Population genetic analyses of Phytophthora cinnamomi reveals three lineages and movement between natural vegetation and avocado orchards in South Africa

Phytophthora cinnamomi is the causal agent of root rot, canker and dieback of thousands of plant species around the globe. This oomycete not only causes severe economic losses to forestry and agricultural industries, but also threatens the health of various plants in natural ecosystems. In this study, 380 isolates of P. cinnamomi from four avocado production areas and two regions of natural vegetation in South Africa were investigated using 15 microsatellite markers. These populations were found to have a low level of genetic diversity and consisted of isolates from three lineages. Shared genotypes were detected between isolates from avocado orchards and natural vegetation, indicating the movement of isolates between these areas. The population from the Western Cape natural vegetation had the highest genotypic diversity and unique alleles, indicating this could be the point of introduction of P. cinnamomi to South Africa. Index of association analysis suggested that five out of six populations were under linkage disequilibrium suggesting a clonal mode of reproduction whereas genotypes sampled from a recently established avocado orchard in the Western Cape were derived from a randomly recombined population. This study provided novel insights on the genetic diversity and spread of P. cinnamomi in South Africa. It also reported on the predominance of triploidy in natural occurring populations and provided evidence for recombination of P. cinnamomi for the first time. The presence of two dominant genotypes in all avocado production areas in South Africa highlight the importance of considering them in disease management and resistance breeding programmes.

Characterization and fungicides sensitivity of Phytophthora cinnamomi isolates causing avocado root rot in Zitácuaro, Michoacán

<em>Phytophthora cinnamomi</em> is the pathogen most frequently associated with avocado root rot. In Zitácuaro, Michoacán, production has increased by 19.8%; however, there are no studies of root rot in this area. The objective of the study was to characterize the isolates obtained from avocado roots and assess the sensitivity to fungicides. Samples from 5 avocado orchards were collected, sampling 5 trees per orchard (a total of 25 samples). The samples isolated were characterized morphological and molecularly. Mating type was analyzed using reference isolates of<em> P. cinnamomi</em> A1 (isolate from camelia) and A2 (isolate from avocado). To confirm the pathogenicity, tests were performed on avocado fruits with the isolates. The sensitivity of 15 isolates to potassium phosphite and to metalaxyl-M at different concentrations was evaluated<em> in vitro</em>. In a subgroup of six isolates, it was evaluated whether there was a relationship between growth rate and potassium phosphite sensitivity. Fifteen isolates were obtained with coenocytic coraloid mycelium, chlamydospores, sporangia without papilla, ovoid to ellipsoid, with internal proliferation, heterothallic with mating type A2, with amphigynous antheridia and plerotic oospores, characteristics consistent with <em>P. cinnamomi</em>. The inoculated isolates were pathogenic on avocado fruits. The isolates were more sensitive to potassium phosphite than to metalaxyl-M, with mean EC50 values of 24.62 and 0.215 ?g mL-1 of i.a., respectively. No relationship was observed between growth rate and potassium phosphite sensitivity. It is necessary to obtain a greater number of<em> P. cinnamomi</em> isolates for virulence studies.

Novel MicroRNAs and Their Functional Targets from Phytophthora infestans and Phytophthora cinnamomi

Background: Even though miRNAs play viral roles in developmental biology by regulating the translation of mRNAs, they are poorly studied in oomycetes, especially in plant pathogen Phytophthora. Objective: The study was aimed to predict and identify the putative miRNAs and their targets in Phytophthora infestans and Phytophthora cinnamomi. Methods: Homology based comparative method was used to identify the unique miRNA sequences in P. infestans and P. cinnamomi with 148,689 EST and TSA sequences of these species. Secondary structure prediction of sRNAs for the 76 resultant sequences has been performed with MFOLD tool and their targets were predicted using psRNAtarget. Result: Novel miRNAs, miR-8210 and miR-4968 were predicted from P. infestans and P. cinnamomi, respectively along with their structural features. The newly identified miRNAs were identified to play important roles in gene regulation, with few of their target genes predicted as transcription factors, tumor suppressor genes, stress responsive genes, DNA repairing genes etc. Conclusion: The miRNAs and their targets identified have opened new interference and editing targets for the development of Phytophthora resistant crop varieties.

Interactive effects of biotic stressors and provenance on chemical defence induction by holm oak (Quercus ilex)

Key Message The patterns of induced chemical defences in Quercus ilex leaves are specific to the biotic stress factor that causes them. Interactive effects between stressors depend on provenance. Abstract Quercus forests are suffering serious decline worldwide, closely linked to the consequences of climate change. The increase of biotic stressors threatens the survival of the holm oak (Quercus ilex), a dominant tree species in the Mediterranean Basin. A better understanding of its resistance mechanisms is urgently required to enable a better control of its decline. In this work, the ability of holm oaks from six Iberian provenances to respond to multiple biotic damage is studied through an analysis of their induced chemical defence patterns. Using 2016 seedlings established in a common garden trial (6 regions × 12 families/region × 7 seedlings/family × 4 treatments), biotic damage was induced at the root level (by infection with the widespread pathogen Phytophthora cinnamomi) and at the above-ground level (by mechanical defoliation). The levels of constitutive and induced total phenols, total tannins and condensed tannins were measured. Results showed that (1) the defensive chemical patterns present significant local and geographical variation, (2) survival to stress is more related to constitutive defences than induced ones, (3) the induced response is stressor-specific, and (4) there is an interactive effect amongst stressors whose sign (induction/inhibition) depends on the provenance. These findings on biotic stressor effects on the chemical defences and survival of holm oak can contribute to the development of genetic material selection programs in the integrated control of the widespread decline of Quercus.

Micropropagation, Characterization, and Conservation of Phytophthora cinnamomi-Tolerant Holm Oak Mature Trees

Holm oak populations have deteriorated drastically due to oak decline syndrome. The first objective of the present study was to investigate the use of axillary budding and somatic embryogenesis (SE) to propagate asymptomatic holm oak genotypes identified in disease hotspots in Spain. Axillary budding was achieved in two out of six tolerant genotypes from the south-western region and in two out of four genotypes from the Mediterranean region. Rooting of shoots cultured on medium supplemented with 3 mg L−1 of indole-3-acetic acid plus 0.1 mg L−1 α-naphthalene acetic acid was achieved, with rates ranging from 8 to 36%. Shoot cultures remained viable after cold storage for 9–12 months; this procedure is therefore suitable for medium-term conservation of holm oak germplasm. SE was induced in two out of the three genotypes tested, by using nodes and shoot tips cultured in medium without plant growth regulators. In vitro cloned progenies of the tolerant genotypes PL-T2 and VA5 inhibited growth of Phytophthora cinnamomi mycelia when exposed to the oomycete in vitro. Significant differences in total phenol contents and in the expression profiles of genes regulating phenylpropanoid biosynthesis were observed between in vitro cultured shoots derived from tolerant trees and cultures established from control genotypes.

Detecting Phytophthora cinnamomi associated with dieback disease on Carya cathayensis using loop-mediated isothermal amplification

Chinese hickory (Carya cathayensis Sarg.) is an economically and ecologically important nut plant in China. Dieback and basal stem necrosis have been observed in the plants since 2016, and its recent spread has significantly affected plant growth and nut production. Therefore, a survey was conducted to evaluate the disease incidence at five sites in Linan County, China. The highest incidence was recorded at the Tuankou site at up to 11.39% in 2019. The oomycete, Phytophthora cinnamomi, was isolated from symptomatic plant tissue and plantation soil using baiting and selective media-based detection methods and identified. Artificial infection with the representative P. cinnamomi ST402 isolate produced vertically elongated discolorations in the outer xylem and necrotic symptoms in C. cathayensis seedlings in a greenhouse trial. Molecular detections based on loop-mediated isothermal amplification (LAMP) specific to P. cinnamomi ST402 were conducted. Result indicated that LAMP detection showed a high coherence level with the baiting assays for P. cinnamomi detection in the field. This study provides the evidence of existence of high-pathogenic P. cinnamomi in the C. cathayensis plantation soil in China and the insights into a convenient tool developed for conducting field monitoring of this aggressive pathogen.

Soil Biodiversity and Root Pathogens in Agroecosystems

Soil ecosystem is a living and dynamic environment, habitat of thousands of microbial species, animal organisms and plant roots, integrated all of them in the food webs, and performing vital functions like organic matter decomposition and nutrient cycling; soil is also where plant roots productivity represent the main and first trophic level (producers), the beginning of the soil food web and of thousands of biological interactions. Agroecosystems are modified ecosystems by man in which plant, animal and microorganisms biodiversity has been altered, and sometimes decreased to a minimum number of species. Plant diseases, including root diseases caused by soil-borne plant pathogens are important threats to crop yield and they causes relevant economic losses. Soil-borne plant pathogens and the diseases they produce can cause huge losses and even social and environmental changes, for instance the Irish famine caused by Phytophthora infestans (1845–1853), or the harmful ecological alterations in the jarrah forests of Western Australia affected by Phytophthora cinnamomi in the last 100 years. How can a root pathogen species increase its populations densities at epidemic levels? In wild ecosystems usually we expect the soil biodiversity (microbiome, nematodes, mycorrhiza, protozoa, worms, etc.) through the trophic webs and different interactions between soil species, are going to regulate each other and the pathogens populations, avoiding disease outbreaks. In agroecosystems where plant diseases and epidemics are frequent and destructive, soil-borne plant pathogens has been managed applying different strategies: chemical, cultural, biological agents and others; however so far, there is not enough knowledge about how important is soil biodiversity, mainly microbiome diversity and soil food webs structure and function in the management of root pathogens, in root and plant health, in healthy food production, and maybe more relevant in the conservation of soil as a natural resource and derived from it, the ecosystem services important for life in our planet.

Detecting Phytophthora cinnamomi associated with dieback disease on Carya cathayensis using loop-mediated isothermal amplification

Chinese hickory (Carya cathayensis Sarg.) is an economically and ecologically important nut plant in China. Dieback and basal stem necrosis have been observed in the plants since 2016, and its recent spread has significantly affected plant growth and nut production. Therefore, a survey was conducted to evaluate the disease incidence at five sites in Linan County, China. The highest incidence was recorded at the Tuankou site at up to 11.39% in 2019. The oomycete, Phytophthora cinnamomi, was isolated from symptomatic plant tissue and plantation soil using baiting and selective media-based detection methods and identified. Artificial infection with the representative P. cinnamomi ST402 isolate produced vertically elongated discolorations in the outer xylem and necrotic symptoms in C. cathayensis seedlings in a greenhouse trial. Molecular detections based on loop-mediated isothermal amplification (LAMP) specific to P. cinnamomi ST402 were conducted. Result indicated that LAMP detection showed a high coherence level with the baiting assays for P. cinnamomi detection in the field. This study provides the evidence of existence of high-pathogenic P. cinnamomi in the C. cathayensis plantation soil in China and the insights into a convenient tool developed for conducting field monitoring of this aggressive pathogen.

Synthesis of N-Methylmorpholinium Derivatives Possessing a 1,3,4-Oxadiazole Core as Feasible Antibacterial Agents against Plant Bacterial Diseases

To develop a kind of quaternary ammonium compounds that can safely apply in agriculture for managing the plant bacterial diseases, herein, a series of N-methylmorpholinium derivatives possessing a classical 1,3,4-oxadiazole core were prepared and the antibacterial activities both in vitro and in vivo were screened. Bioassay results revealed that compounds 3l and 3i showed the strongest antibacterial activity toward pathogens Xanthomonas oryzae pv. oryzae and X. axonopodis pv. citri with the lowest EC50 values of 1.40 and 0.90 μg/mL, respectively. Phytotoxicity test trials indicated that target compounds bearing a bulky N-methylmorpholinium pendant are safe for plants. The following in vivo bioassays showed that compound 3l could control the rice bacterial blight disease, thereby affording good control efficiencies of 55.95% (curative activity) and 53.09% (protective activity) at the dose of 200 μg/mL. Preliminary antibacterial mechanism studies suggested that target compounds had strong interactions with the cell membrane of bacteria via scanning electron microscopy imaging. Additionally, this kind of framework also displayed certain antifungal activity toward Fusarium oxysporum and Phytophthora cinnamomi. Given the above privileged characteristics, this kind of 1,3,4-oxadiazole-tailored N-methylmorpholinium derivatives could stimulate the design of safe quaternary ammonium bactericides for controlling plant bacterial diseases.