Isolation and Characterization of Xanthomonas spp. from Brassicaceae

Abstract

The present investigation was designed to isolate, characterization of pathogenic bacteria from black rot disease of mustard and cabbage and evaluation of its biological control management. Xanthomonas campestris is a Gram-negative bacterium that causes black rot, the most important disease of mustard and cabbage of brassica crops in India as well as in the world. Black rot is a systemic vascular disease. Typical disease symptoms include V-shaped yellow lesions starting from the leaf margins and blackening of the veins. Black rot is a seedborne disease. The disease is favored by warm, humid conditions and can spread rapidly from rain dispersal and irrigation water. The disease affects primarily above ground parts of the plant at any stage of growth and causes high losses, in yield and quality especially in tropical and subtropical regions. The control of black rot is difficult and relies on the use of pathogen-free planting material and the elimination of other potential inoculum sources. An inventory of candidate effector leaves samples with disease symptoms were collected for isolation, identification and biochemical characterization of causing bacterial disease in the juridication of Sardar Vallabhbhai Patel University of Agriculture & Technology, Meerut (U.P.). Present study was undertaken during Rabi 2018-19 to establish an understanding of how biochemical characterization could be practiced to control black rot of brassica family. Bacteria were isolated on nutrient agar by spread plate method by incubating for 24-48 hrs at 28 ºC yellow color, smooth and shiny bacterial colonies were isolated. Moreover, suspected yellow color colonies of Xanthomonas were streaked on phyto buffered starch agar i.e., selective media for xcc, giving characteristic bluish green/bluish white color for xcc after 24 hrs at 28 ºC. finally the Xanthomonas was isolated and purified on YDC agar. After 24-48 hrs at 28 C֯, yellow colored, mucoid, slimy, glistening, convex, colourless and round in shape colonies were isolated. The isolated bacterium was gram negative rod shaped and yellow in color. Several biochemical tests including string test, gram staining, SIM test, catalase test, starch hydrolysis test, gelatin hydrolysis test oxidase test, 97 nitrate reduction test, TSI test and fluorescent test were done for characterization. Results obtained that Xanthomonas is positive for Ryu’s test, starch hydrolysis, gelatin test, hydrogen sulphide production, motility test, citrate test, catalase test. Based on the physiological, biochemical and morphological characteristics, out of 29 isolates 10 were the identified as Xanthomonas campestris. The antibiotic and antibacterial sensitivity was determined by disc diffusion method. Among all the antibiotics, particularly ciprofloxacin produced highest inbition zone of 32 in diameter followed by gentamicin & oflaxacin and least effective for co-trimoxazole and nitrofurantoin. All the isolates were susceptible particularly to gentamicin, levofloxacin, netillin, ofloxacin, amikacin, ciprofloxacin, streptomycin and kanamycin. Antibiotics nitrofurantoin, co-trimoxazole and tetracycline were intermediately effective against Xanthomonas. All of the isolates were resistant, particularly to antibiotics viz., ceftriaxone(Ci), amoxyclav(Ac) and carbenicillin(Cb), co-trimazine(Cm).