Quantitative paper chromatography and radioactive tracers were used to study the flux of alanine through the sporocysts of Cercaria emasculans in vitro. The following evidence indicates that simple diffusion is the main mechanism of permeation: (1) the sporocysts cannot accumulate alanine against a concentration gradient; (2) the concentration of alanine in the sporocysts is a direct linear function of the concentration of alanine in the external environment; (3) addition of 10−2 M glucose to the medium does not enhance permeation; (4) neither heat-killing the sporocysts nor treating them with 10−3 M iodoacetate slows the rate of permeation; (5) the Q10's for the permeation are similar to those to be expected in a process involving simple diffusion; (6) the amino acid molarities in the parasite and host tissues in vivo are strikingly similar. No utilization of alanine during the course of 3 h incubations could be detected by autoradiography.

The dynamic nature of the equilibrium between the sporocysts and the external environment is emphasized by the exchange of alanine that occurred even under isotonic conditions.

Fresh sporocysts leaked alanine and proline during 1 and 4 h incubations in vitro. The significance of this is not known.

An electron microscope study has been made of some of the events which occur during the beginning of the relationship between Moniliformis dubius and its intermediate host, Periplaneta americana. The acanthor penetrates through the intestinal wall and is encapsulated by host haemocytes. This encapsulation is typical of the haemocytic reaction of insects to foreign bodies. A proliferation of the acanthor's surface membrane then takes place and forms a coat of microvilli. The formation of this coat by the parasite is accompanied by the dispersion or disintegration or both of the encapsulating haemocytes. The coat of microvilli constitutes a major component of the envelope surrounding early larval stages of M. dubius. There is no evidence for the presence of host connective tissue in the envelope.

We are grateful to Dr A. V. Grimstone, Department of Zoology, for use of the electron microscope, and to Mr David Barnard and Miss Valerie Furber for technical assistance. We thank Dr George Salt, F.R.S. for his criticism of the manuscript.

The anatomy of the choriothete and its relation to certain features of the tsetse uterus, such as the previously unnoticed radial, uterine ridges, are described. The results of this study suggest that the choriothete is an organ for support of the developing embryo or larva and this role is taken over by the uterine ridges when the larva reaches a sufficient size. The choriothete then appears to degenerate due to stretching of the uterus until the beginning of the next breeding cycle.

Hatching takes place with the aid of a labral egg-tooth as in some other dipteran larvae and does not require the additional presence of the choriothete.

The uterine ridges also help to form a pool of milk gland secretion around the anterior end of the second-instar larva by locking its body wall to that of the uterus.

I am grateful to Dr E. J. Popham and Col. H. W. Mulligan for reading the text and would like to thank Dr T. A. M. Nash of the Tsetse Research Laboratory, Bristol, for the provision of tsetse material. The work was carried out during tenure of a grant from the Ministry of Overseas Development.

The binding of chloroquine to DNA isolated from P. knowlesi has been investigated. In the presence of increasing concentrations of malarial DNA, the absorption spectrum of chloroquine exhibited strong hypochromism in the range 325–350 nm, small red shifts of the absorption maxima and an isosbestic point at 300 nm. The fluorescence emission of the drug at 380 nm (excitation 330 nm) was quenched five-fold by malarial DNA. Chloroquine itself protected malarial DNA from thermal denaturation. These data are consistent with binding of the drug to malarial DNA as has been suggested from studies with DNA from bacteria and mammalian cells. The binding affinity for malarial DNA is of the same order as that for mammalian DNA.

The effect of chloroquine on the metabolism of P. knowlesi in culture was also investigated. There was a direct correlation between concentrations of drug which affect the morphology of parasites and those which affect markedly the incorporation of 3H-adenosine into DNA and RNA, 14C-isoleucine incorporation into protein and the formation of lactate as a result of respiration. However, all processes were inhibited to the same extent, indicating that binding to DNA is not the only action of the drug.

The mode of action of chloroquine and the question of resistance to it are discussed.

One of us (P. I. T.) received financial assistance from the World Health Organization. We thank Dr F. Hawking and Dr J. Williamson for many helpful discussions, Dr K. N. Brown for reading through the manuscript, and Miss Jane Dunnett, Mrs A. C. Gutteridge and Mr T. Scott-Finnigan for skilled technical assistance.

Histochemical analyses of the pigment accompanying lung mite infections in Old World monkeys indicate that, contrary to other endogeneous pigments, this pigment is not homogenous, but consists of at least three components. One fraction is formed by fine granular or amorphous cleavage products of mucin substances, the second portion appears to be large granules of haemosiderin and/or lipofuscin, and the third component consists of needle-like crystals of unknown chemical nature. The source and nature of the pigment are discussed.

I am grateful for the technical assistance of Mrs J. A. Silva and the support by Drs R. T. Damian, S. S. Kalter, R. E. Kuntz and B. J. Myers; Southwest Foundation for Research and Education, San Antonio, Texas.

This study was funded in part by U.S.P.H.S. Grants RR-00361, RR-00278 and RR-00451 and WHO Grant Z2/181/27.

It has been demonstrated that under normal laboratory conditions Lymnaea truncatula infected with Fasciola hepatica are not able to shed all their cercarial loads before their death. Cercariae are irregularly released over an extended period and considerable variations (4–1789) occur in their output.

Laboratory studies have revealed that the green parts of grass (Dactylis glomerata) are preferred to the brown, dead parts by the cercariae of Fasciola hepatica for metacercariae formation. More cysts are formed on the lower surface of submerged leaves than on the upper surface.

There is vertical zonation of metacercariae on grass in water. More cysts occur on the upper parts of submerged grass than on the lower parts.

Vertical migration of cercariae after settling does not usually occur before cyst formation.

The results are tabulated and discussed and correlated with conditio s in the field.

This work was undertaken in the Department of Zoology, University of Bristol, Bristol, under the supervision of Dr H. D. Crofton, to whom I wish to express my gratitude.

Microfilariae belonging to the genera Brugia, Loa and Wuchereria have been stained by a wide variety of histochemical methods. The results are recorded in eight Tables.

The sheath of the microfilaria contains polyanionic material, probably polysulphate, periodate reactive mucosubstance, sulphur-protein and complex lipids. The fluid within the sheath is also polyanionic.

The cuticle of the microfilaria is stabilized by disulphide bonds possibly in a collagen but it was not possible to demonstrate collagen by staining. The cuticle also contains a periodate reactive G mucosubstance and complex lipids. The annular grooves of the cuticle also contain a periodate unreactive mucosubstance. Certain areas of the cuticle, around the excretory pore, the pores of the Schwanzgebilde, and the cephalic structures - oral ring, hook and spines - are hardened by deposition of a keratinous scleroprotein. Some reactions of the cuticle may be masked by the presence of basic protein.

The excretory vesicle contains polyanionic material but there was no clear evidence that the polyanionic material within the fluid in the sheath came from the excretory vesicle. The excretory vesicle, the anal vesicle and the subcuticular cells contain periodate reactive G mucosubstance. Most RNA staining was found in the excretory vesicle, the excretory cell, the anal vesicle, the rectal cells and the G cell.

The pharyngeal thread, the Innenkörper, the rectal cells and the anal vesicle represent the modified gut of this first stage larva. The pharyngeal thread appears to consist largely of a protein-mucosubstance complex that can be unmasked by oxidation. The Innenkörper contains a basic protein and acidic mucosubstance, possibly sialomucin, complex that can be unmasked by adjusting the electrolyte concentration of the staining media.

The relevance of some of these techniques in the analysis of possibly unique invertebrate structures is discussed briefly.

We thank Dr D. A. Denham, Mr P. R. N. Pester and Dr T. Ponnudurai in particular, and many other correspondents, for supplying us with microfilariae for staining. This work was supported by a grant from the Medical Research Council of Great Britain.

An experiment was performed to test whether the larvae of the sciomyzid fly Dichetophora biroi are able to ‘recognize’ and reject Lymnaea tomentosa snails infected with Fasciola hepatica.

The analysis of the results indicates that the larvae do not show any obvious preference when given the simultaneous choice of non-infected and infected snails, and also that, under laboratory conditions, the latter are not more vulnerable than the former to the larval predation.

I wish to express my sincere gratitude to Dr R. P. Herd for his help and encouragement during this work. I am also grateful to Professor C. O. Berg, Cornell University (U.S.A.); Dr J. J. Lynch, C.S.I.R.O. Pastoral Research Laboratory, Armidale (N.S.W.) and Dr M. D. Rickard, University of Melbourne, for valuable advice and fruitful discussions. Dr J. A. Thomson, University of Melbourne, critically read the manuscript.

A quantitative study has been made of the relationship between Moniliformis dubius and Periplaneta americana.

1. Eggs removed from the body cavity of the worm hatch at a higher rate after dessication and after a short period of time following removal.

2. There is an exponential decline in the additional number of cystacanths recovered as the dose is increased linearly.

3. Female cockroaches are more susceptible to high levels of infection than males.

4. Direct injection into the haemocoele produces a similar recovery rate to that from oral infection.

5. Previous infection does not seem to affect the success of a superimposed infection.

6. Rates of development and growth are described.

7. The effect of the parasite on the concentration of free amino acids in the haemolymph was found to be negligible.

I would like to thank Miss S. Arnold and Mr D. Barnard for technical assistance, Mr E. Walters for help with statistical analysis, Dr D. W. T. Crompton for many helpful discussions and Dr P. J. Whitfield for reading this paper. This work has been carried out during the tenure of a Science Research Council Studentship.

The ability of various mammals to transmit Dracunculus medinensis has been investigated. The most suitable definitive host for laboratory transmission was the rhesus monkey (Macaca mulatta) and, in 11 out of 14 of these animals female worms emerged 321 to 371 days after first infection. The parasite has now been maintained in monkeys for four laboratory cycles. Five of 11 dogs and 17 of 30 cats had developing parasites in their tissues when autopsied, but no worms emerged from two dogs left for over a year and only a single worm from one of nine cats. Monkeys showed a seemingly haphazard pattern of susceptibility to infection and reinfection, although none of the 17 animals exposed was completely refractory. Cyclops vernalis americanus (Marsh) was an easily maintained and efficient intermediate host; a 90% infection rate with an average of two infective larvae per Cyclops was obtained.

This work was supported by a grant from the Tropical Medicine Research Board through the Ministry of Overseas Development. Thanks are due to Mrs Vivian Duckett for technical assistance.

Maximum levels of protective antibody activity were reached in the circulation of rats given a primary infection or one or two re-infections of Plasmodium berghei at the time the infection was finally eradicated. This association of maximum protective antibody activity with complete clearance of the parasite was also found in recipients of lymphoid cells from immune donor rats and suggests that a high level of protective antibody activity is one essential factor in complete elimination of the parasite.

Excellent technical assistance was given at different times by Mrs J. Page, Mr A. J. Edwards and Miss D. Leader.

Development of cercariae within intramolluscan germinal sacs is recognized to be in seven stages; in brief, these stages are: 1, germinal cell; 2, naked cell aggregate; 3, germ ball prior to elongation; 4, elongating germ ball; 5, tail bud stage; 6, tail elongation stage; and 7, fully developed cercaria.

Schistosoma mansoni cercariae at each of the developmental stages is described.

The dimensions of both germinal and somatic cells comprising the bodies of developing S. mansoni cercariae decrease with maturation.

The surfacial membrane of Stage 3 cercarial embryos is formed by a number of small somatic cells which migrate to the surface and coalesce.

The chronologic and partially synchronized development of S. mansoni cercariae is presented.

The body-tail ratio during S. mansoni cercarial development is presented.

Thirty-five lambs (14 weeks old), the progeny of Border Leicester rams mated with Border Leicester × Merino ewes, were divided at random into two groups. Seventeen lambs in Group A received 3500 Haemonchus contortus larvae weekly and 18 lambs in Group B received 500 larvae each day: the dosage was decreased to 1400 larvae weekly and 200 larvae per day after 8 weeks.

Pathogenicity was found to be higher when the lambs were infected with a small number of larvae daily (daily group) than when the same number of larvae were given in one dose at weekly intervals (weekly group). The period of survival was consistently shorter in the former group.

No significant difference in faecal egg counts was observed between the two groups.

Reduction in food intake per head per day and in live weight were more pronounced in the ‘daily group’ than in group dosed weekly. The discrepancy in food consumption and live weight between these two treatments was more apparent at the end of the experiment.

Some resistance was observed in both groups but the level of resistance was not high.

Accumulation of inhibited larvae was apparent after 104 days in the ‘weekly group’ and after 78 days in the ‘daily group’. An infection arising from a large number of larvae given at weekly intervals induced a mild form of resistance leading to an accumulation of inhibited larvae at the end of the experiment. This was less apparent in lambs infected with the same number of larvae given each day.

Interference with ovulation in adult females was recorded in both treatments. Sex discrimination against the female population both in the extent of elimination of adults and in the retardation of development of fourth-stage larvae was recorded and the intensity of discrimination was more pronounced with the weekly regime of larger doses than with the same number of larvae but given in daily doses. The worms were eliminated sporadically and not continuously. A challenge infection of 10000 larvae given to two lambs in the ‘weekly’ regime treatment on the 107th day initiated a partial selfcure reaction demonstrated by an immediate fall in faecal eggs per gram (i.e. Epg) but the egg count returned to the original value in 28 days.

Thirty-five lambs (14 weeks old), the progeny of Border Leicester rams mated with Border Leicester × Merino ewes were divided at random into two groups. Seventeen lambs received 3500 Haemonchus contortus larvae at weekly intervals (weekly group) and 18 lambs received 500 larvae each day (daily group). The dosage was decreased to 1400 and 200 larvae respectively after 8 weeks.

The daily infection with a small number of larvae induced a severe anaemic condition with the blood picture falling to 7% PCV; 2% Hb conc. and 1·6m/mm3 RBC count in 78 days. The anaemia with weekly dosing of larvae was less severe with equivalent figures of 10–12% PCV; 4% Hb conc and 3·6m/mm3 RBC count.

Two phases were observed in the changes in the sodium (Na+e) and potassium (K+e) electrolyte concentrations. In the first phase (K+e) increased and (Na+e) decreased, while in the second phase both electrolytes increased. The influx of young red blood cells into the circulation is considered as the factor responsible for these changes.

Adult desert locusts were experimentally infected with a dose of 50 M. nigrescens ova and larval nematodes removed 14, 17, 21 and 24 days afterwards for in vitro radiotracer studies. These nematodes were incubated under controlled conditions either in a nutrient medium or in insect saline, which contained the appropriate 14C or 3H-labelled radioisotope. The rate of incorporation of the radioisotope into the proteins of the nematode was measured.

The rate of incorporation of 14C-leucine into proteins was greatest for the 17- day-old larvae and this accorded with the in vivo growth pattern of the nematode. This isotope was incorporated less rapidly into proteins by the 14-, 21 and 24-day- old larvae, which synthesized proteins at approximately the same rates as each other. A non-linear rate of incorporation was manifest for each of the larval stages.

Total dry weight and protein determinations showed that the nematode synthesizes relatively large quantities of non-proteinaceous reserves during the third week of infection, whilst proteins are preferentially synthesized between the 21st and 24th days of infection.

The 17-day-old nematode larvae were able to incorporate six 14C-amino acids into proteins to varying degrees. Leucine was incorporated most effectively into nematode proteins, whilst glutamic acid was the least incorporated of the six amino acids.

These nematode larvae could not effectively metabolize the 3H-dipeptide histidyl-leucine and incorporate its component amino acid residues into proteins. Similarly, a preparation of 3H-haemolymph proteins were not significantly metabolised and incorporated into proteins by the nematode. However, the nematode metabolized and incorporated an exogenous source of 14C-glucose into proteins at relatively high levels.

Incorporation of 14C-luecine into proteins was seven times more rapid for nematodes incubated in buffered insect saline than for those incubated in a more complete nutrient medium.

The dietary requirements of the nematode for protein synthesis were discussed in relation to their associated effects upon the host metabolism and an active transport of dietary amino acids into the nematode was postulated.

We wish to thank Dr H. L. Speer and Dr K. K. Nair for their helpful suggestions regarding radioisotope techniques. Thanks are also due to the National Research Council of Canada for providing financial assistance (Grant No. A4679).