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Zhan'ao Deng, Fred G. Gmitter Jr., Shunyuan Xiao, and Shu Huang

Citrus tristiza virus (CTV) is the most-significant viral pathogen of citrus in the world. Rapid decline of trees on sour orange and stem pitting of grapefruit and sweet orange, two diseases induced by CTV, severely jeopardize citrus production worldwide. It is recognized that all future rootstocks should be resistant to this virus, and scion resistance to stem pitting stains is desirable. To facilitate introgression of the CTV resistance gene from Poncirus trifoliata and development of CTV-resistant varieties in citrus, gene mapping projects have been initiated and more than a dozen RAPD markers have been identified with tight linkage to the resistance gene. As part of our efforts to use marker-assisted selection with a large number of crosses, and ultimately to accomplish map-based cloning of the CTV resistance gene, we have been converting the most tightly linked RAPD markers into SCAR (sequence characterized amplified region) markers by cloning, sequencing the marker fragments, and designing locus-specific primers. One codominant and several dominant SCARs have been developed thus far. The updated progress and utilization of these SCARs in marker-assisted selection and possibly in characterization of a BAC library will be presented and discussed.

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L. Gene Albrigo and Jude W. Grosser

In Florida, pesticides, nutritional and growth regulators are often sprayed in tank mixes to reduce sprayer use. Many individual spray components are phytotoxic and result in spray burns in combination or if applied with adjuvants. The toxicity level of standard spray materials is not known and new product testing for phytotoxicity is not routine. Three tests were developed to allow testing of cellular and whole fruit susceptibility to spray chemicals. Cell suspension cultures initiated from `nucellar derived' embryonic callus of `Hamlin' sweet orange were grown in log phase for 2 weeks with various levels of test chemicals. Fresh weight increase was measured. Peel disks of orange or grapefruit were grown for 4 weeks on solid media. Color changes and callus growth were used to evaluate phytotoxicity. Dilute sprays and droplet applications to on-tree-fruit were used to evaluate individual and combinations of chemicals with and without spray adjuvants. The 3 tests combined effectively demonstrated levels of phytotoxicity and are useful for testing new citrus production chemicals.

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Eliezer Louzada, Omar Vazquez, Sandy Chavez, Mamoudou Sétamou, and Madhurababu Kunta

Citrus Huanglongbing (HLB, also known as “citrus greening”), an important disease worldwide, is associated with three species of phloem-limited Candidatus liberibacter, of which Candidatus L. asiaticus (CLas) is the predominant one that has severely affected citrus production. TaqMan real-time polymerase chain reaction (PCR) (TM) has been the standard and very efficient method to diagnose several strains of Candidatus Liberibacter in citrus; however, it detects total bacteria and is unable to differentiate dead from live Liberibacter. The detection of only live bacteria is essential for testing methods of control for this important citrus disease. It is well known that ethidium monoazide and propidium monoazide (PMA) are compounds that supposedly enter only dead or membrane-damaged bacteria, intercalate the DNA strand, and make the DNA unavailable for amplification by PCR. These compounds are widely used when extracting the plant DNA to detect only live bacteria. In this research, we tested primers amplifying products from 79 to 1160 bp in TM and SYBR Green real-time PCR (SG) and PMA as DNA intercalating compound. Specifically, primers amplifying a 500-bp amplicon in SG provided the most reliable live-only detection, whereas those producing a smaller amplicon were unable to distinguish between live and dead. This is the first report of testing primers amplifying various amplicon sizes for the detection of only live CLas cells in citrus.

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Kim D. Bowman, Jeffrey P. Shapiro, and Stephen L. Lapointe

. Some of the seeds for this work were provided by the USDA Clonal Germplasm Repository for Citrus and Dates, Riverside, Calif. The described research was funded, in part, by grant No. 951-6 from the Florida Citrus Production Research Advisory Council

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T.L. Weinert, S.A. White, T.L. Thompson, and M.A. Maurer

Citrus production in the southwestern U.S. is highly dependent on inputs of irrigation and N fertilizer to achieve optimum fruit yield and quality. Microsprinkler irrigation may allow for substantial increase in efficiency of N and water application. However, best management practices have not yet been developed for microsprinkler use, particularly on newly established citrus trees. Experiments were conducted during 1997–98 in central Arizona to evaluate the effects of various N rates and fertigation frequencies on growth and N partitioning in young `Newhall' navel oranges planted in Apr. 1997. Two experiments were conducted, each with factorial combinations of N rate and fertigation frequency. In one experiment, non-labeled N fertilizer was used and in the other 15N-labeled N fertilizer. Trunk diameter, leaf N, and 15N partitioning in the trees were measured. During 1997, neither trunk diameter or leaf N were affected by N rate or fertigation frequency. No more than 6% of the N applied was taken up by the trees, and about 50% of the fertilizer N taken up was found in the leaves. Trees grew much more rapidly in 1998. Leaf N in fertilized plots was significantly higher than in control plots, but leaf N in all trees remained above the critical level of 2.5%. Despite rapid tree growth during 1998, no more than 25% of the fertilizer N applied was taken up by the trees. About 60% of the fertilizer N taken up was found in the leaves. Results suggest that N applications are not needed during the first growing season after planting for microsprinkler-irrigated citrus trees in the Southwest. Only modest rates (68 to 136 g/tree) will be needed during the second season to maintain adequate tree reserves.

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Megh Singh, Mayank Malik, Analiza H.M. Ramirez, and Amit J. Jhala

Citrus (Citrus spp.) is one of the most important crops in Florida agriculture. Weed control is a major component in citrus production practices. If not controlled, weeds may compete with citrus trees for nutrients, water, and light and may also increase pest problems. Herbicides are an important component of integrated weed management program in citrus. Saflufenacil, a new herbicide registered for broadleaf weed control in citrus, can be applied alone or in a tank mix with other herbicides to improve weed control efficacy. A total of six field experiments were conducted in 2008 and 2009 to evaluate the efficacy of saflufenacil applied alone or in a tank mix with glyphosate and pendimethalin for weed control. In addition, experiments were also conducted to evaluate phytotoxicity of saflufenacil applied at different rates and time intervals in citrus. The results suggested that saflufenacil applied alone was usually effective for early season broadleaf weed control; however, weed control efficacy reduced beyond 30 days after treatment (DAT) compared with a tank mix of saflufenacil, glyphosate, and pendimethalin. For example, control of weeds was ≤70% when saflufenacil or glyphosate applied alone compared with tank mix treatments at 60 and 90 DAT. Addition of pendimethalin as a tank mix partner usually resulted in better residual weed control compared with a tank mix of saflufenacil and glyphosate, and this herbicide mixture was comparable with grower's adopted standard treatment of a tank mix of glyphosate, norflurazon, and diuron and several other tank mix treatments. Saflufenacil applied once in a season at different rates or even in sequential applications did not injure citrus trees when applied according to label directions. It is concluded that with its novel mode of action, saflufenacil tank mixed with glyphosate and pendimethalin would provide citrus growers with another chemical tool to control broadleaf and grass weeds.

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Raquel L. Boscariol, Mariza Monteiro, Elizabete K. Takahashi, Sabrina M. Chabregas, Maria Lucia C. Vieira, Luiz G.E. Vieira, Luiz F.P. Pereira, Francisco de A.A. Mourão Filho, Suane C. Cardoso, Rock S.C. Christiano, Armando Bergamin Filho, Janaynna M. Barbosa, Fernando A. Azevedo, and Beatriz M.J. Mendes

Citrus canker, caused by Xanthomonas axonopodis Starr and Garces pv. citri (Hasse) Vauterin et al., is one of the main problems affecting citrus production. In order to obtain resistance to phytopathogenic bacteria, insect genes, coding for antimicrobial proteins, have been used in plant genetic transformation. In this study, transgenic Citrus sinensis (L.) Osb. `Hamlin' plants expressing the antimicrobial insect-derived attacin A gene (attA) were obtained by Agrobacterium tumefaciens (Smith and Towns.) Conn-mediated transformation. Initially, the cDNA clone was used to construct a binary plasmid vector (pCattA 2300). The construction included the native signal peptide (SP) responsible for directing the insect protein to the extracellular space where bacteria is supposed to accumulate in vivo. In order to investigate the native SP effectiveness in a plant model system, onion (Allium cepa L.) epidermal cells were transformed, via biobalistics, using plasmids containing the attA gene with or without SP, fused with the green fluorescent protein gene (pattA 1303 and pSPattA 1303). Fluorescence accumulation surrounding the cells was observed only in tissues transformed with the plasmid containing the gene with SP, indicating the protein secretion to the apoplast. Citrus transformation was confirmed by PCR and Southern blot hybridization analysis in 12 regenerated plants. Transcription of attA gene was detected by Northern blot analysis in all transgenic plants. Eight selected transgenic lines were propagated and inoculated with a 106 cfu/mL suspension of the pathogen X. axonopodis pv. citri. Compared to control (non-transformed plant), seven transgenic lines showed a significant reduction in susceptibility to citrus canker. The results obtained here indicate the potential use of antibacterial proteins to protect citrus from bacterial diseases.

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Thomas A. Obreza and Arnold Schumann

article are to describe how N and P are currently managed to grow citrus in Florida, to provide an accounting of the citrus N budget, to describe N leaching and P runoff from citrus production, to outline best management practices to improve citrus

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Leo G. Albrigo and Ed W. Stover

The severe citrus (Citrus sp.) disease Huanglongbing (HLB), associated with Candidatus Liberibacter asiaticus, has resulted in widespread tree decline in Florida and overall citrus production is now the lowest it has been in 50 years. More than 80% of Florida citrus trees are HLB affected, and most growers attempt to sustain production on infected trees through good asian citrus psyllid (Diaphorina citri) control and enhanced fertilization and irrigation management. Although production appears to benefit from these treatments, preharvest fruit drop is considerably greater than on uninfected trees. U.S. Department of Agriculture (USDA) data indicate that Florida statewide fruit drop has increased by 10% to 20% of the entire crop in the last three growing seasons, essentially doubling the historical levels. Extensive research is underway to identify solutions to HLB, but it is essential to maintain production on existing trees to sustain the industry in the near term. For decades, several plant growth regulators (PGRs) have been labeled to reduce preharvest fruit drop in commercial citrus. Trials of these materials, other nonlabeled PGRs, and some fungicides were conducted in two seasons to determine if fruit drop could be reduced. Randomized complete block design experiments were established using four to six replications of four- to six-tree groups as experimental units, blocked spatially. In 2013–14, sprays of gibberellic acid (GA), 2,4-dichlorophenoxyacetic acid (2,4-D), 1-naphthaleneacetic acid (NAA), S-abscisic acid (S-ABA), aminoethoxyvinylglycine (AVG), and 1-methylcyclopropene (1-MCP) were applied once or twice alone or in some combinations at standard rates to trees in various mature blocks of ‘Valencia’ and ‘Pineapple’ sweet orange (Citrus sinensis), ‘Star Ruby’ grapefruit (Citrus paradisi), or ‘Murcott’ tangor (Citrus reticulata ×C. sinensis) in central Florida in the Indian River area. Only 1 of the 10 individual trials had treatments with significantly lower drop rates than controls; and when pooled across all experiments, GA + 2,4-D reduced number of fruit dropped per tree 4%, but only at P = 0.10. NAA, S-ABA, AVG, and 1-MCP had no effect and were not used the following year. Starting in 2014, treatments were initiated earlier in the season with greater effort to minimize variability: GA; 2,4-D; GA + 2,4-D; a natural GA, indolebutyric acid, cytokinin mix; and strobilurin fungicides were applied to 22 mature blocks of ‘Hamlin’ and ‘Valencia’ sweet orange trees. In 2014–15, only three of the 11 individual ‘Hamlin’ trials and one of the 11 ‘Valencia’ trials included a treatment with significant drop reduction compared with controls. However, when all the tests on ‘Hamlin’ were pooled, there was a significant 5% reduction in total crop drop for GA + 2,4-D and significant reductions with many of these PGRs alone, but in only one case with fungicide treatments. When all tests on ‘Valencia’ were combined, 2,4-D reduced drop significantly but only by 2% of the total crop (14% drop vs. 16% drop), but fruit drop in ‘Valencia’ blocks was near the historical average in control trees. Soil conditions and tree conditions were similar across all test sites and there were no apparent relationships between product efficacy and observed tree condition or any other grove characteristics. In addition, four ‘Hamlin’ and four ‘Valencia’ blocks were treated with 1/4 rates of 2,4-D + 6-benzyladenine every 45 days during the growing season (six sprays) and three of the eight individual trials showed significant reductions in drop: when pooled, these treatments reduced drop by 3% in ‘Valencia’ and 6% in ‘Hamlin’. At this time, PGRs cannot be recommended as a consistent way to reduce fruit drop related to HLB, but further work needs to be conducted to refine the most promising treatments.

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Kelly T. Morgan, Lincoln Zotarelli, and Michael D. Dukes

centers of citrus production in the United States are located in Florida (67%), California (28%), Texas (4%), and Arizona (1%) and total 9.4 million tons or 13% of world production. Florida citrus production in 2006–2007 was orange ( Citrus sinensis ) (80