Zucchini (Cucurbita pepo ssp. pepo) has been shown to accumulate high concentrations of DDE and was proposed for phytoremediation of contaminated soils. We performed a field trial covering a full plant life cycle. C. pepo plants inoculated with the plant growth-promoting endophytic strains Sphingomonas taxi UH1, Methylobacterium radiotolerans UH1, Enterobacter aerogenes UH1, or a consortium combining these 3 strains were grown on a DDE-contaminated field for 100 days. The effects of these inoculations were examined at both the plant level, by evaluating plant weight and plant DDE-content, and at the level of the cultivable and total endophytic communities. Inoculating plants with S. taxi UH1, M. radiotolerans UH1, and the consortium increased plant weight. No significant effects of the inoculations were observed on DDE-concentrations in plant tissues. However, the amount of DDE accumulated by C. pepo plants per growing season was significantly higher for plants that were inoculated with the consortium of the 3 strains. Therefore, inoculation of C. pepo with DDE-degrading endophytes might be promising for phytoremediation applications.

Exposure of Cucurbita pepo to DDE-contamination alters the endophytic community: A cultivation dependent vs a cultivation independent approach.

2,2-bis(p-chlorophenyl)-1,1-dichloro-ethylene (DDE) is the most abundant and persistent degradation product of the pesticide 2,2-bis(p-chlorophenyl)-1,1,1-trichloroethane (DDT) and is encountered in contaminated soils worldwide. Both DDE and DDT are classified as Persistent Organic Pollutants (POPs) due to their high hydrophobicity and potential for bioaccumulation and biomagnification in the food chain. Zucchini (Cucurbita pepo ssp. pepo) has been shown to accumulate high concentrations of DDE and other POPs and has been proposed as a phytoremediation tool for contaminated soils. The endophytic bacteria associated with this plant may play an important role in the remedial process. Therefore, this research focuses on changes in endophytic bacterial communities caused by the exposure of C. pepo to DDE. The total bacterial community was investigated using cultivation-independent 454 pyrosequencing, while the cultivable community was identified using cultivation-dependent isolation procedures. For both procedures, increasing numbers of endophytic bacteria, as well as higher diversities of genera were observed when plants were exposed to DDE. Several bacterial genera such as Stenotrophomonas sp. and Sphingomonas sp. showed higher abundance when DDE was present, while, for example Pseudomonas sp. showed a significantly lower abundance in the presence of DDE. These findings suggest tolerance of different bacterial strains to DDE, which might be incorporated in further investigations to optimize phytoremediation with the possible use of DDE-degrading endophytes.

Comparison between cultivated and total bacterial communities associated with Cucurbita pepo using cultivation-dependent techniques and 454 pyrosequencing.

Endophytic bacteria often have beneficial effects on their host plants that can be exploited for bioremediation applications but, according to the literature, only 0.001-1% of all endophytic microbes should be cultivable. This study compared the cultivated endophytic communities of the roots and shoots of Cucurbita pepo with the total endophytic communities as determined by cultivation-dependent techniques and 454 pyrosequencing. The ten most abundant taxa of the total communities aligned well with the cultivated taxa; however, the abundance of these taxa in the two communities differed greatly. Enterobacter showed very low presence in the total communities, whereas they were dominantly present in the cultivated communities. Although Rhizobium dominated in total root and shoot communities, it was poorly cultivable and even then only in growth media containing plant extract. Since endophytes likely contribute to plant-growth promotion, cultivated bacterial strains were tested for their plant-growth promoting capabilities, and the results were correlated with their abundance in the total community. Bacillus and Pseudomonas showed promising results when considering cultivability, abundance in the total community and plant-growth promoting capability. This study demonstrated that, although a limited number of bacterial genera were cultivable, current cultivation-dependent techniques may be sufficient for further isolation and inoculation experiments that aim to improve phytoremediation efficiency.

Optimization of isolation and cultivation of bacterial endophytes through addition of plant extract to nutrient media.

Many endophytes have beneficial effects on plants and can be exploited in biotechnological applications. Studies hypothesize that only 0.001-1% of all plant-associated bacteria are cultivable. Moreover, even after successful isolations, many endophytic bacteria often show reduced regrowth capacity. This research aimed to optimize isolation processes and culturing these bacteria afterwards. We compared several minimal and complex media in a screening. Beside the media themselves, two gelling agents and adding plant extract to media were investigated to enhance the number and diversity of endophytes as well as the growth capacity when regrown after isolation. In this work, 869 medium delivered the highest numbers of cultivable bacteria, as well as the highest diversity. When comparing gelling agents, no differences were observed in the numbers of bacteria. Adding plant extract to the media lead to a slight increase in diversity. However, when adding plant extract to improve the regrowth capacity, sharp increases of viable bacteria occurred in both rich and minimal media.