Abstract
Tomato and brinjal are prone to attack by number of bacterial pathogens
including Ralstonia solanacearum which is destructive and harmful resulting in
serious loss of the crops. Altogether 29 seed samples of tomato and 24 seed
samples of brinjal of different varieties were collected from different sources
viz. government, private and farmer's level of Bangladesh for evaluating seed
quality, seed borne Ralstonia solanacearum with its genetic diversity. Purity
analysis of tomato and brinjal seeds resulted 98.00 to 100.00% pure seeds.
Germination of tomato seeds varied from 73.40 to 97.00% and brinjal from 31.00
to 98.00%. Seed borne bacteria varied from 10.00 to 72.00% in tomato and 11.00
to 89.00% in brinjal on Nutrient Agar. Total 428 bacterial isolates were collected
from different seed samples of tomato and brinjal. The pathogenic isolates of all
groups of bacteria produced pink or light red color colonies or colonies with
characteristics red center and whitish margin on Triphenyl Tetrazolium
Chloride (TTC) medium indicated bacterial isolates of R. solanacearum were
virulent and confirmed by biochemical tests. Post-emergence death, seedling
mortality and chlorosis/necrosis were observed by pathogenicity and
hypersensitivity tests and the isolates were identified as R. solanacearum.
Species-specific confirmatory test of R. solanacearum was also performed by Y2/ OLI-1 primer and the tested isolates produced 288 bp fragment length. The
genetic variation of 20 isolates of tomato seed and 20 isolates of brinjal seed R.
solanacearum was analyzed using the primers (rep-PCR) by Polymerase Chain
Reaction (PCR). The bacterial isolates were then identified by sequencing of
these PCR products using primers specific to 16SrDNA genes of bacteria. The
nucleotide sequences of 16SrDNA of the bacterial isolates were compared with
16SrDNA sequences of other bacterial spp. available in the NCBI database using
Basic Local Alignment Search Tool (BLAST) algorithm. The results revealed that
Ralstonia solanacearum (BDISO02) identified in the present study was closely
related with Pseudomonas sp. and Pseudomonas stutzeri strains reported from
China (KF056821.1) and (MH283838.1) which was supported by bootstrap value
94%. However, it was distantly related with Pseudomonas sp. strains reported
from India (MF574413.1). Phylogenetic analyses of Ralstonia solanacearum
isolates obtained from brinjal by rep-PCR [REP, ERIC and BOX-IR] revealed
two distinct phylogenetic groups. Among the 3 primers REP showed higher
polymorphism than ERIC and BOXIR primers in the isolates of Ralstonia
solanacearum of tomato samples indicating REP is better than ERIC and BOXIR
for genetic diversity analysis. The BOXIR primer showed higher polymorphism
than ERIC and REP primers in seed borne Ralstonia solanacearum isolates
obtained from brinjal samples indicating BOXIR is better than ERIC and REP
for genetic diversity analysis.