Two groups of goldfish maintained at 20°C were experimentally infected with two different sized inocula of Trypanosoma danilewskyi. One group was inoculated with 380000 trypanosomes/fish and the other with 3800. Forty and 90% of the fish, respectively, survived. All surviving fish challenged 190 days post-infection were protected from re-infection. The plasma of these fish when inoculated into another group of fish also protected them from infection. Finally, trypanosomes incubated in vitro in immune plasma at 20°C for 2 h were not infective; however, incubations at 5°C or for 1 h were not as effective.

An evaluation of the genetic diversity within Fasciola hepatica (liver fluke) may provide an insight into its potential to respond to environmental changes, such as anthelmintic use or climate change. In this study, we determined the mitochondrial DNA haplotypes of >400 flukes from 29 individual cattle, from 2 farms in the Netherlands, as an exemplar of fasciolosis in a European context. Analysis of this dataset has provided us with a measure of the genetic variation within infrapopulations (individual hosts) and the diversity between infrapopulations within a herd of cattle. Temporal sampling from one farm allowed for the measurement of the stability of genetic variation at a single location, whilst the comparison between the two farms provided information on the variation in relation to distance and previous anthelmintic regimes. We showed that the liver fluke population in this region is predominantly linked to 2 distinct clades. Individual infrapopulations contain a leptokurtic distribution of genetically diverse flukes. The haplotypes present on a farm have been shown to change significantly over a relatively short time-period.

Leishmania (Viannia) braziliensis is a protozoan that causes mucocutaneous leishmaniasis, which is an infectious disease that affects more than 12 million people worldwide. The available treatment is limited, has side-effects or is inefficient. In a search for alternative compounds of natural origin, we tested the microbicidal activity of Piper aduncum essential oil (PaEO) on this parasite. Our data showed that PaEO had an inhibitory effect on the growth of L. braziliensis promastigotes with an IC50/24 h=77·9 µg mL−1. The main constituent (nerolidol: 25·22%) presented a similar inhibitory effect (IC50/24 h = 74·3 µg mL−1). Ultrastructural observation of nerolidol-treated parasites by scanning and transmission electron microscopies revealed cell shrinkage and morphological alterations in the mitochondrion, nuclear chromatin and flagellar pocket. Flow cytometry analysis showed a reduction in the cell size, loss of mitochondrial membrane potential, phosphatidylserine exposure and DNA degradation, which when associated with the morphological changes indicated that nerolidol induced incidental cell death in the L. braziliensis promastigotes. The results presented here indicate that nerolidol derivatives are promising compounds for further evaluation against Leishmania parasites.

An entire 18S rRNA gene sequence from Schistosoma spindale (1990 bases) and partial 18S rRNA gene sequences from S. haematobium (1950 bases) and S. japonicum (1648 bases) have been determined. Together with the previously published sequence for the S. mansoni 18S rRNA gene, these data encompass the 4 recognized Schistosoma species groups. Although Schistosoma 18S rRNA genes are highly conserved, the sequences permit a preliminary molecular phylogeny to be established for the genus. This identifies S. haematobium and S. spindale as sister taxa in a clade with S. mansoni. S. japonicum does not appear to be closely related to this clade. Much of the observed variation occurs within a ‘hypervariable’ stretch of the gene corresponding to part of the V4 region of 18S rRNA. Despite this variation, the 3 new sequences fit models of 18S rRNA secondary structure predicted from the S. mansoni sequence.

The sequences reported in this paper have been submitted to the EMBL Database under accession numbers Z11976 (S. haematobium), Z11590 (S. japonicum) and Z11979 (S. spindale).

Immunochemical techniques were used to determine the distribution, chemical characteristics and relative abundance of immunoreactivity (IR) to two native platyhelminth neuropeptides, neuropeptide F (NPF) (Moniezia expansa) and the FMRFamide-related peptide (FaRP), GNFFRFamide, in the trematodes, Fasciola hepatica and Schistosoma mansoni; the larger S. margrebowiei was used in the chemical analysis. Extensive immunostaining for the two peptides was demonstrated throughout the nervous systems of both F. hepatica and S. mansoni, with strong IR also in the innervation of muscular structures, including those associated with the egg-forming apparatus. The patterns of immunostaining were similar to those previously described for the vertebrate neuropeptide Y superfamily of peptides and for FMRFamide. Ultra-structurally, gold labelling of NPF- and GNFFRFamide-IRs was localized exclusively to the contents of secretory vesicles in the axons and somatic cytoplasm of neurones. Double-labelling experiments showed an apparent homogeneity of antigenic sites, in all probability due to the demonstrated cross-reactivity of the FaRP antiserum with NPF. Radio-immunoassay of acid-ethanol extracts of the worms detected 8·3 pmol/g and 4·7 pmol/g equivalents of NPF- and FMRFamide-IRs, respectively, for F. hepatica, and corresponding values of 4·9 pmol/g and 4·3 pmol/g equivalents for S. margrebowiei. Gel-permeation chromatography resolved IR to both peptides in discrete peaks and these eluted in similar positions to synthetic NPF (M. expansa) and GNFFRFamide, respectively.

Studies of foraging strategies are often complicated by competing goals of the forager. In contrast, non-feeding infective juvenile entomopathogenic nematodes forage exclusively for a single host. Two questions were posed: (1) what is the relationship between metabolic rate, energy reserves and foraging strategy and (2) when a foraging strategy fails, will an infective-stage parasite switch strategies? Three species of entomopathogenic nematodes were stored in water and changes in their behaviour, metabolic rate, energy reserves, and infectivity were measured throughout the storage period. Steinernema carpocapsae ambushes insect hosts, whereas S. glaseri and Heterorhabditis bacteriophora cruise forage. Steinernema carpocapsae was least active and had the lowest metabolic rate. Heterorhabditis bacteriophora was more active and had the highest metabolic rate. Steinernema glaseri was most active and had an intermediate metabolic rate. Neither cruising species changed foraging strategy. Steinernema carpocapsae decreased nictation (a behaviour associated with ambushing only) and increased their locomotory rate. Any change in searching strategy occurred without assessment of the profitability or distribution of potential hosts, but the advantage this confers is unknown.

Climate change in the Arctic is anticipated to alter the ecology of northern ecosystems, including the transmission dynamics of many parasite species. One parasite of concern is Ostertagia gruehneri, an abomasal nematode of Rangifer ssp. that causes reduced food intake, weight loss, and decreased pregnancy rates in reindeer. We investigated the development, availability, and overwinter survival of the free-living stages of O. gruehneri on the tundra. Fecal plots containing O. gruehneri eggs were established in the Northwest Territories, Canada under natural and artificially warmed conditions and sampled throughout the growing season of 2008 and the spring of 2009. Infective L3 were present 3–4 weeks post-establishment from all trials under both treatments, except for the trial established 4 July 2008 under warmed conditions wherein the first L3 was recovered 7 weeks post-establishment. These plots were exposed to significantly more time above 30°C than the natural plots established on the same date, suggesting a maximum temperature threshold for development. There was high overwinter survival of L2 and L3 across treatments and overwintering L2 appeared to develop to L3 the following spring. The impact of climate change on O. gruehneri is expected to be dynamic throughout the year with extreme maximum temperatures negatively impacting development rates.

In a manner similar to many other groups of organisms, the tapeworm order Proteocephalidea poses a difficult phylogenetic problem if treated on the basis of single-gene analysis. Since the biogeography and host distribution of proteocephalideans make these tapeworms a potentially interesting model for evolutionary and co-evolutionary studies, we tried to resolve their phylogenetic relationships by applying a multi-gene approach. The ITS2 sequences and V4 hypervariable loop of 18S rRNA were obtained for 43 and 35 proteocephalidean taxa, respectively, and combined with other sequences available in the GenBank. The phylogenetic analysis of the combined DNA set was confronted with characters derived from ITS2 secondary structures. Using this approach, a species-rich Neotropical lineage of proteocephalideans could be reliably resolved. The phylogenetic relationships within this group show a high degree of phylogeny-independent host distribution. The reconstruction of ITS2 secondary structure revealed a universal 4-domain arrangement, which is conserved across a wide range of Neodermata. Several motifs of the secondary structure could be mapped to the phylogenetic tree as possible clade synapomorphies.

We have performed restriction endonuclease digestion of nuclear DNAs, combined with gel electrophoresis and molecular hybridization to characterize different Trypanosoma brucei brucei stocks and to identify trypanosome species. Cloned DNA complementary to the messenger RNAs (mRNAs) for different variable surface glycoproteins (VSGs) of T. b. brucei stock LUMP 227 was used to examine differences in genomic DNAs from T. b. brucei stocks and from various other trypanosome species. These cDNA reagents differentiated the stocks of T. b. brucei and also distinguished the trypanosome species. Our data also show that a T. b. brucei nuclear DNA fragment used as a probe in molecular hybridization analysis provides an appropriate marker in determining trypanosome species. We have extended our analysis of trypanosome nuclear DNA by developing a simple and rapid spot test, for the identification of trypanosome species, which can be carried out using crude materials such as isolated parasites, infected whole blood and tsetse saliva. The procedure is species-specific, sensitive to the level of 104 organisms and is suitable for use under field conditions. Thus, with appropriate sequences for hybridization, the procedure has obvious applications for the diagnosis of African trypanosomiasis.

The nematodes described here were collected by biologists of the Australian National Antarctic Research Expedition based at the subantarctic stations at Macquarie Island and Heard Island, over the years 1948–51.

Up to the present very few authors have directed their attention to factors influencing the emergence of larval trematodes from their snail hosts. Cort (1922) has made a study of the escape of cercariae from their hosts and the writer (1931) has examined the effect of light and temperature on the emergence of three species of larval trematodes from three species of fresh-water molluscs. More recently, Giovannola (1936) has investigated the effect of a reversal of light and darkness on the periodicity of emission of cercariae from three species of host snails. All these studies have been made on freshwater molluscs. In the case of molluscs living between tide-marks on the shore, in addition to variation in temperature there exists another variable factor, namely, the salinity of the water. Many rock-pools situated in the intertidal zone are small and will therefore be subject to considerable variations in temperature and salinity at different times of the year and even for short periods of up to 12 hr. during individual days. In the summer months the temperature of the water in these pools may rise to a high degree and the high temperature would be maintained for a longer period near high-water mark than near low-water mark, where the period of exposure is less. Similarly, the decrease in temperature would be very marked during the cold winter months. As the water in the exposed pools evaporates the salinity increases and high up on the shore, between successive tides on hot summer days, it may become twice that of normal sea water. The pools are also exposed to rain and in the same period the salinity of the water may decrease considerably if the precipitation is heavy. The light factor would be the same for molluscs living in fresh and in salt water, but the reaction of various species of larval trematodes to the presence or absence of light varies. Some emerge from their hosts in larger numbers during the daytime, others during the night and some indiscriminately during periods of light and darkness.

Density-dependent constraints on parasite growth, survival or reproduction are thought to be important in preventing the unchecked increase in parasite numbers within individual hosts or host populations. While it is important to know where, and with what severity, density dependence is acting within the parasite life-cycle, interpretation of data from natural infections is difficult. In this paper, we present a Monte Carlo simulation technique for examining such data for evidence of density dependence. We also describe how this technique may be used to distinguish among mechanisms hypothesized to generate density-dependent phenomena.

Genetic proximity between two karyotypic groups of Trypanosoma congolense was investigated using as hybridization probes: (i) total genomic DNA, (ii) a 35 nucleotide long synthetic oligonucleotide, and (iii) non-variant antigen type (non-VAT) specific complementary DNAs. The phylogenetic relationship between Trypanosoma brucei and T. evansi, both of which are accepted species in the subgenus Trypanozoon, was used as a reference to assess the phylogenetic proximity of the two groups of T. congolense. Results indicate that some morphologically indistinguishable T. congolense populations differ in a variety of molecular and genetic properties: molecular karyotypes, majority of the DNA sequences, and the restriction enzyme sites in the genomic environments of various conserved genes. The implications of these findings for trypanosome evolution and T. congolense epidemiology are discussed.

Two types of tetraphyllidean merocercoids, Phyllobothrium delphini and Monorygma grimaldii, are well known from most cetaceans world-wide. The role of cetaceans in the life-cycle of these merocercoids is unclear because their specific identity is as yet unknown. The problem is compounded by poor descriptions of both merocercoids. We used light and scanning electron microscopy, and histological techniques to provide a thorough description of merocercoids collected from 11 striped dolphins, Stenella coeruleoalba, from the Spanish Mediterranean. We also described, for the first time, specimens of P. delphini with immature proglottides. Our merocercoids were morphologically similar to those described previously, except in the structure of the apical organ. Intra- and inter-sample variability in the morphology of the apical organ suggested that it degenerates during larval development. A subsample of 16 specimens of P. delphini and M. grimaldii was characterized for the D2 variable region of the large subunit ribosomal RNA gene (LSU) and compared with published tetraphyllidean cestode LSU sequences. P. delphini showed 2 unique signatures that differed from one another by a single base, whereas all sequences of M. grimaldii were identical. This suggests that each type may represent a single species, contrary to previous speculations based on morphological data. All merocercoid specimens formed a clade together with Clistobothrium montaukensis. Based on the low degree of divergence, all specimens of this clade are predicted to be congeneric.

In this study, we addressed the phylogenetic and taxonomic relationships of Trypanosoma vivax and related trypanosomes nested in the subgenus Duttonella through combined morphological and phylogeographical analyses. We previously demonstrated that the clade T. vivax harbours a homogeneous clade comprising West African/South American isolates and the heterogeneous East African isolates. Herein we characterized a trypanosome isolated from a nyala antelope (Tragelaphus angasi) wild-caught in Mozambique (East Africa) and diagnosed as T. vivax-like based on biological, morphological and molecular data. Phylogenetic relationships, phylogeographical patterns and estimates of genetic divergence were based on SSU and ITS rDNA sequences of T. vivax from Brazil and Venezuela (South America), Nigeria (West Africa), and from T. vivax-like trypanosomes from Mozambique, Kenya and Tanzania (East Africa). Despite being well-supported within the T. vivax clade, the nyala trypanosome was highly divergent from all other T. vivax and T. vivax-like trypanosomes, even those from East Africa. Considering its host origin, morphological features, behaviour in experimentally infected goats, phylogenetic placement, and genetic divergence this isolate represents a new genotype of trypanosome closely phylogenetically related to T. vivax. This study corroborated the high complexity and the existence of distinct genotypes yet undescribed within the subgenus Duttonella.

Two nucleoproteins with similar chemical composition have been isolated from the leaves of tobacco plants infected with tobacco necrosis virus. One of these is crystalline and has a sedimentation constant of 130 × 10−13; the other is amorphous and its principal component has a sedimentation constant of 58 × lO−13.

Each preparation will infect plants at a dilution of 1 in 108 and will precipitate specifically with antiserum at a dilution of 1 in 3·2 × 105.

The nature of the difference between preparations in the two states is obscure and it has not proved possible to convert the one into the other.

Peritoneal hydatids of Echinococcus granulosus equinus of 9 months standing in BALB/c mice occurred as free cysts or cysts within cyst masses. Both showed wide variation in size and in host tissue response, and all had a well-developed laminated layer separating the host tissue response from the germinal layer. In the smallest cyst-mass cysts the host tissue response was present as remnants of the initial cellular attack involving eosinophils. Slightly larger cyst-mass cysts possessed a primary macrophage invasion which phagocytosed the remnants of the initial attack and also, though to little effect, the laminated layer material. In the largest cyst-mass cysts a second macrophage invasion, of monocyte origin, had commenced and transformation stages of these cells to macrophages were observed. No fibroblasts surrounded individual cyst-mass cysts but they were present around the cyst mass, encapsulating it and possibly preventing further host cell invasion. Thus, the host tissue response around individual cyst-mass cysts remained ‘preserved’ at an early stage such as existed at the time of encapsulation. Small free cysts showed a primary macrophage invasion and transformation stages of cells of a secondary infiltration of peritoneal origin. Peripheral to the macrophages were fibroblasts demonstrating limited fibrinogenesis, and each cyst was surrounded by a layer of mesothelial cells. Large free cysts, also delimited by a mesothelial layer, possessed peripheral connective tissue, a deep fibrous layer and a monolayer of very compressed macrophages lying adjacent to the laminated layer. It is emphasized that an understanding of the host tissue response in cysts of different sizes and from different locations is an essential pre-requisite for the design of experimental studies.

The course of single infections of Nematodirus battus was studied and the developmental phases of the fourth larval stage described. The third moult occurred between days 2 and 4 after infection and the fifth between days 8 and 10. The period of maximum larval penetration of the intestinal mucosa occurred around day 4 and most of the larvae had returned to the mucosal surface by day 6, during the middle of the fourth-stage. Some larvae remained deep in the mucosa and moulted to the fifth-stage in this site.

The relationship between growth and morphological development was not constant and was different in worm populations from different sites. Some larvae grew very rapidly during the early fourth-stage and then entered a period of lethargus while continuing to develop sexually, while others grew and developed throughout the larval stage.

The worms recovered from the second 30% of the small intestine were consistently smaller and less advanced morphologically than those from the first 30%. The worms in the posterior region of the small intestine were preferentially lost from the mucosa in the third- and fourth-stages between days 2 and 4 and from the mucosal surface as adults between days 12 and 16.

A microsporidian infection was discovered in laboratory cultures of Drosophila species. Ultrastructural examination suggested it belonged to the poorly characterized species Tubulinosema kingi, and morphological and sequence data are presented. We explored how T. kingi affected the fitness of Drosophila melanogaster and D. subobscura, as well as the fitness of 2 of their parasitoids, Asobara tabida and Pachycrepoideus vindemiae. In Drosophila, infections caused changes in most of the traits we looked at that were associated with fitness, in particular causing a 34–55% reduction in early-life fecundity. Parasitoid fitness was affected more severely by infection than that of their hosts, with pupal mortality in particular increasing by 75–89%. We investigated the most important routes of transmission for T. kingi in a laboratory setting. Letting Drosophila larvae feed on medium contaminated with spores from infected dead flies resulted in 100% infection. Low levels of transmission (<10%) were found between larvae, and vertically between mothers and their offspring. Parasitoids developing in infected hosts all became infected, but infected adults were neither able to transmit the pathogen to their offspring nor to their offspring's Drosophila host, either directly, or via contamination of the ovipositor or other body parts. A field survey of Drosophila and their parasitoids in southern England revealed no natural infections. We discuss the potential importance of Microsporidia in parasitoid-host interactions, and for those working with Drosophila in the laboratory.

Immune responses that kill pathogens or reduce their reproductive rate are generally important in protecting hosts from infection and disease. Pathogens that escape the full impact of such responses will survive, and any heritable genetic basis of this evasion will be selected. Due to the memory component of vertebrate immune responses, pathogens with rare alleles of a target antigen can have an advantage over those with common alleles, leading to the maintenance of a polymorphism. At the genetic level, there ought to be detectable signatures of balancing selection in the genes encoding these antigens. Here, methods for identifying these selective signatures are reviewed. Their practical utility for identifying which antigens are targets of protective immune responses is discussed.