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Detection of Ralstonia solanacearum from Asymptomatic Tomato Plants, Irrigation Water, and Soil Through Non-selective Enrichment Medium with hrp Gene-Based Bio-PCR
作者：Dinesh Singh ,Shweta Sinha ,D. K. Yadav;
公佈日期：105 年 12 月 30 日
研究報告
747
Bacterial wilt of tomato caused by Ralstonia
solanacearum (Smith) Yabuuchi et al. (Microbiol Immunol
39:897–904, 1995) is a serious disease, which causes losses
up to 60 % depending on environmental conditions, soil
property, and cultivars. In present investigation, nucleotide
sequences of virulence, hypersensitive response and pathogenicity
(hrp) gene were used to design a pair of primer
(Hrp_rs 2F: 50-AGAGGTCGACGCGATACAGT-30 and
Hrp_rs 2R: 50-CATGAGCAAGGACGAAGTCA-30) for
amplification of bacterial genome. The genomic DNA of
27 isolates of R. solanacearum race 1 biovar 3 & 4 was
amplified at 323 bp. The specificity of primer was tested on
13 strains of R. solanacearum with other group of bacteria
such as Xanthomonas oryzae pv. oryzae, X. campestris pv.
campestris, and X. citri subsp. citri. Primer amplified DNA
fragment of R. solanacearum at 323 bp. The sensitivity of
the primer was 200 cfu/ml and improved further detection
level by using non-specific enrichment medium casamino
acids-pepton-glucose broth followed by PCR (BIO-PCR).
Out of 130 samples of asymptomatic tomato plants, irrigation
water, and soil collected from bacterial wilt infested
field in different agro-climatic regions of India, R. solanacearum
was detected from 86.9, 88.5, and 90.9 per cents
samples using BIO-PCR, respectively. The primer was
found specific for detecting viable and virulent strains of R.
solanacearum and useful for the diagnosis of R. solanacearum
in tomato seedlings and monitoring of pathogen in
irrigation water and soil.
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