title: Sybr Green- and TaqMan-Based Quantitative PCR Approaches Allow Assessment of the Abundance and Relative Distribution of Frankia Clusters in Soils.
abstract: The nodule-forming actinobacterial genusFrankiacan generally be divided into 4 taxonomic clusters, with clusters 1, 2, and 3 representing nitrogen-fixing strains of different host infection groups and cluster 4 representing atypical, generally non-nitrogen-fixing strains. Recently, quantitative PCR (qPCR)-based quantification methods have been developed for frankiae of clusters 1 and 3; however, similar approaches for clusters 2 and 4 were missing. We amended a database of partial 23S rRNA gene sequences ofFrankiastrains belonging to clusters 1 and 3 with sequences of frankiae representing clusters 2 and 4. The alignment allowed us to design primers and probes for the specific detection and quantification of theseFrankiaclusters by either Sybr Green- or TaqMan-based qPCR. Analyses of frankiae in different soils, all obtained from the same region in Illinois, USA, provided similar results, independent of the qPCR method applied, with abundance estimates of 10 × 105to 15 × 105cells (g soil)-1depending on the soil. Diversity was higher in prairie soils (native, restored, and cultivated), with frankiae of all 4 clusters detected and those of cluster 4 dominating, while diversity in soils underAlnus glutinosa, a host plant for cluster 1 frankiae, orBetula nigra, a related nonhost plant, was restricted to cluster 1 and 3 frankiae and generally members of subgroup 1b were dominating. These results indicate that vegetation affects the basic composition of frankiae in soils, with higher diversity in prairie soils compared to much more restricted diversity under some host and nonhost trees.IMPORTANCERoot nodule formation by the actinobacteriumFrankiais host plant specific and largely, but not exclusively, correlates with assignments of strains to specific clusters within the genus. Due to the lack of adequate detection and quantification tools, studies onFrankiahave been limited to clusters 1 and 3 and generally excluded clusters 2 and 4. We have developed tools for the detection and quantification of clusters 2 and 4, which can now be used in combination with those developed for clusters 1 and 3 to retrieve information on the ecology of all clusters delineated within the genusFrankiaOur initial results indicate that vegetation affects the basic composition of frankiae in soils, with higher diversity in prairie soils compared to much more restricted diversity under some host and nonhost trees.
authors.affiliation: Texas State University, Department of Biology, San Marcos, Texas, USA.
authors.forename: Seifeddine
authors.initials: S
authors.lastname: Ben Tekaya
authors.affiliation: Texas State University, Department of Biology, San Marcos, Texas, USA.
authors.forename: Abirama Sundari
authors.initials: AS
authors.lastname: Ganesan
authors.affiliation: Texas State University, Department of Biology, San Marcos, Texas, USA.
authors.forename: Trina
authors.initials: T
authors.lastname: Guerra
authors.affiliation: University of Illinois at Urbana-Champaign, Department of Natural Resources and Environmental Sciences, Urbana, Illinois, USA.
authors.forename: Jeffrey O
authors.initials: JO
authors.lastname: Dawson
authors.affiliation: Texas State University, Department of Biology, San Marcos, Texas, USA.
authors.forename: Michael R J
authors.initials: MRJ
authors.lastname: Forstner
authors.affiliation: Texas State University, Department of Biology, San Marcos, Texas, USA dh49@txstate.edu.
authors.forename: Dittmar
authors.initials: D
authors.lastname: Hahn
mesh_terms: D029661:Alnus / Q000382:microbiology; D029662:Betula / Q000382:microbiology; D017931:DNA Primers; D015342:DNA Probes; D004269:DNA, Bacterial; D040161:Frankia / Q000235:genetics* / Q000254:growth & development / Q000302:isolation & purification*; D005798:Genes, Bacterial / Q000235:genetics; D014644:Genetic Variation; D005810:Multigene Family / Q000235:genetics*; D010802:Phylogeny; D018517:Plant Roots / Q000382:microbiology; D012338:RNA, Ribosomal, 23S / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D053204:Root Nodules, Plant / Q000254:growth & development / Q000382:microbiology; D016415:Sequence Alignment; D017421:Sequence Analysis; D012987:Soil; D012988:Soil Microbiology; D013559:Symbiosis; D014197:Trees / Q000382:microbiology
chemical_list: D017931:DNA Primers; D015342:DNA Probes; D004269:DNA, Bacterial; D012338:RNA, Ribosomal, 23S; D012987:Soil
keywords: alder; birch; qPCR; quantification; saprotrophic; soil