Evaluation of amplified ribosomal DNA restriction analysis as a method for the identification of Botryosphaeria species

Abstract

The polymerase chain reaction was used to amplify a rDNA fragment containing the internal transcribed spacers (ITS1-5.8S-ITS2) and the D1/D2 variable domains of the 28S rDNA from 10 species of the genus Botryosphaeria (Fungi, Ascomycota). Restriction analysis of the amplicons with frequent-cutting endonucleases (amplified ribosomal DNA restriction analysis) allowed the definition of 12 rDNA haplotypes. Each of the rDNA haplotypes could be unambiguously assigned to a single Botryosphaeria species, thus allowing clear identification of all the species tested. Intraspecific polymorphism was very low and detected only in Botryosphaeria parva and Botryosphaeria dothidea. Cluster analysis of banding patterns of the isolates corresponded well with known species delineations. The method described in this paper provides a simple and rapid procedure for the differentiation and identification of Botryosphaeria isolates at the species level.

A diverse assemblage of Botryosphaeriaceae infectEucalyptusin native and non-native environments

“…Mohali et al (2006) used the same technique with CfoI on ITS rDNA amplicons, as well as an unknown locus, to distinguish six species of Botryosphaeriaceae from Eucalyptus in Venezuela. Alves et al (2005) were able to distinguish 10 Botryosphaeriaceae species using a larger PCR amplicon of the rDNA operon and treating this with six restriction enzymes.…”

Conidial production by <i>Botryosphaeriaceae</i> species from grapevine shoot lesions in Marlborough vineyards

“…The liquid was then spread on the surface of the agar with a sterile metal 'hockey stick' . One representative colony characteristic of a Botryosphaeriaceae species was sub-cultured from each isolation plate onto a fresh PDA plate and identified using amplified ribosomal DNA restriction analysis (ARDRA) as described by Alves et al (2005) and Baskarathevan et al (2012). To confirm the species determined by ARDRA, sequencing of the ribosomal internally transcribed spacer region (ITS) of randomly selected isolates ( 33%) was done at the Lincoln University Sequencing facility using an ABI PRISM® 310 Genetic Analyser (Applied Biosystems, Foster City, California).…”

“…They cause a range of symptoms, including leaf spots, fruit rots, dieback, perennial cankers, and eventual death of the plants, although symptoms vary according to the host species, cultivar and pathogen species (Alves et al 2005;Úrbez-Torres et al 2010). Yield losses caused in vineyards were estimated to be 4-20% in France and 25-30% in South Africa (Pearson & Goheen 1998;Larginon et al 2001).…”

Detection of Neofusicoccum nonquaesitum causing dieback and canker in highbush blueberry from Southern Chile

“…There are more than 18 anamorph genera associated to Botryosphaeria, most of which have been reduced to synonyms of Diplodia (pigmented and ovoid conidia, with thick walls) or Fusicoccum (hyaline and fusiform conidia, with thin walls) (Perez et al, 2008;Phillips, 2010). Nevertheless, conidia are also continuously detected with intermediate morphometric characteristics, which is why the genus Neofusicoccum was proposed, by DNA sequencing, distinct from Fusicoccum sensu stricto (F. aesculi) and which produces two different types of conidia; as a result, many species in Fusicoccum sensu lato were situated in the genus Neofusicoccum, based on morphometric characteristics but no by genetic analysis (Alves et al, 2005;Crous et al, 2006).…”