RNA interference (RNAi) has emerged as an invaluable gene-silencing tool for functional analysis in a wide variety of organisms, particularly the free-living model nematode Caenorhabditis elegans. An increasing number of studies have now described its application to plant parasitic nematodes. Genes expressed in a range of cell types are silenced when nematodes take up double stranded RNA (dsRNA) or short interfering RNAs (siRNAs) that elicit a systemic RNAi response. Despite many successful reports, there is still poor understanding of the range of factors that influence optimal gene silencing. Recent in vitro studies have highlighted significant variations in the RNAi phenotype that can occur with different dsRNA concentrations, construct size and duration of soaking. Discrepancies in methodology thwart efforts to reliably compare the efficacy of RNAi between different nematodes or target tissues. Nevertheless, RNAi has become an established experimental tool for plant parasitic nematodes and also offers the prospect of being developed into a novel control strategy when delivered from transgenic plants.

The hard tick Ixodes ricinus (Ixodidae) is the sole animal thus far shown to harbour an intra-mitochondrial bacterium, which has recently been named Midichloria mitochondrii. The objectives of this work were (i) to screen ixodid ticks for Midichloria-related bacteria and (ii) to determine whether these bacteria exploit the intra-mitochondrial niche in other tick species. Our main goal was to discover further models of this peculiar form of symbiosis. We have thus performed a PCR screening for Midichloria-related bacteria in samples of ixodid ticks collected in Italy, North America and Iceland. A total of 7 newly examined species from 5 genera were found positive for bacteria closely related to M. mitochondrii. Samples of the tick species Rhipicephalus bursa, found positive in the PCR screening, were analysed with transmission electron microscopy, which revealed the presence of bacteria both in the cytoplasm and in the mitochondria of the oocytes. There is thus evidence that bacteria invade mitochondria in at least 2 tick species. Phylogenetic analysis on the bacterial 16S rRNA gene sequences generated from positive specimens revealed that the bacteria form a monophyletic group within the order Rickettsiales. The phylogeny of Midichloria symbionts and related bacteria does not appear completely congruent with the phylogeny of the hosts.

The effect of haemoparasites on the physiology and behaviour traits of their hosts was examined using Haemogregarina sp., a parasite of the common lizard, Lacerta vivipara, from the south of France. Infection with haemogregarines was associated with a reduced haemoglobin concentration and an increased number of immature red blood cells. Parasitized individuals also showed a reduced oxygen consumption at rest and a lower locomotor speed. We also found that the multiplication rate of the parasite depended on the temperature at which the lizard was maintained. Between 21 and 28 °C the multiplication rate of the parasite was significantly lower than between 29 and 35 °C. This suggests that the parasites may suffer reproductive costs when hosts reduce their body temperature.

Malnutrition and helminth infection are amongst the most prevalent chronic conditions affecting human health globally. It is estimated that parasitic helminths infect more than 1 billion people, and that more than 2 million clinical cases occur each year (Peters, 1978; Walsh, 1984). Estimates of the incidence of clinical malnutrition suggest that between 5 and 8 million cases occur annually. In many parts of the developing world malnutrition and infection conjointly are the most serious health problem in children, acting as primary or more often as secondary factors in mortality (Puffer & Serrano, 1973). The impact on health is exacerbated because both conditions are chronic, are most common in growing children and, most importantly, tend to occur together in the same individuals (Pawlowski, 1984; Chandra & Newberne, 1977).

Giardia that infect humans are known to be heterogeneous but they are assigned currently to a single species, Giardia intestinalis (syn. G. lamblia). The genetic differences that exist within G. intestinalis have not yet been assessed quantitatively and neither have they been compared in magnitude with those that exist between G. intestinalis and species that are morphologically similar (G. duodenalis) or morphologically distinct (e.g. G. muris). In this study, 60 Australian isolates of G. intestinalis were analysed electrophoretically at 27 enzyme loci and compared with G. muris and a feline isolate of G. duodenalis. Isolates of G. intestinalis were distinct genetically from both G. muris (approximately 80% fixed allelic differences) and the feline G. duodenalis isolate (approximately 75% fixed allelic differences). The G. intestinalis isolates were extremely heterogeneous but they fell into 2 major genetic assemblages, separated by fixed allelic differences at approximately 60% of loci examined. The magnitude of the genetic differences between the G. intestinalis assemblages approached the level that distinguished the G. duodenalis isolate from the morphologically distinct G. muris. This raises important questions about the evolutionary relationships of the assemblages with Homo sapiens, the possibility of ancient or contemporary transmission from animal hosts to humans and the biogeographical origins of the two clusters.

Most, if not all, organisms face attack by natural enemies and will be selected to evolve some form of defence. Resistance may have costs as well as its obvious benefits. These costs may be associated with actual defence or with the maintenance of the defensive machinery irrespective of whether a challenge occurs. In this paper, the evidence for costs of resistance in insect-parasite and insect-parasitoid systems is reviewed, with emphasis on two host-parasitoid systems, based on Drosophila melanogaster and pea aphids as hosts. Data from true insect-parasite systems mainly concern the costs of actual defence; evidence for the costs of standing defences is mostly circumstantial. In pea aphids, the costs of standing defences have so far proved elusive. Resistance amongst clones is not correlated with life-time fecundity, whether measured on good or poor quality plants. Successful defence by a D. melanogaster larva results in a reduction in adult size and fecundity and an increased susceptibility to pupal parasitoids. Costs of standing defences are a reduction in larval competitive ability though these costs only become important when food is limited. It is concluded that costs of resistance can play a pivotal role in the evolutionary and population dynamic interactions between hosts and their parasites.

Cystic echinococcosis, caused by the larval stage of Echinococcus granulosus, is a global public health problem. Whilst in a few localities, such as New Zealand, the parasite has been effectively controlled or even eradicated, in most endemic regions it remains a persistent problem. In some areas, such as the former Soviet Union, the disease incidence in humans has increased rapidly in recent years. It is important to have an understanding of the transmission dynamics, both between dogs and domestic livestock where the parasite maintains itself and from dogs to people. It is from this knowledge that effective control measures can be devised to reduce the prevalence of the parasite in animals and hence reduce the incidence of human disease. Mathematical models to describe the transmission of the parasite and the effects of different control strategies were first proposed over twenty years ago. Since then further information has been acquired, new technology has been developed and better computing technology has become available. In this review, we summarise these developments and put together a theoretical framework on the interpretation of surveillance information, how this affects transmission and how this information can be exploited to develop new intervention strategies for the control of the parasite. In particular, the parasite remains a persistent or re-emerging problem in countries of low economic output where resources for an intensive control programme, that has been successful in rich countries, are not available. By understanding of the transmission biology, including mathematical modelling, alternative and cost-effective means of control can be developed.

Parasitic protozoa comprise diverse aetiological agents responsible for important diseases in humans and animals including sleeping sickness, Chagas disease, leishmaniasis, malaria, toxoplasmosis and others. They are major causes of mortality and morbidity in tropical and subtropical countries, and are also responsible for important economic losses. However, up to now, for most of these parasitic diseases, effective vaccines are lacking and the approved chemotherapeutic compounds present high toxicity, increasing resistance, limited efficacy and require long periods of treatment. Many of these parasitic illnesses predominantly affect low-income populations of developing countries for which new pharmaceutical alternatives are urgently needed. Thus, very low research funding is available. Amidine-containing compounds such as pentamidine are DNA minor groove binders with a broad spectrum of activities against human and veterinary pathogens. Due to their promising microbicidal activity but their rather poor bioavailability and high toxicity, many analogues and derivatives, including pro-drugs, have been synthesized and screened in vitro and in vivo in order to improve their selectivity and pharmacological properties. This review summarizes the knowledge on amidines and analogues with respect to their synthesis, pharmacological profile, mechanistic and biological effects upon a range of intracellular protozoan parasites. The bulk of these data may contribute to the future design and structure optimization of new aromatic dicationic compounds as novel antiparasitic drug candidates.

The relationship between faecal examination for egg output and worm burden of Opisthorchis viverrini in man of 181 autopsy cases from Northeast Thailand is described. Diagnosis of the parasite infection by stool examination for the presence of eggs was less sensitive than the worm recovery technique. Using Stoll's dilution and formalin–ether technique, no eggs were detected in the faeces of 20 cases harbouring low worm burdens (< 20 worms). The quantitative faecal egg count by Stoll's dilution technique showed a strikingly close positive correlation with the number of worms recovered (r = 0·96, P < 0·001) indicating a strong linear association between eggs per gram of faeces (epg) and worm burden. The number of epg per worm was inversely correlated to the worm burden (P < 0·001), suggesting that density-dependent constraints on fecundity could operate to restrict egg output in heavy infections. The accuracy of egg counts for estimating worm burden and its relevance to parasite epidemiological research are discussed.

In the last 30 years, at various laboratories in the United States, France and England, many strains of small holotrichous ciliates have been established in axenic culture for use in a widening variety of investigations of a physiological or biochemical nature. Since few of these organisms have been adequately described or properly identified by the workers employing them, thus making difficult meaningful comparison of the various experimental results reported, the need for morphological and taxonomical study has become increasingly apparent. At least twenty-nine such axenic strains of ciliates, considered members of a ‘Colpidium-Glaucoma-Leucophrys-Tetrahymena group’ because it has been principally to these four genera that the protozoa have been assigned in the past, are still in existence and twenty-six of them were collected for use in the present investigation. For accurate identification of such relatively undifferentiated ciliates the use of silver nitrate impregnation techniques, the value of which was recognized some years ago by Klein (1926) and stressed by Furgason (1940), is re-emphasized as being nearly indispensable. Details of buccal organelles and differentiated pellicular structures in other parts of the body are revealed precisely and with constancy.

The tick-borne bacterium Rickettsia rickettsii is the aetiological agent of Brazilian spotted fever (BSF). The present study evaluated tick infestations on wild and domestic animals, and the rickettsial infection in these animals and their ticks in 7 forest areas adjacent to human communities in the São Paulo Metropolitan Area (SPMA). The results were compared to ecological traits of each sampled area. Two main tick species, Amblyomma aureolatum and Rhipicephalus sanguineus, were collected from dogs. The major ticks found on small mammals and birds were Ixodes loricatus and Amblyomma longirostre, respectively. Both anti-R. rickettsii antibodies and R. rickettsii-infected ticks were detected on dogs from only 2 areas in the southern part of the SPMA, which were considered to be endemic for BSF; the remaining 5 areas were considered to be non-endemic. Ecologically, the BSF-endemic areas clearly differed from the non-endemic areas by the presence of significantly more degraded forest patches in the former. The present results corroborate historical observations that have indicated that all human cases of BSF in the SPMA were contracted in the southern part of this metropolitan area. However, not all forest patches in the southern part of the SPMA were shown to be associated with BSF endemism.

Helminth infections have large negative impacts on production efficiency in ruminant farming systems worldwide, and their effective management is essential if livestock production is to increase to meet future human needs for dietary protein. The control of helminths relies heavily on routine use of chemotherapeutics, but this approach is unsustainable as resistance to anthelmintic drugs is widespread and increasing. At the same time, infection patterns are being altered by changes in climate, land-use and farming practices. Future farms will need to adopt more efficient, robust and sustainable control methods, integrating ongoing scientific advances. Here, we present a vision of helminth control in farmed ruminants by 2030, bringing to bear progress in: (1) diagnostic tools, (2) innovative control approaches based on vaccines and selective breeding, (3) anthelmintics, by sustainable use of existing products and potentially new compounds, and (4) rational integration of future control practices. In this review, we identify the technical advances that we believe will place new tools in the hands of animal health decision makers in 2030, to enhance their options for control and allow them to achieve a more integrated and sustainable approach to helminth control in support of animal welfare and production.

The present study describes a novel heat-stable, water-stress-related protein with a molecular mass of 47 kDa (designated Desc47) in the entomopathogenic nematode Steinernema feltiae (IS-6). The protein was accumulated about 10-fold (from 7·84±1·85 to 74·09±4·35% relative content level [RCL]) in dehydrated clumps of infective juveniles (IJs), which had lost 34·4% of their initial water content (from 65·1±1·7% to 42·7±0·72%) in a desiccation-tolerance-inducing treatment (97% relative humidity [RH] for 3 days). The appearance of Desc47 was accompanied by trehalose accumulation (from 300 to 600 mg trehalose/g protein) during the process of inducing the IJs into a quiescent anhydrobiotic state. A second cycle of IJ dehydration did not alter the RCL of Desc47 (79·3% for the first cycle and 73·3% for the second cycle). Desc47 retained its high RCL (69·7%) in rehydrated active IJs for 3 days, reaching 51·2% of its initial RCL only after a week. No homology to other known proteins was found by mass-spectrometry electrospray-ion-trap analysis. However, of the 5 sequences obtained from the protein (ranging from 11 to 21 amino acids), the 21-amino-acid peptide NVASDAVETVGNAAGQAG(D/T)AV showed excellent homology (74% identity in 19 amino acids) to the cold-responsive protein COR14b (g6564861) from Triticum aestivum. In the Caenorhabditis elegans predicted proteome database search, the N21 yielded the first-best identity score (59% identity in 17 amino acids) to the CE-LEA homologue protein (g2353333). In plants, COR and LEA are related proteins, heat-stable, which are expressed in response to both dehydration and cold acclimation. The implication of the involvement of Desc47 and the osmoprotectant trehalose in the desiccation-tolerance mechanisms of S. feltiae is discussed.

Mathematical models often propose that within-host competition between parasites can be a major factor in the evolution of increased parasite virulence. Kin selection predicts that as the coefficient of relatedness between infecting parasites decreases, the benefits of competition to individual genotypes increases. Thus where parasites can adjust their behaviour in response to current conditions, higher virulence is predicted in multiple genotype infections. There is limited experimental data, however, regarding the effects of mixed strain infections on host and parasite fitness. We investigated, for a snail–schistosome system, whether a conditional increase in replication rates occurred in mixed genotype infections and resulted in increased virulence. Four groups of Biomphalaria glabrata snails were exposed to 1 or 2 laboratory strains of Schistosoma mansoni. Mixed genotype infections were observed to be more virulent than single genotype infections, in terms of reductions in host reproductive success and survival. Parasite reproductive rate was also increased in mixed strain groups. Reduced host reproductive success was suggested to be directly due to the genetic heterogeneity of the parasitic infections resulting in increased host defence costs. Reduced host survival was consistent with an adaptive conditional parasite response.

Variant antigenic types (VATs) represented in a total of 47 first relapse populations of 6 clones of Trypanosoma brucei LUMP 227 were identified by immunofluorescent staining of living trypanosomes, using antiserum raised against purified surface antigens. The relative growth rates of these 6 clones were measured both individually and when grown together in a mixed population, and were found to be different under these two sets of conditions. A pattern emerged in the VATs represented in relapses of each clone, with some types being expressed more frequently than others and certain VATs being only very rarely expressed. It is suggested that new VATs are expressed according to a statistically definable order of priority which is different for each parent VAT, and that some VATs may be able to change to certain others only after passing through an intermediate VAT. The order of priority of appearance of VATs does not appear to correlate with growth rate measured either in individual clones or when clones are grown in a mixed population.

Two experiments are described in each of which three groups of calves received infective larvae of Ostertagia ostertagi daily, each group at a different rate. Changes in worm burdens were followed by the periodic slaughter of calves.

Numbers of adult worms quickly rose to and remained at levels which bore a positive relation to the infection rate. There was evidence that a turnover of worms was occurring and it appeared that burdens of adult worms were regulated by a loss of worms which depended on the number present. As the host's experience of infection increased, the adult worms present were smaller and an increasing proportion of females lacked a fully developed vulval flap. Evidence is presented which suggests that these were effects of resistance and that they depended on separate mechanisms.

Faecal egg counts of groups carrying different numbers of worms rose to the same peak and then declined according to the same logarithmic curve. This finding, together with data concerning the number of eggs in the uteri of the worms, suggested that the egg output of the entire population was restricted by the host to a limit which depended on previous egg-laying.

Infection did not have an adverse effect on the growth of the calves unless the infection rate exceeded a critical value. The resistance of calves to infection was impaired by clinical ostertagiasis.

Infection of suckling mice with Giardia trophozoites recovered from the intestines of 11 dogs autopsied in Central and Southern Australia in each case produced an established isolate. In contrast, only 1 isolate was obtained by inoculation of faecal cysts. The organisms grew poorly in comparison with isolates from humans or non-canine animal hosts. Light microscopy revealed that the trophozoites had median bodies with the ‘claw hammer’ appearance typical of G. intestinalis (syn. G. duodenalis, G. lamblia) but that they differed in shape and nuclear morphology from axenic isolates of human or canine origin. Allozymic analysis of electrophoretic data representing 26 loci and phylogenetic analysis of nucleotide sequences obtained from DNA amplified from the glutamate dehydrogenase locus showed that the 11 isolates examined from Australian dogs were genetically distinct from all isolates of G. intestinalis that have been established previously from humans and animals, and also from G. muris. Both analytical methods placed 10 of the Australian canine isolates into a unique genetic lineage (designated Assemblage C) and the eleventh into a deep-rooted second branch (designated Assemblage D), each well separated from the 2 lineages (Assemblages A and B) of G. intestinalis that encompass all the genotypes known to infect humans. In contrast, 4 axenic isolates derived from dogs in Canada and Europe (the only other isolates to have been established from dogs) have genotypes characteristic of genetic Assemblages A or B. The findings indicate that the novel Giardia identified in these rural Australian dogs have a restricted host range, possibly confined to canine species. The poor success rate in establishing Giardia from dogs in vitro suggests, further, that similar genotypes may predominate as canine parasites world-wide. The absence of such organisms among isolates of Giardia that have been established from humans by propagation in suckling mice indicates that they are unlikely to infect humans. However, infection of humans by those dog-derived genotypes that grow in vitro cannot be excluded.

The occurrence of multiple-clone Plasmodium falciparum haploid blood-stage infections is a pre-requisite for cross-fertilization and genetic exchange at the diploid stage in the mosquito. Using monoclonal antibodies against 3 polymorphic blood-stage antigens, a method of two-colour differential immunofluorescence allowed the resolution of between 1 and 4 clones/isolate. A mean of 2 P. falciparum clones was detected in the blood of malaria patients in The Gambia. The mean number of clones/patient showed no correlation with age, parasitaemia, or disease severity. There was a slight difference in mean number between sample periods, probably reflecting temporal differences in transmission intensity. A statistical analysis of 2-locus genetic diversity of clones within isolates concludes that not all multiple-clone infections result from superinfection, but that some are due to single multiple-clone inoculations.

Neospora caninum is an apicomplexan parasite that is closely related to Toxoplasma gondii, the causative agent of toxoplasmosis in humans and domestic animals. However, in contrast to T. gondii, N. caninum represents a major cause of abortion in cattle, pointing towards distinct differences in the biology of these two species. There are 3 distinct key features that represent potential targets for prevention of infection or intervention against disease caused by N. caninum. Firstly, tachyzoites are capable of infecting a large variety of host cells in vitro and in vivo. Secondly, the parasite exploits its ability to respond to alterations in living conditions by converting into another stage (tachyzoite-to-bradyzoite or vice versa). Thirdly, by analogy with T. gondii, this parasite has evolved mechanisms that modulate its host cells according to its own requirements, and these must, especially in the case of the bradyzoite stage, involve mechanisms that ensure long-term survival of not only the parasite but also of the host cell. In order to elucidate the molecular and cellular bases of these important features of N. caninum, cell culture-based approaches and laboratory animal models are being exploited. In this review, we will summarize the current achievements related to host cell and parasite cell biology, and will discuss potential applications for prevention of infection and/or disease by reviewing corresponding work performed in murine laboratory infection models and in cattle.

1. The egg-shell in many trematodes and pseudophyllidean cestodes is a quinone-tanned protein; and histochemical methods for polyphenol oxidase or for phenols may be used to demonstrate regions producing or containing shell material (i.e. the vitellaria, its ducts and the uterus) in whole mount preparations.

2. Specimens so treated may be further stained by routine whole mount methods. Gower's carmine is particularly recommended.

3. 70% alcohol was found to be the best general fixative for these methods.

4. These histochemical methods—with certain modifications—may also be used for demonstrating the egg-shell producing regions in sections.

5. Because of their affinity for basic proteins, bromo-phenol-blue and malachite green are suitable for staining these regions in sections. These stains are unsuitable for whole mounts.

6. There is some evidence to suggest that all trematodes and pseudophyllidean cestodes do not form and harden their egg-shell by quinone tanning; consequently, some forms may give negative or unsatisfactory results with these methods.