A. K. Miles

The distribution and spread of citrus canker in Emerald, Australia

Citrus canker is a disease of citrus and closely related species, caused by the bacterium Xanthomonas citri subsp. citri. This disease, previously exotic to Australia, was detected on a single farm [infested premise-1, (IP1). IP is the terminology used in official biosecurity protocols to describe a locality at which an exotic plant pest has been confirmed or is presumed to exist. IP are numbered sequentially as they are detected] in Emerald, Queensland in July 2004. During the following 10 months the disease was subsequently detected on two other farms (IP2 and IP3) within the same area and studies indicated the disease first occurred on IP1 and spread to IP2 and IP3. The oldest, naturally infected plant tissue observed on any of these farms indicated the disease was present on IP1 for several months before detection and established on IP2 and IP3 during the second quarter (i.e. autumn) 2004. Transect studies on some IP1 blocks showed disease incidences ranged between 52 and 100% (trees infected). This contrasted to very low disease incidence, less than 4% of trees within a block, on IP2 and IP3. The mechanisms proposed for disease spread within blocks include weather-assisted dispersal of the bacterium (e.g. wind-driven rain) and movement of contaminated farm equipment, in particular by pivot irrigator towers via mechanical damage in combination with abundant water. Spread between blocks on IP2 was attributed to movement of contaminated farm equipment and/or people. Epidemiology results suggest: (i) successive surveillance rounds increase the likelihood of disease detection; (ii) surveillance sensitivity is affected by tree size; and (iii) individual destruction zones (for the purpose of eradication) could be determined using disease incidence and severity data rather than a predefined set area.

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.

Infection, colonisation and sporulation by Pseudocercospora macadamiae on macadamia fruit

The structures and manner with which Pseudocercospora macadamiae penetrates, colonises and proliferates from the pericarp of macadamia fruit was studied using scanning electron microscopy and fluorescence light microscopy. Germ tubes arising from conidia penetrated open stomata within 20 h of inoculation, without observation of specialised infection structures such as appressoria. Colonisation of the pericarp was intercellular, without observation of specialised intracellular infection structures such as haustoria, and was complete from the epidermis to the mesocarp. The fungus proliferated at the epidermis by the formation of conidiophores and conidia on substomatal and protuberant subepidermal stromata. These structures were not observed on the mesocarp surface. The onset of visual husk spot symptoms coincided with an increase in pathogen biomass on the pericarp surface. The progression of symptoms from tan-coloured spots to larger red-brown lesions coincided with the production of conidiophores from substomatal and protuberant subepidermal stromata. The darker the colour of the husk spot lesion, the more frequently protuberant subepidermal stromata were observed. These findings are discussed in the context of observation of other cercosporoid fungi.

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

Husk spot, caused by Pseudocercospora macadamiae is amajor 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.

Timing of infection of macadamia fruit by Pseudocercospora macadamiae and climatic effects on growth and spore germination.

Pseudocercospora macadamiae causes husk spot of macadamia. Husk spot control would be improved by verifying the stages in fruit development susceptible to infection, and determine some of the climatic conditions likely to lead to high disease pressure periods in the field. Our results showed that the percent conidia germination and growth of germ tubes and mycelia of P. macadamiae were greatest at 26°C, with better conidia germination associated with high relative humidity and free water. The exposure of match-head-sized and pea-sized fruit stages to natural P. macadamiae inoculum in the field led to 2–5-fold increases in husk spot incidence, and up to 8.5-fold increases in premature abscission, compared with unexposed fruit. Exposure of fruit stages later than match-head-sized and pea-sized fruit generally caused no further increases in disease incidence or premature abscission. Climatic conditions were found to have a strong influence on the behaviour of P. macadamiae, the host, oil accumulation, and the subsequent impact of husk spot on premature abscission. Our findings suggest that fungicide application should target fruit at the match-head-sized stage of development in order to best reduce yield losses, particularly in seasons where oil accumulation in fruit is prolonged and climatic conditions are optimal for P. macadamiae.

Phyllosticta spp. on cultivated Citrus in Australia

The occurrence of pathogenic and endophytic species of Phyllosticta on cultivated Citrus in Australia was investigated by DNA sequence analysis of specimens held in plant pathology herbaria and culture collections. Sequences of the internal transcribed spacer region (ITS1, 5.8S, ITS2), and partial translation elongation factor 1-alpha (TEF) gene of 41 Phyllosticta-like isolates from Citrus were compared to those sequences from the type specimens of Phyllosticta recorded from around the world. Phylogenetic analysis resolved all the sequences of Australian accessions into two major clades. One clade corresponded to P. citricarpa, which causes citrus black spot disease. The other clade contained P. capitalensis, which is a known endophyte of Citrus and many other plant species. All included herbarium accessions previously designated as Guignardia mangiferae are now designated P. capitalensis. No Australian isolates were identified as the newly described pathogens of citrus P. citriasiana or P. citrichinaensis, or the endophytes Guignarida mangiferae, P. brazilianiae, or P. citribraziliensis.

A Global Perspective on the Population Structure and Reproductive System of Phyllosticta citricarpa

The citrus pathogen Phyllosticta citricarpa was first described 117 years ago in Australia; subsequently, from the summer rainfall citrus-growing regions in China, Africa, and South America; and, recently, the United States. Limited information is available on the pathogens population structure, mode of reproduction, and introduction pathways, which were investigated by genotyping 383 isolates representing 12 populations from South Africa, the United States, Australia, China, and Brazil. Populations were genotyped using seven published and eight newly developed polymorphic simple-sequence repeat markers. The Chinese and Australian populations had the highest genetic diversities, whereas populations from Brazil, the United States, and South Africa exhibited characteristics of founder populations. The U.S. population was clonal. Based on principal coordinate and minimum spanning network analyses, the Chinese populations were distinct from the other populations. Population differentiation and clustering analyses revealed high connectivity and possibly linked introduction pathways between South Africa, Australia, and Brazil. With the exception of the clonal U.S. populations that only contained one mating type, all the other populations contained both mating types in a ratio that did not deviate significantly from 1:1. Although most populations exhibited sexual reproduction, linkage disequilibrium analyses indicated that asexual reproduction is important in the pathogens life cycle.

Sexual Reproduction in the Citrus Black Spot Pathogen, Phyllosticta citricarpa

Citrus black spot (Phyllosticta citricarpa) causes fruit blemishes and premature fruit drop, resulting in significant economic losses in citrus growing areas with summer rainfall across the globe. The mating type locus of P. citricarpa has recently been characterized, revealing the heterothallic nature of this pathogen. However, insight into the occurrence of mating and the impact of completing the sexual cycle of P. citricarpa was lacking. To investigate the occurrence and impact of sexual reproduction, we developed a method to reliably, and for the first time, produce ascospores of P. citricarpa on culture media. To demonstrate meiosis during the mating process, we identified recombinant genotypes through multilocus genotyping of single ascospores. Because the process of fertilization was not well understood, we experimentally determined that fertilization of P. citricarpa occurs via spermatization. Our results demonstrate that P. citricarpa is heterothallic and requires isolates of different MAT idiomorphs to be in direct physical contact, or for spermatia to fulfill their role as male elements to fertilize the receptive organs, in order to initiate the mating process. The impact of mating on the epidemiology of citrus black spot in the field is discussed.

Novel Pathotypes of Elsinoë australis Associated with Citrus australasica and Simmondsia chinensis in Australia

Molecular phylogenetic analysis, morphology and pathogenicity to citrus fruit were used to study two isolates of Elsinoë australis associated with scab-like symptoms on a fruit of Citrus australasica (finger lime) and Simmondsia chinensis (jojoba) in Australia. In addition to being associated with finger lime, the isolate from finger lime could cause scab symptoms on C. × aurantium cv. Murcott tangor in pathogenicity tests, but could not cause scab symptoms on the other orange, mandarin, lemon or grapefruit tested. Pathogenicity tests also support previous studies showing the isolate from jojoba could not produce symptoms on fruit of C. natsudaidai. Based on the findings of this study, two novel pathotypes of E. australis are designated from Australia; namely the Finger Lime (FL) pathotype associated with finger lime, and the Jojoba Black Scab (JBS) pathotype associated with black scab of jojoba. The significance of these novel E. australis pathotypes on market access and biosecurity issues for citrus are briefly discussed.

Does Fungicide Run-off From Citrus Delay Leaf Litter Decomposition?

Leaf litter is a major inoculum source for citrus diseases such citrus black spot caused by Phyllosticta citricarpa, and greasy spot caused by Mycosphaerella citri. In order to reduce this inoculum source, the efficacy of urea, dolomitic lime, a commercial compost accelerator, and an organic mulch, was assessed for enhanced leaf decomposition and reduction in sporocarps. However, due to the potential for run-off from high volume fungicide applications to disrupt leaf decomposition and microbial antagonism, the amendments were compared with and without simulated fungicide run-off. Mature green leaves of Citrus sinensis were removed from trees and placed inside mesh bags before being pinned to the orchard floor. The amendments were applied, and then simulated run-off from a typical citrus black spot fungicide program (copper, mancozeb, azoxystrobin) was applied. Leaf degradation was assessed every 2-3 weeks by visual ratings and dry weight. No direct effects on sporocarps could be observed due to insufficient infection. The results showed that the organic mulch was the most effective at enhancing decomposition, while there was significantly (P < 0.05) less decomposition in the presence of fungicide run-off.