Technology transfer to the developing world: does new technology have any relevance for developing countries?

Technology is not limited to equipment and commodities but includes know-how, understanding and the ability to control and exploit underlying principles and processes. Diverse technologies, not only those termed 'biomedical', affect the incidence and control of all diseases including tuberculosis. 'New technology' implies something recently developed, but any technology is new to those without prior experience. For developing countries, technologic novelty is far less important than relevance, which encompasses, among other things: direct application to reducing risk of infection and disease; affordability and cost-effectiveness; saving foreign exchange; satisfying public demand with political benefit to the government; and promotion of social equity. The value of health gained by the new technology should exceed its cost, but this is difficult to measure. It is usually presumed that industrial countries are eager to export technologies, but intellectual property and patient regulations of the importing country may inhibit such transfers. Similarly, ethical issues involving protection of human subjects and informed consent may complicate clinical trials and technology assessment in the developing country environment.

Antischistosomal vaccines: beyond the laboratory.

In the effort to develop vaccines against schistosomiasis, insufficient attention has been given to the field evaluation of their safety and efficacy, and to conditions of routine use. Evaluation demands a clear case definition and precise diagnostic tools to determine the number, condition, and pathological effects of the schistosomes present in each subject. These are difficult to achieve for schistosomiasis. The trial should also assess the reduction in community transmission attributable to immunization. Ethical problems in efficacy trials include the need for subjects to be exposed to infection when means are available to minimize it, and the need to define the point at which chemotherapy must be given to subjects found to be infected. The epidemiological and economic consequences of vaccine use must be compared with alternative methods of control and with natural low-level transmission in the community. The characteristics of a desirable vaccine should be defined in advance by modelling and/or a consensus development process to provide guidance to bench researchers and to identify those candidate vaccines that should advance to field trials.

A historical perspective on international health.

Between about 1875 and the early twentieth century, most major disease pathogens were identified and their epidemiology clarified. The technical and material developments of the nineteenth century led to a form of practical internationalism that was expressed also in the health sector. The Pan American Health Organization was established in 1902 to coordinate health issues in the Western Hemisphere. The Health Office of the League of Nations, now little known, played an important role in the 1920s and 1930s, particularly in the control of epidemic diseases in Eastern Europe and later in nutrition. The general framework of the League Health Office became the basis of the World Health Organization, founded in 1946. Certain international agencies play a special role in health; of these, United Nations Children's Fund (UNICEF) and the World Bank have a worldwide role. The Institute of Nutrition of Central America and Panama (INCAP) and the International Centre for Diarrhoeal Diseases Research, Bangladesh (ICDDR,B) are examples of essentially regional organizations with considerable influence. Private voluntary organizations, missions, and foundations have also had a substantial effect on health in certain circumstances.

Why do schistosomes have separate sexes?

Paul Basch postulates that the familiar Schistosoma of humans evolved from hermaphroditic blood flukes of Mesozoic reptiles as those host became warm-blooded. The reproductive superiority that accompanied tendencies to protandry and protogyny in hermaphroditic blood flukes has led to subsequent sexual separation and dimorphism but substantial fragments of the ancestral contrasex genome persist in present-day males, as shown by common tendencies toward hermaphroditism. In present-day females the loss of the male-specific genome is far more complete and in the process of optimizing reproductive efficiency, present-day females have sacrificed many structural elements including locomotory and pharyngeal muscles. These losses have created dependency on the well-muscled male, whose primary functions seem to be compensatory; ie., physical transport of the female from the point of pairing to the point of egg deposition, stimulating growth and development by pumping blood into the female, who unpaired would starve, plus, less importantly, fertilization of the oocytes.

Schistosoma mansoni: effect of transferrin and growth factors on development of schistosomula in vitro.

We have studied the role of human transferrin and of exogenous iron on early growth and development of schistosomula of Schistosoma mansoni in vitro, as determined by developmental achievement and thymidine incorporation. To help define an adequate serumless medium we also utilized fibroblast growth factor, platelet-derived growth factor, epidermal growth factor, and multiplication-stimulating activity, all present in the hepatic environment of developing schistosomula, as well as certain commercial serum substitutes. Supplementation of basal medium with transferrin in the range of 250-1,000 micrograms/ml provided development equal or superior to serum-supplemented medium, at least within the first 2 wk of culture. Serum-free medium supplemented with Nutridoma-HU or -SP stimulated development to a lesser extent. The stimulatory effect of iron (as ferric sulfate) on thymidine incorporation was removed by deferroxamine, a chelating agent. Competitive inhibition studies with 125I-labeled and unlabeled transferrin indicated the presence of nonsaturable binding sites on schistosomula.

Schistosoma mansoni: insulin independence.

Several authors have reported that schistosomes are affected by host hormones, including insulin. We found that insulin in a 10,000-fold range of concentrations failed to affect glucose consumption of males, females, or pairs of Schistosoma mansoni incubated for periods up to 24 hr. Insulin, vasopressin, or tetradecanoyl phorbol acetate (an insulin mimic) did not affect the uptake and incorporation of [14C]glucose and [14C]deoxyglucose over the active transport range. Competitive binding studies using 125I-insulin and 125I-insulin-like growth factor II, with varying concentrations of unlabeled hormones, demonstrated only nonspecific binding to intact worms or subcellular fractions. This generalized nonspecific uptake of label was also shown by autoradiography, suggesting that specific insulin receptors are absent from S. mansoni. We conclude that plasma glucose levels are sufficient to supply the metabolic requirements of schistosomes by diffusion, and that no insulin-dependent mechanisms have evolved in these parasites.

Schistosoma mansoni: pairing of male worms with artificial surrogate females.

To determine whether male worms provide any specific polypeptides to females, we produced extruded alginate fibers that resembled the size and shape of mature female worms. Males clasped these surrogate "female worms" and remained "paired" with them for long periods. On polyacrylamide gel electrophoresis, fibers clasped for several days showed bands at approximately 40 and 46 kDa which were never found in fibers incubated in the same medium but not clasped by males. We believe that these may be substances transferred from male worms during normal pairing. Males biosynthetically labeled with [14C]leucine were permitted to pair with fibers, which took up a broad range of polypeptides visualized on long autoradiographic exposure to gels.

Schistosoma mansoni: reversible destruction of testes by procarbazine.

1. The anticancer drug procarbazine is profoundly damaging to the testes of Schistosoma mansoni when administered at 200 mg/kg or more to the mouse host. Somatic tissues appear entirely unaffected. 2. Within 2 days the meiotic process is disrupted, and primary and secondary spermatocytes and spermatids are destroyed and replaced by amorphous granular material. 3. The testes regenerate within about 15 days, apparently from surviving spermatogonial resting cells near the germinal epithelium of the testis. 4. Livers of mice treated once 7 weeks earlier have numerous egg granulomas and give rise to many miracidia, suggesting that full testis function is regained. 5. Male worms given the drug at 200 mg/kg at 19 days of age and fixed 7 weeks later have on average one testis less than control worms, indicating that about 15% of immature testes are unable to regenerate, whereas mature worms given the drug regenerate the normal number of testes. 6. The drug did not have significant antispermatogenic effects when male worms were incubated for a week with various concentrations in vitro, suggesting that a host metabolite is the active agent.

Schistosoma mansoni: nucleic acid synthesis in immature females from single-sex infections, paired in vitro with intact males and male segments.

1. The object of this study was to see whether stimulation of nucleic acid synthesis in immature females by male Schistosoma mansoni is mediated locally by contact, or is propagated systemically in the female. 2. Immature females perfused from single-sex animal infections were paired for one week in vitro with segments of males cut transversely into thirds; others were paired with intact males, or maintained without males; all were then incubated with [3H]-thymidine or tyrosine. 3. Washed females were bisected transversely and isotope uptake counted separately in the anterior and posterior halves. 4. The halves in contact with cut male segments showed significantly higher uptake of [3H]-thymidine than the non-contact halves, indicating increased DNA synthesis and cell division, but non-contact halves had greater uptake of [3H]-tyrosine. 5. Dot-blot hybridization with a female specific single stranded cDNA failed to detect production of the corresponding mRNA in females paired with male segments.

Immunocytochemical localization of regulatory peptides in six species of trematode parasites.

1. Frozen and paraffin sections of six species of trematodes: Schistosoma mansoni, S. mattheei, S. japonicum, Schistosomatium douthitti, Echinostoma paraensei and Fasciola hepatica have been incubated with antisera against leu-enkephalin, FMRF-amide, gastrin-17, luteinizing hormone releasing hormone, neurotensin, oxytocin, prolactin, substance P, thyroid stimulating hormone and cholecystokinin, using indirect immunofluorescence and biotin-avidin horseradish peroxidase detection systems. 2. Of the ten antisera tested, six (leu-enkephalin, FMRF-amide, gastrin-17, luteinizing hormone releasing hormone, substance P and cholecystokinin) showed significant immunoreactivity, primarily in the central and peripheral nervous system, and also perhaps in the osmoregulatory system of the three species of Schistosoma. 3. Immunopositive nerve fibers extended from ganglia to gut wall, uterus and vitelline follicles, and especially from subtegumental nerve plexi to sensory receptors on the surface or in dorsal nippled tubercles.

The role of Schistosoma mansoni males in feeding and development of female worms.

Female Schistosoma mansoni from unisexual infections have scant pharyngeal musculature, thin intestinal cecal walls, pale and scanty intestinal contents, and lack acidic thiol proteinase digestive enzyme as determined by indirect immunofluorescence using a monoclonal antibody. Their intake of host erythrocytes, measured by 51Cr labeling, is about one-fourth that of paired adult females, and they appear to be starved. In contrast, paired adult females have heavier pharyngeal musculature and intestinal cecal walls and abundant digestive enzyme in the anterior third of their intestinal tract. Females in worm pairs surgically transplanted into uninfected mice continued to feed, but separated females were carried into the liver and deteriorated. Adult female S. mansoni, newly separated from their male partners and incubated in vitro with labeled erythrocytes, ingested marginally fewer cells than did still-paired females, indicating their ability to continue feeding almost normally at least for a period after separation. Paired and ex-paired adult females declined similarly in feeding rate with increased time in vitro. In Schistosomatium douthitti, females grow and mature without males, the pharyngeal musculature and cecal walls are well developed, the gut is full of ingested blood, and the acidic thiol proteinase is present in both unisexual and paired female worms. There are different stimulatory pathways for growth and for reproductive maturation in S. mansoni, although both processes require physical contact with the male. We believe that the growth-stimulating function results from the muscular action of the clasping male, which helps the immature female to pump blood into her intestine, thereby overcoming a state of relative starvation.

An alginate matrix double-embedding method for paraffin sectioning of minute specimens.

A method has been devised for placing minute specimens in viscous aqueous sodium alginate solution, which is polymerized in 0.25 molar calcium chloride. The resulting matrix can then be dehydrated, embedded, and sectioned, maintaining the orientation of the specimens. The sectioned alginate can be dissolved in 0.1 molar EDTA if desired. Tissue sections respond to normal staining procedures.

Schistosoma mansoni: cholesterol uptake by paired and unpaired worms.

Mature males and females of Schistosoma mansoni were incubated for 24 hr in medium containing [3H]cholesterol. Worms thus labeled were paired for 24 hr with unlabeled partners, in vitro or in vivo by surgical implantation into hamsters. Controls consisted of additional unlabeled worms that did not pair. Scintillation counting of thin layer chromatographic separations of lipid extracts of schistosome tissues and of the culture medium indicated that [3H]cholesterol underwent no major metabolic changes during the course of the experiment. During the period of time allowed for pairing, labeled worms lost up to 65% of their [3H]cholesterol, which was detected in the pairing medium. In both unlabeled males and females which had paired with labeled partners, levels of [3H]cholesterol were higher than in unpaired controls. This suggests that normal cholesterol transfer in worm pairs is bidirectional and that it is facilitated by physical contact between juxtaposed membranes. Cholesterol exchange in schistosome worm pairs may be partly or wholly a consequence of normal tegumental turnover of the molecule.

Evidence that the reduced surface antigenicity of developing Schistosoma mansoni schistosomula is due to antigen shedding rather than host molecule acquisition.

Antibody and lectin binding characteristics of Schistosoma mansoni schistosomula maturing in vivo and in vitro were quantitatively assessed and compared in order to investigate the basis of the reduced surface antigenicity of host derived larval schistosomes. Quantitative indirect immunofluorescence assays showed that schistosomula recovered from mice at 24 h and 5-10 days post infection bound low or insignificant amounts of a variety of anti-schistosome antibodies including those from chronically infected and radiation attenuated cercariae-vaccinated mice, a vaccinated rabbit and rabbits hyper-immunized with non-living larval and adult schistosome antigen preparations. In contrast, parasites maturing in vitro continued to bind highly significant levels of each of these antibody preparations until at least 10 days post transformation. To investigate the basis of the decreased surface antigenicity of parasites maturing in vivo, 6-day-cultured parasites were injected intravenously into mice and recovered from the lungs at various times thereafter and examined for their ability to bind both anti-parasite and anti-host antibodies. After 30 min in vivo, cultured schistosomula exhibited a significantly decreased capacity to bind anti-parasite antibodies and concanavalin A (Con A), and by 16 h had lost their binding sites for fucose binding protein (FBP) as well. That this reduction in antigenicity was due to shedding of surface antigens was suggested by the observation that the reduced ability of these parasites to bind anti-parasite antibodies coincided closely with the loss of 125I-labelled surface proteins. Furthermore unlike 6 day schistosomula which had developed wholly in vivo, 6-day-cultured parasites recovered after 30 min in vivo failed to bind anti-host antibodies suggesting that in these organisms parasite antigens were not masked by host molecules. These data argue that surface antigen shedding may explain the reduced surface antigenicity of schistosomula developing in vivo. While this surface modulation apparently occurs independently of host antigen uptake, it is dependent upon an as yet unidentified host factor.

Immunocytochemical localization of ecdysteroids in the life history stages of Schistosoma mansoni.

Sporocysts, cercariae and adults of S. mansoni exhibit focal immunoreactivity against anti-ecdysone serum in an indirect immunofluorescence assay. In adult males immunoreactivity is limited to cell bodies and linear connections in the parenchyma surrounding the intestinal caeca. In both unisexual and paired mature females part of the lining of the ootype is reactive, especially near the entrance of the vitelline duct; this demonstrates that females can make ecdysteroids without mal contact. Adult worms cultured completely in vitro show a similar pattern of reactivity. Immunoreactivity is strong in cercariae, but is essentially absent in miracidia.