The technique of Random Amplification Polymorphic DNA allows fragments of the genome to be amplified by means of polymerase chain reaction (PCR) without previous knowledge of their sequences. The protozoa of the genus Leishmania present great genetic variability, making it difficult to characterize the different species. A method is developed with a single 10-mers long primer, which allows the species L. braziliensis, L. mexicana, L. infantum, L. tropica, L. chagasi, L. amazonensis and L. major to be differentiated. These products amplified by RAPD have also facilitated the design of some primers that amplify L. braziliensis DNA exclusively.

A female-specific sequence was isolated from a cDNA library of Schistosoma mansoni and further characterizedSequences reported here are available from DDBJ/EMBL/GenBank under the Accession number Z27402.. Expression of the corresponding gene (p19) depends on pairing with a male. In situ hybridization and immunohistology experiments revealed exclusive expression of the gene in the cells of the vitellarium, suggesting a function in egg formation. In addition, experimental evidence for cross-linking of the protein under oxidative conditions supports the assumption that the p19 gene may code for an egg-shell precursor protein.

The Kato-Katz method is widely used for diagnosing helminth infections in epidemiological surveys, but is known to have a low sensitivity. In the case of Schistosoma mansoni, statistical methods have been developed to compensate for the poor sensitivity, but the same is not true of any other helminth parasite, or infections with multiple-helminth species. We screened 101 schoolchildren from a rural area of Côte d'Ivoire over 5 consecutive days and made 5 Kato-Katz readings from each stool specimen. We estimated single and dual-species infections with S. mansoni and hookworm based on raw egg count data and after developing a latent-class model. The cumulative prevalence of co-infections was estimated at 9·9% after reading slides on the first day, and 57·0% after reading all 25 slides per person. The latent class model yielded a co-infection prevalence estimate of 79·6%, with marginal prevalence estimates for hookworm and S. mansoni infections of 83·9% and 91·6% respectively. The sensitivities of a single Kato-Katz thick smear for detection of S. mansoni alone, hookworms alone, or S. mansoni plus hookworms were 22·4%, 8·0% and 17·7%, respectively. In the current setting this could be attributable to low infection intensities of both parasites, combined with intra-specimen and day-to-day variation in egg output. If confirmed in other settings, these findings have implications for estimating the prevalence of multiple species helminth infections, and hence the design and implementation of efficacious and cost-effective control programmes.

Biomphalaria glabrata embryonic (Bge) cells have been shown to be a valuable in vitro cellular model for the study of snail host–parasite interactions. They both promote the growth and differentiation of various trematode species including Schistosoma mansoni, and Echinostoma caproni and share some morphological and functional features with circulating haemocytes. As an approach to investigate snail genes potentially regulated following exposure to trematode excretory–secretory (ES) products, we compared gene expression profiles of Bge cells exposed to saline solution, or saline solution containing ES products from S. mansoni or E. caproni, two trematode species parasitizing B. glabrata. Following differential display RT-PCR analysis we characterized 23 differentially displayed cDNAs and we focussed on the 5 cDNAs showing sequence similarity to known genes for expression validation. Using RT-PCR, we confirmed that ES products from S. mansoni and E. caproni differentially affect the expression levels of 4 out of the 5 transcripts. These partial transcripts corresponded to novel B. glabrata sequences, and showed significant sequence similarity to genes coding for (i) cytochrome C, (ii) methyl-binding proteins, (iii) glutamine synthetases, and (iv) protease inhibitors from the Kunitz family. The possible significance of these gene expression changes in host–parasite molecular interactions is discussed.

Studies of the kinetics of adenosine transport were carried out on the P1 and P2 transporters of drug-sensitive Trypanosoma evansi and its cloned derivative, resistant to the melaminophenyl arsenical Mel CY. Characterization of adenosine uptake was made by estimation of the maximum concentration taken up at time infinity (Amax). Amax on the P2 transporter of sensitive T. evansi was greater than Amax on the P1 transporter. Amax of the P2 transporter was significantly decreased in drug-resistant trypanosomes. The effect of adenosine concentration and inhibitors, on the rate of adenosine uptake, was described by Michaelis-Menten equations. In sensitive T. evansi, the maximum velocity of adenosine uptake (Vmax) of the P2 transporter was 2-fold greater than Vmax of the P1 transporter. The Vmax of the P2 transporter in resistant parasites was reduced 9-fold. The binding constants Km and Ki on the P2 transporter of resistant T. evansi, showed that resistance was associated with an increased affinity for adenosine, and a decreased affinity for adenine and Berenil. We suggest that resistance to melaminophenyl arsenicals in T. evansi, occurs via 2 mechanisms: (1) a reduction in the number of expressed P2 transporter molecules resulting in decreased uptake of melaminophenyl arsenicals; (2) a change in the binding properties of the P2 transporter.

The host–parasite interaction between the rainbow trout Oncorhynchus mykiss and the fish louse Argulus japonicus was investigated by administering low levels of dietary cortisol before infecting the fish with low numbers of the parasite. After 24 h, the dietary cortisol treatment elevated blood cortisol and glucose levels and stimulated the synthesis of secretory granules in the upper layer of skin cells. Infection with 6 lice per fish caused skin infiltration by lymphocytes, also in areas without parasites. The lymphocyte numbers in the blood at 48 h post-parasite infection were reduced. Other changes, typical for exposure to many stressors and mediated by cortisol, were also found in the epidermis of parasitized fish, although neither plasma cortisol nor glucose levels were noticeably affected. Glucocorticoid receptors were localized immunohistochemically and found in the upper epidermal layer of pavement and filament cells, and in the leucocytes migrating in these layers. Cortisol-fed fish had reduced numbers of parasites and the changes in the host skin are likely involved in this reduction. Thus a mild cortisol stress response might be adaptive in rejecting these parasites. Further, the data suggest that this effect of cortisol is mediated by the glucocorticoid receptor in the skin epidermis, as these are located directly at the site of parasite attachment and feeding in the upper skin cells that produce more secretory granules in response to cortisol feeding.

This study examined the route of infection by free-swimming larvae of 2 monocotylid monogeneans that inhabit the gills (Neoheterocotyle rhinobatidis) and the nasal tissue (Merizocotyle icopae) of the shovelnose ray, Rhinobatos typus, from Heron Island on the Great Barrier Reef, Australia. Larvae of N. rhinobatidis and M. icopae attached directly to the gills and the nasal tissue of the ray, respectively, and did not first settle on the skin. Initial development of the post-oncomiracidium of N. rhinobatidis was rapid and hamuli formed between 6 and 24 h p.i. at a mean temperature of 26 °C. However, growth then slowed markedly and was variable; only 2 fully mature individuals were found 20 days p.i. at a mean temperature of 24·5 °C. Development of M. icopae was slow and variable throughout; hamuli did not appear until 10 days p.i. and no mature individuals were obtained even 22 days p.i. at a mean temperature of 24·5 °C. No character could be found as an indicator of parasite age for N. rhinobatidis or M. icopae due to the high variability in development in both species.

Manipulative endoparasites can alter the behaviour and the physiology of their intermediate hosts in ways that increase the probability of successful transmission to the final host. This requires that the parasite is able to circumvent its host's immune defence. Successful immune evasion may depend on host–parasite coevolutionary history and the appearance of new hosts invading the local host population may promote local parasite maladaptation. To test this hypothesis, we examined the effect of 2 acanthocephalan parasites, Pomphorhynchus laevis and Polymorphus minutus, on the immunity of their local and new invasive gammarid intermediate hosts, respectively Gammarus pulex and Gammarus roeseli. We found that infection by each parasite was correlated with a decrease, at different degrees, of the standing level of immune defence in local hosts – measured as the phenoloxidase (PO) enzyme activity – whereas invasive hosts infected by P. laevis had their PO-enzyme activity enhanced. These results suggest that these acanthocephalans evade their local host immune response through immunosuppression but cannot evade the immune response of their new invasive host. The potential role of this maladaptation on the success of invasive species is discussed.

Two full-length beta-tubulin cDNAs, representing isotypes 1 and 2, were cloned from the cattle nematode Cooperia oncophora. The predicted protein sequences span 448 amino acids, and show a high degree of identity to beta-tubulins from other nematodes. While C. oncophora isotype 1 sequence had the highest identity to Haemonchus contortus isotype 1 and Teladorsagia circumcincta sequences (95% identity), the C. oncophora isotype 2 sequence was most similar to H. contortus isotype 2 and Trichostrongylus colubriformis (92% identity). Alignment of the two C. oncophora sequences with other trichostrongylid beta-tubulins deposited in GenBank showed a clear distinction between isotype 1 and 2 beta-tubulin classes. The two classes differed at 19 amino acid positions, most notably at the carboxy terminus. These isotype-defining residues were conserved among different trichostrongylid species within a class. Analysis of fragments of both genes revealed a high degree of genetic variability in coding and non-coding regions. However, all nucleotide differences detected in the coding region were silent, as they did not result in any amino acid substitution. Analysis of 2 groups of worms for the codon 200 polymorphism associated with benzimidazole resistance revealed a proportion of worms in 1 of the groups bearing a tyrosine at this position.

Howardula helenoschini sp. n. (Tylenchida: Allantonematidae) is described from 40 million year-old Baltic amber. The nematodes are parasitic in the body cavity of an adult fly of the genus Triphleba (Phoridae: Diptera). Characteristics of the fossil species are the large size of the pre-adult juveniles and a small number of progeny compared with extant Howardula parasites of flies. Reasons are given why these features suggest that this species was at an early stage of adapting, evolutionarily, to its insect host.

The population dynamics of Toxocara canis in pigs, and their immune response to a primary and a challenge infection, were studied by parasitological, haematological and serological parameters. Seventy pigs were divided into 4 groups; 35 pigs received a primary infection (group A), 15 pigs received both a primary and a challenge infection (group B), 15 pigs received the challenge infection only (group C), and 5 pigs served as helminth-free controls (group NC). A dose of 50000 eggs was administered for the primary infection (day 0) and a dose of 10000 eggs was given for the challenge infection (day 28). On days 7, 14, 21 and 28 p.i., 5 pigs of group A, and on days 35, 42 and 49 p.i., 5 pigs from each of groups A, B and C were necropsied. Numbers of recovered larvae varied widely among the 5 pigs of each group on all days of necropsy. Toxocara canis larvae were recovered predominantly from the lungs; migration of larvae to other organs or tissues from the lungs was restricted. In group A, the larval burden in the lungs peaked on day 14 p.i., and the larval densities decreased significantly over time. Thereafter, the majority of larvae were recovered from the lungs until the end of the experiment (day 49 p.i.). A few larvae were found in the muscles and brain until day 42 p.i., and 2 larvae were found in the eyes of 2 pigs on day 35 p.i. There was little evidence of protective immunity to a challenge infection in this experiment. The eosinophil levels tended not to increase in pigs receiving a challenge infection, in contrast to the challenge control pigs. The fact that T. canis larvae migrate and persist in the tissues of pigs for more than 1 month suggests a zoonotic risk in infected pigs. The relevance of these data to the population biology and immunology of porcine and human toxocarosis is discussed.

Analyses of small subunit ribosomal RNA gene sequences of representatives of major taxa of Monopisthocotylea were performed to identify the sister group of Gyrodactylus. Nuclear ribosomal DNA sequencesNucleotide sequence data reported in this paper are available in the GenBank, DDBJ and EMBL databases under the accession numbers AJ566375-79, AJ566768, AJ567670-74, AJ581657. from the complete internal transcribed spacer (ITS) region were used to infer phylogeny of 37 Gyrodactylus species and Gyrodactyloides bychowskii, Macrogyrodactylus polypteri and Gyrdicotylus gallieni, using maximum likelihood, parsimony and Bayesian inference. The genus Gyrodactylus appeared to be a monophyletic group in all analyses, based on the present data set. Within the genus, there were 3 major groups recognized by high bootstrap values and posterior probabilities. None of the 6 subgenera appeared to be monophyletic, and the most basal subgenus G. (Gyrodactylus) was paraphyletic. Characteristics of the excretory system of Gyrodactylus do not seem to be conservative enough to reveal subgenera within Gyrodactylus and we suggest abandoning existing subgenera as indicators of phylogeny. The grouping of species based on the morphology of the ventral bar and marginal hooks seems to have sufficient power to infer relationships between the Gyrodactylus species.