Prevalence of anthelmintic resistance of gastrointestinal nematodes in Polish goat herds assessed by the larval development test.

BACKGROUND: Helminthic infections, in particular those caused by gastrointestinal nematodes (GIN), are found worldwide and are among the most economically important diseases of goats. Anthelmintic resistance (AR) in GIN of goats is currently present worldwide, and single- or multidrug resistant species are widespread. The aim of this study was to determine the prevalence of AR to benzimidazoles (BZ), macrocyclic lactones (ML) and imidazothiazoles represented by levamisole (LEV) in the Polish goat herds by using an in vitro larval development test, which is useful especially in large-scale epidemiological surveys. RESULTS: This cross-sectional study was conducted from September 2018 to June 2019 and enrolled 42 dairy goat herds scattered over the entire country. The most commonly used anthelmintic class in goat herds in Poland were BZ (92%), followed by ML (85%) and LEV (13%). BZ-resistant GIN populations were found in 37 herds (88%, CI 95%: 75 to 95%), ML-resistant GIN populations in 40 herds (95%, CI 95, 84 to 99%), and LEV-resistant GIN populations in 5 herds (12%, CI 95%: 5 to 25%). Multidrug resistance involving all three anthelmintic classes was found in 5 herds (12%, CI 95, 5 to 25%). Based on the morphological features of stage 3 larvae the main resistant GIN turned out to be Haemonchus contortus and Trichostrongylus spp. The use of BZ and frequency of anthelmintic treatments were significantly related to the presence of AR to BZ in Polish goat herds. CONCLUSIONS: This cross-sectional study demonstrates the existence of AR to BZ, ML and LEV on Polish goat farms. Resistance to BZ and ML is widespread, while AR to LEV is currently at a low level. A considerable proportion of herds harbours multidrug resistant GIN, which requires further consideration. An effective anthelmintic treatment strategy, reasonable preventive measures and better understanding of the resistance-related management practices by farmers and veterinarians may delay further development of AR.

Chemical characterization and in vitro biological activity of Cymbopogon citratus extracts against Haemonchus spp. and Trichostrongylus spp. nematodes from sheep.

Medicinal plants have been the focus of several studies due to their nematicide properties which can be used to control nematodes in sheep. No study has examined the morphological effects of Cymbopogon citratus on nematodes. Thus, this study evaluated the chemical composition, nematicidal activity and effects of C. citratus extracts on the morphology of eggs and infective larvae (L3) of sheep. Aqueous and methanolic extracts and fractions of C. citratus were obtained and analysed in vitro. The C. citratus extracts were effective against Haemonchus spp. and Trichostrongylus spp. larvae and eggs. Ten fractions were obtained from C. citratus, six of which had high ovicidal activity at 1000 µg mL-1, and two fractions had high activity at all tested concentrations. The phytochemical analysis identified the presence of compounds such as terpenoids, various ketones, esters, and fatty acids. The ultrastructural analysis showed deformations of the cuticle and wilting along the body of the nematodes at all concentrations. The muscular layer, intestinal cells and the mitochondria profile showed damage compared to the typical pattern. Ultra-thin sections of eggs treated with methanolic fractions of C. citratus presented modifications. This study showed the biological activity and effects of C. citratus on the gastrointestinal nematodes in sheep.

Host immunity shapes the impact of climate changes on the dynamics of parasite infections.

Global climate change is predicted to alter the distribution and dynamics of soil-transmitted helminth infections, and yet host immunity can also influence the impact of warming on host-parasite interactions and mitigate the long-term effects. We used time-series data from two helminth species of a natural herbivore and investigated the contribution of climate change and immunity on the long-term and seasonal dynamics of infection. We provide evidence that climate warming increases the availability of infective stages of both helminth species and the proportional increase in the intensity of infection for the helminth not regulated by immunity. In contrast, there is no significant long-term positive trend in the intensity for the immune-controlled helminth, as immunity reduces the net outcome of climate on parasite dynamics. Even so, hosts experienced higher infections of this helminth at an earlier age during critical months in the warmer years. Immunity can alleviate the expected long-term effect of climate on parasite infections but can also shift the seasonal peak of infection toward the younger individuals.

In vitro anthelmintic effect of Melia azedarach L. and Trichilia claussenii C. against sheep gastrointestinal nematodes.

The control of parasitic diseases in small ruminants is mainly done with the use of synthetic anthelmintics. However, incorrect and indiscriminate use of these products has caused the emergence of parasite resistance. Plants with anthelmintic activity are used in folk veterinary medicine, but it is necessary to investigate and scientifically validate low-cost phytotherapeutic alternatives for future use to control gastrointestinal nematodes in small ruminants by family farmers. Thus, the aim of this study was to evaluate the in vitro anthelmintic effect of plant extracts from Melia azedarach and Trichilia claussenii by the egg hatch test (EHT) and larval development test (LDT) against sheep gastrointestinal nematodes. The hexane extract of M. azedarach fruits was extracted through cold percolation and the methanol extract of T. claussenii leaves was obtained by extraction at room temperature in solvents in order of increasing polarity. The efficacy results were analyzed using the Probit program of SAS. The M. azedarach extract showed a LC(50) of 572.2 µg/mL and LC(99) of 1137.8 µg/mL in the EHT, and LC(50) of 0.7 µg/mL and LC(99) of 60.8 µg/mL in the LDT. In turn, the T. claussenii extract presented a LC(50) of 263.8 µg/mL and LC(99) of 522.5 µg/mL in the EHT and LC(50) of 1.1 µg/mL and LC(99) of 26.4 µg/mL in the LDT. Comparing the extracts of the species from the Meliaceae family, T. claussenii showed greater anti-parasite potential in vitro than M. azedarach. However, studies on the isolated compounds, toxicity and administration forms to animals are also needed to validate low-cost alternative herbal remedies for use to control gastrointestinal nematodes by family farmers.

Animal grazing selectivity and plant chemistry issues impact on the potential of Rhagodia preissii as an anthelmintic shrub.

Rhagodia preissii had shown significant in vitro anthelmintic activity in a previous study, we examined the effect of including this shrub in the diet of sheep infected with Trichostrongylus colubriformis. Worm-infected merino wethers were grazed for 7 weeks on either R. preissii or annual pasture, and faecal egg counts (FECs) were conducted weekly. Plant material was collected weekly from eaten and uneaten plants, and analysed for levels of plant secondary metabolites (tannins, oxalates, saponins) and in vitro anthelmintic activity. While mean FECs were consistently lower in sheep grazing R. preissii compared to pasture (reductions of 20-74%), the differences were not significant. There was no relationship between grazing preference (eaten or uneaten) and in vitro anthelmintic activity of plant extracts. The levels of saponins and oxalates did not correlate with grazing preference or in vitro anthelmintic activity, while tannins were not responsible for the anthelmintic activity. While the identity of the grazing deterrent and in vitro anthelmintic compounds remain unknown, the presence of plants which were both highly preferred by the sheep and showed in vitro anthelmintic activity indicates a potential to develop the species as an anthelmintic shrub through selection of shrub populations dominated by such plants.

Disclosing the bioactive metabolites involved in the in vitro anthelmintic effects of salt-tolerant plants through a combined approach using PVPP and HPLC-ESI-MSn.

Strategies to reduce dependence on synthetic drugs for the treatment of gastrointestinal nematodes (GIN) infections in ruminants include the search for novel anthelmintic scaffolds on plants, yet salt-tolerant plants remain overlooked. This study aims to evaluate the in vitro anthelmintic properties of selected salt-tolerant plants against GIN, and identify the potential bioactive secondary metabolites involved. For that purpose, 80% acetone/water extracts were prepared from dried biomass of aerial organs of nine salt-tolerant plant species and tested against Haemonchus contortus and Trichostrongylus colubriformis by the Larval Exsheathment Inhibition Assay (LEIA) and Egg Hatching Inhibition Assay (EHIA). Pistacia lentiscus, Limoniatrum monopetalum, Cladium mariscus and Helychrisum italicum picardi were the most active in both GIN and life stages. To investigate the role of polyphenols in the anthelmintic activity, four selected extracts were treated with polyvinylpolypyrrolidone (PVPP), and non-treated and treated samples were further characterized by high-performance liquid chromatography with electrospray ionization mass spectrometric detection (HPLC-ESI-MSn). While polyphenols seem responsible for the EHIA properties, they are partially accountable to LEIA results. Several phenolics involved in the anthelmintic effects were identified and discussed. In sum, these species are rich sources of anthelmintic compounds and, therefore, are of major interest for nutraceutical and/or phytotherapeutic applications against GIN in ruminants.

Cyclotide interactions with the nematode external surface.

Cyclotides are a large family of cyclic cystine knot-containing plant peptides that have anthelminthic activities against Haemonchus contortus and Trichostrongylus colubriformis, two important gastrointestinal nematodes of sheep. In this study, we investigated the interaction of the prototypic cyclotide kalata B1 with the external surface of H. contortus larvae and adult worms. We show that cyclotides do not need to be ingested by the worms to exert their toxic effects but that an interaction with the external surface alone is toxic. Evidence for this was the toxicity toward adult worms in the presence of a chemically induced pharyngeal ligature and toxicity of cyclotides toward nonfeeding larval life stages. Uptake of tritiated inulin in ligated adult worms was increased in the presence of cyclotide, suggesting that cyclotides increase the permeability of the external membranes of adult nematodes. Polyethylene glycols of various sizes showed protective effects on the nonfeeding larval life stage, as well as in hemolytic activity assays, suggesting that discrete pores are formed in the membrane surfaces by cyclotides and that these can be blocked by polyethylene glycols of appropriate size. This increased permeability is consistent with recently reported effects of cyclotides on membranes in which kalata B1 was demonstrated to form pores and cause leakage of vesicle/cellular contents. Our data, together with known size constraints on the movement of permeants across nematode cuticle layers, suggest that one action of the cyclotides involves an interaction with the lipid-rich epicuticle layer at the surface of the worm.

Bayesian paternity analysis and mating patterns in a parasitic nematode, Trichostrongylus tenuis.

Mating behaviour is a fundamental aspect of the evolutionary ecology of sexually reproducing species, but one that has been under-researched in parasitic nematodes. We analysed mating behaviour in the parasitic nematode Trichostrongylus tenuis by performing a paternity analysis in a population from a single red grouse host. Paternity of the 150 larval offspring of 25 mothers (sampled from one of the two host caeca) was assigned among 294 candidate fathers (sampled from both caeca). Each candidate father's probability of paternity of each offspring was estimated from 10-locus microsatellite genotypes. Seventy-six (51%) offspring were assigned a father with a probability of >0.8, and the estimated number of unsampled males was 136 (95% credible interval (CI) 77-219). The probability of a male from one caecum fathering an offspring in the other caecum was estimated as 0.024 (95% CI 0.003-0.077), indicating that the junction of the caeca is a strong barrier to dispersal. Levels of promiscuity (defined as the probability of two of an adult's offspring sharing only one parent) were high for both sexes. Variance in male reproductive success was moderately high, possibly because of a combination of random mating and high variance in post-copulatory reproductive success. These results provide the first data on individual mating behaviour among parasitic nematodes.

Optimal age of Trichostrongylus colubriformis larvae (L3) for the in vitro larval exsheathment inhibition test under tropical conditions.

This study identified the optimal age of Trichostrongylus colubriformis larvae (L3) under tropical conditions for the in vitro evaluation of plant extracts using the larval exsheathment inhibition test (LEIT). Two T. colubriformis isolates with different anthelmintic (AH) susceptibility status were used for this study. The L3 of both isolates were maintained on refrigeration (4-5 °C) until use. For the LEIT, the isolates were tested every week during 16 weeks, using a stock solution of Acacia pennatula acetone:water extract at different dilutions (80-1200 µg/mL). Respective positive controls (levamisole 12.5 mM) and negative controls (PBS) were included. Effective concentrations 50 % (EC50), 90 % (EC90) and respective 95 % confidence intervals (95 %CI) for exsheathment inhibition were calculated at 60 min after exposure to chlorine solution. Motility of L3 (migration percentages (M%)) was recorded weekly using the larval migration test (LMT) as an indicator of L3 fitness over time. No correlation was found between L3 age or M% and the extract's EC50 or EC90 values. However, the EC50 values for the A. pennatula extract ranged from 80 to 200 µg/mL from weeks 2-10. Beyond week 12, larval exsheathment was irregular, with higher EC50 and EC90 values and wider 95 %CI. The M% decreased below 85 % on week 7 for Paraiso isolate, and on week 10 for FMVZ isolate. A linear negative relationship was observed between the age of L3 and M% for both T. colubriformis isolates. The relationship (slope) for both isolates was similar therefore a single linear equation was estimated describing all M% data (r2 = 0.771, df = 164, P < 0.05). Thus, when using these T. colubriformis isolates under our tropical conditions for the evaluation of AH activity of plant extracts with LEIT, the optimal age of L3 is between weeks 2-7, when M% remained above 85 %. The latter may ensure consistent and reproducible exsheathment results for T. colubriformis. Each laboratory must identify optimal conditions to perform the LEIT.

Effects of host characteristics and parasite intensity on growth and fecundity of Trichostrongylus retortaeformis infections in rabbits.

Host-mediated responses and parasite density-dependent processes can have a major influence on the growth and fecundity of parasitic nematodes. However, host characteristics and parasite intensity consistently change during the course of an infection and these could affect worm length and number of eggs in a non-constant way. We used a free-living population of rabbits naturally infected with Trichostronglyus retortateformis and examined how adult nematode length and the number of eggs in utero were affected by host characteristics (i.e. age, sex, breeding status) and parasite intensity, in a seasonal environment, between 2004 and 2007. Nematode length and number of eggs in utero decreased exponentially with host age; in contrast, parasite intensity increased, peaked in juveniles and decreased in older hosts. These patterns were consistent between rabbit cohorts. A negative relationship was observed between parasite intensity and nematode length, as well as number of eggs. Nematode length was strongly affected by nematode sex and host age, while the number of eggs was mainly influenced by nematode length. The direct influence of host-mediated effects appeared quantitatively more important than parasite density dependence in controlling length and egg production in naturally infected wild rabbits. However, their relative contribution changed during the course of the infection such that, while host immunity still influenced worm numbers, the direct effect of density-dependent interactions contributed the most at high parasite intensities.

The experimental establishment of ruminant nematodes in European hares (Lepus europaeus).

The factors that control the demography of European hare Lepus europaeus populations are poorly understood, but it has been recognized that the decline of hares in Europe is associated with an increasing intensity of agricultural activity. Many mechanisms have been suggested. We propose another mechanism; a negative impact arising from ingestion of the infective larvae of ruminant livestock. We dosed juvenile hares from a worm-free colony with a conservative dose from a mixed culture of infective larvae of the nematode parasites of sheep and cattle. We examined the hares post-mortem for the establishment of those ruminant nematodes, differences in weight changes, and the shedding of eggs. We found that under the circumstances of our trial, Trichostrongylus colubriformis and to a lesser extent T. rugatus, T. vitrinus, and Teladorsagia circumcincta were able to establish as adults in the dosed animals. We found strongyle eggs in the faeces of the dosed hares, and were able to culture larvae from those eggs. However, the ecological significance of our findings, if any, remains to be elucidated. Because of their mobility, hares may transmit resistant strains of parasites between grazing properties.

The anthelmintic activity of the cyclotides: natural variants with enhanced activity.

The cyclotides are a family of backbone-cyclised cystine-knot-containing peptides from plants that possess anthelmintic activity against Haemonchus contortus and Trichostrongylus colubriformis, two important gastrointestinal nematode parasites of sheep. In the current study, we investigated the in vitro effects of newly discovered natural cyclotides on the viability of larval and adult life stages of these pests. The natural variants cycloviolacin O2, cycloviolacin O3, cycloviolacin O8, cycloviolacin O13, cycloviolacin O14, cycloviolacin O15, and cycloviolacin O16 extracted from Viola odorata showed up to 18-fold greater potency than the prototypic cyclotide kalata B1 in nematode larval development assays. Cycloviolacin O2 and cycloviolacin O14 were significantly more potent than kalata B1 in adult H. contortus motility assays. The lysine and glutamic acid residues of cycloviolacin O2, the most potent anthelmintic cyclotide, were chemically modified to investigate the role of these charged residues in modulating the biological activity. The single glutamic acid residue, which is conserved across all known cyclotides, was shown to be essential for activity, with a sixfold decrease in potency of cycloviolacin O2 following methylation. The three lysine residues present in cycloviolacin O2 were acetylated to effectively mask the positive charge, resulting in a 18-fold decrease in anthelmintic activity. The relative anthelmintic activities of the natural variants assayed against nematode larvae correlated with the number of charged residues present in their sequence.

Effects of copper oxide wire particle bolus therapy on trichostrongyle fecal egg counts in exotic artiodactylids.

Four species of artiodactylids (scimitar-horned oryx [Oryx dama]), roan antelope [Hippotragus equinus], blackbuck [Antilope cervicapra]), and blesbok [Damaliscus pygargus phillipsi]) totaling 13 animals were treated with a one-time 12.5-g dose of copper oxide wire particles (COWPs) in a bolus form. Pretreatment, individual trichostrongyle fecal egg counts (FECs) were performed using the McMaster technique. Individual posttreatment FECs were performed every 7 days for 35 days beginning 7 days after bolus administration, and FEC reduction ratios (FECRRs) expressed as percentage reductions from pretreatment values were calculated every 7 days. Mean FECRRs for the 13 animals were 93% +/- 16%, 98% +/- 7%, 91% +/- 28%, 94% +/- 16%, and 90% +/- 13% at 7, 14, 21, 28, and 35 days posttreatment, respectively. These data demonstrate that COWPs in a bolus form were an effective method for reducing FEC in exotic artiodactylids. Based on this limited data, COWPs show promise as an anthelmintic alternative for exotic artiodactylids in zoologic collections.

Anthelmintic resistance in goat herds-In vivo versus in vitro detection methods.

Anthelmintic resistance (AR) is a serious threat to animal health and has a major economic impact worldwide due to production and financial losses. The aim of this study was to determine the occurrence of AR on 30 goat farms in Slovakia during the pasturing seasons and to compare three widely used in vitro and in vivo methods for detecting AR in field conditions. A three-year survey was conducted during the pasturing seasons of 2014-2016. Goats on each farm were split into treated and control groups and were treated by recommended (5 mg/kg body weight) and double doses (10 mg/kg b.w.) of albendazole. Comparisons between percent reduction in a faecal egg count reduction test (FECRT) and an egg hatch test (EHT) and the presence of L3 larvae in a larval development test (LDT) using resistant concentrations of benzimidazole (BZ) were monitored after treatment. The FECRT indicated percent reductions of 69.2-86.2% for the single dose and of 36.3-45.4% for the double dose. The EHT indicated that all farms had BZ-resistant nematodes. Low (<15% hatching) and high (>15% hatching) levels of resistance were detected on 13 and 17 farms, respectively. The LDT failed to detect resistant larvae on seven farms but detected low and high levels of resistance on seven and 14 farms, respectively. The data indicate a moderate correlation between in vitro and in vivo tests for detecting BZ resistance among the 30 goat farms. The hatching detected by the EHT and the presence of L3 larvae by the LDT at resistant BZ concentrations provided reasonable identification of low levels of resistance in the parasite populations, but the use of a double dose for a treatment may underestimate the real occurrence of low levels of resistant parasites on goat farms.

Genomic characterization of Tv-ant-1, a Caenorhabditis elegans tag-61 homologue from the parasitic nematode Trichostrongylus vitrinus.

A full-length cDNA (Tv-ant-1) encoding an adenine nucleotide translocator (ANT or ADP/ATP translocase) (Tv-ANT-1) was isolated from Trichostrongylus vitrinus (order Strongylida), an economically important parasitic nematode of small ruminants. The uninterrupted open reading frame (ORF) of 894 nucleotides encoded a predicted protein of 297 amino acids, containing characteristic motifs [RRRMMM] and PX(D,E)XX(K,R). Comparison with selected sequences from the free-living nematode Caenorhabditis elegans, cattle and human showed that Tv-ANT-1 is relatively conserved. Sequence identity was the greatest in and near the consensus sequence RRRMMM, and in the six hydrophobic regions predicted to be associated with alpha-helices and to traverse the cell membrane. Phylogenetic analyses of selected amino acid sequence data, using the neighbor-joining and maximum parsimony methods, revealed Tv-ANT-1 to be most closely related to the molecule (Ce-ANT-3) inferred from the tag-61 gene of C. elegans. Comparison of the genomic organization of the full-length Tv-ant-1 gene was similar to that of tag-61. Analysis of the region (5'-UTR) upstream of Tv-ant-1 identified some promoter components, including GATA transcription factor, CAAT and E-box elements. Transcriptional analysis by reverse transcription polymerase chain reaction (RT-PCR) showed that Tv-ant-1 was transcribed in all developmental stages of T. vitrinus, including the first- to fourth- stage larvae (L(1)-L(4)) as well as female and male adults. RNA interference, conducted by feeding C. elegans with double-stranded RNA (dsRNA) from Tv-ant-1 cDNA (using the homologous gene from C. elegans as a positive control), revealed no gene silencing. In spite of nucleotide identities of 100% in 23-30 bp stretches of sequence between the genes Tv-ant-1 and tag-61, these identities seem to be insufficient to achieve effective silencing in C. elegans using the parasite homologue/orthologue Tv-ant-1. This first insight into an ANT of T. vitrinus provides a foundation for exploring its role in developmental and/or survival processes of trichostrongylid nematodes.

Biological control of gastrointestinal parasitic nematodes using Duddingtonia flagrans in sheep under natural conditions in Mexico.

This investigation was aimed to evaluate the use of an oral bio-preparation containing Duddingtonia flagrans chlamydospores for the control of sheep gastrointestinal parasitic nematodes under the Mexican cold high plateau conditions. Two groups of gastrointestinal parasitic nematode naturally infected sheep, were randomly selected and located into two free-gastrointestinal nematode larvae paddocks. Group 1 received once a week a supplement containing D. flagrans chlamydospores mixed with oats and molasses. Group 2 received a similar supplement without any fungal material. After 5 months grazing animals were discarded from the experiment and two groups of free-nematode "tracer" sheep were located into the same paddocks to collect larvae from the contaminated pastures. Animals were slaughtered and necropsied and the nematodes were obtained and counted. A screening of the number of gastrointestinal nematode larvae present on the grass was performed and compared between the two grazing areas. The results showed 56% reduction in the Ostertagia (Teladorsagia) circumcincta and 94% reduction in the Nematodirus sp. population of the "tracer" sheep who grazed on the D. flagrans-treated sheep area, compared to the nematode population in animals grazed on the non-treated area. The results of the number of larvae on the grazing pastures showed a 51.1% reduction for H. contortus, and 100% for Cooperia sp. in the area with fungi. In the case of Trichostrongylus sp. no reduction was observed, when compared to the control group.

Ecology of the free-living stages of major trichostrongylid parasites of sheep.

Significant developments over recent decades make it timely to review the ecology of the major gastrointestinal nematode (GIN) species of sheep. These include the relentless development and spread of anthelmintic resistance in all of the major sheep production regions of the world, and the consequent drive towards integrated parasite management (IPM) systems incorporating non-chemotherapeutic strategies such as grazing management. The success of such programs is dependent on a detailed understanding of the environmental influences on the free-living stages of the nematode lifecycle. Major reviews of the subject were conducted prior to 1980, however considerable work has been completed since, including the development of mathematical models describing the epidemiology of GIN infection. Knowledge of the temperature thresholds for free-living development has also improved, while investigations of moisture influences and interactions with temperature have allowed more effective exploitation of environmental effects for IPM. This review re-evaluates our understanding of the factors that determine the success or failure of the free-living phases of the lifecycle in light of these developments. Temperature and moisture are the dominant influences on the free-living stages of Haemonchus contortus, Teladorsagia circumcincta and Trichostrongylus colubriformis, with the effects of pasture conditions playing a significant modulating role. Early in the free-living phase, the developmental success of the three GIN species is limited by susceptibility to cold temperatures. In general, H. contortus is most susceptible, followed by T. colubriformis and then T. circumcincta. The length of the development cycle is dependent largely on temperature, with development rate increasing at warmer temperatures. However, in order for development to proceed to the infective larval stage, addition of moisture is generally required. There has been considerably less work quantifying the effects of moisture on free-living development, although it is clear that H. contortus is most susceptible to desiccation during the pre-infective stages. Once the infective stage is reached, the influences of temperature and moisture on survival are less important, resulting in considerable survival times under conditions lethal to pre-infective stages. However, hot, dry conditions can be lethal for infective larvae of all three species, while extreme cold is also lethal with significant species variation. While the existing body of knowledge is substantial, the interpretation of many studies and comparison between them is complicated by inadequate description of, or variation in, the environmental measurements used. Confounding the effects of environmental variables on development to infective stage is the migration of larvae from the faeces and subsequent survival on pasture. There is a need to build on recent efforts to explore interaction between the effects of temperature and moisture, and also the trend to more closely simulate field conditions in laboratory studies. We propose a logical framework for future ecological investigations to overcome some of these problems, facilitate the development of a more integrated dataset on the subject and improve prediction of free-living development.

Haemonchus contortus and Trichostrongylus colubriformis in pen-trials with Javanese thin tail sheep and Kacang cross Etawah goats.

Weight gain costs due to infection were higher in sheep than goats, 28 and 17.5%, respectively, for Trichostrongylus colubriformis and 48.7 and 32.2%, respectively, for Haemonchus contortus. The extent of bodyweight cost attributed to anorexia in sheep infected with H. contortus was higher (13.5 g/day) than in sheep infected with T. colubriformis (2.3 g/day). On the other hand, bodyweight cost due to the other pathogenic effects in sheep infected with T. colubriformis were higher (35.6 g/day) compared to sheep infected with H. contortus (10.9 g/day). A strong relationship between faecal egg count and worm count (r=0.79, P=0.006) was shown only in sheep infected with T. colubriformis. About half of the infected sheep and goats had low or zero faecal egg counts throughout the study. In about 40% the egg count rose initially but became low by weeks 10-16, whereas in about 10% counts increased progressively throughout the period of observation and these animals also had the highest numbers of worms at slaughter. Packed cell volume was reduced in sheep and goats infected with H. contortus but serum protein and haemoglobin levels were unaffected. Sheep infected with T. colubriformis had a higher level of eosinophilia after 8 weeks (18.4%) than sheep infected with H. contortus (11.4%), whereas this pattern was reversed in goats and levels were also lower (4.1 and 8.9%, respectively). There was no apparent relationship between eosinophilia and resistance to infection with H. contortus or T. colubriformis.

Experimental concurrent infection of sheep with Oestrus ovis and Trichostrongylus colubriformis: effects of antiparasitic treatments on interactions between parasite populations and blood eosinophilic responses.

The purpose of this experiment was to determine if an earlier infection with Oestrus ovis would down regulate an infection with Trichostrongylus colubriformis when the larvae of O. ovis were expelled from the nasal cavities of sheep by a specific treatment. Three groups of five lambs were used: group 1 was artificially infected with O. ovis larvae and later with T. colubriformis, group 2 received O. ovis larvae and later was treated with ivermectin 14 days before being infected with T. colubriformis. Group 3 was infected with T. colubriformis only. The criteria examined were: the effects on nematode egg excretion, worm fecundity, nematode burdens and the kinetics of blood eosinophils. Significant decreases of nematode egg excretion, worm fecundity, nematode burdens were observed in group 1 compared to group 3. However, no changes were observed in either group 2 or 3. In group 2 it was noted that antiparasitic treatment induced a rapid decrease in blood eosinophils to a range close to the non-infected control group and this was associated with the removal of the down regulation effects of nematode burdens. This experiment showed that there is no cross immunity between O. ovis and T. colubriformis and that eosinophils may act against any parasite without specific priming.

In vitro activity of Peltophorum africanum Sond. (Fabaceae) extracts on the egg hatching and larval development of the parasitic nematode Trichostrongylus colubriformis.

Trichostrongylus colubriformis is an important cause of parasitic gastroenteritis in ruminants, where it causes protracted diarrhoea, rapid loss of weight, loss of production and death. The in vitro efficacy of extracts of Peltophorum africanum was determined against this parasitic nematode. Eggs and larvae of T. colubriformis were incubated at 23 degrees C in the extracts of the leaf, bark and root of P. africanum at concentrations of 0.008-25 mg ml-1 for 2 and 5 days, respectively. Thiabendazole and water were used as positive and negative controls, respectively. Inhibition of egg hatching and larval development increased significantly (P<0.05) with increasing concentrations of the extracts. Concentrations of 0.2-1.0 mg ml-1 of the extracts of leaf, stem bark, and root bark of P. africanum completely inhibited the hatching of eggs and development of larvae. No eggs and larvae of T. colubriformis could be observed in wells incubated with all the three extracts at concentrations of 5 and 25 mg ml-1. The in vitro model results support the traditional use of P. africanum against nematode parasites. Further research is required to isolate and structurally identify the active anthelmintic compounds, and to improve methods of plant extraction of the effective anthelmintic components that will be readily adaptable for use by rural communities against helminthosis.