In this study we evaluated the effects of the anti-microsporidial exospore monoclonal antibody 3B6, recognizing 3 Encephalitozoon species, Encephalitozoon intestinalis (Syn. Septata intestinalis), Encephalitozoon cuniculi, and Encephalitozoon hellem on microsporidial growth in vitro. Pre-treatment of spores for 24 h with mAb 3B6 resulted in 21–29% fewer infected host cells 4 days after inoculation of the cultures compared to cultures pre-treated with medium or an irrelevant isotype control mAb (P<0·001). Fewer intracellular spores (1·2±0·2) in infected cells were found when mAb 3B6 was present in cultures compared to cultures with medium alone (4·3±0·8) or an irrelevant isotype control mAb (4·2±0·9; P<0·001). This decrease appeared not to be dependent on time of exposure, mAb concentration, or presence of complement. It is concluded that antibodies, particularly those directed to potential neutralizing-sensitive epitopes on spores, may have a role in the control of microsporidial growth in vitro.

Schistosomula of Schistosoma mansoni were incubated in RPMI 1640 medium containing 10% fetal calf serum, 10% human portal venous or 10% human peripheral venous sera in the presence of bromodeoxyuridine (BrdU) in order to measure differences in cell proliferation. The rates of cell proliferation as expressed by BrdU labelling indices (BLI) were determined as a function of time of incubation by immunohistochemistry using monoclonal antibody to BrdU. Compared to schistosomula cultured in the presence of RPMI plus 10% fetal calf serum, BLIs were increased by 60% in the presence of human portal, but not in peripheral serum. This stimulatory effect was substantially reproduced by a fraction of portal serum with a molecular weight range between 1 and 50 kDa. However, in the presence of human peripheral venous serum, either whole or fractionated, schistosomula showed no significant difference compared to RPMI plus 10% fetal calf serum alone. Furthermore, human portal serum fractions of molecular weight greater than 50 kDa also revealed no significant difference relative to control. The results indicate that portal venous serum component(s) of a molecular weight range higher than most simple nutrients can greatly stimulate the rate of cell proliferation of Schistosoma mansoni schistosomula.

Observations that growth of Plasmodium falciparum in vitro is inhibited by high temperatures have led to hypotheses that malaria fever may influence the parasite population dynamics, regulating parasite density and synchronizing parasite growth. In order to investigate the fever hypotheses, we have developed an age-structured coupled Markov chain model that describes the parasite erythrocyte cycle and its interaction with the host fever response. We estimated the model parameters using data collected from laboratory parasite cultures that were exposed to febrile or normal temperature. Using the experimental parameter values, quantitative predictions were made of the effect of fever in determining the parasite population dynamics. It was concluded from the model behaviour that, during the primary infection of a non-immune host, a typical episode of fever can effect density-dependent regulation of the parasite population, maintaining cycles of parasitaemia and promoting synchronous parasite growth.

The mathematical model of the in-host dynamics for malaria parasites of Anderson, R. M., May, R. M. & Gupta, S. (1989), Parasitology99 (Suppl.) S59–S79, and subsequently used by other authors, contains an error. This leads to very substantial over-estimates in parasite growth rates. A corrected form of the model is presented in this paper.

Prion diseases of humans and other animals have clinical, economic and political significance. Pre-mortem diagnosis is therefore of great importance. Clinical diagnostic criteria and the current status of the available diagnostic tests are reviewed and possible future developments discussed. Presently, most diagnostic tests are indirect, relying on findings that may not be confined to prion diseases rather than on their particular intrinsic nature. They are therefore not absolutely specific nor are they 100% sensitive. The electroencephalogram (EEG), cerebral imaging and cerebrospinal fluid (CSF) analysis are the main techniques employed. However, there is hope that useful blood tests could be developed which would be simpler and less invasive. Also, there is hope that more specific tests will become available.

In the present study the gp63 gene locus was used as a target for genetic characterization of Leishmania parasites by 2 methods: (i) RFLP analysis with several restriction enzymes (gp63–RFLP), and (ii) intra-genic PCR amplification coupled with restriction analysis (PCR–RFLP). Both methods were applied to a large number of natural isolates belonging to 4 species of the subgenus Viannia, namely L. (V.) braziliensis, L. (V.) peruviana, L. (V.) guyanensis and L. (V.) lainsoni: reference stocks of subgenus Leishmania were included as outgroups. Multilocus isoenzyme typing (MLEE) was used as a reference. On the one hand gp63–RFLP evidenced an extensive polymorphism and revealed specific markers for subgenus, species and geographical populations: congruence with MLEE was demonstrated statistically. The particular interest of gp63–RFLP was illustrated by infra-specific polymorphism, because of the possible relationship with phenotype diversity. On the other hand intra-genic amplification was less resolutive than gp63–RFLP, but also allowed discrimination of the 2 subgenera (PCR alone) and all the species tested in the subgenus Viannia (PCR–RFLP). PCR–RFLP presents an important operational advantage as it allows genetic characterization of minute amounts of parasites, using Leishmania specific primers. The polymorphism revealed by gp63–RFLP and PCR–RFLP illustrates the very high genomic and genetic plasticity of gp63 genes.

A specific extracellular proteinase, degrading selectively the cecropin-based defence system of insects, is secreted into the larval body during parasitism of the greater wax moth by the Heterorhabditis bacteriophora/Photorhabdus luminescens complex and by phase 1 of P. luminescens. The proteolytic digestion of insect inducible cecropin-like immune molecules was demonstrated by the disappearance of the Galleria mellonella cecropins and purified Hyalophora cecropin B peptide PAGE bands upon exposure to infected extracts, and a similar abrogation of antibacterial activity using an agar diffusion assay. Proteolytic activity of infected extracts produced by nematode/bacterial complex and phase 1 variant of P. luminescens was shown to be correlated with cecropin-inhibitory activity, suggesting that this anti-cecropin agent may be responsible for the ability of bacteria to establish infection and the insecticidal nature of H. bacteriophora. Antibacterial activity of Galleria lysozyme and that of chicken egg-white lysozyme to which P. luminescens is insensitive, was unaffected by H. bacteriophora proteinase.

Antibodies to a non-repeat region of the Plasmodium falciparum antigen Pf155/RESA were investigated for their capacity to inhibit parasite cytoadherence to melanoma cells and parasite growth in vitro. The activities of these antibodies were studied since the target region in Pf155/RESA includes a cytoadherence-related motif also found in loop 3 and 7 of human erythrocyte band 3 protein. Overlapping multiple antigen peptides (MAPs) together spanning residues 199–220 of Pf155/RESA were used to raise antibodies in rabbits. Analysis of the fine specificity of these antibodies revealed that antibodies raised against largely overlapping sequences displayed highly different specificity patterns. Similarly, striking differences were seen when analysing the biological effect of antibodies to these MAPs. Antibodies to the cytoadherence-related motif of Pf155/RESA, as well as antibodies raised against a MAP based on a corresponding band 3 motif, inhibited cytoadherence but not parasite growth. In contrast, antibodies to sequences adjacent to the Pf155/RESA cytoadherence motif inhibited parasite growth in vitro but had no effect on cytoadherence.

A single exposure to radiation-attenuated cercariae of Schistosoma mansoni induces a high level of protective immunity in C57BL/6 mice, which is mediated by Th1 responses. Events in the skin and/or draining lymph nodes early after exposure are crucial for the induction of protection, and we have investigated the interactions of vaccinating parasites with host leucocytes in these 2 locations. We observed extensive lateral spreading of cercarial secretions along layers of the stratum corneum but not between keratinocytes. There was little direct contact with host leucocytes during the first 1–2 days when the parasites lay at the base of the epidermis, but cells accumulated in the underlying dermis. In contrast to normal parasites, attenuated larvae persisted in the dermis for >10 days, often surrounded by aggregates of macrophages/dendritic cells. Whilst cells bearing MHC II, CD11b or CD11c markers were present in the lymph nodes, particularly in the periphery and paracortical areas, no obvious redistribution was seen as a result of parasite residence there for 5–15 days. However, ultrastructural observations revealed numerous cells with macrophage/dendritic morphology in the vicinity of parasites, in some instances closely adherent to the tegument. The observations strongly suggest that the tegument is a potent source of the antigens which prime the immune system in the lymph nodes of vaccinated mice for a protective response.

In the accompanying manuscript Saul (1998) points out that a model of within-host malaria population dynamics (Anderson, May & Gupta, 1989) can exhibit unrealistically large growth rates. He suggests that this error can be avoided by replacing the parameter r, which is the number of merozoites produced by each parasite at schizogony, with the value ln (r)+1. This substitution does not, however, address the true underlying problem with the model, namely that whilst in reality there is small variation in the distribution of Plasmodium spp. life-spans, the use of the constant rate α assumes an exponential, and hence much more variable, distribution. This can allow the population to increase, over the time period of the average life-span (48 h in the case of Plasmodium falciparum), by factors considerably larger than r. Saul identifies this assumption as unrealistic in terms of the biology of malaria, but it is not remedied in the proposed model.

Within the structure of the original model there are two ways of addressing the growth rate problem. Firstly, the ‘growth constant’ r can be replaced by the value ln (r)+1, so that if all parasites reinvade, the model increases by a factor r over 1 generation. As pointed out by Saul, this is an artificial device since it means that each parasite produces a reduced number of merozoites. Alternatively, a parasite can produce the observed number of merozoites, r, many of which do not reinvade. This is the situation in the original paper and Gravenor, McLean & Kwiatkowski (1995). In these papers the model does grow at a reasonable rate because the parameter β is estimated directly from observed growth rates. These points, however, are only an aside. Both the original and Saul's modified model can grow at the same rate and they both have the same distributional assumptions concerning parasite life-span.

Since they were first utilized, immunoassays have witnessed phenomenal growth in the range and scope of their applications. A vast array of different labels and assay strategies has been developed to meet the requirements of sensitivity, accuracy and convenience. The development of increasingly sensitive labels and detection equipment has seen a drastic improvement in the sensitivity of immunoassay systems, allowing an ever-increasing range of analytes to be measured accurately. At the same time, simple to use, inexpensive assay systems have been developed with the necessary reliability, accuracy and sensitivity to bring immunoassay technology to much more diverse areas such as home testing, near-patient monitoring, and large screening programmes in developing countries. Recent developments in molecular biology techniques have made possible the production of fusion antibody conjugates, which can lead to further improvements in sensitivity and cost of reagents, as well as possibly revolutionizing the production of monoclonal antibodies. However, dissatisfaction with various aspects of existing immunoassay technology will necessarily lead to the continued development of this already widely diverse subject.

We have investigated the influence of heteroxenous coccidia (Apicomplexa: Eimeriorina) on the predation risk of intermediate hosts. Voles infected with Frenkelia spp. were found more frequently in buzzards' (Buteo buteo) prey than among snap-trapped rodents. To eliminate the possibility of traps selecting for uninfected rodents, a laboratory experiment was performed. Mice experimentally infected with Sarcocystis dispersa seemed to be more likely caught by the final host, the long-eared owl (Asio otus); this result was confirmed by a mathematical model. Field data confirmed the adaptive value of parasite-induced changes. The increase of predation is directed towards the specific final host only or is non-specific. In the populations studied the probability of predation of parasitized individuals by the specific predator was increased.

After infection with sporozoites of the protozoon Theileria parva (Tp) bovine T cells are readily transformed to permanent growth in vivo and in vitro. Their transformed state depends on the constant presence of the parasite but membrane signals remain important. Non-receptor tyrosine kinases play a critical role in the transduction of membrane signals in haematopoietic cells. We have investigated Src-family kinases in bovine T cells transformed by Tp. The T cell receptor-associated tyrosine kinase p60fyn had high activity in all cell lines tested. In addition, weak phosphorylation of 2 novel bands was observed associated with Fyn. In contrast to Fyn, enzymatic activity of p56lck, which in T cells has an essential role in signalling, was low. Furthermore, 1 of 3 Tp transformed cell lines was completely devoid of p56lck indicating that the enzyme is not necessary for the Tp dependent growth of the T cells. In addition to p60fyn and p56lck weak enzymatic activity of 1 splice variant of p53/56lyn was observed after infection of T cells with Tp. These data show that growth transformation by Tp influences kinase activity in bovine T cells. However, they also prove that p56lck does not play an essential role in the transformation mechanism.

Parasitic sex ratio distorters were artificially transferred within and between crustacean host species in order to study the effects of parasitism on host fitness and sex determination and to investigate parasite–host specificity. Implantation of Nosema sp. to uninfected strains of its Gammarus duebeni host resulted in an active parasite infection in the gonad of recipient females and subsequent transovarial parasite transmission. The young of artificially infected females were feminized by the parasite, demonstrating that Nosema sp. is a cause of sex ratio distortion in its host. In contrast, we were unable to cross-infect Armadillidium vulgare with the feminizing microsporidian from G. duebeni or to cross-infect G. duebeni with the feminizing bacterium Wolbachia sp. from A. vulgare.

Diseases caused by viruses are a constant and major problem for livestock production world wide. The diseases range from highly contagious acute forms with high mortality, to chronic disabling diseases with an insidious effect on production. Such diseases cannot be regarded as static in nature due to the highly mutable nature of viruses and their direct selection at the host level and indirect selection in vectors, inducing changes in pathogenicity. Considerable efforts are needed to control these diseases including accurate and rapid diagnosis using both classical and emerging technologies. The methods used are based on both serological and molecular biological based methods. The ELISA and use of monoclonal antibodies are significant in the serological field whereas the Polymerase Chain reaction (PCR) and its direct and indirect uses for identifying (sequencing) and amplification of gene products, is vital to both research and applied fields. Both areas have to be used in a complementary way in the diagnosis of virus diseases.

The amphipod Gammarus duebeni harbours several species of vertically transmitted, feminizing microsporidian parasites. G. duebeni were collected from 3 localities in the UK. Animals from Budle Bay, Northumberland, were infected with Octosporea effeminans, and those from Millport, Isle of Cumbrae and Fenham Flats, Northumberland were infected with microsporidia of the genus Nosema. We derived expected distributions of parasites per host embryonic cell by modelling parasite transmission as a multitype, Galton–Watson branching process. Parasite prevalence (proportion of females infected) was significantly heterogeneous among localities. Parasite burden in zygotes was much higher for females infected with Nosema than in animals infected with O. effeminans. There was no significant difference between localities in the number of Nosema in the zygotes. Comparison of models and data from 64-cell host embryos showed that the distributions of parasites per cell were consistent with the hypothesis that sorting of parasites into daughter cells is biased for at least 1 cell lineage. Host embryos infected with O. effeminans could expect to contain a growing number of parasites in each cell generation within such biased cell lineages; similar estimates for Nosema predict a decline in the number of parasites per cell within a biased lineage. We discuss the possibility that the 2 species of parasite may be employing different strategies in order to ensure transmission to the next host generation.

An enzyme presenting kallikrein-like activity (designated sK1) was purified from the supernatant of Schistosoma mansoni adult worm homogenate. The enzyme cleaves bradykinin from purified rat plasma kininogen. Activity was optimal at pH 9·0 and the enzyme showed amidolytic activity, since it hydrolysed the kallikrein synthetic substrate d-Pro-Phe-Arg-p-nitroanilide. The activity of sK1 upon rat plasma kininogen was strongly inhibited by the serine proteinase inhibitors phenylmethanesulfonyl fluoride, aprotinin or soybean trypsin inhibitor, but not by ethylenediaminetetraacetic acid or sodium tetrathionate. The molecular mass of sK1, as determined by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, was 66 kDa and the pI value, estimated by analytical chromatofocusing, was 4·2. Physical and chemical properties suggest that sK1 is a serine proteinase of the kallikrein family. Evidence is presented which suggests that sK1 is a component of the tegumental surface of the parasite and the levels of its activity in the male adult worm are approximately 21 times higher than those in the female adult worm. The intravenous injection of 3 μg of sK1 into an anaesthetized rat induced a drastic reduction in the arterial blood pressure of the animal. This effect lasted for about 1 min, and was followed by a progressive recovery of the arterial pressure. Neither bradycardia nor cardiac arrhythmias were noticed, suggesting a peripheral vasodilation effect. The presence of sK1 on the surface of adult male worms could play an important role in the wandering capacity of coupled worms into the visceral vasculature of the host.

Stochastic processes play a vital role in the early stages of the evolution of drug-resistant malaria. We present a simple and flexible method for investigating these processes and understanding how they affect the emergence of drug-resistant malaria. Qualitatively different predictions can be made depending on the biological and epidemiological factors which prevail in the field. Intense intra-host competition between co-infecting clones, low numbers of genes required to encode resistance, and high drug usage all encourage the emergence of drug resistance. Drug-resistant forms present at the time drug application starts are less likely to survive than those which arise subsequently; survival of the former largely depends on how rapidly malaria population size stabilizes after drug application. In particular, whether resistance is more likely to emerge in areas of high or low transmission depends on malaria intra-host dynamics, the level of drug usage, the population regulation of malaria, and the number of genes required to encode resistance. These factors are discussed in relation to the practical implementation of drug control programmes.

A decline in serum thyroxine (T4) occurs in Nya[ratio ]NYLAR female mice infected with Toxoplasma gondii. To ascertain whether the hypothyroxinaemia might be the result of primary thyroid dysfunction, 2 parameters of thyrofollicular cell function were monitored to determine (a) if the cell surface membrane receptors for thyroid-stimulating hormone (TSH) were operative, and (b) whether the cyclic adenosine monophosphate (cAMP)-dependent cascade of intracellular events leading to the release of T4 was responsive to exogenous cAMP. Our results indicated that both parameters were intact and functional in the infected-mouse thyrocytes. However, the elicited T4 responses were distinctly diminished in magnitude, reflecting a lack of readily available thyroidal T4 reserves. Because the continuing synthesis, storage, and release of T4 is dependent on the pulsatile stimulation of the thyroid by TSH, we suggest that the depletion of T4 reserves is likely due to perturbation of the pulsatile release of TSH from the pituitary, rather than to primary thyroid malfunction.

Failure of snail reinfection by Schistosoma mansoni has been demonstrated in susceptible Biomphalaria glabrata infected with 1 miracidium and subsequently re-exposed to 1 or 5 homologous parasite larvae. The acquisition of ‘resistance’ to secondary parasite infection was time dependent since complete inhibition was observed at 2 weeks and longer following monomiracidial exposure. This phenomenon was still observed in snails challenged 8 weeks after primary infection. Histological observations revealed that sporocysts from the challenge infection were free of encapsulation, their development was stopped and they degenerated slowly in the absence of haemocytic reaction of the host. Under the hypothesis of an acquired homologous resistance mechanism, this strongly suggests that 1 or several unidentified humoral factors are responsible for the non-development of the sporocysts from the challenge infection. However, considering the time-dependent nature of the phenomenon, an intraspecific larval antagonism process between sporocysts resulting from the primary infection and those from the challenge infection may be involved.