Haemonchus contortus infections in small ruminants are of major economic importance worldwide. Heat shock proteins (HSPs) are a family of molecular chaperones that play important roles in the process of invasion and survival of nematodes. Although HSP70 has been identified in several parasitic nematodes, little is known of its distribution and function in Haemonchus contortus. The aims of this study were to characterize HSP70 from Haemonchus contortus (designed as Hc-hsp70), express Hc-hsp70 and analyse the promoter activity in Caenorhabditis elegans. Bioinformatic analysis revealed that the open reading frame of the Hc-hsp70 cDNA encodes a 646-amino acid peptide, which is highly conserved in comparison to HSP70 in other nematodes. Phylogenetic analysis indicated that H. contortus is closely related to Caenorhabditis. The 5′-flanking region promoted green fluorescence protein (GFP) expression in the intestine in all larval stages and adult with 2 expression patterns in C. elegans. Expression of Hc-hsp70 mRNA transcripts in C. elegans increased following 2, 4, 6 h of heat shock and peaked at 4 h. However, its expression induced down-regulation of hsp-1 of C. elegans. These results suggest that the H. contortus hsp70 might have a similar function to that of C. elegans hsp-1.

We characterized partial sequences of 18S rDNA from sedge warblers infected with a parasite described previously as Hepatozoon kabeeni. Prevalence was 47% in sampled birds. We detected 3 parasite haplotypes in 62 sequenced samples from infected animals. In phylogenetic analyses, 2 of the putative Hepatozoon haplotypes closely resembled Lankesterella minima and L. valsainensis. The third haplotype grouped in a wider clade composed of Caryospora and Eimeria. None of the haplotypes showed resemblance to sequences of Hepatozoon from reptiles and mammals. Molecular detection results were consistent with those from microscopy of stained blood smears, confirming that the primers indeed amplified the parasite sequences. Here we provide evidence that the avian Hepatozoon-like parasites are most likely Lankesterella, supporting the suggestion that the systematic position of avian Hepatozoon-like species needs to be revised.

Biting midges of the genus Culicoides (Diptera: Ceratopogonidae) are vectors of a number of viral diseases worldwide. Following the unforeseen outbreak of bluetongue in northern Europe (2006–2009) there was a need to clarify on-farm breeding substrates utilized by temperate Culicoides spp. Six substrates (cow dung, cow slurry, horse dung, sheep dung, maize silage and soil) were investigated for Culicoides spp. emergence over a 31-week period. Overall, most Obsoletus group Culicoides emerged from the cow dung and the most Pulicaris group Culicoides emerged from the sheep dung. Furthermore, Culicoides of the Obsoletus group were found to be abundant in cow slurry and sheep dung. Temperature played a significant role in the emergence times of adult Culicoides. The Obsoletus group appear to have undergone 3 generations during the experimental period. The sex ratio of emergent Obsoletus group Culicoides was affected by substrate type, with a greater proportion of males emerging from cow dung and slurry compared with the other substrates.

Human–wildlife interactions have reached unprecedented levels, and humans are influencing the earth's ecosystems more rapidly and extensively than ever before. This situation is cause for serious concern, especially since disease interactions between wildlife and humans have been recognized as major conservation threats. In this study, long-tailed macaques, Macaca fascicularis, from 2 forest parks located in north-eastern Thailand were investigated to determine the influence of habitat modification by humans on helminth parasite associations in non-human primates. Macaque populations with contact to anthropogenically modified environments were compared with sylvatic groups in nearby natural environments. In order to test for human–non-human primate transmission of parasites, the local human populations were also examined. Humans were infected with a number of potentially pathogenic parasites, including Opisthorchis viverrini and Strongyloides stercoralis. However, eggs of these helminths were not detected in macaque feces. Thus, no direct parasite transfer from humans to non-human primates could be confirmed. However, macaque groups with more frequent contact with human-modified habitats, and a higher portion of human-provided food in their diet, had significantly higher prevalences and intensities of Strongyloides fuelleborni and of an intestinal fluke (probably Haplorchis sp.) than sylvatic groups. Positive correlations were found between the time foraging on the ground and infection with S. fuelleborni, and the amount of human-provided food and intestinal fluke infection. Human alteration of habitat and associated modifications in non-human primate behaviour are likely to play a role in determining the occurrence, prevalence and intensity of zoonotic helminth infection of wild non-human primates.

Recently, a novel human rickettsiosis, namely Atlantic rainforest spotted fever, was described in Brazil. We herein report results of a survey led around the index case in an Atlantic rainforest reserve in Peruibe municipality, southeastern Brazil. A Rickettsia parkeri-like agent (Rickettsia sp. Atlantic rainforest genotype) and Ricketsia bellii were isolated from adult Amblyomma ovale ticks collected from dogs. Molecular evidence of infection with strain Atlantic rainforest was obtained for 30 (12·9%) of 232 A. ovale adult ticks collected from dogs. As many as 88·6% of the 35 examined dogs had anti-Rickettsia antibodies, with endpoint titres at their highest to R. parkeri. High correlation among antibody titres in dogs, A. ovale infestations, and access to rainforest was observed. Amblyomma ovale subadults were found predominantly on a rodent species (Euryoryzomys russatus). From 17 E. russatus tested, 6 (35·3%) displayed anti-Rickettsia antibodies, with endpoint titres highest to R. parkeri. It is concluded that Atlantic rainforest genotype circulates in this Atlantic rainforest area at relatively high levels. Dogs get infected when bitten by A. ovale ticks in the forest, and carry infected ticks to households. The role of E. russatus as an amplifier host of Rickettsia to A. ovale ticks deserves investigation.

Cryptosporidium is a protozoan parasite of humans and animals and has a worldwide distribution. The parasite has a unique epidemiology in Middle Eastern countries where the IId subtype family of Cryptosporidium parvum dominates. However, there has been no information on Cryptosporidium species in Yemen. Thus, this study was conducted in Yemen to examine the distribution of Cryptosporidium species and subtype families. Fecal samples were collected from 335 patients who attended hospitals in Sana'a city. Cryptosporidium species were determined by PCR and sequence analysis of the 18 s rRNA gene. Cryptosporidium parvum and C. hominis subtypes were identified based on sequence analysis of the 60 kDa glycoprotein (gp60) gene. Out of 335 samples, 33 (9·9%) were positive for Cryptosporidium. Of them, 97% were identified as C. parvum whilst 1 case (3%) was caused by C. hominis. All 7 C. parvum isolates subtyped belonged to the IIaA15G2R1 subtype. The common occurrence of the zoonotic IIa subtype family of C. parvum highlights the potential occurrence of zoonotic transmission of cryptosporidiosis in Yemen. However, this postulation needs confirmation with future molecular epidemiological studies of cryptosporidiosis in both humans and animals in Yemen.

There is an alarming rate of human African trypanosomiasis recrudescence in many parts of sub-Saharan Africa. Yet, the disease has no successful chemotherapy. Trypanosoma lacks the enzymatic machinery for the de novo synthesis of purine nucleotides, and is critically dependent on salvage mechanisms. Inosine 5′-monophosphate dehydrogenase (IMPDH) is responsible for the rate-limiting step in guanine nucleotide metabolism. Here, we characterize recombinant Trypanosoma brucei IMPDH (TbIMPDH) to investigate the enzymatic differences between TbIMPDH and host IMPDH. Size-exclusion chromatography and analytical ultracentrifugation sedimentation velocity experiments reveal that TbIMPDH forms a heptamer, different from type 1 and 2 mammalian tetrameric IMPDHs. Kinetic analysis reveals calculated Km values of 30 and 1300 μm for IMP and NAD, respectively. The obtained Km value of TbIMPDH for NAD is approximately 20–200-fold higher than that of mammalian enzymes and indicative of a different NAD binding mode between trypanosomal and mammalian IMPDHs. Inhibition studies show Ki values of 3·2 μm, 21 nM and 3·3 nM for ribavirin 5′-monophosphate, mycophenolic acid and mizoribine 5′-monophosphate, respectively. Our results show that TbIMPDH is different from its mammalian counterpart and thus may be a good target for further studies on anti-trypanosomal drugs.

The calcium-dependent protein kinases (CDPKs) are unique enzymes found only in plants, green algae, ciliates and apicomplexan parasites. In this study, a novel CDPK gene of Eimeria tenella, designed EtCDPK3, was cloned using rapid amplification of cDNA ends (RACE) based on the expressed sequence tag (EST). The entire cDNA of EtCDPK3 contained 1637 nucleotides encoding 433 amino acids and the deduced EtCDPK3 protein had canonical characteristic domains identified in other CDPKs, including a well-conserved amino-terminal kinase domain and a carboxy-terminal calmodulin-like structure with 4 EF-hand motifs for calcium binding. The expression profiles of the EtCDPK3 gene in different development stages were investigated by real-time quantitative PCR. Messenger RNA levels from the EtCDPK3 gene were higher in sporozoites than in other stages (unsporulated oocysts, sporulated oocysts and merozoites). Western blot analysis showed that rabbit antiserum against recombinant EtCDPK3 could recognize a native 49 kDa protein band of parasite. Indirect immunofluorescent antibody labelling revealed dispersed localization of EtCDPK3 during the first schizogony and intense specific staining. EtCDPK3 was located at the apical end of the sporozoites after early infection of DF-1 cells and the protein was highly expressed. Inhibition of EtCDPK3 function using specific antibodies reduced the ability of E. tenella to invade host cells. These results suggested that EtCDPK3 may be involved in invasion and survival of the parasite intracellular stages of E. tenella. Because this kinase family is absent from hosts, it represents a valid target that could be exploited for chemotherapy against Eimeria spp.

The life cycle of the parasitic copepod Lernaeocera branchialis involves 2 hosts, typically a pleuronectiform host upon which development of larvae and mating of adults occurs and a subsequent gadoid host, upon which the adult female feeds and reproduces. Both the copepodid and adult female stages must therefore locate and identify a suitable host to continue the life cycle. Several mechanisms are potentially involved in locating a host and ensuring its suitability for infection. These may include mechano-reception to detect host movement and chemo-reception to recognize host-associated chemical cues, or kairomones. The aim of this study was to identify the role of kairomones in host location by adult L. branchialis, by analysing their behaviour in response to fish-derived chemicals. Experiments demonstrated that water conditioned by immersion of whiting, Merlangius merlangus, elicited host-seeking behaviour in L. branchialis, whereas cod- (Gadus morhua) conditioned water did not. Lernaeocera branchialis are considered a genetically homogeneous population infecting a range of gadoids. However, their differential response to whiting- and cod-derived chemicals in this study suggests that either there are genetically determined subspecies of L. branchialis or there is some form of environmental pre-conditioning that allows the parasite to preferentially recognize the host species from which it originated.

The lack of an adequate model for Leishmania (Viannia) braziliensis infection is a limiting factor for studying American tegumentary leishmaniasis (ATL). The golden hamster (Mesocricetus auratus) is a promising model because besides being highly susceptible to dermotropic Leishmania infection, the lesions are very similar to cutaneous leishmaniasis (CL) in humans. However, different Leishmania isolates or species and/or protocols have resulted in different outcomes, whereas no study has evaluated the reproducibility of L. braziliensis infection in this model. The natural history of L. braziliensis infection in 34 hamsters was evaluated by using a single parasite isolate in 8 independent experiments under similar experimental conditions. Clinical, histological and immunological analyses were performed. The hamsters presented skin ulcers similar to those observed in ATL. The intra-experiment lesion increment tended to show an intermediary variance. Histological analysis of infected skins showed granulomatous reaction, scarce amastigotes, and Schaumann's bodies. Blood lymphocytes proliferated in response to leishmanial antigens. The severity of the infection was positively correlated to spleen weight, and the titres of anti-Leishmania IgG antibodies. Our findings indicate that the hamster is an appropriate model for immunopathogenesis studies of CL caused by L. braziliensis, supporting its use in clinical, vaccine and chemotherapy experimental protocols.

Refugia-based treatment strategies aim to prolong anthelmintic efficacy by maintaining a parasite population unexposed to anthelmintics. Targeted selective treatment (TST) achieves this by treating only animals that will benefit most from treatment, using a determinant criterion (DC). We developed a mathematical model to compare various traits proposed as DC, and investigate impacts of TST and drenching frequency on sheep performance and anthelmintic resistance. Short term, decreasing the proportion of animals drenched reduced benefits of anthelmintic treatment, assessed by empty body weight (EBW), but decreased the rate of anthelmintic resistance development; each consecutive drenching had a reduced impact on average EBW and an increased impact on the rate of anthelmintic resistance emergences. The optimal DC was fecal egg count, maintaining the highest average EBW when reducing the proportion of animals drenched. Long-term, reducing the proportion of animals drenched had little impact on total weight gain benefits, across animals and years, whilst reducing drenching frequency increased it. Decreasing the frequency and proportion of animals drenched were both predicted to increase the duration of anthelmintic efficacy but reduce the total number of drenches administered before resistance was observed. TST and frequency of drenching may lead to different benefits in the short versus long term.

The crayfish plague pathogen, Aphanomyces astaci, is one of the most serious threats to indigenous European crayfish species. The North American invasive spiny-cheek crayfish, Orconectes limosus, is an important source of this pathogen in central and western Europe. We evaluated potential changes in A. astaci spore release rate from infected individuals of this species by experiments investigating the pathogen transmission to susceptible noble crayfish, Astacus astacus. We filtered defined volumes of water regularly to quantify spore concentration, and sampled crayfish tissues at the end of the experiment. The filters and tissues were then tested for the presence of A. astaci DNA by species-specific quantitative PCR. Additionally, we tested the efficiency of horizontal transmission to apparently uninfected O. limosus. The experiments confirmed that A. astaci can be transmitted to susceptible crayfish during intermoult periods, and that the pathogen was more frequently detected in noble crayfish recipients than in American ones. The pathogen spore concentrations substantially varied in time, and significantly increased during moulting of infected hosts. Our study strengthens the evidence that although the likelihood of crayfish plague transmission by water transfer from localities with infected American crayfish might increase when these are moulting or dying, no time-periods can be proclaimed safe.