Introduce of Rhizotron for assessment of the effects of soil treatments on growth and distribution of plant roots

Introduction: Due to the difficulties in observing root growth in soil, our knowledge regarding soil-root system is limited. The roots are the hidden half of the plants but our knowledge of root’s growing is limited. Now, there are some methods and devices that have been used to analyze and monitoring roots architecture and growth and their relation with soil. The assessment of the root growth of plants is possible with some photographic techniques such as neutron radiography and tomography, as well X-Ray imaging, but the use of these methods for root studies is very costly especially in Iran. The use of the rhizotron can also be one of the most practical and cost effective methods. The rhizotron is a box with a transparent side and uses to study the roots growing by photography or drawing roots on transparent acetate sheets. Here we aimed to introduce the rhizotron as a technique for studying plant roots, and conducted this study to investigate the effects of heterogeneous petroleum pollution in soil and endophytic fungus on growth and distribution of maize root. Materials and Methods: In order to bring rhizotron forward as a method for in situ assessment of growth, establishment and distribution of plant roots a greenhouse experiment was performed. The effect of Piriformospora indica and soil petroleum polluted layers on the growth and distribution of maize (Zea mays L.) roots was studied. The rhizotrons had a wooden frame and back plate, and a removable front cover made of a 4 mm thick glass plate. The inner space was 30 cm high, 20.5 cm wide, and 1.5 cm thick (Figure 1a). The rhizotrons were placed on a rack with a 45˚ inclination to induce roots growing along the front glass to enable visual growth monitoring. The front glass plate was covered with an opaque black plastic to prevent light entering except for the times of observation. Two different patterns of soil-petroleum contamination layering were generated in the packings of the rhizotrons. 1) a shallow layer of 2.5 cm thick petroleum-contaminated soil, underlying of a 2.5 cm and above a 22.5 cm layer of uncontaminated soil (NSC), and 2) 27.5 cm of uncontaminated soil (‘control’). The packing procedure was layer by layer using uniform filling in all three cases. The contaminated soil layers were covered with a 2.5 cm layer of uncontaminated soil in order to facilitate plant establishment. The experiment with three replications was included as the packing methods each for growing maize plants inoculated with and without P. indica, and plant-free controls. The root development was recorded 12, 16, 22, 26, 33, and 45 days after transplanting by tracing all roots that were visible through the front glass on acetate transparencies, then were scanned at 300 dpi and analyzed for parameters such as root length, number of root tips and depth of rooting using the SmartRoot plugin of the software package ImageJ. After the last recording, the experiment was terminated. Roots and shoots were separated after harvesting, weighed and oven-dried. Rhizosphere soil samples were taken from the layer 2.5–5.0 cm below the soil surface for total petroleum hydrocarbons analysis. Results and Discussion: The results well showed that root length, root depth and number of root tips could be monitored by this method. The presence of petroleum pollution in the soil significantly decreased the growth and distribution of roots but inoculated plants had more root length and root depth than uninoculated plants. The number of root tips which representing lateral distribution of roots had similar trend with root the length and they were significantly increased in the inoculated plants as compared with uninoculated ones. Results showed that inoculation of maize by P. indica increases root biomass more than the aboveground biomass. Conclusion: Despite the limitation in the study of the effect of soil treatments on the growth and distribution of plant roots, the use of rhizotron could be a technique to solve this limitation. Our study proved that by using of rhizotron we can show the growth and 2-dimension distribution of plant roots, while the effects of treatment and root-soil interaction could also be assessing by this device.