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David Douches, Walter Pett, Diedrich Visser, Joseph Coombs, Kelly Zarka, Kimberly Felcher, Gurling Bothma, Johan Brink, Muffy Koch and Hector Quemada

Potato tuber moth is a serious pest of potato in South Africa. This insect causes damage during the growing season and in potato storage. The life cycle of potato tuber moth can be completed within 20 to 30 d and there may be as many as 12

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Kelly A. Zarka, Ria Greyling, Inge Gazendam, Dean Olefse, Kimberly Felcher, Gurling Bothma, Johan Brink, Hector Quemada and David S. Douches

Potato tuber moth is a serious pest of potatoes in South Africa, causing losses as much as $5.4 million per annum to the South African potato industry ( Visser and Schoeman, 2004 ). Larval mining on the plant results in the loss of leaf tissue

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Melanie M. Davidson, Jeanne M.E. Jacobs, Jill K. Reader, Ruth C. Butler, Christina M. Frater, Ngaire P. Markwick, Steve D. Wratten and Anthony J. Conner

Transgenic potato (Solanum tuberosum L.) lines of two cultivars, Ilam Hardy and Iwa, were developed using Agrobacterium-mediated transformation to transfer a cry1Ac9 gene under the transcriptional control of the CaMV 35S promoter. PCR confirmed the presence of the nptII selectable marker gene in all recovered lines. All ten lines of Ilam Hardy and 14 of 15 Iwa lines were PCR-positive for the cry gene. In greenhouse trials, all Ilam Hardy transgenic lines produced phenotypically normal plants and significantly inhibited larval growth of potato tuber moth (Phthorimaea operculella Zeller). In contrast, only 60% of the Iwa transgenic lines produced phenotypically normal plants, but all lines positive for the cry gene significantly inhibited larval growth. All transgenic lines with a greenhouse appearance equivalent to the nontransgenic controls and improved resistance to potato tuber moth larvae were planted in the field. Three of the ten Ilam Hardy lines and two of the eight Iwa lines retained phenotypically normal appearance in the field and produced tuber yields equivalent to the nontransgenic controls. All five of these transgenic lines significantly inhibited larval growth of potato tuber moth on excised field-grown leaves. A high correlation was established between larval growth indices from the greenhouse and the field. A transgenic line from each cultivar inhibited larval growth by over 40%, and the line derived from Ilam Hardy prevented pupation of all larvae. Southern analysis on these five elite lines revealed that they contained either one or two copies of the cry1Ac9 gene. The amount of Cry protein in all transgenic lines tested was less than 60 ng·g-1 of fresh leaf tissue. A transgenic line from each cultivar was identified with comparable phenotypic appearance and yield to their parent cultivars coupled with high resistance to PTM.

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D.S. Douches, W. Li, K. Zarka, J. Coombs, W. Pett, E. Grafius and T. El-Nasr

The potato tuber moth (Phthorimaea operculella Zeller) is the primary insect pest of cultivated potato (Solanum tuberosum L.) in tropical and subtropical regions, causing both foliar and tuber damage. In contrast, the Colorado potato beetle (Leptinotarsa decemlineata Say) is the most important insect pest in the northern potato production latitudes. The codon-modified Bacillus thuringiensis Bt-cry5 gene (revised nomenclature cry1IaI), specifically toxic to Lepidoptera and Coleoptera, was transformed into cultivar Spunta using an Agrobacterium vector to provide resistance to both potato tuber moth and Colorado potato beetle. The Bt-cry5 gene was placed downstream from the constitutive CaMV35S promoter. Two transgenic 'Spunta' clones, G2 and G3, produced high levels of mortality in first instars of potato tuber moth in detached-leaf bioassays (80% to 83% mortality), laboratory tuber tests (100% mortality), and field trials in Egypt (99% to 100% undamaged tubers). Reduced feeding by Colorado potato beetle first instars was also observed in detached-leaf bioassays (80% to 90% reduction). Field trials in the United States demonstrated that the horticultural performance of the two transgenic lines was comparable to 'Spunta'. These Bt-cry5 transgenic potato plants with high potato tuber moth resistance have value in integrated pest management programs.

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Wenbin Li, Kelly A. Zarka, David S. Douches, Joseph J. Coombs, Walter L. Pett and Edward J. Grafius

The codon-modified cryV-Bt gene (cryV-Bt) from Bacillus thuringiensis subsp. kurstaki Berliner, which is specifically toxic to Lepidoptera and Coleoptera insects, and a potato virus Yo coat protein gene (PVYocp), in which the aphid transmission site was inactivated, were cotransformed into potato (Solanum tuberosum L.) `Spunta' via Agrobacterium tumefaciens Conn. We demonstrated the integration and expression of both genes by molecular analysis and bioassays. All cryV-Bt/PVYocp-transgenic lines were more resistant to potato tuber moth (Phthorimaea operculella Zeller) and PVYo infection than nontransgenic `Spunta'. Four cryV-Bt/PVYocp transgenic lines were equal in potato tuber moth mortality to a cryV-Bt transgenic line, but of these four only two lines were equivalent in PVYo titer levels to a PVYocp-transgenic line. We identified two transgenic lines, 6a-3 and 6a-5, which showed greater resistance to potato tuber moth and PVYo than the other cryV-Bt/PVYocp transgenic lines. This study indicated that multiple genes, conferring insect pest resistance and virus resistance, could be engineered into and expressed simultaneously in a potato cultivar.

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D.S. Douches, A.L. Westedt, K. Zarka, B. Schroeter and E.J. Grafius

tuber moth cultures. The cost of publishing this paper was defrayed in part by the payment of page charges. Under postal regulations, this paper therefore must be hereby marked advertisement solely to indicate this fact.

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Hector Quemada, Kelly Zarka, Walter Pett, Gurling Bothma, Kimberly Felcher, Hope Mirendil, Muffy Koch, Johan Brink and David Douches

‘SpuntaG2’, a transgenic potato resistant to potato tuber moth, has been previously described by Douches et al. (2002) . ‘SpuntaG2’ contains two foreign genes: cry1Ia1 , a gene derived from the soil-borne bacterium Bacillus thuringiensis and

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Maria A. Estrada, Kelly Zarka, Susannah Cooper, Joseph Coombs, David S. Douches and Edward J. Grafius

Chakrabarti, S.K. Lutz, K.A. Lertwiriyawong, B. Svab, Z. Maliga, P. 2006 Expression of the cry9Aa2 B.t . gene in tobacco chloroplasts confers resistance to potato tuber moth Transgen. Res. 15 481 488

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John L. Coffey, Alvin M. Simmons, B. Merle Shepard, Yaakov Tadmor and Amnon Levi

. ( Seenivasan et al., 2005 ), and the potato tuber moth, Phthorimaea operculella Zell. ( Sharaby et al., 2009 ). There has not been any report of olfactometer assays of C. colocynthis and whiteflies. The objective of this study was to evaluate selected

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Richard O. Nyankanga, Ocen Modesto Olanya, Hans C. Wien, Ramzy El-Bedewy, John Karinga and Peter S. Ojiambo

). Similarly, abrasion of tubers after harvest ( Lambert et al., 1998 ) or wounds created by insects such as potato tuber moth or pathogen infections by bacterial soft rot ( Pectobacterium carotovora ) may also induce wounds and rot in tubers, which may