Molecular characterization of Ralstonia solanacearum infecting Eucalyptus spp. in Brazil

Abstract

Among the bacterial pathogens of Eucalyptus in Brazil, Ralstonia solanacearum is considered one of the most important because of the characteristics of the pathogen, like the high diversity among the strains related to host range, high virulence, broad geographical distribution and its damage to the crop in recent years. Given its importance and the lack of research on this pathosystem, the present study aimed to perform a molecular characterization of different strains of infected Eucalyptus plants in Brazil. A total of 19 bacterial cultures isolated from Eucalyptus in different regions of Brazil were analysed. A 372-bp product generated by multiplex-PCR amplification using Nmult primers identified all the strains analysed as belonging to phylotype II. Eighteen strains were grouped into subclade IIA and one into subclade IIB. The phylogenetic tree generated from the gene sequences of endoglucanase (egl) confirmed the classification of the strains into phylotype II and separated the strains into sequevars. Strains AMC22, IBSBF2568 and IBSBF2576 were grouped into a single clade, as were strains UFV18 and UFV20, with 89% and 78% a posteriori probability, respectively, forming two new potential sequevars not yet defined. We also identified strains belonging to sequevars 41 (100% probability) and 37 (88% probability). However, most of the strains did not fit into any previously described sequevar and did not form distinct clades. The results of the analysis of fragments amplified using the ERIC-PCR technique indicated the existence of genetic diversity among the strains studied, with a generally high correlation between similarity and the geographical origin of the strains.

Two Ralstonia species associated with bacterial wilt of Eucalyptus

“…Differences were seen for each gene region when those sequences from Colombia were compared to the sequences from those isolates used by Fonseca et al . () but the differences were less than 5%.…”

“…Fonseca et al . () used phylotyping to classify R. solanacearum isolates that are associated with bacterial wilt of Eucalyptus in Brazil. Their results showed that the Brazilian isolates grouped into phylotype II.…”

Occurrence and diversity of Ralstonia solanacearum populations in Brazil

“…It is interesting to highlight that, in this review, some studies about the genetic variability of this bacterial population in Brazil report that the existence of diversity among strains oftentimes is correlated with its geographical origin (SILVEIRA et al, 2005;FONSECA et al, 2014), similarly to what was established for the classification into phylotypes. Therefore, the lower local bacterial variability allows disease control through the use of resistant cultivars recommended for each region of the country, although care should be taken against the dissemination of strains via propagation material.…”

“…Classification into phylotypes is done by PCR Multiplex with the Nmult series primers (based on ITS region) and the classification into sequevar by partially sequencing gene egl (encoding the enzyme endoglucanase). Presently, four phylotypes and 51 sequevares of R. solanacearum are described (XU et al, 2009;FONSECA et al, 2014). Analysis of genetic diversity can be conducted based on repetitive sequences (rep-PCR), comprised by the elements BOX, ERIC, and REP, by randomly amplified DNA (RAPD), by amplification of restriction fragments (AFLP), repeated simple sequences (SSR) and by polymorphisms based on restriction fragment size (RFLP) (JAUNET;WANG, 1999;VANDEWALLE;LUISETTI, 1999;COENYE;VANDAMME, 2003;YU et al, 2003;KUMAR;SARMA;ANANDARAJ, 2004;SILVEIRA et al, 2005;LOPES, 2007;IVEY et al, 2007).…”

“…This observation was also reported by Albuquerque et al (2014) that described a new sequevar associated to Moko, named IIA-53. Strains from eucalypt plants were not grouped into known sequevars neither (FONSECA et al, 2014).…”