Using laboratory-bred natural rodent hosts that had been castrated and then implanted with either testosterone or inert oil, we have shown that testosterone causes prolonged and more intense infections of a tick-borne piroplasm, Babesia microti. This will result in more ticks becoming infected while feeding. Sexually active male rodents with high testosterone levels are also known to show increased locomotory activity and reduced innate and acquired resistance to tick feeding, so that more ticks are likely to be picked up and then fed successfully by these hosts. As a result, the transmission potential of B. microti is significantly increased via hosts with high rather than low testosterone levels. It is argued that testosterone helps to generate the observed aggregated distributions of parasites amongst their hosts, which also enhances parasite persistence.

To understand possible factors controlling transmission of trematode larvae between first and second intermediate hosts we examined the impact of ambient fauna on parasite transmission in a marine intertidal parasite-host association. Cockle hosts (Cerastoderma edule) kept together with selected co-occurring macrozoobenthic species in mesocosms acquired a lower parasite load compared to cockles kept alone, when targeted by cercariae of the trematode Himasthla elongata. The reduction of parasite load in the cockles differed between the 7 macrozoobenthic species tested and was between 35 and 91%. Three different types of reduction could be distinguished: (1) predators (Carcinus maenas, Crangon crangon) actively preying upon cercariae, (2) non-host filter feeders (Crepidula fornicata, Mya arenaria, Crassostrea gigas) filtering cercariae but not becoming infected and (3) alternative hosts (Mytilus edulis, Macoma balthica) becoming infected by the cercariae and thus distracting cercariae from the target hosts. In addition, interference competition may occur in the form of disturbance of cockles by ambient organisms resulting in lower filtration rates and subsequently lower parasite loads. Our results suggest that the species composition and relative abundance of the ambient fauna of parasite-host systems play an important role in controlling trematode transmission rates in benthic marine systems.

Opisthorchis viverrini infection is an endemic disease that causes a serious public health problem in southeast Asia, especially in northeast Thailand. We have developed a PCR method using a pair of primers named OV-6F/OV-6R for detecting O. viverrini eggs in stool samples and compared it with Stoll's egg-count method. The primers were designed based on the pOV-A6 specific DNA probe sequence which gave a 330 base pair product. The PCR method can detect a single egg in artificially inoculated faeces or as little as 2×10-17ng of O. viverrini genomic DNA. The method gave 100% sensitivity in all hamster groups except in animals exposed to the lowest intensity of infection (1 metacercaria/hamster). In the first month of infection, the PCR method was more sensitive than using the egg-count method in all infected groups especially in the light infections. The PCR method was also successfully used in monitoring a therapeutic study. Since the PCR method showed no cross-reaction with Heterophyid flukes, it can be useful for specific identification of O. viverrini eggs in stool samples without the risk of false positives. It also has great potential for application in clinical epidemiological studies.

An assessment is made of the repeatability of parasite community structure in space for a marine fish, and in space and time for a freshwater fish from south-eastern Mexico. The marine fish species was the red grouper, Epinephelus morio (collected from 9 localities), and the freshwater species was the cichlid, Cichlasoma urophthalmus (collected from 6 localities: including monthly at 2 localities for 1 year, and bimonthly at 1 locality in 1990 and 1999). Pairwise interspecific associations and analyses of nested patterns in the distributions of parasite species among hosts were used in both fish species, with comparisons over time made only with the cichlid. Positive interspecific associations, and nested patterns were noted in some localities for both fish species, and/or at some sampling times for the cichlid fish. However, non-random patterns in the structure of parasite communities in these 2 host species only were observed sporadically. When present, nestedness in both fish species was apparently linked with a positive association between total infection intensities and fish size. Additionally, adjacent localities were more likely to display similar parasite community structure than distant ones. This preliminary result suggests that distance between localities is an important determinant of predictability in parasite community structure.

Neospora caninum is a cyst-forming parasite that causes abortion in cattle. Despite this parasite's ubiquitous distribution and wide host range, the number of N. caninum isolates obtained to date is limited. In vitro isolation of the parasite is arduous and often unsuccessful. In addition, most isolates have been obtained from clinically affected hosts and therefore could be biased towards more virulent isolates. In this report, an improved isolation approach from transplacentally infected newborn calves was undertaken and 9 new isolates were obtained. Moreover, a microsatellite technique was applied to investigate the genetic diversity of these isolates. Most isolates showed specific genetic profiles. However, the Nc-Spain10 isolate was identical to the previously described Nc-Spain1H isolate and Nc-Spain3H was identical to Nc-Spain4H. These isolates were likely to have identical genotypes because they were isolated from distinct calves of the same herd. Future pathogenic characterization of these isolates will contribute to the investigation of the relationship between isolate virulence and the outcome of infection, as well as other epidemiological features, such as transmission.

Although Nematodirus battus (Nematoda: Trichostrongyloidea) is an economically important and highly pathogenic parasite of sheep in the temperate regions, very little is known about the population dynamics of its free-living stages and their relationship with ambient temperature. Here we describe the temperature-related vital rates and thresholds of egg development, hatching and larval survival for the first time. N. battus eggs were able to develop between 11·5 and 27°C, but development at the lower end of this range was more successful. Embryonated eggs did not hatch below 11°C or above 17°C. This is the first description of an upper threshold for hatching in trichostrongyloids. In contrast with most previous studies, although some eggs hatched only after being chilled, substantial proportions of eggs were also found to hatch without the need for chilling. These proportions were lower with increasing, within-hatching range, temperatures. Larval death rates significantly increased at temperatures towards and above the upper hatching threshold. The peculiar hatching behaviour of N. battus may therefore be explained in terms of optimization of larval survival. We argue that our findings confirm the likelihood of an arctic origin of the parasite. Probable changes and adaptations of parasite behaviour in the temperate regions, and the driving forces behind them, are discussed, as well as factors affecting persistence and geographical spread against a background of climate change.

1. It has been shown that there is a difference between the resistance of male and female mice to infection with Nematospiroides dubius.

2. More parasites were harboured, during both the larval and adult parasitic phases, by male mice.

3. These worms were found to occupy a similar relative length of the intestine between the stomach and the caecum in male and female mice infected for either 5 or 10 days.

4. The relative length of the intestine infected on the fifth day was significantly greater than that infected on the tenth day.

This investigation was carried out during the tenure of a Research Studentship from the Department of Scientific and Industrial Research. I should like to thank Professor I. Chester Jones, in whose department the work was done, for the facilities provided and Dr E. T. B. Francis for his helpful and critical supervision.

Trypanosoma cruzi is easily cultivated in tissue cultures of chick embryos, but it was not possible to infect chicks with trypanosomes cultivated for more than 2 years in chicken cells.

The trypanosome cycle takes place in one cell unit.

The number of trypanosomes formed in one cycle depends on the volume of the host cell.

The transformation of the leishmania form into the flagellate form begins when the cell is full of parasites, and occurs in 12–24 hr.

The intracellular trypanosome, immediately after the crithidia form, is broad and short, the thin form probably being a later phase of the evolution of the parasite.

Division of the adult trypanosome, or mating between the thin and broad forms, was never seen.

At the beginning of the infection the cells with-stand the parasites and continue to proliferate, but finally they are destroyed.

Isolation of blood and intracellular forms of Trypanosoma cruzi was made mainly from rats (90–110 g) which had received 580 rad of whole-body γ-irradiation not more than 24 h before subcutaneous inoculation with 107 trypomastigotes of the Sonya strain of T. cruzi. Unirradiated chinchillas (250–350 g) were, however, used for some experiments. Blood forms were isolated using a technique involving differential centrifugation to remove most of the erythrocytes and DEAE–cellulose chromatography to remove the remaining blood cells. Overall recoveries were usually in the range 30–70%. Parasites were mainly (approximately 98%) broad forms and were motile, metabolically active (as judged by respiratory and radio-tracer incorporation studies) and had lost none of their infectivity for mice. Intracellular forms were isolated from hind-limb muscle tissue. This was disrupted in an MSE tissue homogenizer and the homogenate incubated with DNase, collagenase and trypsin. Parasites, contaminated only by a few blood cells, were then obtained by differential centrifugation. For purer preparations, a terminal sucrose gradient step was used. Recoveries ranged between 40 and 70%. About 1–3% of the parasites isolated were epimastigotes and trypomastigotes; the remainder are probably best collectively termed ‘amastigotes’, though they were pointed and most had a short, free flagellum. They were undamaged as judged by light and electron microscopy and metabolically active as judged by respiratory and radio-tracer incorporation studies. However, the infectivity for mice of both these purified preparations and the initial cell homogenates could be accounted for by the epimastigotes and trypomastigotes present in them. Preliminary biochemical studies with isolated parasites have shown that blood, intracellular and culture forms of T. cruzi have a respiratory system which is in part sensitive to CN- and that all forms synthesize nucleic acids and proteins when incubated in vitro. There appears, however, to be a lack of DNA synthesis in blood stages, and thus it is not surprising that these forms do not divide.

Genetic diversity of Trypanosoma cruzi may play a role in pathogenesis of Chagas disease forms. Natural populations are classified into 6 Discrete Typing Units (DTUs) Tc I-VI with taxonomical status. This study aimed to identify T. cruzi DTUs in bloodstream and tissue samples of Argentinean patients with Chagas disease. PCR-based strategies allowed DTU identification in 256 clinical samples from 239 Argentinean patients. Tc V prevailed in blood from both asymptomatic and symptomatic cases and Tc I was more frequent in bloodstream, cardiac tissues and chagoma samples from immunosuppressed patients. Tc II and VI were identified in a minority of cases, while Tc III and Tc IV were not detected in the studied population. Interestingly, Tc I and Tc II/VI sequences were amplified from the same skin biopsy slice from a kidney transplant patient suffering Chagas disease reactivation. Further data also revealed the occurrence of mixed DTU populations in the human chronic infection. In conclusion, our findings provide evidence of the complexity of the dynamics of T. cruzi diversity in the natural history of human Chagas disease and allege the pathogenic role of DTUs I, II, V and VI in the studied population.

West-central China is an important endemic focus of both alveolar and cystic echinococcosis where several species of intermediate host are commonly infected with Echinococcus granulosus and E. multilocularis. Isolates of E. granulosus were collected from humans and other animals from different geographical areas of Qinghai, Ningxia, Gansu and Sichuan, and genotyped using the mitochondrial DNA marker ATP synthase subunit 6 gene (atp6). The sheep strain (G1 genotype) of E. granulosus was shown to be the only genotype present in sheep, cattle, goats, yaks and humans in the study areas. However, some heterogeneity in the atp6 sequence was evident in a number of the isolates with the most frequent change being a silent substitution (G/A) at position 360 compared with the G1 reference sequence representing isolates collected from the majority of hosts except humans. Two E. multilocularis isolates examined also had sequences that varied from each other and from the reference E. multilocularis atp6 sequence. The genotypic variation we report may reflect phenotypic differences with important consequences in terms of increased host infectivity for hosts by local Echinococcus strains, possibly impacting on the epidemiology and control of echinococcosis. Such adaptations may also result in different sensitivity to drugs or increased virulence for hosts that will impede control efforts and even affect vaccination strategies against Echinococcus.

Haemonchus contortus worm populations isolated from naturally infected sheep at the Pastoral Research Laboratory, Armidale, N.S.W., were found to contain approximately 20% of worms resistant to a 50 mg/kg dose of thiabendazole. Following 3 generations of selection with 50 mg/kg thiabendazole the number of worms removed by the anthelmintic was too small to detect differences between treated and control groups. After more than 15 generations of selection, matings between males from the selected strain and non-resistant females produced resistant males and females in equal numbers. Thus, thiabendazole resistance does not appear to be sex-linked. A dose–response assay on the F2 adults indicated that worms from female resistant × male non-resistant crosses were more resistant than F2 adults of the reciprocal cross. An in vitro technique that identified thiabendazole-resistant eggs by their ability to hatch in a solution containing thiabendazole and 0·1% NaCl solution was also used to study the inheritance of resistance. F1 eggs had similar LC50's to the resistant parents. F2 and back-cross eggs from an original mating of thiabendazole-resistant females × non-resistant males had a higher LC50 than F2 and back-cross eggs from the reciprocal mating, indicating a degree of matroclinous inheritance of resistance. However, the resistant parents had tolerances to thiabendazole exceeding those of F2. F3 eggs had a resistance distribution that ranged from that of the resistant to the non-resistant parent. No significant deviation from linearity was observed in any of the dose–response lines. These results indicate that thiabendazole resistance in H. contortus worms is inherited as an autosomal and semi-dominant trait.

Eavesdrop morning coffee at any major centre of evolutionary theory today, and you will find ‘parasite’ to be one of the commonest words in the language. Parasites are touted as prime movers in the evolution of sex, promising the final solution to that problem of problems, the puzzle that led G. C. Williams to proclaim in 1975 ‘a kind of crisis’ at hand in evolutionary biology (Hamilton, 1980; Tooby, 1982; Seger & Hamilton, 1988). Parasites seem to offer a plausible justification for the otherwise futile effort females put into choosing among posturing males (Hamilton & Zuk, 1982; but see Read, 1990). Frequency-dependent selection exerted by parasites is, according to one admittedly minority view, largely responsible for the high levels of diversity found in gene pools (Clarke, 1979). One might even extrapolate to a time when the entire metazoan body could come to be seen as a gigantic adaptation against microscopic pathogens.

Two morphologically novel Kudoa species are characterized from brain tissue of fish, Kudoa chaetodoni n. sp. from Chaetodon unimaculatus (Chaetodontidae) and Kudoa lethrini n. sp. from Gymnocranius audleyi and Lethrinus harak (Lethrinidae). Additionally we characterized a 5-spore valve (SV) Kudoa species from the brain of Sillago ciliata (Sillaginidae). Intriguingly, its 18S rDNA sequence was identical to that of the 7 SV Kudoa yasunagai extracted from the brain of a paralichthyid halibut in Japan. These 2 species may either prove to be con-specific, even though morphology and distribution differ, or demonstrate the limit of specific resolution in the small subunit rDNA gene region. Small subunit rDNA sequences from these new species were used in molecular phylogenetic analyses of kudoids to examine congruence of phylogeny with tissue tropism, geographical distribution, and host specificity. There was significant correlation between tissue tropism in the form of well-supported brain and heart-infecting clades. Host specificity and geographical distribution showed some correlations with genotype.

The nematode Angiostrongylus vasorum is becoming more widely recorded globally, and is of increasing concern as a cause of disease in dogs. Apparent geographic spread is difficult to confirm due to a lack of standardized disease recording systems, increasing awareness among veterinary clinicians, and recent improvements in diagnostic technologies. This study examines the hypothesis that A. vasorum has spread in recent years by repeating the methods of a previous survey of the fox population. The hearts and lungs of 442 foxes from across Great Britain were collected and examined by dissection and flushing of the pulmonary circulation and microscopic inspection of tracheal scrapes. Sampling and parasite extraction methods were identical to an earlier survey in 2005 to ensure comparability. Prevalence of A. vasorum was 18·3% (exact binomial confidence bounds 14·9–22·3), compared with 7·3% previously (5·3–9·9, n = 546), and had increased significantly in most regions, e.g. 7·4% in the Northern UK (previously zero) and 50·8% in the south-east (previously 23·2%). Other nematodes identified were Crenosoma vulpis (prevalence 10·8%, CI 8·1–14·2) and Eucoleus aerophilus (31·6%, CI 27·3–36·2). These data support the proposal that A. vasorum has increased in prevalence and has spread geographically in Great Britain.

The epidemiological role of the crab-eating fox Cerdocyon thous in the transmission of Leishmania infantum is assessed in a longitudinal study in Amazon Brazil. A total of 37 wild-caught foxes were immunologically and clinically monitored, and 26 foxes exposed to laboratory colonies of the sandfly vector Lutzomyia longipalpis, over a 15-month period. In total 78% (29/37) of foxes were seropositive for anti-Leishmania IgG on at least 1 occasion, and 38% (8/37) had infections confirmed by PCR and/or by culture. Point prevalences were 74% (serology), 15% (PCR), and 26% (culture). No signs of progressive disease were observed. None of the foxes were infectious to the 1469 sandflies dissected from 44 feeds. A conservative estimate of the possible contribution of foxes to transmission was 9% compared to 91% by sympatric domestic dogs. These results show that crab-eating fox populations do not maintain a transmission cycle independently of domestic dogs. The implication is that they are unlikely to introduce the parasite into Leishmania-free dog populations.

A Polymerase Chain Reaction (PCR) and Southern blot hybridization for the detection of Theileria annulata are described. The PCR used primers amplifying a 785 base-pair fragment of the T. annulata gene which encodes the 30 kDa major merozoite surface antigen, Tams1. The sensitivity of the PCR in bovine blood was 1 piroplasm in 1 μl of blood. T. buffeli, T. parva, Babesia bigemina, B. bovis and B. divergens were not detected. The PCR detected down to 1 infected acinus/tick in resting and partially fed adult Hyalomma anatolicum anatolicum ticks and was negative for T. lestoquardi and T. equi, which are transmitted by this tick but are not infective to cattle. The specificity of the PCR was checked using 30 stocks of T. annulata, all of which were detected. Three stocks of T. lestoquardi, 4 of T. equi and 1 each of T. buffeli, T. parva, B. bigemina, B. bovis and B. divergens were used to ascertain there were no cross-reactions. A nested PCR using separate primers for the first reaction and the same primers for the second reaction detected T. annulata to the same sensitivity and specificity in saponin-extracted DNA samples stored for long periods at −20 °C.

Epidemiological factors of tick-borne equine piroplasmoses, caused by Theileria equi and Babesia caballi, were investigated using logistic regression (GLM) and general additive models (GAM) based on the prevalences determined in 510 domestic horses and in ticks in S.W. Mongolia by indirect immunofluorescence antibody test (IFAT) and/or multiplex PCR. Prevalences of T. equi and B. caballi in horses were 66·5% (95% CI: 62·1–70·7) and 19·1% (15·6–22·9), respectively by PCR and 78·8% (74·9–82·3) and 65·7% (61·3–69·9) by IFAT. Of 166 ticks analysed from PCR- and IFAT-negative horses 1 was PCR positive for B. caballi and none for T. equi. GAM demonstrated non-linear increasing proportions of T. equi-PCR and -IFAT positive horses with age suggesting persistent infection. In contrast, the B. caballi-PCR prevalence decreased with age despite a concurrent increase in the proportion of IFAT-positive animals suggesting parasite elimination. The tick (Dermacentor nuttalli) burden of the horses increased with age and decreased with advancing season. Geldings were more likely to be infected with, and seroconvert to, T. equi. Neither herd affiliation, date of sample collection nor abundance of tick infestation had a significant influence on parasite prevalence.

Anopheles stephensi mosquitoes showed a reduction in fecundity over 3 successive gonotrophic cycles, after becoming infected with Plasmodium yoelii nigeriensis. This effect could be observed at high oocyst burdens (> 75) or at low oocyst burdens (mean of 4·36). Mean bloodmeal size of the infected mosquitoes was significantly reduced only when feeding upon a mouse with a high gametocytaemia and the conversion of the bloodmeal into eggs by the infected mosquitoes was disrupted. Patterns of infected mosquito mortality, over the 3 gonotrophic cycles, varied with severity of infection. Although in 1 case increased mortality and decreased bloodmeal size may have affected fecundity, this could not have accounted for all of the observed fecundity reduction. We propose that other, unknown parasite related factors, are involved.

Three groups of stocks of Trypanosoma brucei ssp, defined by the criteria of host, human serum resistance and place of isolation as T. b. gambiense, T. b. brucei (Nigeria) and T. b. brucei/rhodesiense (non-gambiense, Uganda) were screened for electrophoretic variation at 20 enzyme loci. One enzyme (Peptidase C) was found to differentiate all T. b. gambiense stocks from the other T. brucei stocks and, taken together with specific variants of 5 other enzymes, could be used to unambiguously define T. b. gambiense stocks. Using a population genetics approach, the frequencies of the different variants in the three groups of stocks were estimated and from them the average similarity and difference between groups were measured using the statistics of genetic identity (I) and genetic distance (D). These results show firstly, that T. b. gambiense is more different from the other two groups than they are from each other and, secondly, that the values of I and D obtained are consistent with T. b. gambiense constituting a sub- or sibling species of T. brucei. Three domestic animal isolates from Zaire and Cameroun were also screened for enzyme variation and two of these identified as T. b. gambiense, thereby establishing the existence of an animal reservoir host. In parallel, these stocks were tested for human serum resistance resulting in the same identification. Studies of the antigen repertoires and antigen gene structure were carried out by other workers on all the T. b. gambiense stocks reported here and the same conclusions reached as to the identification and ability to discriminate this subspecies from other groups of T. brucei stocks. The results presented here are discussed in relation to other published data on enzyme variation in T. brucei.