Leucocytozoon sakharoffi of English rooks (Corvus f. frugilegus) underwent sporogony in female Simulium (Eusimulium) angustitarse, the cycle taking 3–4 days. Sporozoites from the salivary glands of S. angustitarse, both naturally and experimentally infected, gave rise to L. sakharoffi infections when injected to aparasitaemic rooks, with a minimum pre-patent period of 8 days.

I am very grateful to the following persons, for invaluable help in many ways: Mr C. Swan and Mr A. Ponchaud (Ministry of Agriculture, Fisheries and Food); Dr J. Ash, Mr N. A. Wood and Mr M. Holland (Game Research Association); Mr N. A. Hancock (Smith, Kline and French); Mr N. Biddlecombe (Breamore Estate); Dr L. Davies (Durham University); Professor D. S. Bertram, Mr E. J. Blackie, Miss L. Selden, Miss J. Freeman and Mr P. Mellor (London School of Hygiene and Tropical Medicine).

This work was generously supported by a grant from the Natural Environment Research Council.

In 1977, a parasitological survey was undertaken of stool samples collected from 335 people from 5 villages in semi-arid regions of peninsular India. Stools, which were collected from various individuals in a stratified sample of 40 matched households from each village, were examined by standard diagnostic procedures. Eggs of Ascaris lumbricoides and protozoan cysts, which were identified on morphological grounds as belonging to Entamoeba spp., were most common. Other protozoan cysts were considered to be those of Balantidium coli, and eggs of hookworm and Hymenolepis nana were also observed in some of the stool specimens. The prevalence of parasitic infections varied between villages. The village with the highest prevalence of Ascaris (70%) appeared to have the greatest demands made on its meagre drinking-water supply. This village is located in a dry, windy and dusty region and it is speculated that, in addition to well-established transmission mechanisms, the villagers may be exposed, perhaps through contamination of uncovered drinking-water wells and inhalation, to wind-borne Ascaris eggs. In 1967, a WHO Expert Committee Report on the Control of Ascariasis pointed out that information on the frequency of dust-borne infection with Ascaris in man was inadequate.

Smallpox has been eradicated by immunization with live vaccinia virus. Now, using genetic engineering, infectious vaccinia virus recombinants are being constructed which express genes from other pathogens. These viruses can stimulate specific immunological responses against the foreign antigen in vaccinated animals and can protect against disease caused by the corresponding pathogen. In this paper we describe how recombinant vaccinia viruses are constructed and illustrate the potential of this vector system for expression of genes from parasitic pathogens.

A copper/zinc superoxide dismutase (Cu/ZnSOD) gene and a manganese superoxide dismutase (MnSOD) gene of the human parasite Clonorchis sinensis have been cloned and their gene products functionally characterized. Genes Cu/ZnSOD and MnSOD encode proteins of 16 kDa and 25·4 kDa, respectively. The deduced amino acid sequences of the two genes contained highly conserved residues required for activity and secondary structure formation of Cu/ZnSOD and MnSOD, respectively, and show up to 73·7% and 75·4% identities with their counterparts in other animals. The genomic DNA sequence analysis of Cu/ZnSOD gene revealed this as an intronless gene. Inhibitor studies with purified recombinant Cu/ZnSOD and MnSOD, both of which were functionally expressed in Escherichia coli, confirmed that they are copper/zinc and manganese-containing SOD, respectively. Immunoblots showed that both C. sinensis Cu/ZnSOD and MnSOD should be antigenic for humans, and both, especially the C. sinensis MnSOD, exhibit extensive cross-reactions with sera of patients infected by other trematodes or cestodes. RT-PCR and SOD activity staining of parasite lysates indicate that there are no significant differences in mRNA level or SOD activity for both species of SOD, indicating cytosolic Cu/ZnSOD and MnSOD might play a comparatively important role in the C. sinensis antioxidant system.

Nymphal Rhipicephalus appendiculatus (Trans-Mara) were fed on a steer infected with a Theileria parva parva (Kilae 1) stock isolated from an indigenous steer in the Trans-Mara Division, Kenya, which had a high piroplasm parasitaemia. A total of 5000 engorged nymphs which had dropped on one day were enclosed in elongated nylon bolting silk tubes in groups of 200–300 and were transported immediately to the Trans-Mara where they were suspended vertically in the grass cover with one end touching the ground. Over 98% of the nymphs moulted into adult ticks and 50% moult occurred by day 28 after exposure. The ticks showed over 80% survival up to 308 days post-exposure but thereafter showed a marked mortality so that only 22·5 % of the ticks were alive after 439 days. Theileria parasites were detected in the salivary glands by day 35 post-exposure and infection rates and levels increased markedly between 180 and 235 days post-exposure. Thereafter, the infection rates and levels generally decreased. Groups of ticks were triturated and the resultant supernatant fluid inoculated into pairs of susceptible cattle, and these proved infective from day 44 to 145 after exposure. Three subsequent attempts to induce infections with supernatant fluid were unsuccessful. From 294 days after exposure, groups of 50 ticks were applied to cattle and caused lethal T. p. parva infections up to 439 days post-exposure. Climatic observations showed a relatively even monthly rainfall as well as mean maximum and minimum monthly temperatures. Clean nymphal R. appendiculatus were applied to a steer infected by adult ticks exposed for 405 days and the resultant adults were incubated at 37 °C for 6 days. Supernatant fluid produced from these ticks caused a lethal T. parva infection in a susceptible steer.

(A) Enteromonas hominis Fonseca, 1915 (emend. Dobell, 1921) is the correct designation of the flagellate first adequately described as Tricercomonas intestinalis by Wenyon and O'Connor in 1917.

Organisms morphologically indistinguishable, at all stages in the life-cycle, inhabit various macaques (M. rhesus, M. sinicus, M. nemestrinus). I have grown the simian species in “pure” cultures, and have obtained all stages in vitro and in vivo: and I have also obtained evidence—not “experimental ”, but from observed spontaneous spread of infection—that the species in M. rhesus and M. sinicus, at least, are identical.

An attempt to infect a man experimentally, by oral administration of cysts from a pure culture of the simian species (from M. sinicus), was unsuccessful.

(B) Embadomonas intestinalis [ = Retortamonas Wenrich nec Grassi] is probably the correct designation of the flagellate described by Wenyon and O'Connor (1917) as Waskia: and it is the only intestinal protozoon of Man which can be readily cultivated at all temperatures from 15° to 38° C. This species closely resembles several others described from other vertebrates—both warm-blooded and cold-blooded—but the identity of them all is still questionable.

A species similar to E. intestinalis has been observed in macaques by several workers, and has been assumed to be identical on insufficient evidence. Two attempts which I have made to infect macaques (M. rhesus and M. sinicus) experimentally, by feeding on cysts in pure cultures of E. intestinalis from Man, were completely negative.

(C) Consequently, while it seems probable on general grounds that both the Enteromonas and the Embadommas of men and macaques are specifically identical, it is still premature—with only negative evidence for their transmissibility from host to host—to assert this conclusion as a fact.

A brief summary of previous work done on the life history and the biology of fleas is given. The duration of various well-defined periods during the course of metamorphosis of the flea at temperature 23° C. and 90 per cent relative humidity is recorded.

It is experimentally proved that blood, although the most important part of the nutritional requirements of the flea larvae, is by itself not sufficient for their normal and successful development. The flea larvae require an additional food which in nature is supplied by the organic refuse present in the bed of the host of the adult fleas.

Experiments were carried out to determine the nutritional value of different constituents of the blood and it was found that the fibrin has no nutritive value to the flea larvae, while the serum and red corpuscles have equal nutritive value. Lagging larvae are the result of some deficiency in the food. The serum possesses all the essential proteins for the normal development of flea larvae but lack of the required amount of iron in it increases the percentage of mortality and delays the development. On the other hand the red corpuscles or haemoglobin, though rich in proteins containing iron but on account of the poverty of some other proteins necessary for the normal development prolong the development and increase the percentage of mortality. It was further experimentally proved that flea larvae require blood for their development in order to obtain the necessary supply of iron for their normal growth. The denatured proteins of the blood and its constituents have no nutritive value for the flea larvae.

RNA with messenger activity has been extracted from both blood and culture (insect mid–gut) forms of Trypanosoma brucei and translated in a reticulocyte cell–free system. The products of this cell–free system have been compared, and many common polypeptides demonstrated. A major polypeptide of 58000–65000 molecular weight was made when both blood and culture from RNA was added to the cell–free system.

Antiserum raised against purified variant antigen from a cloned variant (MIAG 099) was used to detect specific products of this system. A major polypeptide of approximately 58000–65000 molecular weight was precipitated when the homologous trypanosome (MIAG 099) blood form RNA was used in the cell–free system. No such polypeptide was precipitated when RNA from a heterologous strain culture or blood form was used in the system. Competition experiments, in which excess purified variant antigen was added after incubation but before addition of specific antiserum, confirmed that the polypeptide of 58000–65000 molecular weight is the variant antigen.

Poly(A)–containing RNA from trypanosomes was shown to contain the mRNA for the variant antigen.

Isolates of avian trypanosomes from nestling sparrowhawks (Accipiter nisus) and from Simulium latipes were compared by isoenzyme electrophoresis with the previously described avian trypanosomes Trypanosoma corvi and T. everetti. Simulium isolates developed into trypomastigotes in semi-defined medium at 37 °C confirming that they belong to the genus Trypanosoma. Cellulose acetate electrophoresis (CAE) and polyacrylamide gel electrophoresis (PAGE) methods were employed. A total of 11 enzymes was examined, of which 8 gave satisfactory results for all lysates (MDH, ICD, PGM, 6PGD, SOD, PEP I, PEP II and PEP D). The zymograms of 3 isolates from Simulium and 4 from A. nisus were similar indicating that they were the same organism and the isoenzyme patterns coincided with those of T. corvi. It is concluded that A. nisus and S. latipes are infected with T. corvi. Flagellate infections were found in the midgut, hindgut and rectum of Simulium where large numbers of epimastigotes were found in the lumen and attached to the cuticular intima of the rectal ampullae by hemidesmosomes at the distal end of an expanded flagellum. Out of 285 nestling sparrowhawks 13 (5%) were infected with trypanosomes. In view of their age and inability to preen themselves it is suggested they became infected via infected prey.

The question whether a stress event can have a long-lasting effect on susceptibility to parasites was studied using a freshwater bivalve clam and its crustacean parasite as a model system. Anodonta piscinalis clams were collected from 2 populations during August–September 2002. Clams were transported to the laboratory and marked. The stressed clams were subjected to low oxygen for 25 days, while the unstressed control clams were caged in their lakes of origin for the same period. Then the clams were transported to a third lake where they were exposed to natural infections by the ergasilid copepod, Paraergasilus rylovi, 11 months after the stress event. The stressed clams were more intensively parasitized. They also showed lower growth, lower reproduction and lower survival than the unstressed control clams. The results indicate that susceptibility of A. piscinalis to P. rylovi infection may be condition dependent, and that stress may have a long-lasting, increasing effect on host susceptibility to parasitism in natural populations.

Variation in host susceptibility causes significant differences in infection rates between hosts living in a semi-natural situation. Such knowledge has implications for population dynamics and evolutionary models of host–parasite interactions as well as for estimations of parasite abundance. Infection rates by Lernaeocera branchialis (L.) were measured through time and space on caged Atlantic cod (Gadus morhua L.). One group of hosts, identified by their infection history, developed significantly higher infection rates than the others. These were fish which had been infected previously, but had lost their infection. Differences between groups were consistent through both time and space. Two types of cod seem to have been present in the caged population; a small group of inherently susceptible fish, which were infected, and reinfected if the parasite was lost, and another group of resistant hosts with a small chance of becoming infected.

Macrolecithus papilliger sp.nov. from the stomach and intestine of Phoxinus phoxinus (L.) is described and its systematic position discussed. The life span in the fish, in captivity during January, is 5–6 weeks. The sequence of events in release of oocytes from the ovary and egg capsule formation has been observed in living specimens. Mature oocytes, in the ovary, are enclosed in groups of eight in a transparent membrane which ruptures, releasing them singly into the ovicapt. The egg capsule is formed in two stages: a transparent membrane is secreted, probably by Mehlis's gland, around the oocyte and vitelline cells in the ootype; later this is reinforced by vitelline globules released from the vitelline cells. These coalesce on the inner side of the membrane. The operculum is formed in the ootype. Histochemical tests support the observations.

I would like to express my gratitude to Mr S. Prudhoe, British Museum (Nat. Hist.), for helpful comments on this species and to Mr M. C. Bibby and Mrs M. Fisher for technical assistance.

The work was supported by a grant from the Science Research Council, for which I am most grateful.

1. Embadomonas blattae a new species from the hind gut of Blatta orientalis is described. The flagellate measures 6–9 μ × 5–7 μ and the cysts 5–6 μ × 4–5 μ.

2. An Embadomonas from Rana temporaria, Bufo vulgaris, and Salamandra maculosa has been cultivated and described. I propose to call it E. dobelli. It differs from other known species of this flagellate in shape, and in absence of a definite karyosome. The flagella and their basal granules are duplicated first at division, and a centrodesmose is formed between one of each pair of basal granules. The cytostome disappears and new ones are reorganised after nuclear division is completed. Definite chromosomes were never seen. Double individuals and occasionally individuals with three nuclei were seen.

3. Cysts of E. dobelli were obtained in culture. They are uninucleate. A karyosome is formed; the peripheral chromatin of the nucleus stains intensely and becomes elongated. The lip of the cytostome can often be distinguished, and sometimes the glagella. These cysts are the same as those described originally by Dobell as cysts of Eutrichomastix batrachorum and Trichomonas batrachorum.

4. Cysts of Trichomonas batrachorum have never been found in faeces or in cultures.

Mice selected as liable (L) and refractory (R) over 10 generations voided significantly more and less Nematospiroides dubius eggs compared with randomly mated (Rd) mice after primary infection with 100 larvae. Strong positive correlation was found between the numbers of N. dubius eggs in mouse faeces and the numbers of adult N. dubius recovered from mice culled from the R, Rd and L colonies. Selection limit based on a faecal e.p.g. was reached in the L mice after 9 generations whereas the faecal N. dubius e.p.g. voided by the R mice continued to decline throughout selection. In contrast, no change in worm numbers was found in L or R mice after 6 generations. The refractory state of the trait, liability to infection with N. dubius, was inherited as a dominant character with a realized heritability value of ~ 0∣2. Differential correlated responses from N. dubius infectivity compared with N. dubius fecundity and growth indicated murine genetic control of this trait by 2 major genetic units. In general, there were negative phenotypic and genetic correlations between the faecal N. dubius e.p.g. of mice and murine morphological conformation, but positive correlations between the faecal N. dubius e.p.g. of selected mice and establishment, growth and fecundity of N. dubius populations in these mice. There was little correlation between the faecal N. dubius e.p.g. after primary infection and anti-N. dubius antibody titres and parasite female/male sex-ratio.

Ornithodoros moubata ticks were fed on blood infected with Babesia equi. However, the parasites were quickly cleared as evidenced by the disappearance of B. equi-specific ribosomal RNA from the ticks. We hypothesized that if the Babesia parasite can escape midgut-associated barriers a non-vector tick can become infected with Babesia. To test this hypothesis, B. equi parasite-infected blood from in vitro culture was injected into the haemocoel of ticks. B. equi-specific rRNA was surprisingly detected 45 days after injection even in the eggs. Babesia-free dogs were infested with O. moubata ticks that were infected by inoculation with B. gibsoni-infected red blood cells. Parasitaemia and antibody production against Bg-TRAP of B. gibsoni increased gradually. These results indicate that O. moubata may be a useful vector model for Babesia parasites and also a very important tool for studies on tick immunity against Babesia parasites and tick-Babesia interactions.

Experimental procedures are described that lead to the formation of sealed red cell ghosts capable of efficient invasion by malaria parasites. Both human and monkey cells have been studied with respect to invasion by Plasmodium falciparum and P. knowlesi respectively. Resealed ghosts containing about half the normal concentration of haemoglobin are prepared by osmotic lysis and resealing at a haematocrit of 70%. When examined in the scanning electron microscope, populations of these ghosts contain few echinocytes, but have an abundance of stomatocytic forms. When undiluted haemolysate is substituted for the aqueous saline diluent, in which the cells are suspended for lysis and resealing, discocytes result, with a morphology very similar to that of the original cells. Invasion is somewhat, but not dramatically higher for this material. An investigation of the effect of intracellular potassium concentration revealed that this had no major effect on invasion. A small increase in invasion resulted from addition of 50 or 100 mg/ml of albumin to the medium in which the cells were resealed, but intra-erythrocytic protein content per se is evidently not a major factor in determining the efficiency of invasion. Augmentation of the cytoplasmic ATP concentration during lysis and resealing increased the level of parasitaemia significantly, and the parasites in these ghosts developed normally and gave rise to viable merozoites. It was found that albumin at certain concentrations passed freely into the lysed cells and reached equilibrium with that in the external medium. By contrast, the high molecular weight solute Blue Dextran 2000 was completely excluded from the lysed cells.

Experimental studies on the survival of Fasciola hepatica miracidia show no evidence that miracidial mortality varies with the pH of the medium, at least in the range 6·0–8·0. On the other hand, miracidial mortality is shown to vary with both the temperature of the medium and the age of the larvae. The mean expected life-span of the miracidium decreases from about 35 h at 6°C to about 6° h at 25° C. The Gompertz survival function provides a good description of the miracidial survivorship curves over the range of temperatures used, and we describe, a maximum likelihood method of estimating the mean values of the parameters of this function, together with their approximate 95% confidence limits.

FK506 and rapamycin (Rap) are immunosuppressive drugs that act principally on T-lymphocytes. The receptors for both drugs are FK506-binding proteins (FKBPs), but the molecular mechanisms of immunosuppression differ. An FK506–FKBP complex inhibits the protein phosphatase calcineurin, blocking a key step in T-cell activation, while the Rap –FKBP complex binds to the protein kinase target of rapamycin (TOR), which is involved in a subsequent signalling pathway. Both drugs, and certain non-immunosuppressive compounds related to FK506, have potent antimalarial activity. There is however conflicting evidence on the involvement of Plasmodium calcineurin in the action of FK506, and the parasite lacks an apparent TOR homologue. We therefore set out to establish whether inhibition of the Plasmodium falciparum FKBP PfFKBP35 itself might be responsible for the antimalarial effects of FK506 and Rap. Similarities in the antiparasitic actions of FK506 and Rap would constitute indirect evidence for this hypothesis. FK506 and Rap acted indistinguishably on: (i) specificity for different intra-erythrocytic stages in culture, (ii) kinetics of killing or irreversible growth arrest of parasites and (iii) interactions with other antimalarial agents. Furthermore, PfFKBP35's inhibitory effect on calcineurin was independent of FK506 under a range of conditions, suggesting that calcineurin is unlikely to be involved in the antimalarial action of FK506.

Northeastern China is a region of high tick abundance, multiple tick-borne pathogens and likely human infections. The spectrum of diseases caused by tick-borne pathogens has not been objectively evaluated in this region for clinical management and for comparison with other regions globally where tick-transmitted diseases are common. Based on clinical symptoms, PCR, indirect immunofluorescent assay and (or) blood smear, we identified and described tick-borne diseases from patients with recent tick bite seen at Mudanjiang Forestry Central Hospital. From May 2010 to September 2011, 42% (75/180) of patients were diagnosed with a specific tick-borne disease, including Lyme borreliosis, tick-borne encephalitis, human granulocytic anaplasmosis, human babesiosis and spotted fever group rickettsiosis. When we compared clinical and laboratory features to identify factors that might discriminate tick-transmitted infections from those lacking that evidence, we revealed that erythema migrans and neurological manifestations were statistically significantly differently presented between those with and without documented aetiologies (P < 0.001, P = 0.003). Twelve patients (6.7%, 12/180) were co-infected with two tick-borne pathogens. We demonstrated the poor ability of clinicians to identify the specific tick-borne disease. In addition, it is necessary to develop specific laboratory assays for optimal diagnosis of tick-borne diseases.

Detecting signals of selection in the genome of malaria parasites is a key to identify targets for drug and vaccine development. Malaria parasites have a unique life cycle alternating between vector and host organism with a population bottleneck at each transition. These recurrent bottlenecks could influence the patterns of genetic diversity and the power of existing population genetic tools to identify sites under positive selection. We therefore simulated the site-frequency spectrum of a beneficial mutant allele through time under the malaria life cycle. We investigated the power of current population genetic methods to detect positive selection based on the site-frequency spectrum as well as temporal changes in allele frequency. We found that a within-host selective advantage is difficult to detect using these methods. Although a between-host transmission advantage could be detected, the power is decreased when compared with the classical Wright–Fisher (WF) population model. Using an adjusted null site-frequency spectrum that takes the malaria life cycle into account, the power of tests based on the site-frequency spectrum to detect positive selection is greatly improved. Our study demonstrates the importance of considering the life cycle in genetic analysis, especially in parasites with complex life cycles.