Yevtushenko, Dmytro

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    A genome resource for 192 Verticillium dahliae isolates infecting potatoes across Canada
    (ACS Publications, 2023) Arseneault, Tanya; Lafond-Lapalme, Joël; Gonzalez, Chary Esteban Quinche; Jordan, Katerina S.; Yevtushenko, Dmytro P.; Barrett, Ryan; Van der Heyden, Hervé; Chen, Dahu; Tenuta, Mario; Mimee, Benjamin
    Verticillium dahliae is an important soilborne pathogen causing Verticillium wilt. It is also the primary causal agent of potato early dying, a disease complex involving the root-lesion nematode. Here, we report the whole-genome sequencing of 192 isolates of V. dahliae originating from the major potato production areas across Canada. Our results yielded a resource of 277,010 genetic variations that will be useful for genetic analyses and revealed the presence of two major lineages, both present in all provinces but exhibiting differences in regional prevalence.
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    A survey of Helicotylenchus, Paratylenchus, Pratylenchus, and Tylenchorhynchus nematodes in potato fields in Alberta, 2018 and 2019
    (Taylor & Francis, 2021) Robertson, C. J.; Yevtushenko, Dmytro P.; Snowdon, E.; Harding, M. W.
    The prevalence of Helicotylenchus, Paratylenchus, Pratylenchus and Tylenchorhynchus genera of nematodes is not well understood across the prairie provinces of Canada. These nematodes can cause economic damage by feeding on the host crop directly or serve as vectors of plant diseases. While conducting a larger project, nematode populations were quantified in the soil of three commercial potato fields planted with cultivar ‘Russet Burbank’; one in 2018 and two in 2019. The nematodes were extracted from soil samples, identified morphologically, and then quantified as numbers per kg of fresh soil. All four genera were detected in all fields, but the population sizes varied between fields and across time within fields.
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    Combining air sampling and DNA metabarcoding to monitor plant pathogens
    (APS Publications, 2023) Reich, Jonathan; Chen, Wen; Radford, Devon; Turkington, Kelly; Yevtushenko, Dmytro P.; Hamelin, Richard; Chatterton, Syama
    Monitoring the air for airborne plant pathogens is an increasingly common method for the management of economically important plant diseases. In Alberta, Canada, several commodity clusters, including dry bean, canola, potato, and wheat, currently support air monitoring research programs for airborne pathogens of interest. In this study, we assessed the feasibility of monitoring for these, and more, plant fungal pathogens simultaneously using two different sampler types (cyclone versus rotation impaction) and by metabarcoding the ITS1 region using the Illumina sequencing platform. We collected air samples from four geographically distant sites across Alberta and monitored four crop types in southern Alberta. Overall, we found weak, but statistically significant, effects of geographic location and crop type on the aeromycobiota community composition. A few common taxa, such as Ramularia, Alternaria, and Epicoccum, constituted the vast majority of reads across all samples. Nevertheless, in each sample, we identified many plant pathogens of interest and organisms that previous research has found antagonistic to those pathogens, highlighting the utility of these approaches in understanding the pathobiome. In assessing the real-world implications of read counts, we discovered that they were only weakly correlated with spore counts quantified by qPCR. The two types of samplers collected different community profiles, reinforcing the importance of carefully considering which sampler type to use in monitoring programs. Taken together, our results show promise for the future of monitoring the air pathobiome, although much more work is required to understand the relationship of airborne communities to their in-field impact on disease development.
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    Description of Filenchus species from agroecosystem of southern Alberta, Canada
    (MDPI, 2022) Munawar, Maria; Castillo, Pablo; Yevtushenko, Dmytro P.
    Understanding the existing nematode biodiversity is of significant concern because nematodes may divert nutrients from plants and use them for their own development and reproduction. The presence and diagnostics of Filenchus species occurring in southern Alberta have not been addressed in previous studies. Herein, we provide a comprehensive characterization of adult females of four known Filenchus species (F. cylindricus, F. hazenensis, F. sheri, and F. thornei) recovered from cultivated fields in southern Alberta. Three of the species are new records in Canada, while one is a native species that was previously described from the Canadian high arctic area. These organisms are mild parasitic species; we describe them here to enhance the visibility of soil nematodes and facilitate accurate species identification. The diagnostic resolution within Filenchus is low, because many species are described without adequate consideration of intra-specific variation. The species descriptions and molecular data obtained during the present study will reduce the confusion in examining the existing lineages among Filenchus species and will aid in improving phylogenetic resolution. Our results suggest that the known diversity of Canadian nemato-fauna has increased. However, more research is needed to further identify other genera and species of phytoparasitic nematodes that may occur in grasses, weeds, and wild plants present in cultivated areas. Moreover, the molecular characterization of these species from Canada, in comparison to a reference dataset (NCBI) of Tylenchidae nematodes, provides insight into the biogeography of nematodes.
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    Discovery and characterization of four Aphelenchid species from cultivated regions of southern Alberta, Canada
    (MDPI, 2024) Munawar, Maria; Castillo, Pablo; Yevtushenko, Dmytro P.
    The nematode family Aphelenchoididiae is considered fungal-feeding, predatory, or root hair feeders. Some members of this family are universally present in agricultural landscapes and are an integral part of soil health and conservation studies. In the present soil nematode biodiversity survey, we detected four species of the genera Aphelenchus, Aphelenchoides, and Robustodorus. Because fungal-feeding nematodes from southern Alberta have not previously been reported, we conducted a detailed morphological and molecular investigation, identifying these species as Aphelenchus avenae, Aphelenchoides limberi, Aphelenchoides prairiensis n. sp. and Robustodorus paramegadorus n. sp. The first two species we document as new records from southern Alberta, whereas A. prairiensis n. sp. and R. paramegadorus n. sp. we describe in detail as new taxa. Briefly, A. prairiensis n. sp. is an amphimictic species having 4 lateral lines; hemispherical anteriorly flattened lip region; delicate stylet and swelling-like stylet knobs; excretory pore at the posterior edge of nerve ring. Female tail conical, gradually tapering towards a truncated end with single mucro. Spicule 23.0 (20.0–25.0) µm long having elongated rounded condylus, small, blunt conical rostrum, and lamina that gradually tapers towards the rounded distal end; three pairs of caudal papillae were present on the male tail. Robustodorus paramegadorus n. sp., is a parthenogenetic species with 3 lines in the lateral fields; lip region rounded, anteriorly flattened; stylet robust, with knobs rounded to bean-shaped; excretory pore located posterior to nerve ring; reproductive components were quite indiscernible with a short 24.0 (18.0–27.0) µm post-vulval uterine sac; tail conical, ending with pointed to wedge-shaped tip. We performed molecular characterizations for each species and constructed phylogenetic trees to study the phylogenetic relationship of these aphelenchid species. The discovery of A. prairiensis n. sp. and R. paramegadorus n. sp. indicates that soil nematode diversity is relatively unexplored in southern Alberta. The findings of this study will significantly enhance the identification processes and may contribute towards future soil health and biodiversity efforts.
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    Douglas-fir LEAFY COTYLEDON1 (PmLEC1) is an active transcription factor during zygotic and somatic embryogenesis
    (Wiley, 2021) Vetrici, Mariana A.; Yevtushenko, Dmytro P.; Misra, Santosh
    Douglas-fir (Pseudotsuga menziesii) is one of the world's premier lumber species and somatic embryogenesis (SE) is the most promising method for rapid propagation of superior tree genotypes. The development and optimization of SE protocols in conifers is hindered by a lack of knowledge of the molecular basis of embryogenesis and limited sequence data. In Arabidopsis, the LEAFY COTYLEDON1 (AtLEC1) gene is a master regulator of embryogenesis that induces SE when expressed ectopically. We isolated the LEC1 homologue from Douglas-fir, designated as PmLEC1. PmLEC1 expression in somatic embryos and developing seeds demonstrated a unique, alternating pattern of expression with the highest levels during early stages of embryogenesis. PmLEC1 protein accumulation during seed development correlated with its transcriptional levels during early embryogenesis; however, substantial protein levels persisted until 2 weeks on germination medium. Treatment of mature, stratified seeds with 2,4-epibrassinolide, sorbitol, mannitol, or NaCl upregulated PmLEC1 expression, which may provide strategies to induce SE from mature tissues. Sequence analysis of the PmLEC1 gene revealed a 5′ UTR intron containing binding sites for transcription factors (TFs), such as ABI3, LEC2, FUS3, and AGL15, which are critical regulators of embryogenesis in angiosperms. Regulatory elements for these and other seed-specific TFs and biotic and abiotic signals were identified within the PmLEC1 locus. Most importantly, functional analysis of PmLEC1 showed that it rescued the Arabidopsis lec1-1 null mutant and, in the T2 generation, led to the development of embryo-like structures, indicating a key role of PmLEC1 in the regulation of embryogenesis.
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    First report of three Tylenchidae taxa from southern Alberta, Canada
    (MDPI, 2021) Munawar, Maria; Yevtushenko, Dmytro P.; Castillo, Pablo
    Members of the family Tylenchidae are highly abundant in soil habitats, including agricultural settings, where they play key ecological roles. In the present study, we identified three Tylenchidae species, namely Basiria bhabi, Coslenchus acceptus, and Filenchus vulgaris, using integrative taxonomy. The detailed morphological and morphometric characteristics, distribution, and host associations of each species were also discussed. Phylogenetic analyses of these populations with other Tylenchidae nematodes indicated the presence of divergent lineages in Filenchus and Basiria, whereas Coslenchus appeared to be a monophyletic genus. Herein, we aim to grow awareness about this common but least studied group of nematodes. The species reported in this study are new records for Canada, revealing that the identified nematode diversity in our cultivated areas is relatively underrepresented. Our analyses also provided greater taxonomic resolution and captured rare taxa that might have been missed or misidentified in prior nematode inventory surveys. These findings will add to our understanding of the nematofauna of southern Alberta, thereby providing a more complete picture of existing nematode diversity present in the fields of this highly cultivated region.
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    Integrative taxonomy, distribution, and host associations of Geocenamus brevidens and Quinisulcius capitatus from southern Alberta, Canada
    (Sciendo, 2021) Munawar, Maria; Yevtushenko, Dmytro P.; Castillo, Pablo
    Two stunt nematode species, Geocenamus brevidens and Quinisulcius capitatus, were recovered from the potato growing regions of southern Alberta, described and characterized based on integrative taxonomy. Morphometrics, distribution, and host associations of both species are discussed. The Canadian populations of both species displayed minor variations in morphometrical characteristics (viz., slightly longer bodies and tails) from the original descriptions. The populations of G. brevidens and Q. capitatus species examined in this study are proposed as standard and reference populations for each respective species until topotype specimens become available and molecularly characterized. Phylogenetic analyses, based on partial 18S, 28S, and ITS sequences, placed both species with related stunt nematode species. The present study updates the taxonomic records of G. brevidens and Q. capitatus from a new location, southern Alberta, Canada, and will aid in the decision whether these stunt nematodes should be included in nematode management programs.
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    Morpho-anatomy, in vitro culture, and phylogenetic studies of two helicotylenchus species from southern Alberta, Canada
    (MDPI, 2024) Munawar, Maria; Konschuh, Michele; Castillo, Pablo; Yevtushenko, Dmytro P.
    Spiral nematodes (Helicotylenchus spp.) are polyphagous parasitic species exhibiting a broad host range and geographical distribution. However, their diversity in the cultivated regions of southern Alberta remains understudied. Hence, we conducted a comprehensive survey of the region’s arable lands for the presence of spiral nematodes and revealed two Helicotylenchus species, H. crassatus and H. oscephalus. H. crassatus consisted of two distinct morphotypes: one morphotype had a conoid tail with slight ventral projection on the distal end, whereas the other had a broadly rounded tail. This study presents the first documentation of H. crassatus and H. oscephalus from southern Alberta, Canada. Molecular characterization was based on the partial 18S rRNA, the D2–D3 of 28S rRNA, ITS rRNA, and COI gene sequences, complemented by detailed morphological studies using scanning electron microscopy. In this work, Helicotylenchus species were often co-detected with root lesion nematodes, which made the evaluation of their role in crop damage more difficult. To meet the requirements for threshold and pathogenicity assessments, we introduced both spiral nematode species to sterile carrot disks and evaluated the feasibility of their multiplication and mass production in vitro. The present findings expand the taxonomic records of Helicotylenchus spp. and improve diagnostics of these morphologically similar species. Furthermore, our in vitro culture technique will provide a reliable source of the initial inoculum for future plant–nematode interaction studies.
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    Morphological and molecular characterization of Nothotylenchus medians and N. similis (Nematoda: Anguinidae) from southern Alberta, Canada
    (MDPI, 2022) Munawar, Maria; Rahman, Atta Ur; Castillo, Pablo; Yevtushenko, Dmytro P.
    The nematode family, Anguinidae, is a diverse group of polyphagous nematodes, generally known as fungal feeders or parasites of aerial plant parts. Here, we present the morphological and molecular characterization of adult females of two Nothotylenchus species, N. medians and N. similis, along with host association and geographical distribution data of the genus. Both species are recorded as new reports from Canada and designated as reference populations for future studies. Morphological or morphometrical variation was not observed in the Canadian population of N. medians and N. similis, in comparison with the original description. Phylogenetic analyses based on 18S and D2–D3 of 28S genes placed both species within Anguinidae. Since the biology of the genus Nothotylenchus has not been rigorously characterized, the habitat and distribution information presented in this study will shed some light on the ecology of these nematodes. Notably, the detection of N. medians and N. similis in our nematode inventory survey indicates that considerable Nothotylenchus diversity is hidden in these soils. Consequently, increased surveys and more in-depth research are needed to explore the full diversity of anguinids inhabiting these cultivated areas.
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    New records of Ditylenchus species from southern Alberta, Canada
    (MDPI, 2023) Munawar, Maria; Rahman, Atta U.; Castillo, Pablo; Yevtushenko, Dmytro P.
    The presence of plant-parasitic nematodes (PPNs) in cultivated areas is a limiting factor in achieving marketable crop yield. To control and alleviate the effects of these nematodes and determine appropriate management strategies, species-level identification is crucial. Therefore, we conducted a nematode diversity survey, which resulted in the detection of four Ditylenchus species in cultivated areas of southern Alberta, Canada. The recovered species had six lines in the lateral field, delicate stylets (>10 µm long), distinct postvulval uterine sacs, and pointed to rounded tail tips. The morphological and molecular characterization of these nematodes revealed their identity as D. anchilisposomus, D. clarus, D. tenuidens and D. valveus, all of which are members of the D. triformis group. All of the identified species were found to be new records in Canada except for D. valveus. Accurate Ditylenchus species identification is crucial because false-positive identification can result in the implementation of quarantine measures over the detected area. Our current study not only documented the presence of Ditylenchus species from southern Alberta, but also described their morpho-molecular characteristics and subsequent phylogenetic relationships with related species. The results of our study will aid in the decision on whether these species should become a part of nematode management programs since nontarget species can become pests due to changes in cropping patterns or climate.
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    Overexpression of Douglas-Fir LEAFY COTYLEDON1 (PmLEC1) in Arabidopsis induces embryonic programs and embryo-like structures in the lec1-1 mutant but not in wild type plants
    (MDPI, 2021) Vetrici, Mariana A.; Yevtushenko, Dmytro P.; Misra, Santosh
    Somatic embryogenesis (SE) is the most promising method for the quick propagation of desirable plant genotypes. However, application of SE to conifers remains challenging due to our limited knowledge about the genes involved in embryogenesis and the processes that lead to somatic embryo formation. Douglas-fir, an economically important lumber species, possesses a homolog of the angiosperm embryo-regulatory LEC1 gene. In the present study, we analyzed the potential of Douglas-fir PmLEC1 to induce embryonic programs in the vegetative cells of a heterologous host, Arabidopsis thaliana. PmLEC1 complemented the Arabidopsis lec1-1 null mutant and led to a variety of phenotypes ranging from normal morphology to developmental arrest at various stages in the T1 generation. PmLEC1 did not affect the morphology of wild type Arabidopsis T1 plants. More profound results occurred in T2 generations. PmLEC1 expression induced formation of recurrent somatic embryo-like structures in vegetative tissues of the rescued lec1-1 mutant but loss of apical dominance (bushy phenotype) in wild type plants. The activation of embryonic programs in the lec1-1PmLEC1 T2 plants was confirmed by the presence of the embryo-specific transcripts, OLEOSIN and CRUCIFERIN. In contrast, no embryo-like structures, and no OLEOSIN or CRUCIFERIN were observed in PmLEC1-expressing bushy wild type T2 plants.
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    Overview of the genus Boleodorus and first reports of Boleodorus thylactus and B. volutus from southern Alberta, Canada
    (MDPI, 2021) Munawar, Maria; Yevtushenko, Dmytro; Castillo, Pablo
    The present study provides the morphological and molecular characterization of Boleodorus thylactus and B. volutus populations, recovered from agricultural fields of southern Alberta. Despite a significant abundance of this group of nematodes, none of the Boleodorus species were previously reported in Canada. Therefore, representative adult specimens of each population were photographed and examined morphometrically. Phylogenetic trees were reconstructed using partial D2–D3 expansion segments of the 28S and 18S rDNA sequences to understand the relationships of Boleodorus species with Tylenchidae-related genera. Boleodorus species are relevant to soil ecological studies and therefore we summarized the important morphological and morphometric characters in tabular form for easy and efficient species identification. Moreover, we discuss the associated hosts and the distribution of all described Boleodorus species. This study will serve as a guide and basic framework for species diagnostics in the genus Boleodorus and will aid in filling the gaps in our knowledge of the species present in our cultivated lands.
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    Predicting airborne ascospores of Sclerotinia sclerotiorum through machine learning and statistical methods
    (Wiley, 2024) Reich, Jonathan; McLaren, Debra; Kim, Yong Min; Wally, Owen; Yevtushenko, Dmytro P.; Hamelin, Richard; Chatterton, Syama
    A main biological constraint of dry bean (Phaseolus vulgaris) production in Canada is white mould, caused by the fungal pathogen Sclerotinia sclerotiorum. The primary infectious propagules of S. sclerotiorum are airborne ascospores and monitoring the air for inoculum levels could help predict the severity of white mould in bean fields. Daily air samples were collected in commercial dry bean fields in Alberta, Manitoba and Ontario and ascospores were quantified using quantitative PCR. Daily weather data was obtained from in-field weather stations. The number of ascospores on a given day was modelled using 63 different environmental variables and several modelling methods, both regression and classification approaches, were implemented with machine learning (ML) (random forests, logistic regression and support vector machines) and statistical (generalized linear models) approaches. Across all years and provinces, ascospores were most highly correlated with ascospore release from the previous day (r ranged from 0.15 to 0.6). This variable was also the only variable included in all models and had the greatest weight in all models. Models without this variable had much poorer performance than those with it. Correlations of ascospores with other environmental variables varied by province and sometimes by year. A comparison of ML and statistical models revealed that they both performed similarly, but that the statistical models were easier to interpret. However, the precise relationship between airborne ascospore levels and in-field disease severity remains unclear, and spore sampling methods will require further development before they can be deployed as a disease management tool.
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    Relationship between desiccation tolerance and biofilm formation in shiga toxin-producing Escherichia coli
    (MDPI, 2024) Javed, Muhammad Qasim; Kovalchuk, Igor; Yevtushenko, Dmytro P.; Yang Xianqin; Stanford, Kim
    Shiga toxin-producing Escherichia coli (STEC) is a major concern in the food industry and requires effective control measures to prevent foodborne illnesses. Previous studies have demonstrated increased difficulty in the control of biofilm-forming STEC. Desiccation, achieved through osmotic stress and water removal, has emerged as a potential antimicrobial hurdle. This study focused on 254 genetically diverse E. coli strains collected from cattle, carcass hides, hide-off carcasses, and processing equipment. Of these, 141 (55.51%) were STEC and 113 (44.48%) were generic E. coli. The biofilm-forming capabilities of these isolates were assessed, and their desiccation tolerance was investigated to understand the relationships between growth temperature, relative humidity (RH), and bacterial survival. Only 28% of the STEC isolates had the ability to form biofilms, compared to 60% of the generic E. coli. Stainless steel surfaces were exposed to different combinations of temperature (0 °C or 35 °C) and relative humidity (75% or 100%), and the bacterial attachment and survival rates were measured over 72 h and compared to controls. The results revealed that all the strains exposed to 75% relative humidity (RH) at any temperature had reduced growth (p < 0.001). In contrast, 35 °C and 100% RH supported bacterial proliferation, except for isolates forming the strongest biofilms. The ability of E. coli to form a biofilm did not impact growth reduction at 75% RH. Therefore, desiccation treatment at 75% RH at temperatures of 0 °C or 35 °C holds promise as a novel antimicrobial hurdle for the removal of biofilm-forming E. coli from challenging-to-clean surfaces and equipment within food processing facilities.
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    Simple sequence repeat analysis of new potato varieties developed in Alberta, Canada
    (Wiley, 2019) Tillault, Anne-Sophie; Yevtushenko, Dmytro P.
    The worldwide demand for potato production requires the constant development of new potato varieties with improved yield, quality, disease resistance, and abiotic tolerance. However, cultivar registration is preceded by a long process to morphologically and physiologically characterize the plants. Notably, this process can be expedited by DNA marker analysis. Simple sequence repeats (SSRs), also known as microsatellites, are the most common reliable DNA markers used to discriminate between genotypes. In this study, 20 potato varieties, including five new genotypes developed in Alberta, Canada, were fingerprinted using 10 SSR markers selected for their high discriminatory power. Different SSRs were amplified from potato DNA using specific primers, and the DNA fragment sizes were analyzed by denaturing polyacrylamide gel electrophoresis. The number of alleles per locus ranged from two for the SSR marker STPoAc58 to six for STM0030 and STM0037 with an average of 4.4. In addition, a total of 77 unique patterns were observed for the 10 SSR markers. The polymorphic information content ranged from 0.477 to 0.802 with an average of 0.675 per locus. In this study, STM0037, STM1016, and STM1104 were found to be the best SSR markers to detect genetic differences between potato varieties. A minimum of two markers was required to distinguish between all 20 genotypes. Most importantly, this highly informative molecular tool confirmed that the developed potato varieties were genetically different from their respective maternal lines and potentially constituted new cultivars.
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    Species diversity of pin nematodes (Paratylenchus spp.) from potato growing regions of southern Alberta, Canada
    (MDPI, 2021) Munawar, Maria; Yevtushenko, Dmytro ; Palomares-Rius, Juan E.; Castillo, Pablo
    Pin nematodes (Paratylenchus spp.) are polyphagous parasitic species with a wide host range and geographical distribution; their diversity is unknown in the potato growing region of Alberta, Canada. The present study aims to provide morphological and molecular characterization of three pin nematode species, namely P. neoprojectus, P. tateae, and a new species, Paratylenchus enigmaticus sp. nov. All of them were recovered from the potato growing region of southern Alberta. The nematodes were isolated using the sieving and flotation-centrifugation method, and their morphology was assessed by light microscopy. Molecular characterization was performed using partial 18S, D2–D3 expansion domains of the 28S and ITS ribosomal genes. This study is the first report of molecular characterization of P. tateae and P. neoprojectus, being new records from southern Alberta, and two Spanish populations of P. tateae comprising the first report of this species in Europe. The phylogenetic analysis of the 18S, D2–D3 expansion domains of the 28S and ITS ribosomal DNA regions underscores the importance of using molecular data for accurate species identification and clarifies the status of P. nanus type B and P. sheri. Moreover, our findings will be useful to determine the impact of pin nematodes on potato production in future field research.
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    Survey of potato early dying disease complex in Alberta's commercial potato fields, 2021-2022
    (Taylor & Francis, 2023) Rahman, A. U.; Munawar, M.; Konschuh, Michele; Tenuta, M.; Harding, M. W.; Yevtushenko, Dmytro P.
    Potato fields were evaluated for inci dence and severity of early dying disease complex in the growing seasons of 2021 and 2022. Disease sure varied among the fields in both years. Fields taining higher inoculum levels of Verticillium dahliae in the fall soil samples as determined by qPCR of soil samples typically showed more potato early dying (PED) symptom development in the subsequent ing season. The incidence of PED was higher in 2022 than in 2021. Results demonstrated the potential of qPCR evaluation of representative soil samples as an aid for risk prediction activities in potato production systems dealing with PED.