Assessing Frankia populations in plants and soil using molecular methods

Abstract

In recent years, molecular approaches have increasingly supplemented nodulation‐dependent detection methods for studying Frankia populations in nature. The new methods are revealing much about the genetic diversity and distribution of Frankia, as well as refining and expanding knowledge about endophyte‐host specificities. PCR‐based approaches have been used to unravel the phylogenetic relationships of isolates, as well as of uncultured endophytes in root nodules of many actinorhizal plants from which no isolates have been obtained. A comparative sequence analysis of PCR‐amplified 16S ribosomal DNA led to the emendation of the family Frankiaceae to contain only the genus Frankia with four main subdivisions: (i) a large group mainly comprising Frankia alni and other typical nitrogen‐fixing strains belonging to the Alnus and the Casuarina host infection groups, respectively, (ii) uncultured endophytes of Dryas, Coriaria and Datisca species, (iii) strains of the Elaeagnus host infection group and (iv) atypical non‐nitrogen‐fixing strains. A considerable diversity among both cultured Frankia strains and uncultured endophytes in nodules was indicated using RFLP analyses of PCR‐amplified fragments of the 16S rRNA gene, the glutamine synthetase II (glnII) gene, the intergenic spacer of the 16S‐23S rRNA operon or the intergenic spacer between the nitrogenase nifH and nifD (nifH‐D) or the nifD and nifK (nifD‐K) genes. The growing database of discriminative target sequences for frankiae is increasingly exploited for studies on the distribution of specific Frankia populations in the environment using PCR or in situ hybridization. Until recently, most studies have focused on the analysis of Frankia populations in root nodules, the natural locale of enrichment for this organism. These populations, however, represent only the fraction of physiologically active, infecting frankiae in soils rather than the total Frankia population. Future approaches to studies of Frankia populations should therefore incorporate the many opportunities for more than just phylogenetic analyses, the description of diversity and studies of Frankia populations in nodules. The molecular approaches open the door to more sophisticated studies of environmental influences on the dynamics of indigenous or introduced Frankia populations in plants and soil. These studies may lead to advancements in the management of actinorhizal plants and Frankia, provided specific Frankia populations can be attributed with silviculturally beneficial features. Such features include persistence and the growth in soil, competition with less efficient Frankia populations for nodule formation, prompt and efficient nodule formation and an ultimately superior nitrogen‐fixing capacity.

Biosynthetic Potential of Phylogenetically Unique Endophytic Actinomycetes from Tropical Plants

“…Also, endophytic actinomycetes may easily gain access to roots through the damaged epidermal layer created when lateral roots grow out from the existing root structure. Such is the case for the most well-studied endophytic actinomycete, Frankia, which forms root nodules in nonleguminous woody plants but also exists in soil in substantial numbers (16). Once inside the roots, it is conceivable that endophytic actinomycetes other than Frankia could benefit plants by fixing nitrogen without forming nodules (37) and/or producing antibiotics or siderophores to protect against infection by soil-borne pathogens.…”

Bacterial diversity in hyperarid Atacama Desert soils

“…Bacteria in the genus Frankia are distinguished by their ability to form a symbiotic relationship with many plant species where they fix nitrogen for use by the plant within nodules that they induce to form on the roots [Benson and Silvester, 1993]. While Frankia are notoriously difficult to culture in the laboratory and are primarily isolated from within the roots nodules they form, Frankia have also been found to exist independently of host plants in soils via molecular methods and culture based techniques and are widespread in soils around the world [Baker and Okeefe, 1984;Hahn et al, 1999]. The clones in this study related to Frankia species had a sequence similarity to known Frankia species ranging from 91-95%.…”

Diversity of Plant Associated Actinobacteria

“…This technique was used to assess diversity within populations of Actinobacteria (Song et al 2004b;Wanner 2009;de Leon et al 2009;Dees et al 2012). Many researches focused on the IGS between functional genes in Actinobacteria such as nif gene (Jamann et al 1993;Navarro et al 1997;Hahn et al 1999;Dai et al 2004). Random amplified polymorphic DNA (RAPD) uses short primers, which anneal randomly at multiple sites on the genomic DNA under low annealing temperature and generates PCR amplicons of various lengths, was also used for some Actinobacteria (Pastrik and Rainey 1999;de Leon et al 2009).…”