Field evaluation of a plant activator, captan, chlorothalonil, copper hydroxide, iprodione, mancozeb and strobilurins for the control of citrus brown spot of mandarin

Brown spot (caused by Alternaria alternata) is a major disease of citrus in subtropical areas of Australia. A number of chemicals, the strobilurins azoxystrobin, trifloxystrobin, pyraclostrobin and methoxycrylate, a plant activator (acibenzolar), copper hydroxide, mancozeb, captan, iprodione and chlorothalonil/pyrimthanil were tested in the field for its control. Over three seasons, trees in a commercial orchard received 16, 14 and 7 fungicide sprays, respectively, commencing at flowering in the first season, and petal fall in the later seasons. In all experiments, the strobilurins used alone, or incorporated with copper and mancozeb, were as effective as, or better than the industry standard of copper and mancozeb alone. The only exception was trifloxystrobin, which when used alone was less effective than the industry standard. Acibenzolar used alone was ineffective. Applying a mixture of azoxystrobin and acibenzolar was found to reduce the incidence of brown spot compared with applying azoxystrobin alone but, in either case, disease levels were not found to be significantly different to the industry standard. Captan, iprodione and chlorothalonil/pyrimthanil were as effective as the industry standard. The incidence and severity of rind damage were significantly lowest in the azoxystrobin, methoxycrylate, iprodione and chlorothalonil/pyrimthanil treatments. Medium and high rates of trifloxystrobin (0.07 g/L, 0 .15 g/L) and pyraclostrobin (0.8 g/L, 1.2 g/L) applied alone were the only treatments found to be IPM-incompatible as shown by the elevated level of scale infection on fruit.

Timing of fungicide applications for control of husk spot caused by Pseudocercospora macadamiae in macadamia

Pseudocercospora macadamiae is an important pathogen of macadamia in Australia, causing a disease known as husk spot. Growers strive to control the disease with a number of carbendazim and copper treatments. The aim of this study was to consider the macadamia fruit developmental stage at which fungicide application is most effective against husk spot, and whether application of copper-only applications at full-size fruit developmental stage toward the end of the season contributed to effective disease control. Fungicides were applied to macadamia trees at four developmental stages in three orchards in two subsequent production seasons. The effects of the treatments on disease incidence and severity were quantified using area under disease progress curve (AUDPC) and logistic regression models. Although disease incidence varied between cultivars, incidence and severity on cv. A16 showed consistent differences between the treatments. Most significant reduction in husk spot incidence occurred when spraying commenced at match-head sized-fruit developmental stage. All treatments significantly reduced husk spot incidence and severity compared with the untreated controls, and a significant positive linear relationship (R2 = 73%) between AUDPC and severity showed that timing of the first fungicide application is important for effective disease control. Application of fungicide at full-size fruit stage reduced disease incidence but had no impact on premature fruit drop.

Alternative fungicides for controlling husk spot caused by Pseudocercospora macadamiae in macadamia

Husk spot, caused by Pseudocercospora macadamiae is a major fungal disease of macadamia in Australia. Chemicals to control the disease are limited and frequent failure to control the disease is a major concern to growers. The overall goal of this research was to improve the chemical control strategy of P. macadamiae through the provision of fungicides with different modes of action to carbendazim, which is the current industry standard. Husk spot incidence, premature fruit abscission, kernel quality and yield were evaluated following application of different fungicide products in replicated field experiments at three different sites. Results showed significant differences in disease incidence and premature fruit abscission between fungicide treatments, field sites and years. Generally, disease incidence and premature fruit abscission on trees treated with fungicide were significantly (P < 0.05) lower than the untreated control. Pyraclostrobin conferred significantly better protection than trifloxystrobin, reducing disease severity by 70% compared with a 50% reduction by trifloxystrobin. The pyraclostrobin treatment had a similar efficacy to the current industry standard (70% reduction cf. 73% reduction by tank-mixed carbendazim and copper). Higher amounts of immature kernels occurred in the untreated control, followed by difenoconazole and trifloxystrobin. Diseased fruit accounted for 78% of premature fruit abscission, which indicates that husk spot enhances fruit abscission in macadamia. Our results suggest that pyraclostrobin provided similar efficacy to the industry standard and could, therefore, play a key role in the management of husk spot.

The effects of holding space on growth and survival of individually reared three-spot crab (Portunus sanguinolentus).

The problem of cannibalism in communally reared crabs can be eliminated by separating the growing crabs into holding compartments. There is currently no information on optimal compartment size for growing crabs individually. 136 second instar crablets (Portunus sanguinolentus) (C2 ca. 7-10 mm carapace width (CW)) were grown for 90 days in 10 different-sized opaque and transparent walled acrylic compartments. The base area for each compartment ranged from small (32 mm × 32 mm) to large (176 mm × 176 mm). Effects of holding space and wall transparency on survival, CW, moult increment, intermoult period and average weekly gain (AWG) were examined. Most crabs reached instars C9-C10 (50-70 mm CW) by the end of experiment. The final survival rate in the smallest compartment was 25% mainly due to moult-related mortality predominantly occurring at the C9 instar. However, crabs in these smaller compartments had earlier produced significantly larger moult increments from instar to instar than those in the larger compartments (P < 0.05). Crabs in the smaller compartments (<65 mm × 65 mm) also showed significantly longer moult periods (P < 0.05). The net result was that AWG in CW was 5.22 mm week-1 for the largest compartment and 5.15 mm week-1 in smallest and did not differ significantly between compartment size groups (P = 0.916). Wall transparency had no impact on survival (P = 0.530) but a slight impact on AWG (P = 0.014). Survival rate was the best indicator of minimum acceptable compartment size (?43 mm × 43 mm) for C10 crablets because below this size death occurred before growth rate was significantly affected. For further growth, it would be necessary to transfer the crablets to larger compartments.

Tan spot: A new disease of pyrethrum caused by Microsphaeropsis tanaceti sp. nov

The isolation frequency of Microsphaeropsis sp. in spring in association with necrotic lesions on leaves in Tasmanian pyrethrum (Tanacetum cinerariifolium) fields has increased substantially since first identification in 2001. Examination of morphological features and sequencing of the internal transcribed spacer region (ITS) resulted in the identification of a new species, herein described as Microsphaeropsis tanaceti sp. nov. The pathogenicity of three M. tanaceti isolates to two pyrethrum cultivars was confirmed by inoculating glasshouse-grown plants in three experiments. No significant differences in the susceptibility of the two cultivars to infection by M. tanaceti were found. Symptoms were tan-coloured spots which coalesced around the margins of the leaves. Therefore, the name 'tan spot' is proposed for this new disease of pyrethrum. The sensitivity of seven M. tanaceti isolates to difenoconazole and azoxystrobin, commonly used fungicides for the management of foliar diseases in spring, was assessed under in vitro conditions. Sensitivity testing for difenoconazole was conducted using a mycelial growth assay on potato dextrose agar, whilst testing for sensitivity to azoxystrobin used a conidial germination assay on water agar. Microsphaeropsis tanaceti was found to be more sensitive to azoxystrobin than difenoconazole, with complete inhibition of conidial germination at concentrations above 0.625 µg a.i. mL-1. By comparison, concentrations of 50 µg a.i. difenoconazole mL-1 or greater were required for significant inhibition of mycelial growth. It therefore appears likely that there is currently some control of tan spot as a result of the use of azoxystrobin and to a lesser extent, difenoconazole, for the control of other diseases.

Field and laboratory evaluations of fungicides for the control of brown spot (Corynespora cassiicola) and black spot (Asperisporium caricae) of papaya in far north Queensland, Australia

Several chemicals including strobilurins (pyraclostrobin and azoxystrobin), triazoles (difenoconazole and tebuconazole), dithiocarbamates (propineb, metiram, ziram and mancozeb) and the phthalimide chlorothalonil were evaluated in three field experiments in north Queensland, Australia, for the control of brown spot (caused by Corynespora cassiicola) and black spot (caused by Asperisporium caricae) of papaya. Chlorothalonil and pyraclostrobin were shown to be more effective than the industry standard, mancozeb, for the control of brown spot. In the black spot experiments, difenoconazole, pyraclostrobin and chlorothalonil used alone or in spray programs were as effective as, or better than, the industry standards, mancozeb and tebuconazole. Plants treated with pyraclostrobin and difenoconazole had more fruit unaffected by black spot (97% and 99% respectively) than plants treated with tebuconazole (51%), mancozeb (20%) and the untreated controls (1%). Laboratory tests also showed that A. caricae was more sensitive to difenoconazole (EC50 of 2ppm) than tebuconazole (EC50 of 14ppm). In 2007, off-label permits were obtained for chlorothalonil for control of brown spot and difenoconazole and chlorothalonil for the control of black spot of papaya.

SSR analysis of cultivated groundnut (Arachis hypogaea L.) germplasm resistant to rust and late leaf spot diseases

Cultivated groundnut (Arachis hypogaea L.) is an agronomically and economically important oilseed crop grown extensively throughout the semi-arid tropics of Asia, Africa and Latin America. Rust (Puccinia arachidis) and late leaf spot (LLS, Phaseoisariopsis personata) are among the major diseases causing significant yield loss in groundnut. The development of varieties with high levels of resistance has been constrained by adaptation of disease isolates to resistance sources and incomplete resistance in resistant sources. Despite the wide range of morphological diversity observed in the cultivated groundnut gene pool, molecular marker analyses have thus far been unable to detect a parallel level of genetic diversity. However, the recent development of simple sequence repeat (SSR) markers presents new opportunities for molecular diversity analysis of cultivate groundnut. The current study was conducted to identify diverse disease resistant germplasm for the development of mapping populations and for their introduction into breeding programs. Twenty-three SSRs were screened across 22 groundnut genotypes with differing levels of resistance to rust and LLS. Overall, 135 alleles across 23 loci were observed in the 22 genotypes screened. Twelve of the 23 SSRs (52%) showed a high level of polymorphism, with PIC values ≥0.5. This is the first report detecting such high levels of genetic polymorphism in cultivated groundnut. Multi-dimensional scaling and cluster analyses revealed three well-separated groups of genotypes. Locus by locus AMOVA and Kruskal-Wallis one-way ANOVA identified candidate SSR loci that may be valuable for mapping rust and LLS resistance. The molecular diversity analysis presented here provides valuable information for groundnut breeders designing strategies for incorporating and pyramiding rust and late leaf spot resistances and for molecular biologists wishing to create recombinant inbred line populations to map these traits.

Outbreak of angular leaf spot, caused by Xanthomonas fragariae, in a Queensland strawberry germplasm collection.

Strawberry (Fragaria (x) ananassa) plants exhibiting leaf lesions consistent with angular leaf spot (ALS, caused by Xanthomonas fragariae Kennedy and King 1962) were identified in the Queensland strawberry germplasm at Bundeberg in May 2010. Water suspensions of bacterial ooze tested positive using a previously described primer set. However, the slow growth rate of X. fragariae and the presence of a fast-growing, non-pathogenic, undescribed Xanthomonas species presented problems that were overcome by dilution plating and DNA sequence analysis. Sequencing of the gyrB locus of putative colonies of X. fragariae indicated 100% sequence similarity to other X. fragariae isolates. A new set of diagnostic primers for X. fragariae based on the gyrB locus is presented.

Alternaria Fruit Spot: New Directions

This project determined the identity of causal pathogens, epidemiology and disease cycle of Alternaria leaf blotch and fruit spot in Australian apples and provided a management strategy for both diseases for inclusion in the integrated fruit production manual.

Spot Form of Net Blotch Resistance in a Diverse Set of Barley Lines in Australia and Canada.

The responses of 95 barley lines and cultivars to spot form of net blotch (SFNB) caused by Pyrenophora teres f. maculata were analyzed as seedlings and adults in Australia and Canada. Cluster analyses revealed complex reaction responses. Only 2 lines (Esperance Orge 289 and TR3189) were resistant to all isolates at the seedling stage, whereas 15 lines and cultivars (81-82/033, Arimont, BYDV-018, CBSS97M00855T-B2-M1-Y1-M2-Y-1M-0Y, C19776, Keel, Sloop, Torrens, TR326, VB0111, Yarra, VB0229, WI-2477, WI2553, and Wisconsin Pedigree) were resistant toward the two Canadian isolates and mixture of Australian isolates at the adult stages. In Australian field experiments, the effectiveness of SFNB resistance in three barley cultivars (Barque. Cowabbie, and Schooner) and one breeding line (VB9104) with a different source of resistance was tested. Barque, which possessed a resistance gene that provided complete resistance to SFNB, was the most effective and showed no effect on grain yield or quality in the presence of inoculum. Generally, cultivars with seedling or adult resistance had less disease and better grain quality than the susceptible control. Dash, but they were not as effective as Barque. A preliminary differential set of 19 barley lines and cultivars for P teres I. maculata is proposed.

Morphological and molecular diversity of Colletotrichum spp. causing pepper spot and anthracnose of lychee (Litchi chinensis) in Australia

Since the 1980s a new disease has been affecting Australian lychee. Pepper spot appears as small, black superficial lesions on fruit, leaves, petioles and pedicels and is caused by Colletotrichum gloeosporioides, the same fungus that causes postharvest anthracnose of lychee fruit. The aim of this study was to determine if a new genotype of C.gloeosporioides is responsible for the pepper spot symptom. Morphological assessments, arbitrarily-primed PCR (ap-PCR) and DNA sequencing studies did not differentiate isolates of C.gloeosporioides from anthracnose and pepper spot lesions. The ap-PCR identified 21 different genotypes of C.gloeosporioides, three of which were predominant. A specific genotype identified using ap-PCR was associated with the production of the teleomorph in culture. Analysis of sequence data of ITS and -tubulin regions of representative isolates did not group the lychee isolates into a monophyletic clade; however, given the majority of the isolates were from one of three genotypes found using ap-PCR, the possibility of a lychee specific group of C.gloeosporioides is discussed.

In planta localization and interactions of impatiens necrotic spot tospovirus proteins

Impatiens necrotic spot tospovirus (INSV) is a significant pathogen of ornamentals. The tripartite negative- and ambi-sense RNA genome encodes six proteins that are involved in cytoplasmic replication, movement, assembly, insect transmission and defence. To gain insight into the associations of these viral proteins, we determined their intracellular localization and interactions in living plant cells. Nucleotide sequences encoding the nucleoprotein N, non-structural proteins NSs and NSm, and glycoproteins Gn and Gc of a Kentucky isolate of INSV were amplified by RTPCR, cloned, sequenced and transiently expressed as fusions with autofluorescent proteins in leaf epidermal cells of Nicotiana benthamiana. All proteins accumulated at the cell periphery and co-localized with an endoplasmic reticulum marker. The Gc protein fusion also localized to the nucleus. N and NSm protein self-interactions and an NSm-N interaction were observed by using bimolecular fluorescence complementation. A tospovirus NSm homotypic interaction had not been reported previously.

Pericarps retained in the tree canopy and stomatal abundance are components of resistance to husk spot caused by Pseudocercospora macadamiae in macadamia

Pseudocercospora macadamiae Beilharz, Mayers and Pascoe infects macadamia fruit via stomata causing husk spot disease. Information on the variability of fruit stomatal abundance, its association with diseased fruit pericarps (sticktights) that are retained in the tree canopy, and its influence on the husk spot intensity (incidence, severity and lesion number) among macadamia genotypes is lacking. We examined a total of 230 macadamia trees comprising 19 cultivars, 56 wild germplasm accessions and 40 breeding progeny, for the prevalence of sticktights and husk spot intensity over three production seasons. We observed a strong association between the prevalence of sticktights and disease intensity indicating its usefulness as a predictor of husk spot and as a useful phenotypic trait for husk spot resistance selection in breeding programmes. Similarly, stomatal abundance varied among macadamia genotypes, and a significant linear relationship (P < 0.001; 93%) was observed between fruit stomatal abundance and husk spot for all the macadamia genotypes analysed, confirming the utility of that trait for disease resistance screening. The genotypes were grouped into disease resistance groups. Correlations between fruit stomatal abundance, disease intensity and prevalence of sticktights revealed that the numbers of sticktights, and relative stomatal abundance were the main factors influencing the intensity of husk spot among macadamia genotypes. This is the first comprehensive study of natural variation of stomatal abundance in Macadamia species that reveals genetic variation, and provides relevant relationships with disease intensity and the prevalence of sticktights. The phenotypic plant traits indentified in this study may serve as selection tools for disease resistance screening in macadamia breeding programmes.

Reduced fungicide use on a new Australian peanut cultivar, highly resistant to the late leaf spot and rust pathogens

Rust (caused by Puccinia arachidis) and late leaf spot (LLS, caused by Mycosphaerella berkeleyi) can cause significant yield losses in Australian peanut crops. Until recently, all commercial peanut varieties were highly susceptible to these pathogens, but the new Australian cultivar Sutherland has significantly higher levels of resistance than the older cultivars. Field trials were conducted at two sites in Queensland to (a) confirm the improved resistance of cv. Sutherland over another commercial cultivar, Menzies, (b) study the effects of timing of first spray, spray interval and cultivar on disease severity and yield, and (c) develop a suitable fungicide management program for cv. Sutherland. In the 2006 and 2007 trials, rust and LLS developed slower and had lower final disease ratings and AUDPC values on unsprayed plots of cv. Sutherland than on cv. Menzies. The timing of the first spray is critical in managing both rust and late leaf spot, with the results demonstrating that the first fungicide spray on cv. Sutherland should be applied as soon as rust and LLS are first seen on cv. Menzies. In most trials spray intervals of 14 days or 21 days were suitable to effectively control rust and LLS. In years with low disease pressure, few, if any, fungicide applications will be needed to manage the diseases, but in other years up to four sprays may be necessary. © Australasian Plant Pathology Society Inc. 2012.

Sources and seasonal dynamics of Alternaria inoculum associated with leaf blotch and fruit spot of apples

Alternaria leaf blotch and fruit spot caused by Alternaria spp. cause annual losses to the Australian apple industry. Control options are limited, mainly due to a lack of understanding of the disease cycle. Therefore, this study aimed to determine potential sources of Alternaria spp. inoculum in the orchard and examine their relative contribution throughout the production season. Leaf residue from the orchard floor, canopy leaves, twigs and buds were collected monthly from three apple orchards for two years and examined for the number of spores on their surface. In addition, the effects of climatic factors on spore production dynamics in each plant part were examined. Although all four plant parts tested contributed to the Alternaria inoculum in the orchard, significant higher numbers of spores were obtained from leaf residue than the other plant parts supporting the hypothesis that overwintering of Alternaria spp. occurred mainly in leaf residue and minimally on twigs and buds. The most significant period of spore production on leaf residue occurred from dormancy until bloom and on canopy leaves and twigs during the fruit growth stage. Temperature was the single most significant factor influencing the amount of Alternaria inoculum and rainfall and relative humidity showed strong associations with temperature influencing the spore production dynamics in Australian orchards. The practical implications of this study include the eradication of leaf residue from the orchard floor and sanitation of the canopy after harvest to remove residual spores from the trees.