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Title: First report of BLTVA phytoplasma in Capsicum annuum and Circulifer tenellus in Mexico

item Swisher Grimm, Kylie
item Munyaneza, Joseph - Joe
item VELASQUEZ-VALLE, R - Instituto Nacional De Investigaciones Forestales Y Agropecuarias (INIFAP)
item MENA-COVARRUBIAS, J - Instituto Nacional De Investigaciones Forestales Y Agropecuarias (INIFAP)

Submitted to: Plant Disease
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/25/2017
Publication Date: 6/1/2017
Citation: Swisher, K.D., Munyaneza, J.E., Velasquez-Valle, R., Mena-Covarrubias, J. 2017. First report of BLTVA phytoplasma in Capsicum annuum and Circulifer tenellus in Mexico. Plant Disease. 101(6):1032. doi:10.1094/PDIS-12-16-1723-PDN.

Interpretive Summary: Plant pathogens of pepper and other solanaceous crops like potato and tomato are a serious threat to production in the United States and around the world. Researchers at USDA-ARS Wapato in Washington, in collaboration with scientists at INIFAP in Mexico, assessed whether pathogens were associated with diseased pepper plants found in commercial fields in the states of Durango and Zacatecas, Mexico. A specific strain of group 16SrVI-A phytoplasma, known as BLTVA, was identified in the symptomatic pepper plants. BLTVA is transmitted to pepper by the beet leafhopper insect, and samples of this insect collected in and around pepper fields in Zacatecas, Mexico, were also positive for BLTVA. This pathogen is one of economic importance in solanaceous crops in North America, and its presence in peppers and beet leafhoppers suggests that growers will need to effectively monitor and control the insect vector to decrease overall disease pressure and the economic losses that

Technical Abstract: Pepper (Capsicum annuum) plants in Durango and Zacatecas, Mexico, in September and October, 2014, had small, chlorotic, curled leaves, plant stunting, and/or big bud symptoms characteristic of phytoplasma infection (Lee et al. 2004). Samples from symptomatic pepper fields included 33 collected near Poanas, Durango, 7 near Sombrerete, Zacatecas, 35 near Villa de Cos, Zacatecas, 40 near Laguna Seca, Zacatecas, and 41 near Calera, Zacatecas. Total DNA was extracted from foliar tissue of all samples with the CTAB extraction method (Munyaneza et al. 2010), and tested using universal phytoplasma nested PCR with primers P1/P7 followed by FU5/RU3 (Crosslin et al. 2006). Of 156 samples, 104 (67%) were positive for phytoplasma, including 3 from Poanas, 7 from Sombrerete, 27 from Villa de Cos, 31 from Laguna Seca, and 36 from Calera. All positive samples were tested for Beet Leafhopper Transmitted Virescence Agent (BLTVA) phytoplasma of the Clover proliferation group 16SrVI, subgroup A, using nested PCR with primers P1/P7 and BLTVA-specific FU5/BLTVA-int (Crosslin et al. 2006); all were positive for BLTVA. DNA was extracted from samples of the BLTVA beet leafhopper vector, Circulifer tenellus (Baker), collected from pepper fields and weeds adjacent to pepper fields in Zacatecas, Mexico, between September and October, 2014, using the CTAB method (Crosslin et al. 2006) and tested for BLTVA using nested PCR with primers P1/P7 and FU5/BLTVA-int. Of 52 samples, 23 (44%) were positive for BLTVA. Sequence analysis of the FU5-BLTVA-int amplicon from 6 peppers (2 each) from Poanas, Durango, Sombrerete, Zacatecas, and Laguna Seca, Zacatecas, and 3 C. tenellus samples from Zacatecas, produced two identical consensus sequences (basepairs 224-1415, GenBank accession KY047614 and KY047615, respectively). BLAST analysis of this region of 16S rRNA gene was 100% identical to Columbia Basin purple top phytoplasma (also known as BLTVA) (Crosslin et al. 2006), from the 16SrVI Clover proliferation group (GenBank accession KR072666.1). To verify phytoplasma infection, 6 plants and 3 insect samples were subjected to PCR using universal phytoplasma nested primers P1/P7 and R16F2n/R16R2 (Lee et al. 2004), and the amplicons were sequenced. Two identical consensus sequences resulted from the plant and insect samples (basepairs 1-1250, GenBank accession KY047614 and KY047615, respectively). Restriction Fragment Length Polymorphism analysis was conducted using iPhyClassifier analysis tool (Zhao et al. 2009). Both sequences were identified as 16Sr group VI, subgroup A, showing 99.7% similarity with ‘Candidatus Phytoplasma trifolii’ (GenBank accession AY390261). Candidatus Phytoplasma trifolii was reported in peppers in Mexico (Mauricio-Castillo et al. 2015), but the specific strain of 16SrVI-A group was not determined. Therefore, this is the first report of the Clover proliferation group strain BLTVA phytoplasma in peppers and its insect vector in this region of Mexico. BLTVA is an economically important disease in solanaceous crops in the US (Lee et al. 2004), and poses a threat to the pepper region in Mexico, stressing the need for control of the beet leafhopper vector of BLTVA phytoplasma.