The combined effect of the application of a biocontrol agent Paecilomyces lilacinus, with various practices for the control of root-knot nematodes

The effectiveness of a formulated product containing spores of the naturally occurring fungus Paecilomyces lilacinus, strain 251, was evaluated against root-knot nematodes in pot and greenhouse experiments. Decrease of second-stage juveniles hatching from eggs was recorded by using the bio-nematicide at a dose of 4 kg ha(-1), while a further decrease was recorded by doubling the dose. However, the mortality rate decreased by increasing the inoculum level. Application of P. lilacinus and Bacillus firmus, singly or together in pot experiments, provided effective control of second-stage juveniles, eggs or egg masses of root-knot nematodes. In a greenhouse experiment, the bio-nematicide was evaluated for its potential to control root-knot nematodes either as a stand-alone method or in combination with soil solarization. Soil was solarized for 15 d and the bio-nematicide was applied just after the removal of the plastic sheet. Soil solarization for 15 d either alone or combined with the use of P. lilacinus did not provide satisfactory control of root-knot nematodes. The use of oxamyl, which was applied 2 weeks before and during transplanting, gave results similar to the commercial product containing P. lilacinus but superior to soil solarization. (C) 2007 Elsevier Ltd. All rights reserved.

Effects of a non-chemical nematicide combined with soil solarization for the control of root-knot nematodes

The effectiveness of a formulated bio-nematicide product containing lyophilized bacteria spores of Bacillus firmus was evaluated against root-knot nematodes (RKN) in greenhouse and field experiments. A decrease of second stage juveniles hatching from eggs was recorded by using the bio-nematicide at a dose of 0.9 g kg(-1) of soil while further a decrease was recorded by doubling the dose. However, the mortality rate decreased as the inoculurn level increased. Exposure of either second stage juveniles or egg masses to temperatures of 35-40 degrees C for 1-4 weeks had a marked effect on their survival. In a field experiment, the bio-nematicide was evaluated for its potential to control RKN either as a stand-alone method or in combination with soil solarization. The latter was tested for 15-30 days and the bionematicide was applied just before soil coverage with the plastic sheet or just after its removal. Soil solarization either for 15-30 days provided satisfactory control of RKN. The combination of soil solarization with the bio-nematicide improved nematode control and gave results similar to the chemical treatment. (c) 2007 Elsevier Ltd. All rights reserved.

The potential of combining Pasteuria penetrans and neem (Azadirachta indica) formulations as a management system for root-knot nematodes on tomato

Initial applications of 10(4) spores g(-1) of Pasteuria penetrans, and dried neem cake and leaves at 3 and 2% w:w, respectively, were applied to soil in pots. Juveniles of Meloidogyne javanica were added immediately to the pots (500, 5,000 or 10,000) before planting 6-week-old tomato seedlings. The tomatoes were sampled after 64 days; subsequently a second crop was grown for 59 days and a third crop for 67 days without further applications of P. penetrans and neem. There was significantly less root-galling in the P. penetrans combined with neem cake treatment at the end of the third crop and this treatment also had the greatest effect on the growth of the tomato plants. At the end of the third crop, 30% of the females were infected with P. penetrans in those treatments where spores had been applied at the start of the experiment. The effects of neem leaves and neem cake on the nematode population did not persist through the crop sequences but the potential for combining the amendments with a biological control agent such as P. penetrans is worthy of further evaluation.

Efficacy of neem (Azadirachta indica) formulations on biology of root-knot nematodes (Meloidogyne javanica) on tomato

Second stage juveniles of Meloidogyne javanica were exposed to aqueous extracts of neem crude formulations (leaves and cake) at 10%, 5%, and 2.5% w/v and a refined product, Aza at 0.1% w/v. The 10% extracts of neem leaf and cake caused 83% and 85% immobility and 35% and 28% mortality, respectively. Aza caused neither immobility or mortality of juveniles. When egg masses were placed in extracts of these formulations, hatching did not occur at all the concentrations (10%, 5%, 2.5% and 1.25% w/v) of the crude formulations. When the treated egg masses were returned to water, the eggs resumed hatching. Aza did not affect the nematode hatching. In glasshouse experiments, soil application of neem formulations significantly reduced the invasion of tomato roots by root-knot nematodes but once the nematodes managed to invade them, no effect detected on their development. Soil applications of Aza at 0.05% and 0.1% w/v significantly reduced the invasion and delayed development of nematodes within tomato roots whereas 0.025% did not. There were significantly fewer egg masses on tomato roots exposed to single egg mass in neem amended soil as compared to control. (C) 2007 Elsevier Ltd. All rights reserved.

Systemic and persistent effect of neem (Azadirachta indica) formulations against root-knot nematodes, Meloidogyne javanica and their storage life

Neem leaves, neem cake (a by-product left after the extraction of oil from neem seed) and a commercially refined product aza (azadirachtin) extracted from seed were evaluated. Aqueous extracts of crude neem formulations used as a seedling dip treatment significantly reduced the number of females and egg masses in roots whereas the refined one did not. A split-root technique was used to demonstrate the translocation of active compounds within a plant and their subsequent effect on the development of nematodes. When applied to the root portion all formulations significantly reduced the number of egg masses and eggs per egg mass. Whereas on the untreated root portion, neem cake at 3% w/w and aza at 0.1% w/w significantly reduced the number of egg masses as compared with neem leaves at 3% w/w, aza at 0.05% and control. All the neern formulations significantly reduced the number of eggs per egg mass on' the untreated root portion. The effect of neem leaves and cake on the development of root-knot nematodes was tested at 2, 4, 6, 8, and 16 weeks after their application to soil. Even after 16 weeks all the treatments significantly reduced the galling index and number of egg masses but their effectiveness declined over time. After storing neem leaves, cake and aza for 8 months under ambient conditions the efficacy of neem leaves and aza, against root-knot nematodes, remained stable whereas that of cake declined. (c) 2006 Elsevier Ltd. All rights reserved.

The control of root-knot nematodes (Meloidogyne spp.) by Pseudomonas oryzihabitans and its immunological detection on tomato roots

Pseudomonas oryzihabitans, a bacterium associated with the entomopathogenic nematode Steinernema abbasi, was evaluated for its potential to colonise roots and thereby control a field population of root-knot nematodes. Immunological techniques were developed to detect root colonisation of P. oryzihabitans on tomato roots using a specific polyclonal antibody raised against vegetative bacterial cells. In vitro, bacterial cell filtrates were also shown significantly to inhibit juveniles hatching. In a glasshouse pot experiment, there were 22 and 82% fewer females in roots of plants treated with suspensions containing 10(3) and 10(6) cells ml(-1) of P oryzihabitans, respectively. In addition, there were significantly fewer egg masses produced; however, the numbers of eggs per egg mass did not differ significantly. The relationship between root colonisation and nematode control is discussed.

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.

Morphological and molecular characteristics of a new species of Pasteuria parasitic on Meloidogyne ardenensis

A species of the hyper-parasitic bacterium Pasteuria was isolated from the root-knot nematode Meloidogyne ardenensis infecting the roots of ash (Fraxinus excelsior). It is morphologically different from some other Pasteuria pathogens of nematodes in that the spores lack a basal ring on the ventral side of the spore and have a unique clumping nature. Transmission electron microscopy (TEM) showed that the clumps of spores are not random aggregates but result from the disintegration of the suicide cells of the thalli. Sporulation within each vegetative mycelium was shown to be asynchronous. In addition to the novel morphological features 16S rRNA sequence analysis showed this to be a new species of Pasteuria which we have called P. hartismeri. Spores of P. hartismeri attach to juveniles of root-knot nematodes infecting a wide range of plants such as mint (Meloidogyne hapla), rye grass (unidentified Meloidogyne sp.) and potato (Meloidogyne fallax). (c) 2007 Elsevier Inc. All rights reserved.

Factors affecting biological control effectiveness of Pasteuria penetrans in Meloidogyne javanica and the bacterial development in the nematode body

The invasion and infectivity of Meloidogyne javanica juveniles (J2) encumbered with spore of Pasteuria Penetrans were influenced by the temperature and the time J2 were in the soil before exposure to roots. The percentage of infected females decreased as the time juveniles spent in soil increased. When spore encumbered J2 were maintained at 30 degrees C the decrease in infection was greater than that at 18 degrees C. The thermal time requirements and the base temperature for P. penetrans development were estimated. The rate of development followed an exponential curve between 21 and 36 degrees C and the base temperature for development was estimated by extrapolation to be 18.5 degrees C. The effect of integrating a nematode resistant tomato cultivar with the biocontrol agent P. penetrans also was investigated. The ability of the biocontrol agent to reduce numbers of root-knot nematodes was dependent on the densities of the nematode and P. penetrans spores in the soil.

Genotypic differences in the growth of bananas (Musa spp.) infected with migratory endoparasitic nematodes. 1. Roots

The effects of nematodes on root morphology and the association of root characteristics with resistance to nematodes of seven banana varieties were investigated in two experiments. Banana plants were grown in controlled conditions within polytunnels and harvested on three occasions for the measurement of root morpholopy, and biomass. Varieties differed in their resistance to nematodes, from resistant (Yg Km5, FHIA 17, FHIA 03) and partly resistant (FHIA 01, FHIA 25) to not resistant ((FHIA 23, Williams). Nematodes reduced the root dry weight of FHIA 01, FHIA 17 and FHIA 23 at some harvests. Primary root number was on average 9.5% lower in nematode-infected plants than controls, with no differences among the varieties. Thus, there was no simple association between the resistance of these varieties and their tolerance to nematodes. Varieties differed in root morphology. Root dry weight was greatest for resistant varieties Yg Km5 and FHIA 03, and least for non-resistant varieties FHIA 23 and Williams. Thus, resistance to nematodes was associated with varieties with greater root mass and more and larger primary roots.

Genotypic differences in the growth of bananas (Musa spp.) infected with migratory endoparasitic nematodes. 2. Shoots

In order to identify the effect of burrowing nematodes on the shoots (pseudostem and leaves) of banana plants and to determine whether or not shoot characteristics are associated with plant resistance to nematodes two experiments were conducted in controlled conditions within polytunnels. The banana plants were harvested on three occasions for the measurement of root morphology and biomass. Varieties differed in their resistance to nematodes from resistant (Yg Km5, FHIA 17, FHIA 03) and partly resistant (FHIA 01, FHIA 25) to not resistant (FHIA 23, Williams). Nematodes reduced total plant dry weight at the first harvest in Experiment 1 and by an average of 8.8% in Experiment 2, but did not affect leaf area in either experiment. The ratio of above-ground Weight to total plant weight was reduced from 75% to 72% in nematode-infected plants compared with the control plants for all varieties tested in Experiment 1, but was only reduced in FHIA 25 and FHIA 23 in Experiment 2. Varieties differed in above-ground growth. The FHIA varieties had greater shoot weights and leaf area than YgKm5 and Williams. Overall, resistance to nematodes was associated with the partitioning of a greater proportion of biomass to the roots than to above-ground parts.

Using historical lesion volume data in the design of a new phase II clinical trial in acute stroke

Background and Purpose-Clinical research into the treatment of acute stroke is complicated, is costly, and has often been unsuccessful. Developments in imaging technology based on computed tomography and magnetic resonance imaging scans offer opportunities for screening experimental therapies during phase II testing so as to deliver only the most promising interventions to phase III. We discuss the design and the appropriate sample size for phase II studies in stroke based on lesion volume. Methods-Determination of the relation between analyses of lesion volumes and of neurologic outcomes is illustrated using data from placebo trial patients from the Virtual International Stroke Trials Archive. The size of an effect on lesion volume that would lead to a clinically relevant treatment effect in terms of a measure, such as modified Rankin score (mRS), is found. The sample size to detect that magnitude of effect on lesion volume is then calculated. Simulation is used to evaluate different criteria for proceeding from phase II to phase III. Results-The odds ratios for mRS correspond roughly to the square root of odds ratios for lesion volume, implying that for equivalent power specifications, sample sizes based on lesion volumes should be about one fourth of those based on mRS. Relaxation of power requirements, appropriate for phase II, lead to further sample size reductions. For example, a phase III trial comparing a novel treatment with placebo with a total sample size of 1518 patients might be motivated from a phase II trial of 126 patients comparing the same 2 treatment arms. Discussion-Definitive phase III trials in stroke should aim to demonstrate significant effects of treatment on clinical outcomes. However, more direct outcomes such as lesion volume can be useful in phase II for determining whether such phase III trials should be undertaken in the first place. (Stroke. 2009;40:1347-1352.)