Long-Term Hatching Dynamics of the Sugar Beet Cyst Nematode, <i>Heterodera schachtii</i>（Rhabditida: Hoplolaimidae）, in Japan

Summary The hatching and emergence of juveniles from cysts of Heterodera schachtii in response to various stimuli are well understood. These responses relate to means of optimising the chances of successful reproduction and survival of populations. However, the effect of different cultivars of host species is not so well understood. To gain a further understanding of any differences between cultivars of host species, we conducted a number of experiments using root leachates obtained from a range of cultivars of sugar beet (Beta vulgaris), radish (Raphanus sativus) and white mustard (Sinapis alba). Heterodera schachtii cysts were exposed to root leachate solutions in in vitro laboratory assays during which emergence of second-stage juveniles (J2) from cysts was monitored weekly. Significant differences in the stimulation of hatch of J2 were found in response to different sugar beet cultivars and these differences may be driven by differing growth habits of such varieties. However, results obtained from experiments using brassica varieties showed contrasting responses. In one experiment, root leachates appeared to inhibit hatch of J2. However, two experiments did show a stimulation of hatch of J2 and also significant differences in hatch responses due to the age of the plant. These findings may help develop future screening procedures for brassica trap crops and help identify those that show the greatest potential for H. schachtii control.

The soybean cyst nematode (SCN) Heterodera glycines continues to be a major threat to soybean production worldwide. Morphological discrimination between SCN and other nematodes of the Heterodera schachtii sensu stricto group is not only difficult and time-consuming but also requires high expertise in nematode taxonomy. Molecular assays were developed to differentiate SCN from sugar beet cyst nematode (SBCN) and other nematodes and to quantify SCN directly from DNA extracts of field soils. SCN- and SBCN-specific quantitative real-time PCR (qPCR) primers were designed from a nematode-secreted CLAVATA gene and used for these assays. The primers were evaluated on the basis of target specificity to SCN or SBCN using DNA from 20 isolates of SCN and 32 isolates of other plant-parasitic nematodes. A standard curve relating threshold cycle and log values of nematode numbers was generated from artificially infested soils and was used to quantify SCN in naturally infested field soils. There was a high correlation between the SCN numbers estimated from naturally infested field soils by conventional methods, and the numbers quantified using the SYBR Green I-based qPCR assay. The qPCR assay is highly specific and sensitive and provides improved SCN detection sensitivity down to 1 SCN egg in 20 g of soil (10 eggs/200 g soil). This assay is useful for efficient detection and quantification of SCN directly from field soil. Species-specific conventional PCR assays were also developed each for SCN and SBCN, alongside a qPCR assay that simultaneously discriminates SCN from SBCN. These assays require no expertise in nematode taxonomy and morphology, and they may serve as useful diagnostic tools in research, diagnostic laboratories, and extension services for SCN management. Sensitive and accurate detection and quantification of SCN are essential for recommending effective management measures against SCN. We also investigated the impact of soil texture and nematode life stage on molecular quantification of SCN.

<i>Heterodera glycines</i>

Cyst nematodes remain a major threat to global agricultural production, causing huge losses. To understand the parasitism of the cyst nematodes Heterodera trifolii (HT) and Heterodera schachtii (HS), we constructed whole-genome assemblies using short- and long-read sequencing technologies. The nematode genomes were 379 Mb and 183 Mb in size, with the integrated gene models predicting 40,186 and 18,227 genes in HT and HS, respectively. We found more than half of the genes predicted in HT (64.7%) and HS (53.2%) were collinear to their nearest neighbor H. glycines (HG). Large-scale duplication patterns in HT and segmental duplications of more than half of the orthologous genes indicate that the genome of HT is polyploid in nature. Functional analysis of the genes indicated that 65.6% of the HG genes existed within the HT genome. Most abundant genes in HT and HS were involved in gene regulation, DNA integration, and chemotaxis. Differentially expressed genes showed upregulation of cuticle structural constituent genes during egg and female stages and cytoskeletal motor activity-related genes in juvenile stage 2 (J2). Horizontal gene transfer analyses identified four new vitamin biosynthesis genes, pdxK, pdxH, pdxS, and fabG, of bacterial origin, to be first reported in HT and HS. Mitogenomes of HT, HS, and HG showed similar structure, composition, and codon usage. However, rates of substitution of bases in the gene nad4l were significantly different between HT and HS. The described genomes, transcriptomes, and mitogenomes of plant-parasitic nematodes HT and HS are potential bio-resources used to identify several strategies of control of the nematode.

SUMMARY Taxonomy: Superkingdom Eukaryota; kingdom Metazoa; phylum Nematoda; class Chromadorea; order Tylenchida; suborder Tylenchina; superfamily Tylenchoidea; family Heteroderidae; subfamily Heteroderinae; main genera Heterodera and Globodera. Cyst nematodes comprise approximately 100 known species in six genera. They are pathogens of temperate, subtropical and tropical plant species and the host range of many species is narrow. The most economically important species are within the Globodera and Heterodera genera. Globodera pallida and G. rostochiensis are important pathogens of potato crops. There are many economic species in the Heterodera genus, including Heterodera glycines (soybean cyst nematode), H. avenae (cereal cyst nematode) and H. schachtii (sugar beet cyst nematode), the last of which attacks a range of Chenopodiaceae and Cruciferae, including Arabidopsis thaliana. Disease symptoms: Field symptoms of severe cyst nematode infection are often stunting, wilting and chlorosis, but considerable yield loss can occur without obvious symptoms. The only unique indicator of cyst nematode infection is the presence of adult female nematodes attached to host roots after several weeks of parasitism. Disease control: This is usually achieved by using integrated pest management involving cultural practices such as crop rotation, resistant cultivars if available and chemical control when economically justified.

Sugar beet cyst nematode (SBCN), Hederodera schachtii Schmidt. 1871, marked as one of the most damaging disease of sugar beet fields worldwide. Non chemical controls measures were carried out, using organic fertilizers including farm, poultry and green manure (waste cabbage leaves), compost (municipal’s waste products, 0.15 and 0.08 mm in size) and vermicomposting as the soil treatments at various rates against control (no treatments) in glass house and field highly infested with the nematode, SBCN in Isfahan, Iran. The related data on cysts final population, number of eggs and second larvae per cyst and per gram of soil were subjected to statistical analysis. The reproduction factor and the percent reduction and or increasing in population of nematode, SBCN were taken into consideration in comparison to initial populations. The results revealed that, poultry manure at the rate of 40 t ha -1 had the most and highly significant effects on the final population reduction in contrast to other ones, including controls. Followed by 0.15mm (60 t ha -1 ), compost 0.08 (60 t ha -1 ), poultry manure at the rate of 20 t ha -1 , with no significant differences respectively. Vermicomposting, cabbage leaves and form manure treatments had the least effects on the final population reduction in sugar beet cyst nematode populations. Keywords: compost municipality wastes, cyst nematodes, organic matters, Isfahan, Iran

The vegetative mycelium of Pleurotus ostreatus (oyster mushroom) exhibits the ability to reduce nematode populations. This property may be utilized in integrated management programs targeting harmful nematodes such as Heterodera schachtii, a major pest of sugar beet crops. In addition to sugar beet, many other plant species serve as hosts for this nematode; susceptible plants promote H. schachtii development and population growth. Current control strategies rely on integrated plant protection methods, including the use of tolerant cultivars, fallowing, and trap crops such as oilseed radish and white mustard. This study aimed to determine whether sugar beet cv. Janetka or nematocidal plants—oilseed radish cv. Romesa and white mustard cv. Bardena—affect the nematocidal activity of P. ostreatus mycelium when applied together. Specifically, the influence of root or seed secretions from these plants on the activity of ten P. ostreatus mycelial strains was assessed using the model nematode Caenorhabditis elegans and the target pest H. schachtii. Experiments were conducted under laboratory conditions on water agar media colonized by P. ostreatus mycelium. Seeds or root exudates of the tested plants were applied to the mycelial surface. Following incubation, nematode mobility (C. elegans) and cyst entwining by the mycelium (H. schachtii) were evaluated, along with the ability of the mycelium to produce toxocysts. The results indicate that trap plants did not significantly alter the nematocidal activity of the mycelium. However, certain mycelial strains were slightly stimulated by seed diffusates or root exudates. Oilseed radish moderately influenced the nematocidal activity of four mycelial strains against C. elegans, whereas in the case of H. schachtii, similar effects were observed with white mustard. The mycelial elimination of H. schachtii occurred through cyst entwining, which was generally more effective in the presence of plant exudates. Overall, the findings demonstrate that incorporating trap crops such as oilseed radish cv. Romesa or white mustard cv. Bardena, as green manure in crop rotation systems, does not interfere with the nematocidal activity of P. ostreatus mycelium and simultaneously may enrich the soil with nutrients. The study further confirms that P. ostreatus maintains its ability to effectively entwine and eliminate H. schachtii cysts even in the presence of sugar beet, supporting its potential role as a biological control agent. To our knowledge, this is the first experiment that integrates the activities of trap plants and sugar beet with the nematocidal effects of P. ostreatus mycelium.

In a previous paper published in 1937 Carroll &amp; McMahon discussed the possibility of effecting control of potato root eelworm (Heterodera schachtii) by means of trap cropping with potatoes. During the past two seasons further experiments, designed with the same object in view, have been carried out. The additional information obtained from these experiments and from the experience of the past two years is presented in the present paper.

An integrated strategy is required to control the sugar beet cyst nematode Heterodera schachtii which causes severe yield losses in sugar beet. This study aimed at evaluating the effect of trap crop cultivation (nematode resistant mustard, crop mixture (Trifolium alexandrinum L., Lupinus angustifolius L., Pisum sativum L., Phacelia tanacetifolia Benth., Guizotia abyssinica (L.F.) Cass., Avena strigosa Schreb., Vicia sativa L.), straw mulch as control), nematicide application before sugar beet sowing and sugar beet variety (susceptible, tolerant or resistant to H. schachtii) on population dynamics of H. schachtii and sugar yield. Field experiments were conducted in eight environments (site×year) located in Northern Germany in 2012–2013 and 2013–2014. An insufficient trap crop dry matter yield (1.1–2.5tha−1) mostly led to non-significant trap crop effects on the nematode population whereas in one environment a high dry matter yield of mustard (3.3tha−1) resulted in a nematode reduction of 40%. However, there were no significant differences to the straw mulch control. The nematicide application had no effect on nematode reproduction. In contrast, population dynamics of H. schachtii were strongly influenced by the sugar beet variety and the initial nematode population (PiSB). The highest reduction of 70% was achieved when a resistant sugar beet variety was grown, while the tolerant and susceptible varieties increased the nematode population in most environments. There was evidence that the sugar beet harvest date can highly influence population dynamics of H. schachtii. Sugar yield was influenced by variety and PiSB, but not by trap crop cultivation or nematicide application. Sugar yield decreased with increasing PiSB for all varieties. The resistant and tolerant varieties did not differ in sugar yield and response to H. schachtii, while the susceptible showed the steepest decline in sugar yield with increasing PiSB. The cultivation of the resistant sugar beet variety can be clearly suggested with respect to sugar yield and nematode control because all other tested factors failed to control the nematode population.

Beet cyst nematode-resistant cultivars, which were introduced recently, originated from the homozygous inbred line B883. This translocation product was unstable and the transmission of resistance when crossed with a susceptible cultivar did not exceed 94%. Tests with the resistant cultivars in climate cabinets showed a wide variety of resistance against Heterodera schachtii and beet necrotic yellow vein virus (BNYVV), expressed as average numbers of infective units per plant and percentages of resistant plants. In a series of field trials at different levels of infection of H. schachtii, their multiplication rates on all resistant cultivars depended on the initial density, which was caused by the presence of small numbers of susceptible plants. Since tolerance to wilting was also incorporated in B883, reasonable yields were obtained in the presence of H. schachtii. However, at increasing initial densities of H. schachtii, yields decreased considerably, since penetrating juveniles cause a hypersensitivity reaction in resistant plants. Based upon the results of three series of field trials, it was concluded that resistant cultivars should preferably be applied at population densities between 500 and 2000 eggs and juveniles of H. schachtii per 100ml of soil. Cultivars with double resistance against H. schachtii and BNYVV behaved like those with H. schachtii resistance in soils infected with beet cyst nematodes, but not with BNYVV. In soils with a combined infection of H. schachtii and BNYVV double resistant cultivars were far superior to single resistant ones, since damage caused by BNYVV was far more serious than damage caused by H. schachtii. No substantial interaction between soil pathogens nor types of resistance could be detected.

Climate change is expected to cause a mean annual temperature increase in Germany of 2 °C by 2050 and up to 4 °C by 2100. This is likely to have effects on crop development and pathogen development as well. For this Ph.D. thesis, experiments were conducted to investigate the effect of increasing temperatures on the beet cyst nematodes Heterodera schachtii and Heterodera betae and thus study the changes of their interaction with their host plant the sugar beet (Beta vulgaris subsp.). Differences in hatch between the beet cyst nematode species were assessed at constant temperatures in incubators as well as simulated temperature conditions set to be 4 °C higher than the standard temperature regime. The optimal temperatures for hatch were found to be different for both cyst nematode species. The optimal temperature range for hatching of H. schachtii was found to be between 15 and 30 °C and for H. betae between 20 and 30 °C. Emergence of juveniles of both beet cyst nematode species began at 5 °C, however in very low percentages. For both beet cyst nematode species no significant differences were found in the final cumulative hatch percentages when comparing the standard temperature regime with the by 4°C increased temperature regime. In the climate chambers, the influence of temperature on the interspecific competition between both beet cyst nematode species was studied. Both beet cyst nematode species performed better at higher temperatures. But there were no clear indications that under competition one of the two species will profit more from higher temperatures under the predicted climate change. In conclusion, the damage done by cyst nematodes in sugar beet is expected to increase with global warming. H. schachtii and H. betae are likely to continue to cause damages. But in competition, neither of the two species will profit more from rising temperatures compared to the other. The effect of experimental soil warming on H. schachtii population development and sugar beet performance was assessed for sugar beet cultivars that were susceptible, tolerant or resistant to H. schachtii. In this study, soil heating lead to a significant increase in the final number of recovered cysts on the tolerant cultivar and susceptible cultivar. The resistant cultivar did not allow nematode reproduction at all. Therefore no effect of soil heating could be detected. Plant-nematode interaction varies greatly depending on the cultivar. Thus cultivar choice is an important element when trying to prevent nematode infestation or controlling nematode populations in the field. In case of high nematode population densities and with the expected increasing soil temperatures, planting nematode-resistant cultivars will become even more important in the future, as an effective tool to reduce nematode populations and prevent damages.

The integration of oilseed rape (OSR) into sugar beet rotation systems is restricted due to the very good host status of OSR for the beet cyst nematode (BCN) Heterodera schachtii. In contrast to sugar beet, the cultivation of winter OSR covers a longer period, but at a lower soil temperature regime. Thus, presumably one or two generations of BCN may develop during the cultivation of winter OSR, resulting in moderate multiplication rates of 1–2 in the present study. This multiplication rate was year-dependent, but not affected by different sowing times. For the first time, the present study identified volunteer OSR emerging in high densities post-harvest as a major risk for a high multiplication of BCN at optimum temperatures. The emergence of BCN females with offspring was observed very early, resulting in a significant population increase before 350-degree days (&gt;8 °C) in inoculation experiments and in field investigations. Conducting treatment trials with glyphosate to control volunteer OSR in micro-plots and field experiments confirmed effective suppression of BCN reproduction when growth of volunteer OSR was interrupted at 250–350-degree days. Thus, data gained from BCN reproduction studies under controlled and field conditions provided a unique basis for the development of a trap crop method. The degree day model has been successfully implemented as part of an open access management tool.

SummaryA field experiment in which main‐crop potatoes were grown every other year was conducted on a sandy soil from 1994 to 1999. The aim of the experiment was to control soil‐borne pathogens of potato with ecologically sound methods. Potato grown as a trap crop from the end of April to the end of June (8 wk) was used to control potato cyst nematodes (PCN) (Globodera pallida), and its effects on other important soil pathogens and on the growth of a subsequent potato crop were also assessed. Additional experimental treatments were a potato crop from which the haulm was removed and a green manure crop. Three potato cultivars with different degrees of resistance to PCN were grown as the main crop. Duplicate sets of the experiment were run concurrently.The PCN were effectively controlled by the potato trap crop. When a highly resistant potato cultivar was grown as a main crop after the trap crop, the post‐harvest soil infestation was very low. When a moderately resistant cultivar was grown after the trap crop the soil infestation also remained low. When the trap crop was alternated with a susceptible potato cultivar as a main crop, soil infestation increased slightly, but the degree of control when compared with no trap crop averaged 96%.Soil infestation with root‐knot nematodes (mainly Meloidogyne hapla) increased when potato was grown as a trap crop, but soil infestation with the root‐lesion nematode Pratylenchus crenatus was not affected. Stem canker caused by Rhizoctonia solani was not affected by the trap crop but black scurf (sclerotia of R. solani) on tubers was reduced. Soil infestation with Verticillium dahliae declined in one of the duplicate sets of the experiment but not in the other. However, stem infections by V. dahliae were significantly decreased in both sets, although the effect depended on the PCN‐resistance level of the potato cultivar. When a highly resistant potato cultivar was grown Verticillium stem infections were not significantly affected, they were decreased with a moderately resistant cultivar but the decrease was most pronounced with a PCN‐susceptible cultivar.Senescence of a following potato crop was not influenced by the trap crop when a highly PCN‐resistant cultivar was grown, but it was delayed in the case of a moderately resistant or a susceptible cultivar, resulting in higher tuber yields for those cultivars.The experiment proved that a trap crop can be an alternative to chemical soil disinfection but, for several reasons, the potato itself is not an ideal crop for this purpose; a trap crop other than potato must be developed.

BackgroundSugar beet (Beta vulgaris subsp. vulgaris) is an economically important crop that provides nearly one third of the global sugar production. The beet cyst nematode (BCN), Heterodera schachtii, causes major yield losses in sugar beet and other crops worldwide. The most effective and economic approach to control this nematode is growing tolerant or resistant cultivars. To identify candidate genes involved in susceptibility and resistance, the transcriptome of sugar beet and BCN in compatible and incompatible interactions at two time points was studied using mRNA-seq.ResultsIn the susceptible cultivar, most defense-related genes were induced at 4 dai while suppressed at 10 dai but in the resistant cultivar Nemakill, induction of genes involved in the plant defense response was observed at both time points. In the compatible interaction, alterations in phytohormone-related genes were detected. The effect of exogenous application of Methyl Jasmonate and ET-generator ethephon on susceptible plants was therefore investigated and the results revealed significant reduction in plant susceptibility. Genes putatively involved in the resistance of Nemakill were identified, such as genes involved in phenylpropanoid pathway and genes encoding CYSTM domain-containing proteins, F-box proteins, chitinase, galactono-1,4-lactone dehydrogenase and CASP-like protein. Also, the transcriptome of the BCN was analyzed in infected root samples and several novel potential nematode effector genes were found.ConclusionsOur data provides detailed insights into the plant and nematode transcriptional changes occurring during compatible and incompatible interactions between sugar beet and BCN. Many important genes playing potential roles in susceptibility or resistance of sugar beet against BCN, as well as some BCN effectors with a potential role as avr proteins were identified. In addition, our findings indicate the effective role of jasmonate and ethylene in enhancing sugar beet defense response against BCN. This research provides new molecular insights into the plant-nematode interactions that can be used to design novel management strategies against BCN.

Sugar beet cyst nematode (SBCN), Heterodera schachtii, was first detected in 2017 from a highland vegetable production area of Hara village, Nagano Prefecture, central Japan.We should consider two important characteristics of nematode biology for application of control measures to prevent resurgence after emergency control by fumigation: 1) the local SBCN population can parasitize not only Brassicaceae and Amaranthaceae crops but also tomatoes; 2) its seasonal occurrence pattern could be complex due to single adult females producing eggs with different hatching properties, and to the possibility of two generations per year in the infestation area.We should also consider the local farming system: the size of an individual crop field is small, e.g., mean 1.5 ha for broccoli, a good host of SBCN, and the farmers have a tendency to produce a single crop item exclusively.Among the crop items cultivated in the infestation area, celer y, parsley, lettuce, maize, and zucchini were determined nonhosts.However, resistant host crop varieties are not available cur rently.Fumigation, organophosphate application, 2 month-cultivation of trap crops, 2 month-fallow, and 4 month-cultivation of nonhost crops reduced nematode population densities by m i n i mu m e s t i m at e s of 60, 40, 40, 30 a nd 10% , respectively.Here, we propose crop rotation systems integrated with these individual control measures to keep SBCN at an undetectably low density after emergency control.Nematol.Res. 54, 33-46.(2024)