The amounts of ammonia excreted by two endoparasitic flatworms have been compared and, weight for weight, the sheep liver fluke Fasciola hepatica produces ten times as much ammonia as the rat tapeworm Hymenolepis diminuta. This is thought to reflect a difference in diet, the fluke feeding mainly on the blood and tissue proteins of its host, and the tapeworm mainly on carbohydrates from the food materials in the gut of the host. However, the addition of glucose to the saline in which the flukes are cultured produces a marked decrease in the ammonia and an increase in the lactic acid excreted, suggesting that flukes are not obligatory protein feeders but that they may utilize carbohydrate when it is available. The relatively low production of lactic acid when proteins are being metabolized suggests that the flukes are producing an alternative end-product of respiratory metabolism. It is possible that this is the neutral lipid found in the excretory system of Fasciola and the possibility of an alternative metabolic pathway for the production of such lipid is discussed. The respiratory metabolism appears to be independent of oxygen, and the possible role of oxygen in other aspects of the metabolism of the fluke is discussed.

I should like to express my sincere gratitude to Dr J. Llewellyn for his constant advice and encouragement during the course of this work and the preparation of the manuscript. I also thank Professor 0. E. Lowenstein, F.R.S., for allowing me the use of the facilities in his department during the tenure of an S.R.C. Research Studentship. Finally, my thanks are due to Mr E. Wilson, the Veterinary Officer at the Birmingham City Meat Market, for his constant co-operation in obtaining supplies of liver flukes.

(1) Light-microscopic observations were made on B. pahangi larvae at various stages of development in stained serial sections of A. aegypti thoraces, and on 8-to 9-day 2nd-stage live, unstained larvae.

(2) Six-day larvae situated in A. aegypti flight muscle, and isolated 8- to 9-day 2nd-stage larvae, were examined with the electron microscope.

(3) Results showed that host-muscle mitochondria which appeared to be morphologically intact aggregated outside the larvae at the head and tail ends and were present from 6 days onwards in the oesophagus and in the midgut. Those in the oesophagus were unchanged; some of those in the midgut were intact, others showed varying degrees of disintegration suggestive of digestion. Host-muscle mitochondria are therefore likely to be a source of nutrients for B. pahangi larvae during the last third of their intramuscular developmental phase.

Dr W. W. Macdonald, of the Liverpool School of Tropical Medicine, kindly supplied the infected mosquitoes, and offered valuable advice in the course of this work. Miss M. A. Johnson assisted with the care of the mosquitoes.

We are particularly indebted to Mr David John for his patient work in preparing the filariae for electron-microscopic observation. Mrs J. Oliver assisted with some of the histological preparations.

We are pleased to acknowledge the financial assistance of the Royal Society towards the purchase of the ultramicrotomes used in the course of this work.

An account is given of the morphological characteristics of larvae of four species of the Ixodes ricinus complex, namely I. ricinus, I. persulcatus, I. pacificus and I. nipponensis. Particular reference is made to chaetotaxy and its importance in differentiating larvae of closely related species is emphasized.

We are grateful to Dr G. Owen Evans for discussion on matters on chaetotaxy.

Electron-microscope investigations on the outer layers of parasitic platyhel-minths have so far neglected the monogeneans and have been confined to the endoparasitic digeneans and cestodes (Braten, 1968; Burton, 1966; Charles & Orr, 1968; Erasmus, 1967; Lumsden, 1966; Morris & Threadgold, 1967; Race, Larsh, Esch & Martin, 1965; Rothman, 1968; Threadgold, 1965). For this reason the covering layer of monogeneans is still referred to as a ‘cuticle’ even though it is unlikely to be so since the surface of the endoparasitic platyhelminths has been shown to be a living cytoplasmic ‘epidermis’ (see Lee, 1966) which, as pointed out by Erasmus (1967) and Lumsden (1966), constitutes a metabolically active inter-face with host tissues and fluids. Two main considerations prompted the present investigations of the covering layer of monogeneans. First, it was hoped that a comparison of the covering layer of these ectoparasites with that of the digeneans and cestodes might help to define particular specializations of this layer associated specifically with the ectoparasitic or endoparasitic habit; secondly, because ectoparasitism can, in an evolutionary sense, be considered as a transitional state between a free-living and an endoparasitic existence (see Llewellyn, 1965), it was felt that this work on monogeneans could assist an understanding of the evolution of the platyhelminth outer layer.

1. The weight of blood ingested by all stages of H. anatolicum anatolicum has been determined and from the results a direct relationship appears to exist between the weights of replete adults and their blood content. In engorged nymphs the relationship was subject to some variations, while in fully fed larvae no such correlation was apparent, indicating the ingestion of a large proportion of extravascular fluids.

2. It was found that all instars concentrate the imbibed blood by the elimination of water. In larvae the concentration is by a factor of at least 5·25, in nymphs 3·4, in females 2·1 and in males 1·4.

3. A correlation was found between the external colour of fully engorged larvae and the quantity of blood imbibed.

I wish to thank Dr R. P. Chaudhuri for providing the original specimens of H. anatolicum anatolicum from which the culture was started, and Professor Don. R. Arthur for reading the manuscript and for his advice and assistance.

Studies have been made of the growth and excystment of a trematode, Acanthoparyphium spinulosum, in chicks fed on various denned diets.

Day-old chicks were fed metacercariae and maintained on deficient as well as complete rations for periods up to 20 days. The excystment rate was determined by the number of worms that were recovered from the metacercariae. The largest number of worms were recovered from chicks that received a normal diet with starch as the sole carbohydrate source. The smallest percentage of parasites was found in birds that had received a normal diet with sucrose serving as the carbohydrate source. About 25 % were recovered from chicks fed several other diets.

The trematodes developed and grew well in vitamin A deficient chicks; there was a large number of intra-uterine ova observed in worms from these hosts. Acanthoparyphium spinulosum from chicks fed a ‘no vitamin D’ diet grew at a normal rate until the 6th day; after this they grew more slowly than 6-day-old worms in chicks fed the ‘no vitamin A’ diet.

The lengths of worms from birds deprived of all vitamins differed little from those of the parasites recovered from the chicks deficient in vitamin D. The largest worms were obtained from chicks fed a complete diet with starch as the sole carbohydrate source.

Parasites from chicks fed a diet 50 % deficient in carbohydrates (with sucrose as the carbohydrate) developed normally.

Worms also developed in chicks fed a ‘no protein’ diet with starch and a ‘no protein’ diet with sucrose. A deleterious effect on ova production was observed in worms from both these groups.

Metacercariae developed in chicks fed a ‘no carbohydrate’ diet, but no mature worms were recovered due to the reduced life of the host. Starvation of the hosts produced small worms containing few intra-uterine ova.

The growth curve of A.spinulosum was plotted and compared with Echino-paryphium flexum.

This investigation was supported in part by U.S. Public Health Service Grant UI-00202, and by a grant-in-aid of research by the Society of the Sigma Xi, New Haven, Connecticut.

The structure and function of the vacuoles in the eggs of Schistosoma mansoni is unknown. In this study the eggs have been immersed in serum albumin and examined by phase-contrast optics. Under these conditions the vacuoles exhibit a definite internal structure, suggesting that they may be or may have been cellular. Histochemical staining techniques to detect the nature of the vacuolar contents revealed that the vacuoles stained heavily with iodine and gave a positive PAS reaction. This suggested the presence of carbohydrate. Stains for nucleic acids, protein and lipid were not taken up by the vacuoles. The egg shell stained with o–toluidine blue and with basic fuchsin uniformly, except for a thin irregular band which might indicate a possible line of weakness. The vacuolar membranes seem to be similar to the vitelline membrane in permeability to hydroxyl ions and it is possible that these membranes have a common origin. Plasmolysis studies with salts, urea, glycerol and sucrose showed that the vacuolar membranes have about the same permeability properties to these compounds as the other egg membranes. Glycerol treatment of the eggs until equilibrium is attained, followed by transfer of the eggs to water or saline results in a hatching of the eggs. In this, a dead mira-cidium is extruded from the shell. Similar treatment of eggs equilibrated with sucrose does not result in this hatching process. It is postulated that glycerol solubilizes a factor which aids the hatching process.

I should like to thank the following: Dr S. A. Ibrahim, in whose department this work was carried out; Dr S. Dawood of the Stack Laboratory, Khartoum, who kindly made his microscope available for use; Mr J. R. Lauder for valuable discussions and suggestions; and many young patients in Khartoum Civil Hospital for stool samples and smiles.

1. The percentage conversion of cercariae into schistosomula, when the former were penetrating human epidermal sheets, was found not to be significantly affected by the underlying media 1—5 (Table 1). The percentage conversion varied from 16·4% (in normal saline underlying skin) to 26.2 %. When distilled water was the underlying medium significantly fewer cercariae penetrated the sheets.

2. There was a statistically significant decrease in the percentage conversion of cercariae to schistosomula when sodium chloride final concentration 0·67 % was placed in the exposure tube with the cercariae (1·6 % P = < 0·02 when compared with La Ye medium); 0·43 % sodium chloride also significantly decreased penetration.

3. There was a statistically significant decrease in the percentage conversion of cercariae to schistosomula when the orientation of the human epidermis was reversed, i.e. the biologically inner surface was exposed to the cercariae (2.6 %, P = < 0.02, when compared with La Ye medium).

4. Schistosomula agglutinated in the presence of normal or immune human serum or horse serum. In human serum to which donor red blood cells had been added, many red cells adhered to the schistosomula. The number adhering was less in inactivated serum. The adhesion was specific for red cells, the effect not occurring with either yeast cells or silicic acid particles. The red cells were shed from the surface of the schistosomulum after 4—12 h.

I should like to thank the following: Dr S. A. Ibrahim, in whose laboratory this work was carried out; Dr S. Dawood of the Stack Laboratory, Khartoum for the use of his microscope; Mr T. R. Melrose and Mr J. R. Lauder for valuable discussions; the Medical Research Council, U.K., and the University of Khartoum for research grants. I am very grateful to the Pathology Department Faculty of Medicine, University of Khartoum, for preparing the sections of human epidermis.

Several species of Australian python, Morelia spilotes variegatus, Liasis amethy-stinus, L. fuscus and L. childreni, were experimentally infected with eggs of Ophidascaris moreliae and the development of the larvae described. The first moult occurred in the egg, which survived and remained infective for over 7 years in moist conditions. The first moult occurred in 12 days and the second-stage larva in the egg was infective to mice in 14–21 days. After ingestion the larvae migrated to liver and lungs and in 21 days were mainly distributed throughout the subcutaneous tissues, particularly in the neck region and behind the ears. Growth occurred to about 8 mm; the larvae attained their infectivity for pythons after 5 weeks and retained it for more than a year. The second moult was not observed but it was assumed that the third stage was infective for pythons. Development to about the same length occurred in guinea-pigs and indigenous and laboratory rats and in the bandicoot (Isoodon obesulus). In the possum (Tricho-surus sp.) a more rapid growth occurred to about the same length. No growth occurred in tadpoles.

After ingestion by pythons larvae migrated to the lungs, where they remained for 3 months or more, grew to a length of 45 mm and underwent the third moult. Larvae removed from the lungs and reingested by pythons returned to the lungs if they had been in the first python for less than 60 days. After this time, they remained in the stomach of the second python. During the third moult larvae underwent a marked constriction of the anterior end, moved up the trachea and became attached in the upper oesophagus as fourth-stage larvae. In mouse-infected M. spilotes the third moult occurred at 117 days at a length of 23–45 mm; fourth-stage larvae measured 34–50 mm. The fourth moult occurred in the stomach at 318 days at a length of 37–55 mm. The smallest adult specimens found were a 55 mm male and a 63 mm female. Development to the adult stage was only observed in M. spilotes variegatus. In other python species only pre-adult forms were recovered. It appeared that development may be retarded at the fourth moult during the winter. The morphology of the larvae during the five stages of development is described. It is concluded that the main growth phases are the third stage in the lungs of the python and the adult stage in the stomach of the python. Encapsulated forms were found only in the tissues of intermediate hosts. The larvae from mouse tissues emerged from the capsules during putrefaction and survived for several weeks in water.

Acknowledgement is made to the able technical assistance of Miss Marian Hollis. Financial assistance for this study was provided under grants from the Australian Research Grants Committee and the United States Department of Health, Education and Welfare no. A 107023–02.

Cysteine (20–100 mM) and reduced glutathione (20–80 mM) potentiated the development of the exsheathed third-stage larva of Haemonchus contortus to the fourth stage. The potentiating effect of cysteine was greatest during the short period before and after the exsheathment of the third-stage larva. Two hundred mM cysteine and 1 mM p–chloro-benzoate inhibited development. This inhibition was reversible. Ten mM iodoacetate and 0·5 and 5 mM n–ethyl-maleimide inhibited development of the larvae. The larvae appeared to be most susceptible to the inhibiting effects of these reagents during the period of development approaching the third moult. Development was either little affected or potentiated, when larvae were incubated with these reagents for short periods immediately after exsheathment.

Daily 1 mg doses of a cortisone derivative, betamethasone, caused an increase in oocyst production and extended the patent period of E. mivati infections in partially immunized chickens. The more prolonged the course of cortisone injections, the greater the effect, although this was substantial when only a single dose was given, especially if this was at or near to the time of oocyst inoculation. E. mivati infections in susceptible chickens were similarly affected but not to such a great extent.

The possible means whereby cortisone treatment reduces acquired immunity to E. mivati infection are discussed.

Thanks are due to Mrs P. Hesketh and to other members of the parasitology department for technical assistance, also to the senior poultry attendant for care of the animals. Mr C. C. Wannop examined several of the birds post mortem.

Cortisone treatment of BL chickens given oocysts of Eimeria mivati resulted in the production of greater numbers of oocysts in the faeces and a prolonged patent period.

Schizonts were seen in sections of the intestines of treated birds up to the 24th day after oocyst inoculation although none was seen in control chickens after the 8th day. These late schizonts resembled those of the 3rd and 4th generations recorded for this species but may be those of a generation not yet described.

The depletion of intestinal lymphoid tissue in cortisone-treated birds is noted. The findings are discussed with reference to host-parasite relationships.

We wish to thank the scientific assistants and senior poultry attendant of the department for their help throughout the work.