The effect of different growing substrates on the development and quality of Phasmarhabditis hermaphrodita (Nematoda: Rhabditidae)

The effect of different growing substrates on the development of the slug parasite Phasmarhabditis hermaphrodita has been studied in a series of laboratory experiments. Wild, laboratory and Nemaslug strains of P. hermaphrodita and their monoxenic counterparts with Moraxella osloensis (CCM 5605T) were reared on homogenised pig kidney, Deroceras reticulatum, Arion lusitanicus, and Galleria mellonella, the faeces of D. reticulatum and A. lusitanicus, or leaf compost. Development time, yield, lipid reserves and the body length of nematodes were assessed. All P. hermaphrodita strains were able to grow and reproduce on all tested substrates; however, yields were markedly higher on invertebrate media. Lipid content and body size varied across the substrates and strains. A uniform response of all nematode strains and the most dramatic differences among substrates were recorded in yields, which could indicate that the quality of the substrate is expressed mainly in yield. Monoxenized strains had higher yield, length, lipid content and shorter development time. In general, the strain differences between the observed parameters were most likely due to different bacterial associates. The dramatic differences in yields on invertebrate substrates, in comparison to the others, illustrate the evolutionary advantage of the association of nematodes with invertebrates.     

Biological control of slugs in winter wheat using the rhabditid nematode Phasmarhabditis hermaphrodita

A field experiment on winter wheat in autumn 1991 investigated the effect of the rhabditid nematode, Phasmarhabditis hermaphrodita, applied to soil at five dose rates (10⁸ - 10¹⁰ infective larvae ha^-1) immediately after seed sowing, on slug populations and damage to seeds and seedlings. The nematode was compared with methiocarb pellets broadcast at recommended field rate immediately after drilling and no molluscicide treatment. Slug damage to wheat seeds and seedlings was assessed 6 and 13 wk after drilling. Seedling survival increased and slug grazing damage to seedlings declined linearly with increasing log nematode dose. These two measures of slug damage were combined to give an index of undamaged plant equivalents, which also increased linearly with increasing log nematode dose. ANOVA showed that, after 6 wk, there were significantly more undamaged plant equivalents on plots treated with the two highest nematode doses (3 × 10⁹ and 1 × 10¹⁰ ha^-1) than on untreated plots, but the number of undamaged plant equivalents on methiocarb-treated plots was not significantly greater than that on untreated plots. Slug populations were assessed by refuge trapping and soil sampling. Deroceras reticulatum was the commonest of several species of slugs recorded. During the first 4 wk after sowing, significantly more slugs were found under refuge traps on plots treated with certain doses of P. hermaphrodita than under traps on untreated plots and more showed signs of nematode infection than expected from the prevalence of infection in slugs from soil samples, suggesting that the presence of P. hermaphrodita altered slug behaviour. Application of P. hermaphrodita had no significant impact on numbers or biomass of slugs in soil during a 27 wk period after treatment, except after 5 wk when slug numbers were inversely related to log nematode dose. However, by this time, numbers in soil samples from untreated plots had declined to levels similar to those in plots treated with the highest dose of nematodes. During the first 5 wk after treatment, c. 20% of slugs in soil samples from untreated plots showed symptoms of nematode infection. It is suggested that this represented the background level of infection in the experimental field rather than spread of infection from treated plots. The apparent lack of impact of P. hermaphrodita on slug numbers and biomass in soil suggests that its efficacy in protecting wheat from slug damage was through inhibition of feeding by infected slugs.     

Susceptibility of neonate Lissachatina fulica (Achatinidae: Mollusca) to a US strain of the nematode Phasmarhabditis hermaphrodita (Rhabditidae: Nematoda)

ABSTRACT

The nematode Phasmarhabditis hermaphrodita is a lethal parasite of many pest slug and snail species. It is used as a commercial biological control agent (Nemaslug®) in Europe but a number of key gastropod pests (e.g. Lissachatina fulica) are thought to be unsusceptible. Our data, however, demonstrated that a recently discovered U.S. strain of P. hermaphrodita causes statistically significant mortality to neonate L. fulica and consequently this nematode could be an important IPM tool for managing this snail pest in areas where P. hermaphrodita has been discovered. Soil moisture content appears to play an important role in determining nematode efficacy.     

Pathogenicity of the rice root nematode,Hirschmanniella oryzae to rice plants in relation to nematode reproduction,plant growth,grain yield and biochemical changes

Influence of different inoculum levels of 0, 10, 100, 1000 and 10,000 individuals of Hirschmanniella oryzae on nematode reproduction and plant growth of rice cv. Giza171 and biochemical changes of infected plants was studied under screen-house conditions. Rate of nematode build up (P_f/P_i) was negatively correlated with the progressive increase in nematode inoculum levels. The percentage reduction in growth parameters, rice grain yield and the amount of total and reducing sugars were markedly affected showing a negative correlation with the tested inocula. The conspicuous reductions of plant growth, yield and total and reducing sugar contents were obtained by using 1000 and 10,000 nematodes per pot. The inoculum level of 1000 nematodes per pot was identified as critical population at which control programme must be started.     

Size-specific susceptibility of the pest slugs Deroceras reticulatum and Arion lusitanicus to the nematode biocontrol agent Phasmarhabditis hermaphrodita

The nematode Phasmarhabditis hermaphrodita is a commercially available biocontrol agent against slugs. This product is especially interesting for use in organic farming, where products containing metaldehyde or carbamates cannot be used for controlling pest slugs. We investigated the potential of P. hermaphrodita for the control of the pest slugs Deroceras reticulatum and Arion lusitanicus. These two species are the most harmful slug pests in Switzerland. At different times of the year, we collected slug specimens of different weight and assessed their susceptibility to P. hermaphrodita in the laboratory. Batches of five slugs were subjected to five different doses of nematodes plus an untreated control and replicated three times. During six weeks, feeding and survival of the slugs were recorded. D. reticulatum was strongly affected by increasing nematode doses, irrespective of the slugs' body weight. In small specimens of A. lusitanicus, feeding and survival were strongly affected by the nematodes, while larger specimens remained almost unaffected. Because A. lusitanicus has an asynchronous development in Switzerland, it seems difficult to control the entire population with a single nematode application. To what extent nematodes will be used in practice for slug control depends on their effectivity against the pest slugs of major importance, on the longevity of the molluscicidal effect and on the price of nematodes.     

Pathogenicity of root-knot nematode,Meloidogyne incognita on Lens culinaris (Medik.)

Effects of the root-knot nematode (Meloidogyne incognita) on lentil (Lens culinaris) were studied under greenhouse conditions. The plants were inoculated with 250, 500, 1000, 2000 and 4000 J₂ per plant. Plant growth, yield, nodulation, seed weight, chlorophyll, nitrogen, phosphorus and potassium, (NPK) contents, as compared to control, were found decreased in all the nematode infected plants. The extent of reduction increased with an increase in inoculum levels. The reductions were significant at 500 J₂ and at higher inoculum levels, i.e. 1000, 2000 and 4000 J₂ per pot over the control. An increase in inoculum level caused enhancement in galling, egg mass production and nematode population. At higher inoculum levels, the population of the nematode in the root as well as in the soil increased to a greater magnitude than at lower inoculum levels. On the contrary, reproduction factor (RF) and rate of population increase (RPI) decreased with increasing inoculum levels.     

Pathogenicity of Moraxella osloensis, a Bacterium Associated with the Nematode Phasmarhabditis hermaphrodita, to the Slug Deroceras reticulatum

Moraxella osloensis, a gram-negative bacterium, is associated with Phasmarhabditis hermaphrodita, a nematode parasite of slugs. This bacterium-feeding nematode has potential for the biological control of slugs, especially the grey garden slug, Deroceras reticulatum. Infective juveniles of P. hermaphrodita invade the shell cavity of the slug, develop into self-fertilizing hermaphrodites, and produce progeny, resulting in host death. However, the role of the associated bacterium in the pathogenicity of the nematode to the slug is unknown. We discovered that M. osloensis alone is pathogenic to D. reticulatum after injection into the shell cavity or hemocoel of the slug. The bacteria from 60-h cultures were more pathogenic than the bacteria from 40-h cultures, as indicated by the higher and more rapid mortality of the slugs injected with the former. Coinjection of penicillin and streptomycin with the 60-h bacterial culture reduced its pathogenicity to the slug. Further work suggested that the reduction and loss of pathogenicity of the aged infective juveniles of P. hermaphrodita to D. reticulatum result from the loss of M. osloensis from the aged nematodes. Also, axenic J1/J2 nematodes were nonpathogenic after injection into the shell cavity. Therefore, we conclude that the bacterium is the sole killing agent of D. reticulatum in the nematode-bacterium complex and that P. hermaphrodita acts only as a vector to transport the bacterium into the shell cavity of the slug. The identification of the toxic metabolites produced by M. osloensis is being pursued.     

Influence of inoculum density on population dynamics and dauer juvenile yields in liquid culture of biocontrol nematodes Steinernema carpocapsae and S. feltiae (Nematoda: Rhabditida)

For improvement of mass production of the rhabditid biocontrol nematodes Steinernema carpocapsae and Steinernema feltiae in monoxenic liquid culture with their bacterial symbionts Xenorhabdus nematophila and Xenorhabdus bovienii, respectively, the effect of the initial nematode inoculum density on population development and final concentration of dauer juveniles (DJs) was investigated. Symbiotic bacterial cultures are pre-incubated for 1 day prior to inoculation of DJs. DJs are developmentally arrested and recover development as a reaction to food signals provided by their symbionts. After development to adults, the nematodes produce DJ offspring. Inoculum density ranged from 1 to 10 × 10³ DJ per milliliter for S. carpocapsae and 1 to 8 × 10³ DJs per milliliter for S. feltiae. No significant influence of the inoculum density on the final DJ yields in both nematode species was recorded, except for S. carpocapsae cultures with a parental female density <2 × 10³ DJs per milliliter, in which the yields increased with increasing inoculation density. A strong negative response of the parental female fecundity to increasing DJ inoculum densities was recorded for both species with a maximum offspring number per female of >300 for S. carpocapsae and almost 200 for S. feltiae. The compensative adaptation of fecundity to nematode population density is responsible for the lack of an inoculum (or parental female) density effect on DJ yields. At optimal inoculation density of S. carpocapsae, offspring were produced by the parental female population, whereas S. feltiae always developed a F1 female population, which contributed to the DJ yields and was the reason for a more scattered distribution of the yields. The F1 female generation was accompanied by a second peak in X. bovienii density. The optimal DJ inoculum density for S. carpocapsae is 3–6 × 10³ DJs per milliliter in order to obtain >10³ parental females per milliliter. Density-dependent effects were neither observed on the DJ recovery nor on the sex ratio in the parental adult generation. As recovery varied between different batches, assessment of the recovery of inoculum DJ batches is recommended. S. feltiae was less variable in DJ recovery usually reaching >90%. The recommended DJ inoculum density is >5 × 10³ DJs per milliliter to reach >2 × 10³ parental females per milliliter. The mean yield recorded for S. carpocapsae was 135 × 10³ and 105 × 10³ per mililiter for S. feltiae.     

Influences of preceding cover crops on slug damage and biological control using Phasmarhabditis hermaphrodita

In a replicated field experiment, ryegrass, vetch and red clover were grown or the soil was kept bare over a 2–month period in summer to compare the effects of these treatments on slug damage to the following crop (Chinese cabbage) and on the efficacy of nematodes (Phasmarhabditis hermaphrodita) applied as biological control agents to the soil at planting time to protect this crop. Slug damage was significantly (c. two times) greater after red clover or vetch than after ryegrass. Damage on plots without cover crop was intermediate and not significantly different from either extreme. Slug damage was reduced by about one‐third by the nematode treatment. The preceding cover crop did not influence nematode efficacy. Numbers of slugs on harvested plants (mainly Deroceras reticulatum and Deroceras panormitanum) were influenced by an interaction between cover crop and nematode treatment. On subplots without nematodes, more slugs were recorded with than without a preceding cover crop. No such differences were found on nematode‐treated subplots. Soil samples were collected at intervals from 0–99 days after nematode treatment to monitor nematode survival and infectivity in bioassays with D. reticulatum. No significant effects of cover crops were detected in bioassays. Moreover, there were no significant effects of nematodes on slug survival. Their effects on slug food consumption were mostly insignificant and any effects were transient and not consistent. However, significantly more slug cadavers contained nematodes when slugs were exposed to nematode‐treated soil. The implications of these results are discussed.     

Nematode associates and susceptibility of a protected slug (Geomalacus maculosus) to four biocontrol nematodes

The impact of selected entomopathogenic nematodes and Phasmarhabditis hermaphrodita on the European-Union-protected slug Geomalacus maculosus and the sympatric Lehmannia marginata was investigated. There was no significant difference in mortality between slugs treated with nematodes and their controls. The presence of P. hermaphrodita in two G. maculosus cadavers may be the result of necromenic behaviour. This study constitutes the first record of P. californica in Europe.     

Pathogenicity,reproduction and foraging behaviours of some entomopathogenic nematodes on a new turf pest,Dorcadion pseudopreissi (Coleoptera: Cerambycidae)

Entomopathogenic nematodes (EPNs) in the families Heterorhabditidae and Steinernematidae have considerable potential as biological control agents of soil-inhabiting insect pests. In the present study, the control potential of the EPNs Steinernema carpocapsae (TUR-S4), S. feltiae (Nemaplus), S. carpocapsae (Nemastar), S. feltiae (TUR-S3) and Heterorhabditis bacteriophora (Nematop) against a new longicorn pest, Dorcadion pseudopreissi Breuning, 1962 (Coleoptera: Cerambycidae), on turf was examined in laboratory studies. Pathogenicity tests were performed at the following doses: 50, 100 and 150 Dauer Juveniles (DJs)/larva at 25°C. Highest mortalities (75–92%) of the larvae were detected at the dose of 150 DJs/larva for all nematodes used. Reproduction capabilities of the used EPNs were examined at doses of 50, 75, 100 and 150 DJs/larva at 25°C. S. carpocapsae (TUR-S4) had the most invasions (32 DJs/larva) and reproduction (28042 DJs/larva) at the dose of 100 DJs, and the highest reproduction (per invaded DJ into a larva) was observed in H. bacteriophora (Nematop) (2402.85 DJs) at a dose of 50 DJs. The foraging behaviour of the nematodes in the presence of D. pseudopreissi and Galleria mellonella L. (Lepidoptera: Galleriidae) larvae was studied using a Petri dish filled with sand at 20°C. All of the used nematodes accumulated near the larvae section of both insect species (32–53% of recovered DJs) with a higher percentage of S. carpocapsae (TUR-S4) (53%) and H. bacteriophora (48%) (Nematop) moving towards larvae of D. pseudopreissi, than the S. feltiae strains.     

Pathogenicity and life cycle of Meloidogyne javanica on broccoli

A pathogenicity trial conducted against root-knot nematode, Meloidogyne javanica on broccoli indicated that a gradual increase in the nematode inoculum from 500 to 8000 juveniles/kg soil was associated with a progressive decline in all the plant growth parameters and reproduction factor of the nematode. Although 8000 juveniles/kg soil showed maximum plant growth reduction and root knot index, statistical analysis of the data revealed that the population of 1000 juveniles/kg soil was associated with a significant decline in plant growth. Hence, this level was indicative of being the pathogenic level. The significant reduction in seedling emergence was recorded at and above 2000 juveniles/kg soil and it decreased further with increasing inoculum levels. Meloidogyne javanica required 27 days to complete the life cycle on broccoli at a temperature range of 28–35°C.     

Meloidogyne incognita Inoculum Source Affects Host Suitability and Growth of Yellow Nutsedge and Chile Pepper

Meloidogyne incognita (Mi) reproduction and host plant responses in chile pepper (Capsicum annuum) and yellow nutsedge (Cyperus esculentus = YNS) to three sources of inoculum obtained by rearing a single Mi population on chile, YNS, and tomato were evaluated in two factorial greenhouse experiments. The interactive effects of Mi inoculum source and crop-weed competition were determined. In the absence of YNS competition, chile growth was reduced less by Mi inoculum from chile than by inoculum from YNS or tomato. When YNS was present, chile root weight was not affected and shoot weight increased with Mi initial inoculation, regardless of inoculum source. Chile plants inoculated with Mi from tomato exhibited double the nematode reproduction observed with inoculum from chile or YNS. With chile present, Mi reproduction on YNS was nearly three times greater with inoculum from tomato, but reproduction was similar among inoculum sources when chile was absent. Reductions in YNS root mass due to competition from chile failed to reduce the total number of Mi eggs produced on YNS plants. Differences in total Mi reproduction among inoculum sources were not attributable to differences in root growth or plant competition. This study illustrates the influence of Mi-YNS interactions and previous hosts on severity of Mi infection.     

Behavioural avoidance by slugs and snails of the parasitic nematode Phasmarhabditis hermaphrodita

The nematode Phasmarhabditis hermaphrodita has been developed as a biological control agent for slugs and snails. Slugs avoid areas where P. hermaphrodita is present. We investigated whether behavioural avoidance of P. hermaphrodita is a common feature of slugs and snails by exposing eight species to P. hermaphrodita. We showed that slugs generally avoided P. hermaphrodita, whereas snails did not. We also showed that slugs specifically avoided the commercial strain and a natural isolate of P. hermaphrodita and were not deterred by other nematodes such as Steinernema kraussei or Turbatrix aceti. We also showed that slugs avoided the dauer stage of P. hermaphrodita and not mixed-stage cultures. Furthermore, slugs do not avoid dead P. hermaphrodita or exudates from live nematodes. Taken together, we have unravelled further factors that are essential for slugs to avoid P. hermaphrodita in soil, which could have important implications for the biological control of slugs and snails.     

Comparison of concomitant and sequential inoculation of Steinernema sp. in the management of root-knot (Meloidogyne incognita) nematode infecting eggplant (Solanum melongena)

Culture of Steinernema sp. was maintained on Corcyra cephalonica larvae. Steinernema sp. (at 50, 500, 1000, 2500, 5000, 10,000 and 20,000 ij’s /500?g soil) was concomitantly inoculated with 500 J₂ of Meloidogyne incognita/500?g soil to the eggplant seedlings in the pots filled with 4?kg sterilised soil. The simultaneous inoculation of M. incognita with either of the inoculum levels (1000, 2500, 5000 and 10,000 J₃/500?g soil) of Steinernema sp. significantly reduced the damage caused by M. incognita in terms of plant growth parameters, viz. plant length, dry weight, number of flowers and weight of fruits. Moreover, the highest improvement in plant growth parameters, viz. plant length, dry weight, number of flowers and weight of fruits, was recorded in plants inoculated with 5000 J₃ of Steinernema sp./500?g soil followed by 2500, 1000 and 10,000 J₃/500?g soil. The highest reduction in the reproduction factor and number of galls/root system was recorded in the plants treated with 5000 J₃ Steinernema sp./500?g soil followed by 2500, 1000 and 10,000 J₃/500?g soil. Comparison of concomitant and sequential inoculations showed that the sequential inoculation (both prior and after) of Steinernema sp. at different inoculum levels (1000, 2500, 5000, 10,000 and 20,000 ij’s/500?g soil) was more effective in the management of root-knot nematode than the concomitant inoculation. Therefore, the application of Steinernema sp. might be useful for suppression of nematode pest on eggplant and may be used as an alternative for chemicals.     

Spatiotemporal Analysis of Predation by Carabid Beetles (Carabidae) on Nematode Infected and Uninfected Slugs in the Field

The dynamics of predation on parasites within prey has received relatively little attention despite the profound effects this is likely to have on both prey and parasite numbers and hence on biological control programmes where parasites are employed. The nematode Phasmarhabditis hermaphrodita is a commercially available biological agent against slugs. Predation on these slugs may, at the same time, result in intraguild predation on slug-parasitic nematodes. This study describes, for the first time, predation by carabid beetles on slugs and their nematode parasites on both spatial and temporal scales, using PCR-based methods. The highest nematode infection levels were found in the slugs Deroceras reticulatum and Arion silvaticus. Numbers of infected slugs decreased over time and no infected slugs were found four months after nematode application. The density of the most abundant slug, the invasive Arion vulgaris, was positively related to the activity-density of the carabid beetle, Carabus nemoralis. Predation on slugs was density and size related, with highest predation levels also on A. vulgaris. Predation on A. vulgaris decreased significantly in summer when these slugs were larger than one gram. Predation by C. nemoralis on slugs was opportunistic, without any preferences for specific species. Intraguild predation on the nematodes was low, suggesting that carabid beetles such as C. nemoralis probably do not have a significant impact on the success of biological control using P. hermaphrodita.     

Studies on the Pathogenic Potential and Life Cycle of Reniform Nematode on Broccoli

The significant reduction in the plant growth was observed at an initial inoculum level of 2 immature females of Rotylenchulus reniformis/g soil and hence this inoculum level was considered as the minimum damaging threshold level for broccoli. The population of reniform nematode increased with the increasing levels of inoculum. Though, the maximum nematode population was recorded at the highest level of inoculum, the rate of reproduction of nematode was found to be highest at the lowest level of initial inoculum. Symptoms like chlorosis, stunted growth, shedding of leaves, early sprouting of inflorescence and sparsely developed roots were found during the experimental studies. Rotylenchulus reniformis required 30 days to complete the life cycle on broccoli.     

Effects of Caenorhabditis elegans (Nematoda: Rhabditidae) on yield and quality of the cultivated mushroom Agaricus bisporus

Thirteen species of saprobic rhabditid nematodes (11 genera) were identified from samples of compost and casing material collected from mushroom farms in the British Isles. Caenorhabditis elegans, the most frequently found saprobe, was mass-produced monoxenically and its effects on the cultivated mushroom, Agaricus bisporus (strain U3) were studied. C. elegans did not multiply in well-prepared, pasteurised, spawned compost, whereas casing material proved to be a highly suitable environment for its reproduction. An initial casing inoculum of 10⁶ nematodes/crate of compost (7.5 kg), caused a significant reduction in mushroom yield. Losses in total mushroom yields of 11%, 20% and 26% were caused by initial inoculum rates of 10⁶, 10⁷and 2 × 10⁷ nematodes/crate, respectively. Yields were negatively correlated with the initial nematode inoculation level and regression equations were derived. The nematode treatments caused fewer mushrooms to be produced and an absence of the usual distinctive flushing patterns. C. elegans caused considerable deterioration in mushroom quality and characteristic distortion of mushrooms. Individual sporophores were mis-shapen, notched and had brown or violet coloured grills. Up to 3.8%, 6.7% and 10.8% of total weight and 3.5%, 5.4% and 8% of total numbers of mushrooms were distorted at the three highest nematode inoculum rates tested. Weights and numbers of distorted mushrooms were positively correlated with the initial nematode population. C. elegans commonly colonised sporophores.     

First record for the slug parasitic nematode,Phasmarhabditis hermaphrodita (Schneider) in Egypt

The rhabditid nematode, Phasmarhabditis hermaphrodita is a lethal parasite of certain terrestrial gastropods and has been shown as a biocontrol agent under laboratory and field conditions. In Egypt, P. hermaphrodita was isolated for the first time from different species of terrestrial gastropods found associated with various crops at Aga and Mansoura districts of Dakhalia Governorate during the year 2000/2001. Females and dauer larvae (IJs) were described and illustrated based on the light microscope. Males are not found as this species seemed to be protandrous. PCR analysis confirmed nematode identification. The Egyptian isolate of P. hermaphrodita was found to be shorter and lower in width than the British isolate. V%, a, b and c parameters showed detectable variations between two isolates with values of 54%, 17.7, 4.28, 13.7 in the Egyptian isolate, and 51%, 19.5, 7.2 and 15.8 for the British isolate, respectively.     

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.