Species Of Root-lesion Nematodes Research Articles

Development of a Recombinase Polymerase Amplification Assay for Rapid Detection of the New Root-lesion Nematode Pratylenchus dakotaensis on Soybean.

Root-lesion nematodes, Pratylenchus spp. are reported to cause serious yield losses in various crops including soybean. A new root-lesion nematode species was recently detected in a soybean field in North Dakota (ND) and named Pratylenchus dakotaensis. Nematode detection and differentiation from other species are critical in management strategies. Thus, a recombinase polymerase amplification (RPA) assay was developed for rapid detection of this nematode from field soils under isothermal conditions. New primers and probes were designed from ITS-rDNA region of the nematode genome and tested for both specificity and sensitivity. The RPA assay was able to detect DNA from a single adult nematode at 39.5°C in 20 minutes using both Basic and Exo kits. The specificity of the primers was initially confirmed through in silico analyses, followed by laboratory tests. The assay successfully amplified DNA from the target species, while no amplification occurred for other Pratylenchus spp. and non-Pratylenchus control species. Sensitivity testing with real-time RPA revealed its ability to detect DNA in dilutions equivalent to 1/32 of a single nematode from DNA extracted from inoculated sterile soil. To further validate the assay, it was tested with 19 field soil samples collected in ND. This assay amplified soil DNA extracts of all P. dakotaensis-infested field samples confirmed through conventional PCR. It did not amplify DNA from 13 other field soils infested with other Pratylenchus spp. This is the first report of RPA develoment for detecting a root-lesion nematode species. The RPA assay developed can help in the rapid detection of this nematode species for effective nematode management.

Distribution, morphological and molecular characterization of root-lesion nematodes (Pratylenchus spp.) (Tylenchida: Pratylenchidae) in maize (Zea mays L.) (Poales: Poaceae) in Türkiye.

Root-lesion nematodes (RLN) are the most economically important pathogenic nematodes attacking maize. Significant economic losses due to lesion nematodes have been reported in maize producing countries in the world. This study was conducted to determine the distribution and identity of root-lesion nematodes (Pratylenchus spp.) (Tylenchida: Hoplolaimidae) in maize (Zea mays L.) (Poales: Poaceae) fields of the Black Sea region of Türkiye. For this purpose, 39 locations were surveyed and soil samples were taken from 17 regional provinces. Nematodes were extracted using the modified Baerman funnel technique. The species were identified based on sequences of the Internal Transcribed Spacer (ITS) region of ribosomal DNA, as well as morphological characters and morphometrics. In addition, species identifications were confirmed using species-specific primers in the D3 expansion region of 26S rDNA. At the end of the study, 51.3% of the maize production areas sampled in the region were infected with root-lesion nematode species. Pratylenchus agilis, P. mediterraneus, P. neglectus, P. penetrans, P. thornei, and P. vulnus were identified, and were present in 25%, 5%, 25%, 10%, 15%, and 20% of samples, respectively. To our knowledge, this is the first report of P. agilis in Türkiye. The present study concluded that the molecular analysis of Pratylenchus sequences based on the ITS and D3 region of ribosomal RNA genes allowed the identification of six root lesion nematode species. This study is of great importance in terms of adding additional species to the root-lesion nematode fauna in Turkey and will provide data for future research on the management of these nematodes.

Prevalence of Verticillium spp. and Pratylenchus spp. in Commercial Potato Fields in Atlantic Canada

A survey of New Brunswick (NB) and Prince Edward Island (PEI) potato fields in crop rotation phase prior to potato production was conducted in fall (October and November) between 2017 and 2021. A total of 113 and 126 fields for NB and PEI, respectively, were surveyed with 20 to 35 fields each year tested in each province. Root lesion nematodes (RLN, Pratylenchus spp.) were detected in 99 and 98% of the fields for NB and PEI, respectively, and two root lesion nematode species, P. crenatus and P. penetrans, were identified in both provinces from 2017 to 2021. Based on 2019 and 2020 results, all surveyed fields in NB and PEI were detected with P. crenatus, while only 29 and 43% of the fields in NB and PEI were detected with P. penetrans, respectively. P. crenatus accounted for 96 and 89% of the populations for NB and PEI, respectively, while P. penetrans accounted for 4 and 11% in commercial fields, respectively. In a single in-depth sampled experimental field with a history of severe potato early dying complex in 2018 in NB, P. crenatus accounted for 88% and P. penetrans was 12%. Verticillium dahliae was detected in 94 and 92% of potato fields in NB and PEI, respectively. All isolates obtained from potato cv. “Russet Burbank” in a baiting trial were V. dahliae, belonging to two lineages. V. albo-atrum was detected in a few fields at very low level, except two fields in NB where V. albo-atrum was predominating over V. dahliae. Rotation crops did not affect V. dahliae population densities for NB and PEI, and did not affect RLN population in NB, but significantly affected RLN in PEI. Fall green cover crop did not affect the populations of RLN and V. dahliae in PEI. The present study revealed that the potato pathogenic root lesion nematode P. penetrans was present in less than 50% of surveyed fields and accounted for around 10% of root lesion nematode population in NB and PEI, and V. dahliae was the dominant species and was present in greater than 90% of surveyed fields in both provinces.

Development and validation of four TaqMan real-time PCR diagnostics for the identification and quantification of Pratylenchus crenatus, Pratylenchus neglectus, Pratylenchus penetrans and Pratylenchus thornei

Summary Pratylenchus crenatus, P. neglectus, P. penetrans and P. thornei are globally the most commonly occurring species of root-lesion nematodes (RLN). Correct identification and quantification of these nematodes is important for strategic management interventions such as rotation choice and nematicide use. A real-time quantitative PCR can provide a fast and reliable alternative to morphological identification, which requires significant taxonomic experience. A TaqMan hydrolysis probe method based on the 28S rDNA D2-D3 expansion region was developed and validated for the identification and quantification of these four root-lesion nematode species. Standard curves for each target RLN species were generated by plotting known gene copy number, obtained by a ten-fold serial dilution of purified plasmids, with corresponding Ct values. Each standard curve had a strong linear correlation () between Ct value and gene copy number. There was consistent amplification of samples with target species from different geographic locations within the UK, whereas a lack of amplification was noted for selected non-target species: P. coffeae, P. pseudocoffeae, P. vulnus, P. fallax, Globodera rostochiensis, Meloidogyne hapla, Trichodorus primitivus and Bitylenchus hispaniensis. Specificity and sensitivity of the methods were confirmed by three experiments that explored different life stages, increasing the number of target species, and had mixed Pratylenchus samples. Finally, estimates obtained by qPCR methods were compared with counting carried out by microscopy showing a good correlation (). The TaqMan real-time PCR developed in this study provides a specific, fast and accurate quantification of P. crenatus, P. neglectus, P. penetrans and P. thornei.

Evaluating the densities and distribution of root-lesion nematodes (Partylenchus spp.) in wheat grown in Canterbury, New Zealand

Species of root-lesion nematode (Pratylenchus spp.) are associated with significant reductions in wheat yield in wheat-growing regions around the world. Of these, Pratylenchus thornei and P. neglectus are known to cause the highest damage to the Australasian wheat industry. New Zealand is known to produce high wheat yields on a per-hectare basis yet little research has been conducted to date to determine the effects of Pratylenchus spp. on the production of wheat in New Zealand. Therefore, as the first step towards filling this knowledge gap, the current research focused on conducting surveys to determine the population densities and distribution of Pratylenchus spp. in wheat-growing regions in Canterbury, South Island, New Zealand. Surveys were conducted at ten selected sites that were geographically distinct from each other. At six of the ten sites, lesion nematode populations were reported to be above the recorded Australian threshold of 2000 nematodes per kg of soil. In Australia, it’s been recorded that around 50% yield reductions can occur in intolerant wheat varieties when population densities reaches this number. Differences in population density within each location was also observed indicating the uneven distribution of lesion nematodes within a field. Morphological measurements of the nematodes collected from multiple sites during this study confirmed the presence of P. thornei and P. neglectus in Canterbury wheat-producing areas indicating a potential threat to the New Zealand wheat industry by root-lesion nematodes. Further studies need to be conducted to fully understand the situation and to develop management strategies to mitigate threats from nematodes.

Multi-locus genome-wide association study of chickpea reference set identifies genetic determinants of Pratylenchus thornei resistance.

Pratylenchus thornei is an economically important species of root-lesion nematode adversely affecting chickpea (Cicer arietinum) yields globally. Integration of resistant crops in farming systems is recognised as the most effective and sustainable management strategy for plant-parasitic nematodes. However, breeding for P. thornei resistance in chickpea is limited by the lack of genetic diversity. We deployed a genome-wide association approach to identify genomic regions and candidate genes associated with P. thornei resistance in 285 genetically diverse chickpea accessions. Chickpea accessions were phenotyped for P. thornei resistance in replicated glasshouse experiments performed for two years (2018 and 2020). Whole genome sequencing data comprising 492,849 SNPs were used to implement six multi-locus GWAS models. Fourteen chickpea genotypes were found to be resistant to P. thornei. Of the six multi-locus GWAS methods deployed, FASTmrMLM was found to be the best performing model. In all, 24 significant quantitative trait nucleotides (QTNs) were identified, of which 13 QTNs were associated with lower nematode population density and 11 QTNs with higher nematode population density. These QTNs were distributed across all of the chickpea chromosomes, except chromosome 8. We identified, receptor-linked kinases (RLKs) on chromosomes 1, 4 and 6, GDSL-like Lipase/Acylhydrolase on chromosome 3, Aspartic proteinase-like and Thaumatin-like protein on chromosome 4, AT-hook DNA-binding and HSPRO2 on chromosome 6 as candidate genes for P. thornei resistance in the chickpea reference set. New sources of P. thornei resistant genotypes were identified that can be harnessed into breeding programs and putative candidate P. thornei resistant genes were identified that can be explored further to develop molecular markers and accelerate the incorporation of improved P. thornei resistance into elite chickpea cultivars.

Population Development of the Root-Lesion Nematode Pratylenchus dakotaensis on Soybean Cultivars.

Two greenhouse experiments were conducted with the new species of root-lesion nematode, Pratylenchus dakotaensis, discovered in North Dakota, USA in 2017 and named in 2021. Experiment 1 was conducted to ascertain resistance levels of soybean (Glycine max [L.] Merr.) cultivars to the new species P. dakotaensis and Experiment 2 was conducted to evaluate the population levels of P. dakotaensis in roots versus soil. A total of 20 soybean cultivars, and the positive control, cultivar Barnes, accompanying a nonplanted control, were evaluated in both experiments; they each had five replicates and the experiments were repeated. Among the 20 soybean cultivars evaluated, a cultivar with the ID# 13 consistently produced the greatest population densities of P. dakotaensis across all the experiments and therefore it was selected as the susceptible check. The ratio of the final nematode population density of a test cultivar relative to the final population density of the susceptible check was used to scale resistance rating. Combined results of all the trials indicated that seven of the cultivars were moderately resistant, nine of the cultivars plus 'Barnes' were moderately susceptible, and four of the cultivars were susceptible. However, none of the cultivars tested were classified as resistant. Analysis of P. dakotaensis population levels revealed that ≥50% of the population resided in the root habitat and the remaining nematodes were recovered from soil at 9 weeks after planting for a majority of the cultivars tested. These results provide an insight into the virulence of P. dakotaensis on commercial soybean cultivars.

Interaction effect of Root Lesion Nematodes and Fusarium culmorum Sacc. on disease complex in some wheat cultivars

The study was conducted to assess a possible interaction between fungal pathogen of Fusarium culmorum Sacc. and Root lesion nematodes at sequentially and simultaneously inoculation. Fusarium culmorum spore suspension effect evaluated on plant disease severity and nematode density on moderate resistant wheat line to F. culmorum (2-49) and susceptible wheat variety (İkizce) under controlled condition. The disease severity was higher on İkizce cv. than 2-49. This indicates that there is a considerable important pathogen resistance. Simultaneous and sequential inoculation of Pratylenchus thornei and F. culmorum reduced the final nematode density and reproduction rate more than inoculation with only nematode on İkizce cv. The simultaneously P. penetrans and F.culmorum inoculation (N+F) affected the positively of final density on İkizce cv. The N+ F treatment affected the positively of P. neglectus density on İkizce cv., whereas negative effect was found in pre or post inoculation of F. culmorum treatments. No synergistic interactions were detected on 2-49 when plants were co-infected by the root lesion nematode and fungus. It was determined that P. thornei had a positive effect on disease severity when it entered the plant simultaneously and before the F. culmorum on İkizce cv.. The disease severity decreased in F. culmorum was applied four weeks after the P. neglectus treatment on 2-49 wheat line and İkizce cv. In the interaction with F. culmorum on İkizce cv., differences were determined between the species of root lesion nematodes.

Development of Real-Time and Conventional PCR Assays for Identifying a Newly Named Species of Root-Lesion Nematode (Pratylenchus dakotaensis) on Soybean.

A rapid and accurate PCR-based method was developed in this study for detecting and identifying a new species of root-lesion nematode (Pratylenchus dakotaensis) recently discovered in a soybean field in North Dakota, USA. Species-specific primers, targeting the internal transcribed spacer region of ribosomal DNA, were designed to be used in both conventional and quantitative real-time PCR assays for identification of P. dakotaensis. The specificity of the primers was evaluated in silico analysis and laboratory PCR experiments. Results showed that only P. dakotaensis DNA was exclusively amplified in conventional and real-time PCR assays but none of the DNA from other control species were amplified. Detection sensitivity analysis revealed that the conventional PCR was able to detect an equivalent to 1/8 of the DNA of a single nematode whereas real-time PCR detected an equivalent to 1/32 of the DNA of a single nematode. According to the generated standard curve the amplification efficiency of the primers in real-time PCR was 94% with a R2 value of 0.95 between quantification cycle number and log number of P. dakotaensis. To validate the assays to distinguish P. dakotaensis from other Pratylenchus spp. commonly detected in North Dakota soybean fields, 20 soil samples collected from seven counties were tested. The PCR assays amplified the DNA of P. dakotaensis and discriminated it from other Pratylenchus spp. present in North Dakota soybean fields. This is the first report of a species-specific and rapid PCR detection method suitable for use in diagnostic and research laboratories for the detection of P. dakotaensis.

Pratylenchusvovlasi sp. Nov. (Nematoda: Pratylenchidae) on Raspberries in North Italy with a Morphometrical and Molecular Characterization.

Root-lesion nematode species rank third only to root-knot and cyst nematodes as having the greatest economic impact on crops worldwide. A survey of plant-parasitic nematodes associated with decaying raspberries (Rubus sp.) in northern Italy revealed that root-lesion nematodes were the most frequently occurring species among other phytonematodes. Several Pratylenchus species have been associated with Rubus sp. in Canada (Quebec, British Columbia) and USA (North Carolina, Maryland, New Jersey) including P. penetrans and P. crenatus. In the roots and rhizosphere of symptomatic raspberries, nematodes of two Pratylenchus spp. were detected. Detailed morphometrics of the two root-lesion nematode isolates were consistent with Pratylenchus crenatus and with an undescribed Pratylenchus species. The extracted nematodes were observed and measured as live and fixed materials and subsequently identified by integrative taxonomy (morphometrically and molecularly). The latter species is described herein as Pratylenchus vovlasi sp. nov., resulting morphometrically closest to P. mediterraneus and phylogenetically to P. pratensis. The molecular identification of Pratylenchus vovlasi sp. nov. was carried out by sequencing the ITS region, D2-D3 expansion domains of the 28S rRNA gene and a partial region of the nuclear hsp90 gene. ITS-RFLP and sequence analyses revealed that Pratylenchus vovlasi sp. nov. had species-specific restriction profiles with no corresponding sequences present in the database. The phylogenetic relationships with ITS and D2-D3 sequences placed the Pratylenchus vovlasi sp. nov. in a clade with P. pratensis and P. pseudopratensis. This research confirms the occurrence of cryptic biodiversity within the genus Pratylenchus as well as the need for an integrative approach to the identification of Pratylenchus species.

Morphometric and Molecular Diversity among Seven European Isolates of Pratylenchus penetrans.

Pratylenchus penetrans is an economically important root-lesion nematode species that affects agronomic and ornamental plants. Understanding its diversity is of paramount importance to develop effective control and management strategies. This study aimed to characterize the morphological and genetic diversity among seven European isolates. An isolate from the USA was included in the molecular analyses for comparative purposes. Morphometrics of the European P. penetrans isolates generally were within the range of the original descriptions for this species. However, multiple morphometric characteristics, including body length, maximum body width, tail length and length of the post-vulval uterine sac showed discrepancies when compared to other populations. Nucleotide sequence-based analyses revealed a high level of intraspecific diversity among the isolates. We observed no correlation between D2-D3 rDNA- and COXI-based phylogenetic similarities and geographic origin. Our phylogenetic analyses including selected GenBank sequences also suggest that the controversy surrounding the distinction between P. penetrans and P. fallax remains.

Morphological and Molecular Characterization of Pratylenchus dakotaensis n. sp. (Nematoda: Pratylenchidae), a New Root-Lesion Nematode Species on Soybean in North Dakota, USA.

Root-lesion nematodes (Pratylenchus spp.) of the genus Pratylenchus Filipjev, 1936, are among the most important nematode pests on soybean (Glycine max (L.) Merr.), along with soybean cyst and root-knot nematodes. In May 2015 and 2016, a total of six soil samples were collected from a soybean field in Walcott, Richland County, ND and submitted to the Mycology and Nematology Genetic Diversity and Biology Laboratory (MNGDBL), USDA, ARS, MD for analysis. Later, in 2019, additional nematodes recovered from a greenhouse culture on soybean originally from the same field were submitted for further analysis. Males, females, and juveniles of Pratylenchus sp. were recovered from soil and root samples and were examined morphologically and molecularly. DNA from single nematodes were extracted, and the nucleotides feature of three genomic regions targeting on the D2–D3 region of 28S rDNA and ITS rDNA and mitochondrial cytochrome oxidase subunit I (COX1) gene were characterized. Phylogeny trees were constructed to ascertain the relationships with other Pratylenchus spp., and polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was performed to provide a rapid and reliable differentiation from other common Pratylenchus spp. Molecular features indicated that it is a new, unnamed Pratylenchus sp. that is different from morphologically closely related Pratylenchus spp., including P. convallariae, P. pratensis, P. fallax, and P. flakkensis. In conclusion, both morphological and molecular observations indicate that the North Dakota isolate on soybean represents a new root-lesion nematode species which is named and described herein as Pratylenchus dakotaensis n. sp.

First report of root-lesion nematode, Pratylenchus oleae from pistachio in Iran.

Pistachio, Pistacia vera is one of the most important cash crops in Iran that is scattered in arid and semi-arid regions. During a survey of plant-parasitic nematodes of pistachio in Ardakan city in Yazd Province, a species of root-lesion nematode was isolated and identified by morphological, morphometrical, and molecular methods as Pratylenchus oleae Palomares-Rius et al., 2014. This species was isolated from several pistachio trees rhizosphere regarding to molecular analysis, D2–D3 expansion segments of 28S rRNA was amplified by PCR and sequenced. The sequence was deposited in GenBank (Accession No. MW338666). Along with the related phylogenetic analysis, placed this species in a monophyletic clade with other Pratylenchus oleae isolates, based on Bayesian inference (BI) phylogeny. A PCR reaction with the P. oleae specific primer set produced a 547 bp fragment. This is the first report of P. oleae infecting Pistachio tree in the world.

Pratylenchus smoliki, a new nematode species (Pratylenchidae: Tylenchomorpha) from the Great Plains region of North America.

Pratylenchus smoliki is a new species of root-lesion nematode described from corn-soybean production fields in the Central Great Plains of North America. It is characterized by populations with relatively abundant males, two lip annuli, females with a round functional spermatheca and a conoid to subcylindrical tail with a non-crenate, smooth terminus. In host preference tests, corn and wheat produce the largest nematode populations, whereas sorghum and soybeans produce less than 20% the numbers observed on corn. Scanning electron microscopy reveals that the en face patterns compare to those seen in Pratylenchus pseudocoffeae, P. scribneri, P. hexincisus, and P. alleni. The pattern is described as rectangular to trapezoidal subdorsal and subventral lips adjoining oral disc, but with a clear demarcation between the oral disc and the subdorsal and subventral sectors. A Maximum Likelihood COI tree recognizes P. smoliki as a moderately-well-supported clade with several haplotype subgroups. A Maximum Likelihood partial 28S tree provides strong support for the P. smoliki clade and reinforces the close relationships between species with similar en face patterns. Topotype specimens of P. alleni were demonstrably different from P. smoliki using DNA markers. The geographic range of P. smoliki overlaps with the ranges of P. alleni, P. scribneri, P. neglectus, P. hexicisus, and P. dakotaensis. The observed host range (corn, rye, sunflower, and wheat) suggests that P. smoliki may be native to the tallgrass prairie region of the Great Plains.

Occurrence and abundance of nematodes on onion in Turkey and their relationship with soil physicochemical properties

Summary The distribution of plant-feeding and free-living nematodes in large scale onion production areas in five geographical regions in Turkey was investigated in 2016 and 2017. Ditylenchus spp. and Tylenchus spp. were widely distributed. The stem and bulb nematode, Ditylenchus dipsaci, was found in 48 locations from 13 provinces. Other plant-feeding nematode genera were Pratylenchus, Paratylenchus and Pratylenchoides. Pratylenchus thornei was the most widely distributed root-lesion nematode species in onion fields in 11 locations from seven provinces. Pratylenchus neglectus was present in three locations and P. vulnus was in four locations. Aphelenchus spp. and Aphelenchoides spp. were the principal fungal-feeding nematodes in onion-growing areas. The most abundant bacterial-feeding nematode genera were Acrobeloides, Cephalobus, Eucephalobus and Rhabditis. Acrobeles and Wilsonema genera were low in occurrence and abundance. Nematodes from Dorylaimida and predator nematodes, Mononchus spp., were also found. The numbers of Ditylenchus from plant samples were significantly correlated positively to silt content, and significantly correlated negatively to organic matter and calcium content.

Identification of candidate effector genes of Pratylenchus penetrans.

Pratylenchus penetrans is one of the most important species of root lesion nematodes (RLNs) because of its detrimental and economic impact in a wide range of crops. Similar to other plant-parasitic nematodes (PPNs), P.penetrans harbours a significant number of secreted proteins that play key roles during parasitism. Here, we combined spatially and temporally resolved next-generation sequencing datasets of P.penetrans to select a list of candidate genes aimed at the identification of a panel of effector genes for this species. We determined the spatial expression of transcripts of 22 candidate effectors within the oesophageal glands of P.penetrans by insitu hybridization. These comprised homologues of known effectors of other PPNs with diverse putative functions, as well as novel pioneer effectors specific to RLNs. It is noteworthy that five of the pioneer effectors encode extremely proline-rich proteins. We then combined insitu localization of effectors with available genomic data to identify a non-coding motif enriched in promoter regions of a subset of P.penetrans effectors, and thus a putative hallmark of spatial expression. Expression profiling analyses of a subset of candidate effectors confirmed their expression during plant infection. Our current results provide the most comprehensive panel of effectors found for RLNs. Considering the damage caused by P.penetrans, this information provides valuable data to elucidate the mode of parasitism of this nematode and offers useful suggestions regarding the potential use of P.penetrans-specific target effector genes to control this important pathogen.

First Report of the Root-lesion Nematode Pratylenchus neglectus on Wheat (Triticum aestivum) in North Dakota

Root-lesion nematodes (Pratylenchus spp.) are important nematode pests that invade roots of plants and restrict productivity of wheat (Smiley et al. 2005). In August 2015, a soil sample was collected from a harvested wheat field in Walsh County, ND, and was found to contain 1,044 root-lesion nematodes per kg of soil using the sugar centrifugal flotation method. In October 2015, four soil samples were collected from the same field and had root-lesion nematodes ranging from 125 to 1,000/kg soil. One soil sample with 500 lesion nematodes/kg soil was used to inoculate hard red spring wheat cvs. Glenn and Faller. After 10 weeks of growth in a greenhouse room maintained at 22°C, wheat roots were harvested and washed, and light brown lesions were observed on lateral roots. The washed roots were cut into 1-cm segments for nematode extraction using the Whitehead tray method. After 48 h, lesion nematodes were recovered from the root tissues. Averages of 24 and 20 root-lesion nematodes per gram were found in the roots of Glenn and Faller, respectively. Nematodes from soil and wheat roots were examined morphologically and molecularly for species identification. Morphological measurements of adult females (n = 10) included body length (range = 402.0 to 495.0 μm, mean = 446.0 μm), stylet (15.0 to 17.5, 16.4), tail length (16.0 to 22.0, 18.8), body width (17.0 to 22.0, 19.6), anterior end to basal bulb (90.0 to 115.0, 101.6), a = (21.2 to 24.7, 23.1), b = (3.7 to 4.8, 4.4), c = (19.6 to 27.1, 23.8), and V (81.0 to 85.0%, 82.8%). The lip region had two annules, and was not set off, with anterior margins of apical lip annule convex, second annule was slightly wider than the first; lateral field had four lines with central zone of lateral field having oblique striae; tail terminus was smooth, rounded, or slightly oblique. The nematode species was identified as Pratylenchus neglectus (Rensch, 1924) Filipjev & Schuurmans Stekhoven, 1941 based on morphological and morphometric characteristics (Castillo and Vovlas 2007). DNA was extracted from single nematodes (n = 11) isolated from soil and wheat roots, and ITS region of rDNA was amplified (Tanha Maafi et al. 2003). PCR products from three nematodes were cloned using pGEM-T easy vector and sequenced, and the resulting ITS sequences were identical. The consensus sequence (GenBank Accession No. KU705392, 684 bp) was 99% homologous with one population of P. neglectus from China (JX046941) and 83% or less identical to other Pratylenchus spp. including P. thornei, a species closely related to P. neglectus. The specific primers from D3 28S rRNA and ITS rDNA were used to amplify DNA of eight nematodes and produced single bands specific for P. neglectus (Yan et al. 2008, 2013). The combination of the molecular tests confirmed the target species as P. neglectus. Two species of root-lesion nematodes, P. neglectus and P. thornei, were reported as damaging pathogens affecting wheat production in the Pacific Northwest (Smiley et al. 2005). To our knowledge, this is the first report of P. neglectus on wheat in North Dakota. The resistance levels of various wheat cultivars to this lesion nematode are being identified.

Morphometric and molecular characterization of populations of Pratylenchus kumamotoensis and P. pseudocoffeae (Nematoda, Pratylenchidae) newly recorded in Korea.

At least 70 species of root-lesion nematodes, Pratylenchus spp., have been identified worldwide, many of which are serious pests of various agricultural crop plants. In Korea, only 14 species have been recorded in vegetable and fruit fields. Here, we report two new records of root-lesion nematode species in Korea based on morphometric and molecular methods. Soil samples were collected from chrysanthemum cultivars in various regions. Morphometric diagnosis showed that two new records for Korea: Pratylenchus kumamotoensis in Chilgok County and Pratylenchus pseudocoffeae in Geumsan County. In addition, molecular diagnosis using the two sequences of the internal transcribed spacer (ITS) and the D2–D3 region of ribosomal DNA showed that these two species were most similar with those from Japan, Costa Rica and USA. The similarities of the ITS and D2–D3 sequences were 99.85 and 99.74%, respectively, for Pratylenchus kumamotoensis and 99.99 with Costa Rica populations and 99.86% with USA populations, respectively, for Pratylenchus pseudocoffeae. To our knowledge, this is the first report of two species in Korea.

The Pratylenchus penetrans Transcriptome as a Source for the Development of Alternative Control Strategies: Mining for Putative Genes Involved in Parasitism and Evaluation of in planta RNAi.

The root lesion nematode Pratylenchus penetrans is considered one of the most economically important species within the genus. Host range studies have shown that nearly 400 plant species can be parasitized by this species. To obtain insight into the transcriptome of this migratory plant-parasitic nematode, we used Illumina mRNA sequencing analysis of a mixed population, as well as nematode reads detected in infected soybean roots 3 and 7 days after nematode infection. Over 140 million paired end reads were obtained for this species, and de novo assembly resulted in a total of 23,715 transcripts. Homology searches showed significant hit matches to 58% of the total number of transcripts using different protein and EST databases. In general, the transcriptome of P. penetrans follows common features reported for other root lesion nematode species. We also explored the efficacy of RNAi, delivered from the host, as a strategy to control P. penetrans, by targeted knock-down of selected nematode genes. Different comparisons were performed to identify putative nematode genes with a role in parasitism, resulting in the identification of transcripts with similarities to other nematode parasitism genes. Focusing on the predicted nematode secreted proteins found in this transcriptome, we observed specific members to be up-regulated at the early time points of infection. In the present study, we observed an enrichment of predicted secreted proteins along the early time points of parasitism by this species, with a significant number being pioneer candidate genes. A representative set of genes examined using RT-PCR confirms their expression during the host infection. The expression patterns of the different candidate genes raise the possibility that they might be involved in critical steps of P. penetrans parasitism. This analysis sheds light on the transcriptional changes that accompany plant infection by P. penetrans, and will aid in identifying potential gene targets for selection and use to design effective control strategies against root lesion nematodes.

NEMATÓIDE DAS LESÕES ASSOCIADO A MUDAS DE MANGUEIRA EM ASSU-RN

RESUMO: O objetivo do presente trabalho foi relatar, pela primeira vez, população de Pratylenchus brachyurus associada a raízes de mangueira cv. Tommy Atkins no estádio de muda em solo naturalmente infestado no município de Assú, Rio Grande do Norte. A caracterização da referida população do nematóide foi fundamentada em estudo morfométrico realizado em 20 fêmeas coletadas das raízes. Os dados morfométricos foram comparados aos dados publicados por diferentes autores em trabalhos sobre a ocorrência da referida espécie de nematóide das lesões e permitiram a identificação da população como pertencente à P. brachyurus.