Rice, the second major cereal crop of Pakistan after wheat is suffering from destructive Bacterial Leaf Blight Disease (BLB) caused by Xanthomonas oryzae Pv. oryzae. Approximately 50 % incidence of BLB disease has been reported in Kaller, a high quality rice producing belt of Pakistan.
The present study was conducted to investigate the antimicrobial activity of different plant species extracts through in-vivo and in-vitro assays, and to isolate, purify and characterize the bioactive compounds exhibiting maximum anti-bacterial activity against Xanthomonas oryzae Pv. oryzae. The isolates of Xanthomonas oryzae Pv. oryzae were obtained from infected rice foliar samples collected from all agro ecological zones of Pakistan. Out of 123 isolates, 105 were found hypersensitive. The virulence and aggressiveness of seven hypersensitive isolates, (Xoo 20, Xoo 36, Xoo 51, Xoo 65, Xoo 74, Xoo 99 and Xoo 105) tested on seven rice varieties (Basmati 385, IRRI 26, Basmati 386, Dilroosh 97, JP 5, Super Basmati, Basmati 2000 and Ks 282), Xoo 99 showed most aggressive reaction on detached leaves and Xoo 105 exhibited more aggressive reaction on potted plant leaves as compared to other isolates. Moreover, Basmati 2000, Basmati 385, Super Basmati and IRRI 6 were more susceptible to BLB disease. On the basis of virulent and a-virulent reaction, pathogenicity test, colony morphology and bio-chemical tests, four isolates Xoo 51, Xoo 75, Xoo 99 and Xoo 105 were selected for antibacterial susceptibility test.
A total of 387 aqueous and non-aqueous plant extracts were screened for antibacterial susceptibility test through hole plate diffusion method. Out of 150 cold water diffusates only ten (Thuja orientalis, Prunus domestica, Prunus armanica, Citrus limon, Citrus aurantium, Allium sativum, Vitis vinefera, Phyllantus emblica, Terminali arjuna, Mangifera indica) exhibited maximum activity against Xanthomonas oryzae. Out of 140 hot water diffusates tested only eight (Linum usitatissimum, Phyllanthus emblica, Prunus domestica, Terminalia chebule, Citrus limon, Mangifera indica, Terminalia arjuna, Tamarindus indica) showed maximum inhibition against test bacterium. The decoction of 84 plants was tested and only 8 (Vernonia anthelmentica, Azadirachata indica, Terminalia chebula, Terminalia belerica, Antheum sp, Dodonea sp, Amomum subulatum, Piper nigrum, Curcuma Longa) gave maximum efficacy. Oil extracts of Eruca sativa or organic solvent extracts of Phyllanthus emblica showed maximum efficacy.
In comparison with the standard streptomycin drug, aqueous extracts of Terminalia chebula, Allium sativum and Tamarindus indica and non-aqueous extracts of Phyllanthus emblica, and Eruca sativa showed maximum activity index. The most potential aqueous extracts of Allium sativum, Terminalia chebula and Tamarindus indica showed maximum efficacy against BLB disease of rice when evaluated through detached leaf, glass house and field assays. The non-aqueous plant extracts of Linum usitatissum in detached leaf assay and Eruca sativa in glass house assay gave maximum efficacy in terms of controlling BLB disease lesions.
The plant extracts showing high antibacterial activity were further screened for bioactive compounds through chromatographic techniques. Through phytochemical studies, four bioactive compounds viz; 8-amino (8-butoxyoct)-2-enyldisulfanyl) (-3-(2 hydroxy-4-methoxy phenyl) prop-l-enyl) benzene-I, 3-diol, 1- Tigloyl-3-acetyl-l, 1-hydroxymeliacarpin (Azadirachtin D) Chebulinic acid and curcumin were isolated from Allium sativum, Azadirachata indica, Terminalia chebule and Curcuma longa respectively. These compounds were found effective against the test bacterium through autographic method. The present findings suggest that botanicals can effectively control BLB disease both in-vitro and in- vivo however, further studies or scaling up is needed.