Nigel A. Harrison¹*, Maria Narváez², Humberto Almeyda³, Ivan Cordova², Monica L. Carpio¹ and Carlos Oropeza²

¹ University of Florida, Research and Education Center, 3205 College Avenue, Fort Lauderdale, FL 33314, USA
² Centro de Investigación Cientifíca de Yucatán, Merida, Yucatán, Mexico
³ INIFAP, Monterrey, Nuevo León, Mexico

*naha@ufl.edu

Accepted for publication 22/07/02

Concern about the possible westward spread of lethal yellowing (LY) disease of coconut (Cocos nucifera L.) palm, from the LY-endemic Yucatan peninsula to the centre of coconut production in southwestern Mexico, prompted surveys of the latter region for this phytoplasma disease. Coconut palms displaying prominent leaf yellowing symptoms suggestive of LY were identified during these surveys, at separate locations in the states of Oaxaca in 1997 and Guerrero in 1998 (Fig. 1). Although nutfall and inflorescence necrosis symptoms, which invariably precede foliar yellowing on LY-diseased palms, were atypically absent or delayed, affected palms were removed as a precautionary measure against further disease spread.

Figure 1. A local tall-type coconut palm in Puerto Escondido, Oaxaca displaying prominent leaf yellowing without nutfall (A); and a local tall-type coconut palm in Monte Alto, Guerrero displaying advanced leaf yellowing (B) that was followed by partial nutfall and inflorescence necrosis (C).

Leaf or inflorescence tissues from eleven symptomatic palms were assayed for phytoplasma infection using a polymerase chain reaction (PCR) assay employing phytoplasma universal rRNA primer pair P1/P7 as previously described (Harrison et al., 2002). An rDNA product (about 1.8 kb) was amplified from nine leaf yellowing (LfY)-affected palms. Reassessment of all samples by PCR employing LY-specific nonribosomal primer pair LYF1/LYR1 (Harrison et al., 1994), failed to amplify a DNA product from any LfY-diseased palm samples, or from DNAs of known lethal yellowing (16SrIV) group phytoplasmas coconut lethal decline (LDY) and Carludovica palmata yellows (CPY), or symptomless coconut included as negative experimental controls. A 1 kb DNA product was generated from DNA of LY reference phytoplasmas from Yucatan and Florida, indicating that LfY-affected palms harbored phytoplasmas differing from the LY agent, a subgroup 16SrIV-A strain (Lee et al., 1998).

Figure 2. AluI endonuclease digests of phytoplasma rDNA products amplified by nested PCR employing rRNA primer pairs P1/P7 followed by P1m/LY16-23Sr. Samples for comparison consisted of coconut leaf yellowing (LfY)-affected palms in Guerrero (LfY1 and LfY2) and in Oaxaca (LfY3, LfY4 and LfY5); and phytoplasmas associated with Carludovica palmata yellows (CPY); coconut lethal decline (LDY) and lethal yellowing (LYM) in Mexico; or coconut lethal yellowing in Florida (LYF).

AluI endonuclease digests of PCR products (1.7 kb) reamplified from primary P1/P7 products by P1m (5’-TCCTGGCTCAGGATTAC-3’) and LY16-23Sr (5’-TTGAGAATTTACGT TGTTTATCTAC-3’)-primed PCR, yielded two distinct fragment patterns among diseased palms (Fig. 2). One pattern obtained for four palms in Guerrero and three palms in Oaxaca is illustrated by samples LfY1, LfY2 and LfY3. A second ambiguous pattern was obtained for two other palms (LfY4 and LfY5) in Oaxaca. From these products, six unique 16S rDNA sequences were obtained (GenBank accessions AF500329, AF500330, AF500331, AF500332, AF500333, AF500334), including four dissimilar sequences cloned from the two co-infected Oaxaca palms (LfY4 and LfY5). Subsequent phylogenetic analysis determined that three sequences were most similar, albeit distinct from LDY, a subgroup 16SrIV-B phytoplasma, whereas the remaining sequences were similar to or indistinguishable from CPY, a subgroup 16SrIV-C strain (Harrison et al., 2002).

This is the first report of 16SrIV group phytoplasmas infecting coconut palms on the Pacific coast of the Americas.

Florida Agricultural Experiment Journal Series No. R-08768

References

Harrison NA, Richardson PA, Kramer JB, Tsai JH, 1994. Detection of the phytoplasma associated with lethal yellowing disease of palms in Florida by polymerase chain reaction. Plant Pathology 43, 998-1008.

Harrison, NA, Womack M, Carpio ML, 2002. Detection and characterization of a lethal yellowing (16SrIV) group phytoplasma in Canary Island date palms affected by lethal decline in Texas. Plant Disease 86, 676-681.

Lee I-M, Gundersen-Rindal DE, Davis RE, Bartoszyk IM, 1998. Revised classification scheme of phytoplasmas based on RFLP analyses of 16S rRNA and ribosomal protein gene sequences. International Journal of Systematic Bacteriology 48, 1153-1169.

The British Society for Plant Pathology