Cryptosporidium are parasitic protozoans that cause gastrointestinal disease and represent a significant risk to public health. Cryptosporidium oocysts are prevalent in surface waters as a result of human, livestock and native animal faecal contamination. The resistance of oocysts to the concentrations of chlorine and monochloramine used to disinfect potable water increases the risk of waterborne transmission via drinking water. In addition to being resistant to commonly used disinfectants, it is thought that oocysts can persist in the environment and be readily mobilized by precipitation events. This paper will review the critical processes involved in the inactivation or removal of oocysts in the terrestrial and aquatic environments and consider how these processes will respond in the context of climate change.

Ascaris is a large parasitic roundworm (nematode) of the small intestine of humans and pigs, which causes the socio-economically important disease, ascariasis. To better understand the relationship of Ascaris between the 2 host species, recent studies in China have focused on investigating the genetics and epidemiology of Ascaris from humans and pigs using a mutation scanning-based approach. Findings provided support for a low level of gene flow between the human and porcine Ascaris populations. Extending the studies of genotypic variability within Ascaris from humans and pigs, experimental infections of mice and pigs with selected genotypes of Ascaris were carried out. Initial results indicate that there is a significant difference in the ability of Ascaris eggs of genotype G1 (derived from human) and G3 (derived from pig) to infect and establish as adults in pigs, supporting the difference in the frequencies of these genotypes in natural Ascaris populations between pigs and humans in China. Taken together, current information supports that there is limited cross-infection of Ascaris between humans and pigs in endemic regions and that pigs are not a significant reservoir of human infection with the adult nematode in such areas.

Multiplication of Plasmodium parasites within human erythrocytes is essential to malarial disease. The cell-division cycle of this organism, however, is still poorly understood. In other eukaryotes, various techniques for (apparent) cell-cycle synchronization have been used to shed light on the mechanisms involved in cell division and its control. Thus far there is no technique for cell-cycle synchronization (as opposed to selection of parasites of a limited age-range) in Plasmodium. We therefore investigated the possibility that inhibitors of DNA synthesis, the mitotic spindle, or cell-cycle control elements (such as cyclin-dependent kinases) could be used to synchronize P. falciparum cultures to a particular cell-cycle phase. Surprisingly, most of these compounds did not cause a block at a specific phase. Three compounds, Hoechst 33342, roscovitine and L-mimosine, did block development at the trophozoite–schizont transition (S or G2 phase). The block caused by the latter 2 inhibitors was reversible, suggesting that they might be used as synchronizing agents. However, a consideration of the perturbing effects of inhibitors and problems with ‘batch’ synchronization techniques in general lead us to believe that any results obtained using roscovitine- or L-mimosine-treated parasites may not be reflective of the normal cell cycle.

The genetic diversity of Cryptosporidium spp. and Giardia duodenalis from dairy cattle and pigs in Denmark was determined in the present study. Faecal samples from 1237 pigs and 1150 cattle originating from 50 sow herds and 50 dairy herds, respectively, were analysed for the presence of the two parasites by immunofluorescence microscopy. A large proportion of the (oo)cyst containing samples were selected for molecular characterization. Sequencing and phylogenetic analysis of the 18S rDNA locus and/or the HSP70 gene of 183 pig and 154 cattle isolates of Cryptosporidium revealed the presence of C. suis, pig genotype II, C. parvum (cattle genotype), C. bovis, Cryptosporidium deer-like genotype and a novel C. suis-like genotype. For both cattle and pigs, a host age-related change in distribution of species/genotypes was observed. The zoonotic C. parvum (cattle genotype) was most prevalent in young calves. For Giardia, 82 and 145 isolates from pigs and cattle, respectively, were analysed at the 18S rDNA locus and/or the gdh gene. Giardia isolates belonging to the zoonotic Assemblage A was found in both young and older calves, as well as in weaners and piglets, whereas cows seemed to be infected purely by isolates of the livestock group, Assemblage E.

Cryptosporidium parvum is composed of multiple subtypes, which appear to be zoonotic or anthroponotic. Therefore, in order to understand the molecular epidemiology of this species and to control the zoonotic transmission, it is essential to identify the isolates at the subtype level. In the present study, 60·2% of 103 examined dairy calves from 10 farms in the Belgrade area were positive for Cryptosporidium infection by PCR, and all of them were identified as C. parvum by PCR-RFLP of SSUrRNA and COWP genes. Eighteen C. parvum isolates selected randomly from 9 positive farms were classified by molecular phylogenetic analysis of the 60 kDa glycoprotein gene sequences into 3 subtype allele families, IIa and IId, which are known to be zoonotic, and a new family IIj. Furthermore 10 isolates in IIa were classified into 3 subtypes IIaA16G1R1b, IIaA18G1R1, IIaA20G1R1, 2 in IId into IIdA18G1b, and 6 in IIj into IIjA16R2 and IIjA17R2. The last 2 subtypes in IIa and 2 subtypes in IIj were new subtypes. These results suggest that C. parvum isolates harboured in calves in the Belgrade area have zoonotic potential and C. parvum is genetically diverse only in a limited area. This is the first molecular epidemiological report about Cryptosporidium infection in Serbia and Montenegro.

In vitro propagation followed by PCR, and a PCR-based method capable of the direct detection of Blastocystis in faeces were utilized to detect Blastocystis from various hosts in Australia, including primates and their handlers from the Perth Zoo. In addition, Blastocystis isolates from dogs and humans living in a localized endemic community in Thailand were also characterized genetically. PCR-based detection directly from faeces was shown to be more sensitive compared with in vitro culture for the detection of Blastocystis. Moreover, phylogenetic analysis of Blastocystis isolates amplified utilizing in vitro techniques prior to PCR revealed that this method favoured the preferential amplification of Blastocystis subtype 5 over subtype 1. This study is the first to provide molecular-based evidence supporting the zoonotic potential of Blastocystis in dogs, possums and primates in a natural setting.

Heat shock is believed to be a developmental inductor of differentiation in Leishmania. Furthermore, heat shock genes are extensively studied as gene models to decipher mechanisms of gene regulation in kinetoplastids. Here, we describe the organization and expression of the HSP70 loci in representative Leishmania species (L. infantum, L. major, L. tropica, L. mexicana, L. amazonensis and L. braziliensis). With the exception of L. braziliensis, the organization of the HSP70 loci was found to be well conserved among the other Leishmania species. Two types of genes, HSP70-I and HSP70-II, were found to be present in these Leishmania species except for L. braziliensis that lacks HSP70-II gene. Polymorphisms in the HSP70 locus allow the differentiation of the Old and New World species within the subgenus Leishmania. A notable discrepancy between our data and those of the L. major genome database in relation to the gene copy number composing the L. major HSP70 locus was revealed. The temperature-dependent accumulation of the HSP70-I mRNAs is also conserved among the different Leishmania species with the exception of L. braziliensis. In spite of these differences, analysis of the HSP70 synthesis indicated that the HSP70 mRNAs are also preferentially translated during heat shock in L. braziliensis.

Leishmania infantum and Leishmania donovani both pertain to the L. (L.) donovani complex and are responsible for visceral leishmaniasis. To explore the L. donovani complex, we focused our study on cysteine protease B (cpb) and especially on 2 cpb copies: cpbE and cpbF. We selected cpb genes because of their phylogenetic interest and host–parasite interaction involvement. Sequencing these 2 copies revealed (i) that cpbE is specific to L. infantum and cpbF is specific to L. donovani and (ii) that these 2 copies are different in length and sequence.Nucleotide sequence data reported in this paper are available in the GenBank database under Accession numbers AY896776 AY896777, AY896778, AY896779, AY896780, AY896781, AY896782, AY896783, AY896784, AY896785, AY896786, AY896787, AY896788, AY896789, AY896790, AY896791. Phylogenetic analysis and protein predictions were carried out in order to compare these copies (i) with other trypanosomatid cpb, especially L. mexicana, and (ii) within the L. donovani complex. Our results revealed patterns specific to the L. donovani complex such as the COOH-terminal extension, potential epitopes and N-glycosylation sites. Moreover, phylogenetic analysis revealed different levels of polymorphism between L. infantum and L. donovani and confirmed the ancestral status of the latter. L. infantum has a shorter sequence and a deleted sequence responsible for modifications in protein conformation and catalytic triad. Considering the clinical aspect, L. infantum dermotropic strains appeared more polymorphic than L. infantum viscerotropic strains.

Wildlife can act as reservoir of different tick-borne pathogens of veterinary and zoonotic importance. To investigate the role of wild ruminants as reservoir of piroplasm infection, 28 red deer, 69 roe deer and 38 chamois from Northern Spain were examined by reverse line blot (RLB) hybridization. The survey detected a prevalence of 85·7% in red deer, 62·3% in roe deer and 28·9% in chamois. Four different piroplasms were identified: Theileria sp. OT3 (previously described in sheep) as the most prevalent (85·7% in red deer, 46·4% in roe deer and 26·3% in chamois); Theileria sp. 3185/02 (previously described in a red deer in Central Spain) more abundant in red deer (53·6%) than in roe deer (10·1%) but absent from chamois; Babesia divergens detected in 6 roe deer; Theileria ovis present in 1 chamois. Mixed infections (Theileria sp. OT3 and Theileria sp. 3185/02) were only found in red and roe deer. Sequencing analysis of the 18S rRNA gene confirmed the RLB results and showed 99·7% identity between Theileria sp. 3185/02 and T. capreoli, suggesting that they are the same species. Tick distribution and contact of wild ruminants with domestic animals are discussed in terms of piroplasm infection. The results suggest that a considerable number of wildlife ruminants are asymptomatic carriers that may serve as reservoirs of the infection posing a serious concern in terms of piroplasmosis control.

In this study we examine the population biology of Brueelia apiastri, a chewing louse living on the European bee-eater (Merops apiaster). We investigate the relationships between parasite intensity of infestation, sex ratio, reproductive output, parasite size and their environment i.e. the morphology, condition, age and sex of the host. Chewing lice were collected, their sex and age (developmental stage) identified and parasite body size determined as a measure of parasite condition (larger individuals consume larger meals and larger females may produce larger clutches). The data show that there is variation in intensity as well as body size of B. apiastri between individual bee-eaters and this variation is independent of the sex of the birds. However, size, condition and age of the birds seem to influence the infestation rates with B. apiastri. The study suggested size-dependent depredation, since more, smaller chewing lice (usually nymphs) living on birds in better condition and birds having longer bills. Furthermore, more male chewing lice (males are smaller than females) live on older birds. Intraspecific competition between parasites seems to have a negative effect on female but not male body size but this result could be also explained by size-dependent depredation.

Varroa mites (Varroa destructor) are serious ectoparasites of honey bees (Apis mellifera). This research addresses the impact of varroa mites on survivorship, viral incidence, and physiological traits of newly-emerged worker bees. RT-PCR confirmed our previous finding that varroa parasitization was linked to high levels of deformed wing virus (DWV). In non-treatment bees, varroa parasitization combined with increased viral levels altered survivorship curves from long-survival to shorter-survival types. After challenge with live Escherichia coli, the survivorship of mite-parasitized bees was significantly lower than mite-free bees. Deformed-wing, mite-parasitized bees died on average within 1 day, even without E. coli challenge. This was correlated with the absence of an important enzyme activity in insect immunity, phenol oxidase, lacking even in those bees challenged with immuno-elicitors. The lack of inducible phenol oxidase activity indicated that the bee immune system is not fully competent upon adult emergence. Varroa parasitism also significantly reduced body weight of the parasitized bees, but body weight was not significantly correlated with the survivorship of mite-parasitized bees. Our research indicates that the combination of mite parasitization, the interaction of DWV and microbes, and a developmental immune incompetency attribute to decreased worker survivorship and have a negative impact on colony fitness.

Many zoonotic disease agents are transmitted between hosts by arthropod vectors, including fleas, but few empirical studies of host-vector-microparasite dynamics have investigated the relative importance of hosts and vectors. This study investigates the dynamics of 4 closely related Bartonella species and their flea vectors in cyclic populations of field voles (Microtus agrestis) over 3 years. The probability of flea infestation was positively related to field vole density 12 months previously in autumn, but negatively related to more recent host densities, suggesting a dilution effect. The 4 Bartonella species exhibited contrasting dynamics. Only B. grahamii, showed a distinct seasonal pattern. Infection probability increased with field vole density for B. doshiae, B. taylorii and BGA (a previously unidentified species) and with density of coexisting wood mice for B. doshiae and B. grahamii. However, only the infection probability of BGA in spring was related to flea prevalence. B. doshiae and BGA were most common in older animals, but the other 2 were most common in non-reproductive hosts. Generally, host density rather than vector abundance appears most important for the dynamics of flea-transmitted Bartonella spp., possibly reflecting the importance of flea exchange between hosts. However, even closely related species showed quite different dynamics, emphasising that other factors such as population age structure can impact on zoonotic risk.

Hookworms feed on blood, utilizing haemoglobin for nutrition, growth and reproduction. The haemoglobin digestion cascade has been partially elucidated, but the process immediately preceding this event, haemolysis, has received considerably less attention. We have cloned and expressed Ancylostoma caninum mRNAs encoding 2 proteins belonging to the saposin-like protein (SAPLIP) family, termed Ac-slp-1 and Ac-slp-2. The open reading frames of SLP-1 and SLP-2 were used to identify expressed sequence tags encoding SAPLIPs from the 4 major clades of animal parasitic nematodes. Both Ac-slp-1 and slp-2 mRNAs were shown to be expressed in all life stages assessed, with slp-1 predominantly being expressed in third-stage larvae (L3) before and after activation with dog serum. Recombinant SLP-1 and SLP-2 were expressed in insect cells and used to raise specific antisera in mice. These antisera were used as probes in fluorescence microscopy to localize the anatomic expression sites of both proteins to small, punctate organelles or vesicles within the intestinal cells of adult worms; weak staining was detected on the microvillar brush border of the intestine. Using transmission electron microscopy, both proteins were localized to similar vesicles in the intestinal cells of the L3. Recombinant proteins contained C-terminal purification tags that potentially precluded dimerization and possibly interfered with the subsequent detection of haemolytic activity. Their expression in the gut of the L3 and adult stages suggests a role for these hookworm SAPLIPs in the lysis of host cells during tissue migration and/or feeding.

We studied patterns of variation in parasite communities of 2 closely related species of Mastomys rodents. These 2 species live in sympatry in South-eastern Senegal, but differ drastically in their habitat choice. We asked (a) whether the host species have the same parasites; (b) whether there is any observable pattern relative to the host species/habitat type in the structure of parasite communities; (c) whether the variability in parasite community for each host species is related to habitat characteristics. We analysed 220 and 264 individuals of each host species, sampled respectively in 10 and 11 trap sites. Twenty parasite taxa were recorded, and the majority were nematodes. Between-host species comparisons showed that helminth communities were slightly more diversified in M. natalensis. Many parasite species were found in both Mastomys. However, various helminth taxa varied in frequency and abundance between host species. Within each host species, helminth diversity, prevalence and/or abundance of some parasites were correlated with habitat or host population factors that may influence parasite life-cycles, such as village structure, or the presence/absence of a pool. Our results suggest that habitat characteristics have a strong impact on helminth community structure.