Preliminary Analysis of Citrus Greening (Huanglungbin) Epidemics

Gottwald, T. R., Aubert, B., and Xue-Yuan, Z. 1989. Preliminary analysis of citrus greening (Huanglungbin) epidemics in the People's Republic of China and French Reunion Island. Phytopathology 79:687-693. Epidemics of citrus greening disease in sweet orange groves were severity in the Reunion plot. Ordinary runs analysis indicated aggregation monitored for several years at two sites in the Guangxi Province of the of diseased trees in all plots during all years except in the case of one People's Republic of China and one site in French Reunion Island. Both plot in China when disease incidence exceeded 0.99. Aggregation along the exponential and the logistic models adequately described the increase rows of the Reunion plot was associated with prevailing winds. Isopaths of greening expressed as disease incidence over time in two plots in of greening disease incidence demonstrated the highest level of disease China and when the disease was expressed as both disease incidence and was present in the western half of the Reunion plot. This spatial pattern disease severity for the Reunion Island plot. The logistic apparent infection suggested the pathogen was introduced from the west against the prevailing rates, r, were 2.53, 1.87, and 0.99/yr for disease incidence at the two winds, probably by psyllid vectors. Chinese and one Reunion plots, respectively, and 1.04/yr for disease Additional keywords: likubin, citrus yellow shoot, citrus vein phloem degeneration, Diaphorina citri, Triosa erytreae, Lloyd's index of patchiness, spatial lag order autocorrelation. Citrus greening disease (greening) is devastating on most major and poorly colored, and seeds frequently abort. In Southeast Asia citrus cultivars in many parts of Southeast Asia, China, Taiwan, citrus tristeza virus (CTV) and citrus canker pathogen severely Indonesia, Philippines, Indian Ocean Islands, Africa, India, and affect limes, lemons, grapefruit, and oranges, thus restricting the the Arabian Peninsula (2,5,9,10,11,17,32,39). The disease is also cultivated citrus mostly to mandarins. Mandarins and oranges known as yellow shoot, huanglungbin, likubin, leaf mottling, are considered the most sensitive citrus trees to greening. citrus dieback, vein phloem degeneration, blotchy-mottle, and Citrus greening disease is caused by a fastidious, phloemyellow dragon. Citrus affected by the disease initially shows leaf limited, gram-negative bacterium (21-23,38). Although attempts mottling and chlorosis symptoms similar to zinc pattern to culture the bacterium are in progress, to date no confirmed deficiency, followed by retarded growth, often restricted to one isolate of the organism exists (19). Therefore, the disease organism branch or side of the tree (5,27,47). Eventually, twig dieback and is poorly characterized, although electron micrographs and a general decline ensue due to its severe effect on the phloem sensitivity to penicillin indicate it has a cell wall (4,20,37). Attempts of the host. Fruit on diseased trees are small, often asymmetrical, to produce monoclonal antibodies to the causal organism are in progress (24). __________________________________________ The greening pathogen is transmissible by grafting and This article is in the public domain and not copyrightable. It may be freely propagating with infected plant material (2,35,43). Thus, in areas reprinted with customary crediting of the source. The American in which greening is endemic, new groves are often established Phytopathological Society, 1989. with infected trees. The greening pathogen is vectored by two Vol. 79, No. 6, 1989 687 psyllid species, Diaphorina citri Kuwayama and Triosa erytreae established in the spring of 1969 and consisted of 126, 153, and Del Guercio (1,2,9,15,16,26,33,34). D. citri is widespread through195 trees of Xinhuicheng and Anliucheng sweet orange, Tankan out southern Asia and India, the Arabian peninsula, Reunion, mandarin (C. tankan Hayata), and Ponkan tangerine (C. reticulata and Mauritius, whereas T. erytreae is found primarily in Africa, Blanco), respectively, on various rootstocks. All trees were planted the southeastern part of the Arabian peninsula, Madagascar, on a 3.3X 4-m spacing. The two Liuzhou, China, plots were Reunion, and Mauritius (2,9,14,15). D. citri is more tolerant of probably established from greening-infected planting material, warm temperatures and less tolerant of cooler temperatures than although no greening symptoms were noted at planting. D. citri T. erytreae but is hindered by prolonged high rainfall and high was the only vector present in all three plots. Prevailing wind humidity (8,25). Both psyllid vectors are limited to hosts in the direction in the two China plots was variable; however, prevailing Rutaceae. In addition to citrus species, Murraya paniculata (L.) winds in the Reunion Island plot were east to west. Jack, a common ornamental shrub in Southeast Asia, is a Disease assessment. Disease severity and disease incidence were preferred host for both vectors but has not been shown to harbor visually assessed for each tree in RI during 1975, 1977, and 1979. the pathogen. Transmission appears to be related to high vector To estimate disease severity, each tree was divided into an upper populations and an extensive inoculum reservoir (2). Psyllid and lower hemisphere by an imaginary horizontal plane at about migrations appear to be highest when host plants are flushing midcanopy height. Each hemisphere was subdivided into four and psyllid populations sedentary and feeding when foliage is equal quadrants by two imaginary perpendicular planes passing mature (2). Thus, natural spread is probably greatest in late spring through the axis of the tree trunk. The resulting eight sections when new flush is available and psyllid populations are highest were scored individually on a 0-5 scale that was indicative of (2). Both psyllid species are parasitized by hymenopterous the proportion of limbs expressing disease symptoms within each ectoparasites Tetrastichus dryi Waterston and T. radiatus section (0 = no limbs, 5 = all limbs). The summation of the Waterston, a fact that has been used to accomplish biological eight scores for each tree resulted in a severity rating of 0-40 control of vector populations in Reunion Island and attempted for each tree on each survey date (3). This type of canopy rating elsewhere (2,6,12,18,43). was originally designed to check the recovery of trees injected Two forms of greening have been identified. The African form with antibiotics in separate experiments. Disease incidence was expresses symptoms best at temperatures between 20-24 C, calculated as the number of trees expressing disease symptoms whereas the Asian form expresses symptoms at temperatures up divided by the total number of trees in the plot. to 32 C (9,13). Therefore, although both psyllid species can Disease confirmation. Because citrus greening symptoms can transmit either form (25,26,33), African greening is most often be confused with those caused by other phloem pathogens such associated with T. erytreae and Asian greening with D. citri (2). as tristeza virus, samples taken from the plots were indexed for For instance, in Reunion, D. citri is established preferentially greening several times during the duration of the experiments on the dry leeward side of the island, prone to the Asian form (3). Indexing was accomplished by side grafting suspect shoots of greening. from the field onto seedling mandarin indicator trees in the Several analytical models have been proposed to quantify plant greenhouse and incubating these grafted trees for several months. disease epidemics by expressing the disease as a function of time Seedling mandarin trees are tolerant of tristeza virus but display (7,28,29,40,44,46). Epidemics of greening often require several greening symptoms within 2 mo of inoculation. Plants expressing years to closely approach an asymptotic level. The temporal such symptoms were taken as positive determination of the dynamics of greening in mature groves has not previously been presence of greening in trees from which the original samples investigated. Such information is essential to the understanding were collected. In the Reunion plots, additional samples were of the epidemic potential of greening in citrus groves. These studies confirmed as positive for greening by examining prepared foliar presented an opportunity to investigate the disease in citrus groves specimens via transmission electron microscopy (3). Bacterialike in two situations, where inoculum was introduced by infected organisms in the phloem sieve tubes provided further evidence planting stock, and where clean stock was used and the pathogen of the presence of the greening pathogen. was introduced by the ingress of contaminated psyllid vectors. Analysis of disease progression. The goodness-of-fit of the linear This study was conducted to establish preliminary rates of disease forms of the monomolecular, logistic, exponential, and Gompertz increase of citrus greening under endemic conditions in the models (7,28,29,46) to disease progress data of each plot was presence of vector populations and to develop preliminary examined by least squares regression analysis. Standard residual temporal models to estimate the expected longevity of sweet plots were examined to test the appropriateness of each model orange groves in China. (28). Predicted values were detransformed and correlated with observed, nontransformed values to test the efficiency of each MATERIALS AND METHODS model for describing disease progress within each plot. Models with larger correlation coefficients were considered superior (29). Field plots. Citrus greening disease was monitored by visual Analysis of spatial data. Ordinary runs analysis was used to assessment in three citrus plots over a period of several years determine if diseased trees were aggregated or occurred at random in two locations. The Reunion Island plot (RI) was located at within each plot (30). Analysis was performed by examining the Le Gol Etang Sale in the coastal leeward side of the island where data along rows and across rows. A nonrandom aggregation of D. citri was endemic. The plot was established in 1970 by IRFAdiseased or healthy trees was assumed if the expected number Reunion (Institut de Recherches Fruitibres Outre Mer), from of runs differed from the observed values at P-= 0.05 (30). disease-free planting material originating from Station de Lloyd's index of patchiness was calculated from quadratized Recherches Agronomiques SRA, Corsica. It consisted of 220 data for all three plots as a further indicator of aggregation (45). Valencia sweet orange (Citrus sinensis (L.) Osbeck) trees planted Indices of patchiness < 1 indicate a regular distribution, = 1 in 11 rows of 20 plants per row on a 6X 6-in pattern. The indicate a random distribution, and > 1 indicate aggregation of Liuzhou Citrus Farm Plot (LCF) was located 20 km from Liuzhou diseased individuals. City, Guangxi Province, People's Republic of China. This plot Spatial lag correlation analysis was performed on each plot was established in October 1953 and consisted of 405 trees, of for each year to assess the relationship among individual 3 >K 3 which 162 were Xinhuicheng and 243 Anliucheng sweet orange, tree quadrats for LCF and LARI and among individual greening two very similar cultivars, on rangpur lime (C. limonia Osbeck) diseased trees for RI (36). Correlation matrices were generated rootstock planted in 15 rows of 27 trees on a 4X 5-in spacing. in which the (x,y) element represented the correlation coefficient The Liuzhou Agricultural Research Institute plot (LARI) was between all tree pairs or quadrats that were x units apart in the located 22 km from Liuzhou City. The western part of the plot, east-west direction and y units apart in the north-south direction. which was originally designed as a rootstock trial, was established Significant correlations indicated the diseased trees or quadrats in spring 1968 and consisted of 108 trees of various citrus scion! were spatially autocorrelated (31 ,42). Cluster size of diseased trees rootstock combinations. The eastern part of the plot was or quadrats was estimated by interpreting the correlation matrices.