Molecular diagnostic assays have been developed and utilized to diagnose and to confirm the diagnoses of many plant-parasitic nematodes. We screened several gene sequences of soybean cyst nematode (SCN) [Heterodera glycines, Ichinohen] for their use as molecular markers. A methodology then was developed to use them to detect and quantify the number of H. glycines directly from Mississippi soil. A novel procedure utilizing multiple databases containing nematode DNA and EST sequences was developed to assist in the selection of SCN primers used in the PCR and qPCR assays. In vitro testing demonstrated that the DNA primers and probes developed from the novel procedure for the qPCR assays could accurately detect the presence of SCN. Subsequent testing resulted in a trend of increasing observed numbers of SCN contributing to increasing estimates by qPCR.

Soybean (Glycine max) yield is significantly affected by soybean cyst nematode (SCN), Heterodera glycines, and causes an annual loss of billions of US dollars. In this study, Glycine max xyloglucan endotransglycosylase/hydrolase gene (Gm-XTH52) was transformed into a nematode susceptible G. max [Williams 82/PI 518671] variety of soybean to test whether the protein expression has a role in resistance to H. glycines, and possible chemical changes the expression may cause in the plant composition. Expression level of the Gm-XTH52 gene was three times higher than in controls. Significant reduction in the number of SCN cysts suggested suppression of H. glycines parasitism upon transformation. While total sugar amounts did not significantly differ between the transformed and control plants, xyloglucan amounts of loosely bound sugars of genetically mosaic plants were significantly lower in comparison to controls. Control plants showed lower molecular weight sugars than the transformed plants not subjected to H. glycines infection.

Plant MAP3Ks have expanded significantly compared to their metazoan counterparts. A new, sequential workflow combining multispecies ortholog clustering and newly built, family-specific HMMs is used to identify the MAP3K gene family within seven plant species, allowing for a refinement of previously proposed gene family cladding and the novel identification of the MAP3K gene families in the allotetraploid cotton Gossypium hirsutum and newly sequenced monocot seagrass Zostera marina. The MAP3K gene family architecture is further refined and validated using bioinformatics analyses before the recently characterized Arabidopsis Raf-like MAP3K ILK1 is identified and characterized in upland cotton. Transient gene silencing reveals an increase in RKN susceptibility following GhILK1.1 silencing in the susceptible TM1 cultivar. No changes in susceptibility were seen in the resistant M240 cultivar or against reniform nematodes. GhILK1.1 is only the second cotton gene characterized to have a direct role in mediating RKN resistance.

The infection of plants by Heterodera glycines, commonly known as soybean cyst nematode (SCN), is a serious agricultural problem of worldwide extent. Meanwhile, it provides an excellent experimental model to study basic aspects of how cells function, in particular, during biotic challenge. Heterodera glycines challenges plant cells by initiating, developing and sustaining an interaction that results in the formation of a nurse cell from which the nematode derives nourishment. The presented experiments examine (1) how a cell can be de-differentiated and reprogrammed to perform a much different biological role and (2) how a cell’s immune responses can be engaged or suppressed to accomplish that goal. The observation of alpha soluble N-ethylmaleimide-sensitive factor attachment protein (alpha-SNAP) expression, its location within the rhg1 locus and known involvement in the vesicular transport machinery relating to defense made it a strong candidate for further functional analysis. Functional studies demonstrated that overexpression of alpha-SNAP in the susceptible G. max[Williams 82/PI 518671] genotype that lacks its expression results in the partial suppression of H. glycines infection. This indicated that the vesicles could be delivering cargo to the site of infection to engage a defense response. High levels of expression of a cell wall modifying gene called xyloglucan endotransglycosylase also occur during defense. XTHs associate with vesicles, act in the apoplast outside of the cell, and have a well-known function in cell wall restructuring. These observations indicated that alterations in the cell wall composition of nurse cells could be important for the successful defense response. Overexpression of a G. max xyloglucan endotransglycosylase (Gm-XTH) in the susceptible G. max[Williams 82/PI 518671] genotype resulted in a significant negative effect on H. glycines as well as R. reniformis parasitism. The results, including preliminary experiments on components of the vesicle transport system, identify a potent mechanism employed by plants to defend themselves from two types of plant-parasitic nematodes.

No description available.

<p>  <em>Heterodera glycines</em> or soybean cyst nematode (SCN) is one of the most yield-limiting soybean plant pathogens. The HG (<em>Heterodera glycines</em>) type test is a greenhouse test to determine the virulence of SCN populations to seven soybean “indicator” lines. Since there has not been a survey of SCN virulence phenotypes in Indiana in over a decade, a total of 124 samples were collected, and all tested positive for SCN. Of 42 selected populations for the HG type test, 88% were virulent to the soybean breeding line PI 88788, from which most SCN-resistant cultivars derive their resistance. All populations were virulent to at least one indicator line, including PI 548402 (‘Peking’), which pose problems for the future of soybean breeding for SCN. <em>Meloidogyne</em> spp. or root-knot nematodes (RKNs) are extremely damaging to agriculture, especially to specialty crops. RKN species are attracted by plant root exudates. The gene expression of <em>M. javanica</em> second-stage juveniles was determined after exposure to tomato root exudates, and the differentially expressed genes were analyzed.</p>

Thesis (M.A. Agricultural Management (Plant Production)) -- University of Limpopo, 2017 / Nemafric-BL and Nemarioc-AL phytonematicides, which are being researched and developed to serve as alternatives to methyl bromide, have not been tested against plant growth and accumulation of essential nutrient elements in watermelon (Citrullus lanatus) cultivars. The objectives of this study were two fold, to determine the (1) overall growth responses and accumulation of selected essential nutrient elements in watermelon cultivars ‘Congo’ and ‘Charleston Gray’ and suppression of Meloidogyne javanica in response to increasing concentrations of Nemafric-BL and Nemarioc-AL phytonematicides, and (2) efficacy of Velum, Nemafric-BL and Nemarioc-AL phytonematicides on growth of watermelon, accumulation of selected essential nutrient elements in leaf tissues of cv. ‘Congo’ and the suppression of population densities of Meloidogyne species. Objective 1 and Objective 2 were achieved under greenhouse and field conditions, respectively. At 56 and 90 days after initiating the greenhouse and field treatments, respectively, the data were collected and subjected to statistical analyses. Nemafric-BL phytonematicide significantly affected growth of watermelon cultivars ‘Charleston Gray’ and ‘Congo’, whereas Nemarioc-AL phytonematicide did not have any significant effects on the plant variables of both cultivars. Generally, plant growth variables and increasing concentrations of Nemafric-BL phytonematicide had density-dependent growth (DDG) patterns, which were quantified through either positive or quadratic equations (Chapter 3). In contrast, both phytonematicides had significant effects on selected nutrient elements in leaf tissues of both watermelon cultivars. The affected nutrient elements, P, Mn and Na versus increasing concentrations of phytonematicides exhibited the DDG patterns, which were also quantified through either positive or quadratic equations (Chapter 3). The phytonematicides were consistent in suppressing nematode numbers in both watermelon cultivars. Comparison of synthetic nematicide Velum and the two phytonematicides under field conditions suggested that, relative to untreated control, the three products each stimulated growth of watermelon cv. ‘Congo’. The efficacy of Nemafric-BL and Nemarioc-AL phytonematicides on suppression of population densities of Meloidogyne species was comparable to that of Velum. In conclusion, Nemafric-BL and Nemarioc-BL phytonematicides were highly effective in managing population densities of Meloidogyne species in watermelon cultivars and also affected the partitioning of selected nutrient elements in tissues. The study provided essential information that could assist in decision-making in nematode management in watermelon production, particularly in fertiliser application.

Thesis (M.Sc. Agriculture (Horticulture)) -- University of Limpopo, 2017 / Phytonematicides, mainly due to their allelopathic nature, might be highly phytotoxic to crops protected against nematode damage. Phytotoxicity issues are compounded by the fact that the efficacy of plant extracts on nematode suppression depended much on their concentration and duration of exposure to the nematodes. Phytotoxicity could result in low crop yield and/or even in the eventual death of the protected crops. Concentrations that were suppressive to nematode numbers, but phytotoxic to the tested crop would not be useful when applied as a post-planting phytonematicides. The Mean Concentration Stimulation Point (MCSP) values were developed from the Curve fitting Allelochemical Response Dosage (CARD)computer based model to ensure that a non-phytotoxic concentration was applied for each crop. The objective of this study was to determine whether a series of Nemarioc-AL and Nemafric-BL phytonematicide concentrations would provide the MCSP for beetroot (Beta vulgaris) under greenhouse, microplot and field conditions. The greenhouse treatments included 0, 2, 4, 8, 16 and 32% for each phytonematicide. The microplot treatments were 0, 0.8, 1.6, 3.2, 6.4 and 12.8% concentrations, whereas in the field trial treatments were 0, 2.4, 4.8, 9.6, 19.2 and 38.4%. Under each condition, treatments of Nemarioc-AL and Nemafric-BL phytonematicides were, in separate experiments arranged in a randomised complete block design, with the greenhouse, microplot and field trials having 15, 10 and 9 replications, respectively. In the greenhouse, seedlings were raised in 20cm diameter plastic pots, containing pasteurised river sand and commercial seedling growing medium Hygromix® at 3:1 (v/v) ratio. Each seeding was inoculated with 5000 eggs and second-stage juveniles (J2) of Meloidogyne incognita. Seedlings were irrigated with chlorine free tapwater every other day using 250 ml/plant, with irrigation substituted by the treatment once weekly. On the microplot, the procedures were as in the greenhouse except that they growing mixture comprised pasteurised soil collected from the site. In the field, seedlings were directly transplanted into the soil. At 56 days after inoculation, in the greenhouse trial, the effects of Nemarioc-AL and Nemafric-BL phytonematicides were highly significant (P ≤ 0.01) on root galls, contributing 77 and 72% in total treatment variation (TTV) of root galls, respectively. Relative to untreated control, the respective products reduced root galls by 28-72% and 43-67%. Nemarioc-AL and Nemafric-BL phytonematicides had MCSP values on beetroot of 18.1 and 6.4%, respectively, with overall sensitivity values of 0 and 1, respectively.There was no treatment effect on nematode since there was absence of nematode in untreated control.Under microplottrials,Nemafric-BL phytonematicide had significant (P≤ 0.05) effects on fresh root mass, dry root mass and root galls, contributing 20, 19 and 57% in TTV of the three variables, with relative increases for fresh root mass and dry root mass of 65-159% and 63-143%, respectively, whereas root galls were reduced by 82-100%. Nemafric-BL phytonematicide had MCSP value on beetroot of 10.2%, with overall sensitivity value of 4 units. There was no treatment effect on nematode since there was absence of nematode in untreated control. In microplots, Nemarioc-AL phytonematicide did not have significant effects on all plant variables. Under field conditions, the treatments did not have significant effects on plant variables. In conclusion, results of the current study suggested that under greenhouse and microplot conditions the MCSP values of the phytonematicides ranged from 6.4 to 18.1%, with a wide range of overall sensitivities of phytonematicides to the test of beetroot cultivar / National Research Foundation (NRF) and Land Bank Chair of Agriculture - University of Limpopo and the Agricultural Research Council Universities Collaboration Centre

Many diseases, such as Parkinson's disease and heavy metal poisoning, are associated with impaired or aberrant locomotion. Because the underlying mechanisms are difficult to study in humans, simpler metazoans like Caenorhabditis elegans are commonly employed to model these diseases. C. elegans is especially useful in this respect because its innate electrotactic behaviour allows instantaneous manipulation of its locomotion using mild electric fields in a microfluidic environment, the results of which can be captured on video. However, extraction of locomotory data from these videos is a major bottleneck to the throughput of the microfluidic electrotaxis platform. In the present study, we describe the development of novel software to analyze electrotaxis videos in an automated fashion. The software, dubbed the Automated Nematode Tracking System (ANTS), uses efficient, parameterless computer vision techniques to simultaneously track and assess movement characteristics of ambulating animals. In combination with the previously described microfluidic electrotaxis platform, ANTS promises to accelerate research with C. elegans models of locomotory dysfunction. / Thesis / Master of Applied Science (MASc)

Thesis (M.Sc.(Plant Protection)) -- University of Limpopo, 2022 / Runner beans are extremely sensitive to root-knot (Meloidogyne species) nematodes. Phytonematicides had been consistently used in managing population densities of Meloidogyne species in various crops, with the application technologies being restricted to the ground leaching technology (GLT) and botinemagation technology, each with its own advantages and disadvantages. The use of seeds as carriers of active ingredients of phytonematicides and then drying prior to sowing, is being considered as another potential application strategy in seeds with hypogeal germination. In such seeds, during seedling emergence the seed cover and the endosperm remain below the soil surface, just above the developing root system. As a result, in phytonematicide-primed seeds, the seed structures could serve as carriers for the active ingredients of phytonematicides. In cucurbitacin phytonematicides, Nemarioc-AL and Nemafric-BL phytonematicides contain cucurbitacin A and B, respectively as active ingredients. The objectives of the study were two-fold, namely, to determine whether runner bean (Phaseolus coccineus L.) seeds would (1) serve as carriers of active ingredients of cucurbitacin-containing phytonematicides without affecting seed germination under in vitro conditions, (2) serve as carriers of cucurbitacins intended for suppression of M. incognita population densities under greenhouse and microplot conditions. Two separate studies were conducted under laboratory conditions, with seven treatment solutions at 0, 2, 4, 8, 16, 32 and 64% Nemafric-BL or Nemarioc-AL phytonematicide. After exposure to separate solutions for 2 h, runner bean seeds were dried on laboratory benches for 72 h. Treatments were arranged in a completely randomised design (CRD), with 8 replications. Two layers of filter papers were placed inside each Petri dish seeded with 10 primed and dried seeds. Petri dishes were incubated inside LABCON growth chamber at 25ºC and 75% relative humidity. Successful seed germination, viewed as emergence of radicle from the testa, was recorded daily for a period of 10 days, with each count being removed from Petri dish to avoid re-counting. Under greenhouse and microplot conditions, primed-and dried seeds were sown in plastic pots containing 2 700 ml steam-pasteurised sandy loam soil, arranged in a randomised complete block design, replicated six times and eight times, respectively. Each seedling was inoculated by distributing 5 000 eggs and second-stage juveniles (J2) of M. incognita race 4 using a 50 ml plastic syringe. Originally, pots were irrigated using 500 ml chlorine-free tapwater, which was reduced to half after seedling emergence at every other day. Plant variables were collected at 56 days after inoculation and data were subjected to the Curve-fitting Allelochemical Response Dose algorithm model. In vitro, germination percentage (R 2 = 0.96), radicle length (R 2 = 0.89) and plumule diameter (R 2 = 0.96) versus Nemarioc-AL phytonematicide exhibited positive quadratic relations. Similarly, the variables versus Nemafric-BL phytonematicide, exhibited positive quadratic relations. In vitro, Mean Concentration Stimulation Point (MCSP) value of Nemarioc-AL phytonematicide on runner bean seeds was 1.05%, whereas for Nemafric-BL phytonematicide MCSP value was 0.58%. Under greenhouse conditions, plant height (R 2 = 0.97), chlorophyll content (R 2 = 0.92), dry shoot mass (R 2 = 0.98), dead nodule number (R 2 = 0.90), total pod number (R 2 = 0.97) and active nodule number (R 2 = 0.93) versus Nemarioc-AL phytonematicide exhibited positive quadratic relations., Similarly, chlorophyll content (R 2 = 0.95), gall rating (R 2 = 0.82), dry shoot weight (R 2 = 0.69), stem diameter (R 2 = 0.85) and total nodule number (R 2 = 0.86) versus Nemafric-BL phytonematicide exhibited positive quadratic relations. Under greenhouse conditions, MCSP values for Nemarioc-AL and Nemafric-BL phytonematicides were 4.18 and 3.69%, respectively. Under microplot conditions, total number of nodules (R 2 = 0.88), number of nodules dead (R 2 = 0.99), number of nodules active (R 2 = 0.95), fresh root mass (R 2 = 0.99), and fresh pod mass (R 2 = 0.99) versus Nemarioc-AL phytonematicide, exhibited positive quadratic relations, whereas plant height (R 2 = 0.85), number of nodules dead (R 2 = 0.87), dry shoot mass (R 2 = 0.97), fresh root mass (R 2 = 0.97) and total number of nodules (R 2 = 0.63) versus Nemafric-BL phytonematicide exhibited positive quadratic relations. Under microplot conditions, MCSP values for Nemarioc-AL and Nemafric-BL phytonematicides were 3.76 and 3.93%, respectively, each with ∑k = 0. All degrees of Nemarioc-AL and Nemafric-BL phytonematicides profoundly reduced nematode numbers under greenhouse and microplot trials. Based on the information obtained from this study, it was confirmed that runner bean (P. coccineus) is sensitive to Nemafric-BL and Nemarioc-AL phytonematicides supported by the Curve-fitting Allelochemical Response Dose (CARD) model results due to most plant variables that had sensitivity values of zero. In conclusion, the priming technology should be developed further since it has the potential of being successful in nematode management in seeds with hypogeal germination / National Research Foundation (NRF) and Potatoes South Africa