Almond Leaf Research Articles

Green Analytical Methodology Using Indian Almond (Terminalia Catappa L.) Leaf Extract for Determination of Aluminum Ion in Waste Water from Ceramic Factories

The use of natural reagents from plant extracts for chemical analysis is one of the approaches in the development of low cost and environmentally friendly green analytical chemistry methodology. Here, crude extract from Indian Almond (Terminalia Catappa L.) leaves was used for colorimetric determination of aluminum by monitoring the absorbance of the Al(3+)-extract complex at 435 nm. Dry leaves and freeze-dried fresh leaf extract can be kept for extended use. A simple flow injection analysis (FIA) system was also employed for rapid analysis (approximately 180 injections/h). The linear working range up to 100 mg L(-1) was established with a detection limit (blank + 3SD) of 0.8 mg L(-1), a limit of quantitation (blank + 10SD) of 2.4 mg L(-1), and a relative standard deviation of 3-5%. This simple green analytical chemistry methodology was applied for the determination of Al(3+) in waste water samples from ceramic factories. The results agreed well with the results obtained from the ICP-OES technique.

Almond Leaf Scorch Disease Development on Almond Branches High-Grafted on Peach Rootstock

Development of almond leaf scorch (ALS) disease was monitored on young almond (Prunus dulcis 'NePlus' and 'Peerless') shoots produced from almond scion wood that was high-grafted on peach rootstocks (P. persica 'Queencrest/Nemaguard'), after the almond shoots were mechanically inoculated with Xylella fastidiosa. The objective of this study was to evaluate the potential movement of X. fastidiosa through almond-peach graft unions. ALS symptoms developed on both X. fastidiosa-inoculated and uninoculated almond shoots that were high-grafted on different peach limbs of the same tree in September following inoculations with X. fastidiosa made in June and July 2002, respectively, when the average distance in peach rootstock between the two almond-peach graft unions was 30.5 cm or shorter. No ALS symptoms were observed on uninoculated almond shoots late in the growing season of the second year. The incidence of ALS-affected leaves on shoots inoculated with X. fastidiosa decreased in the second year on the inoculated shoots of Peerless as determined by the number of inoculated shoots showing ALS symptoms in 2002 versus 2003. No visible ALS symptoms were observed in NePlus late in the growing season of the second year, suggesting that survival of X. fastidiosa in NePlus was less than in Peerless. These data demonstrate that movement of X. fastidiosa through two almond-peach graft unions was possible during the year of inoculation when the peach rootstock stem distance between the two almond-peach graft unions was minimal. However, X. fastidiosa may not survive the winter season or survived so poorly as not to be able to produce ALS symptoms in the second growing season on the uninoculated shoots that previously showed ALS symptoms in the fall of the year when an adjacent shoot was inoculated. The results suggest that high-grafting almond scion wood on multiple peach rootstock limbs may provide a means to limit movement of X. fastidiosa from one almond limb to another on the same tree.

Determination of the Antibacterial Effects of Prunus amygdalinus (Almond Leaf) and Ocimum Gratissimum (Basil Leaf) on Selected Bacteria

antimicrobial acts of leaf extracts of Ocimumgratissimum and Prunusamygdalinus were checked on Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa and Salmonella typhi using agar cup plate method. The outcomes demonstrated that the chosen organisms were susceptible to the plant extracts at concentrations of 500mg, 50mg and 5mg respectively with diameter zone of inhibition (DZI) of about 12.00± 2.00, 13.00± 2.00, 11.60±3.40, and 12.30±3.30. Phytochemical screening of the leaf extract of Ocimumgratissimum manifested the existence of alkaloid, phenols, glycosides, saponin, steroids and triterpenes while the leaf extract of Prunusamygadalinus demonstrated the existence of saponin, flavonoid and triterpenes. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) assayed reveals MIC of 5mg and MBC of 50mg respectively. Ocimumgratissimum and Prunusamygdalinus extracts may be applied to treat disorder caused by these organisms.

Phenology of Xylella fastidiosa and Its Vector Around California Almond Nurseries: An Assessment of Plant Vulnerability to Almond Leaf Scorch Disease.

Management of almond leaf scorch disease requires knowledge of all possible infection pathways. The disease is caused by the xylem-limited bacterium Xylella fastidiosa, which is transmitted by several species of sharpshooters. The objectives of this research were to elucidate the fate of bacteria in planta after inoculations in almond nursery plants and to determine patterns of insect vector population dynamics and temporal distribution of X. fastidiosa-infected plants relative to host plant assemblages in habitats surrounding commercial nurseries. In an experimental nursery, disease incidence was markedly affected by rootstock type. Prior to bud grafting, 'Nemaguard' rootstock seedlings were not susceptible to bacterial infection. After bud grafting with a susceptible scion ('Sonora'), scions were susceptible to infection regardless of rootstock genotype. Surveys near commercial nurseries revealed that only habitats with permanent grass cover sustained vector populations throughout the season. A total of 87 plant samples tested positive for X. fastidiosa (6.3%) using enzyme-linked immunosorbent assay (ELISA), with a higher number of X. fastidiosa-infected plants found in weedy alfalfa fields than in other habitat types. Among plant species infected by X. fastidiosa, 33% were winter annuals, 45% were biennials or perennials, and 22% were summer annuals. Collectively, these findings identified a potential pathway for X. fastidiosa infection of almonds in nursery situations.

Management of Almond Leaf Scorch Disease: Long-Term Data on Yield, Tree Vitality, and Disease Progress.

Almond leaf scorch disease (ALSD) has been a chronic problem for California almond growers. This disease is caused by the bacterial pathogen Xylella fastidiosa and is transmitted by xylem-feeding insects. Previous research suggested that retaining, rather than roguing, ALSD-affected trees may be more economically beneficial because ALSD-affected trees produced a reasonable yield and did not die over a 3-year period. Because almond orchards are kept in production for approximately 25 years, longer-term data are needed to fully evaluate the merits of retaining ALSD-affected trees. Extension of yield evaluations from 3 to 5 years demonstrated that yield loss due to ALSD was consistent over 5 years, with yields of ALSD-affected trees reduced by 20 and 40% compared with unaffected trees for 'Nonpareil' and 'Sonora', respectively. To assess risk of ALSD-affected trees serving as a source of inocula for secondary (tree-to-tree) spread and to evaluate vitality of ALSD-affected trees, previous surveys of two orchards were extended from 3 to 6 or 7 years. The relationship between disease incidence (percentage of trees infected) and survey year was linear for all cultivars examined at both orchards. Furthermore, at each orchard, the spatial location of infections detected after the first survey was random with respect to the spatial location of infections identified during the first survey, suggesting that ALSD-affected trees retained in orchards did not serve as a source for secondary spread. Over the 6- to 7-year study period, death of ALSD-affected trees was rare, with only 9% of ALSD-affected trees dying. Because orchards used in this study had relatively high disease incidence, 61 orchards containing Sonora were surveyed to determine typical levels of ALSD incidence. ALSD was widespread, with at least one infected tree in 56% of orchards surveyed, but incidence was typically low (mean incidence = 0.47%). Collectively, the results suggest that retaining ALSD-affected trees may be economically beneficial in older orchards.

DETECTION AND IDENTIFICATION METHODS AND NEW TESTS AS USED AND DEVELOPED IN THE FRAMEWORK OF COST 873 FOR BACTERIA PATHOGENIC TO STONE FRUITS AND NUTS

The quarantine pathogen Xylella fastidiosa (EPPO A1 list), a xylem-inhabiting, vector-transmitted, Gramnegative, very slow growing bacterium, was cultured and properly described in 1987 in the USA as the cause of Pierce’s disease (PD) of grapevine (Vitis vinifera) and of phony peach disease (PPD) in peach (Prunus persica) and in 1993 in Brazil as the cause of citrus variegated chlorosis (CVC) in sweet orange (Citrus sinensis). Moreover, it was found that the bacterium also causes a number of so-called leaf scorch diseases in Prunus spp. [including almond leaf scorch (ALS) in P. armeniaca and plum leaf scald (PLS) in P. domestica], ornamental trees, Coffea arabica (CLC, in Brazil, isolated in 1995), and many other plant species (see under ‘Host range’). Many wild plants, including grasses, sedges and trees may carry the pathogen without showing symptoms. All these diseases are quite destructive, giving substantial yield losses. They occur mainly in tropical/subtropical areas, although leaf scorch diseases also occur in much colder climates, e.g. oak leaf scorch in eastern North America, as far north as Canada. Several pathogenic genotypes have been described, which are often host-specific (e.g., the strain will not cause disease if introduced to peach or plum). The following subspecies are currently known: (i) Xylella fastidiosa subsp. fastidiosa (erroneously named X. f. subsp. piercei) strains from cultivated grape, alfalfa, almond (two), and maple; (ii) X. f. subsp. multiplex, strains from peach, elm, plum, pigeon grape, sycamore, and almond; (iii) X. f. subsp. pauca, strains from citrus and probably those of coffee; (iv) X. f. subsp. sandyi, strains from Nerium oleander. X. f. subsp. taske was described from the ornamental tree Chitalpa tashkentensis on the basis of one gene sequence only and its status as a separate subspecies is therefore not ultimately established. Vectors are sharpshooter leafhoppers (Hemiptera:Cicadellidae) and froghoppers or spittlebugs (Hemiptera: Cercopidae), the most important of which in the USA include Cuerna costalis and Graphocephala versuta (PPD), Draculacephala minerva (green sharpshooter); G. atropunctata (blue-green sharpshooter, formerly Hordnia circellata) and Homalodisca vitripennis (formerly H. coagulata, glassy-winged sharpshooter, GWSS) (PD). CVC vectors in Brazil include Acrogonia terminalis, Dilobopterus costalimai and Oncometopia fascialis. Local possible vectors for Europe are Cicadella viridis and Philaenus spumarius (meadow spittle bug), as well as all other members of the family Cercopidae and the leafhopper subfamily Cicadellinae. The bacterium is, as far it is known, still absent from Europe. For more general information on the bacterium, epidemiology, control, etc. see Purcell (1997), Hopkins and Purcell (2002), EPPO/OEPP (1992), Chatterjee et al. (2008), Janse and Obradovic (2010 ), and http://www.cnr. berkeley.edu/xylella/ In order to study the disease and train on diagnosis in a pro-active way, the EU COST873 project organised in October 2010 a workshop specifically on X. fastidiosa detection and identification with excellent support and participation of two outstanding experts in the field, Prof. A.H. Purcell and Prof. R.P.P. Almeida from the University of California at Berkeley (USA). The workshop was hosted by IAM, Bari, Italy (http://cost873.ch/5_activites/ meeting_detail.php?ID=28). Most of the methods presented here have been found useful by the US experts over the years and were practised during the workshop.

Biosorption of palladium(II) and platinum(IV) from aqueous solution using tannin from Indian almond (Terminalia catappa L.) leaf biomass: Kinetic and equilibrium studies

A feasibility study was performed on Indian almond leaf biomass (Terminalia catappa L.) to remove palladium (Pd(II)) and platinum (Pt(IV)) ions from aqueous solution by biosorption. The biosorption characteristics of Pd(II) and Pt(IV) ions were investigated in terms of equilibrium, kinetics and thermodynamics. Optimum biosorption conditions were determined as a function of pH, biomass dosage, contact time, and temperature. Langmuir, Freundlich, and Dubinin–Radushkevich (D–R) models were applied to describe the biosorption isotherm. The Langmuir model fitted the equilibrium data better than the Freundlich isotherm. Palladium is more preferable with T. catappa L. than platinum. The maximum biosorption capacity (qm) of T. catappa L. biomass for Pd(II) and Pt(IV) ions were 41.86 and 22.50, respectively. The mean free energy values evaluated from the D–R model indicated that the biosorption of Pd(II) and Pt(IV) onto T. catappa L. biomass took place by chemical ion-exchange. The calculated thermodynamic parameters ΔG0, ΔH0 and ΔS0 indicate that the biosorption of Pt(II) and Pd(II) ions onto T. catappa L. biomass is feasible, spontaneous and exothermic. Biosorption kinetics using pseudo-first-order and pseudo-second-order kinetic models were also examined. Experimental data was found to be in good agreement with pseudo-second-order kinetics.

ASSESSMENT OF DROUGHT TOLERANCE IN ALMOND VARIETIES

Over three years (2004-2006), the chlorophyll concentration, surface and specific leaf weight, density, surface area, and stomate opening sensitivity were measured on different almond varieties growing in different soils and climates and with varying water availability. In almond, different anatomic and physiologic leaf behaviours are observed in response to specific water stress situations. Varieties such as ‘Guara’ have good cuticle isolation while others, such as ‘Masbovera’, have a high sensitivity to stomate closure. These preliminary results show a correlation between the chlorophyll meter (SPAD) reading and specific leaf weight. Also, leaf surface is a genetic character, which suffers slight modifications due to environmental conditions. There is an inverse correlation between stomate density and stomate surface. Likewise, a correlation between leaf water potential and leaf saturation permits calculation of the stomate closure sensitivity for each cultivar.

Expression of almond Knotted1 homologue (PdKn1) anticipates adventitious shoot initiation

The transcription factor encoded by the gene Knotted1 is a nuclear homeodomain protein, regulating meristematic cells at the shoot apical meristem. It has been demonstrated that Knotted1 (KN1) expression specifies stem cell fate in adventitious shoot induction in herbaceous plants. This gene may thus potentially identify the initiation of meristem development in adventitious shoot induction in difficult-to-regenerate plants such as Prunoideae. We isolated an almond (Prunus dulcis Mill.) KN1-type gene using degenerate primers targeting the most conserved regions of Knotted1 gene. The 5′ and 3′ ends of the isolated sequence were obtained by rapid amplification of cDNA ends-polymerase chain reaction (RACE-PCR), and the gene was named P. dulcis Knotted1-like (PdKn1). PdKn1 transcripts were detected by reverse transcription (RT)-PCR mainly in shoot apical and axillary meristems. The RT-PCR and RT-quantitative PCR PdKn1 expression in almond leaf explants was found to anticipate the organization of adventitious shoot meristems. Apricot RNA isolated from induced leaf explants cross-hybridized with the almond probe PdKn1 in Northern blotting. We suggest that almond PdKn1 may be a useful marker to study the adventitious regeneration system by revealing the potential organogenic conditions, not only in almond but also in other Prunoideae.

Seasonal Abundance of Draeculacephala minerva and Other Xylella fastidiosa Vectors in California Almond Orchards and Vineyards

Almond leaf scorch (ALS) disease is caused by the bacterium Xylella fastidiosa and transmitted by xylem-feeding insects. Reports of increased incidence of ALS-diseased trees in California prompted surveys in three almond [Prunus dulcis (Mill.) D. A. Webb]-growing regions, from June 2003 to September 2005, to determine insect vector species composition and abundance. For comparison, sampling in and near vineyards in the San Joaquin Valley, California, also was completed. Sampling in or near almond orchards collected >42,000 Cicadomorpha of which 4.8% were xylem feeders, including 1912 grass sharpshooter, Draeculacephala minerva Ball; five Xyphon fulgida Nottingham; and a single spittlebug, Philaenus spumarius L. The most abundant vector was D. minerva. Season-long sampling indicated that D. minerva was a year-round resident in and/or near almonds in the Sacramento Valley, but not in the San Joaquin Valley. Similarly, D. minerca was rare in vineyards in the San Joaquin Valley, but was abundant in irrigated pastures near vineyards. D. minerva was most frequently collected along orchard margins, and peak densities were observed in summer, the period of time when bacterial titers are reported to increase in infected trees. Screening of D. minerva for presence of X.fastidiosa found that 1.1% of insects collected near almond orchards and 4.5% of insects collected from pastures tested positive. The X. fastidiosa subspecies and genotype detected in insects collected from orchards matched those collected from ALS-diseased almond trees in the same orchard. Of the few X. fulgida and P. spumarius collected, none tested positive for X. fastidiosa. Results are discussed with respect to X. fastidiosa vector control and detection methods.

Influence of Inoculation Date on the Colonization of Xylella fastidiosa and the Persistence of Almond Leaf Scorch Disease Among Almond Cultivars.

Field grown 2-year-old almond trees (Prunus dulcis cvs. Butte, Carmel, Mission, Ne Plus Ultra, Padre, Peerless, Price, Solano, Sonora, and Thompson) were mechanically inoculated with Xylella fastidiosa in the growing seasons of 2002 and 2003 to study the effect of inoculation date on the movement and colonization of X. fastidiosa and the overwintering persistence of almond leaf scorch disease (ALS) in these cultivars. X. fastidiosa was inoculated into the base of current-season growing shoots in April, May, June, July, August, September, and October. Almond trees inoculated in spring months developed more ALS-symptomatic leaves and more extensive within-plant spread of X. fastidiosa by the end of the current growing season compared with trees inoculated in July, August, September, and October. Trees inoculated in June developed the most severe ALS symptoms during the season in which they were inoculated. Trees inoculated in June and July 2002 had significantly higher disease ratings in 2003 than inoculations made in August and October 2002. Based on disease ratings observed in 2003, 1 year after inoculation, Sonora and Solano were the most ALS susceptible, Mission and Price intermediate, and Carmel, Padre, Ne Plus Ultra, Butte, Peerless, and Thompson were the least susceptible cultivars for allowing X. fastidiosa to overwinter and cause disease the following year. Assessment of all trees in August 2004 indicated that trees inoculated in June and July 2002 had a significantly higher amount of ALS-infected branches than trees inoculated in other months. Butte, Carmel, Padre, and Thompson cultivars had no symptomatic branches, while X. fastidiosa infections persisted or colonized new branches in Sonora, Solano, Peerless, Price, Mission, and Ne Plus Ultra. Based on the 2004 assessment, Sonora was the most susceptible cultivar. Surveys of a diseased orchard in Chico, CA showed large differences in ALS incidence in four almond cultivars. Nonpareil and Peerless had significantly greater incidence of disease than Butte and Carmel over the 2 years surveyed. These data suggest that cultivar susceptibility and the time of X. fastidiosa infection are important factors in determining the persistence of ALS in almond trees.

Analysis of Xylem Fluid Components in Almond Cultivars Differing in Resistance to Almond Leaf Scorch Disease.

Almond leaf scorch (ALS) is caused by the pathogenic bacterium Xylella fastidiosa and poses a threat to the California almond industry. Almond cultivars are differentially resistant or susceptible to ALS. X. fastidiosa can infect but does not overwinter in resistant cultivars in sufficient numbers to cause symptoms or be detected by polymerase chain reaction. To better understand the biochemical or morphological factors mediating resistance, we extracted and analyzed almond xylem fluid from four almond cultivars differing in ALS susceptibility, including Butte and Carmel cultivars that are field resistant and Peerless and Sonora that are ALS susceptible. Xylem fluid was collected over winter months in 2007 to 2009, as well as July 2008 and April 2009, and analyzed for the following: pH, osmolarity, concentrations of sugars, calcium, magnesium, organic acids, and total phenolics. For most of these analyses, we found no clear differences in xylem fluid from resistant and susceptible almond cultivars. However, during the winter months, resistant cultivars tended to have higher concentrations of total phenolic compounds compared with susceptible cultivars (P = 0.05). In February 2009, Carmel had the highest total phenolic concentration measured, 233 μg/ml of gallic acid equivalents. The lowest phenolic concentrations occurred in April 2009. The cross-sectional areas of xylem vessels in Butte (resistant) and Peerless (susceptible) trees were not significantly different between cultivars.

XYLELLA FASTIDIOSA: ITS BIOLOGY, DIAGNOSIS, CONTROL AND RISKS

XYLELLA FASTIDIOSA: ITS BIOLOGY, DIAGNOSIS, CONTROL AND RISKS

Whole Genome Sequences of Two Xylella fastidiosa Strains (M12 and M23) Causing Almond Leaf Scorch Disease in California

Xylella fastidiosa is a Gram-negative plant-pathogenic bacterium causing many economically important diseases, including almond leaf scorch disease (ALSD) in California. Genome information greatly facilitates research on this nutritionally fastidious organism. Here we report the complete genome sequences of two ALSD strains of this bacterium, M12 and M23.

Epidemiology of Diseases Caused by Xylella fastidiosa in California: Evaluation of Alfalfa as a Source of Vectors and Inocula.

Pierce's disease and almond leaf scorch disease have been chronic problems for California grape and almond growers, respectively. Both diseases are caused by the xylem-limited, bacterial pathogen Xylella fastidiosa, which is transmitted by xylem-feeding insects. We evaluated the potential for alfalfa to serve as a source of vectors and inocula in California. Analysis of Geographic Information Systems maps on the distribution and abundance of grape, almond, and alfalfa plantings determined that 94,521 ha of almond and grape were planted within 1.6 km of an alfalfa field. Seasonal trends of X. fastidiosa detection were monitored outdoors and in the greenhouse in five needle-inoculated alfalfa cultivars (CUF101, Moapa69, WL342, WL530HQ, and WL625HQ) over 2 years. Results suggest that cool winter temperatures reduced X. fastidiosa populations to undetectable levels but did not eliminate infections. Sampling of alfalfa fields to assess incidence of X. fastidiosa corroborated this result, with positive samples detected in summer only. Incidence of X. fastidiosa in alfalfa during summer was low, with only 6 positive samples out of 1,156 samples collected over 3 years. Insect trapping in alfalfa fields over 3 years found that the green sharpshooter (Draeculacephala minerva) was the most abundant vector. Within alfalfa fields, green sharpshooter abundance was highest in weedy areas, suggesting a preference for weeds over alfalfa. These results confirm that weedy alfalfa fields can serve as an important source of vectors. Incidence of X. fastidiosa in alfalfa was low, possibly due to preference of vectors for weeds over alfalfa.

Differential Susceptibility of Prunus Germplasm (Subgenus Amygdalus) to a California Isolate of Xylella fastidiosa

Seedling peach (Prunus persica Batsch) and clonal peach–almond hybrids are popular rootstock choices for commercial almond growers in California. In this study, clonal replicates of peach and almond [P. dulcis (Mill.) D.A. Webb] rootstock germplasm and a first-generation peach–almond hybrid created from them were challenged with Xylella fastidiosa isolate M23. Clonal replicates were needle-inoculated with M23 and maintained in a greenhouse environment for a growing season. Typical almond leaf scorch disease symptoms began to develop on M23-inoculated almonds 11 weeks after inoculation. No leaf scorch symptoms were observed on M23-inoculated peach or peach–almond hybrids. Quantitative real-time polymerase chain reaction revealed consistent levels of X. fastidiosa DNA among inoculated almond replicates, whereas X. fastidiosa DNA was undetectable in replicates of peach–almond hybrids. A trace level of X. fastidiosa DNA was detected in a single peach replicate, and statistical analysis demonstrated that this level differed significantly (P < 0.001) from that detected in almond replicates. Selected almonds were further sampled sequentially along their meristematic axes to examine bacterial titer throughout the trees. Selected almond trees differed significantly (P = 0.036) in bacterial titer, but no significant differences were noted in levels of X. fastidiosa from different vertical sections of the main growth axes. The results suggest that peach and peach–almond hybrid rootstock germplasm used by commercial almond tree nurseries in California are not primary inoculum sources for X. fastidiosa-induced diseases.

Leaf Scorch of Purple-Leafed Plum and Sweetgum Dieback: Two New Diseases in Southern California Caused by Xylella fastidiosa Strains with Different Host Ranges.

Sweetgum dieback and leaf scorch of purple-leafed plum are two new diseases of southern California landscape ornamentals. Samplings were conducted in 2003 and 2004 and 28 of 105 sweetgum (Liquidambar styraciflua) and 38 of 62 purple-leafed plum (Prunus cerasifera) plants tested positive for Xylella fastidiosa by enzyme linked immunosorbent assay. In all, 3 strains of X. fastidiosa were isolated from sweetgum and 13 from purple-leafed plum. All sweetgum strains and some purple-leafed plum strains grew on PW but not PD3 media. Strain PC045 from purple-leafed plum and strain LS022 from sweetgum were inoculated into their original hosts in addition to almond, oleander, and grapevine plants. Sweetgum plants also were inoculated with strains causing Pierce's disease, almond leaf scorch, and oleander leaf scorch. Strain PC045 caused symptoms in purple-leafed plum and almond plants within 6 months, and the pathogen was recovered from 93 and 100% of inoculated plants, respectively. Inoculation of grapevine and oleander plants with PC045 did not result in disease or recovery of the pathogen. In all, 5 of 25 sweetgum plants inoculated with LS022 showed symptoms after 9 months, and the pathogen was recovered from 3 of these plants. Inoculation of grapevine, oleander, and almond with LS022 resulted in no disease or recovery of the pathogen from the plants. A strain of Pierce's disease, a strain of oleander leaf scorch, and two strains from almond did not cause disease in sweetgum. These results confirm the role of X. fastidiosa strains as pathogens of purple-leafed plum and sweetgum, and that strains from sweetgum are unique in their host range.

Winter curing of Prunus dulcis cv ‘Butte,’ P. webbii and their interspecific hybrid in response to Xylella fastidiosa infections

Clonal replicates of Prunus dulcis cv ‘Butte,’ P. webbii and their interspecific hybrid P 63-61 were inoculated with Xylella fastidiosa strain M23 and evaluated for almond leaf scorch disease and subsequent winter curing of infections during three growing seasons. Initial inoculations established greater than 90% infection in each of the accessions, based on PCR diagnoses from petiole tissues sampled near the inoculation site. Classic leaf scorch symptoms were evident in each population during the first growing season in a controlled greenhouse environment. Trees were removed from the greenhouse during the winters to accumulate chill hours and to provide the possibility of winter curing X. fastidiosa infections. Both PCR diagnostics and in vitro cultivation were used during the second and third growing seasons to determine the persistence of X. fastidiosa in clones among the three populations. Tree survival and the degree of winter cured infections differed among the three populations, with P. webbii and P 63-61 demonstrating enhanced levels of survivorship over ‘Butte.’ After two cycles of ambient winter temperatures and subsequent growth, ‘Butte’ averaged 21.2% winter cured trees with 73.1% mean survival. Tree survival and winter cured infections were nearly 100% for both P. webbii and P 63-61, demonstrating the utility of P. webbii in almond breeding efforts aimed at reducing tree vulnerability to X. fastidiosa infections.

Seasonal Behavior of Xylella fastidiosa Causing Almond Leafscorch Disease under Field Conditions and Improved Detection of the Bacteria by Means of Array-PCR

Almond leaf scorch disease (ALSD) caused by Xylella fastidiosa is potentially a serious threat to the almond industry in San Joaquin Valley of California. Knowledge of X. fastidiosa behaviour in the plant host under field conditions is important for disease control and this issue is being addressed in this project. Occurrence of ALSD is strongly influenced by environmental factors. In 2006, the earliest leaf scorching symptoms were observed in June, whereas in 2007, the earliest occurrence of leaf scorching symptoms was in July, a delay of 1 month. In both years, PCR detected X. fastidiosa 1 month before of symptom expression. PCR was slightly more sensitive than cultivation method for early bacterial detection. However, uneven bacterial distribution and random sampling errors may have contributed to the differences among the assays. Correlation between cultivation and PCR detection was greater than 90%. During the processing of a large number of samples, we noticed occasional failures in PCR amplifications of some samples, interfering result interpretation. We developed an array-PCR protocol using primers from seven housekeeping genes to correct the deficiency.

Temporal Distribution of Egg Hatch for TwoHomalodiscaSpp. (Hemiptera: Cicadellidae) Under Constant Temperatures

The glassy-winged sharpshooter (GWSS), Homalodisca vitripennis (Germar) (=H. coagulata (Say)) (Takiya et al. 2006), is an exotic pest in California and an important vector of the xylem-limited bacteria, Xylella fastidiosa Wells et al., the causal agent of several plant diseases including Pierce's disease of grapes, phony peach disease, almond leaf scorch, alfalfa dwarf, and oleander leaf scorch (Blua et al. 1999; UCOP 2000; Varela et al. 2001). The closely related smoke-tree sharpshooter (STSS), H. liturata Ball (=H. lacerta (Fowler)) (Burks & Redak 2003), is native to California and also a vector of Pierce's disease and oleander leaf scorch (Freitag et al. 1952; Purcell et al. 1999). Eggs of both Homalodisca spp. are laid just below the epidermis of leaves and other plant parts including stems and fruits in clusters, with eggs oriented nearly parallel to one another. Al-Wahaibi & Morse (2003, 2009) described the relationship between the rate of embryonic development of if. vitripennis and temperature in the range of 16.7-35.0°C. We present here a follow-up study aimed to elucidate the distribution of emergence of first instars of if. vitripennis and if. liturata across time, and to determine whether different constant egg incubation temperatures would modify that distribution. In order to satisfy these objectives, 2 experiments were conducted. The first examined the daily distribution of egg hatch of H. vitripennis and H. liturata at 8 constant temperatures. The second evaluated the diurnal distribution of hatching of H. vitripennis at 3 constant temperatures. For the first experiment, egg masses were collected by caging field-collected adults ofH. vitripennis on potted, rooted cuttings of chrysanthemum, Dendranthema grandiflorum (Kitam) (White Diamond cultivar, Growlink Co., Ventura, CA), in sleeve cages which were held at 23°C, 5070% RH, and 14:10 L:D photoperiod inside an insectary room. Separate plants were exposed to colonies of the 2 leafhopper species for 24 h. Plants with egg masses of the 2 species were then incubated at the same time in situ (inside leaves intact on plant) in growth chambers (Percival Scientific, Inc., Perry, LA) at 13.0, 16.7, 19.7, 25.6, 31.2, 32.9, 33.4, and 35.0°C (mean temperatures based on HOBO data loggers, Onset Computer Co., Bourne, MA). For all temperature treatments, relative humidity varied between 50-70%, and the light regime was set at 14:10 L:D photoperiod. When hatching was imminent (presence of large dark eye-spots), leaves containing egg masses were excised and placed inside 100-mm diameter Petri dishes on top of moist tissue paper. This allowed easier and more accurate observation of emergence of nymphs from individual eggs. Egg masses inside Petri dishes were retained at the same incubation temperatures as the egg masses were exposed to prior to excision of leaves. Thereafter, egg hatch was monitored twice daily, in the morning (8-10 AM) and afternoon (4-6 PM). The proportion of eggs hatching during the first day of hatch and during the following 6 d (i.e., 7 d in total) was calculated (out of the total number of hatched eggs per egg mass). Each egg mass was used as a replicate. Following this, the proportion of eggs hatching during each of d 1, 2, 3, and 4-7 were compared among temperature treatments by ANOVA. Means shown to be significantly different by ANOVA were further compared to each other and separated with the Tukey-Kramer HSD procedu e. In addition, the proportion of eggs hatching (pooled across temperature treatments) during ach of dl, 2, 3, and 4-7 were compared between the 2 Homalodisca spp. by £-tests. All statis ical tests were performed with JMP IN (SAS Institute 1996). Most hatching occurred during the first day (79%). The percentage of eggs hatching during the second day averaged 19%, with 2% on the third day. Combined egg hatch from d 4 to 7 eggs averaged 0.2%. There was no significant difference beween the 2 Homalodisca species for the proportion of hatched eggs on each of d 1, 2, 3, or 4-7 (all £-tests, P > 0.05). Temperature had a slightly significant effect on first day hatching (ANOVA, P = 0.0247) (Fig. 1). Generally, greater first day hatch was observed at higher temperatures, with hatching at 33.4°C being significantly higher than at 19.7°C. Temperature had a marginally significant effect on the proportion of second day-egg hatch (P = 0.0502), with slightly higher hatching at 16.7 and 19.7°C than at the higher temperatures. Third day hatching was significantly higher at 13.0°C than at all other temperatures (P = 0.0003). There was no significant effect of temperature on egg hatch over d 4-7 (P = 0.1553).