Artemisinin (QHS) and its derivatives are new antimalarials which are effective against Plasmodium falciparum parasites resistant to chloroquine (CQ). As these drugs are introduced it is imperative that resistance is monitored. In this paper we demonstrate that the inoculum size used in in vitro testing influences the measured in vitro susceptibility to QHS and its derivative dihydroartemisinin (DHA) and to mefloquine (MEF) and CQ over the range of parasitaemias routinely used in testing with the WHO in vitro microtest. An increase in parasitaemia and/or haematocrit was accompanied by a decrease in the measured sensitivity of 2 laboratory lines. In the context of a field study testing in vitro susceptibility of parasite isolates from patients with uncomplicated malaria in Fajara, The Gambia we demonstrate that failure to control for inoculum size significantly overestimates the level of resistance to QHS and DHA as well as MEF, halofantrine (HAL) and quinine (QUIN). When controlling for the inoculum effect, cross-resistance was observed between QHS, MEF and HAL suggesting the presence of a multidrug resistance-like mechanism. These studies underline the importance of inoculum size in in vitro susceptibility testing.

The ability to increase the chances that infectious prey are taken by predators is an observed feature of many parasites that rely on one or more predator-prey relationships to complete their life-cycle. In the sylvatic life-cycle of Echinococcus multilocularis – the causative agent of human alveolar echinococcosis – foxes are the final host, with voles acting as intermediate hosts. Here we review the evidence that E. multilocularis causes increased susceptibility to predation and present a general mathematical model for the sylvatic life-cycle. The ability to increase susceptibility to predation in the intermediate host reduces the sensitivity of the parasite population to adverse conditions. For example, there is no critical density of foxes below which the parasite is expected to die out, even if the effect of the parasite on infected prey is very small. We suggest that increased susceptibility to predation is a plausible explanation for the observed resilience of E. multilocularis during and following field trials of praziquantel baiting.

Rabbit polyclonal and mouse monoclonal antibodies (Mabs) prepared against synthetic peptides patterned on exofacialloops 3 (amino acids 546–555) and 7 (821–834) of the human anion transport protein band 3 inhibited the cytoadherence of Plasmodium falciparum-infected erythrocytes to C32 amelanotic melanoma cells. Mabs directed against exofacial loop4 (amino acids 628–642) did not inhibit adherence to a significant degree. The murine Mabs recognized only P. falciparum- infected erythrocytes suggesting that the epitopes of loops 3, 4 and 7 are normally cryptic in uninfected erythrocytes.

Inhibited development of Haemonchus contortus was studied in single experimental infections of worm-free lambs. Chilling of the infective larvae at + 4°C was without effect on the percentage of larvae subsequently becoming inhibited and a period of exposure to autumnal conditions was unnecessary to induce a high rate of inhibition. It was concluded that seasonal inhibition of H. contortus in East Anglia is brought about primarily by an environmental stimulus acting upon the preparasitic stages but that, unlike Obeliscoides cuniculi and Ostertagia ostertagi, this was not cold. It could be provided in a culture kept in the dark at 25 °C for 12 days. While the age of the host did influence the phenomenon, in that larvae were less inclined to inhibition in very young animals, it was concluded that this was not a primary factor in the aetiology.

The study was conducted to evaluate a mAb-based coproantigen detection assay for monitoring fox prevalence of Echinococcus multilocularis infections in the Shiretoko National Park in Hokkaido, Japan. Eight fox families, each consisting of 2–4 adults and their pups, were selected because their territories have been determined. Faeces of each family (total n=537) were collected monthly from April to October, 1994. Detection of coproantigen and taeniid eggs was recognized in 58 and 27 faeces, respectively, but mostly restricted in 1 family, in which coproantigen ELISA OD values had dual peaks, one in June and the other in October, whereas taeniid eggs were detected only from April to July. Fox pup faeces (n=51) collected around the dens used by the positive family were all coproantigen and taeniid egg negative except for 1 faeces. These results suggest that the prevalence and infection pressure in the study area were low and the infected family terminated the infection in the summer and acquired a new one in the early autumn, 1994. This monitoring method for fox infection will be a practical management tool for E. multilocularis infection, especially at the tourist spots in Hokkaido where close contact of fox with humans is frequently observed.

Intra-clonal phenotypic (antigenic) variation is used by many pathogens to evade the consequences of immune-mediated killing by mammalian hosts. In this substantially theoretical article, I emphasise that antigenic variation (sensu stricto) involves no change in genotype; its importance as a mechanism for promoting pathogen transmission and its polyphyletic origin. From a functional perspective, antigenic variation is constrained by the requirement to meet five conditions. These are: capability to express several antigens against which functional immunity predominates; capability to interact with the environment; mutually exclusive expression of variable antigens in each cell within an infection; mutually exclusive expression in the within-host pathogen population and the capability for population growth within a host. Meeting these conditions leads to chronicity of infection and high rates of hierarchical and reversible switching of expression between variable antigens. The organisation of hierarchical expression is discussed in some detail.

Activation of casein kinase II (CK2) was one of the first observations made on how Theileria parasites manipulate host cell signal transduction pathways and we argue that CK2 induction may in fact contribute to many of the different activation events that have been described since 1993 for Theileria-infected lymphocytes such as sustained activation of transcription factors c-Myc and NF-κB. CK2 also contributes to infected lymphocyte survival by inhibiting caspase activation and is probably behind constitutive PI3-K activation by phosphorylating PTEN. Finally, we also discuss how CK2A may act not only as a kinase, but also as a stimulatory subunit for the protein phosphatase PP2A, so dampening down the MEK/ERK and Akt/PKB pathways and for all these reasons we propose CK2 as a central player in Theileria-induced lymphocyte transformation.

Only with the completion of the life cycles of Fasciola hepatica in 1883 and 30 years later those of Schistosoma japonicum (1913), Schistosoma haematobium and Schistosoma mansoni (1915) did research on schistosomiasis really get underway. One of the first papers by Cawston in 1918, describing attempts to establish the means of transmission of S. haematobium in Natal, South Africa, forms the historical perspective against which to judge where we are now. Molecular biology techniques have produced a much better definition of the complexity of the schistosome species and their snail hosts, but also revealed the extent of hybridization between human and animal schistosomes that may impact on parasite adaptability. While diagnostics have greatly improved, the ability to detect single worm pair infections routinely, still falls short of its goal. The introduction of praziquantel ~1982 has revolutionized the treatment of infected individuals and led directly to the mass drug administration programmes. In turn, the severe pathological consequences of high worm burdens have been minimized, and for S. haematobium infections the incidence of associated squamous cell carcinoma has been reduced. In comparison, the development of effective vaccines has yet to come to fruition. The elimination of schistosomiasis japonica from Japan shows what is possible, using multiple lines of approach, but the clear and present danger is that the whole edifice of schistosome control is balanced on the monotherapy of praziquantel, and the development of drug resistance could topple that.

Network theory is gaining momentum as a descriptive tool in community ecology. Because organisms with the same lifestyle can still exhibit ecological differences, it is crucial to determine the scale at which networks should be described. Here we show that networks of hosts (mammals) and parasites (ectoparasitic gamasid mites) differ when either facultative or obligatory parasites only are considered. More importantly, the structure of these networks is opposed, with obligatory parasites networks being more modular, and facultative parasites networks being more nested. Our results have consequences for the way we define which species to include in ecological networks, which we discuss in the light of community ecology and epidemiology.

Host–parasite interactions were studied for the microsporidium Pleistophora intestinalis and its host, Daphnia magna. Two host clones were established from the same population from which the parasites were taken (home-1 and 2), and two clones from two other ponds (neighbour and Munich clone). With increasing clutch number infected females from home-1 clone produced relatively smaller clutches than uninfected females. Age and body length at maturity were not affected by the infection, but body length of the sixth adult instar was reduced. In an experiment including all four host clones, the parasite reproduced well in the two home clones and in the neighbour clone, but poorly in the Munich clone. Juvenile growth and age at maturity was not affected in the two home clones, but for the neighbour and the Munich clone age was delayed by 2·2 days and 4·1 days, and juvenile growth reduced by 16 and 23%, respectively. Significant host-clone x parasite-treatment interactions were also found for size at maturity and clutch size. This pattern of host-parasite interactions suggests that there is no general positive relation between disease severity and parasite multiplication rate.

Fasciola hepatica secretes proteolytic enzymes and other molecules that are essential for host penetration and migration. This mixture may include enzymes required for the degradation of supramucosal gels, which defend epithelial surfaces against pathogen entry. These contain hydrated mucins that are heavily glycosylated. Excretory-secretory products (ES) from F. hepatica were examined for a range of glycosidase activities, using synthetic 4-methylumbelliferyl glycosides as substrates. The ES product contained at least 8 different glycosidase activities, the most abundant of which were β-N-acetylhexosaminidase, β-galactosidase and β-glucosidase. Alpha-fucosidase, β-glucuronidase, α-galactosidase, α-mannosidase and neuraminidase were also present. β-N-acetylhexosaminidase and β-galactosidase were present in multiple isoforms (at least 4), whereas β-glucosidase appeared to exist as one isoenzyme with a pI <3·8. All three enzymes had acidic pH optima (4·5–5·0). Ovine small intestinal mucin was degraded by ES at pH 4·5 or 7·0, with or without active cathepsin L, the major protease found in F. hepatica ES. The ability of F. hepatica ES to degrade mucin in the presence or absence of active cathepsin L suggests that cathepsin L is not essential for mucin degradation. The abundance of β-galactosidase and β-hexosaminidase in ES supports a role for these enzymes in mucin degradation.

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.

The effects of cold storage of infective third-stage larvae (L3) of different isolates of the parasitic nematode Haemonchus contortus were studied with respect to infectivity, pre-patent period and propensity for larval arrestment. Two complementary experiments were conducted with 2 groups of lambs, each animal being inoculated with 2000 L3 of either Swedish or Kenyan origin. In a first experiment, L3s were cold treated at 5 °C for 9 months prior to infection, whereas in a second experiment larvae were newly hatched. Individual faecal egg counts (FECs), and worm burdens were determined for each experiment. The results showed that the greatest differences were associated with the pre-treatment of larvae. The pre-patent period and the FECs differed significantly between the experiments but not between the isolates used in each experiment. However, the extent of hypobiosis was significantly different between the two isolates when fresh larvae were used (36% Kenyan isolate and 70% Swedish). The storage of H. contortus at 5 °C had no apparent effect on the infectivity of L3s, as high establishment ranging from 43 to 74% were observed, irrespective of isolates used. This study showed that H. contortus exhibited similar phenotypic traits regardless of geographical origin. Thus, there was limited evidence for adaptations to temperate climatic conditions.

1. The genus Meteterakis is reconstituted with eight species: M. govindi, M. louisi, M. baylisi, M. longispiculata, M. cophotis, M. japonica, M. mabuyi and M. triaculeata.

2. M. cophotis, M. japonica and M. longispiculata were previously placed in the genus Spinicauda.

3. M. mabuyi was previously in the genus Africana.

4. M. triaculeata was previously in the genus Ganguleterakis.

5. M. baylisi and M. louisi are new species.

6. Spinicauda bufonis and, probably, Africana varani are considered synonyms of M. govindi.

7. Africana howardi is considered a synonym of M. japonica.

8. A list of the remaining species in the genera Africana and Spinicauda is given.

A comparison of the oncomiracidia of the monogenean skin parasites Entobdella hippoglossi from the halibut (Hippoglossus hippoglossus), E. diadema from the sting ray (Dasyatis pastinaca) and E. soleae from the skin of the common sole (Solea solea) has revealed the presence of ‘body gland cells’, all of which have openings on the ventral or lateral surfaces of the larvae. In all three species there are two pairs of glands lying in the posterior region of the body proper with long ducts extending into the haptor and opening on its ventral surface. In addition to these two pairs of glands there are seven other pairs of body glands in E. hippoglossi, six pairs in E. diadema and two pairs in E. soleae. In E. soleae all the ‘body gland cells’ are poorly developed in the oncomiracidium and are more conspicuous in post-oncomiracidia. The fate of the ‘body gland cells’ during post-oncomiracidial development has been studied in E. soleae, and their possible functions are discussed. The glands in the head region (anterior median, posterior median and lateral head glands) are similar in all three species. Gland cells associated with the alimentary system are less easy to observe but differences between the three species appear to be small.

In the oncomiracidia of all three species the number of flame cells and their distribution are the same, but the oncomiracidia of E. hippoglossi and E. diadema have loops in the excretory ducts anterior to the bladder and the bladders of E. diadema contain refringent bodies.

The oncomiracidial eyes of E. hippoglossi and E. soleae have lenses but the lenses are greatly reduced in the oncomiracidia of E. diadema. In the three larvae there are small but characteristic differences in the shapes of the accessory sclerites.

The oncomiracidium of E. hippoglossi has not previously been described.

The distribution and fine structure of gland cells in the miracidium of Fasciola hepatica is described. There is a large flask-shaped, multinucleate, apical gland lying ventrally in the anterior. On each side of this are pairs of uninucleate accessory gland cells. Both apical gland and accessory gland cells communicate with the tip of the apical papilla. A third group of vesiculated gland cells opens by ducts at the base of the apical papilla. The miracidium is covered with a surface mucous coat and this appears to be directly extruded from the underlying ciliated epithelial cells. The findings are discussed with relevance to attachment to, and penetration of, the snail intermediate host.

Prevalence of infection with intestinal helminths is usually assessed through stool examination. There are limitations in using this technique to measure infection status, since some infected individuals have no patent infection. The relationship between infection assessed by stool examination and actual infection prevalence is investigated for the three major nematode species-Ascaris lumbricoides, Trichuris trichiura and the bookworms-using a model which describes the presence of non-egg producing worm combinations. The analysis demonstrates that stool examination under-estimates the actual infection prevalence, and that the degree of under-estimation is dependent on the level of infection, the nematode species and the parasite sex ratio. These findings have implications for the validity of epidemiological surveys and the evaluation of control programmes.

A monoclonal antibody against the circumsporozoite proteins of the Plasmodium berghei sporozoite was used to trace the synthesis and expression of these proteins, via the Lowicryl immunogold technique, within the developing oocyst. The proteins were detected on the endoplasmic reticulum of the oocyst and were present in the sporozoite membranes at the point of their formation.

Gregariousness can be advantageous in interspecific competition while intraspecific competition may favour solitude. We examined feeding behaviour of the ixodid tick, Rhipicephalus appendiculatus, in the context of interspecific (tick–host) and intraspecific (tick–tick) competition. Such competition is mediated through host rejection responses to tick infestation to which ticks respond by secreting immunodulatory saliva. We observed that group feeding adults increased their blood-feeding rate, reducing the time to mating and repletion, compared with individual feeding of paired adults. The benefits of feeding aggregation indicate direct reciprocity between ticks, most likely resulting from the shared activities of their bioactive saliva. However, fast-feeding ticks appeared to impair blood-feeding success of slow-feeding females during group feeding. This may be explained by the faster feeders exacerbating host responses on detachment that are then directed against the slower feeders. As female fecundity is generally proportional to the size of the bloodmeal, there will be a selection pressure to feed gregariously. Greater understanding of the benefits and costs of feeding aggregation may help to improve tick control strategies.

One α- and 2 β-tubulin isotypes (isotypes 1 and 2) from the parasitic nematode Haemonchus contortus were artificially expressed in E. coli and purified to obtain tubulin that was capable of polymerizing into microtubules. Binding of [14C] mebendazole (MBZ), a benzimidazole compound, to each individual unpolymerized isotype and to microtubules polymerized from recombinant α- and β-tubulin was assessed and Kd and Bmax values determined. Mebendazole bound to the individual tubulin isotypes with a stoichiometry of 1:1. Binding occurred with highest affinity to α-tubulin followed by β-tubulin isotype 2 and β-tubulin isotype 1 indicating that α-tubulin may play a role in benzimidazole binding to microtubules. Upon polymerization of α- and β-tubulin isotype 2 into microtubules the stoichiometry of binding increased to 2:1 (mebendazole:tubulin) while binding affinity remained the same. Mebendazole binding to α/β-isotype 1 microtubules remained unchanged following polymerization. The increase in the number of benzimidazole receptors on α/β-isotype 2 microtubules suggests the formation of a new benzimidazole receptor upon polymerization.