The cellular mucosal and systemic effectors of gilthead sea bream (GSB) (Sparus aurata) involved in the acute immune response to the intestinal parasite Enteromyxum leei were studied in fish experimentally infected by the anal route. In the intestinal inflammatory infiltrates and in lymphohaematopoietic organs (head kidney and spleen) of parasitized fish, the number of plasma cells, B cells (IgM immunoreactive) and mast cells (histamine immunoreactive) were significantly higher, whereas the number of acidophilic granulocytes (G7 immunoreactive) decreased, compared with non-parasitized and unexposed fish. These differences were stronger at the posterior intestine, the main target of the parasite, and no differences were found in the thymus. In non-parasitized GSB, the percentage of splenic surface occupied by melanomacrophage centres was significantly higher. These results suggest that the cellular response of GSB to E. leei includes proliferation of leukocytes in lymphohaematopoietic organs and recruitment into intestines via blood circulation involving elements of innate and adaptive immunity. Acidophilic granulocytes and mast cells presented opposite patterns of response to the parasite infection, with an overall depletion of the former and an increased amount of the latter. Some differences between both cell types were also detected in regard to their granule density and cell morphology.

Rats thymectomized at birth were infected with Nippostrongylus brasiliensis at 6 weeks of age. In some, but not all, rats egg production was greatly prolonged after the primary infection. Thymectomized rats were less resistant than controls to second infections.

In rats infected at birth the development of resistance was significantly delayed. Neonatally infected rats, when they were 6 weeks old, were less resistant to reinfection than adult controls. Rats infected three times in the neonatal period were more resistant to challenge at 6 weeks of age than rats infected once at birth, but less resistant than adult controls. The degree of immunity induced by a single neonatal infection was variable; some rats showed little or no resistance to reinfection 6 weeks later.

Antibody formation, measured by the occurrence of reagins, was altered in the same way by both neonatal thymectomy and neonatal infection. In the thymectomized (immunologically deficient) and newborn (immunologically immature) rats, reagin formation following initial infection was delayed; there was a reduced anamnestic response after challenge; mean titres were lower and more variable than in control animals; and in some rats circulating reagins were never detected.

In thymectomized rats the capacity to demonstrate anaphylaxis was as variable as in control rats, but in the few thymectomized rats in which egg production was prolonged, anaphylaxis was slight at a time when most intact rats had developed the capacity to show anaphylaxis. Rats infected at birth and adult rats infected at the same time were equally susceptible to systemic anaphylaxis 6 weeks later, although by this time there was a marked difference in the degree of resistance to the parasite developed by the two groups.

In adult rats splenectomy before, during or after initial infection had no effect either on the rate of development, or the degree of immunity induced by primary or secondary infections, or on antibody formation.

We would like to thank Miss W. Griffiths and A. J. Edwards for their willing technical assistance. This work was done whilst one of us (B.M.O.) was the holder of a Wellcome Fellowship of the Animal Health Trust.

The results are presented of an experimental investigation of the relationship between faecal egg counts and worm burden in outbred MF1 mice infected with Heligmosomoides polygyrus (Nematoda). The sex-ratio of the adult parasites was found to be significantly female biased, but independent of parasite burden. Evidence was also found to suggest a significant association between parasite density and mating success. The linear logarithmic relationship (gradient of approximately 2) between the variance and mean of repeated egg counts from individual mice could not be disrupted by controlled experimental conditions, and is therefore concluded to be an inherent feature of the biology of helminth-host interactions. A significant positive association was detected between host faecal output and parasite fecundity; further investigation of the cause of this association is now required. Average faecal production was found to be inversely related to worm burden. In combination, the above two factors account partly, but not completely, for the observed density dependence in the absolute rate of parasite egg production. Two epidemiological consequences of the relationship between faecal production and parasite fecundity are discussed. First, absolute rates of egg production can only be estimated by temporal egg counts (epd). Secondly, measurements of egg production/g of faecal material (epg) are more reliable indicators of worm burden than measurements of epd. Both faecal output and worm fecundity respond as might be predicted to a period of host food deprivation; faecal egestion and measurements of epd are signifcantly depressed, and measurements of epg are significantly increased. The data are consistent with the hypothesis that egg production and faecal output are causally associated, but that other factors relating to parasite density also influence egg production. Of the possible causes for the high variability in parasite egg production between mice harbouring low number of worms, the most likely explanation seems to be host heterogeneity (possible genetic in origin). This is discussed in relation to similar patterns observed in data collected from human populations in areas of endemic helminth infection.

The behaviour of oncomiracidia of the monogenean skin parasite Entobdella soleae, after hatching without chemical stimulation, has been studied in vertical, 4 cm high chambers illuminated from above. Most freshly hatched larvae are photo-positive and swim to the top of the vessel, where they remain actively swimming. Occasionally larvae become photo-negative and make a brief excursion to the bottom of the vessel. As the larvae grow older, these photo-negative excursions become more frequent and larvae spend progressively longer in the photo-negative phase of behaviour, but, even after many hours, photo-positive vertical excursions are still occurring. If this pattern of alternating photo-positive and photo-negative vertical movements occurs in the natural environment, these vertical movements, coupled with horizontal transport of larvae by water currents, would provide a search pattern for the host, the diurnally-inactive, bottom-living flatfish Solea solea. When eggs are stimulated to hatch in the chambers with urea, large numbers of larvae emerge within a few min, and it was observed that during the period of about 30 min after stimulation, many larvae are photo-negative. When eggs on the sea bottom are stimulated to hatch by mucus or urine from a nearby host, such behaviour would greatly enhance chances of contact with the fish. When eggs are stimulated to hatch by means of urea in the absence of light, there is evidence that the oncomiracidia are geo-negative. It has been shown that larvae are able to extricate themselves in total darkness from sediment containing buried eggs and that, in an experimental situation illuminated only by an infra-red beam, larvae readily attach themselves to the lower skin of soles within 2 min of hatching. There is evidence that, in aquaria, some invasion of the host takes place from below.

The prevalence of the caecal threadworm Trichostrongylus tenuis in red grouse in the north of Scotland was high despite low grouse densities. Prevalence, intensity and aggregation of threadworms was higher in old than in young grouse. Infections were long-lasting: populations of adult worms could survive for over 2 years in grouse, with little mortality. Parasite egg output decreased with the age of a worm population, largely as a result of a decrease in the fecundity of ageing female worms. Seasonal variations in worm fecundity were also evident.However, there was no evidence of an intensitydependent decrease of worm fecundity with increasing worm numbers in either captive or wild grouse. The long life and high reproductive capacity of T. tenuis probably contribute to its effective transmission and high prevalence.

Of two groups of calves, one was regularly treated with the cortisone-derivative, β-methazone. This treatment greatly reduced the ability of the calves to produce antibody and gave rise to a number of side effects. Calves of both groups were given 1500 infective larvae of Ostertagia ostertagi daily. A milder régime of cortisone administrations using prednisolone was employed in a second experiment in which two comparisons were made. Worm burdens of calves infected with 1500 larvae daily and receiving cortisone were compared with those of control calves infected at the same rate and cortisone-treated calves receiving 1500 larvae per day were compared with cortisone-treated calves infected at twice this rate.

By periodic post-mortem examination it was shown that even in cortisone-treated animals the worm burden was regulated at a level only slightly higher than in control calves, apparently by a density-dependant loss of worms. A more rapid loss of worms which occurred for a period in the control calves was prevented in cortisone-treated animals unless the infection rate was high.

The limit set by the normal host to the total egg output of its worm population was raised by cortisone treatment, very greatly in the first experiment, rather less in the second in which the cortisone treatment was less severe. The effect of host resistance on the length of worms and on the incidence of females without vulval flaps was greatly reduced by cortisone. The inhibition of development was not affected.

We studied the pattern of infection and the inter-annual variation and individual factors affecting the infection of 2 species of nasopharyngeal bot flies, Cephenemyia auribarbis and Pharyngomyia picta (Diptera: Oestridae), in a population of Iberian red deer (Cervus elaphus hispanicus) from south central Spain (10 annual periods between 1990 and 2003). Mean prevalence±S.E.95%CI of infection was 35·19±4·24% (n=486). The frequency distribution of the parasites was markedly aggregated (K: 0·213, mean abundance±S.D.: 5·49±12·12). Parasite load of Oestridae peaked at calf and subadult age groups and declined thereafter, which suggests that acquired immunity may be acting. In common with other host–parasite relationships, male hosts were found to have higher prevalence and abundance levels than females. The prevalence of P. picta was positively affected by the presence of C. auribarbis whereas the intensity of infection of P. picta was negatively affected by the presence of C. auribarbis. Intensity of P. picta in concomitant infections with C. auribarbis was lower than in pure P. picta infections, whilst the intensity of C. auribarbis infections did not change. This provides good evidence of interspecific competence, which could be dealt with by parasites by means of asynchronous life-cycles and different maturation periods. Weather also affects the dynamics and transmission rates of these parasites. Previous annual rainfalls positively affected the level of infection with oestrids. Yearly autumn rainfalls affected positively P. picta, possibly due to an effect on the pupal stage survival. Infection of Oestridae affected body condition in calves and subadults, suggesting that oestrids could have sublethal effects on Iberian red deer. Future research is needed to investigate the effect of parasites on the dynamics of the Iberian red deer.

The discrete typing units (DTUs) of Trypanosoma cruzi that infect domestic dogs and cats have rarely been studied. With this purpose we conducted a cross-sectional xenodiagnostic survey of dog and cat populations residing in 2 infested rural villages in Pampa del Indio, in the humid Argentine Chaco. Parasites were isolated by culture from 44 dogs and 12 cats with a positive xenodiagnosis. DTUs were identified from parasite culture samples using a strategy based on multiple polymerase-chain reactions. TcVI was identified in 37 of 44 dogs and in 10 of 12 cats, whereas TcV was identified in 5 dogs and in 2 cats –a new finding for cats. No mixed infections were detected. The occurrence of 2 dogs infected with TcIII –classically found in armadillos– suggests a probable link with the local sylvatic transmission cycle involving Dasypus novemcinctus armadillos and a potential risk of human infection with TcIII. Our study reinforces the importance of dogs and cats as domestic reservoir hosts and sources of various DTUs infecting humans, and suggests a link between dogs and the sylvatic transmission cycle of TcIII.

The relationship between cell-mediated granulomatous inflammation and transmission of disease in schistosomiasis and tuberculosis has been explored. In 2 experiments involving Schistosoma mansoni-infected normal and T cell-deprived mice, and infected deprived mice that had been variously reconstituted with immune or normal lymphocytes or immune serum, there was a significant positive numerical correlation between mean liver granuloma diameters and faecal egg counts in individual animals. Lymphocytes from donors with recently patent infections were more active than cells from chronically infected or uninfected donors in reconstituting egg excretion rates in deprived recipients, and mesenteric lymph node (MLN) cells were more active than spleen cells. Modulation of granulomatous activity with increasing chronicity of infection in the donors, resulting in a decrease in granuloma size around freshly produced tissue-bound eggs, was paralleled by a waning of the capacity of transferred lymph node cells to reconstitute egg excretion in the recipients. Serum taken from chronically infected donor mice over the same period and transferred to infected deprived recipients became more active in enhancing egg excretion in the recipients as the cell-mediated activity declined. A recent study in Kenya has found that S. mansoni-infected patients with concurrent human immunodefficiency virus (HIV) infection excrete fewer eggs than patients exposed to the same levels of schistosome infection, but who are not HIV-infected, thus indicating that schistosome egg excretion in humans is also immune-dependent. Attention is drawn to an apparently parallel situation in human tuberculosis, another pathogen which induces a cell-mediated granulomatous immune response. Several studies have shown that patients with tuberculosis who are also HIV-seropositive tend to have fewer tubercle bacilli detectable in their saliva than those with tuberculosis, but who are HIV-negative. This discrepancy, associated with differences in lung pathology in HIV-positive patients, suggests that in tuberculosis immune cell-mediated granulomatous inflammation causes the destruction of host tissue in a manner which facilitates onward transmission of the bacterial pathogen.

Immunity to Eimeria maxima, judged by oocyst production resulting from a low challenge inoculum, was evident when the challenge was given as early as 3 days after the immunizing dose of 250 or 500 oocysts. Immunity was complete, i.e. no oocysts were produced, to challenge at 7 days after an initial inoculum of 500 oocysts, or at 14 days after 250 oocysts. Judged by the above criteria 250 oocysts of E. tenella did not induce immunity to challenge given at 3, 5, 7 or 14 days. The findings are discussed.

A mathematical model of the parasitic phase of the life-cycle of Ostertagia ostertagi in calves is described. The model is used in the re-analysis of previously published data from a long-term trickle infection experiment in which groups of calves were infected daily with graded doses of 3rd-stage (L3) larvae. The results of the analysis are consistent with the hypothesis that the observed changes in the intensity of infection in the calves were the result of a decline in the proportional establishment of ingested L3 larvae, and a rise in the death rate of the 5th-stage worms as the duration of exposure to infection increased. The proportion of ingested L3 larvae that become established in the mucosa can be described as an exponential decay function of the duration of the infection. Within the range of trickle intensities investigated, the function appears to vary independently of the level of exposure to infection. In contrast, the rate of mortality of the 5th-stage worms appears to be an increasing linear function of the total cumulative number of 3rd-stage larvae administered.

A 3000 bp cDNA insert (G6/1) from Giardia duodenalis, cloned into Escherichia coli is located on chromosome 3 of Giardia stocks which have 3 chromosomes detectable by field-inversion gel electrophoresis in the range 650–800 kb and on chromosome 3 and/or 4 of stocks with 4 chromosomes in this size range. The loss of this sequence from chromosome 4 but not chromosome 3 was associated with the induction of drug resistance in a previously sensitive laboratory stock. G6/1 appears to represent a single copy gene in Giardia as determined by hybridization of the probe to cleaved genomic DNA. Furthermore, the sequences flanking at least 12 kb of G6/1 are the same when G6/1 appears on both chromosomes 3 and 4. The cDNA encodes a protein associated with the nuclei of trophozoites during some stages of growth of the parasite. In a non-dividing culture, the antigen is associated with the nuclei of about 30% of trophozoites and fewer in a dividing culture. Three Giardia stocks with obvious chromosome rearrangements, which grow poorly and fail to divide normally, apparently lack the DNA sequence G6/1.

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.

The effect of waterborne zinc (Zn) on Gyrodactylus population dynamics was studied on isolated guppies maintained at concentrations ranging from 0 to 240 μg Zn/l. After 1 week pre-exposure to Zn, each fish was experimentally infected with 3 gyrodactylids and parasite numbers were recorded daily on each fish until the fish either died or recovered from infection. Parasite establishment was most successful at 0 and 240 μg Zn/l (97%) compared with the intermediate Zn concentrations. Low to moderate concentrations of Zn were beneficial to the parasite, as evidenced by the concentration-dependent increase in peak parasite burden on recovered fish up to 120 μg Zn/l. In contrast, 240 μg Zn/l may have been toxic to the parasite, as both peak parasite burden (in fish that recovered from infection), and maximum rate of increase of the parasite population (in fish that died) declined at this concentration. The combined effect of infection and Zn is harmful to the fish, because mortality of infected fish (but not uninfected fish) increased with increasing Zn concentrations. We suggest that the observed mortality occurs because of the inability of fish to continuously produce mucous that is a key factor for protecting fish from both waterborne Zn and ectoparasites.

The effects of a range of FMRFamide-related peptides (FaRPs) on isometric contractility were tested using isolated muscle strips from the liver fluke, F. hepatica. The neuropeptides tested were the molluscan FaRPs, FMRFamide and FLRFamide, the turbellarian FaRPs, RYIRFamide and GYIRFamide, the cestode peptides, NPF and GNFFRFamide, and the nematode FaRPs, AF-1 (KNEFIRFamide), AF-2 (KHEYLRFamide), AF-8 (KSAYMRFamide), and PF-4 (KPNFIRFamide). Dose-response experiments were undertaken at a concentration range of 5 nM–5 μM for all of the neuropeptides tested. FMRFamide and AF-8 caused statistically significant increases in the amplitude and frequency of contractions at concentrations of 0·5 μM and 5 μM. FLRFamide and AF-2 also caused significant increases in contraction frequency at concentrations of 0·5 μM and 5 μM, although a significant increase in amplitude of contraction was observed only at a concentration of 5 μM. GYIRFamide increased both amplitude and frequency significantly at concentrations of 50 nM, 0·5 μM and 5 μM. RYIRFamide significantly increased frequency of contractions at concentrations of 0·5 μM and 5 μM, but failed to have a significant effect on contraction amplitude. AF-1 at a concentration of 5 μM increased contraction amplitude, but failed to have an effect on frequency at any of the concentrations used. PF-4 caused a statistically significant increase in both the amplitude and frequency of contractions at a concentration of 5 μM. NPF and GNFFRFamide had no effect on the in vitro motility of F. hepatica over the range of concentrations tested. The results are discussed in the light of possible structure–activity relationships in the FaRPs tested.

The development of the exsheathed third-stage larva of Haemonchus contortus to the fourth stage in vitro has been investigated in half-strength Ringer's solution and in solutions with ionic concentrations similar to those found in the abomasum during normal secretion of acid and during heavy infestations of Haemonchus. The concentration of carbon dioxide required for the optimum development of the larva has been shown to be dependent on the pH of the medium. At a low pH the optimum pC02 is low and the larva is less sensitive to a range of carbon dioxide concentrations than at a pH near neutrality. Gastric mucin and histamine have been shown to have an inhibiting and a potentiating effect, respectively, on development, in media at pH 6.

Following a 2–5 ml meal containing 2 g (80%) glucose and 30 mg methionine administered by stomach tube at 09.00 h, H. diminuta was found to change its position in the rat's intestine. Usually while the stomach contained food ≤ 45 %, by weight, of the tapeworm lay in the posterior 40 % of the small intestine, after the stomach was empty the reverse was true.

The effect of hyperosmotic fluid entering the duodenum was determined by injecting, following laparotomy, varying amounts of 80 % glucose and 1 ml of varying concentrations. A volume ≤ 0-4 ml of 80 % led to the death and expulsion of worms within 7 h, and 1 ml of < 50 % glucose also killed the worms.

The expulsion of 15 % of the tapeworms from rats given 2–5 ml hypertonic meal by mouth was correlated with the failure of the stomach in some rats to regulate the stomach emptying. Worms in these rats were found from 4 h onwards in the caecum and large intestine, and by 9 h in the faeces.

That the movement of worms, other than those expelled, was not caused by a sub-lethal flux of hyperosmotic fluid was shown by feeding an iso-osmotic meal to rats; the worms moved to a more posterior position during the day, but none was expelled.

Single worm infections in rats kept on a normal feeding regime until 07.00 h and then starved throughout the day also showed a posterior migration. Using a method which involved the simultaneous cutting of the small intestine into 5 cm sections, it was possible to show that this movement involved the whole worm, the scolex moving from a mean position 10–15 cm to one 20–25 cm from the stomach, and the terminal proglottid moving from approximately 75 cm to 95 cm from the stomach.

Worms in Wistar rats responded similarly to those in SPF CFE (Sprague Dawleys) with the exception that there was some evidence that the stomach emptying of a hyperosmotic meal was better controlled and hence a smaller percentage of the worms were expelled.

The results are discussed in relation to diurnal changes in the level of the external site location stimulus which other workers have shown to exist by surgically transplanting H. diminuta into the ileum.

The enthusiastic assistance of Miss Helen Stallard, B.Sc., and Miss Patricia Grant is gratefully acknowledged, and financial support from the Medical Research Council, London (Grant No. G967/164/T).

The development of the free-living stages of Ostertagia ostertagi incubated at constant temperatures from 5 to 27 °C and in dung pats in the field was observed. The death rates were adequately described by a single exponential function and the distribution of times for development by a delayed gamma density function. The times of appearance of selected development stages such as maximum numbers of embryonated eggs and half the maximum number of infective larvae, were highly dependent on temperature. The relationships between these times and temperature were adequately described by the Arrhenius equation. The parameters of this equation and mean hourly temperatures recorded in dung pats in the field were used to predict development times in the field. A comparison of predicted and observed times showed that an initial delay accounted for a large proportion of the time required for development in dung pats, particularly the embryonation of eggs. This delay was attributed to lack of aeration associated with high moisture content of the dung pat.

Arecoline hydrobromide was tested for its efficiency in causing dogs to purge and in removing hydatid worms by infecting, dosing and autopsying 157 dogs. Doses were given by mouth in water at various rates, by mouth in meat, and by subcutaneous injection.

The tabulated results indicate that the median efficiency of hydatid-worm removal, with doses of or above grain/10 lb. weight, are 95% or more. Dosing in meat is almost equally efficient but the action is slower, taking usually over 1 hr. instead of 15–30 min. Injections of arecoline caused purgation but not worm removal. Increasing the dosage rate above grain/10 lb. body weight increased the proportion of dogs that vomited and caused convulsions in some, but did not markedly increase efficiency in worm removal. The dose recommended for general use is grain/10 lb. body weight; though half or double this dose is about equally suitable. None of the dogs tested was killed by arecoline.

Repeated doses were given to see how many were required to ‘clear‘ a dog. Dogs usually gave negative results after three doses that caused purging.

Arecoline showed a similar high degree of efficiency against Taenia species and Dipylidium; usually removed about half the Toxocara specimens present, and had no significant effect against hookworms.

Solutions of arecoline were tested in vitro on isolated Taenia segments arranged to record. Solutions of 0.001% or more in Ringer's solution almost invariably caused extreme relaxation of the segment, its normal rhythmical contractions ceasing. The segments were not killed.

Arecoline hydrobromide appears to exert its anthelminthic action: first, by causing the muscles of cestodes to relax, so that the hold on the host's intestine is lost; secondly, by causing the host to purge, so that the detached worms are removed from the host's body.

The writer wishes to thank Sir Louis Barnett, Dr Hercus and other members of the Hydatid Committee for their interest and support. She especially thanks Miss D. Medley, Hydatid Assistant, who has carried out the heavy share of the infecting of dogs and handling of faeces. Thanks are also given to Mr L. K. Whitten, Agricultural Parasitologist, for technical advice; to Mr Grimmett, for analysing arecoline-content of pills; and to the various members of the Medical School staff who have given helpful advice.

A Polymerase Chain Reaction (PCR)- based assay developed for the specific identification of Thelazia gulosa, Thelazia rhodesi and Thelazia skrjabini (Nematoda, Spirurida), which cause bovine ocular thelaziosis, was evaluated for its usefulness in detecting the intermediate hosts and in estimating the infection prevalence of vectors in field conditions throughout 5 years (from 1997 to 2001). A total of 5190 flies were captured and identified as Musca larvipara, Musca osiris, Musca autumnalis, Musca tempestiva or Musca domestica. Genomic DNA was extracted from pools constituted by heads, thoraces, abdomens and wings of 10 flies of each species, and 2076 samples were subjected to a PCR assay to specifically detect the ribosomal ITS-1 sequence of bovine Thelazia. Amplicons were sequenced and subjected to digestion with CpoI restriction enzyme. M. autumnalis, M. larvipara, M. osiris and M. domestica species were shown to be PCR positive. T. gulosa was specifically detected by PCR in M. autumnalis, M. larvipara, M. osiris and M. domestica, whereas T. rhodesi is in M. autumnalis and M. larvipara. Of 27 positive samples, 23 were positive for T. gulosa and 4 for T. rhodesi, with a mean prevalence of 2·86% in the whole fly population collected. The highest mean prevalence values of infection were detected in M. autumnalis (4·46%) and M. larvipara (3·21%), and the former species was confirmed to be the vector of T. gulosa and T. rhodesi. This study is the first report of M. osiris as a vector of T. gulosa and M. larvipara as a vector of T. gulosa and T. rhodesi under natural conditions. The occurrence of Thelazia in fly populations in the Apulia region of Italy (in the 5 grazing seasons considered) indicates that cattle thelaziosis is enzootic in southern Italy. This molecular assay should be a useful epidemiological tool for assessing the role of different species of flies as intermediate hosts of thelaziae.