Comparison of four routinely used methods for assessing root colonization by arbuscular mycorrhizal fungi

Quantifying the proportion of roots colonized by arbuscular mycorrhizal fungi (AMF) is routine work for researchers conducting AMF studies. However, in practice, the methods are always misused, with their adaptability to different conditions neglected. In this study, four frequently used methods (root segment ±, root segment estimation, grid-line intersect, and magnified intersections) were evaluated and compared. Using the light microscopy based staining technique, we assessed AMF colonization of the roots of five plant species (Trifolium repens Linn., Zea mays Linn., Robinia pseudoacacia Linn., Populus simonii Carr., and Caragana korshinskii Kom.). The results revealed that a root length of at least 150 cm (rather than the usual 30 or 50 cm or 100 to 150 intersections generally used when following these four methods) should be examined to represent a single root sample whatever the method used. All four methods had good reproducibility, even though there was a high level of divergence among the results obtained using the different methods to assess the same root sample. We concluded that when assessing the AMF colonization of roots from the same species, all methods except the root segment ± method can be used; however, when assessing root samples from different species, the root segment estimation and magnified intersections methods give more reliable results. We suggest that the root segment ± method is an effective method for revealing the uniformity of AMF distributed in host roots of a certain length.