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3566

In August of 2011, potato (Solanum tuberosum) tubers grown in the lower Columbia Basin of southern Washington State and northern Oregon were observed with internal discolorations suggestive of zebra chip disease (ZC). Symptoms included brown spots, streaks, and stripes in and near the vascular tissue, typical of ZC (1). Symptoms were observed in cvs. Alturas, Russet Norkotah, Pike, Ranger Russet, Umatilla Russet, and Russet Burbank. Foliar symptoms on plants that produced symptomatic tubers included purple discoloration in upper leaves, leaf rolling, axial bud elongation, chlorosis, leaf scorch, and wilt. Tissue was taken from two symptomatic tubers each of cvs. Alturas and Russet Norkotah, three tubers of cv. Umatilla Russet, and one tuber of cv. Pike. These tubers were tested by PCR for “Candidatus Liberibacter solanacearum”, an unculturable alphaproteobacterium associated with ZC (1,4). Primers specific for the 16S rDNA were CLipoF (4) and OI2c (3), and primers OMB 1482f and 2086r were specific for the outer membrane protein (2). All of these samples, except one Umatilla tuber, were positive for the bacterium. The 16S rDNA and OMB amplicons from one symptomatic tuber each of Alturas (from Washington) and Pike (from Oregon) were cloned and three clones of each were sequenced. BLAST analysis of the consensus sequences confirmed “Ca. L. solanacearum”. The 16S sequences (1,071 bp) from the two tubers were identical and showed 99 to 100% identity to a number of 16S rDNA sequences of “Ca. L. solanaceaum” in GenBank (e.g., Accession Nos. HM246509 and FJ957897). The 16S rDNA sequences were deposited in GenBank as Accession Nos. JN848751 and JN848753. Consensus sequences of the two OMB clones (605 bp; deposited in GenBank as Accession Nos. JN848752 and JN848754) were identical and showed 97% identity to the two “Ca. L. solanacearum” OMB sequences in GenBank (Accession Nos. CP002371 and FJ914617). Potato psyllids (Bactericera cockerelli Sulc), the vector of “Ca. L. solanacearum”, were present in ZC-affected fields in Oregon and Washington and the bacterium was confirmed by PCR in 5 to 10% of 128 adult psyllids collected from two fields. On the basis of foliar and tuber symptoms, specific PCR amplification with two primer pairs, sequence analyses, and the presence of Liberibacter-infected potato psyllids, ZC and “Ca. L. solanacearum” are present in potatoes in Oregon and Washington State. Washington and Oregon together grow ~80,000 ha of potatoes. ZC has caused significant economic damage to potatoes in Texas, Mexico, Central America, and New Zealand (1). Therefore, ZC may pose a risk to agriculture in Oregon, Washington, and neighboring states. However, the potential for development of widespread and serious disease will depend upon the arrival time and number of infective potato psyllids entering the region. References: (1) J. M. Crosslin et al. Online publication. doi:10.1094/PHP-2010-0317-01-RV, Plant Health Progress, 2010. (2) J. M. Crosslin et al. Southwest. Entomol. 36:125, 2011. (3) S. Jagoueix et al. Mol. Cell. Probes 10:43, 1996. (4) G. A. Secor. Plant Dis. 93:574, 2009.

Carrot (Daucus carota) plants with symptoms resembling those associated with the carrot psyllid Trioza apicalis and the bacterium “Candidatus Liberibacter solanacearum” (1–4) were observed in 70% of commercial fields in southern Sweden in August 2011, with approximately 1 to 45% symptomatic plants per field. T. apicalis, a pest of carrot in northern and central Europe, including Sweden, can cause as much as 100% crop loss and is associated with “Ca. L. solanacearum” (1–4). Symptoms on affected plants include leaf curling, yellow and purple discoloration of leaves, stunted growth of shoots and roots, and proliferation of secondary roots (3). Carrot plant and psyllid samples were collected from fields in the province of Halland. Total DNA was extracted from petiole and root tissues of 33 symptomatic and 16 asymptomatic plants (cvs. Nevis and Florida), with the cetyltrimethylammonium bromide (CTAB) buffer extraction method (2,3). DNA was also extracted from 155 psyllids (3). DNA samples were tested by PCR using primer pairs OA2/OI2c (5′'-GCGCTTATTTTTAATAGGAGCGGCA-3′/5′-GCCTCGCGACTTCGCAACCCAT-3′) and CL514F/R (5′-CTCTAAGATTTCGGTTGGTT-3′/5′-TATATCTATCGTTGCACCAG-3′), to amplify a portion of 16S rDNA and rplJ/rplL ribosomal protein genes, respectively, of “Ca. L. solanacearum” (2,3). A 1,168-bp 16S rDNA fragment was detected in the DNA from all 33 symptomatic and two asymptomatic plants, and a 668-bp rplJ/rplL fragment was amplified from the DNA of all 33 symptomatic and four asymptomatic plants, indicating the presence of liberibacter. DNA from 23 and 49 psyllid samples yielded similar amplicons with OA2/OI2c and CL514F/R primer pairs, respectively. Amplicons from the DNA of four carrot roots and three T. apicalis with each primer pair were cloned (pCR2.1-TOPO; Invitrogen, Carlsbad, CA) and three clones of each of the 14 amplicons were sequenced (MCLAB, San Francisco, CA). BLAST analysis of the 16S rDNA consensus sequences from carrot (GenBank Accession No. JN863095) and T. apicalis (GenBank Accession No. NJ863096) showed 100% identity to those of “Ca. L. solanacearum” previously amplified from carrot (GU373048 and GU373049) and T. apicalis (GU477254 and GU477255) from Finland (2,3). The rplJ/rplL consensus sequences from carrot (GenBank Accession No. JN863093) and T. apicalis (GenBank Accession No. JN863094) were 99% identical to the sequences of rplJ/rplL “Ca. L. solanacearum” ribosomal protein gene from carrots in Finland (GU373050 and GU373051). To our knowledge, this is the first report of “Ca. L. solanacearum” associated with carrot and T. apicalis in Sweden. The disease associated with this bacterium caused millions of dollars in losses to potato and several other solanaceous crops in North and Central America and New Zealand (1). This plant pathogen is also associated with significant economic damage to carrot crops observed in Finland (2,3). References: (1) J. E. Munyaneza. Southwest. Entomol. 35:471, 2010. (2) J. E. Munyaneza et al. Plant Dis. 94:639, 2010. (3) J. E. Munyaneza et al. J. Econ. Entomol. 103:1060, 2010. (4) A. Nissinen et al. Entomol. Exp. Appl. 125:277, 2007.

ABSTRACT We present the genome sequence of “ Candidatus Mycoplasma haemominutum” strain Birmingham 1, a low-pathogenicity feline hemoplasma strain.