Meng-Ling Wu

Identification of the Chinese Box Orange (Severinia Buxifolia) as an Alternative Host of the Bacterium Causing Citrus Huanglongbing

The Chinese box orange (Severinia buxifolia) was shown by graft-inoculation and psyllid-transmission tests to be an alternative host of the bacterium causing citrus Huanglongbing (HLB). A PCR-based assay for detection of the HLB bacterium (HLBB) was used to monitor HLBB. In graft-inoculation tests, the Chinese box orange (CBO) grafted with HLBB-infected scions of Luchen sweet orange (LSO) were positive for HLBB, 2–3 months after grafting. The back-grafting test demonstrated that HLBB-infected CBO scions could transmit HLBB back to LSO hosts via grafting. In psyllid-transmission tests, psyllids (insect vectors) transmitted HLBB to CBO plants, in which HLBB could be detected 3–4 months after inoculation. Acquisition-access tests of psyllids revealed that HLBB-free psyllids can acquire HLBB from diseased CBO hosts and can transmit HLBB back to the LSO plants. A field survey verified the presence of HLBB-infected CBO plants in the vicinity of citrus orchards. In this paper, CBO is shown to be a susceptible host plant in which HLBB can exist and replicate. It is also a donor plant from which HLBB can be transmitted to citrus hosts by grafting or by psyllid vectors.

Multiplex detection, distribution, and genetic diversity of Hop stunt viroid and Citrus exocortis viroid infecting citrus in Taiwan

BackgroundTwo citrus viroids, Citrus exocortis viroid (CEVd) and Hop stunt viroid (HSVd), have been reported and become potential threats to the citrus industry in Taiwan. The distributions and infection rates of two viroids have not been investigated since the two diseases were presented decades ago. The genetic diversities and evolutionary relationships of two viroids also remain unclear in the mix citrus planted region.MethodsMultiplex RT-PCR was used to detect the two viroids for the first time in seven main cultivars of citrus. Multiplex real-time RT-PCR quantified the distributions of two viroids in four citrus tissues. Sequence alignment and phylogenetic analysis were performed using the ClustalW and MEGA6 (neighbor-joining with p-distance model), respectively.ResultsHSVd was found more prevalent than CEVd (32.2% vs. 30.4%). Both CEVd and HSVd were commonly found simultaneously in the different citrus cultivars (up to 55%). Results of the multiplex quantitative analysis suggested that uneven distributions of both viroids with twig bark as the most appropriate material for studies involving viroid sampling such as quarantine inspection.Sequence alignment against Taiwanese isolates, along with analysis of secondary structure, revealed the existence of 10 and 5 major mutation sites in CEVd and HSVd, respectively. The mutation sites in CEVd were located at both ends of terminal and variability domains, whereas those in HSVd were situated in left terminal and pathogenicity domains. A phylogenetic analysis incorporating worldwide viroid isolates indicated three and two clusters for the Taiwanese isolates of CEVd and HSVd, respectively.ConclusionsModerately high infection and co-infection rates of two viroids in certain citrus cultivars suggest that different citrus cultivars may play important roles in viroid infection and evolution. These data also demonstrate that two multiplex molecular detection methods developed in the present study provide powerful tools to understand the genetic diversities among viroid isolates and quantify viroids in citrus host. Our field survey can help clarify citrus-viroid relationships as well as develop proper prevention strategies.

First Report of Banana Streak Virus Infecting Banana Cultivars (Musa spp.) in Taiwan

Banana (Musa sapientam L.) is an economically important crop for both export and local consumption in Taiwan. Recently, leaf symptoms characteristic of banana streak disease (1) were found on banana cv. Mysore (AAB group) introduced from Australia in the germ plasm collection of the Taiwan Banana Research Institute. The citrus mealybug (Planococus citri) has been shown to transmit banana streak virus (BSV) but not banana bunchy top virus or cucumber mosaic virus (CMV) (2). When mealybugs were fed on leaves of diseased Mysore banana and transferred to healthy banana cv. Cavendish seedlings in a growth chamber, the latter developed fine chlorotic streaks characteristic of symptoms caused by BSV within 1 to 3 months. Some chlorotic streaks became necrotic. BSV was detected in diseased but not healthy leaves of Mysore and Cavendish bananas by polymerase chain reaction (PCR) with primer pairs of BSV provided by J. E. Thomas of Queensland Department of Primary Industries. Subsequently, fine chlorotic streaks were observed in leaves of Cavendish banana in several fields in southern Taiwan. Some of these diseased plants developed severe leaf necrosis, causing heart rot of spindle leaves characteristic of symptoms caused by CMV. Presence of BSV in these plants was verified by PCR assay. However, CMV was also detected by double antibody sandwich-enzyme-linked immunosorbent assay with a monoclonal antibody to CMV, indicating that these plants were simultaneously infected by both viruses. This is the first report of BSV infecting Musa spp. in Taiwan. References: (1) B. E. L. Lockhart. Phytopathology 76:995, 1986. (2) B. E. L. Lockhart. 1995 Food & Fertilizer Technol. Center (ASPAC) Tech. Bull. 143. 11 pp.