Nematodes associated with terrestrial gastropod molluscs in Belgium and additional characterisation of Pellioditis californica and P. hermaphrodita.

A survey for slug- and snail-associated nematodes was conducted in forests, parks, botanical gardens, and nature reserves at 13 localities in Belgium to uncover more diversity of gastropod mollusc-associated nematodes and to characterise Pellioditis populations found in the country. A total of 319 slugs and snails belonging to nine species were examined. Arion vulgaris was the most commonly found mollusc species in this study (eight locations), and 19.4% of the examined mollusc specimens were found infected by nematodes. The highest prevalence of nematodes was observed in Cornu aspersum (60%) followed by A. vulgaris (34.8%), Limax maximus (28.6%), and Cepaea sp. (20%). Eleven nematode species belonging to eight families were isolated and identified from the mollusc hosts including Alloionema appendiculatum, Angiostoma dentiferum, A. gandavense, Angiostrongylus vasorum, Cosmocerca longicauda, Panagrolaimus cf. subelongatus, Pellioditis californica, P. hermaphrodita, Rhabditis sp., Tetrameres cf. fissispina, and Troglostrongylus cf. brevior.Pellioditis was the most commonly found nematode genus (at nine localities) and C. longicauda and P. californica were reported in Belgium for the first time. Co-infections of more than one nematode species were observed in eight (2.5%) molluscs specimens. Most co-infections consisted of two nematode species. In one A. vulgaris specimen, a co-infection of three nematode species (A. vasorum, P. hermaphrodita, and Tetrameres cf. fissispina) was observed. Four ex vivo cultures of P. californica and six ex vivo cultures of P. hermaphrodita were established from single hermaphrodites, and both species were described based on light microscopy, scanning electron microscopy, and morphometric, morphological, and molecular data.

Interactions between the oomycete Pythium arrhenomanes and the rice root-knot nematode Meloidogyne graminicola in aerobic Asian rice varieties.

BACKGROUND: Aerobic rice fields are frequently infested by pathogenic oomycetes (Pythium spp.) and the rice root-knot nematode Meloidogyne graminicola. Here, the interaction between Pythium arrhenomanes and Meloidogyne graminicola was studied in rice roots of two aerobic rice varieties. In different experimental set-ups and infection regimes, plant growth, rice yield, Pythium colonization, as well as establishment, development and reproduction of M. graminicola were studied. RESULTS: In this study, it is shown that the presence of P. arrhenomanes delays the establishment, development and reproduction of M. graminicola compared to single nematode infected plants. The delay in establishment and development of M. graminicola becomes stronger with higher P. arrhenomanes infection pressure. CONCLUSIONS: Our data indicate that P. arrhenomanes antagonizes M. graminicola in the rice root and that the plant benefits from this antagonism as shown by the yield data, especially when either of the pathogens is present in high levels.

Management of root knot nematode, Meloidogyne incognita in cucumber (Cucumis sativus) using silicon.

Silicon (Si) has been reported to effectively manage some pests and diseases of plants. This study was conducted to determine the effect of Si concentration, mode, and frequency of application in managing Meloidogyne incognita in cucumber. A susceptible cultivar of cucumber (cv. Cyclone) was planted in pots containing heat-sterilized soil. Three weeks after planting, the plants were inoculated with 1,000 juveniles/ pot. Uninoculated plants were provided to serve as control. Three concentrations of Si in the form of sodium metasilicate was applied on the leaves and roots alone and also on both the leaves and roots. Application was done once during the growing period and weekly until seven days before harvest. Leaf and root application of Si was found to significantly increase (p = 0.0029) the fresh top weight of inoculated and uninoculated plants. On the other hand, inoculation of root-knot nematode significantly increased the fresh root weight of cucumber which could be due to enlargement of roots or formation of galls. Interestingly, the inoculated plants gave significantly higher marketable yield than uninoculated ones. Application of Si at the rate of 200 ppm significantly increased the marketable yield compared to the higher rate of Si (400 ppm). At 200 ppm, one application of Si both on the leaves and roots significantly reduced the number of galls in inoculated plants. This was comparable to the same concentration applied continuously on the roots and at higher concentration (400 ppm) applied continuously on the leaves and on the roots. On the other hand, single root application of Si at the rate of 400 ppm gave the lowest number of eggmasses, however, it was comparable to the same Si concentration applied singly on the leaves and applied continuously both on the leaves and roots. These treatments, however, were at par with continuous application of the lower rate of Si (200 ppm) on the leaves and both leaves and roots.

AMF-induced biocontrol against plant parasitic nematodes in Musa sp.: a systemic effect.

Although mycorrhizal colonization provides a bioprotectional effect against a broad range of soil-borne pathogens, including plant parasitic nematodes, the commercial use of arbuscular mycorrhizal fungi (AMF) as biocontrol agents is still in its infancy. One of the main reasons is the poor understanding of the modes of action. Most AMF mode of action studies focused on AMF-bacterial/fungal pathogens. Only few studies so far examined AMF-plant parasitic nematode interactions. Therefore, the aim of the study was to determine whether the AMF Glomus intraradices was able to incite systemic resistance in banana plants towards Radopholus similis and Pratylenchus coffeae, two plant parasitic nematodes using a split-root compartmental set-up. The AMF reduced both nematode species by more than 50%, even when the AMF and the plant parasitic nematodes were spatially separated. The results obtained demonstrate for the first time that AMF have the ability to induce systemic resistance against plant parasitic nematodes in a root system.

Nematode infection and reproduction in transgenic and mutant Arabidopsis and tobacco with an altered phenylpropanoid metabolism.

Transgenic and mutant Arabidopsis and tobacco plants with altered phenylpropanoid metabolism were infected with the plant parasitic root knot nematode Meloidogyne incognita to assess the effect of the transgene or mutation on nematode infection and reproduction. Modifications in the lignin biosynthetic pathway which alter lignin composition in roots affected reproduction. In Arabidopsis with increased levels of syringyl lignin, reproduction was lower than in wild-type plants, while in tobacco with reduced levels of syringyl lignin, life cycle progression was stimulated. Overexpression of a MYB transcription factor of phenylpropanoid metabolism in tobacco significantly stimulated reproduction of M. incognita, while overexpression of L-phenylalanine ammonia-lyase had no effect. Arabidopsis transparent testa mutants with deficiencies in flavonoid pathway enzymes did not affect reproduction of M. incognita in the present infection tests.

Aseptic Culture Systems of Radopholus similis for In Vitro Assays on Musa spp. and Arabidopsis thaliana.

Radopholus similis is one of the most damaging nematodes in bananas. Chemical control is currently the most-used method, but nematode control through genetic improvement is widely encouraged. The objective of this study was to establish an aseptic culture system for R. similis and determine whether R. similis can infect and reproduce on in vitro banana plantlets and in vitro Arabidopsis thaliana. In the study's first part, a suitable aseptic culture system was developed using alfalfa callus. Radopholus similis could penetrate and reproduce in the callus. Six weeks after inoculation with 25 females, the reproduction ratio was 26.3 and all vermiform stages were present. The reproduction ratio increased to 223.2 after 12 weeks. Results of a greenhouse test showed that R. similis did not lose its pathogenicity after culturing on alfalfa callus. In the study's second part, the infection and reproduction of the nematodes cultured on the callus were studied on both in vitro banana plantlets and A. thaliana. Radopholus similis infected and reproduced on both banana and A. thaliana. Furthermore, nematode damage was observed in the root systems of both hosts. These successful infections open new perspectives for rapid in vitro screening for resistance in banana cultivars and anti-nematode proteins expressed in A. thaliana.

Germination of Bacillus thuringiensis spores in bacteriophagous nematodes (Nematoda: Rhabditida).

During screening of Bacillus thuringiensis isolates for nematicidal activity it was observed that spores of B. thuringiensis germinated in the intestine of bacteriophagous nematodes in the presence of antibiotics. This phenomenon was studied more closely by scanning electron microscopy. The nematodes were fed with bacterial spore-crystal mixtures in axenic culture medium supplemented with tetracyclin and chloramphenicol. Germination of spores was rare but was more frequently observed in Panagrellus redivivus than in other nematode species investigated. Germination of spores in the nematode intestine resulted in the colonization of the entire nematode within 24 hr. Crude nematode tissue preparations supported germination and subsequent growth of B. thuringiensis spores and vegetative cells. The mechanism for the loss of antibiotic activity in the nematode intestine is unknown. Since B. thuringiensis requires a nutrient-rich environment for reproduction, e.g., a cadaver, bacteriophagous nematodes may serve as suitable hosts for B. thuringiensis.

Histological Studies of Ditylenchus africanus Within Peanut Pods.

Ditylenchus africanus entered the immature pegs and pods of peanut (Arachis hypogaea cv. Sellie) at the peg-connection and subsequently invaded the parenchymatous regions of the hull exocarp and endocarp, and eventually the seed testa. The nematode caused malformations of the cells of infected tissues, cell wall breakage, and cell collapse. The damage appeared to be due to enzymatic activity. In some testae the entire parenchyma region, which aids in protection of the seed, was destroyed. In immature pods, the nematodes moved across the fibrous region of the mesocarp into the hull endocarp. In mature pods, however, the fibrous mesocarp of the hull was lignified and apparently was a barrier to penetration of the inner pod tissues. In late-harvested pods, increased numbers of eggs and anhydrobiotes were found in the hull tissues, and eggs in the seed testa, suggesting the onset of winter survival mechanisms of the nematode.

Tissue treatment for whole mount internal lectin staining in the nematodes Caenorhabditis elegans, Panagrolaimus superbus and Acrobeloides maximus.

Four different fixation schemes, using ten fluorescent-labelled lectins, were investigated for whole mount internal staining of three rhabditid nematodes: Caenorhabditis elegans, Panagrolaimus superbus and Acrobeloides maximus. Acetone-only fixation was found to give strong and reproducible staining, which could be prevented either by periodate treatment of the organisms or by specific inhibitory sugars of the lectins under investigation. Whereas the use of either phosphate or TRIS buffers had no effect on the staining pattern or the fluorescence intensity, the incubation time as well as the incubation temperature affected the staining reaction. The best results were obtained upon overnight incubation at 4 degrees C: the lectin staining could be inhibited in all cases, except for the intestinal brush border of C. elegans by the lectin of Lens culinaris.

Reproductive and Damage Potential of Ditylenchus destructor on Six Peanut Cultivars.

Commercial peanut cultivars were evaluated for host suitability and sensitivity to Ditylenchus destructor. All cultivars were susceptible. Approximately 94% of the final population were in the pods. Highest Pf occurred at harvest on early maturing cultivars. Damage occurred on four of six cuttivars at Pi = 100/3 liters of soil and all six cultivars at Pi = 1,000. Norden and Selmani were the most susceptible cultivars. Sellie was the most tolerant and highest yielding cultivar. This cultivar may be the most profitable one for growers.

Effect of Host Plant Age on Population Development and Pathogenicity of Ditylenchus destructor on Peanut.

The effect of inoculating peanut, Arachis hypogaea cv. Sellie, with Ditylenchus destructor at timed intervals after planting and with different initial nematode population densities (Pi) was tested in greenhouse experiments. Final nematode population densities (Pf) in hulls and seeds were greater (Pf < 0.001) in plants inoculated at or before 9 weeks after planting. Pod disease symptoms correlated positively with the Pf in the pods. The seedgrade of peanuts inoculated at or before 9 weeks after planting was reduced, whereas grade of peanuts from plants inoculated at 15 weeks or later was not reduced. Peanut plants inoculated 12 weeks after planting with a Pi of 10-100 had a lower Pf (P < 0.05) than plants with a Pi of 250 to 8,000. Seed of plants with a Pi of 250 or less could be marketed as choice edible seed, whereas those with a Pi of 500 or more were of reduced seedgrade. These results suggest that as few as 500 nematodes per plant at 12 weeks after planting can build up to injurious levels before harvest. A nematicide should therefore be active for longer than 12 weeks after planting to sufficiently suppress the population.

Minimizing damage by Ditylenchus destructor to Peanut Seed with Early Harvest.

Greenhouse and microplot experiments were conducted to evaluate the damage potential of Ditylenchus destructor on four South African commercial peanut cultivars as influenced by harvest date. The cultivars Sellie and Harts should be harvested by 150 and 120 days after planting, respectively. Losses were 12-13% with early harvest, but a 15-day delay resulted in losses of 45-49%. Harvest of Natal Common and Norden at 125 and 145 days after planting, respectively, resulted in the highest seed grade. By normal harvest time (140 and 160 days, respectively) these two cultivars were downgraded to crushing seed quality. Even though seed weight increases with time, a net loss occurs if harvest is delayed.

Reproductive and Damage Potential of Ditylenchus destructor on Peanut.

The reproductive and damage potential of Ditylenchus destructor on peanut, Arachis hypogaea cv. Sellie, was determined in greenhouse tests. Final nematode population densities (Pf) in roots, hulls, and seeds increased (P = 0.01) as a function of increasing initial population (Pi). Final population densities were higher in hulls than in seeds and roots. Final densities in hulls and seeds were positively (P = 0.01) correlated. Fresh root and hull weight and number of pods and seeds per plant were not affected by D. destructor. Second generation germination and pod and seed disease severity increased (P = 0.01), whereas fresh seed weight decreased (P = 0.01) as a function of increasing Pi, and Pf in seeds and Pf in hulls. At Pi 250 and higher, 10-25% of seeds germinated into second generation seedlings before harvest. At Pi 250 and higher, fresh weight of harvested seed was suppressed 20-50%. At Pi 50 or Pf greater than 20 per seed, pod disease severity was 3-7 (on a scale of 1 to 10) and 15-80% of seeds were blemished or unsound.