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Hühnerhirse (Echinochloa crus-galli) gewinnt als Ungras im Maisanbau zunehmend an Bedeutung. Einseitiger Einsatz von ALS-Inhibitoren erzeugt einen Selektionsdruck, der zu vermehrtem Auftreten von Resistenzen führt. Häufig handelt es sich um Target-Site-Resistenzen (TSR) mit Punktmutationen an den Positionen Ala-122, Pro- 197 oder Trp-574 des ALS-Gens. Dieses liegt in der hexaploiden Hühnerhirse in drei homologen Varianten vor. Gängige PCR-Verfahren ermöglichen nur eine gemeinsame Analyse aller Genvarianten, wodurch ein großer Teil des Informationsgehalts verloren geht. Für eine fundierte Aussage müssen die Genvarianten getrennt analysiert werden. Ein methodischer Ansatz war die allel-spezifische PCR. Hierfür wurden DNA-Polymorphismen zwischen den drei ALS-Genvarianten genutzt, um diese separat bei Position 574 hochspezifisch zu amplifizieren und zu sequenzieren. Die schwierige Struktur und geringe Polymorphismen verhinderten den Einsatz der allelspezifischen PCR an den Positionen 122 und 197. Hier wurden die drei Genvarianten gemeinsam amplifiziert und anschließend in einem mehrstufigen Ansatz mit Restriktionsenzymen spezifisch verdaut. Nach Separation, Aufreinigung und Reamplifikation der Fragmente wurden die einzelnen Genvarianten durch Pyrosequenzierung analysiert. So konnten an Position Ala-122 in allen ALS-Homologen heterozygote Mutationen nachgewiesen werden, bei Pro-197 und Trp-574 dagegen heterozygote bzw. homozygote Mutationen nur bei ALS3. Durch die Etablierung dieser Methoden können nun Resistenzmuster in Populationen von Echinochloa crus-galli detaillierter untersucht und wichtige Daten zu deren Ausbreitungsdynamik gewonnen werden.

BACKGROUND: Brome grass (Bromus diandrus Roth) is prevalent in the southern and western cropping regions of Australia, where it causes significant economic damage. A targeted herbicide resistance survey was conducted in 2020 by collecting brome grass populations from 40 farms in Western Australia and subjecting these samples to comprehensive herbicide screening. One sample (population 172‐20), from a field that had received 12 applications of clethodim over 20 years of continuous cropping, was found to be highly resistant to the acetyl‐CoA carboxylase (ACCase)‐inhibiting herbicides clethodim and quizalofop, and so the molecular basis of resistance was investigated. RESULTS: All 31 individuals examined from population 172‐20 carried the same resistance‐endowing point mutation causing an aspartate‐to‐glycine substitution at position 2078 in the translated ACCase protein sequence. A wild‐type susceptible population and the resistant population had similar expression levels of plastidic ACCase genes. The level of resistance to quizalofop, either standalone or in mixture with clethodim, in population 172‐20 was lower under cooler growing conditions. CONCLUSION: Target‐site resistance to ACCase‐inhibiting herbicides, conferred by one ACCase mutation, was selected in all tested brome plants infesting a field with a history of repeated clethodim use. This mutation appears to have been fixed in the infesting population. Notably, clethodim resistance in this population was not detected by the farmer, and a high future incidence of quizalofop resistance is anticipated. Herbicide resistance testing is essential for the detection of evolving weed resistance issues and to inform effective management strategies. © 2023 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

The paper presents reduced order results for three different realistic scenarios with respect to the damage tolerance behaviour. The scenarios are dedicated to stiffened structures and feature the fuselage side panel, the upper fuselage panel and a lower wing panel. A wide range of parameter variations is discussed and the influence on the inspection interval is shown. Results may be used both in preliminary aircraft design and structural optimisation. [ABSTRACT FROM AUTHOR]

Downy mildews caused by obligate biotrophic oomycetes result in severe crop losses worldwide. Among these pathogens, Pseudoperonospora cubensis and P. humuli , two closely related oomycetes, adversely affect cucurbits and hop, respectively. Discordant hypotheses concerning their taxonomic relationships have been proposed based on host–pathogen interactions and specificity evidence and gene sequences of a few individuals, but population genetics evidence supporting these scenarios is missing. Furthermore, nuclear and mitochondrial regions of both pathogens have been analyzed using microsatellites and phylogenetically informative molecular markers, but extensive comparative population genetics research has not been done. Here, we genotyped 138 current and historical herbarium specimens of those two taxa using microsatellites (SSRs). Our goals were to assess genetic diversity and spatial distribution, to infer the evolutionary history of P. cubensis and P. humuli , and to visualize genome-scale organizational relationship between both pathogens. High genetic diversity, modest gene flow, and presence of population structure, particularly in P. cubensis , were observed. When tested for cross-amplification, 20 out of 27 P. cubensis -derived gSSRs cross-amplified DNA of P. humuli individuals, but few amplified DNA of downy mildew pathogens from related genera. Collectively, our analyses provided a definite argument for the hypothesis that both pathogens are distinct species, and suggested further speciation in the P. cubensis complex. [ABSTRACT FROM AUTHOR]

Abstract: Alternaria alternata is the most common fungal pathogen of tomatoes in Upper Egypt. Morphological identification of this fungus is challenging; therefore, this study searched for new classification tools based on molecular techniques. Using a dilution plating method, 67 strains of A. alternata were isolated from 34 samples of rotten tomato fruits representing the Giza 80 and Edkawy cultivars. The collected strains were identified using the amplification products of the internal transcribed spacer (ITS) region, glyceraldehyde 3‐phosphate dehydrogenase (Gpd) and Alt a1, which is a gene involved in the production of most of the allergens produced by A. alternata. The screening revealed that A. alternata constituted more than half of the total fungi recovered from rotten tomatoes in this study. According to the phylogenetic analysis using these three loci, the collected strains clustered in accordance with the host cultivar type from which they had been isolated. Specific gene random primer polymerase chain reaction (SGRP‐PCR) techniques indicated that the A. alternata population in the tested region has a high genetic diversity. The pathogenicity test showed that most of the A. alternata isolates (67.2%) were highly pathogenic, and no correlation was found between the phylogenetic analysis and pathogenicity. In addition, the influence of the fungicide Disan 80% on the collected strains showed significant differences that were attributed to the source of isolation. [ABSTRACT FROM AUTHOR]

Microthlaspi is a predominantly Eurasian genus which also occurs in the northernmost parts of Africa (Maghreb). The most widespread species of the genus is M. perfoliatum , which can be found from Sweden to Algeria and from Portugal to China. The other species are thought to have much more confined distribution ranges, often covering only a few hundred kilometres. This is also believed for the diploid M. erraticum , which was recently re-appraised as a taxon independent from the tetra- to hexaploid M. perfoliatum . Previously, M. erraticum was believed to be present only in Central Europe, from the East of France to Slovenia. In order to gain a deeper understanding of the ecology, evolution and migration history of Microthlaspi it was the focus of the current study to investigate, if M. erraticum is present in habitats outside Central Europe, but with microclimates similar to Central Europe. It is demonstrated that M. erraticum is much more widespread than previously thought, while other lineages apart from M. perfoliatum s.str. and M. erraticum seem to have restricted distribution ranges. The latter species was observed from the Alps and their foreland, the Balkans, the mountainous areas around the Black Sea, Southern Siberia, as well as the Altai and Tien Shan mountains. This demonstrates a widespread occurrence of this easily-overlooked species. [ABSTRACT FROM AUTHOR]

Pseudoperonospora cubensis, an obligate biotrophic oomycete causing devastating foliar disease in species of the Cucurbitaceae family, was never reported in seeds or transmitted by seeds. We now show that P. cubensis occurs in fruits and seeds of downy mildew-infected plants but not in fruits or seeds of healthy plants. About 6.7% of the fruits collected during 2012–2014 have developed downy mildew when homogenized and inoculated onto detached leaves and 0.9% of the seeds collected developed downy mildew when grown to the seedling stage. This is the first report showing that P. cubensis has become seed-transmitted in cucurbits. Species-specific PCR assays showed that P. cubensis occurs in ovaries, fruit seed cavity and seed embryos of cucurbits. We propose that international trade of fruits or seeds of cucurbits might be associated with the recent global change in the population structure of P. cubensis. [ABSTRACT FROM AUTHOR]

Smut fungi are well-suited to investigate the ecology and evolution of plant pathogens, as they are strictly biotrophic, yet cultivable on media. Here we report the genome sequence of Melanopsichium pennsylvanicum, closely related to Ustilago maydis and other Poaceae-infecting smuts, but parasitic to a dicot plant. To explore the evolutionary patterns resulting from host adaptation after this huge host jump, the genome of Me. pennsylvanicum was sequenced and compared with the genomes of U. maydis, Sporisorium reilianum, and U. hordei. Although all four genomes had a similar completeness in CEGMA (Core Eukaryotic Genes Mapping Approach) analysis, gene absence was highest in Me. pennsylvanicum, and most pronounced in putative secreted proteins, which are often considered as effector candidates. In contrast, the amount of private genes was similar among the species, highlighting that gene loss rather than gene gain is the hallmark of adaptation after the host jump to the dicot host. Our analyses revealed a trend of putative effectors to be next to another putative effector, but the majority of these are not in clusters and thus the focus on pathogenicity clusters might not be appropriate for all smut genomes. Positive selection studies revealed that Me. pennsylvanicum has the highest number and proportion of genes under positive selection. In general, putative effectors showed a higher proportion of positively selected genes than noneffector candidates. The 248 putative secreted effectors found in all four smut genomes might constitute a core set needed for pathogenicity, whereas those 92 that are found in all grass-parasitic smuts but have no ortholog in Me. pennsylvanicum might constitute a set of effectors important for successful colonization of grass hosts. [ABSTRACT FROM AUTHOR]

Before the advent of molecular phylogenetics, species concepts in the downy mildews, an economically important group of obligate biotrophic oomycete pathogens, have mostly been based upon host range and morphology. While molecular phylogenetic studies have confirmed a narrow host range for many downy mildew species, others, like Pseudoperonospora cubensis affect even different genera. Although often morphological differences were found for new, phylogenetically distinct species, uncertainty prevails regarding their host ranges, especially regarding related plants that have been reported as downy mildew hosts, but were not included in the phylogenetic studies. In these cases, the basis for deciding if the divergence in some morphological characters can be deemed sufficient for designation as separate species is uncertain, as observed morphological divergence could be due to different host matrices colonised. The broad host range of P. cubensis (ca. 60 host species) renders this pathogen an ideal model organism for the investigation of morphological variations in relation to the host matrix and to evaluate which characteristics are best indicators for conspecificity or distinctiveness. On the basis of twelve morphological characterisitcs and a set of twelve cucurbits from five different Cucurbitaceae tribes, including the two species, Cyclanthera pedata and Thladiantha dubia, hitherto not reported as hosts of P. cubensis, a significant influence of the host matrix on pathogen morphology was found. Given the high intraspecific variation of some characteristics, also their plasticity has to be taken into account. The implications for morphological species determination and the confidence limits of morphological characteristics are discussed. For species delimitations in Pseudoperonospora it is shown that the ratio of the height of the first ramification to the sporangiophore length, ratio of the longer to the shorter ultimate branchlet, and especially the length and width of sporangia, as well as, with some reservations, their ratio, are the most suitable characteristics for species delimitation. [ABSTRACT FROM AUTHOR]

Pseudoperonospora cubensis causes some of the most devastating diseases of cucurbitaceous crops, while P. humuli is an important pathogen of hop (Cannabaceae). Although parasitic to different Angiosperm orders, these pathogens are highly similar, both in morphology and based on molecular comparisons. Considering the close relationship of P. humuli and P. cubensis, it was hypothesized that cross infectivity of the pathogens between their optimum hosts might be possible. Two strains of P. humuli and one of P. cubensis used in this study were able to reproduce on the two wild cucurbit relatives Bryonia dioica and Sicyos angulatus. Interestingly, limited infectivity of the P. cubensis strain to hop was also observed, and the P. humuli strain was also able to infest Cucumis sativus. The cross infections were verified by comparing the nuclear ribosomal-internal transcribed spacer sequences of newly produced sporangiophores from the different hosts. Morphologically the two pathogens could be distinguished by statistical analyses on their original hosts but these differences diminished on some alternate hosts. The ability of P. cubensis to infest the perennial Bryonia dioica and Humulus lupulus might offer a limited possibility for overwintering in temperate regions and needs to be evaluated in future studies. The ability of P. humuli to complete its life cycle on all the tested cucurbits provides evidence that the host jump across orders may continue and points to the high virulence potential of this pathogen. [ABSTRACT FROM AUTHOR]

Peronospora tabacina is an obligate parasite that causes blue mold of tobacco. The pathogen reproduces primarily by sporangia, whereas the sexual oospores are rarely observed. A collection of 122 isolates of P. tabacina was genotyped using nine microsatellites to assess the population structure of individuals from subpopulations collected from central, southern, and western Europe; the Middle East; Central America; North America; and Australia. Genetic variations among the six subpopulations accounted for -8% of the total variation, including moderate levels of genetic differentiation, high gene flow among these subpopulations, and a positive correlation between geographic and genetic distance (r = 0.225; P < 0.001). Evidence of linkage disequilibrium (P < 0.001) showed that populations contained partially clonal subpopulations but that subpopulations from Australia and Mediterranean Europe did not. High genetic variation and population structure among samples could be explained by continuous gene flow across continents via infected transplant exchange and/or long-distance dispersal of sporangia via wind currents. This study analyzed the most numerous P. tabacina collection and allowed conclusions regarding the migration, mutation, and evolutionary history of this obligate biotrophic oomycete. The evidence pointed to the species origin in Australia and identified intracontinental and intercontinental migration patterns of this important pathogen. [ABSTRACT FROM AUTHOR]

Highlights • Southeastern Europe and Anatolia were the source for recolonisation of Central Europe after the most recent glacial period. • Northwestern Europe was re-colonised from refugia in southwestern France. • Human activity might have caused the postglacial migrations of Microthlaspi to southern Sweden. • Pleistocene glacial refugia were in all of the Mediterranean peninsulas, southwestern France, Transcaucasia and Central Asia. Abstract Quaternary climatic oscillations had strong effects on Palearctic biodiversity. Although Transcaucasia is hypothesised to be a centre of diversity for many taxa, relatively few studies have investigated its potential role as such, or as a Pleistocene refugium. Using AFLP and sequence data (ITS, mat K, trn L-F) from samples collected across the entire range of Microthlaspi perfoliatum , from the Iberian peninsula to Kazakhstan, and including the Mediterranean Basin and Central Europe, in this study it is examined how the interplay of Pleistocene climatic oscillations and historical demography may have shaped the current genetic structure of the species. The results of this study provide evidence that M. perfoliatum survived Pleistocene glaciation in five major refugia, Iberia, southwestern France, Italy, the Balkans and Anatolia/Transcaucasia and a sixth, unknown refugium in the disjunct distribution area in Central Asia. Our analyses support an ancient colonisation of M. perfoliatum towards the western and central Palearctic, perhaps, during the early Pleistocene or late Pliocene, likely starting from Transcaucasia. However, postglacial recolonisation of western and Central Europe has taken place from two distinct refugia, namely southwestern France/northern Italy, and Anatolia/Southern Balkans, respectively. The Iberian populations apparently did not contribute to postglacial recolonisation of Europe, perhaps because the Pyrenees acted as a significant barrier to dispersal. It is also conceivable that a rapidly expanding population from southwestern France and the southeastern part of the range effectively blocked the establishment of latecomers. The present study thus reveals a complex pattern of colonisation of Europe from Transcaucasia, with an interplay of climate and physical geography as main factors shaping the present-day genetic structure of M. perfoliatum. [ABSTRACT FROM AUTHOR]

The range-wide genetic diversity of Microthlaspi erraticum (Brassicaceae), a Eurasian calcareous annual plant, was investigated with respect to its migration and post-glacial re-colonisation of Central Europe. Both AFLP and sequence data (ITS, mat K, trn L-F) were used to analyse 85 populations of the species sampled across its range, including Europe and Anatolia as well as the disjunct populations in Central Asia. A substantial portion of the explainable genetic variation was found to be correlated with climatic heterogeneity. A low level of within-population genetic diversity throughout the range, and a clear genetic differentiation among populations is consistent with the mainly selfing nature and fragmented distribution of the species. The highest within-population genetic diversity was found in Central Europe. However, the number of rare and private fragments was substantially higher in southern Europe and Anatolia, the inferred ancestral range of M. erraticum , whereas populations found in Kazakhstan originated from Anatolia, following a profound population bottleneck. The diversity patterns found in Central Europe and migration models support postglacial re-colonisation from two regions; the south-western Alps and the Balkans. Hence, our results provide molecular evidence for a suture zone in Central Europe, where advancing genotypes from different refugia meet and mix. [ABSTRACT FROM AUTHOR]

Microthlaspi erraticum is widely distributed in temperate Eurasia, but restricted to Ca 2+ -rich habitats, predominantly on white Jurassic limestone, which is made up by calcium carbonate, with little other minerals. Thus, naturally occurring Microthlaspi erraticum individuals are confronted with a high concentration of Ca 2+ ions while Mg 2+ ion concentration is relatively low. As there is a competitive uptake between these two ions, adaptation to the soil condition can be expected. In this study, it was the aim to explore the genomic consequences of this adaptation by sequencing and analysing the genome of Microthlaspi erraticum . Its genome size is comparable with other diploid Brassicaceae, while more genes were predicted. Two Mg 2+ transporters known to be expressed in roots were duplicated and one showed a significant degree of positive selection. It is speculated that this evolved due to the pressure to take up Mg 2+ ions efficiently in the presence of an overwhelming amount of Ca 2+ ions. Future studies on plants specialized on similar soils and affinity tests of the transporters are needed to provide unequivocal evidence for this hypothesis. If verified, the transporters found in this study might be useful for breeding Brassicaceae crops for higher yield on Ca 2+ -rich and Mg 2+ -poor soils., (Copyright © 2020 Mishra, Ploch, Runge, Schmuker, Xia, Gupta, Sharma and Thines.)