Fresh (3 h) schistosomula of Schistosoma mansoni were obtained by the skin penetration method and injected intravenously into groups of mice either immediately or after 48 h incubation in tissue culture. Schistosomula were then recovered from the lungs 1 to 2 days after injection. The recovery results showed that the ability of uncultured schistosomula to emerge from lung tissue increased with time, became maximal on the 3rd day and diminished thereafter. In contrast, cultured schistosomula were able to emerge from chopped lung at a much earlier time after injection.

Fresh schistosomula, as well as those recovered from mouse lungs, were assayed by indirect immunofluorescence for the presence of parasite and host antigens. Similarly, fresh schistosomula which had been incubated in foetal calf serum, with or without mouse red blood cells, were assayed for parasite and host antigens after 3–24 h of culture. Fresh schistosomula expressed parasite antigen with greater intensity than those recovered from mouse lungs and showed no trace of host antigen. Lung schistosomula, on the other hand, expressed less parasite antigen but more red blood cell antigen with increasing time in the host. Schistosomula cultured in the absence of mouse red blood cells expressed only parasite antigen throughout the period in culture, while those cultured in the presence of mouse red blood cells also expressed parasite antigen during the entire period in culture but, in addition, by 24 h host antigen could also be detected.

These developmental and adaptational changes may play a role in determining the survival of parasites in the normal or immune host.

The uninfected digestive gland of Bithynia tentaculata consists of three main cell types: ‘absorptive’ cells, ‘thin’ cells and ‘secretory’ cells. With infection by sporocysts, rediae, or metacercaria, pressure effects caused a reduction in the lumen of the digestive gland tubules and this probably resulted in a starvation autolysis. Some tubules recovered from infection and in such cases there was a reduction in the height of the epithelium. A migration of the digestive gland nuclei was only evident in redial infections and this may have been due to the toxic effects of rediae. In addition, rediae directly ingested host digestive gland tissue.The digestive gland was reduced to about two-thirds of its normal size in all heavily infected snails. Secretory cells of the gland were more resistant to infection than were the absorptive or thin cells.

I am indebted to Dr F. R. Stranack for her supervision and guidance throughout this investigation, and to Dr G. Charles and Dr T. Jenkins for their continued help and advice. This study was carried out during the tenure of a Research Assistantship awarded by the Governors of Portsmouth Polytechnic.

In the male reproductive system of Nematodirus battus the spermatogonia and spermatocytes are arranged upon a central rachis and the spermatocytes are characterized by the presence of cup-shaped membranous organelles which are formed in association with Golgi complexes and granular endoplasmic reticulum. Spermatozoa of N. battus in the male nematode have an electron-dense, conical tail-like region which contains the nuclear material and an elongate cytoplasmic region which contains the cell organelles. In the uterus of the female, however, the cytoplasmic region of the spermatozoon has an amoeboid appearance and in the seminal receptacle the cytoplasmic region is dome-shaped and many of the membranous organelles have connexions with the outer membrane of the spermatozoa. Movement was not observed in spermatozoa released from the seminal vesicle of the male worm.

The study was designed to test the hypothesis that the majority of individuals in a population of Anguillicola crassus can tolerate seawater by osmoconformation with the blood plasma of the eel host. The osmolality of the pseudocoelomic fluid of pre-adult and adult A. crassus was compared to the osmolality of eel plasma after eels were maintained in laboratory freshwater for 2 weeks (short-term transfer) or 3 months (long-term transfer) or in natural seawater for 2 h (acute transfer), 2 weeks or 3 months. The majority of A. crassus (at least 90% of the tested population) osmoconform with their hosts in seawater within ±30 mOsm/kg of host osmolality. Some pre-adults and adults (15–21% of the total population) were unable to withstand osmotic stress resulting in vacuolation of the hypodermis and intestinal wall, and cuticular detachment. The reasons for variation in the tissue tolerance of A. crassus to increased osmolality of host plasma are unknown and are not related to maturity, sex or conditions in the swimbladder. Osmoconformation in the majority of the population, however, enables parasites to survive and reproduce for long periods in seawater eels. The adaptation of A. crassus to its euryhaline host has played an important part in the rapid spread of the swimbladder nematode in populations of the European eel.

1. Dirofilaria aethiops Webber, 1955 has been discovered in African monkeys (Cercopithecus aethiops). Adult worms were found in the interfascicular connective tissue, especially in the thighs; microfilariae were found in the blood.

2. The adult worms are morphologically similar to other members of the genus, from which they differ principally in the structure of the spicules of the male worms. The thickening of the left spicule is complex at the twist; the latter is nearer to the proximal than to the distal end. The right spicule is robust and ends in a cap-like structure.

3. The microfilariae are indistinguishable from those of other members of the genus.

4. In Dirofilaria schoutedeni Vuylsteke & Rodhain, 1938 the left spicule is simpler in structure than the left spicule of D. aethiops, and the twist is nearer to the distal end than to the proximal end. The distal part of the right spicule of D. schoutedeni has longitudinal markings but no cap.

5. D. aethiops has been recorded from Tanganyika, Uganda and Gambia, in Cercopithecus and Colobus monkeys.

6. The anatomy of the female D. aethiops, as it is shown in serial sections, is briefly described.

β–glucans are immunomodulators able to activate innate immunity and to potentiate acquired immune reactions. We investigated the impact of co-administration of liposomized β-glucan on the larvicidal effect of the anthelmintic praziquantel (PZQ) in the livers and peritoneal cavities in mice infected with Mesocestoides vogae (M. corti). Also, within 2 weeks following therapy (up to day 29 p.i.) we examined collagen synthesis in the livers of mice by means of biochemical determination of hydroxyprolin concentration, total mast cell counts and cell proliferative capacity using immunohistochemical and radiometrical methods. After co-administration of liposomized glucan (LG) and PZQ efficacy (%) was significantly higher than after treatment with either compond alone, particularly in the peritoneal cavity compared to the liver. In comparison with the control, more intense collagenesis was found in the B-liver parts (high intensity of infection) and lowering of collagen content in the A-parts (very weak infection). This effect was strongest after LG treatment and co-administration of PZQ abolished the pro-fibrotic effect of LG. In all groups, mast cell counts were higher in the B-liver parts than in the A-parts and the dynamics of mastocytosis was profoundly modulated following therapy. Whereas the effect of PZQ was only moderate, early and very strong onset was seen after LG treatment. Administration of PZQ supressed LG induced-elevation of mast cells counts in both liver parts. Using DNA S-phase markers (BrdU and 3H-thymidine) the proliferative capacity was shown to be associated with several kinds of liver cells. Therapy significantly stimulated [3H]-thymidine incorporation (cell proliferation) only in the A-parts over that in control, the most after LG administration. In summary (i) the anthelmintic effect of PZQ could be enhanced after simultaneous administration of the immunomodulator β-glucan entrapped in a liposomal carrier, (ii) intense mastocytosis seen after treatment with LG seems to have a direct role in the glucan′s pro-fibrotic activity and can be abolished after co-administration of PZQ in a time-dependent manner, (iii) the pattern of cell proliferation indicates that in the case of PZQ treatment, the reparative processes of liver parenchyma are enhanced in an inverse correlation with the intensity of infection.

1. A new monocystid gregarine is reported from the seminal vesicles of brandling worms, Eisenia foetida Sav. The morphology and movements of the trophozoite are described, and the stages of gametogony and sporogony are outlined.

2. The chromosome cycle is investigated. The haploid number of chromosomes is four, and the reduction division is shown to be zygotic.

3. The systematic position of the gregarine is discussed, and it is placed in the genus Apolocystis as A. elongata n.sp.

Genetic variation was examined in the anoplocephalid cestode Progamotaenia festiva, from Australian marsupials, in order to test the hypothesis that P. festiva, is a complex of sibling species and to assess the extent of host switching reported previously based on multilocus enzyme electrophoresis (MEE). Polymerase chain reaction (PCR)-based single-strand conformational polymorphism (SSCP) was used for the analysis of sequence variation in the cytochrome c oxidase subunit 1 (cox1) gene among 179 specimens of P. festiva (identified based on morphology and predilection site in the host) from 13 different host species, followed by selective DNA sequencing. Fifty-three distinct sequence types (haplotypes) representing all specimens were defined. Phylogenetic analyses of these sequence data (utilizing maximum parsimony and neighbour-joining methods) revealed 12 distinct clades. Other heterologous species, P. ewersi and P. macropodis, were used as outgroups and the remaining bile-duct inhabiting species, P. diaphana and P. effigia, were included in the analysis for comparative purposes. The latter 2 species were nested within the clades representing P. festiva. Most clades of P. festiva identified were restricted to a single host species; one clade primarily in Macropus robustus was also found in the related host species M. antilopinus in an area of host sympatry; another clade occurring primarily in M. robustus occurred also in additional kangaroo species, M. rufus and M. dorsalis. High levels of genetic divergence, the existence of distinct clades and their occurrence in sympatry provide support for the hypothesis that P. festiva represents a complex of numerous species, most of which, but not all, are host specific. Three distinct clades of cestodes were found within a single host, M. robustus, but there was no evidence of within-host speciation.

Stage-specific alternative splicing of the heat-shock transcription factor of Schistosoma mansoni (SmHSF) generates isoforms with structural diversity that may modulate the activity of SmHSF at different life-stages, and thus may regulate the expression of different genes at different developmental stages. RT-PCR, cloning and DNA-sequence analyses showed stage-specific alternative splicing inside the DNA-binding domain (DBD) involving introns I1 and I2, and beyond the DBD involving introns I4a and I7. Retention of introns I2 and I4a would inactivate SmHSF since they contain termination codons. Retention of intron I1 would add 11 amino acids inside the DBD and may change the DNA-binding specificity of SmHSF; intron I7 would add 13 amino acids to the effector region of HSF. Retention of introns was more pronounced in cercariae (larval stage living in water) than in adult worms (parasitic form in mammals). The isoforms were expressed in bacteria, but functional evaluation was not feasible, because only the isoform lacking introns was soluble while isoforms with introns were insoluble. However, stage-specific alternative splicing that changed HSF function in vivo was evidenced in intact cercariae. The cercarial SmHSF mRNA was enriched with introns I2 and I4a that contain termination codons. Therefore, translation of the SmHSF mRNA was impaired, and the SmHSF protein was undetectable. Consequently, the HSP70 gene could not be transcribed, and the HSP70 mRNA was missing. Alternative splicing was observed for short DNA segments (33–45 bp) bound by splice signals, located in the coding region. These are not bona fida exons since they are not flanked by introns. Yet, they are not regular introns since they are often found in mature mRNA. Alternative splicing of these DNA segments caused structural diversity that could modulate the function of the gene product.

Calculating epidemiological measures of infection by Trypanosoma cruzi, the causative agent of Chagas disease, is complex, because it involves several species, different stages of infection in humans and multiple transmission routes. Using the next-generation matrix method, we analysed a model which considers the three stages of human infection, triatomines and dogs (the main domestic reservoirs of T. cruzi when triatomines are present) and the main transmission routes. We derived R0 and type-reproduction numbers T. We deduced formulas for the number of new infections generated through each transmission route by each infected individual. We applied our findings in Argentine Gran Chaco. The expressions achieved allowed quantifying the high infectivity of dogs and emphasizing the epidemiological importance of the long and asymptomatic chronic indeterminate stage in humans in the spread of the infection. According to the model, it is expected that one infected human infects 21 triatomines, that 100 infected triatomines are necessary to infect one human and 34 to infect a dog, and that each dog infects on average one triatomine per day. Our results may allow quantifying the effect of control measures on infected humans, triatomines and dogs (or other highly infected vertebrate) or on a specific route of transmission, in other scenarios.

Studies have been undertaken to examine the alternative hypotheses of a Poisson or a negative binomial distribution of the numbers of strongyloid nematode eggs in deposits of faeces produced by flocks of grazing sheep, which were proposed in certain mathematical models for the distribution of strongyloid infective larvae on pastures.

Observations were made on the distribution of the weights of faecal deposits produced by flocks of ewes and lambs. In all cases the distribution was found to be significantly positively skewed. A possible fundamental basis for this skewness has been suggested.

The distribution of strongyloid egg concentration in random samples of faecal deposits collected from pasture, and the distribution of egg concentration for the single species N. battus and N. filicollis in rectal samples of faeces collected from a lamb flock, have both shown highly significant departures from a Poisson distribution. This, coupled with the fact that the weights of individual deposits vary widely, provide strong empirical grounds for rejecting the Poisson distribution as a model for the distribution of egg numbers per deposit.

An examination of the distribution of egg counts for N. battus and N. filicollis in samples of faeces collected from a lamb flock at weekly intervals through the spring and summer has suggested that the distribution tends to become more overdispersed with the passage of time and as the mean egg count is falling.

Fundamental grounds for preferring a more general model, such as the negative binomial, to describe the distribution of strongyloid egg output in flocks of sheep have been discussed.

These studies were carried out under the supervision of Dr H. D. Crofton, to whom I am deeply grateful for his stimulating encouragement and constructive criticism. My thanks go also to Professor J. E. Harris, C.B.E., F.R.S., for the provision of facilities in the Department of Zoology, and to Professor T. K. Ewer and Mr M. R. McCrea for the provision and handling of pastures and sheep at the School of Veterinary Science, Langford. This work was undertaken during tenure of a C.S.I.R.O. Overseas Research Studentship.

The comparative nutritive value of dried horse blood, highly milled wheat flour devoid of bran, a mixed diet of blood and wheat flour and that of blood and yeast for the larvae of Xenopsylla cheopis, X. brasiliensis and X. astia was ascertained experimentally. Pure blood proved inadequate, probably because it is deficient in accessory food factors. The growth of their larvae on wheat flour alone was erratic; only partial success was obtained, and the adults emerged after long and irregular intervals. This I attribute to the association of micro-organisms, possibly fungi, with this food. A mixture of blood and wheat flour quickened their larval development, but it was not a satisfactory larval diet. Blood and yeast form an ideal food for all flea larvae.

It is concluded that larval diets, containing blood or wheat proteins and vitamins of the B group, are essential for the successful rearing of these rat-fleas, and that the proper sclerotization of the adult is due to the presence of haemoglobin in the larval food. The available data on the effects of diverse diets on the growth of flea larvae lend strong support to the conclusion that successful development depends on the presence of these vitamins in the food; it also leads me to think that their source in nature may be the association of micro-organisms with the food.

The larval food appears to be an important factor that governs the distribution and host preferences of different species of flea. The larvae of X. astia require the most nutritive diet. If a rich larval food is present in a rodent burrow, X. astia flourishes, as in the burrows of Tatera indica and Bandicota malabarica, and even in those of the domestic rats in certain regions. In contrast, the nutritional requirements of the larvae of Xenopsylla cheopis and X. brasiliensis are simple; thus they prosper readily in a burrow of the domestic rat, even where the nutritive value of the larval food is very low. As the temperature tolerance of X. brasiliensis is the lowest, this species is confined to some of the cooler regions.

The irregular distribution of the three species of rat-fleas inside India may be related to differences in the nutritional value of the varied organic sub stances found in rat burrows in different plaoes. The fact that the distribution of X. cheopis and X. brasiliensis is wider than that of X. astia is attributed to the ability of the larvae of the first two species to grow better on flour alone; this possibly enables them to survive transport in grain, even without rats, to places far from their original home.

In recent years the speciation of parasites as related to their host relationships has presented some of the most interesting and at the same time most difficult problems in the field of parasitology. These problems are by no means of purely academic interest but are of prime importance in connection with the control and prophylaxis of a number of important infections of man and domestic animals.

1. Eleven strains of Entamoeba histolytica were isolated from contact carriers and all proved non-invasive to rats.

2. Three strains from cases showing symptoms proved invasive to rats.

3. The treatment of murine infection with non-invasive strains with emetine hydrochloride shows that most of them were successfully cured with six daily oral doses of 2 mg./kg., though one was not susceptible to this dose. Another proved refractory to 4 mg./kg.

4. The results of previous work with kittens are discussed in the light of the present observations.

The time-course for the development of infectivity was studied in Hyalomma detritum ticks fed as pre-imagoes on calves infected with Theileria annulata. Unfed adults derived from infected nymphs were non- infectious when inoculated into susceptible calves, whereas ticks of both sexes that had fed for 2–3 days or longer on calves or rabbits were always infectious. Some adults fed for only 1–2 days were infectious, whereas others were not. As few as 2 ticks were capable of infecting susceptible calves.

RNA polymerase III (Pol III) synthesizes small RNA molecules that are essential for cell viability. Accurate initiation of transcription by Pol III requires general transcription factor TFIIIB, which is composed of three subunits: TFIIB-related factor BRF1, TATA-binding protein and BDP1. Here we report the molecular characterization of BRF1 in Trypanosoma brucei (TbBRF1), a parasitic protozoa that shows distinctive transcription characteristics. In silico analysis allowed the detection in TbBRF1 of the three conserved domains located in the N-terminal region of all BRF1 orthologues, namely a zinc ribbon motif and two cyclin repeats. Homology modelling suggested that, similarly to other BRF1 and TFIIB proteins, the TbBRF1 cyclin repeats show the characteristic structure of five α-helices per repeat, connected by a short random-coiled linker. As expected for a transcription factor, TbBRF1 was localized in the nucleus. Knock-down of TbBRF1 by RNA interference (RNAi) showed that this protein is essential for the viability of procyclic forms of T. brucei, since ablation of TbBRF1 led to growth arrest of the parasites. Nuclear run-on and quantitative real-time PCR analyses demonstrated that transcription of all the Pol III-dependent genes analysed was reduced, at different levels, after RNAi induction.

1. Shortly before the imago emerges, air accumulates under the pupal skin. This air is swallowed by the imago and passes directly into the stomach; the resulting distention of the abdomen helping to push the thorax forward so as to rupture the pupal skin.

2. After rupture of the pupal skin, the imago continues to swallow air, which causes the stomach (and therefore the abdomen) to expand further and thereby facilitate the emergence of the imago.

3. Within an hour after complete emergence of the imago, the air in the mid-gut commences to pass forward into the (hitherto empty) oesophageal diverticula, and after 12–22 hours the mid-gut no longer contains air. (Note. We find that the process of emergence here described for Culicidae also applies to some other diptera which, like mosquitoes, live for a part of their lives in water.)

4. In mosquitoes fed on liquid (e.g. raisin juice) other than blood, the liquid goes direct into the oesophageal diverticula and displaces the air from them more or less completely (the experiments of Nuttall and Shipley (1903, p. 177) showed the air to be thus displaced, but this point was not specially mentioned by them).

5. When mosquitoes feed on blood (whether previously fed on raisin juice or not) all the blood goes direct into the mid-gut. (Note. Traces of blood, negligible in quantity but sufficient to cause visible coloration, are occasionally to be observed in the diverticula. This appears more likely to happen in cases where no previous meal of raisin juice has been taken.)

6. When a mosquito is fed on raisin juice and subsequently starved for a few days, the fluid contents of the diverticula are entirely replaced by air. The experiments indicate that this air reaches the diverticula via the cardia, in the same manner as during their initial inflation.

7. In the case of “domestic” mosquitoes, the air in the diverticula becomes reduced in quantity (sometimes disappearing altogether) prior to hibernation, and increases in amount towards the end of the hibernating period.

8. In the case of male mosquitoes, the transference of air from the mid-gut to the oesophageal diverticula proceeds pari passu with the turning of the hypopygium, the two processes terminating at about the same time.

Intact surface membrane vesicles were obtained and purified from the schistosomula of Schistosoma mansoni. The method of preparation of the vesicles involved treating the schistosomula in a buffered high salt medium for 1 h at 4°C (Step 1), and then for 1 h at 25–37°C (Step 2). Vesicles were formed from mechanically- and skin-transformed schistosomula, and their size and number depended on the temperature of Step 2. The majority of the vesicles expressed surface membrane concanavalin A receptors and parasite antigens on their outermost surfaces. When incubated with sensitized peripheral blood mononuclear cell preparations from humans exposed to schistosomal preparations, the vesicles stimulated lymphocyte transformation as effectively as soluble egg and adult worm antigens. Although both preparations contained identical proteins, small vesicles were less effective than large vesicles in stimulating lymphocyte transformation. Hence, vesicles prepared under a variety of conditions might be used to study those factors which influence the presentation of membrane-bound surface antigens to the immune system of the host.

Plasmodium falciparum has developed resistance to most anti-malarials; therefore, an investigation of potential targets should be performed. DNA helicases are enzymes that catalyse the unwinding of double-stranded DNA to provide single-stranded templates for DNA replication, repair and recombination. In this study, a DNA helicase (PfDH A) was purified from a crude extract of Plasmodium falciparum. DNA helicase activity was measured by assaying unwinding activity. The apparent molecular weight of PfDH A as determined by SDS-PAGE was 90 kDa. PfDH A moved unidirectionally in the 3′ -to- 5′ direction along the bound strand and preferred a fork-like substrate structure and could not unwind blunt-ended duplex DNA. Unwinding activity required Mg2+ and could be inhibited by 200 mM NaCl or KCl and was dependent on hydrolysis of ATP or dATP. Anthracyclines, including daunorubicin, nogalamycin, doxorubicin, and aclarubicin, inhibited PfDH A activity with IC50 values of 2, 5, 8 and 9 μM, respectively. Based on the results, PfDH A differs from all known human DNA helicases. However, its function and roles in parasite DNA replication need to be elucidated in the future.