Weedy Hosts and Prevalence of Potential Leafhopper Vectors (Hemiptera: Cicadellidae) of a Phytoplasma (16SrIX group) Associated with Huanglongbing Symptoms in Citrus Groves Author(s): R. N. Marques, D. C. Teixeira, P. T. Yamamoto, and J.R.S. Lopes Source: Journal of Economic Entomology, 105(2):329-337. 2012. Published By: Entomological Society of America DOI: http://dx.doi.org/10.1603/EC11254 URL: http://www.bioone.org/doi/full/10.1603/EC11254 BioOne (www.bioone.org) is a nonprofit, online aggregation of core research in the biological, ecological, and environmental sciences. BioOne provides a sustainable online platform for over 170 journals and books published by nonprofit societies, associations, museums, institutions, and presses. Your use of this PDF, the BioOne Web site, and all posted and associated content indicates your acceptance of BioOne’s Terms of Use, available at www.bioone.org/page/terms_of_use. Usage of BioOne content is strictly limited to personal, educational, and non-commercial use. Commercial inquiries or rights and permissions requests should be directed to the individual publisher as copyright holder. BioOne sees sustainable scholarly publishing as an inherently collaborative enterprise connecting authors, nonprofit publishers, academic institutions, research libraries, and research funders in the common goal of maximizing access to critical research. ARTHROPODS IN RELATION TO PLANT DISEASE Weedy Hosts and Prevalence of Potential Leafhopper Vectors (Hemiptera: Cicadellidae) of a Phytoplasma (16SrIX group) Associated With Huanglongbing Symptoms in Citrus Groves 1,2 3 1 1 R. N. MARQUES, D. C. TEIXEIRA, P. T. YAMAMOTO, AND J.R.S. LOPES J. Econ. Entomol. 105(2): 329Ð337 (2012); DOI: http://dx.doi.org/10.1603/EC11254 ABSTRACT Huanglongbing (HLB) is a severe citrus (Citrus spp.) disease associated with the bacteria genus Candidatus Liberibacter, detected in Brazil in 2004. Another bacterium was found in association with HLB symptoms and characterized as a phytoplasma belonging to the 16SrIX group. The objectives of this study were to identify potential leafhopper vectors of the HLB-associated phytoplasma and their host plants. Leafhoppers were sampled every other week for 12 mo with sticky yellow cards placed at two heights (0.3 and 1.5 m) in the citrus tree canopy and by using a sweep net in the ground vegetation of two sweet orange, Citrus sinensis (L.) Osbeck, groves infected by the HLB-phytoplasma in Sa˜o Paulo state. Faunistic analyses indicated one Agalliinae (Agallia albidula Uhler) and three Deltocephalinae [Balclutha hebe (Kirkaldy), Planicephalus flavicosta (Stål), and Scaphytopius (Convelinus) marginelineatus (Stål)] species, as the most abundant and frequent leaf- hoppers (Hemiptera: Cicadellidae). Visual observations indicated an association of leafhopper species with some weeds and the inßuence of weed species composition on leafhopper abundance in low-lying vegetation. S. marginelineatus and P. flavicosta were more frequent on Sida rhombifolia L. and Althernantera tenella Colla, respectively, whereas A. albidula was observed more often on Conyza bonariensis (L.) Cronq. and B. hebe only occurred on grasses. DNA samples of Þeld-collected S. marginelineatus were positive by polymerase chain reaction and sequencing tests for the presence of the HLB-phytoplasma group, indicating it as a potential vector. The association of leafhoppers with their hosts may be used in deciding which management strategies to adopt against weeds and diseases in citrus orchards. KEY WORDS weed, plant disease, phloem-limited bacteria, sap-sucking insect Many pests and diseases can inßict signiÞcant damage genic agents transmitted by insects (mostly leafhop- to the Brazilian citriculture, including Huanglongbing pers but also planthoppers and psyllids), causing Ϸ700 (HLB), one of the most destructive diseases of citrus diseases in plants, many of which are lethal (Wein- (Citrus spp.). Until recently, this disease was associ- traub and Beanland 2006, Bertaccini 2007). ated only with the bacteria Candidatus Liberibacter Until now, Hemiptera is the only order identiÞed as asiaticus and Ca. L. americanus, both transmitted by having phytoplasma vectors and presenting several the Asian citrus psyllid, Diaphorina citri Kuwayama, characteristics that make its members efÞcient vectors and with Ca. L. africanus, transmitted by the psyllid of these plant pathogens. Adults and nymphs generally Trioza erytreae (Del Guercio) (Bove´ 2006, Yamamoto have similar feeding habits; consequently, both stages et al. 2006, Gottwald 2010). However, in 2008 a new can acquire and transmit phytoplasmas. Some agent was identiÞed in orange trees with HLB symp- hemipteran groups feed speciÞcally and selectively in toms that were negative for the presence of Ca. Li- the phloem of their host plants, allowing acquisition beribacter spp. It was a phytoplasma with a 99% sim- and inoculation of phytoplasmas that inhabit this plant ilarity with Pigeon pea witchesÕ broom phytoplasma of tissue. In addition, the phytoplasmas have a persistent the 16Sr IX group (Teixeira et al. 2008). The phyto- and propagative relationship with insects, similar to plasmas are prokaryotes with no cell wall and size other symbiotic prokaryotes found in hemipterans variation between 200 and 800 nm, important patho- (Weintraub and Beanland 2006, Weintraub 2007). There are more studies on the identiÞcation and 1 Escola Superior de Agricultura “Luiz de Queiroz”/Universidade molecular characterization of phytoplasmas than on de Sa˜o Paulo, Departamento de Entomologia e Acarologia, Av. Pa´dua the determination of vectors for this pathogen group Dias 11, Piracicaba, SP, Brazil 13418900. (Weintraub and Beanland 2006). Because many phy- 2 Corresponding author, e-mail: [email protected]. toplasma vectors are still unknown, as it is the case of 3 Fundo de Defesa da Citricultura, Fundecitrus, Departamento Cientõ´Þco, Av. Adhemar Pereira de Barros 201, Araraquara, SP, Brazil, the phytoplasma associated with HLB, the surveying 14807040. of leafhopper species present in commercial crops is 0022-0493/12/0329Ð0337$04.00/0 ᭧ 2012 Entomological Society of America 330 JOURNAL OF ECONOMIC ENTOMOLOGY Vol. 105, no. 2 fundamental for knowing the potential of some spe- Sticky yellow cards (both sides, measuring 8.5 by 11 cies to transmit these pathogens. Ongoing studies sug- cm) were used in the two study areas to evaluate gest that the diversity of leafhoppers in Brazil guar- leafhopper activity. These traps were set out at 20 antees many options of potential vector species points within each plot, with each point consisting of (Yamamoto and Gravena 2000, Lopes and Oliveira one citrus plant with two traps fastened to the tree 2004, Giustolin et al. 2009, Miranda et al. 2009b, side-branches at heights of 0.3 and 1.5 m. These two Ringenberg et al. 2010). heights were chosen to investigate leafhopper activity Other important information, such as alternative in the canopy of the orange trees or moving within the host plants for the leafhoppers, are necessary for the orchard (1.5 m in height) and also on the ground implementation of management strategies. However, vegetation (0.3 m). Captured leafhoppers were re- these same plants can be hosts to phytoplasmas be- moved from the sticky cards by using a solvent (ker- cause various taxonomic plant groups are infected by osene) to dissolve the glue and were mounted on these pathogens (Montano et al. 2007, Eckstein 2010). insect pins. In the search for information on potential vectors Insect identiÞcation was made under a stereoscopic of the HLB-associated phytoplasma (phytoplasma- microscope, comparing the morphological character- HLB) and their host plants, the aim of the current istics with those of leafhopper specimens already iden- study was to identify the leafhopper species present in tiÞed from a reference collection at the Entomology citrus groves affected by this pathogen; associate the Museum of the Department of Entomology and Ac- presence of weeds with the populations of these in- arology, ESALQ/University of Sa˜o Paulo (USP). Sev- sects, as well as the importance of these plants in the eral specimens of each morphospecies collected in the area; and to evaluate the occurrence of leafhoppers survey were sent to Dr. Keti Zanol (Department of species carrying the pathogen. Zoology, Federal University of Parana´, Parana´, Brazil), a specialist in Deltocephalinae, for taxonomic identi- Þcation. Voucher specimens of each species were de- posited at the Entomology Museum of ESALQ/USP. Materials and Methods Based on the number of leafhoppers collected from Survey of Leafhoppers. The survey of leafhoppers sticky traps in each plot for the May 2008ÐMay 2009 was made in the Nova Granada municipality, northern period, a faunistic analysis was made to determine Sa˜o Paulo state, Brazil. This area was selected by the those species that had the highest indices of con- presence of phytoplasma infected plants of the 16SrIX stancy, frequency, abundance, and dominance (Sil- group, associated with HLB symptoms and conÞrmed veira Neto et al. 1995). by polymerase chain reaction (PCR) and sequencing. Collection and Identification of Weeds in the Plots. Two areas of sweet oranges were sampled [Citrus A botanical survey of weeds was made in the study sinensis (L.) Osbeck], one area composed of ÔValeˆnciaÕ plots. A 1- by 1-m (1-m2) square was thrown Þve times plants grafted onto 14-yr-old rangpur lime root- randomly between the orange tree rows and Þve times stock