title: Genome wide sequence analysis grants unbiased definition of species boundaries in "Candidatus Phytoplasma".
abstract: The phytoplasmas are currently named using the Candidatus category, as the inability to grow them in vitro prevented (i) the performance of tests, such as DNA-DNA hybridization, that are regarded as necessary to establish species boundaries, and (ii) the deposition of type strains in culture collections. The recent accession to complete or nearly complete genome sequence information disclosed the opportunity to apply to the uncultivable phytoplasmas the same taxonomic approaches used for other bacteria. In this work, the genomes of 14 strains, belonging to the 16SrI, 16SrIII, 16SrV and 16SrX groups, including the species "Ca. P. asteris", "Ca. P. mali", "Ca. P. pyri", "Ca. P. pruni", and "Ca. P. australiense" were analyzed along with Acholeplasma laidlawi, to determine their taxonomic relatedness. Average nucleotide index (ANIm), tetranucleotide signature frequency correlation index (Tetra), and multilocus sequence analysis of 107 shared genes using both phylogenetic inference of concatenated (DNA and amino acid) sequences and consensus networks, were carried out. The results were in large agreement with the previously established 16S rDNA based classification schemes. Moreover, the taxonomic relationships within the 16SrI, 16SrIII and 16SrX groups, that represent clusters of strains whose relatedness could not be determined by 16SrDNA analysis, could be comparatively evaluated with non-subjective criteria. "Ca. P. mali" and "Ca. P. pyri" were found to meet the genome characteristics for the retention into two different, yet strictly related species; representatives of subgroups 16SrI-A and 16SrI-B were also found to meet the standards used in other bacteria to distinguish separate species; the genomes of the strains belonging to 16SrIII were found more closely related, suggesting that their subdivision into Candidatus species should be approached with caution.
authors.affiliation: Dipartimento di Scienze Agrarie ed Ambientali, Università di Udine, Udine, Italy; Istituto Nazionale di Biostrutture e Biosistemi, Interuniversity Consortium, Italy. Electronic address: firrao@uniud.it.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Firrao
authors.forename: Marta
authors.initials: M
authors.lastname: Martini
authors.forename: Paolo
authors.initials: P
authors.lastname: Ermacora
authors.forename: Nazia
authors.initials: N
authors.lastname: Loi
authors.forename: Emanuela
authors.initials: E
authors.lastname: Torelli
authors.forename: Xavier
authors.initials: X
authors.lastname: Foissac
authors.forename: Patricia
authors.initials: P
authors.lastname: Carle
authors.forename: Bruce C
authors.initials: BC
authors.lastname: Kirkpatrick
authors.forename: Lia
authors.initials: L
authors.lastname: Liefting
authors.forename: Bernd
authors.initials: B
authors.lastname: Schneider
authors.forename: Cristina
authors.initials: C
authors.lastname: Marzachì
authors.forename: Sabrina
authors.initials: S
authors.lastname: Palmano
mesh_terms: D016000:Cluster Analysis; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D016680:Genome, Bacterial; D058885:Multilocus Sequence Typing / Q000379:methods*; D010802:Phylogeny; D045582:Phytoplasma / Q000145:classification* / Q000235:genetics*
chemical_list: D004269:DNA, Bacterial
keywords: ANI; Candidatus Phytoplasma; Genomics; MLSA; Taxonomy