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Rust fungi (Pucciniales) are a diverse group of plant pathogens in natural and agricultural systems. They pose ongoing threats to the diversity of native flora and cause annual crop yield losses. Agricultural rusts are predominantly managed with fungicides and breeding for resistance, but new control strategies are needed on non-agricultural plants and in fragile ecosystems. RNA interference (RNAi) induced by exogenous double-stranded RNA (dsRNA) has promise as a sustainable approach for managing plant-pathogenic fungi, including rust fungi. We investigated the mechanisms and impact of exogenous dsRNA on rust fungi through in vitro and whole-plant assays using two species as models, Austropuccinia psidii (the cause of myrtle rust) and Coleosporium plumeriae (the cause of frangipani rust). In vitro, dsRNA either associates externally or is internalized by urediniospores during the early stages of germination. The impact of dsRNA on rust infection architecture was examined on artificial leaf surfaces. dsRNA targeting predicted essential genes significantly reduced germination and inhibited development of infection structures, namely appressoria and penetration pegs. Exogenous dsRNA sprayed onto 1-year-old trees significantly reduced myrtle rust symptoms. Furthermore, we used comparative genomics to assess the wide-scale amenability of dsRNA to control rust fungi. We sequenced genomes of six species of rust fungi, including three new families (Araucariomyceaceae, Phragmidiaceae, and Skierkaceae) and identified key genes of the RNAi pathway across 15 species in eight families of Pucciniales. Together, these findings indicate that dsRNA targeting essential genes has potential for broad-use management of rust fungi across natural and agricultural systems.

Austropuccinia psidii, commonly known as myrtle rust, is an obligate, biotrophic rust pathogen that causes rust disease on a broad host range of Myrtaceae species. Eucalyptus grandis, a widely cultivated hardwood Myrtaceae species, is susceptible to A. psidii infection, with this pathogen threatening both their natural range and various forest plantations across the world. This study aimed to investigate the A. psidii transcriptomic responses in resistant and susceptible E. grandis at four time points. RNA-seq reads were mapped to the A. psidii reference genome to quantify expressed genes at 12-hours post inoculation (hpi), 1-, 2- and 5-days post inoculation (dpi). A total of eight hundred and ninety expressed genes were found, of which forty-three were candidate effector proteins. These included a rust transferred protein (RTP1) gene, expressed in susceptible hosts at 5-dpi and a hydrolase protein gene expressed in both resistant and susceptible hosts over time. Functional categorisation of expressed genes revealed processes enriched in susceptible hosts, including malate metabolic and malate dehydrogenase activity, implicating oxalic acid in disease susceptibility. These results highlight putative virulence or pathogenicity mechanisms employed by A. psidii to cause disease and provides the first insight into the molecular responses of A. psidii in E. grandis over time.

Brown root rot disease (BRRD), caused by Phellinus noxius, is an important tree disease in tropical/subtropical areas. To improve chemical control of BRRD and deter emergence of fungicide resistance in P. noxius, this study investigated control efficacies and systemic activities of fungicides with different modes of action. Fourteen fungicides with 11 different modes of action were tested for inhibitory effects in vitro on 39 P. noxius isolates from Taiwan, Hong Kong, Malaysia, Australia, and Pacific Islands. Cyproconazole, epoxiconazole, and tebuconazole (FRAC 3, target-site G1) inhibited colony growth of P. noxius by 99.9 to 100% at 10 ppm and 97.7 to 99.8% at 1 ppm. The other effective fungicide was cyprodinil + fludioxonil (FRAC 9 + 12, target-site D1 + E2), which showed growth inhibition of 96.9% at 10 ppm and 88.6% at 1 ppm. Acropetal translocation of six selected fungicides was evaluated in bishop wood (Bischofia javanica) seedlings by immersion of the root tips in 100 ppm of each fungicide, followed by liquid or gas chromatography tandem-mass spectrometry analyses of consecutive segments of root, stem, and leaf tissues at 7- and 21-days post-treatment. Bi-directional translocation of the fungicides was also evaluated by stem injection of fungicide solutions. Cyproconazole and tebuconazole were the most readily absorbed by roots and efficiently transported acropetally. Greenhouse experiments suggested that cyproconazole, tebuconazole, and epoxiconazole have a slightly higher potential for controlling BRRD than mepronil, prochloraz, and cyprodinil + fludioxonil. Because all tested fungicides lacked basipetal translocation, soil drenching should be considered instead of trunk injection for their use in BRRD control.

Knowledge of breeding systems and genetic diversity is critical to select and combine desired traits that advance new cultivars in agriculture and horticulture. Mushrooms that produce psilocybin, magic mushrooms, may potentially be used in therapeutic and wellness industries, and stand to benefit from genetic improvement. We studied haploid siblings of Psilocybe subaeruginosa to resolve the genetics behind mating compatibility and advance knowledge of breeding. Our results show that mating in P. subaeruginosa is tetrapolar, with compatibility controlled at a homeodomain locus with one copy each of HD1 and HD2, and a pheromone/receptor locus with four homologs of the receptor gene STE3. An additional two pheromone/receptor loci homologous to STE3 do not appear to regulate mating compatibility. Alleles in the psilocybin gene cluster did not vary among the five siblings and were likely homozygous in the parent. Psilocybe subaeruginosa and its relatives have three copies of PsiH genes but their impact on production of psilocybin and its analogues is unknown. Genetic improvement in Psilocybe will require access to genetic diversity from the centre of origin of different species, identification of genes behind traits, and strategies to avoid inbreeding depression.

Austropuccinia psidii is the causal agent of myrtle rust, a fungal disease that infects over 480 species in the Myrtaceae. A. psidii is a biotrophic pathogen that reproduces sexually and asexually. Sexual reproduction has been previously shown on Syzygium jambos and little is known about its reproductive biology on other hosts or whether populations that were formerly structured by host range can outcross on universally susceptible hosts. We investigated if mating genes in three genomes of A. psidii were under selection as a proxy for whether different strains can reproduce sexually on a shared host. We examined three homologs of the STE3.2 gene, sequences of which were near-identical in the three genomes, and the homeodomain locus, which contained two alleles of two homeodomain genes in each genome. A. psidii likely uses tetrapolar mating. Pheromone/receptor loci were distal to homeodomain loci, and based on haplotypes of a phased assembly, mate compatibility is regulated by multiallelic HD genes and biallelic STE3.2 genes; the third homolog of STE3.2 (STE3.2-1) was present in both haplotypes, and our study supports hypotheses this gene does not regulate mate recognition. Populations of A. psidii formerly structured by host range could potentially outcross on universal hosts based on their related mating genes, however this hypothesis should remain theoretical given the implications for biosecurity. Additionally, we searched for core meiotic genes in genomes of A. psidii, four species of Puccinia, and Sphaerophragmium acaciae through comparative genomics based on 136 meiosis-related orthologous genes modeled from Mycosarcoma maydis. Meiotic genes are conserved in rust fungi at family rank. We analyzed the expression of two meiotic and four mitotic genes of A. psidii on E. grandis over a 28-day time course to validate that identified meiotic genes were upregulated in teliospores. Three mitotic genes were significantly downregulated in samples collected 28 days after

Austropuccinia psidii is a fungal plant pathogen that infects species within the Myrtaceae, causing the disease myrtle rust. Myrtle rust is causing declines in populations within natural and managed ecosystems and is expected to result in species extinctions. Despite this, variation in response to A. psidii exist within some species, from complete susceptibility to resistance that prevents or limits infection by the pathogen. Untargeted metabolomics using Ultra Performance Liquid Chromatography with Ion Mobility followed by analysis using MetaboAnalyst 3.0, was used to ex-plore the chemical defence profiles of resistant, hypersensitive and susceptible phenotypes within Melaleuca quinquenervia during the early stages of A. psidii infection. We were able to identify three separate pools of secondary metabolites: (i) metabolites classified structurally as flavonoids that were naturally higher in the leaves of resistant individuals prior to infection, (ii) organoheterocyclic and carbohydrate‐related metabolites that varied with the level of host resistance post‐infection, and (iii) metabolites from the terpenoid pathways that were responsive to disease progression re-gardless of resistance phenotype suggesting that these play a minimal role in disease resistance during the early stages of colonization of this species. Based on the classes of these secondary me-tabolites, our results provide an improved understanding of key pathways that could be linked more generally to rust resistance with particular application within Melaleuca. © 2022 by the authors. Licensee MDPI, Basel, Switzerland.

Sexual reproduction, mutation, and reassortment of nuclei increase genotypic diversity in rust fungi. Sexual reproduction is inherent to rust fungi, coupled with their coevolved plant hosts in native pathosystems. Rust fungi are hypothesised to exchange nuclei by somatic hybridisation with an outcome of increased genotypic diversity, independent of sexual reproduction. We provide criteria to demonstrate whether somatic exchange has occurred, including knowledge of parental haplotypes and rejection of fertilisation in normal rust life cycles.

Brown root rot disease (BRRD), caused by Phellinus noxius, is an important tree disease in tropical/subtropical areas. To improve chemical control of BRRD and deter emergence of fungicide resistance in P. noxius, this study investigated control efficacies and systemic activities of fungicides with different modes of action. Fourteen fungicides with 11 different modes of action were tested for inhibitory effects in vitro on 39 P. noxius isolates from Taiwan, Hong Kong, Malaysia, Australia, and Pacific Islands. Cyproconazole, epoxiconazole, and tebuconazole (FRAC 3, target-site G1) inhibited colony growth of P. noxius by 99.9 to 100% at 10 ppm and 97.7 to 99.8% at 1 ppm. The other effective fungicide was cyprodinil + fludioxonil (FRAC 9 + 12, target-site D1 + E2), which showed growth inhibition of 96.9% at 10 ppm and 88.6% at 1 ppm. Acropetal translocation of six selected fungicides was evaluated in bishop wood (Bischofia javanica) seedlings by immersion of the root tips in 100 ppm of each fungicide, followed by liquid or gas chromatography tandem-mass spectrometry analyses of consecutive segments of root, stem, and leaf tissues at 7- and 21-days post-treatment. Bi-directional translocation of the fungicides was also evaluated by stem injection of fungicide solutions. Cyproconazole and tebuconazole were the most readily absorbed by roots and efficiently transported acropetally. Greenhouse experiments suggested that cyproconazole, tebuconazole, and epoxiconazole have a slightly higher potential for controlling BRRD than mepronil, prochloraz, and cyprodinil + fludioxonil. Because all tested fungicides lacked basipetal translocation, soil drenching should be considered instead of trunk injection for their use in BRRD control.

Exogenous RNA interference (RNAi) is an effective non-genetically modified (GM) control strategy for plant viruses, insect pests, fungal and oomycete pathogens. It involves the exogenous application of doublestranded RNA (dsRNA) to plants to trigger the silencing of targeted pest/pathogen genes. We have been exploring exogenous RNAi as a sustainable control for rust fungi, one the largest groups of plant pathogens, across natural and agricultural systems. A particular focus has been on Austropuccinia psidii, the causal agent of myrtle rust, an aggressive disease of Myrtaceae plants. A. psidii arrived in Australia in 2010 and poses a serious threat to native plant biodiversity. The pathogen has a large host range of >350 species and serious species decline is already underway. Myrtle rust is not only destroying natural ecosystems but is also impacting native plant industries. We synthesised dsRNA molecules targeting rust fungi genes and tested their fungicidal effects in vitro and in planta. We found that urediniospores can take up dsRNA during the early stages of germination and that dsRNA targeting essential rust genes significantly reduced germination and development of infection structures such as appressoria and infection pegs. Exogenous RNAi was also effective in planta, significantly reducing myrtle rust symptoms on one-year-old trees. Our comparative genomics study showed that genes in the RNAi pathway are conserved across eight families of rust fungi, indicating that exogenous RNAi has potential for broad-scale management of rust fungi.

Eucalyptus grandis, in its native Australian range, varies in resistance to Austropuccinia psidii (syn. Puccinia psidii). The biotrophic rust fungus, A. psidii is the causal agent of myrtle rust and poses a serious threat to Australian biodiversity. The pathogen produces yellow pustules of urediniospores on young leaves and shoots, resulting in shoot tip dieback, stunted growth, and death. Dissecting the underlying mechanisms of resistance against this pathogen will contribute to improved breeding and control strategies to mitigate its devastating effects. The aim of this study was to determine the molecular dialogue between E. grandis and A. psidii, using an RNA-sequencing approach. Resistant and susceptible E. grandis seedlings grown from seed collected across its natural range were inoculated with the pandemic biotype of A. psidii. The leaf tissue was harvested at 12-h post inoculation (hpi), 1-day post inoculation (dpi), 2-dpi and 5-dpi and subjected to RNA-sequencing using Illumina 50 bp PE reads to a depth of 40 million reads per sample. Differential gene expression and gene ontology enrichment indicated that the resistant seedlings showed controlled, coordinated responses with a hypersensitive response, while the susceptible seedlings showed no systemic response against myrtle rust. Brassinosteroid signaling was apparent as an enriched term in the resistant interaction at 2-dpi, suggesting an important role of this phytohormone in defense against the pathogen. Brassinosteroid mediated signaling genes were also among the candidate genes within two major disease resistance loci (Puccinia psidii resistance), Ppr3 and Ppr5. While brassinosteroids have been tagged as positive regulators in other plant disease resistance interactions, this is the first report in the Eucalyptus – Austropuccinia psidii interaction. Furthermore, several putative resistance genes, underlying known resistance loci and implicated in the interaction have been identified and highlighted fo

Austropuccinia psidii, cause of myrtle rust, has spread globally where Myrtaceae occur. Multiple strains of A. psidii have been identified, including a unique strain found only in South Africa. The South African strain is a biosecurity concern for species of Myrtaceae worldwide. This is because preliminary testing of South African Myrtaceae suggests it could have a wide host range and thus, has the potential to be invasive. In this study, we assessed the ability of the South African strain to infect other species of Myrtaceae by testing the susceptibility of New Zealand provenance Myrtaceae. Seedlings of four native New Zealand Myrtaceae species (Metrosideros excelsa, Leptospermum scoparium, Kunzea robusta, and Kunzea linearis) were artificially inoculated in South Africa with a single-uredinium isolate of the South African strain. Fourteen days after inoculation, uredinia, and in many cases telia, had developed on the young leaves and stems of all four host species, which led to shoot tip dieback in the more severe cases. When comparisons were made between the species, K. robusta was the least susceptible to the South African strain of A. psidii, while L. scoparium and M. excelsa were the most susceptible. While only a limited number of seed families were tested, only a small proportion of the seedlings showed resistance to infection by the South African strain. This preliminary testing highlights the potential invasive risk the South African strain poses to global Myrtaceae communities, including New Zealand Myrtaceae.

Uromycladium acaciae has damaged plantations of Acacia mearnsii in southern Africa since 2013. Uredinia of a species of Uromycladium have been known on A. mearnsii in South Africa since the 1980s. However, the recent damage is associated with telia and spermogonia. Uredinia and telia were previously treated as conspecific with a phylogenetic species concept. However, uredinia did not form after previous artificial inoculation experiments with teliospores. Controlled studies identified the optimum conditions for basidiospore infection, but the optimum conditions for sporulation and dispersal have not been identified. To investigate the life cycle and field epidemiology of Uromycladium on A. mearnsii, spores were trapped weekly and development of disease symptoms and plant phenology were monitored monthly at three plantations. Telia and spermogonia developed independently from uredinia, and nucleotide polymorphisms between rDNA of uredinia and telia were fixed based on high throughput sequencing and PCRs. All three weather variables measured had a significant effect on teliospore abundance at two of the three sites. Teliospore abundance was greatest during trapping periods when mean relative humidity was high, mean rainfall was 4–5 mm day-1, and mean temperature was 15–16°C. Teliospore counts peaked at the end of summer, potentially the result of epidemic build-up. Results support the hypothesis that despite sharing a most recent common ancestor, uredinia on A. mearnsii in southern Africa are independent to the life cycle of the telial rust, which likely constitutes a new introduction. Furthermore, teliospores of U. acaciae disperse under wet conditions; and, the wet season between October and March is the optimal period for wattle rust development. This article is protected by copyright. All rights reserved.

Resistance to the pandemic strain of Austropuccinia psidii was identified in New Zealand provenance Leptospermum scoparium, Kunzea robusta, and K. linearis plants. Only 1 Metrosideros excelsa-resistant plant was found (of the 570 tested) and no resistant plants of either Lophomyrtus bullata or L. obcordata were found. Three types of resistance were identified in Leptospermum scoparium. The first two, a putative immune response and a hypersensitive response, are leaf resistance mechanisms found in other myrtaceous species while on the lateral and main stems a putative immune stem resistance was also observed. Both leaf and stem infection were found on K. robusta and K. linearis plants as well as branch tip dieback that developed on almost 50% of the plants. L. scoparium, K. robusta, and K. linearis are the first myrtaceous species where consistent infection of stems has been observed in artificial inoculation trials. This new finding and the first observation of significant branch tip dieback of plants of the two Kunzea spp. resulted in the development of two new myrtle rust disease severity assessment scales. Significant seed family and provenance effects were found in L. scoparium, K. robusta, and K. linearis: some families produced significantly more plants with leaf, stem, and (in Kunzea spp.) branch tip dieback resistance, and provenances provided different percentages of resistant families and plants. The distribution of the disease symptoms on plants from the same seed family, and between plants from different seed families, suggested that the leaf, stem, and branch tip dieback resistances were the result of independent disease resistance mechanisms.

Phellinus noxius, the cause of brown root-rot disease, is an invasive fungal pathogen that causes a white rot among woody plants in Asia, Oceania, and Africa. Because the origin and diversity of this pathogen are unknown, it is difficult to predict its behavior and invasive capacity, especially under future climate-change scenarios. We characterized genetic relationships and ecological differences among P. noxius lineages across eastern Asia and Oceania to better understand evolutionary responses of the pathogen to environmental changes. Sequences of four loci (nuclear large subunit, internal transcribed spacers, partial RNA polymerase II, and partial translation elongation factor – 1 alpha) from 95 P. noxius isolates were used for genetic analyses. Our analyses revealed three genetically distinct lineages of P. noxius: 1) eastern Asia (Japan, Taiwan, Hong Kong, and Malaysia); 2) western Oceania/Japan/Taiwan (including Australia, Palau, Guam, Saipan, Yap, Pohnpei, and Kosrae) with some isolates from Japan and Taiwan; and 3) a distinct group from American Samoa. Population genetic analyses highlighted admixture and migration among the three lineages. Climate-based, species distribution models were used to predict ecological patterns of P. noxius for two genetic lineages: eastern Asia and western Oceania/Japan/Taiwan. Contemporary bioclimatic models depicted potential areas at high risk for P. noxius invasion, and predicted that suitable climate space (potential distribution) is lineage specific. Trade of important economic crops worldwide coupled with changing climates could exacerbate the spread of P. noxius into new geographic areas with suitable habitats for brown root-rot disease.

Myrtle rust (Austropuccinia psidii) is an invasive species causing damage to Myrtaceae species in natural and managed ecosystems in many countries. To better understand myrtle rust epidemiology we studied latent period (LP) and ontogenic resistance in relation to temperature on three susceptible hosts (Metrosideros excelsa, Lophomyrtus bullata × L. obcordata and Syzygium jambos). The latent period curve was U-shaped, with latent development >0 from between 8 and 10 °C, depending on the host, to 32 °C. Optimum range was 22–28 °C with minimum LP of 5–7 days. Peak spore production occurred over about 2 weeks, starting about 1 week after the LP ended. Some spore production continued for 1–2 months. Comparison of the LP data with field temperatures indicated that the uredinial stage of A. psidii can overwinter in the latent phase in temperate areas of New Zealand and southern Australia and, therefore, uredinial or telial reinfection is not required during winter. The LP information was used to correct the LP function in a New Zealand myrtle rust climatic risk model. The transition of emergent leaf and stem tissues in susceptible Myrtaceae genotypes from susceptible to immune (ontogenic resistance) was characterized in terms of uredinium density and LP. Onset of ontogenic resistance was closely linked to the degree of leaf expansion, with fully expanded leaves being immune to infection. Because ontogenic resistance restricts infection to periods when growth flushes occur, understanding it is crucial for explaining the seasonality of myrtle rust development in the natural environment.

Two economically, environmentally and culturally important Australian native tree species, Araucaria cunninghamii (hoop pine) and A. bidwilli (bunya pine) are rapidly declining in the Bunya Mountains national park. Some of the affected bunya pines are estimated to be more than 600 years old. Symptoms include yellowing of the crown and rapid dieback, which is consistent with infection by Phytophthora, a fungal-like oomycete. Aerial surveys have shown that symptomatic trees of both species are spread throughout the national park. Phytophthora multivora, a pathogen associated with dieback of Eucalyptus gomphocephala and Pinus radiata in Western Australia, was isolated from the rhizosphere of symptomatic bunya pines. Phytophthora multivora has also been associated together with P. cinnamomi in the dieback of wollemi pine (Wollemia nobilis) in New South Wales, and kauri pine (Agathis australis) in New Zealand. Both pathogens have wide host ranges that include agriculturally and environmentally important plant species. DNA meta-barcoding analysis of the soil detected both pathogens at high numbers. Pathogenicity testing of the organisms is currently being conducted on bunya and hoop pine to confirm aggressiveness. Additional surveys and testing need to be conducted to confirm how wide spread the problem is, and to implement control measures to prevent future damage to the national park and surrounding softwood plantations. From initial observations of the symptoms, spread patterns and environmental history of the affected trees, P. multivora is the most likely causal agent of the dieback.

RNA interference (RNAi)-inducing sprays are a new non-toxic, non-transgenic, environmentally friendly strategy that has the immediate potential to revolutionise crop protection against insect-transmitted plant viruses, and facilitate a transition away from the use of chemical insecticides in agriculture. The approach, which involves spraying plants with virus-specific dsRNA, triggers and augments systemic RNAi, the plant’s natural defence mechanism against viruses. When applied in combination with stabilising nanoclay, in the formulation known as BioClay, dsRNA can provide virus protection to plants for more than 20 days, making a single spray a commercially feasible and lasting approach to protect crops from viruses. The aim of the present work is to develop BioClay-based RNA sprays to protect avocados, pineapples and Australian Myrtaceae species from pre- and post-harvest fungal diseases. This will lead to clean green safe produce from pre- to post-harvest, from field to supermarket trolley, safeguarding Queensland horticulture and ecosystems

Uromycladium acaciae is the cause of a severe wattle rust epidemic in plantations of Acacia mearnsii (black wattle) in southern Africa. Research on the biology of this damaging rust is assisting in the development of control strategies. One strategy under investigation is the identification and deployment of resistant lines of A. mearnsii. Selection of resistant families currently relies on large-scale, time-consuming and expensive field trials. In this study, we present a detailed artificial inoculation protocol for U. acaciae, which can be used to screen for resistance. The results of an experiment that used the protocol to screen the relative resistance of 12 families of A. mearnsii to U. acaciae are also presented. The developed artificial inoculation protocol can also be used to investigate several other aspects of this host–pathogen system. © 2019, © 2019 NISC (Pty) Ltd.

Three resistance phenotypes to the pandemic biotype of the invasive fungal biotrophic pathogen Austropuccinia psidii were identified in artificially inoculated New Zealand provenance Leptospermum scoparium (mānuka) plants grown from seed collected from mother trees growing in different locations within New Zealand. The first two resistance phenotypes, putatively a constitutive response and a hypersensitive response, were leaf resistance phenotypes. On the lateral and main stems a putative constitutive stem resistance was also observed in the trials. Mānuka is the first myrtaceous species where consistent infection of stems in trials was observed, resulting in the development of a new disease severity assessment scale. No individual plants demonstrated both leaf resistances: c. 18% of plants showed both stem resistance and one of the leaf resistances; however c. 26% plants showed either a leaf or a stem resistance but not both, and the remainder of the plants tested (56%) were susceptible, with some plants showing extensive symptom expression. Plant and seed family analysis revealed limited genetic linkage between the putative constitutive leaf and stem resistances suggesting two independent resistance mechanisms. The locale from where the seed were collected was important, as the proportion of leaf and/or stem resistance plants grown from the independent seed families varied by seed provenance.

The rust fungus Austropuccinia psidii has spread globally and naturalized in areas with naïve species of Myrtaceae. Previous studies have revealed multiple strains of A. psidii within South America and two strains outside of its native range. The rust spreads by windborne mitotic urediniospores, which are the dominant spore stage. Teliospores and basidiospores of A. psidii are also formed; however, the biological role of these stages in the life cycle is unknown. Experiments presented here tested whether basidiospores of A. psidii could infect Syzygium jambos. The sori produced by infection with basidiospores were screened with five microsatellite markers to confirm whether they were a product of recombination. The findings showed that basidiospores of A. psidii could cause infection on species of Myrtaceae and the resulting sori were a product of recombination. This has important implications for programmes that breed for resistance to this aggressive pathogen in commercial eucalypt forestry. © 2017 British Society for Plant Pathology

It is challenging to sequence and assemble genomes of obligate plant pathogens and microorganisms because of limited amounts of DNA, comparatively large genomes and high numbers of repeat regions. We sequenced the 1.2 gigabase genome of an obligate rust fungus, Austropuccinia psidii, the cause of rust on Myrtaceae, with a Chromium 10X library. This technology has mostly been applied for single-cell sequencing in immunological studies of mammals. We compared scaffolds of a genome assembled from the Chromium library with one assembled from combined paired-end and mate-pair libraries, sequenced with Illumina HiSeq. Chromium 10X provided a superior assembly, in terms of number of scaffolds, N50 and number of genes recovered. It required less DNA than other methods and was sequenced and assembled at a lower cost. Chromium sequencing could provide a solution to sequence and assemble genomes of obligate plant pathogens where the amount of available DNA is a limiting factor. © 2018 Future Science. All rights reserved.

The complete genome sequence of Cucumber mosaic virus strain K was determined by deep RNA sequencing. The tripartite genome consists of a 3,382-nucleotide (nt) RNA1, a 3,050-nt RNA2, and a 2,218-nt RNA3 segment. Phylogenetic analysis placed RNA1 and RNA2 in subgroup IB. However, RNA3 grouped with subgroup IA isolates, indicating a likely recombination event. © 2018 Moyle et al.

RNA-sequencing technology has been widely adopted to investigate host responses during infection with pathogens. Dual RNA-sequencing (RNA-seq) allows the simultaneous capture of pathogen specific transcripts during infection, providing a more complete view of the interaction. In this review, we focus on the design of dual RNA-seq experiments and the application of downstream data analysis to gain biological insight into both sides of the interaction. Recent literature in this area demonstrates the power of the dual RNA-seq approach and shows that it is not limited to model systems where genomic resources are available. A reduction in sequencing cost and single cell transcriptomics coupled with protein and metabolite level dual approaches are set to enhance our understanding of plant-pathogen interactions. Sequencing costs continue to decrease and single cell transcriptomics is becoming more feasible. In combination with proteomics and metabolomics studies, these technological advances are likely to contribute to our understanding of the temporal and spatial aspects of dynamic plant-pathogen interactions.

Damage caused by Phytophthora cinnamomi Rands remains an important concern on forest tree species. The pathogen causes root and collar rot, stem cankers, and dieback of various economically important Eucalyptus spp. In South Africa, susceptible cold tolerant Eucalyptus plantations have been affected by various Phytophthora spp. with P. cinnamomi considered one of the most virulent. The molecular basis of this compatible interaction is poorly understood. In this study, susceptible Eucalyptus nitens plants were stem inoculated with P. cinnamomi and tissue was harvested five days post inoculation. Dual RNA-sequencing, a technique which allows the concurrent detection of both pathogen and host transcripts during infection, was performed. Approximately 1% of the reads mapped to the draft genome of P. cinnamomi while 78% of the reads mapped to the Eucalyptus grandis genome. The highest expressed P. cinnamomi gene in planta was a putative crinkler effector (CRN1). Phylogenetic analysis indicated the high similarity of this P. cinnamomi CRN1 to that of Phytophthora infestans. Some CRN effectors are known to target host nuclei to suppress defense. In the host, over 1400 genes were significantly differentially expressed in comparison to mock inoculated trees, including suites of pathogenesis related (PR) genes. In particular, a PR-9 peroxidase gene with a high similarity to a Carica papaya PR-9 ortholog previously shown to be suppressed upon infection by Phytophthora palmivora was down-regulated two-fold. This PR-9 gene may represent a cross-species effector target during P. cinnamomi infection. This study identified pathogenicity factors, potential manipulation targets, and attempted host defense mechanisms activated by E. nitens that contributed to the susceptible outcome of the interaction.

Field trials evaluating several parameters of growth, fruit yield and quality of 'Hass' avocado grafted to different rootstocks were established in 2004-2005 in four different growing regions of Australia. Fruit were harvested in three seasons from 2008, ripened and assessed for severity and incidence of anthracnose and stem end rot diseases. Peel samples were collected at harvest and analysed for concentrations of the cations (N, K, Ca, Mg). Rootstock significantly affected marketability of fruit (no stem end rot and less than 5% anthracnose) in 58% of the total number of trials evaluated, with better quality fruit harvested from 'Hass' grafted to Guatemalan or West Indian rootstocks such as 'A10' or 'Velvick'. Fruit quality was frequently poor from trees grafted to Mexican race rootstocks, regardless of growing location. Correlation analyses showed that fruit from rootstocks with superior fruit quality was often associated with lower skin N and higher Ca concentrations. There were significant positive correlations between anthracnose and skin N or N:Ca ratio in 75% of trials evaluated. There was a significant negative correlation between anthracnose and Ca in 42% of trials. The correlations between stem end rot and skin N (positive) or Ca (negative) were each significant in 42% of trials. Based on the results in this project, N:Ca ratios in the skin of unripe avocado fruit at harvest may provide one of the best indicators of potential postharvest disease in ripe fruit, and may have implications for fertiliser regimes.