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The development of techniques to non-destructively monitor allelochemical dynamics in soil using polydimethylsiloxane (PDMS) microtubing (silicone tubing microextraction, or STME) provides a means to test important ecological hypotheses regarding the roles of these compounds in plant-plant interactions. The objective of this study was to investigate the impact of intra- and interspecific competition on the exudation of thiophenes by marigolds (Tagetes patula L.). Marigolds were grown at a density of 1, 3 and 5 plants in pots (8.75 × 8.75 cm) containing two STME samplers. An additional treatment included one marigold surrounded by four velvetleaf (Abutilon theophrasti L.) plants. Marigold roots released two primary thiophenes, 3-buten-1-ynyl)-2,2'-bithienyl and α-terthienyl, which are readily absorbed by silicone microtubing. Thiophene exudation was monitored over the period 15-36 days after planting, at 2-5 day intervals. At the end of the study, root and soil samples were also analyzed for thiophene content. Thiophene production per plant increased over time, and thiophene release was strongly correlated with plant size. These results indicate that thiophene release in this study was passively controlled by resource availability. However, poor growth of velvetleaf plants competing with marigold suggests that thiophenes negatively influenced velvetleaf growth. This study, then, provides indirect evidence that thiophene exudation is insensitive to neighbor identity but differentially effective in inhibiting the growth of heterospecific neighbors.

Understanding allelopathy has been hindered by the lack of methods available to monitor the dynamics of allelochemicals in the soil. Previous work has demonstrated the feasibility of using polydimethylsiloxane (PDMS) microtubing (silicone tubing microextraction, or STME) to construct sampling devices to monitor the release of lipophilic allelochemicals from plant roots. The objective of this study was to use such sampling devices to intensively monitor thiophene fluxes beneath marigolds over several weeks to gain insight into the magnitude of temporal and spatial heterogeneity in these fluxes. Marigolds were grown in rhizoboxes (20.5 x 20.5 x 3.0 cm) with 16 individual STME samplers per box. Thiophene sampling and HPLC analysis began 45 days after planting. At the end of the study, roots around each sampler were analyzed by HPLC. Results confirmed the tremendous spatial and temporal heterogeneity in thiophene production seen in our previous studies. STME probes show that thiophene concentrations generally increase over time; however, these effects were sampling-port specific. When sampling ports were monitored at 12 h intervals, fluxes at each port ranged from 0 to 2,510 ng day(-1). Fluxes measured over daylight hr averaged 29 % higher than those measured overnight. Fluxes were less than 1 % on average of the total thiophene content of surrounding roots. While the importance of such heterogeneity, or "patchiness", in the root zone has been recognized for soil nutrients, the potential importance in allelopathic interactions has seldom been considered. The reasons for this variability are unclear, but are being investigated. Our results demonstrate that STME can be used as a tool to provide a more finely-resolved picture of allelochemical dynamics in the root zone than has previously been available.

Allelopathy, a phenomenon where compounds produced by one plant limit the growth of surrounding plants, is a controversially discussed factor in plant-plant interactions with great significance for plant community structure. Common mycorrhizal networks (CMNs) form belowground networks that interconnect multiple plant species; yet these networks are typically ignored in studies of allelopathy. We tested the hypothesis that CMNs facilitate transport of allelochemicals from supplier to target plants, thereby affecting allelopathic interactions. We analyzed accumulation of a model allelopathic substance, the herbicide imazamox, and two allelopathic thiophenes released from Tagetes tenuifolia roots, by diffusion through soil and CMNs. We also conducted bioassays to determine how the accumulated substances affected plant growth. All compounds accumulated to greater levels in target soils with CMNs as opposed to soils without CMNs. This increased accumulation was associated with reduced growth of target plants in soils with CMNs. Our results show that CMNs support transfer of allelochemicals from supplier to target plants and thus lead to allelochemical accumulation at levels that could not be reached by diffusion through soil alone. We conclude that CMNs expand the bioactive zones of allelochemicals in natural environments, with significant implications for interspecies chemical interactions in plant communities.

The difficulties of monitoring allelochemical concentrations in soil and their dynamics over time have been a major barrier to testing hypotheses of allelopathic effects. Here, we evaluate three diffusive sampling strategies that employ polydimethylsiloxane (PDMS) sorbents to map the spatial distribution and temporal dynamics of root-exuded thiophenes from the African marigold, Tagetes erecta. Solid phase root zone extraction (SPRE) probes constructed by inserting stainless steel wire into PDMS tubing were used to monitor thiophene concentrations at various depths beneath marigolds growing in PVC pipes. PDMS sheets were used to map the distribution of thiophenes beneath marigolds grown in thin glass boxes. Concentrations of the two major marigold thiophenes measured by these two methods were extremely variable in both space and time. Dissection and analysis of roots indicated that distribution of thiophenes in marigold roots also was quite variable. A third approach used 1 m lengths of PDMS microtubing placed in marigold soil for repeated sampling of soil without disturbance of the roots. The two ends of the tubing remained out of the soil so that solvent could be washed through the tubing to collect samples for HPLC analysis. Unlike the other two methods, initial experiments with this approach show more uniformity of response, and suggest that soil concentrations of marigold thiophenes are affected greatly even by minimal disturbance of the soil. Silicone tube microextraction gave a linear response for alpha-terthienyl when maintained in soils spiked with 0-10 ppm of this thiophene. This method, which is experimentally simple and uses inexpensive materials, should be broadly applicable to the measurement of non-polar root exudates, and thus provides a means to test hypotheses about the role of root exudates in plant-plant and other interactions.