Effects of dual inoculation of mycorrhiza and endophytic,rhizospheric or parasitic bacteria on the root-knot nematode disease of tomato

The effects of mycorrhisation and inoculation with soil bacteria on the disease caused by Meloidogyne incognita on tomato were studied in pots under greenhouse conditions. Efficacy in promoting plant growth and reducing disease severity and final nematode densities were evaluated for two arbuscular mycorrhizal fungi (AMF; Funneliformis mosseae and Rhizophagus irregularis), three soil bacteria with different living strategies (the endophyte Bacillus megaterium, a rhizospheric Pseudomonas putida and the hyperparasite of nematodes Pasteuria penetrans) and combinations of the fungi and bacteria. In M. incognita-infested plants, F. mosseae increased tomato growth more than R. irregularis, and plants inoculated with B. megaterium presented higher shoot fresh weight than with P. putida or P. penetrans, but dual inoculation did not improve tomato growth more than single inoculations. Disease severity and final nematode densities were reduced by F. mosseae compared to non-mycorrhizal plants. B. megaterium and P. penetrans reduced both the root galling and the final nematode densities compared to treatments without bacteria. P. penetrans reduced final nematode densities more than B. megaterium or P. putida. Dual inoculation of AMF and P. penetrans showed the highest efficacy in reducing the final nematode densities in tomato.     

Consequences of variation in species diversity in a community of root-feeding herbivores for nematode dynamics and host plant biomass

To date, no study has explicitly addressed effects of variation in species diversity of root‐feeding herbivores on host plant biomass. Root‐feeding nematodes typically occur in multi‐species communities. In a three‐year field experiment, we investigated how variation in species diversity of root‐feeding nematodes affected nematode dynamics and response of the dune grass Ammophila arenaria to root‐feeder activity. This plant species needs regular burial by fresh beach sand to remain vigorous, suggesting that A. arenaria benefits from a temporary escape from root‐feeding soil organisms and that root‐feeders are involved in plant degeneration in stabilized dunes. We created series of ceased and continued sand burial and added the endoparasitic nematodes Meloidogyne maritima, Heterodera arenaria and Pratylenchus penetrans alone or in combination to A. arenaria. We included treatments with and without the whole soil community, measured plant biomass and quantified numbers of nematodes. Addition of H. arenaria and P. penetrans decreased numbers of M. maritima juveniles and delayed the first appearance in time of both juveniles and females, while numbers of males only decreased when plants had been buried. Burial with sand and addition of the other two endoparasites affected numbers of H. arenaria juveniles, while numbers of P. penetrans were low and not affected. Shoot biomass of A. arenaria was lower when M. maritima had been added alone than when the three species had been added together. Addition of root zone soil decreased biomass of all plant parts. Burial with sand decreased aboveground shoot biomass, whereas it increased belowground shoot and root biomass. Our results point at idiosyncratic effects of nematode diversity on A. arenaria biomass. Heterodera arenaria and P. penetrans protected their host by reducing numbers and delaying activity of M. maritima to a later stage in the growth season, when root‐feeding activity was less harmful for plant biomass development.     

Infection of Seedlings of Alfalfa and Red Clover by Concomitant Populations of Meloidogyne incognita and Pratylenchus penetrans

Invasion of 2-day-old seedlings of ''Buffalo'' alfalfa and ''Kenland'' red clover by larvae of M. incognita and adults of P. penetrans, during 1-3 day periods of incubation at 24 C, was investigated in 50-mm petri dishes on 1% agar. Penetration by both nematodes increased arithmetically with increased numbers in inocula. P. penetrans invaded alfalfa more readily than red cover, but M. incognita invaded red clover more readily than alfalfa. Both nematodes inhibited root-elongation of alfalfa more than that of red clover. In combinations of 10 and 50 of both nematodes, invasion of both plants by both nematodes was the same as for each nematode alone. Penetration by M. incognita into alfalfa, but not into red clover, was significantly reduced when combinations of 50 M. incognita and 200 P. penetrans were inoculated simultaneously. In the presence of large numbers of entrant P. penetrans in both plants, penetration by M. incognita was highly significantly reduced. Penetration by P. penetrans was unaffected in the reciprocal situations.     

Pathogenicity of Pratylenchus penetrans,Heterodera glycines,and Meloidogyne incognita on Soybean Genotypes

The pathogenicity of Heterodera glycines, Meloidogyne incognita, and Pratylenchus penetrans on H. glycines-resistant ''Bryan,'' tolerant-susceptible ''G88-20092,'' and intolerant-susceptible ''Tracy M'' soybean cultivars was tested using plants grown in 800 cm³ of soil in 15-cm-diam. clay pots in three greenhouse experiments. Plants were inoculated with 0, 1,000, 3,000, or 9,000 H. glycines race 3 or M. incognita eggs, or vermiform stages of P. penetrans/pot. Forty days after inoculation, nmnbers of all three nematodes, except H. glycines on Bryan, generally increased with increasing inoculum levels in Experiment I. Heterodera glycines and M. incognita significantly decreased growth only of Tracy M. At 45 and 57 days after inoculation with 6,000 individuals/pot in experiments II and III, respectively, significantly more P. penetrans and M. incognita than H. glycines were found on Bryan. However, H. glycines and M. incognita population densities were greater than P. penetrans on G88-20092 and Tracy M. Growth of Tracy M infected by H. glycines and M. incognita and growth of G88-20092 infected by M. incognita decreased in Experiment III. Pratylenchus penetrans did not affect plant growth. Reduction in plant growth differed according to the particular nematode species and cultivar, indicating that nematodes other than the species for which resistance is targeted can have different effects on cultivars of the same crop species.     

Effects of Monoclonal Antibodies,Cationized Ferritin,and Other Organic Molecules on Adhesion of Pasteuria penetrans Endospores to Meloidogyne incognita

The incidence of adhesion of Pasteuria penetrans endospores to Meloidogyne incognita second-stage juveniles (J2) was studied after pretreatment of the latter with monoclonal antibodies (MAb), cationized ferritin, and other organic molecules in replicated trials. Monoclonal antibodies developed to a cuticular epitope of M. incognita second-stage juveniles gave significant reductions in attachment of P. penetrans endospores to treated nematodes. MAb bound to the entire length of J2 except for the area of the lateral field, where binding was restricted to the incisures. Since reductions in attachment with MAb treatment were modest, it is uncertain if these results implicated a specific surface protein as a factor that interacted in binding of the endospore to the nematode cuticle. Endospore attachment was decreased following treatment of the nematode with the detergents sodium dodecyl sulfate (SDS) and cetyltrimethylammonium bromide (CTAB). Endospore attachment to live nematodes was significantly greater than attachment to dead nematodes. Attachment rates of three P. penetrans isolates to M. incognita race 3 varied between isolates. The effects of neuraminidase, pronase, pepsin, trypsin, lipase, and Na periodate on endospore attachment were inconsistent. The cationic dye alcian blue, which binds sulfate and carboxyl groups on acidic glycans, had no consistent effect on endospore attachment. The incidence of endospore attachment was significantly lower but modest, at best, for nematodes that were treated with cationized ferritin alone or cationized ferritin following monoclonal antibody. The lack of consistency or extreme reduction in most experiments suggests that attachment of P. penetrans spores to M. incognita is not specified by only one physico-chemical factor, but may involve a combination of at least two physico-chemical factors (including surface charge and movement of the J2). This points to a need for analysis of combined or factorial treatment effects.     

Elevated CO2 changes the interactions between nematode and tomato genotypes differing in the JA pathway

Interactions between the root‐knot nematode Meloidogyne incognita and three isogenic tomato (Lycopersicon esculentum) genotypes were examined when plants were grown under ambient (370 ppm) and elevated (750 ppm) CO₂. We tested the hypothesis that, defence‐recessive genotypes tend to allocate ‘extra’ carbon (relative to nitrogen) to growth under elevated CO₂, whereas defence‐dominated genotypes allocate extra carbon to defence, and thereby increases the defence against nematodes. For all three genotypes, elevated CO₂ increased height, biomass, and root and leaf total non‐structural carbohydrates (TNC):N ratio, and decreased amino acids and proteins in leaves. The activity of anti‐oxidant enzymes (superoxide dismutase and catalase) was enhanced by nematode infection in defence‐recessive genotypes. Furthermore, elevated CO₂ and nematode infection did not qualitatively change the volatile organic compounds (VOC) emitted from plants. Elevated CO₂ increased the VOC emission rate only for defence‐dominated genotypes that were not infected with nematodes. Elevated CO₂ increased the number of nematode‐induced galls on defence‐dominated genotypes but not on wild‐types or defence‐recessive genotypes roots. Our results suggest that CO₂ enrichment may not only increase plant C : N ratio but can disrupt the allocation of plant resources between growth and defence in some genetically modified plants and thereby reduce their resistance to nematodes.     

Do endogeic earthworms change plant competition? A microcosm study

Plants compete for limited resources. Although nutrient availability for plants is affected by resource distribution and soil organisms, surprisingly few studies investigate their combined effects on plant growth and competition. Effects of endogeic earthworms (Aporrectodea jassyensis), root-knot nematodes (Meloidogyne incognita) and the spatial distribution of ¹⁵N labelled grass litter on the competition between a grass (Lolium perenne), a forb (Plantago lanceolata) and a legume (Trifolium repens) were investigated in the greenhouse. Earthworms promoted N uptake and growth of L. perenne. Contrastingly, shoot biomass and N uptake of T. repens decreased in the presence of earthworms. P. lanceolata was not affected by the earthworms. We suggest that earthworms enhanced the competitive ability of L. perenne against T. repens. Nematodes increased the proportion of litter N in each of the plant species. Litter distribution (homogeneous vs. patch) did not affect the biomass of any plant species. However, P. lanceolata took up more ¹⁵N, when the litter was homogeneously mixed into the soil. The results suggest that endogeic earthworms may affect plant competition by promoting individual plant species. More studies including decomposers are necessary to understand their role in determining plant community structure.     

Polarographische Bestimmung von Chlorat-Rückständen im Boden und in der Pflanze

An increase in the inoculum level of root‐knot nematode, Meloidogyne incognita and the reniform nematode, Rotylenchulus reniformis resulted in a relative decrease in plant growth parameters of chickpea. Consequently water absorption capability of roots was impaired. M. incognita caused greater reduction than R. reniformis at the same inoculum level. In concomitant inoculation of M. incognita and R. reniformis there was greater suppression in plant growth of chickpea. The suppression in concomitant inoculations was less than the sum of the suppression caused by the same levels of inoculations of the individual species. The multiplication rate of the nematodes decreased as the inoculum level increased. The results also suggest competition for feeding sites between the two nematode species. The multiplication rate of one species progressively decrease with the increase in the inoculum levels of the other nematode.     

Evaluation of Perennial Glycine Species for Response to Meloidogyne Incognita,Rotylenchulus Reniformis,and Pratylenchus Penetrans

Root-knot (Meloidogyne incognita (Kofoid & White) Chitwood), reniform (Rotylenchulus reniformis Lindford & Oliveira), and lesion nematodes (Pratylenchus penetrans (Cobb) Filipjev & Schuurmans Stekhoven) are plant-parasitic nematodes that feed on soybean (Glycine max (L.) Merr.) roots, limiting seed production. The availability of resistance in soybeans to these nematodes is limited. However, new sources of resistance can be discovered in wild relatives of agronomic crops. Perennial Glycine species, wild relatives to soybean, are a source of valuable genetic resources with the potential to improve disease resistance in soybean. To determine if these perennials have resistance against nematodes, 18 accessions of 10 perennial Glycine species were evaluated for their response to M. incognita and R. reniformis, and eight accessions of six perennial Glycine species were evaluated for their response to P. penetrans. Pot experiments were conducted for M. incognita and R. reniformis in a growth chamber and in vitro experiments were conducted for P. penetrans. We found both shared and distinct interactions along the resistance-susceptible continuum in response to the three plant-parasitic nematode species. Ten and 15 accessions were classified as resistant to M. incognita based on eggs per gram of root and gall index, respectively. Among them, G. tomentella plant introductions (PIs) 446983 and 339655 had a significantly lower gall index than the resistant soybean check cv. Forrest. Of three R. reniformis resistant accessions identified in this study, G. tomentella PI 441001 showed significantly greater resistance to R. reniformis than the resistant check cv. Forrest based on nematodes per gram of root. In contrast, no resistance to P. penetrans was recorded in any perennial Glycine species.     

Effects of earthworms and organic litter distribution on plant performance and aphid reproduction

Human management practices and large detritivores such as earthworms incorporate plant litter into the soil, thereby forming a heterogeneous soil environment from which plant roots extract nutrients. In a greenhouse experiment we investigated effects of earthworms and spatial distribution of ¹⁵N-labelled grass litter on plants of different functional groups [Lolium perenne (grass), Plantago lanceolata (forb), Trifolium repens (legume)]. Earthworms enhanced shoot and root growth in L. perenne and P. lanceolata and N uptake from organic litter and soil in all plant species. Litter concentrated in a patch (compared with litter mixed homogeneously into the soil) increased shoot biomass and ¹⁵N uptake from the litter in L. perenne and enhanced root proliferation in P. lanceolata when earthworms were present. Growth of clover (T. repens) was rather independent of the presence of earthworms and organic litter distribution: nevertheless, clover took up more nitrogen in the presence of earthworms and exploited more ¹⁵N from the added litter than the other plant species. The magnitude of the effects of earthworms and organic litter distribution differed between the plant species, indicating different responses of plants with contrasting root morphology. Aphid (Myzus persicae) reproduction was reduced on P. lanceolata in the presence of earthworms. We suggest that earthworm activity may indirectly alter plant chemistry and hence defence mechanisms against herbivores.     

Observations on the pathogenicity of Pasteuria penetrans, a parasite of root-knot nematodes

A simple test was used to determine whether or not Pasteuria penetrans spores would attach to 17 species of nematodes. All susceptible individuals had spores attached to their cuticles after 24 h of gentle agitation in suspensions containing 10⁵spores/ml. Spores of P. penetrans from six populations of Meloidogyne only adhered to species of Meloidogyne and they adhered in greatest numbers to the species from which they had been originally isolated. Sonication of spores from infected females increased attachment but the effect was dependent on pH and whether the test was conducted in tap or distilled water. Invasion of tomato roots was reduced by up to 86% when, rather than using healthy juveniles, second-stage juveniles bearing 15 or more spores were added to soil at high densities (1000 or 3000/plant); at low densities (500/plant) invasion was not significantly affected. The rate of development of M. incognita juveniles infected with P. penetrans was slower than that of healthy juveniles. The numbers of second-generation of M. incognita were reduced by 82–93% when juveniles encumbered with 1–15 spores were added to soil instead of those bearing no spores. Pasteuria penetrans populations differed in their aggressiveness and when juveniles encumbered with the same number of spores from two populations were added to soil there were differences in the numbers of females that became infected. The implications of these results for the development of P. penetrans as a biological control agent are discussed.     

Effect of Meloidogyne incognita on Reproduction of Pratylenchus penetrans in Red Clover and Alfalfa

Roots of seedlings of red clover and alfalfa growing on 10⁻¹ Hoagland and Arnon solution agar were inoculated with various combinations of Meloidogyne incognita and Pratylenchus penetrans. Egg-laying by P. penetrans decreased as the number of nematodes, the ratio of entrant M. incognita to entrant P. penetrans, and the priority of invasion of roots by M. incognita increased. Embryogeny and hatching of eggs of P. penetrans, and development of larvae of M. incognita, were not affected. In red clover, the greatest red uction occurred when there were 65 entrant nematodes, the ratio of M. incognita:P. penetrans was 4:1 and M. incognita was inoculated four days prior to P. penetrans. In alfalfa, the less-favorable host for both nematodes, the greatest reduction occurred when there were 45 entrant nematodes, the ratio of M. incognita:P. penetrans was 2:1, and M. incognita was inoculated 4 days prior to P. penetrans.     

Pseudomonas fluorescens mediated suppression of Meloidogyne incognita infection of cowpea and tomato

Plant growth-promoting rhizobacterium, Pseudomonas fluorescens strain BICC602 suppresses root-knot nematode (Meloidogyne incognita) by enhancing defence mechanism leading to induced systemic resistance in cowpea (Vigna unguiculata) cv. L.Walp. and tomato (Solanum lycopersicum) cv. Pusa Ruby. In cowpea, the soil treatment proved more effective than foliar spray on root galling and eggs in roots. However, which factors are necessary in the induction of resistance response in plants against nematodes by BICC602 is not yet known. Salicylic acid (SA) production by some bacteria acts as endogenous signal for the activation of certain plant defence responses. In a split-root trial with tomato as a host plant and M. incognita as challenging parasite, BICC602 induces systemic resistance in tomato plants. Based on the results, it is assumed that P. fluorescens-induced resistance against M. incognita in cowpea and tomato is made either through SA-dependent or SA-independent transduction pathway.     

Effect of inter-cropping bell-pepper with ginger on plant parasitic nematode populations and crop yields

During 1994 studies were undertaken to improve ginger (Zingiber officinale Roscoe) yield against the nematodes Pratylenchus penetrans, Meloidogyne incognita, Helicotylenchus dihystera and Tylenchorhynchus mashhoodi in Himachal Pradesh (HP) (India) by inter-cropping bell-pepper (Capsicum annum L) in eight different sequences (treatments). Inter-cropping of “one rhizome of ginger x one plant of bell-pepper” gave the highest ginger yield (600 g per rhizome). This treatment was completely free from P. penetrans and M. incognita. All treatments with bell-pepper plants equal to or higher in number to that of ginger rhizomes had higher ginger yields than treatments with ginger alone or with fewer bell-pepper plants. In the former, populations of P. penetrans and M. incognita were lower than in the latter treatments. The yield of ginger varied irrespective of population densities of H. dihystera and T. mashhoodi, indicating that P. penetrans and M. incognita are the major nematode problems of ginger in HP. Bell-pepper was a non-host to P. penetrans and non-preferred host to M. incognita. This helped to improve ginger yields by making the rhizosphere unfavourable for the development and multiplication of the major ginger nematode pests.     

Competition between Heterodera glycines and Meloidogyne incognita or Pratylenchus penetrans: Independent Infection Rate Measurements

Competition on soybean between Heterodera glycines (race 3) and Meloidogyne incognita or H. glycines and Pratylenchus penetrans were investigated in greenhouse experiments. Each pair of nematode species was mixed in 3-ml suspensions at ratios of 1,000:0, 750:250, 500:500, 250:750, and 0:1,000 second-stage juveniles or mixed stages for P. penetrans. Nematodes from a whole root system were counted and infection rates standardized per 1,000 nematodes (per replication) prior to testing the null hypothesis through a lack-of-fit F-test. Although the effect of increasing H. glycines proportions on the infection rate of M. incognita was generally adverse, the rate deviated significantly from a trend of linear decline at the 75% H. glycines level in one of two experiments. All lack-of-fit F-tests for the H. glycines and P. penetrans mix were significant, indicating that infection rates for both nematodes varied considerably across inocula. The infection rate of H. glycines decreased with increasing P. penetrans proportions. The rate of P. penetrans infection increased with increasing H. glycines proportions up to the 50% level, but declined at the 75% level. Competition had no effect on nematode development. The general adverse relationships between M. incognita and H. glycines and those between P. penetrans and H. glycines showed a linear trend. The relationship between H. glycines and P. penetrans indicates that the former may be competitive when present at higher proportions than the latter. In this study we have evaluated nematode competition under controlled conditions and provide results that can form a basis for understanding the physical and physiological trends of multiple nematode interactions. Methods critical to data analyses also are outlined.     

Low-level herbivory by root-knot nematodes (Meloidogyne incognita) modifies root hair morphology and rhizodeposition in host plants (Hordeum vulgare)

Low amounts of root infestation by plant parasitic nematodes are suggested to increase nutrient supply and in turn enhance microbial activity and net mineralization rate in the rhizosphere. These effects are generally related to “leakage” of plant-derived metabolites from damaged roots. Besides leakage, the present study examines other nematode–host interactions such as alterations in root exudation and morphology, which were almost not considered yet. This includes undamaged root parts in order to assess systemic plant response. The root-knot nematode Meloidogyne incognita (Kofoid and White 1919; Chitwood 1949) and barley (Hordeum vulgare L. cv. Europa) was used as model system. Host plants were grown in mini-rhizotrons inoculated with 0, 2,000, 4,000 or 8,000 M. incognita for 4 weeks. Root morphology, rhizodeposition (sugars, carboxylates, amino acids), and rhizosphere microbial communities (PLFAs) were assessed. In treatments with 4,000 nematodes, shoot biomass, total N and P content increased by the end of the experiment. Generally, an enhanced release of plant metabolites (sugars, carboxylates, amino acids) from the apical root zone occurred 1 week after inoculation with 4,000 and 8,000 M. incognita, indicating root leakage. Low levels of root herbivory stimulated root hair elongation in both infected and uninfected roots. These systemic changes in root morphology likely contributed to the increased sugar exudation in uninfected roots in all nematode treatments at 3 weeks after inoculation. Root-knots formed a separate microhabitat within the root-system. They were characterised by decreased rhizodeposition and increased fungal to bacterial ratio in the adhering rhizosphere soil. The present study provides the first evidence that, apart from leakage, nematode root herbivory at background levels induces local and systemic effects on root morphology and exudation, which in turn may affect plant performance.     

Inheritance of Resistance to Pratylenchus penetrans in Alfalfa

Fifty-two alfalfa (Medicago sativa L.) clones, randomly selected from the cultivar Baker and the experimental line MNGRN-4, were evaluated for resistance (based on nematode reproduction) to Pratylenchus penetrans in growth chamber tests (25 C). Twenty-five clones, representing the range of nematodes and eggs per plant, were selected and retested. Four moderately resistant and two susceptible alfalfa clones were identified. Inheritance of resistance to P. penetrans was studied in these six clones using a diallel mating design. The S₁, Fl, and reciprocal progenies differed for numbers of nematodes and eggs per g dry root and for shoot and root weights (P < 0.05). Resistance, measured as numbers of nematodes in roots, was correlated between parental clones and their S₁ families (r = 0.94), parental clones and their half-sib families (r = 0.81), and S₁ and half-sib families (r = 0.88). General combining ability (GCA) effects were significant for nematode resistance traits. Both GCA and specific combining ability (SCA) effects were significant for plant size traits, but SCA was more important than GCA in predicting progeny plant size. Reciprocal effects were significant for both nematode resistance and plant size traits, which may slow selection progress in long-term selection programs. However, the GCA effects are large enough that breeding procedures that capitalize on additive effects should be effective in developing alfalfa cultivars with resistance to P. penetrans.     

Techniques for image analysis of movement of juveniles of root-knot nematodes encumbered with Pasteuria penetrans spores

Root-knot nematodes (Meloidogyne spp.) are the most significant plant-parasitic nematodes that damage many crops all over the world. The free-living second stage juvenile (J2) is the infective stage that enters plants. The J2s move in the soil water films to reach the root zone. The bacterium Pasteuria penetrans is an obligate parasite of root-knot nematodes, is cosmopolitan, frequently encountered in many climates and environmental conditions and is considered promising for the control of Meloidogyne spp. The infection potential of P. penetrans to nematodes is well studied but not the attachment effects on the movement of root-knot nematode juveniles, image analysis techniques were used to characterize movement of individual juveniles with or without P. penetrans spores attached to their cuticles. Methods include the study of nematode locomotion based on (a) the centroid body point, (b) shape analysis and (c) image stack analysis. All methods proved that individual J2s without P. penetrans spores attached have a sinusoidal forward movement compared with those encumbered with spores. From these separate analytical studies of encumbered and unencumbered nematodes, it was possible to demonstrate how the presence of P. penetrans spores on a nematode body disrupted the normal movement of the nematode.     

Interactions of Pratylenchus penetrans and Meloidogyne incognita as Coinhabitants in Tomato

Greenhouse experiments on the interactions of Pratylenchus penetrans and Meloidogyne incognita showed that the population densities of both nematode species were depressed whenthey coinhabited tomato roots. Fifty days after inoculation, the population level of a P. penetrans monoculture was about four times higher than when M. incognita was present. Conversely, M. incognita reproduced twice as fast alone as in combination. There were no significant differences in the numbers of P. penetrans when they were inoculated either 10 days prior to or after introduction of M. incognita. Root entry by P. penetrans was significantly inhibited by the presence of M. incognita. Split-root experiments showed that the inhibitory effects of M. incognita upon reproduction of P. penetrans involved factors other than the availability of feeding sites. On the other hand, the inhibitory effects of P. penetrans on M. incognita appeared to be primarily due to the quantity of available roots.