Reduced Egg Hatch Research Articles

Halyomorpha halys mortality and sublethal feeding effects following insecticide exposure

The brown marmorated stink bug, Halyomorpha halys (Stal), is a highly polyphagous invasive pest. Increased use of broad-spectrum insecticides to manage H. halys has resulted in secondary pest outbreaks and disruptions to integrated pest management (IPM) programs. We evaluated H. halys mortality, molting, and feeding after exposure to insecticides in the laboratory. Five insecticides (four active ingredients), considered less risky to natural enemies, were compared to a pyrethroid insecticide and an untreated control. Compared to the control, only azadirachtin + pyrethrins significantly reduced egg hatch, while all insecticides caused significant direct mortality to 1st and 2nd instars 5 days after hatch (DAH). Bifenthrin quickly caused complete mortality of adults, and the only insecticide to statistically match this level of mortality was sulfoxaflor at 14 days after treatment (DAT). Azadirachtin + pyrethrins and sulfoxaflor significantly reduced the proportion of 1st instars that molted compared to the control. Adults that survived sulfoxaflor exposure produced significantly fewer feeding sites than the control. However, when taking into consideration both lethal and sublethal effects, all insecticides, except pyrethrins, resulted in significant reductions in feeding sites/individual compared to the control. This more complete estimate of efficacy (i.e., reduction in injury/insect), confirms the potential of several insecticides to reduce crop injury without the necessity of high direct mortality to H. halys.

Biopotentials of Marine Algae Extracts against Root-Knot Nematode, Meloidogyne incognita

Nematicidal activity of four species of marine algae Ulva fasciata Delile (UF) (green algae), Corallina mediteranea, Corallina officinalis (red algae) and Spirulina platensis (blue-green algae) at concentrations of (125, 250, 500 and 1000 μg/ml) for aqueous and ethanolic extracts were investigated against the knot-root nematode, Meloidogyne incognita in laboratory and greenhouse on tomato plants. In laboratory experiments, all tested algae caused significant mortality of the second larval stage (J2). It was, also, noticed that the tested algae reduced egg hatching. Ethanol extract of all algae possesses highest nematicidal activity than water extract. U. fasciata,ethanolic extracts showed the highest nematicidal activity among the tested algae in vitro. The results of greenhouse experiment clarified that the tested ethanolic extracts of all algae, reduced numbers of root galls, egg masses and population of nematodes of tomato plants at concentration 1000 mg/kg soil compared to the inoculated control. U. fasciata was the most effective treatment in reducing root galls, egg masses and final population of the nematode except for oxamyl. Results revealed that all tested treatments increased various measures of plant growth characters; fresh shoot and root weights and lengths as well as reduced the root- knot infestation. Among all of the tested algae, U. fasciata followed by C. officinalis and C. mediteranea were the most effective treatments to increase both fresh shoot & root weights and lengths. None of the algae was phytotoxic at the tested treatments. A chemical constituent of U. fasciata was recognized by GC-MS. GC-MS analysis, exhibited the presence of organic component such as bis(2-ethylhexyl) phthalate which ranked the first with 63.75%, followed by diethyl phthalate (18.46%). Results stated that marine algae and especially, U. fasciata exhibited significant nematicidal activity in-vitro and in-vivo experiments and could be considered as useful natural namaticidal agents.

Control of Globodera spp. Using Brassica juncea Seed Meal and Seed Meal Extract

The eradication program for the potato cyst nematode (PCN), Globodera pallida, in the Northwest of the United States revolves around the use of soil fumigation. Alternative, integrated strategies are needed to continue to battle this invasive nematode. Laboratory, greenhouse, and field experiments were conducted with G. pallida and another cyst nematode found in the United States, Globodera ellingtonae, to evaluate the efficacy of a new formulated Brassica juncea seed meal extract, as well as a traditional B. juncea seed meal, as alternate eradication strategies. This is the first report on the efficacy of B. juncea seed meal extract against plant-parasitic nematodes. Rates of B. juncea seed meal greater than 2.2 t/ha and 4.5 t/ha for G. pallida and G. ellingtonae, respectively, were required for egg hatch suppression, as determined by a potato root diffusate (PRD) bioassay. Reproduction of G. pallida on potato after exposure to B. juncea seed meal at a rate of 2.2 t/ha was also significantly reduced. In the field, 8.9 t/ha B. juncea seed meal almost eliminated egg hatch of G. ellingtonae. Rates needed for Globodera spp. suppression were greatly reduced when using the B. juncea seed meal extract. When compared side-by-side, half as much B. juncea seed meal extract, 1.1 t/ha, was required to suppress G. ellingtonae egg hatch to the same extent as B. juncea seed meal. Exposure of G. pallida to B. juncea seed meal extract at 4.5 t/ha reduced egg hatch by 90% compared with a nonamended control. The ability to reduce the amount of material being applied to soil by using an extract has the potential for integration into a G. pallida eradication program.

Molecular and functional characterization of Bm05br antigen from Rhipicephalus microplus

Rhipicephalus microplus is a cattle-specific tick, causing considerable losses in the livestock industry. The identification of molecules responsible for modulation of host defenses during different parasite stages can help in the development of alternative methods, such as vaccination, to control tick infestations. Hq05, a protein of unknown function identified in the tick Haemaphysalis qinghaiensis, induced a significant protective immune response when used as a vaccine in sheep. In the present study, we investigated Bm05br, the Hq05 homologous gene from R. microplus. Besides H. qinghaiensis, Bm05br homologous found in other tick species such as Rhipicephalus annulatus, Rhipicephalus sanguineus sensu lato, Haemaphysalis longicornis and Ixodes scapularis were comparatively analyzed. Bm05br expression profile in different R. microplus tissues and life-stages was determined by qRT-PCR and Western blot. Bm05br was detected in ovaries, salivary glands and the fat body of both partially and fully engorged females. The highest transcription levels were observed in partially engorged females fat body and salivary glands. Gene knockdown by RNAi reduced egg hatching rate and the weight of tick larvae obtained from treated group, when compared to controls. These results indicate that Bm05br may be involved in R. microplus reproduction. Together with its distribution and high sequence conservation across different tick species, our data suggest Bm05br as a potential antigen for development of a multispecies anti-tick vaccine.

Effect of heat stress and oil formulation on conidial germination of Metarhizium anisopliae s.s. on tick cuticle and artificial medium

The effect of heat stress (45°C) versus non-heat stress (27°C) on germination of Metarhizium anisopliae sensu stricto (s.s.) isolate IP 119 was examined with conidia formulated (suspended) in pure mineral oil or in water (Tween 80, 0.01%), and then applied onto the cuticle of Rhipicephalus sanguineus sensu lato (s.l.) engorged females or onto culture medium (PDAY). In addition, bioassays were performed to investigate the effect of conidia heated while formulated in oil, then applied to blood-engorged adult R. sanguineus females. Conidia suspended in water then exposed to 45°C, in comparison to conidia formulated in mineral oil and exposed to the same temperature, germinated less and more slowly when incubated on either PDAY medium or tick cuticle. Also, conidial germination on tick cuticle was delayed in comparison to germination on artificial culture medium; for example, germination was 13% on tick cuticle 72h after inoculation, in contrast to 61.5% on PDAY medium. Unheated (27°C) conidia suspended in either water or oil and applied to tick cuticle developed appressoria 36h after treatment; whereas only heat-stressed conidia formulated in oil developed appressoria on tick cuticle. In comparison to conidia heated in mineral oil, there was a strong negative effect of heat on germination of conidia heated in water before being applied to arthropod cuticle. Nevertheless, bioassays [based primarily on egg production (quantity) and egg hatchability] exhibited high percentages of tick control regardless of the type of conidial suspension; i.e., water- or oil-formulated conidia, and whether or not conidia were previously exposed to heat. In comparison to aqueous conidial preparations, however, conidia formulated in oil reduced egg hatchability irrespective of heat or no-heat exposure. In conclusion, mineral-oil formulation protected conidia against heat-induced delay of both germination and appressorium production when applied to the cuticle of R. sanguineus.

English

Present investigation was carried out to assess the biocontrol potential of Rhizobium leguminosarum against root-knot nematode, Meloidogyne incognita on soybean. Effect of Rhizobium strains LSI-21 and CRI-29 was assessed individually and concomitantly on reproduction of M. incognita. In vitro studies were conducted to check the effect of Rhizobium strains on egg hatching and juvenile mortality of M. incognita. Concomitant application of Rhizobium strains was more effective in reducing egg hatching and causing juvenile mortality as compared to individual treatment. In green house, Rhizobium strains were used as soil drench and seed dressing for the management of root knot nematode on soybean. It caused considerable reduction in reproduction of root knot nematode as well as improved plant growth parameters of soybean. Seed dressing showed most significant results than soil drenching. It is interesting to note that Rhizobia not only controlled reproduction of M. incognita but also an increase in plant height, fresh and dry root shoot weight was observed. It would seem that Rhizobium being a good rhizosphere organism for field and vegetable crops presumably prevent the contact of pathogenic organisms on roots.

Long-chain acyl-CoA synthetase 2 knockdown leads to decreased fatty acid oxidation in fat body and reduced reproductive capacity in the insect Rhodnius prolixus

Long-chain acyl-CoA esters are important intermediates in lipid metabolism and are synthesized from fatty acids by long-chain acyl-CoA synthetases (ACSL). The hematophagous insect Rhodnius prolixus, a vector of Chagas' disease, produces glycerolipids in the midgut after a blood meal, which are stored as triacylglycerol in the fat body and eggs. We identified twenty acyl-CoA synthetase genes in R. prolixus, two encoding ACSL isoforms (RhoprAcsl1 and RhoprAcsl2). RhoprAcsl1 transcripts increased in posterior midgut on the second day after feeding, and RhoprAcsl2 was highly transcribed on the tenth day. Both enzymes were expressed in Escherichia coli. Recombinant RhoprACSL1 and RhoprACSL2 had broad pH optima (7.5–9.5 and 6.5–9.5, respectively), were inhibited by triacsin C, and were rosiglitazone-insensitive. Both showed similar apparent Km for palmitic and oleic acid (2–6μM), but different Km for arachidonic acid (0.5 and 6μM for RhoprACSL1-Flag and RhoprACSL2-Flag, respectively). The knockdown of RhoprAcsl1 did not result in noticeable phenotypes. However, RhoprACSL2 deficient insects exhibited a 2.5-fold increase in triacylglycerol content in the fat body, and 90% decrease in fatty acid β-oxidation. RhoprAcsl2 knockdown also resulted in 20% increase in lifespan, delayed digestion, 30% reduced oviposition, and 50% reduction in egg hatching. Laid eggs and hatched nymphs showed remarkable alterations in morphology. In summary, R. prolixus ACSL isoforms have distinct roles on lipid metabolism. Although RhoprACSL1 functions remain unclear, we propose that RhoprACSL2 is the main contributor for the formation of the intracellular acyl-CoA pool channeled for β-oxidation in the fat body, and is also required for normal reproduction.

Ovicidal effects of indigenous plant extracts alone and with triazophos 40 EC on Spodoptera litura (F.)

Against Spodoptera litura (F.) the plant extracts when evaluated neem seed extract 5% recorded minimum egg hatching followed by subza (Ocimum basilicum) leaf extract and these were significantly superior over marigold (Tagetes erecta) leaf extract, orange (Citrus reticulata) peel extract and ginger (Zingiber officinale) rhizome extract @ 5% each. However, none of these alone could surpass the effects of trizophos 40 EC. There was an increase in the ovicidal activity when these extracts were mixed with trizophos 40 EC compared with their individual effects. All the plant extracts and their combinations with trizophos 40 EC were effective in reduced egg hatching but not effective compared to trizophos. The post hatching observations indicated that the moribund larvae were more with leaf extracts.

EFFECT OF FREEZING, HIGH TEMPERATURE AND LASER RADIATION FOR CONTROLLING KHAPRA BEETLE, TROGODERMA GRANARIUM (EVERTS), ON WHEAT GRAINS

khapra beetle, Trogoderma granarium (Everts) (Coleoptera, Dermestidae) is one of primary insects on stored grains. To control this pest with non-chemical methods, all stages (eggs, larvae, pupae and adults) of the pest were exposed to low and high temperature for different periods. Results revealed that mortality of all stages of T. granarium reached 100% after 7 and 5 days freezing at -10 and -15 °C, respectively. Eggs were more sensitive to low temperature than the other stages. On the other hand, high temperatures of 50 and 60 °C caused 100% mortality for all stages of the same pest after 6 hours of exposure. The increase of high temperature to 70 °C, the decrease of exposure period to 4 hours. Both low and high temperatures significantly affected germination and chlorophyll contents of wheat grains. Also laser ray of solid state of diod pump (wave length of 532 nm) reduced egg hatchability and pupation as well as adult emergence of exposed 0-1 day old eggs or 0-1 day old pupae. Laser irradiation significantly reduced germinationrate, but there was no effect on chlorophyll content.

Testing various biocontrol agents against the root-knot nematode (Meloidogyne incognita) in cucumber plants identifies a combination of Syncephalastrum racemosum and Paecilomyces lilacinus as being most effective

The root-knot nematode (RKN) (Meloidogyne incognita) is one of the most important pathogens on greenhouse-cultivated cucumber in northern China, and chemical nematicides are neither ecofriendly nor cost-effective. Thus, more benign and acceptable biological control measures for this destructive pathogen are required. The in vitro ovicidal and larvicidal activities of Syncephalastrum racemosum, Paecilomyces lilacinus and Bacillus cereus were tested on cucumber, and their combined effects against M. incognita were evaluated in pot and greenhouse experiments. The chemical nematicide fosthiazate was evaluated for comparison. All the bioagents displayed nematicidal potential via their ovicidal and larvicidal action, except the B. cereus bioagent at a concentration of 2%. The highest ovicidal activity was recorded for the combination of S. racemosum and P. lilacinus at a concentration of 50%, which resulted in an approximately 70% reduction in egg hatching compared to the control. The best larvicidal activity was observed in the S. racemosum treatment at a concentration of 50%, which caused a 96.7% mortality rate. Under the pot conditions, compared to the untreated control, the application of S. racemosum combined with P. lilacinus significantly stimulated the root length, shoot length and plant weight of cucumber plants, and significantly fewer galls and nematodes were observed. Under the greenhouse conditions, the combination of S. racemosum and P. lilacinus not only decreased the nematode severity but also increased the cucumber yield compared to the untreated control. Among all the treatments, the combined application of S. racemosum and P. lilacinus was more effective at controlling M. incognita than other treatments, except for the fosthiazate treatment. The results indicate that the combined use of S. racemosum and P. lilacinus represents a new biocontrol strategy against parasitic nematodes in protected agricultural areas.

Maternal effect and speciation: maternal effect contributes to the evolution of hybrid inviability between Drosophila simulans and Drosophila mauritiana.

Haldane's rule has been the basis of speciation research during the last 30 years. Most studies have focused on the nature of incompatibilities in the hybrid male, but not much attention has been given to the genetic basis of fertility and inviability in hybrid females. Hybridizations between Drosophila simulans and Drosophila mauritiana produce fertile females and sterile males. Here, we re-examined the level of fertility in reciprocal F1 females of these two species and looked for the presence of maternal effects. Our results show that the reciprocal F1 females of D. simulans and D. mauritiana hybridizations are fully fertile and in fact show a significant level of heterosis in the rate of oviposition but display reduced egg hatching in one direction. Reduced egg hatching was observed in the progenies of F1 hybrid females with D. mauritiana as mother, the same cross that showed a stronger negative effect on F1 male fertility. A review of the literature on the hybridizations in Lepidoptera also showed a maternal effect on inviability when reciprocal crosses produced asymmetric results. Our findings point to the importance of maternal effects in the evolution of embryo inviability and thus enhancing the process of speciation through the evolution of hybrid inviability.

Systemic acquired resistance activation in solanaceous crops as a management strategy against root-knot nematodes.

Activators of systemic acquired resistance (SAR), such as salicylic acid (SA) and its synthetic functional analogues benzo(1,2,3)thiadiazole-7-carbothionic acid-S-methyl ester (BTH) and 2,6-dichloroisonicotinic acid (INA), were tested on tomato, eggplant and pepper for the control of the root-knot nematode Meloidogyne incognita. Effects on plant fitness, nematode reproduction and root galling were screened in relation to different methods of application, to different applied dosages of chemicals and to different plant growth stages. Dosages applied to plants were in relation to plant weights. These chemicals were also tested for their possible nematotoxic activity in vitro. Soil drenches of SA and INA and root dip application of SA and BTH inhibited nematode reproduction, at specific dosage ranges, without affecting plant growth. SA and INA were able to reduce root galling as well. Foliar sprays of both SA and BTH were ineffective against nematode attacks. Plants tolerated SA more than the other chemicals tested. BTH at elevated concentrations increased the mortality of nematode juveniles and reduced egg hatching in vitro. SAR activators at concentrations suitable for different plant growth stages and applied by the proper method can possibly be included in IPM programmes for nematode management.

Haemocyanin is essential for embryonic development and survival in the migratory locust.

Haemocyanins are commonly known as copper-containing oxygen carriers within the haemolymph of arthropods, and have been found in many orders of insects. However, it remains unresolved why haemocyanins persist in insects that possess elaborate tracheal systems for oxygen diffusion to cells. Here we identified haemocyanins in the migratory locust Locusta migratoria that consists of two distinct subunits, Hc1 and Hc2. Genomic sequence analysis indicated that Hc1 and Hc2 have four and three gene copies, respectively, which may have evolved via gene duplication followed by divergent evolution of introns. The two subunits exhibit abundant and embryonic-specific expression at the mRNA and protein level; their expression peaks in the mid-term embryo and is not detectable in the late nymphal and adult stages. A larger proportion of the haemocyanins is present in the yolk compared with that in the embryo. Immunostaining shows that haemocyanins in the embryo are mainly expressed in the epidermis. Knockdown of Hc1 and Hc2 results in significant embryonic developmental delay and abnormality as well as reduced egg hatchability, ie the proportion of hatched eggs. These results reveal a previously unappreciated and fundamental role for haemocyanins in embryonic development and survival in insects, probably involving the exchange of molecules (eg O2 ) between the embryo and its environment.

Ixodicide activity of Eysenhardtia polystachya against Rhipicephalus (Boophilus) microplus.

The objective of this study was to evaluate the acaricidal effect of 2 compounds extracted from Eysenhardtia polystachya in vitro, namely coatlines and matlalines. Each extract was obtained by aqueous extraction in a nitrogen atmosphere. Engorged adult females of an amitraz-resistant strain of Rhipicephalus (Boophilus) microplus were used to produce larvae and to carry out the adult immersion test (AIT), while larvae were assayed using the larval packet test (LPT). After exposure to coatlines or matlalines, the mortality of larvae and engorged females, as well as the reproductive parameters of adult ticks were assessed. Coatlines were not lethal against larvae and did not decrease reproductive parameters. Conversely, matlalines showed a 90 to 100% efficacy against adults and larvae at all concentrations tested and a reduction of 76.41 to 80.64% oviposition and a 48.02 to 54.86% reduction in egg hatchability. Therefore, the acaricidal activity of matlalines was more efficient. Further studies are required to elucidate both the mode of action of matlalines as well the structure-activity relationships responsible for the observed differential efficacy between these 2 related isoflavans against the cattle tick.

Effects of Exposure to Methyl Parathion on Egg Hatchability and Eggshell Chemical Composition in Podocnemis expansa (Testudines, Podocnemididae).

Eggshell evaluation may serve as an indicator of the effect of substances released in the environment, which may change eggshell shape, size, structure, and/or chemical composition. Additionally, exposure may interfere with hatching rates in contaminated eggs. The objective of this study was to better understand how exposure to the insecticide methyl parathion interferes with chemical changes in eggshells of Podocnemis expansa throughout their artificial incubation, as well as with egg hatchability. A total of 343 P. expansa eggs were collected in a natural reproduction area for the species. These eggs were transferred to and artificially incubated in the Wild Animal Teaching and Research Laboratory at Universidade Federal de Uberlândia. On the first day of artificial incubation, 0, 35, 350, and 3500 ppb of methyl parathion were incorporated to the substrate. Eggs were collected every three days for chemical analysis of eggshells. Hatchability was evaluated as the number of hatchlings in each treatment, for the eggs that were not used in the chemical analysis. Student's T-test was used for data on eggshell chemical composition, and the Binomial Test for Two Proportions was used in the hatchability analysis, at a 5% significance level. It was observed that the incorporation of methyl parathion to the substrate on the first day of artificial incubation of P. expansa eggs reduced the levels of total fat in the shells throughout their incubation, besides reducing egg hatchability.

Effects of methoprene, a juvenile hormone analog, on survival of various developmental stages, adult emergence, reproduction and behavior of Asian citrus psyllid,Diaphorina citriKuwayama

The Asian citrus citrus psyllid, Diaphorina citri Kuwayama, transmits a bacterium that causes huanglongbing in citrus. Frequent and repeated use of neurotoxic insecticides against D. citri has resulted in the development of insecticide resistance. We evaluated the effects of the juvenile hormone analog methoprene on egg hatch, nymphal development, adult emergence, reproduction and behavior of D. citri. Methoprene significantly reduced the viability of eggs that were between 0 and 4 days old. Egg hatch of 0-48-h-old and 49-96-h-old eggs was 8 and 9%, respectively, when treated with 320 µg mL(-1) of methoprene. Methoprene caused significant mortality of first-, third- and fifth-instar D. citri nymphs and reduced adult emergence as compared with controls. Methoprene caused less than 5% adult emergence when first- and third-instar stages were treated, respectively, and less than 40% adult emergence when fifth instars were treated. Reduced fertility of females was observed when they emerged from methoprene-treated fifth instars. Methoprene was effective in reducing egg hatch, suppressing nymphal development and decreasing adult emergence of D. citri under laboratory conditions. Treatment of fifth instars reduced the fertility of females. Methoprene might be a possible tool for integrated management of D. citri.

Optimal numbers of matings: the conditional balance between benefits and costs of mating for females of a nuptial gift‐giving spider

In species where females gain a nutritious nuptial gift during mating, the balance between benefits and costs of mating may depend on access to food. This means that there is not one optimal number of matings for the female but a range of optimal mating numbers. With increasing food availability, the optimal number of matings for a female should vary from the number necessary only for fertilization of her eggs to the number needed also for producing these eggs. In three experimental series, the average number of matings for females of the nuptial gift-giving spider Pisaura mirabilis before egg sac construction varied from 2 to 16 with food-limited females generally accepting more matings than well-fed females. Minimal level of optimal mating number for females at satiation feeding conditions was predicted to be 2-3; in an experimental test, the median number was 2 (range 0-4). Multiple mating gave benefits in terms of increased fecundity and increased egg hatching success up to the third mating, and it had costs in terms of reduced fecundity, reduced egg hatching success after the third mating, and lower offspring size. The level of polyandry seems to vary with the female optimum, regulated by a satiation-dependent resistance to mating, potentially leaving satiated females in lifelong virginity.

A single hot event stimulates adult performance but reduces egg survival in the oriental fruit moth, Grapholitha molesta.

Climate warming is expected to increase the exposure of insects to hot events (involving a few hours at extreme high temperatures). These events are unlikely to cause widespread mortality but may modify population dynamics via impacting life history traits such as adult fecundity and egg hatching. These effects and their potential impact on population predictions are still largely unknown. In this study, we simulated a single hot event (maximum of 38°C lasting for 4 h) of a magnitude increasingly found under field conditions and examined its effect in the oriental fruit moth, Grapholitha molesta. This hot event had no impact on the survival of G. molesta adults, copulation periods or male longevity. However, the event increased female lifespan and the length of the oviposition period, leading to a potential increase in lifetime fecundity and suggesting hormesis. In contrast, exposure of males to this event markedly reduced the net reproductive value. Male heat treatment delayed the onset of oviposition in the females they mated with, as well as causing a decrease in the duration of oviposition period and lifetime fecundity. Both male and female stress also reduced egg hatch. Our findings of hormetic effects on female performance but concurrent detrimental effects on egg hatch suggest that hot events have unpredictable consequences on the population dynamics of this pest species with implications for likely effects associated with climate warming.

English

The prevalence of anthelmintic resistance and the consumer demand for alternative farming systems that limit the use of chemical anthelmintics has made the search for alternative gastrointestinal nematode parasites control methods crucial. Traditional medicinal/herbal plants can offer an alternative to the reliance on chemical anthelmintic drugs. This study evaluates the efficacy of crude aqueous extracts of Tephrosia vogelii Hook., Tephrosia villosa Pers., and Carica papaya Linn. leaves and Carica papaya Linn. seeds against gastrointestinal nematodes using in vitro egg hatch and larval development inhibition assays. Rectal faecal samples from sheep were subjected to parasitological examination for faecal egg counts (FEC) using the McMaster counting technique. 100 g of dried and poultice aqueous leaf extract of T. vogelii, T. villosa, C. papaya leaves and seeds was blended into liquefaction in 200 ml of distilled water then boiled at 90-100°C for 1 h and cooled. Levamisol and distilled water were used as positive and negative control in the bioassay. Egg hatch assay revealed more than 95.8% reduction in egg hatch at concentration of 500 mg/ml for dried and poultice paste of T. vogelii leaves and C. papaya seeds. Larval development inhibition assay results showed that both dried and poultice paste of T. vogelii leaves and C. papaya seeds extract yielded more than 98% inhibition at a concentration of 500 mg/ml. Based on the LD50 dried extract of C. papaya seeds was most potent extracts for the inhibition of both egg hatching (49.94 mg/ml) and larval development (49.32 mg/ml). Both poultice and dried extract for all the plants showed significant and dose dependent egg and larval development inhibition. These findings indicate that the evaluated plants have potential anthelmintic effect and could provide viable alternatives for the control of gastrointestinal helminthes in ruminants. Key words: Aqueous extracts, Anthelmintic activity, Medicinal plants, Tephrosia vogelii, Tephrosia villosa, Carica papaya.

Efficacy of the entomopathogenic fungus Metarhizium brunneum in controlling the tick Rhipicephalus annulatus under field conditions

High infectivity of entomopathogenic fungi to ticks under laboratory conditions has been demonstrated in many studies. However, the few reports on their use under field conditions demonstrate large variations in their success, often with no clear explanation. The present study evaluated the factors affecting the efficacy of the fungus Metarhizium brunneum against the tick Rhipicephalus (Boophilus) annulatus. It demonstrates how environmental conditions and ground cover affect the efficiency of the fungus under field conditions. During the summer, 93% of tick females exposed to fungus-contaminated ground died within 1 week, whereas during the winter, only 62.2% died within 6 weeks. Nevertheless, the hatchability of their eggs was only 6.1% during the summer and 0.0% during winter. Covering the ground with grass, leaves or gravel improved fungal performance. Aside from killing female ticks, the fungus had a substantial effect on tick fecundity. Fungal infection reduced the proportion of female ticks laying full-size egg masses by up to 91%, and reduced egg hatchability by up to 100%. To reduce the negative effect of outdoor factors on fungal activity, its conidia were mixed with different oils (olive, canola, mineral or paraffin at 10% v/v) and evaluated in both laboratory and field tests for efficacy. All tested oils without conidia sprayed on the sand did not influence tick survival or weight of the laid eggs but significantly reduced egghatchability. Conidia in water with canola or mineral oil spread on agarose and incubated for 18h showed 57% and 0% germination, respectively. Comparing, under laboratory conditions, the effects of adding each of the four oils to conidia in water on ticks demonstrated no effect on female mortality or weight of the laid egg mass, but the percentage of hatched eggs was reduced. In outdoor trials, female ticks placed on the ground sprayed with conidia in water yielded an average of 175 larvae per female and there was no hatching of eggs laid by females placed on ground sprayed with conidia in water with canola or mineral oils.