Skin schistosomula can be prepared by collecting them after isolated mouse skin have been penetrated by cercariae in vitro. The schistosomula can also migrate out of isolated mouse skin penetrated by cercariae in vitro and from mouse skin penetrated by cercariae in vivo. Schistosomula can also be produced from cercariae applied through a syringe or in a vortex. When certain surface properties of the different forms of schistosomula were compared, those migrating from mouse skin penetrated by cercariae in vivo or in vitro had greatly increased permeability to membrane impermeant molecules such as Lucifer yellow and high molecular weight dextrans. These migrating forms also possessed surfaces which showed greatly enhanced uptake into internal membrane vesicles of the dye FM 143, a marker for endocytosis. This greatly enhanced activity and permeability of the surfaces of tissue migrating schistosomula is likely to be of great importance in the adaptation to the new host.

It has been observed that fluorescent membrane-impermeant molecules can enter the cercariae as they penetrate mouse skin. The hypothesis to be tested was that such molecules, which included Lucifer Yellow and a variety of fluorescent dextrans, entered the parasite through the nephridiopore and excretory tubules as well as through the surface membrane. FITC-labelled poly-L-lysine (molecular weight 10 kDa), added at 4°C during syringe transformation, was found to enter the nephridiopore and labelled the excretory bladder and sometimes the excretory tubules. This finding indicates that macromolecules (10 kDa) can enter the nephridiopore. It was found that linoleic acid (a normal constituent of skin) greatly stimulated uptake of Lucifer Yellow and dextrans into the excretory/subtegumental region of 2-h-old schistosomula. This correlated with an increased uptake of membrane-impermeant propidium iodide at 37°C. Since increased uptake of propidium iodide occurs when membranes become permeable, the surface membrane could also be a pathway of transport of the membrane-impermeant molecules into the schistosomulum.

Fluorescent lipophilic compounds can be used to label the surface membrane of Schistosoma mansoni by adding the compound in small amounts of organic solvents to aqueous medium in vitro. Under these conditions it is difficult to follow routes of distribution of the label. Here we have absorbed nitrobenzoxadiazolamine methylamino–(NBD)–ceramides to positively charged Dowex beads, and incubated the labelled beads with living parasites. The NBD–ceramide transfers to the surface membrane as a patch 50–100 μm in diameter, after which the label can be seen localized in the gut and in a very concentrated form in organelles within the oesophageal gland cells. Subsequently the labelled compound can be found in organelles within other body cells, including subtegumental cells. We show that the labelled ceramide has been transported from the patch in the surface membrane through internal membrane systems to the destination in the gut and oesophageal gland and not transported through the gut via the external medium. A different pattern was observed when NBD–cholesterol was used. The pharynx was rapidly labelled when NBD–cholesterol was added in medium with or without serum or attached to red blood cells only. Diffuse labelling of the surface membrane and oesophageal gland occurred. We have demonstrated a novel route of lipid transport within the parasite. The route requires the surface membrane to have very specialized regions to facilitate such transport.

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.

In this paper we describe the effect of poly-L-lysines of different molecular weight on the schistosomula. In the control sample, the schistosomula of Schistosoma mansoni take up fluorescent Texas Red conjugated to bovine serum albumin (TxR–BSA) into the gut. Following slight damage by 24·0 kDa poly-L-lysine, a high proportion of schistosomula take up fluorescent TxR–BSA into the excretory system. Subsequently, the dye diffused into the bodies of the schistosomula. We suspected that this diffusion involved the process of endocytosis so we investigated this with the use of endocytosis inhibitor, Latrunculin A. Addition of the endocytosis inhibitor Latrunculin A following poly-L-lysine treatment inhibited gut uptake of TxR–BSA as well as the diffusion of excretory-ingested TxR–BSA molecules.

The variability within schistosome populations was explored using mixed populations of cercariae from multimiracidial snail infections and individual clones of Schistosoma mansoni cercariae obtained from monomiracidial snail infections. We investigated the heterogeneity between different clones of S. mansoni with respect to infectivity and metabolism. One difference between clones of cercariae was found in the recovery of adult worms from Balb/C mice. Recovery of adult worms was greater after infections with a mixed population than with a clonal population. To investigate some biochemical features of individuals in clones or mixed populations, the uptake of [35S]methionine into individual parasites and their membrane proteins was measured. Isoelectric focusing of a soluble membrane fraction: the frozen–thawed supernatant extracted from individual clones, showed the presence of proteins of isoelectric point between 7.2 and 8.2 in all clones. These proteins were less labelled with [35S]methionine in the clones than in the mixed population. It was concluded that basic proteins are synthesized by all clones and in the mixed population but at different rates. Differences in the rate of incorporation of [35S]methionine into the surface membranes of schistosomula and adult worms derived from individual clones are reported. In addition, a direct correlation between the percentage of recovery of adult worms from mice infected with individual clones of S. mansoni and the rate of incorporation of [35S]methionine into schistosomula of these particular clones was observed. It is suggested that the high rate of metabolism shown by an individual clone may account for the enhanced survival of the cercariae derived from that clone during penetration of the skin and migration through the vertebrate host. In order to examine individuals in a population of schistosomula, from a clone or mixed population, the lysosome-specific fluorescent probe LysoTracker DND-99 was used to label the parasites and quantitative fluorescent measurements were made on individual parasites. There were significant differences between clones and a mixed population. Furthermore, the variation between individuals from a mixed population was greater than from that in any clone, just as was found in the infectivity studies. Freshly transformed schistosomula of individual clones labelled with the LysoTracker DND-99 showed less variations in the quantitative uptake of the dye within a single clone when compared to the mixed population. We conclude that for any biochemical and biological parameter, a population of cercariae consists of individuals showing a wide range of values, with a much greater range in a mixed population. This variability is likely to have great relevance for infectivity of the final host and the efficacy of drugs and the immune system.