The human malarial parasite Plasmodium falciparum extensively modifies its host erythrocyte, and to this end, is faced with an interesting challenge. It must not only sort proteins to common organelles such as endoplasmic reticulum, Golgi and mitochondria, but also target proteins across the ‘extracellular’ cytosol of its host cell. Furthermore, as a member of the phylum Apicomplexa, the parasite has to sort proteins to novel organelles such as the apicoplast, micronemes and rhoptries. In order to overcome these difficulties, the parasite has created a novel secretory system, which has been characterized in ever-increasing detail in the past decade. Along with the ‘hardware’ for a secretory system, the parasite also needs to ‘program’ proteins to enable high fidelity sorting to their correct subcellular location. The nature of these sorting signals has remained until relatively recently, enigmatic. Experimental work has now begun to dissect the sorting signals responsible for correct subcellular targeting of parasite-encoded proteins. In this review we summarize the current understanding of such signals, and comment on their role in protein sorting in this organism, which may become a model for the study of novel protein trafficking mechanisms.

Antigenic variation of the intestinal protozoan parasite Giardia lamblia is caused by an exchange of the parasite's variant surface protein (VSP) coat. Many investigations on antigenic variation were performed with G. lamblia clone GS/M-83-H7 which produces surface antigen VSP H7. To generate novel information on giardial vsp gene transcription, vsp RNA levels were assessed by quantitative reverse transcription-(RT)-PCR in both axenic VSP H7-type trophozoites and subvariants obtained after negative selection of GS/M-83-H7 trophozoites by treatment with a cytotoxic, VSP H7-specific monoclonal antibody. Our investigation was not restricted to the assessment of the sense vsp transcript levels but also included an approach aimed at the detection of complementary antisense vsp transcripts within the two trophozoite populations. We found that sense vsp H7 RNA predominated in VSP H7-type trophozoites while sense RNA from only one (vsp IVg) of 8 subvariant vsp genes totally analysed predominated in subvariant-type trophozoites. Interestingly, the two trophozoite populations exhibited a similar relative distribution regarding the vsp H7 and vsp IVg antisense RNA molecules. An analogous sense versus antisense RNA pattern was also observed when the transcripts of gene cwp 1 (encoding cyst wall protein 1) were investigated. Here, both types of RNA molecules only appeared after cwp 1 had been induced through in vitro encystation of the parasite. These findings for the first time demonstrated that giardial antisense RNA production did not occur in a constitutive manner but was directly linked to complementary sense RNA production after activation of the respective gene systems.

Parasites inflict fitness costs on their hosts, but often the exact reduction in fitness is not well understood. We investigated the influence of infection by the gregarine genus Monocystis sp. on growth and female investment (cocoon production) of its earthworm host, Lumbricus terrestris. Earthworms (n=81) were observed in a laboratory setting for 8 months, after which parasite load was determined. The results revealed a significant negative relationship between parasite load and growth, yet no association to cocoon production was found. Although the exact nature, strength, and evolutionary consequence of reduced growth are still unclear, the results are the first indication for a clear, albeit weak effect of Monocystis on host fitness.

The taxonomy and phylogenetic relationships of fish trypanosomes are uncertain. A collection of 22 cloned trypanosome isolates from 14 species of European freshwater fish and 1 species of African freshwater fish were examined by molecular phylogenetic analysis. The small subunit ribosomal RNA (ssu rRNA) genes of 8 clones were sequenced and compared with ssu rRNA gene sequences from a wider selection of vertebrate trypanosome isolates by phylogenetic analysis. All trypanosomes from freshwater fish fell in a single clade, subdivided into 3 groups. This clade sits within a larger, robust clade containing trypanosomes from marine fish and various amphibious vertebrates. All 22 trypanosome clones were analysed by random amplification of polymorphic DNA. The resulting dendrogram shows 3 groups, which are congruent with the groups identified in the ssu rRNA gene phylogeny. Two of the groups contain the majority of trypanosome isolates and within-group variation is slight. These groups do not separate purported trypanosome species distinguished by morphology or host origin, and thus these criteria do not appear to be reliable guides to genetic relationships among fish trypanosomes. However, we suggest that the 2 groups themselves may represent different species of fish trypanosomes. The polymorphic DNA markers we have identified will facilitate future comparisons of the biology of these 2 groups of fish trypanosomes.

Blastocrithidia culicis is an insect trypanosomatid that presents bacterial endosymbionts. The cell-associated and secreted proteinases of the endosymbiont-bearing and aposymbiotic strains were compared through the incorporation of proteinaceous substrates into sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Few qualitative changes could be detected in the proteolytic zymograms in the 2 strains studied when gelatin, casein, haemoglobin or bovine serum albumin (BSA) were tested. However, the level of proteolytic activities was significantly higher in the aposymbiotic strain. Some of the B. culicis proteins reacted in Western blots with antibodies raised against gp63, a zinc-metalloproteinase, and cruzipain, a cysteinyl-proteinase, which are virulence factors of the human pathogenic trypanosomatids, Leishmania spp. and Trypanosoma cruzi, respectively. The anti-cross-reacting determinant (CRD) antibody recognized 2 polypeptides (50 and 58 kDa) in the spent culture media and in the supernatant from glycosylphosphatidylinositol-phospholipase C (GPI-PLC)-treated cells, suggesting that these proteins are GPI-anchored to the plasma membrane. In addition, the anti-gp63 reacted with the 50 kDa protein. The identification of protein homologues in trypanosomatids with distinct life-cycles may help to determine the importance of proteinases in trypanosomatids.

In this study with the filarial model Litomosoides sigmodontis, we demonstrate that the worms ingest host red blood cells at a precise moment of their life-cycle, immediately after the fourth moult. The red blood cells (RBC) were identified microscopically in live worms immobilized in PBS at 4 °C, and their density assessed. Two hosts were used: Mongolian gerbils, where microfilaraemia is high, and susceptible BALB/c mice with lower microfilaraemia. Gerbils were studied at 12 time-points, between day 9 post-inoculation (the worms were young 4th stage larvae) and day 330 p.i. (worms were old adults). Only the very young adult filarial worms had red blood cells in their gut. Haematophagy was observed between days 25 and 56 p.i. and peaked between day 28 and day 30 p.i. in female worms. In males, haematophagy was less frequent and intense. Similar kinetics of haematophagy were found in BALB/c mice, but frequency and intensity tended to be lower. Haematophagy seems useful to optimize adult maturation. These observations suggest that haematophagy is an important step in the life-cycle of L. sigmodontis. This hitherto undescribed phenomenon might be characteristic of other filarial species including human parasites.

A high degree of intra- and inter-geographical variation has been demonstrated previously in the excretory/secretory proteinases released by adult Haemonchus contortus. Proteinase activity has also been associated with host-protective ‘hidden’ antigens isolated from the gut of adult H. contortus. If similar geographical strain variation also exists within the gut-associated proteinases, this will have important implications for the development of a globally effective vaccine. The proteinases active in integral-membrane protein extracts from 3 different strains of adult H. contortus were characterized on the basis of their pH optima and molecular size. Although enzyme activity was detected over a wide pH range, the majority of proteinase activity was detected at acidic pH. Differences in specific activity and size of enzymes were observed between the 3 different parasite strains at different pH values. A high degree of conservation in reactive peptides was observed when protein extracts were probed with antisera raised to the protective hidden gut-antigen complexes isolated from the Moredun strain of H. contortus, or to bacterially expressed subcomponents thereof. Therefore, despite the observed differences in membrane-bound proteinase profiles, the similarity of the immunogenic response against these hidden antigens may be sufficient to prove protective against different geographical isolates of H. contortus.

We use results from a simulation-based model of nematode infection of sheep to refine the parameters in a simpler generic model of host–parasite population dynamics. These parameters describe the following host–parasite traits: probability of establishment of ingested larvae, mortality rate of adult parasites, and fecundity of adult female parasites. This simple model is then extended by allowing those parameters to vary amongst individual hosts. A sensitivity analysis is performed to determine which parameters have most influence on host parasite burden. The establishment parameter has the greatest effect on the peak value of parasite burden whilst the other two parameters have more effect on the duration of the burden. A comparison is made with results from the flock model after discussion of the definition of an average host. By allowing these parameters to vary simultaneously within the individual hosts we are able to reproduce the over-dispersed distribution of adult parasites frequently seen in nematode infections of sheep flocks.

During a 3-year trial of the effects of ivermectin (MectizanR) on adult worms and microfilariae of Onchocerca volvulus in the Republic of Cameroon, comparison was made between the percentages of calcified and uncalcified moribund (M) and dead (D) adult female worms dying following (a) the standard dose (150 μg/kg) given annually; (b) high doses (400, then 800 μg/kg) given annually; and (c and d) these same doses given at 3-monthly intervals. In the killing of adult female O. volvulus worms, the relative rôles of (a) natural causes; (b) a presumed, direct, anthelminthic, macrofilaricidal action of ivermectin; and (c) a potentially fatal pleomorphic ovarian neoplasm (PN), of which the incidence is increased by ivermectin treatment, are herein further investigated and discussed. It is concluded that ivermectin per se has a considerable direct macrofilaricidal action against female worms and that this lethal effect is supplemented by the drug's ability in some worms to increase the incidence, and the spread throughout the body of the worm, of the potentially fatal PN ovarian tumour. In moribund and dead ivermectin-treated female worms that were heavily invaded by PN, it is probable that the neoplasm was chiefly responsible for their death, but the additional direct anthelminthic action of the drug, which by itself has been responsible for the death of many other female worms, cannot be excluded as having played a supplementary lethal rôle. Similar problems as to the exact means by which adult female worms are killed may arise now that ivermectin is used in Africa for the mass treatment of lymphatic filariasis; or if and when the macrofilaricidal actions on O. volvulus of other drugs, which are closely related to ivermectin, come to be investigated.

Cystic echinococcosis (CE) is a high prevalence zoonosis among the rural population of Tacuarembó (Uruguay). The correlation between serological data and the incidence of risk factors was studied in a survey carried out in 1998 among rural communities where 480 individuals were examined by means of abdominal sonography (local prevalence=0·8%). Serum samples (305) were analysed by ELISA to determine specific IgG against crude antigens from Echinococcus granulosus. A total of 27 individuals exhibiting no detectable changes in abdominal sonographic examination were found to be seropositive (‘ultrasound normal group’). Of these individuals 9 were seroreactive against purified antigen B. A significant degree of correlation was found between seroreactivity and the incidence of some risk factors (CE antecedent in the family, P<0·005 and use of rural water, P<0·0001) among this group. Follow-up of individuals of the ‘ultrasound normal group’ was carried out after 2 years to evaluate the implications of this serological reactivity. No predictive value for cyst development was assessed with complementary image study; in contrast transient antibodies were observed with both crude and purified antigen as approximately 60% of individuals became negative when re-sampled.

Two types of tetraphyllidean merocercoids, Phyllobothrium delphini and Monorygma grimaldii, are well known from most cetaceans world-wide. The role of cetaceans in the life-cycle of these merocercoids is unclear because their specific identity is as yet unknown. The problem is compounded by poor descriptions of both merocercoids. We used light and scanning electron microscopy, and histological techniques to provide a thorough description of merocercoids collected from 11 striped dolphins, Stenella coeruleoalba, from the Spanish Mediterranean. We also described, for the first time, specimens of P. delphini with immature proglottides. Our merocercoids were morphologically similar to those described previously, except in the structure of the apical organ. Intra- and inter-sample variability in the morphology of the apical organ suggested that it degenerates during larval development. A subsample of 16 specimens of P. delphini and M. grimaldii was characterized for the D2 variable region of the large subunit ribosomal RNA gene (LSU) and compared with published tetraphyllidean cestode LSU sequences. P. delphini showed 2 unique signatures that differed from one another by a single base, whereas all sequences of M. grimaldii were identical. This suggests that each type may represent a single species, contrary to previous speculations based on morphological data. All merocercoid specimens formed a clade together with Clistobothrium montaukensis. Based on the low degree of divergence, all specimens of this clade are predicted to be congeneric.