The inhibitory effects of the molluscicide metaldehyde on feeding, locomotion and faecal elimination of three pest species of terrestrial slug

Single meals on maize flour pellets containing 0, 0·5, 1, 2, 4, 6, and 8% metaldehyde were analysed in Arion hortensis (agg), Deroceras reticulatum and Deroceras caruanae, by attaching the pellets to a mechanical transducer from which signals were ultimately fed into a BBC microcomputer for on-line analysis. Post-feeding tests were used to investigate toxic effects of metaldehyde and the recovery of slugs was examined daily for 7 days. Bite rates were slightly lower and more irregular on metaldehyde meals especially in A. hortensis. The meal length and number of bites per meal were markedly inhibited by metaldehyde and inversely proportional to the concentration. The size of the bites was also reduced, by about 30%, in meals on 6% pellets compared with 0%. Clear symptoms of poisoning became evident during the meal or normally in the 16-min test period immediately afterwards. In these tests slugs which fed on metaldehyde pellets showed a disruption of their ability to right themselves (after being placed on their side), less activity and a reduced probability of accepting a ‘dummy’ pellet containing no metaldehyde. It is hypothesised that feeding is prematurely terminated primarily because of the toxic effects of the metaldehyde and that aversion also acts to terminate the meal early, by decreasing the probability of continuing to feed as the meal proceeds. At higher concentrations of metaldehyde aversion may be more important than the toxic effects in ending the meal. Faecal elimination is delayed after metaldehyde meals and this contributes to mortality. The estimated lethal doses were about 0·2 mg/g body weight of slug in Deroceras spp. A. hortensis was less susceptible. To increase the likelihood of a lethal dose being ingested the inhibition of feeding needs to be overcome, at least partly, and the results presented indicate the need for a bait formulation which reduces the rate of absorption of the molluscicide from the gut.     

EFFECTS OF ORALLY AND DERMALLY APPLIED METALDEHYDE ON MUCUS CELLS OF SLUGS (DEROCERAS RETICULATUM) DEPENDING ON TEMPERATURE AND DURATION OF EXPOSURE

Using light and electron microscopy in combination with histochemicaland immunocytochemical techniques, the impact of orally anddermally applied metaldehyde on mucus cells in the digestivetract, the skin, and the salivary gland of the slug Derocerasreticulatum (Müller) was investigated. The studies showedthat metaldehyde induces severe alterations and damages in mycocyteseven under low temperature and humid conditions when sufficientlyhigh doses were applied. After metaldehyde application, notonly the quantity of mucus produced by slugs but also its qualityis modified. Structural, enzymehistochemical and immunocytochemicalinvestigations revealed metaldehyde-induced effects in mucocytesto be related to influences of the molluscicide on serotoninand on energy metabolism. (Received 29 September 1997; accepted 8 December 1997)     

CELL PROLIFERATION IN THE CASTRATE MOUSE SEMINAL VESICLE IN RESPONSE TO TESTOSTERONE PROPIONATE

The cell proliferation kinetics following induced DNA synthesis in the mouse seminal vesicle were measured after treatment with testosterone propionate. Fraction labelled mitosis curves at 24, 48 and 72 hr after injection gave t ₂ values of 1·5, 2·0 and 1·8 hr respectively, and t _s values of 10·5, 8·0 and 8·0 hr. T _c measured 48 hr after stimulation was 17·5 hr. Growth fraction rose from 0·14 at 24 hr to 0·64 at 48 hr, and fell to 0·32 by 72 hr. A simple model is proposed in which the rise and fall of mitotic index and labelled index is determined by the 'cell distribution ratio'.     

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.     

EFFECTS OF HYDROXYUREA AND RADIATION ON HAIR MATRIX CELLS

Investigations were undertaken in CF₁ mice to study the effects of hydroxyurea (HU) on hair matrix cell kinetics, and to assess the effects of combined administration of HU and irradiation on induction of temporary alopecia. HU 100 mg/kg was administered intraperitoneally and each animal received tritiated thymidine 0·5 μCi/g 30 min before biopsy. Serial biopsies were taken up to 48 hr after drug administration. Autoradiographs of anagen follicle squashes revealed sharp reductions in mitotic and labeling indices within 30 min. Depressed mitotic indices of 0·6–0·9% at 1–4 hr returned to normal (2·3%) after 6 hr, followed by cyclic mitotic 'overshoot', and were preceded by parallel changes in the labeling indices. HU-induced cellular damage was most marked 4 hr after HU injection, with almost complete recovery from injury observed at the 24 hr interval.
The effects of varying the time intervals from 1 to 12 hr between HU administration and irradiation (650 rads) after injection of HU 1200 mg/kg were examined. Hair loss was measured 7 days later by photomicroscopy. Cyclic maximum alopecia was found at the 1–5 and 8–12 hr intervals, with relative 'protection' occurring at the 6–7 hr time periods.     

Differential susceptibility of species of slugs to metaldehyde/bran and to methiocarb baits

Summary Laboratory experiments were designed to study differential susceptibility of nine species of slugs to metaldehyde/bran and to methiocarb baits. Differential susceptibility was found between some of the species for each type of bait, both immediately after baiting and also after a 48 h recovery period. Thus neither type of bait is suitable for use in ecological investigations. This conclusion was supported by evidence that interspecific behavioural interactions occurred when metaldehyde, but not methiocarb, baits were used.At the same time the relative efficiency of the two types of baits as molluscicides was estimated. Methiocarb was shown to be a better molluscicide because, not only did it initially poison more slugs than did metaldehyde, but also a number of the slugs initially poisoned by metaldehyde recovered during the following 48 h period, whereas this did not happen with methiocarb. The similarity between these results and those obtained from field experiments by other workers is discussed.     

Effect of delayed first feeding on development and feeding ability of Paralabrax maculatofasciatus larvae

The impact of delayed feeding early in development on late feeding ability and development of spotted sand bass Paralabrax maculatofasciatus larvae was examined. Larvae were sampled from hatching until day 19 after delayed feeding for zero (control), 1, 2, or 3 days. Feeding incidence was evaluated as the percentage of larvae with food in the gut and feeding intensity was measured by direct counting of prey after dissection of the gut. Delayed feeding effects due to starvation were observed early in development. By day 5, significant differences ( P  < 0·05) were observed between controls and larvae submitted to degrees of delayed feeding, including total length ( L _T), eye diameter, and the size of the head, liver, gut, muscles and body. Differences were still apparent in L _T, and body, liver and muscle heights at the end of the studied period. Larvae under total starvation did not survive beyond day 5. Initial feeding incidence was 35, 60, 90 and 10% for larvae first fed on day 2, 3, 4 or 5, respectively. Mean ±  s . d . feeding intensity was 3·6 ± 0·8, 2·8 ± 1·3, 5·2 ± 0·3, and 10·2 ± 1·5 rotifers per larva depending on whether larvae were initially fed on day 2, 3, 4 or 5, respectively. The point of no return occurred between 4 and 5 days after hatching.     

Laboratory methods for assessing the toxicity of contact poisons to slugs

Two techniques for comparing the activity of different contact poisons to slugs in controlled conditions were used to measure the relative toxicities of five substances. A laboratory immersion test rated their median lethal concentrations as follows: ioxynil 8·3 ppm, sodium pentachlorophenate 22·0 ppm, copper sulphate 68·1–75·3 ppm, acetaldehyde 4822·0 ppm. Metaldehyde gave inconsistent results with this method but, using a dry-contact method, metaldehyde (42370 ppm) was much less toxic than copper sulphate (2027 ppm). The materials giving practical control in the field were not the most toxic of those tested.     

Field Test Using the Nematode Phasmarhabditis hermaphrodita for Biocontrol of Slugs in Spain

An experiment was carried out between May and July 1999 in Galicia (North-West Spain) to test the capacity of the nematode Phasmarhabditis hermaphrodita to protect field grown lettuces from slug damage in our field conditions. The experiment compared a single dose of nematodes (3 ×109 ha -1) with mini-pellets containing 5% metaldehyde, applied at the recommended field rate (3 g pellets m -2), and untreated plots. Slug damage for each lettuce head was estimated on six dates during the first 4 weeks after planting. At harvest, each lettuce head was weighed, scored as marketable or not by weight and external aspect, and inspected for slugs. Metaldehyde significantly reduced slug damage to lettuce plants from the first day after planting to the third week. Nematodes significantly reduced slug damage from the second to the third week. At harvest, 6 weeks after planting, the mean weight of the lettuce heads and the number of marketable heads in the nematode plots were as good as in the metaldehyde plots, and both treatments were significantly better than the untreated plots. The number of slugs within the harvested plants was significantly reduced only with the metaldehyde treatment.     

CYTOKINETIC ANALYSIS OF THE JB-1 ASCITES TUMOUR AT DIFFERENT STAGES OF GROWTH

The cell kinetics of the murine JB-1 ascites tumour have been investigated on days 4, 7 and 10 after transplantation of 2·5 × 10⁶ cells. The experimental data, growth curve, percentage of labelled mitoses curves, continuous labelling curves and cytophotometric determination of single-cell DNA content have been analysed by means of a mathematical model for the cell kinetics. The important result was the existence of 8% non-cycling cells with G₂ DNA content in the 10-day tumour, while only 0·2 and 0% were observed in the 7- and 4-day tumours, respectively. The doubling times determined from the growth curve were 22·8, 70 and 240 hr, respectively, in the 4-, 7- and 10-day tumours. Growth fractions of 76, 67 and 44% were calculated for the same tumour ages. The mean cell cycle time increased from 14 to 44 hr from day 4 to 7 due to a proportional increase in the mean transit time of all phases in the cell cycle. In the 10-day tumour, the mean cell cycle changed to 41 hr and T _G1 decreased to 0·5 hr. The cell production rate was 4·3%/hr in the 4-day tumour, 1·2%/hr in the 7-day tumour and 1·0%/hr in the 10-day tumour. The cell loss rates in the same tumours were 1·3, 0·2 and 0·7%/hr, respectively. The analysis made it probable that the mode of cell loss was an age-specific elimination of non-cycling cells with postmitotic DNA content.     

HYDROXYUREA EFFECTS IN THE C3H MOUSE II. MAMMARY TUMOR CELL KINETICS

The cellular kinetics of C3H mouse mammary tumors were studied following a single dose (3 mg/g body weight) of hydroxyurea (HU). This dose was large enough to cause a significant perturbation in the growth curves of these tumours. This was accomplished by labeling the cells with tritiated 5-iodo-2'-deoxyuridine and performing detailed autoradiographic analysis. This dose of HU caused a temporary inhibition in growth and completely inhibited DNA synthesis for 4–5 hr. The HU-killed cells (pyknotic and karyorrhectic) reach a maximum around 10–12 hr and are apparently all removed in about 1 day. Tumors from a fast-growing line (S102F) showed some evidence for cell synchrony upon recovery from HU inhibition but desynchronization occurred within one cell cycle. The cell generation time was not decreased during the acute recovery phase, but the growth fraction shifted from 0·6 to 1·0, and the data suggested that the normal flow of cells from the proliferating pool to the degenerate pool was temporarily interrupted. The cellular kinetic parameters have probably returned to normal by 48 hr after the HU injection.     

Effects of metaldehyde and methiocarb on feeding preferences and survival of a slug predator (Pterostichus melanarius (F.): Carabidae,Pterostichini)

The potential for reducing slug populations in crops through predation by generalist carabid beetles is well documented. However, few studies have considered interactions between biological and chemical control agents of slugs. Laboratory assays supported previous findings that the consumption of metaldehyde by slugs (Deroceras reticulatum) leads to increased duration of feeding bouts by carabid beetles (such as Pterostichini) on sub‐lethally affected individuals. However, a similar effect was not found for Pterostichus melanarius exposed to slugs fed on the other widely applied pelleted molluscicide formulation (methiocarb). Examination of beetle survival after consumption of slugs containing molluscicides demonstrated the strong biocidal properties of methiocarb, whereas metaldehyde consumption (ingested through slug predation) did not differ from control slugs killed by freezing. Beetle avoidance of slugs containing a more toxic molluscicide compound and the interaction between slug mucus production and beetle attack rates are discussed.     

The Use of Metaldehyde Against Slugs

One part of metaldehyde in thirty parts of bran by volume is the minimum effective concentration for obtaining an economic kill of slugs under average weather conditions. For use against Milax gracilis Leydig. heaps of bait should not be normally more than 2 yd. apart and against Arion hortensis Fer. not more than 3 ft. apart. The mortality of slugs attracted to metaldehyde-bran baits is dependent on the drying property of the soil and atmosphere around the baits. D.D.T. flea-beetle dust and powdered pyrethrum flowers are sufficiently distasteful to slugs to prevent their consuming a lethal dose of the poison mixed with bran. The addition of 10%, by volume, of dried blood to metaldehyde-bran bait increases the catch of some slugs under certain conditions but does not increase the mortality. The addition of 10%, by volume, of dextrose increases the mortality produced by metaldehyde but does not increase the numbers attracted to the bait. A metaldehyde-bran bait compacted into 'biscuit' form with anhydrous calcium sulphate catches more slugs than an ordinary metaldehyde-bran bait, but almost completely counteracts the effect of the metaldehyde. A metal-dehyde-bran-casein glue bait in 'biscuit' form increases the catch and kill of Milax gracilis and Agriolimax reticulatus Müll., but not of Arion hortensis . Metaldehyde-bran-casein glue bait broadcast in 'broken biscuit' form is effective over a longer period, thus producing a greater kill of slugs, than a broadcast metaldehyde-bran bait.     

The fumigant effect of metaldehyde on slugs

The fumigant effect of metaldehyde on slugs is confirmed and shown almost certainly to be caused by impure metaldehyde breaking down to the volatile parent monomer, acetaldehyde. Acetaldehyde was slightly more toxic to Agriolimax reticulatus than to Arion hortensis.     

THE PROLIFERATION OF LYMPHOID CELLS IN GUINEA-PIG BONE MARROW

A double isotope DNA labelling method has been used to determine the duration of DNA synthesis (S) in bone marrow lymphoid cells classified by their nuclear diameters in smears. Incorporation of ³H-thymidine was confined almost entirely to marrow lymphoid cells of 8·0-15·0 μm nuclear diameter (large lymphoid cells). After exposure to ³H-thymidine in vivo and ¹⁴C-thymidine 40-104 min later in vitro , the proportion of cells labelled with ³H alone to those labelled with ¹⁴C(±³H) in radioautographic smears, plotted against time indicated the efflux from S per hour. Collectively, 28·3 ± 1·1% of all large lymphoid cells were in S and the efflux from S was 15·1% per hour. With decreasing cell size (nuclear diameter) the efflux fell progressively from 28·3% per hour (11·0 μm) to 9·2% per hour (8·0-8·9 μm) and the proportion of cells in S declined from 54·9 ± 2·3% to 14·8 ± 1·6%. Influx into S, measured in vitro by reversing the sequence of isotopes, closely resembled the corresponding efflux values in vivo relative to cell size. Most DNA synthesizing marrow large lymphoid cells belonged to a subgroup with deeply basophilic cytoplasm. The results demonstrate that basophilic large lymphoid cells in the marrow are actively proliferating and have a mean S phase duration of 6·6 hr. The largest marrow lymphoid cells (11·0 μm) proliferate most rapidly (S phase, 3·5 hr; maximum cell cycle time, 6·4 hr) while S duration is prolonged progressively to 10·9 hr for the smaller cells (8·0-8·9 μm).     

Overall Versus Band Application of the Nematode Phasmarhabditis hermaphrodita With and Without Incorporation into Soil, for Biological Control of Slugs in Winter Wheat

In two concurrent field experiments, the effects of three types of soil cultivation and two patterns of nematode application were studied in order to investigate their effects on damage to winter wheat by slugs (assessed at Zadoks Growth Stage 12). In experiment 1, infective juveniles (IJs) of the nematode Phasmarhabditis hermaphrodita were applied to soil as an overall spray or as a band spray (8-cm wide), centred on the drill rows (16.7-cm apart). Nematodes were either left undisturbed on the soil surface or harrowed into the soil immediately after application. The control provided by nematodes was compared with that provided by metaldehyde and methiocarb pellets broadcast at the recommended rate immediately after drilling. In this experiment, winter wheat on plots treated with IJs showed significantly less slug damage than on wheat plots treated with metaldehyde or methiocarb pellets or untreated plots. There was no significant difference in plant damage between plots treated with band and overall spray applications of IJs, nor was there any significant difference between plots with and without harrowing. There was also no significant difference between untreated plots and plots treated with metaldehyde or methiocarb pellets, probably because rainfall shortly after treatment rendered the pellets ineffective. In experiment 2, nematodes were applied as an overall spray or plots were not treated with nematodes before soil was cultivated with tines, Roterra or Dutzi cultivators. Nematode application before soil cultivation using tines or Roterra reduced the number of plants damaged significantly. However, nematodes applied before Dutzi cultivation appeared to be rendered ineffective. Damage to winter wheat was lowest in plots that had been sprayed with nematodes and subsequently cultivated with tines or Roterra.     

Schistosoma haematobium: Amoscanate and adult worm ultrastructure

Amoscanate, when administered orally as an aqueous or “formulated” preparation, induced pronounced ultrastructural abnormalities in male and female Schistosoma haematobium. Higher dose levels of the aqueous suspension (300 mg/kg body wt) had to be administered to achieve the full range of effects induced by formulated doses of 2.5–8 mg/kg body wt. Worms were recovered from hamsters between 1 and 120 hr after treatment. Although the amount of amoscanate-induced damage varied considerably between worms, an overall pattern of damage emerged. Initially, 1 hr after treatment, amoscanate caused tegumental vacuolation and oedema. As the drug treatment period was extended to 24 hr, blebbing, exudation, collapse of sensory organelle bases, and abnormal mitochondria became increasingly evident. With exposure to higher drug doses (50–300 mg/kg body wt), the tegument became further distorted with the appearance of necrotic structures and myelin whorls, which appeared to represent various stages in lysosomal formation and digestion. Eventually, erosion of surface layers resulted in the breakdown of tegumental integrity. The caeca and vitellaria were also adversely affected by drug treatment. Basal vacuolation and the formation of myelin whorls occurred in the gastrodermis. In the mature S4 vitelline cells, coalesced vitelline droplets and myelin whorls were evident.     

The genetic architecture of correlations among growth‐related traits and male age at maturation in rainbow trout

Heritabilities and genetic correlations among growth‐related traits of two cultured strains (Rainbow Springs and Spring Valley) of rainbow trout Oncorhynchus mykiss were estimated using restricted maximum likelihood methods with a three‐generation pedigree. Heritability was high (>0·50 ± 0·03) for body mass and condition factor but moderate (0·35 ± 0·04) for age at sexual maturity in males. Body mass and age at sexual maturation were phenotypically correlated in the families of one experimental strain, Rainbow Springs, and had a positive genetic correlation (0·26 ± 0·03) across families from both test strains (Rainbow Springs and Spring Valley). This indicates that faster growing individuals were more likely to mature at 2 years of age than slower growing individuals in the two hatchery strains investigated. Microsatellite markers of body mass quantitative tract loci (QTL) were reconfirmed as being located on linkage groups B, G, N, 5 and new markers on Oi were detected. Some QTL effects were restricted to specific sampling dates suggesting temporal expression of QTL. QTL for condition factor were limited to linkage group G in both strains. Three suggestive QTL for precocious maturation mapped to similar regions as those for body mass in the Rainbow Springs families while no associations were evident in the Spring Valley families. The results suggest that these regions may play a role in the basis for genetic and phenotypic correlations between body mass and precocious maturation in this species.     

TESTING MOLLUSCICIDES AGAINST SLUGS -- THE IMPORTANCE OF ASSESSING THE RESIDUAL POPULATION

The field performance of molluscicidal baits used against slugpests of crops is normally measured by the numbers of dead animalsfound on the surface of plots which have been treated with differentbaits. This paper presents evidence that although some materialsseem to give high kills of the Grey Field Slug, Deroceras reticulatum(Müll.), the numbers of slugs left behind on the plotsafter such treatments, the residual population, are the sameas other plots where apparently fewer slugs have been killed.It thus seems likely that the field performance of baits containingmethiocarb and metaldehyde are very similar, both materialsproducing a similar reduction in the adult slug population. Field trials of mollusdcides should always attempt to measurethe residual population left behind on the plots and this wouldthen eliminate apparent errors in the interpretation of resultscaused by the dissimilar modes of action of the various materialsused as molluscicides. (Received 8 June 1982;     

Caffeine as a novel toxicant for slugs and snails

In this study, caffeine is shown to act as both a repellent and toxicant against slugs and snails. This research is the first to document the potential of caffeine as a molluscicide. A drench treatment using a 1% or 2% solution of caffeine caused 100% of slugs (Veronicella cubensis) to exit treated soil, and the majority of these slugs subsequently died from caffeine poisoning. A 2% solution of caffeine applied to the growing medium of orchids killed 95% of orchid snails, (Zonitoides arboreus), and gave better control than a liquid metaldehyde product representing the standard commercial control for this pest. Using leaf‐dip bioassays, we discovered that slugs tended to avoid feeding on plant material treated with caffeine solutions 0.1%, and caffeine solutions as low as 0.01% significantly reduced overall feeding by slugs. Due to concerns about chemical residues, available molluscicides generally cannot be applied directly to food crops for control of slug and snail pests. Caffeine is a natural product which is approved as a food additive. Therefore, caffeine may prove useful for protecting food crops from slugs and snails.