Diversity of Ralstonia solanacearum Infecting Eggplant in the Philippines.

ABSTRACT The diversity of Ralstonia solanacearum strains isolated from eggplant (Solanum melongena) grown in five provinces of the Philippine island group of Luzon was assessed using a recently described hierarchical system. All strains keyed to race 1, biovar 3 or 4. Phylotype-specific multiplex polymerase chain reaction (PCR) indicated that, like most other strains of Asian origin, all the strains in our Philippine collection belong to phylotype I. Taxometric and phylogenetic analyses of partial endoglucanase gene sequences of strains from this collection and those previously deposited into GenBank revealed at least four subgroups among the otherwise monophyletic phylotype I strains. Nucleotide polymorphisms within each subgroup were infrequent and, among the subgroups identified in this study, variation was always <1.3%, indicating that the large majority of strains could be assigned to a single sequevar. Genomic DNA fingerprinting using enterobacterial repetitive intergenic consensus (ERIC)-PCR revealed additional fine-scale genetic variation that was consistent with the endogluconase sequence data. Whole-pattern and band-based analyses of the genomic fingerprint data revealed four and eight distinct genotypes, respectively, within our collection. Eggplant from infested fields in different provinces tended to harbor mixed populations of ERIC genotypes, with the predominant genotype varying by location.

DNA Analysis of Ralstonia solanacearum and Related Bacteria Based on 282-bp PCR-Amplified Fragment.

Strains of Ralstonia solanacearum, Pseudomonas syzygii, and the blood disease bacterium (BDB) from different countries were tested for polymerase chain reaction amplification of the 282-bp fragment using the primer pair 759 and 760. These 282-bp fragments from 49 strains of R. solanacearum, six strains of P. syzygii, and two strains of BDB were sequenced. A phylogenetic tree was generated based on the sequence alignment. The R. solanacearum strains were divided into three groups. Group I was composed of strains belonging to biovars 3, 4, 5, and biovar N2 from Japan. Most of the strains from this group were of Asian origin except for two strains from Australia and Guyana (GMI 1000), the type strain. Group II was composed of strains belonging to biovars 1 and 2 and biovar N2 from Brazil. Group III was composed of strains belonging to biovar N2 from Japan and the Philippines. All strains of P. syzygii and BDB clustered in group III. Based on nucleotide differences of the 282-bp fragments, restriction enzyme NlaIII was capable of differentiating R. solanacearum strains into the three groups. Restriction analysis of 165 R. solanacearum isolates from the Philippines using NlaIII showed that all biovar 3 and 4 (group 1) strains had restriction fragments of 116 and 166 bp, strains belonging to biovars 1 and 2 (group 2) showed no restriction, and one strain belonging to biovar 2 (group 3) showed restriction fragments of 54 and 228 bp in size. Thus, NlaIII could be used for rapid differentiation of R. solanacearum strains. Additionally, other restriction enzymes, such as McrI, BsiEI, and MnlI could be used to differentiate R. solanacearum strains from P. syzygii strains.