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1 1 INTEGRATED CONTROL of BACTERIAL WILT of POTATO Ralstonia Solanacearum Is the Causal Agent of the Disease Known As Potato

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1 1 INTEGRATED CONTROL of BACTERIAL WILT of POTATO Ralstonia Solanacearum Is the Causal Agent of the Disease Known As Potato 1 INTEGRATED CONTROL OF BACTERIAL WILT OF POTATO INTEGRATED CONTROL OF Authors: S. Priou, P. Aley, E. Chujoy, BACTERIAL WILT OF POTATO B. Lemaga and E. R. French S. Priou, P. Aley, E. Chujoy, B. Lemaga and E. French Ralstonia solanacearum is the causal agent of the disease known as potato brown rot or bacterial wilt (BW). The bacterium affects more than 30 plant species, the most susceptible crops being potato, tomato, eggplant, pepper, banana and groundnut. Spread of the pathogen worldwide has been associated with its dissemination in latently infected planting mate- rial. Bacterial wilt is the second most important constraint to potato production in tropical and sub- tropical regions of the world. Quarantine measures necessary to avoid spread of the disease to BW-free areas often restrict the production of seed potatoes and limit the commercialization of ware potatoes between countries and between infected and non-infected regions within a country. Limited availability of high-quality seed, and lack of farmer knowledge on proper agronomic practices, threaten the sustainability of the potato crop in the developing world. BW incidence can be reduced only if various control components are combined. This involves mainly the planting of 1 healthy seed in clean soil, planting important in Indonesia, the Philip- tolerant varieties, rotation with pines, southern Vietnam, Laos, non-susceptible crops, and, the Japan and southern China. In the application of various sanitation early 1990s, BW became a seri- and cultivation practices. An ous threat to potato production in integrated disease management European countries including approach can lead to significant Belgium, England, France, The reduction, or even eradication of Netherlands, Spain, Italy and BW. Portugal. It has also been reported 2 in Russia. Earlier incidence in WORLDWIDE DISTRIBUTION OF Sweden was followed by eradica- WORLDWIDE DISTRIBUTION OF Ralstonia solanacearum ON POTATO R. solanacearum ON POTATO tion. Bacterial wilt has spread to most 3 potato producing countries. Its SURVEY AND DIAGNOSIS occurrence in Australia and in the Inspection of potato fields is of southeastern United States has prime importance in seed certifica- resulted in concentration of scien- tion schemes. This is the way to tific research on BW in these identify diseased areas where areas. In Latin America, the national crop protection services disease has been reported from all may apply quarantine and control potato producing countries except SURVEY AND DIAGNOSIS measures. Likewise, early disease Ecuador. BW occurs throughout diagnosis allows potato growers to central and southern Africa, and is define an appropriate manage- a serious production constraint in ment strategy. Field diagnosis of Uganda, Ethiopia, Kenya, Mada- bacterial wilt disease can be done gascar, Rwanda, Burundi, Nigeria by identifying the foliage and tuber and Cameroon. Although not a symptoms during the growing serious disease in Egypt, latent season or at harvest. infection of tubers has resulted in 4 a strong decline of potato exports Foliage symptoms to Europe. In South Asia, BW occurs in the mid-hills of the Himalayas in India, Pakistan, Nepal and Bhutan; also in the mid- Foliage symptoms hills and plains of India, where it is effectively controlled. In East and Southeast Asia, bacterial wilt is 2 3 5 Total and unilateral wilting of 8 Bacterial exudate from vascular potato ring Above ground symptoms of BW Tuber symptom is often described include wilting, stunting and yel- as brown rot. Cut tubers show Total wilt lowing of the foliage. The browning brownish discoloration of the of vascular bundles may be seen vascular ring, and slight squeezing Unilateral wilt externally, or when the cortex is forces a pus-like slime out of the peeled. Characteristic too, is the ring, or it may exude naturally. initial wilting of only part of the Bacterial exudate from vascular ring stems of a plant, or even one side of a leaf or stem. If disease devel- 9 Necrosis involving secondary opment is rapid, the entire plant infection wilts quickly, without yellowing. The vascular ring, or the whole Alternatively, the diseased stem tuber, may disintegrate completely can wilt completely and dry up, at more advanced stages of while the remainder of the plant necrosis development. This often appears healthy. involves secondary infection with 6 saprophytic bacteria (Erwinia spp., Tuber symptoms Necrosis involving secundary infection Clostridium sp.) or fungi (Fusarium sp., Pythium sp.). 10 Field diagnosis Tuber symptoms Field diagnosis 7 Bacterial exudate in tuber eyes External symptoms on the tuber are visible at harvest when infec- 11 tion is severe. Bacterial ooze The vascular flow test collects at tuber eyes or on the Vascular flow test A diagnostic test is necessary end of the stolon, causing soil to because plant wilting caused by adhere to the secretions. the bacterium Ralstonia Bacterial exudate in tuber eyes solanacearum can be confused with symptoms induced by other pathogens such as Fusarium 4 5 eumartii, Verticillium sp., Erwinia positive bacterium) the thread is chrysanthemi, or by insect or not produced. mechanical damage at the stem 13 base. Diagnosis in the field can be Laboratory identification easily accomplished by cutting a piece of stem 2-3 cm long from the base, and suspending it in clear water in a glass container. The cut Laboratory identification stem can be held with an opened paper clip to maintain a vertical position. Within a few minutes, the smoke-like milky threads stream downward from the cut stem. This milky slime exuding from the stem 14 Isolation of R. solanacearum on is characteristic of R. modified Kelman’s medium solanacearum present within the The BW agent can be isolated vascular system. from a diseased tuber or stem by 12 plating the bacterial tuber exudate The KOH test or two drops of the suspension The brown rot symptom on tubers obtained in the vascular flow test can be confused with ring rot on modified Kelman’s medium KOH test caused by Clavibacter Isolation of Ralstonia solanacearum (TZC containing only 2.5 g on modified Kelmans medium michiganensis subsp. sepedonicus dextrose). After 48 h incubation at (previously called Corynebacterium 30°C, the typically fluid, slightly sepedonicum). A quick differential red-tinted colonies of virulent R. test can be performed directly on solanacearum are easily distin- the bacterial exudate on the tuber guished from other saprophytic to differentiate between the two bacteria (round shaped, uniformly bacteria. Two drops of 3% dark red-colored colonies). potassium hydroxide (KOH) are 15 placed on the ooze and mixed with Pathogenicity test a laboratory loop or a wooden Potato The purified culture of R. Pathogenicity test toothpick for 10 seconds. The solanacearum on Kelman’s me- formation of a milky thread upon Tomato dium can be inoculated to tomato lifting the toothpick indicates the or potato seedlings to confirm its presence of R. solanacearum (a pathogenicity. Young tomato or Gram-negative bacterium), potato transplants (third to fifth whereas with C. michiganensis true leaf) are inoculated by injecting subsp. sepedonicus (a Gram- 6 7 the stems with 20 µl (e.g., a drop) negative, oxidation of alcohols of a bacterial suspension of R. negative; solanacearum at 108 cells/ml with Bv 2 = utilization of disaccharides a 1 ml-propylene syringe and a positive, oxidation of alcohols hypodermic needle, just above the negative; cotyledons. Another inoculation Bv 3 = utilization of disaccharides method is to insert (at the level of positive, oxidation of alcohols the third leaf axil) a toothpick positive; carrying the bacteria from a Bv 4 = utilization of disaccharides Kelman’s agar plate. Greenhouse negative, oxidation of alcohols conditions that favor disease positive; development are 28 ± 4ºC, 80- Bv 5 = utilization of disaccharides 90% R.H., and natural daylight. positive, oxidation of only mannitol. Plants are irrigated normally, except one day before inoculation. 17 If the isolate is a pathogenic strain Biovar determination of R. solanacearum, the wilting of These biochemical tests can be tomato or potato seedlings may performed together in microtitre begin in less than a week, but will plates. The utilization of the disac- certainly appear within 4 weeks. charide and oxidation of the After symptoms appear, the alcohol result in the acidification of vascular flow test can be per- the medium, expressed by the formed, and the pathogen can be Biovar determination change in the medium color from re-isolated from the vascular flow green (neutral pH) to yellow (acidic as described previously. pH). 16 Differentiation of 18 Differentiation of R. solanacearum R. solanacearum into biovars Equivalence between biovar and into biovars EquivalenceR. between solanacearum biovar race in R. solanacearum BIOCHEMICAL BIOVAR Two classification systems are and race in TESTS 1 2 3 4 5 used for R. solanacearum: the Race BiovarsHosts Location The race system is based on host : Solanaceous Utilization of Lowland tropics 1 1, 3, 4 + many m ore • lactose, maltose -++-+ range under field conditions. Four race and the biovar systems. The and cellobiose Musaceae American and 1, 3 2 = Moko disease Asian tropics Oxidation of : biovar system consists of bio- races can be distinguished: Potato 3 2A Cool climates • mannitol --+++ Tomato • Race 1 affects a wide range of • sorbitol, dulcitol chemical tests based on the ability --++- 4 5 Mulberry China Not Prim arily South solanaceous crops including of the bacterium to utilize three 2T Numerous known A merican low land s disaccharides and/or oxidize three potato and many weeds. hexose alcohols. Biovar classifica- Some strains can affect tion is based on the following: groundnut, ginger and diploid Bv 1 = utilization of disaccharides banana. It is prevalent at lower 8 9 elevations in the tropics and host crops, potato volunteers). R. subtropics. solanacearum is mainly spread by • Race 2 affects plants of the movement of infected seed tubers. musaceae family, such as Contaminated surface water used triploid banana, plantain and for irrigation and infested soil Heliconia spp.
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