Tracking outbreak populations of the pepper weevil Anthonomus eugenii (Coleoptera; Curculionidae) using complete mitochondrial genomes

The pepper weevil, Anthonomus eugenii, is a major pest on Capsicum species. Apart from natural spread, there is a risk of spread via international pepper trade. In the Netherlands, a pepper weevil outbreak occurred in 2012 and affected six greenhouses producing different sweet pepper varieties. The following year, a pepper weevil outbreak occurred in Italy. To trace the origin of the Dutch outbreak and to establish if the Dutch and Italian outbreaks were linked, we determined the mitogenomes of A. eugenii specimens collected at outbreak locations, and compared these with specimens from the native area, and other areas where the pest was introduced either by natural dispersal or via trade. The circular 17,257 bp A. eugenii mitogenome comprises thirteen mitochondrial genes typically found in insect species. Intra-species variation of these mitochondrial genes revealed four main mitochondrial lineages encompassing 41 haplotypes. The highest diversity was observed for specimens from its presumed native area (i.e. Mexico). The Dutch outbreak specimens represented three highly similar haplotypes, suggesting a single introduction of the pest. The major Dutch haplotype was also found in two specimens from New Jersey. As the Netherlands does not have pepper trade with New Jersey, it is likely that the specimens sampled in New Jersey and those sampled in the Netherlands originate from a shared source that was not included in this study. In addition, our analysis shows that the Italian and Dutch outbreaks were not linked. The mitochondrial genome is a useful tool to trace outbreak populations and the methodology presented in this paper could prove valuable for other invasive pest species, such as the African fruit moth Thaumatotibia leucotreta and emerald ash borer Agrilus planipennis.</p

Analysis of Thaumatotibia leucotreta (Lepidoptera: Tortricidae: Olethreutinae) mitochondrial genomes in the context of a recent host range expansion

Abstract Background The false codling moth (FCM), Thaumatotibia leucotreta (Meyrick, 1913), is a significant pest of various important economic crops and is a EU quarantine pest. In the last decade the pest has been reported on Rosa spp. In this study we determined whether this shift occurred within specific FCM populations across seven eastern sub-Saharan countries or whether the species opportunistically switches to this novel host as it presents itself. To achieve this, we assessed the genetic diversity of complete mitogenomes of T. leucotreta specimens intercepted at import and analysed potential linkages with the geographical origin and host species. Results Genomic, geographical and host information were integrated into a T. leucotreta Nextstrain build which contains 95 complete mitogenomes generated from material intercepted at import between January 2013 and December 2018. Samples represented seven sub-Saharan countries and mitogenomic sequences grouped in six main clades. Discussion If host strains of FCM would exist, specialization from a single haplotype towards the novel host is expected. Instead, we find specimens intercepted on Rosa spp. in all six clades. The absence of linkage between genotype and host suggests opportunistic expansion to the new host plant. This underlines risks of introducing new plant species to an area as the effect of pests already present on the new plant might be unpredictable with current knowledge

Synchytrium microbalum RefSeq Genome

Synchytrium microbalum genome reference projec

The linear mitochondrial genome of the quarantine chytrid Synchytrium endobioticum; insights into the evolution and recent history of an obligate biotrophic plant pathogen

Background: Chytridiomycota species (chytrids) belong to a basal lineage in the fungal kingdom. Inhabiting terrestrial and aquatic environments, most are free-living saprophytes but several species cause important diseases: e.g. Batrachochytrium dendrobatidis, responsible for worldwide amphibian decline; and Synchytrium endobioticum, causing potato wart disease. S. endobioticum has an obligate biotrophic lifestyle and isolates can be further characterized as pathotypes based on their virulence on a differential set of potato cultivars. Quarantine measures have been implemented globally to control the disease and prevent its spread. We used a comparative approach using chytrid mitogenomes to determine taxonomical relationships and to gain insights into the evolution and recent history of introductions of this plant pathogen. Results: We assembled and annotated the complete mitochondrial genome of 30 S. endobioticum isolates and generated mitochondrial genomes for five additional chytrid species. The mitochondrial genome of S. endobioticum is linear with terminal inverted repeats which was validated by tailing and PCR amplifying the telomeric ends. Surprisingly, no conservation in organisation and orientation of mitochondrial genes was observed among the Chytridiomycota except for S. endobioticum and its sister species Synchytrium microbalum. However, the mitochondrial genome of S. microbalum is circular and comprises only a third of the 72.9 Kbp found for S. endobioticum suggesting recent linearization and expansion. Four mitochondrial lineages were identified in the S. endobioticum mitochondrial genomes. Several pathotypes occur in different lineages, suggesting that these have emerged independently. In addition, variations for polymorphic sites in the mitochondrial genome of individual isolates were observed demonstrating that S. endobioticum isolates represent a community of different genotypes. Such communities were shown to be complex and stable over time, but we also demonstrate that the use of semi-resistant potato cultivars triggers a rapid shift in the mitochondrial haplotype associated with increased virulence. Conclusions: Mitochondrial genomic variation shows that S. endobioticum has been introduced into Europe multiple times, that several pathotypes emerged multiple times, and that isolates represent communities of different genotypes. Our study represents the most comprehensive dataset of chytrid mitogenomes, which provides new insights into the extraordinary dynamics and evolution of mitochondrial genomes involving linearization, expansion and reshuffling

Spizellomyces sp. 'palustris' RefSeq Genome

Spizellomyces sp. 'palustris' genome reference projec

Chytriomyces confervae RefSeq Genome

Chytriomyces confervae genome reference projec

Comparative genomics of chytrid fungi reveal insights into the obligate biotrophic and pathogenic lifestyle of Synchytrium endobioticum

Synchytrium endobioticum is an obligate biotrophic soilborne Chytridiomycota (chytrid) species that causes potato wart disease, and represents the most basal lineage among the fungal plant pathogens. We have chosen a functional genomics approach exploiting knowledge acquired from other fungal taxa and compared this to several saprobic and pathogenic chytrid species. Observations linked to obligate biotrophy, genome plasticity and pathogenicity are reported. Essential purine pathway genes were found uniquely absent in S. endobioticum, suggesting that it relies on scavenging guanine from its host for survival. The small gene-dense and intron-rich chytrid genomes were not protected for genome duplications by repeat-induced point mutation. Both pathogenic chytrids Batrachochytrium dendrobatidis and S. endobioticum contained the largest amounts of repeats, and we identified S. endobioticum specific candidate effectors that are associated with repeat-rich regions. These candidate effectors share a highly conserved motif, and show isolate specific duplications. A reduced set of cell wall degrading enzymes, and LysM protein expansions were found in S. endobioticum, which may prevent triggering plant defense responses. Our study underlines the high diversity in chytrids compared to the well-studied Ascomycota and Basidiomycota, reflects characteristic biological differences between the phyla, and shows commonalities in genomic features among pathogenic fungi.</p

Comparative genomics of chytrid fungi reveal insights into the obligate biotrophic and pathogenic lifestyle of Synchytrium endobioticum

Synchytrium endobioticum is an obligate biotrophic soilborne Chytridiomycota (chytrid) species that causes potato wart disease, and represents the most basal lineage among the fungal plant pathogens. We have chosen a functional genomics approach exploiting knowledge acquired from other fungal taxa and compared this to several saprobic and pathogenic chytrid species. Observations linked to obligate biotrophy, genome plasticity and pathogenicity are reported. Essential purine pathway genes were found uniquely absent in S. endobioticum, suggesting that it relies on scavenging guanine from its host for survival. The small gene-dense and intron-rich chytrid genomes were not protected for genome duplications by repeat-induced point mutation. Both pathogenic chytrids Batrachochytrium dendrobatidis and S. endobioticum contained the largest amounts of repeats, and we identified S. endobioticum specific candidate effectors that are associated with repeat-rich regions. These candidate effectors share a highly conserved motif, and show isolate specific duplications. A reduced set of cell wall degrading enzymes, and LysM protein expansions were found in S. endobioticum, which may prevent triggering plant defense responses. Our study underlines the high diversity in chytrids compared to the well-studied Ascomycota and Basidiomycota, reflects characteristic biological differences between the phyla, and shows commonalities in genomic features among pathogenic fungi.</p

Molecular characterization and comparisons of potato wart (Synchytrium endobioticum) in historic collections to recent findings in Canada and the Netherlands

Synchytrium endobioticum (Schilb.) Perc. is a chytrid fungus causing potato wart disease and is one of the most important quarantine diseases on cultivated potato. Infected host tissues develop warts rendering the crop unmarketable. Resting spores, that can remain viable and infectious for decades, are formed in warted tissues and are released into the surrounding soil when host tissue decays. To better understand the pathogen’s diversity and to potentially uncover pathways of migrations and introduction events, molecular characterization was performed on the historical S. endobioticum resting spore collection of the Dutch National Plant Protection Organization. Mitochondrial genomes were assembled and annotated, and four novel structural variants were identified from these materials with intronic presence-absence variation in cox1 or cob genes and structural variation in the dpoB – TIR region. Several fungal isolates were shown to contain mixtures of structural variants. We analyzed the mitogenomic sequences obtained from recent potato wart disease findings in Canada and the Netherlands in the context of the historical materials and found that fungal isolates from the new Dutch outbreak contained a specific mixture of mitogenomic variants previously not observed in the Netherlands. Based on the mitogenomic profile, pathotype 38(Nevşehir) was suspected which was later verified with the Spieckermann bioassay. To further facilitate dissemination of data and interactive visual analytics we created a public Nextstrain webpage with S. endobioticum mitogenomic sequences and associated metadata on their geographic origin, pathotype identity and (mixture) of mitogenomic variants (https://nextstrain.nrcnvwa.nl/Sendo)

Guidelines for the reliable use of high throughput sequencing technologies to detect plant pathogens and pests

High-throughput sequencing (HTS) technologies have the potential to become one of the most significant advances in molecular diagnostics. Their use by researchers to detect and characterize plant pathogens and pests has been growing steadily for more than a decade and they are now envisioned as a routine diagnostic test to be deployed by plant pest diagnostics laboratories. Nevertheless, HTS technologies and downstream bioinformatics analysis of the generated datasets represent a complex process including many steps whose reliability must be ensured. The aim of the present guidelines is to provide recommendations for researchers and diagnosticians aiming to reliably use HTS technologies to detect plant pathogens and pests. These guidelines are generic and do not depend on the sequencing technology or platform. They cover all the adoption processes of HTS technologies from test selection to test validation as well as their routine implementation. A special emphasis is given to key elements to be considered: undertaking a risk analysis, designing sample panels for validation, using proper controls, evaluating performance criteria, confirming and interpreting results. These guidelines cover any HTS test used for the detection and identification of any plant pest (viroid, virus, bacteria, phytoplasma, mycetes, nematodes, arthropods, plants) from any type of matrix. Overall, their adoption by diagnosticians and researchers should greatly improve the reliability of pathogens and pest diagnostics and foster the use of HTS technologies in plant health