We tested a theoretical model based on the physics of capillary flow and confirmed that anaemia accelerates blood intake in the yellow-fever mosquito, Aedes aegypti (L.). We also investigated the influence of anaemic blood on egg production of mosquitoes and found that it has a negative influence on fecundity. Based strictly on egg production and the physics of fluid intake, we propose that although anaemia associated with blood-borne parasites may be detrimental to mosquitoes that can engorge to repletion in one session, it may be beneficial to those interrupted before repletion because the greater quantity of the bloodmeal may compensate for its lower quality. Epidemiological consequences are discussed but require further inquiry.

In the third and fourth decades of this century chemotherapy began to be established as one of the greatest success stories in medicine. Now unfortunately severe problems compromise the efficacy of drugs used to treat infectious diseases, two of the most serious handicaps being the rapidity with which target pathogens can develop drug-resistance and the slow rate at which replacement products are appearing on the market. Increased understanding of the ways in which existing drugs act may help both to prolong their usefulness and to generate novel therapeutic strategies.

Recent data indicate that platelets may play an important role in the host defence against Toxoplasma gondii infections. T. gondii-stimulated human platelets release thromboxane A2 (TXA2) and 12-hydroxyeicosatetraenoic acid (12-HETE) from arachidonic acid and 13-hydroxyoctadecadienoic acid (13-HODE) from linoleic acid (Yong et al. 1991; Henderson et al. 1992). We have previously demonstrated that the eicosanoid TXA2 has potent cytotoxic activity against T. gondii trophozoites (Yong et al. 1991). In this study, we examined whether 12-HETE, 13-HODE, and linoleic acid also have toxoplasmacidal activity. 13-HODE at concentrations ≥ 10−8 M rapidly induced cytotoxic changes in T. gondii. Ultrastructural changes induced by 13-HODE in T. gondii included an initial leakage of cytoplasmic contents into a space between the inner and outer parasite bilayer membrane units which was followed by intracellular vacuolation and loss of cytoplasmic contents. In contrast, linoleic acid and 12-HETE lacked toxoplasmacidal activity at 10−10–10−6 M concentrations. These data indicate that 13-HODE, a product of linoleic acid metabolism, has potent cytotoxic activity against T. gondii; this toxoplasmacidal activity may be important in the inflammatory response to this pathogen.

This study was undertaken to assess whether glycolipid antigens (particularly gangliosides) are associated with Pneumocystis carinii obtained from human lungs. Gangliosides were extracted, purified in high performance thin-layer chromatography and stained with resorcinol. Two resorcinol-positive bands, co-migrating with GM1 and GD1a were demonstrated, suggesting the existence of ganglioside molecules on P. carinii. No resorcinol-positive bands were revealed in the pulmonary control tissue. In addition, an antiserum obtained from rabbits immunized with P. carinii antigen reacted with gangliosides GM1, and GD1a, as revealed by a dot immunobinding assay. This reactivity was inhibited by first incubating the antiserum with ganglioside micelles. Furthermore, anti-glycosphingolipid antibodies (aGM1) reacted with the bands of 200 and 55 kDa of P. carinii antigen. These results suggest that ganglioside antigens expressed on P. carinii can trigger specific immune responses.

CBA/T6 and Balb/c mice inoculated with Plasmodium berghei ANKA strain (PbA) died from cerebral malaria 6–8 days post-inoculation. DBA/2J mice similarly inoculated developed a non-fatal cerebral malaria, with mild temporary cerebral symptoms, and died between days 15 and 22 from other malaria-related complications. When inoculated with P. berghei K173 (Pb) these mouse strains did not develop a cerebral malaria but died between days 15 and 22 from other malaria-related complications. These mouse strain/parasite strain combinations allow for detailed examination of factors critical in the pathology of murine cerebral malaria. Monastral Blue, a colloid dye, when injected intravascularly between days 0 and 2 into PbA-inoculated CBA (PbA-CBA) or Balb/c (PbA-Balb/c) mice prevented death from cerebral malaria. There was no evidence of increased vascular permeability at this stage. When Monastral Blue was injected between days 5 and 8, there was increased vascular permeability in the kidney, liver, lung, spleen and brain of PbA-CBA and PbA-Balb/c mice. Injection of Monastral Blue into these animals at this time also precipitated cerebral symptoms and death, but not in Pb-infected mice. Endothelial and mononuclear cells phagocytosed, and were coated with, the Monastral Blue particles when the dye was injected between days 5 and 8 into PbA-CBA and PbA-Balb/c mice. Control, uninfected mice did not demonstrate either of these features. Pb-infected mice only demonstrated coated mononuclear cells. Mononuclear cell attachment to the endothelium, increased vascular permeability and increased association of Monastral Blue particles with monocytes and endothelial cells were correlated with cerebral symptoms and death. Monastral Blue is thus a useful agent for studying the roles of mononuclear cells and endothelium in murine cerebral malaria.

The effect of treatment with monoclonal antibodies (Mabs) which deplete CD4+ or CD8+ T lymphocytes, on infections with Eimeria spp. was examined in NIH mice. Treatment with anti-CD4 Mab increased susceptibility to primary infections with E. vermiformis or E. pragensis and reduced the subsequent resistance of the mice to homologous challenge. Similar treatment of immune mice did not affect their resistance to re-infection but this was reduced in mice depleted of CD8+ T lymphocytes. In mice immunized with E. vermiformis the effect of CD8+-depletion was very slight, apparent only as the presence of small numbers of oocysts in the faeces of some mice; in mice immunized with E. pragensis there was a small, though significant, increase in oocyst production, compared with controls and anti-CD4-treated groups. These results confirm the importance of mechanisms involving the function of CD4+ T lymphocytes in the control of primary infections with Eimeria spp. and indicate that CD8+ cells play some part in the expression of resistance to reinfection. They also show that a major part of this resistance was not affected by either of the treatments given.

The development of cerebral lesions in Plasmodium berghei-infected mice was dependent on the Strain of mice and the size of the infectious inoculum. In particular, C57B1/6J mice develop cerebral lesions when infected with low numbers of parasitized erythrocytes. By increasing the number of parasites in the infectious inoculum, the percentage of animals that develop cerebral malaria is decreased. Varying degrees of protection against the development of cerebral malaria can be obtained by several methods of immunization. (1) Injection of mice with large numbers of disrupted parasitized erythrocytes 1 or 2 weeks before the challenge infection (protection up to 70%). (2) A 2-day immunizing infection given 9 or 14 days before the challenge infection (protection up to 85%). (3) Injection of mice with plasmodial exoantigen preparations 1 week before the challenge infection (variable protection-rate, up to 100%). In all mice protected against cerebral malaria, parasitaemia is not affected by the immunizing treatment, indicating that protective mechanisms against cerebral malaria and parasitaemia are independent.

Exclusion of magnesium ions from resealed ghosts or their extraction from intact human red cells by means of an ionophore results in a reversible drop in susceptibility to invasion by Plasmodium falciparum merozoites in vitro. Resealed ghosts, containing magnesium-ATP and diluted cytosol, are invaded with high efficiency only when the original hypotonic lysis is carried Out in the presence of magnesium ions. This effect is not related to the loss of membrane-associated constituents when magnesium ions are absent. Ghosts containing calcium ions, together with the protective agent, flunarizine, were essentially resistant to invasion; this effect is again at least partially reversible. A possible explanation of these phenomena is that entry of the merozoite may be inhibited by breakdown of the host cell phospholipid asymmetry, with the appearance of aminophospholipids at the outer cell surface.

This paper reviews naturally occurring and synthetic compounds that either enhance immune defences or lower both natural and acquired immunity. Immunomodulatory agents used both for laboratory study and clinically for the management of immunologically based diseases are considered

Babesia hylomysci and B. divergens were studied for superoxide dismutase (SOD) activity by enzyme assay and isoelectric focusing (IEF). In the two Babesia species, parasite-associated SOD is cyanide-insensitive and inhibited by H2 O2, indicating that iron is the cofactor metal. Measurements of SOD activity from purified parasites show that the SOD activity detected in Babesia is, for the main part, due to an endogenous enzyme.

Erythrocyte invasion assays are described for two species of rodent malaria, namely Plasmodium berghei and P. c. chabaudi. These invasion assays are simple, are carried out using a candle jar and allow a number of assays to be performed simultaneously. Our results demonstrate that both rodent malaria species show an in vitro preference for reticulocytes although the preference of P. c. chabaudi for these cells is not as marked as that of P. berghei. The details of our invasion assays and our results obtained are discussed.

Cloned lines of the four rodent Plasmodium species can be differentiated by the RFLP pattern generated following Southern blotting and probing with PCsv4.1, a probe derived from a P. chabaudi chabaudi genomic library. Groups of CBA/Ca mice were inoculated simultaneously with cloned lines from two parasite species or strains. Six mixed species and three mixed strain infections using rodent malaria lines were initiated. The composition of the parasite population in each group was determined qualitatively and semi-quantitatively by analysis of the DNA purified from daily blood samples, thereby providing a dynamic representation of each mixed infection. Effects on the course of parasitaemias are presented and discussed.

Following sequence analysis of a Leishmania donovani kinetoplast DNA (kDNA) minicircle, we have developed synthetic oligonucleotides for use in the polymerase chain reaction (PCR). With these primers, we have amplified L. donovani kDNA from splenic aspirates and blood samples taken from kala-azar patients. Treatment of the samples for PCR requires only limited DNA purification by lysis in SDS, digestion with proteinase K, phenol extraction and ethanol precipitation of the resulting nucleic acid. We have obtained amplified product routinely with DNA prepared from the equivalent of 2·5–25 µl of splenic aspirate or of 50–500 µl of blood from infected patients. In dilution experiments a visible product has been obtained on amplification of DNA from the equivalent of 2·5 × 10−7 µl of splenic material. We therefore propose the amplification of L. donovani kDNA by PCR as a rapid and highly sensitive method for the diagnosis of kala-azar.

A DNA probe PCsv4 and a subclone thereof PCsv4.1, hybridize specifically to rodent malaria DNA. DNA purified from a small volume (10 μ1) of infected mouse blood was used to determine the composition of the parasite population present. The hybridization signal following PCsv4 probing of slot-blotted DNA correlated directly with parasitaemia. The hybridization pattern and Intensity, resulting from probing restriction enzyme digested and Southern-blotted genomic DNA, determined the identity of the infecting parasite line(s), and provided a semi-quantitative measure of parasite burden. Fifteen parasite lines representative of all four Plasmodium species infecting rodents can be differentiated in this way.

Cyclosporin A (CsA), a cyclic undecapeptide with powerful properties of immunosuppression, acts on parasitic infections in laboratory animals in various ways. The outcome of drug administration in vivo varies with timing of treatment relative to infection, route of administration, dose and number of treatments applied. CsA is clearly antiparasitic against malaria, schistosomes, adult tapeworms, metacestodes and filarial nematodes. By contrast, it acts as an immunomodulator against trypanosomes and Giardia, by exacerbating infection; in the case ofLeishmania spp. the drug acts variously. In some other infections CsA acts both as an antiparasite drug and as an immunosuppressant (Toxoplasma, avian coccidiosis and gastrointestinal nematodes).

This study examined whether allelic changes observed when clinical isolates of Giardia intestinalis made in suckling mice were adapted to in vitro growth occurred as a result of gene switching (alternate isoenzymes) or through selection of organisms with different genotypes from mixed infections. Samples were compared electrophoretically at 20 enzyme loci. Marked allelic differences were detected between the uncloned clinical isolates grown in mice and the axenic cultures established from them. Furthermore, the allelic profiles of the uncloned isolates changed during the course of in vivo or in vitro growth. In contrast, all clones produced from each isolate retained identical allelic profiles, regardless of whether they were grown in vivo or in vitro. These findings argue against gene switching as an explanation for the observed allozyme changes and support preferential selection of organisms with specific genotypes by growth conditions. The data indicate the presence of at least 2 and possibly up to 4 distinct genotypes within each clinical isolate. The genetic differences detected between clinical isolates in suckling mice were of similar magnitude to those that separate different axenic isolates of G. intestinalis into cryptic species. Conversely, the genetic differences between the isolates were limited when sampled after establishment in vitro. These findings have significant implications for research on Giardia and other medically important parasites and raise the possibility that culture may exert a similar selective bias on the genotypes isolated from infections with other parasitic protozoa.

A 196 bp fragment of cloned kinetoplast (k) DNA of Leishmania infantum has been sequenced and shown to be diagnostic for this species. The DNA from 104–105 promastigotes was routinely and specifically detected in crude dot blots of whole cells lysed in situ, and there was no cross-hybridization with 106 cells of L. major, L. tropica, L. killicki, L. aethiopica, L. braziliensis and L. mexicana. A low degree of hybridization was found with L. donovani and L. chagasi. This probe will be of particular use in Mediterranean countries, such as Italy, in which dermotropic zymodemes of L. infantum can be difficult to isolate and grow in numbers sufficient for isoenzyme typing.

Host immune responses have been shown to enhance the efficacy of several schistosomicidal drugs. The evidence derives mainly from experiments on Schistosoma mansoni infections in the mouse with their immune status variously modulated; this review emphasises praziquantel (PZQ), which is now the main drug used for treatment of human schistosomiasis. Electron microscopy and indirect immunofluorescence indicate that PZQ disrupts the integrity of the surface membranes of S. mansoni, particularly those covering the dorsal tubercles of adult male worms, and this causes antigens which are the targets of antibody attack to be revealed. We review the evidence that two S. mansoni antigens in particular are implicated in the immune-dependent action of PZQ: a 200 kDa glycoprotein and a 27 kDa antigen with non-specific esterase activity.

A new method is described which has made possible the long-term axenic cultivation of Leishmania mexicana amastigotelike forms in Schneider's Drosophila medium supplemented with 2% (v/v) foetal calf serum. Unlike previous methods, it utilizes direct culture of parasites obtained from the lesions of infected animals rather than adaptation of promastigotes in vitro. Ultrastructural (possession of megasomes), biochemical (cysteine proteinase activity and gelatin SDS-PAGE banding pattern) and infectivity (in vivo) data are presented which show the close similarity of the cultured forms to lesion amastigotes. The axenically cultured forms grew optimally at a temperature of 32–33 °C, providing further evidence for their amastigote nature. It was found that adjustment of the pH of the growth medium to 5·4 was required in order to retain the amastigote morphology of the cultured parasites. This supports the notion that leishmanial amastigotes are acidophiles.