DNA barcoding of arbuscular mycorrhizal fungi
b'Plant beneficial microorganisms, such as arbuscular mycorrhiza fungi (AMF), increasingly attract\\nscientific and agronomic attention due to their capacity to increase nutrient accessibility for plants\\nand to reduce inorganic fertilizer requirements. AMF are thought to form symbioses with most land\\nplants, obtaining carbon from the autotrophic host whilst enhancing uptake of poorly available\\nnutrients.\\nThe species of AMF are mainly identified by spore morphology, which is time consuming, requires\\nexpertise and is rarely applicable to AMF identification in roots. Molecular tools such as analysis of\\nstandardized DNA fragment sequences may allow the recognition of species through a \\u2018DNA\\nbarcode\\u2019, which may partly overcome this problem. The focus of this study was to evaluate\\ndifferent regions of widely used rDNA repeats for their use as DNA barcodes for AMF including the\\nsmall subunit rRNA gene (SSU), the internal transcribed spacer (ITS) and the large subunit rRNA\\ngene (LSU). Closely related species in the genus Ambispora, members of which have dimorphic\\nspores, could not be separated by analysis of the SSU region, but of the ITS region. Consequently,\\nthe SSU was not used for subsequent analysis, but a DNA fragment covering a small part of the\\nSSU, the entire ITS region and about 800 bp of the LSU (SSUmCf-LSUmBr fragment) was\\nanalysed, providing phylogenetic resolution to species. New AMF specific primers for these\\npotential barcoding regions were developed and can be applied, without amplification of non-target\\norganisms, for AMF species determination, including identification from field and root samples.\\nAnalyses based on the application of the SSUmCf-LSUmBr fragment showed that the widely used\\nAMF model organism Glomus sp. DAOM197198 (formerly called Glomus intraradices) is not\\nconspecific with Gl. intraradices. The SSUmCf-LSUmBr fragment clearly provides a much higher\\nspecies resolution capacity when compared with the formerly preferred ITS and LSU regions.\\nFurther study of several groups of AMF species using different regions of the SSUmCf-LSUmBr\\nfragment revealed that only the complete SSUmCf-LSUmBr fragment allowed separation of all\\nanalysed species. Based on these results, an extended DNA barcode covering the ITS region and\\nparts of the LSU region is suggested as a DNA barcode for AMF. The complete SSUmCf-LSUmBr\\nfragment sequences can serve as a database backbone for also using smaller rDNA fragments as\\nbarcodes. Although the smallest fragment (approximately 400 bp) analysed in this study was not\\nable to discriminate among AMF species completely, such short regions covering the ITS2 or LSU\\nD2 regions, respectively, would most likely be suitable for community analyses with 454 GS-FLX\\nTitanium sequencing, providing that the analyses is based on the longer DNA sequences.'