Two mimotopes of Leishmania infantum identified by phage display were evaluated as vaccine candidates in BALB/c mice against Leishmania amazonensis infection. The epitope-based immunogens, namely B10 and C01, presented as phage-fused peptides; were used without association of a Th1 adjuvant, and they were administered isolated or in combination into animals. Both clones showed a specific production of interferon-gamma (IFN-γ), interleukin-12 (IL-12) and granulocyte/macrophage colony-stimulating factor (GM-CSF) after in vitro spleen cells stimulation, and they were able to induce a partial protection against infection. Significant reductions of parasite load in the infected footpads, liver, spleen, bone marrow and paws’ draining lymph nodes were observed in the immunized mice, in comparison with the control groups (saline, saponin, wild-type and non-relevant clones). Protection was associated with an IL-12-dependent production of IFN-γ, mediated mainly by CD8+ T cells, against parasite proteins. Protected mice also presented low levels of IL-4 and IL-10, as well as increased levels of parasite-specific IgG2a antibodies. The association of both clones resulted in an improved protection in relation to their individual use. More importantly, the absence of adjuvant did not diminish the cross-protective efficacy against Leishmania spp. infection. This study describes for the first time two epitope-based immunogens selected by phage display technology against L. infantum infected dogs sera, which induced a partial protection in BALB/c mice infected with L. amazonensis.

The objective of this experiment was to detect a possible interaction between iron deficiency and intestinal nematode infections. We report on a 2×2 study where thirty-one 10-week-old pigs fed a low or a normal iron diet were infected with both Trichuris suis (4500 eggs) and Ascaris suum (1200 eggs). No significant difference was detected between diet groups with respect to parasitological parameters for A. suum or the total number of adult T. suis recovered at necropsy 10 weeks p.i. However, in the low iron group T. suis were located more proximally and the worms were increased in length. A higher proportion of pigs with initial faecal egg excretion at 6 weeks p.i. was observed in the low iron group, indicating a shortened pre-patency period. Worm fecundity and total faecal egg excretion were also highest in the low iron group. A significant correlation was found between female worm length and fecundity. The peripheral eosinophil counts were diminished in the low iron host groups. The infected low iron group experienced more severe pathophysiological changes in terms of hypoalbuminaemia and decreases in erythrocyte volumes. A significant inverse correlation existed between iron content in the bone-marrow and liver (body) store. In conclusion, iron deficiency increased the severity of T. suis infection in pigs.

The apicomplexan protozoan parasites include the causative agents of animal and human diseases ranging from malaria (Plasmodium spp.) to toxoplasmosis (Toxoplasma gondii). The complex life cycle of T. gondii is regulated by a unique family of calcium-dependent protein kinases (CDPKs) that have become the target of intensive efforts to develop new therapeutics. In this review, we will summarize structure-based strategies, recent successes and future directions in the pursuit of specific and selective inhibitors of T. gondii CDPK1.

Due to the efforts to control schistosomiasis transmission in tropical countries, a large proportion of individuals from endemic areas present low parasite loads, which hinders diagnosis of intestinal schistosomiasis by the Kato-Katz (KK) method. Therefore, the development of more sensitive diagnostic methods is essential for efficient control measures. The aim was to evaluate the accuracy of a real-time polymerase chain reaction (RT-PCR) to detect Schistosoma mansoni DNA in fecal samples of individuals with low parasite loads. A cross-sectional population-based study was conducted in a rural community (n = 257) in Brazil. POC-CCA® was performed in urine and feces were used for RT-PCR. In addition, fecal exams were completed by 18 KK slides, saline gradient and Helmintex techniques. The combined results of the three parasitological tests detected schistosome eggs in 118 participants (45.9%) and composed the consolidated reference standard (CRS). By RT-PCR, 117 out of 215 tested samples were positive, showing 91.4% sensitivity, 80.2% specificity and good concordance with the CRS (kappa = 0.71). RT-PCR identified 86.9% of the individuals eliminating less than 12 eggs/g of feces, demonstrating much better performance than POC-CCA® (50.8%). Our results showed that RT-PCR is a valuable alternative for the diagnosis of intestinal schistosomiasis in individuals with very low parasite loads.

Leishmania parasites alternate in their life cycle between promastigote stages that develop in the gut of phlebotomine sand flies and amastigotes residing inside phagocytic cells of vertebrate hosts. For experimental infections of sand flies, promastigotes are frequently used as this way of infection is technically easier although ingestion of promastigotes by sand flies is unnatural. Here we aimed to answer a critical question, to what extent do promastigote-initiated experimental infections differ from those initiated with intracellular amastigotes. We performed side-by-side comparison of Leishmania development in Phlebotomus argentipes females infected alternatively with promastigotes from log-phase cultures or amastigotes grown ex vivo in macrophages. Early stage infections showed substantial differences in parasite load and representation of morphological forms. The differences disappeared along the maturation of infections; both groups developed heavy late-stage infections with colonization of the stomodeal valve, uniform representation of infective metacyclics and equal efficiency of transmission. The results showed that studies focusing on early phase of Leishmania development in sand flies should be initiated with intracellular amastigotes. However, the use of promastigote stages for sand fly infections does not alter significantly the final outcome of Leishmania donovani development in P. argentipes and their transmissibility to the vertebrate host.

The family Hippoboscidae is a less known group of blood-sucking flies. Deer ked are particularly important for animal health; they may act as potential vectors of disease to ungulates, and may transmit pathogens to animals and humans. The aim of this study was to investigate the presence of Trypanosoma (Megatrypanum) DNA in deer keds using molecular methods. Results prove the presence of Megatrypanum trypanosome DNA in the studied winged adult deer keds and this is the first detection of this pathogen in Lipoptena fortisetosa. In addition, this paper evidences the occurrence of L. fortisetosa in two new locations: one in the Białowieża Primeval Forest, and another in the Strzałowo Forest Inspectorate (Piska Forest), both in north-eastern Poland.

New ideas for diagnostics in clinical parasitology are needed to overcome some of the difficulties experienced in the widespread adoption of detection methods for gastrointestinal parasites in livestock. Here we provide an initial evaluation of the performance of a newly developed automated device (Telenostic) to identify and quantify parasitic elements in fecal samples. This study compared the Telenostic device with the McMaster and Mini-FLOTAC for counting of strongyle eggs in a fecal sample. Three bovine fecal samples were examined, in triplicate, on each of the three fecal egg-counting devices. In addition, both manual (laboratory technician) and automated analysis (image analysis algorithm) were performed on the Telenostic device to calculate fecal egg counts (FEC). Overall, there were consistent egg counts reported across the three devices and calculation methods. The Telenostic device compared very favourably to the Mini-FLOTAC and McMaster. Only in sample C, a significant difference (P < 0.05) was observed between the egg counts obtained by Mini-FLOTAC and by the other methods. From this limited dataset it can be concluded that the Telenostic-automated test is comparable to currently used benchmark FEC methods, while improving the workflow, test turn-around time and not requiring trained laboratory personnel to operate or interpret the results.

The Index of Association (IA) has been proposed by Maynard Smith et al. (1993) as a general method for characterizing the population structures of microorganisms as either: clonal, epidemic, cryptic species or panmictic. With reference to the current debate surrounding the mode of reproduction in parasitic protozoa, this study explores (i) the suitability and limitations of the IA for characterizing populations of Trypanosoma brucei s.l., and (ii) the idea that the significance of genetic differences between populations may be better understood if the evolution, spread and temporal stability of certain parasite genotypes are also considered. Four populations of T. brucei from Côte d'Ivoire, Uganda and Zambia are analysed using the IA and a complementary test for linkage disequilibrium, test f of Tibayrenc, Kjellberg & Ayala (1990). The two populations from Uganda are characterized as epidemic, while the others appear more or less clonal; the merits of the two methods are compared. The implications of the various population classifications are discussed with reference to genotype longevity in each region; the evolutionary and biomedical consequences of the genetic non-homogeneity of T. brucei are reviewed.

Praziquantel is widely used for treatment of schistosomiasis. Elimination of intravascular worms involves a combination of drug-induced changes at the worm surface and a host immune response against exposed parasite components. Loss of spines is a typical feature of schistosomes obtained from praziquantel-treated infected animals. The mechanism for this is unknown. We have considered the possibility that praziquantel treatment involves exposure of surface spines consisting of ‘paracrystalline’ actin and a subsequent host response against actin through mechanisms established in the host independently from schistosomal infection. Drug-induced exposure of actin was demonstrated by fluorescence microscopy using anti-actin antibodies and phallacidin, an actin-binding mushroom toxin. Actin spines remained intact at the schistosome surface after in vitro exposure, but spine morphology was lost after in vivo exposure to praziquantel. Disintegration of spines in vivo was associated with binding of host antibodies. In vitro spine destruction could be seen in the presence of normal human serum. The effect was linked to calcium-dependent binding of actin depolymerizing factor, gelsolin.

Three satellite DNAs previously isolated from the entomopathogenic nematodes Steinernema carpocapsae, Heterorhabditis bacteriophora and Heterorhabditis indicus give hybridization signals only with the S. carpocapsae, H. bacteriophora and H. indicus populations tested, indicating that these satellite sequences are species-specific. Because of their reiteration and their variabilities, we have shown that these sequences are able to discriminate at the interspecific level between the Steinernema and Heterorhabditis species, but also at the intraspecific level between S. carpocapsae strains. Furthermore, in simple squashed nematode experiments, we are able to unambiguously identify S. carpocapsae, H. bacteriophora and H. indicus populations. This last procedure is effective even on a single infective juvenile, with the main advantage that it avoids time-consuming DNA extractions.

1. Aphidius matricariae Hal. is an internal parasite of Myzus persicae Sulz.; it was also found parasitizing M. circumflexus Buckt. and Aulacorthum solani Kalt. (Myzus pseudosolani Theob.).

2. The percentage of aphides successfully parasitized increased with an increased number of strikes per host. A single female can lay up to 350 eggs, and one female successfully parasitized 309 M. persicae.

3. The female parasite reaches its maximum activity during the 2nd or 3rd day of oviposition, and this activity has no correlation with light intensity.

4. The disappearance of supernumerary larvae due to cannibalism has been observed.

5. The species is arrhenotokous. Mating takes place from 2 hr. after emergence. In the progeny of early matings the females preponderated; in later matings the sex ratio was approximately 1:1.

6. The parasite appears to fly under the same conditions as those generally favourable for aphides. It is generally disseminated by flight of the adult itself and through localized movements of parasitized apterous aphides.

7. The host is unable to reproduce if ‘struck’ in the 3rd instar or earlier.

An account is given of the spontaneous occurrence in the Anglo-Egyptian Sudan of strains of the camel trypanosome, Trypanosoma evansi, devoid of kinetonuclei.

These cases, of which four are recorded, were observed in widely separated localities within the enzootic area of camel trypanosomiasis.

A critical review is given of the question of “akinetonucleate” trypanosomes in general—in strains produced experimentally and in those occurring spontaneously—with special reference to their mode of origin and their significance in the production of new races.

The prevalence of “akinetonucleate” forms in normal strains of trypanosomes has been estimated in the case of a number of species and races hitherto not examined from this point of view, all the existing data being brought together in tabular form.

While the factors causing the appearance of “akinetonucleate” individuals and strains under natural conditions could not be ascertained, irregular division of the trypanosomes is held to be mainly responsible for the elimination of the kinetonucleus and for the persistence of the aberrant condition.

Leishmania spp. are intracellular protozoan parasites that are delivered within the dermis of their vertebrate hosts. Within this peripheral tissue and the draining lymph node, they find and/or rapidly create dynamic microenvironments that determine their ultimate fate, namely their more or less successful expansion, and favour their transmission to another vertebrate host though a blood-feeding vector. Depending on their genetic characteristics as well as the genetic make-up of their hosts, once within the dermis Leishmania spp. very rapidly drive and maintain sustained T cell-dependent immune responses that arbitrate their ultimate fate within their hosts. The analysis of the parasitism exerted by Leishmania major in mice of different genetic backgrounds has allowed us to recognize some of the early and late mechanisms driven by this parasite that lead to either uncontrolled or restricted parasitism. Uncontrolled parasitism by Leishmania major characterizing mice from a few inbred strains (e.g. BALB/c) is associated with the expansion of parasite reactive Th2 CD4 lymphocytes and results from their rapid and sustained activity. In contrast, restricted parasitism characteristic of mice from the majority of inbred strains results from the development of a polarized parasite-specific Th1 CD4 response. This murine model of infection has already been and will continue to be particularly instrumental in dissecting the rules controlling the pathway of differentiation of T cells in vivo. In the long run, the understanding of these rules should contribute to the rational development of novel immunotherapeutic interventions against severe infectious diseases.

We analysed the patterns of tick distribution on 2274 adult toads from Venezuela and Brazil, to explore whether these ectoparasites have any impact on the survival of Bufo marinus. A maximum-likelihood analysis showed that aggregation levels of ticks decreased significantly with the mean intensity of infection. This decline could be attributed to a density-dependent reduction of ticks within toads, density-dependent tick-induced toad mortality and/or density-dependent tick-induced changes in toad susceptibility. However, the relationship between the rate of change in tick loads and tick burdens from recaptured toads indicated that neither the loss of ticks within toads nor the toad susceptibility to further infection were dependent upon tick burdens. Therefore, we can indirectly infer that density-dependent tick-induced toad mortality is responsible for the observed decline in aggregation levels with tick age and burdens. On the other hand, a significant negative relationship between tick burdens and the size-specific weight of toads suggested that ticks may also have a significant impact on the patterns of weight deposition of adult toads. This evidence suggests that these ectoparasites may play an important role in regulating the densities of B. marinus in native habitats.

Attenuated lines of Eimeria acervulina were isolated between 62 and 72 h post-infection from the Houghton (H) strain. The inoculation of small numbers of oocysts of precocious (HP) lines gave substantial protection to Light Sussex chicks kept on litter against challenge with the virulent H strain. The precocious trait of the 72 h HP line was shown to be stable because the kinetics of oocyst production remained unaltered after 9 consecutive passages through birds in which only late developing oocysts were used for passage. The precocious 62 h HP line was subjected to further selection for 5 consecutive passages. The resultant (62 A HP) line was shown to be more attenuated than the 72 h HP line but remained capable of immunizing chicks against challenge with the H strain.

Young rabbits were infected by mouth with sporulated Eimeria stiedae oocysts and killed at 1, 3, 5, 8, 12, 16, 24, 30, 48, 54, 72, 84, and 96 h after infection. Sporozoites were found in the lamina propria of the duodenum between 5 and 8 h after infection. Sporozoites were present in the mesenteric lymph nodes, both free and within lymphatic monocytes at between 12 and 84 h after infection. No sporozoites were observed in duodenal portal blood. These observations provide morphological evidence compatible with a lymphatic route of migration between duodenum and mesenteric lymph nodes, probably within lymphatic monocytes. The route to the bile duct epithelium is discussed and a subsequent migration in the portal blood suggested.

I wish to express my gratitude to Dr D. F. Kelly of the School of Veterinary Medicine, Cambridge, for his guidance and constructive criticism, and to Dr Elaine Rose of the Houghton Poultry Research Station, St Ives, for her invaluable advice on technical problems and for donating the oocysts for the study. I should also like to thank Dr E. A. Pierce of the Department of Animal Physiology, Babraham, and Mr P. Long of Houghton for their advice.

This study was carried out as a Research Project during the Natural Sciences Tripos, Part II Pathology course 1965, in the Department of Pathology, Cambridge.

Community data on the prevalence of helminth infections is important for guiding health policy, but expensive to collect. As a result most surveys focus on school-aged children, frequently using schools as a sentinel population. Since there already exists a vast amount of data on infection levels in school-aged children, but limited community-based data, we undertook a literature search on age-stratified infection data for intestinal nematode infections and schistosomiasis in Africa, to investigate whether estimates of the prevalence of infection in school-aged children could provide an index for determining community prevalence. The observed data on prevalence of infection in infants, school-aged children and adults were fitted using linear and logistic regression models which take into account variation in sample prevalences. Despite the wide variation in study sites, the observed relationship between community prevalence and school-aged prevalence was remarkably consistent for each parasite species. The prevalence of infection in school-aged children alone was shown to be higher than the predicted prevalence in the community, but the degree of overestimation was dependent on the parasite species and the level of infection. The results suggest that the prevalence of infection in school-aged children could provide a cost-effective predictive tool which can be used at a district/national level to identify target areas for control and to evaluate the numbers at risk of infection.

A new haemogregarine, Schellackia landauae sp.nov., is described in the Brazilian lizard Polychrus marmoratus (Linn) from Pará State, north Brazil. Sporozoites are found principally in the red blood cells (84%) in the peripheral blood but also occur in lymphocytes and monocytes. Experimental transmission to three uninfected P. marmoratus was achieved after feeding them with laboratory-bred Culex pipiens fatigans which had engorged on an infected lizard 14 days previously. The cycle of development in the small intestine of P. marmoratus takes approximately 30 days: schizogony, gametogony and fertilization of the macrogametocytes is in the epithelial cells of the gut, with zygotes penetrating the lamina propria, where the mature oocysts develop. Living oocysts average 14·3 × 13·3 μm, and are approximately 10·0 μm as seen in histological sections. During the period of intestinal development, the parasite also undergoes asexual multiplication within cells of the spleen and liver by a process which appears to be endodyogeny. The exact time required before the first invasion of the peripheral blood by the sporozoites remains to be ascertained, but is some time within 30–45 days after the lizard ingests the infected mosquitoes. Morphology of the sporozoite in the vertebrate host is variable and depends on the host cell occupied; there may be 1 or 2 refractile bodies. Reduction or fusion of these to a single refractile body in those sporozoites within the gut cells of the infected mosquito suggests that the bodies may represent some form of energy source that is used up during this latent phase in the insect vector. Accumulation of sporozoites takes place in the reticulo-endothelial cells of the viscera, in particular the pigmentladen cells of the liver and lung. Attempts to infect other species of lizards, Tropidurus torquatus (Iguanidae) and Ameiva ameiva (Teiidae) failed.

The amount of [14C]methionine in the stomach, anterior and posterior intestine wall and lumen, blood and caecum of a rat, and in the anterior and posterior parts of the tapeworm Hymenolepis diminuta in the intestine, was measured at intervals from 10 min to 10 h after feeding a meal of 2 g glucose and 37.5 mg methionine by stomach tube.

Nearly all the meal left the stomach within 5 h; the amount entering the tapeworm depended on the position of the worm. That part lying in the anterior half of the intestine was subjected to a rapid influx reaching a maximum concentration of at least 8 mM within 1–3 h after the meal was administered. The level in the worm then dropped to about 20 % of the peak level, although the stomach was still emptying and the level of methionine in the intestine wall was still rising. This was a stable state, and equal to the concentration in the posterior part of the worm which was reached 2 h after the rat was fed.

The reason why the anterior part of the worm should show a big influx and efflux is discussed and related to changes in concentration of methionine in the lumen. The lack of such a flux into and out of the worm further down the intestine is related to the host's homeostatic control of amino acid balance in the intestine.

The results prove that dietary imbalance of amino acids affect the amino acid pool in a tapeworm for about 3 h, if it lies in the anterior of the gut. A dietary regime is suggested which should produce a prolonged effect and thereby retard growth of the worm.

The weights of the parts of the worms found in the anterior and posterior half of the small intestine revealed an unexpected phenomenon; the majority of the tapeworm, by weight, lies in the anterior of the intestine for about 4 h following a meal, but in the posterior of the intestine after the stomach has emptied.

It is a great pleasure to thank Miss Gillian Moore and Miss Patricia Grant for technical assistance, and Shell International Petroleum Company, London, for financial support.

This study applies flexible statistical methods to morphometric measurements obtained via light and scanning electron microscopy (SEM) to discriminate closely related species of Gyrodactylus parasitic on salmonids. For the first analysis, morphometric measurements taken from the opisthaptoral hooks and bars of 5 species of gyrodactylid were derived from images obtained by SEM and used to assess the prediction performance of 4 statistical methods (nearest neighbours; feed-forward neural network; projection pursuit regression and linear discriminant analysis). The performance of 2 methods, nearest neighbours and a feed-forward neural network provided perfect discrimination of G. salaris from 4 other species of Gyrodactylus when using measurements taken from only a single structure, the marginal hook. Data derived from images using light microscopy taken from the full complement of opisthaptoral hooks and bars were also tested and nearest neighbours and linear discriminant analysis gave perfect discrimination of G. salaris from G. derjavini Mikailov, 1975 and G. truttae Gläser, 1974. The nearest neighbours method had the least misclassifications and was therefore assessed further for the analysis of individual hooks. Five morphometric parameters from the marginal hook subset (total length, shaft length, sickle length, sickle proximal width and sickle distal width) gave near perfect discrimination of G. salaris. For perfect discrimination therefore, larger numbers of parameters are required at the light level than at the SEM level.