Partial mitochondrial cytochrome c oxidase subunit I (mt COI) sequences were generated from: Toxoplasma gondii (strains CTG, GTI, MAS, ME49, PTG, TgCatBr5, TgCat, Br64, TgCgCal, TgToucan); Neospora caninum (Strain NC1); Hammondia hammondi (Strain H.H–20); H. heydorni; H. cf. triffittae; Cystoisospora felis; C. suis; C. canis; C. rivolta; C. cf. ohioensis; Caryospora bigenetica; Sarcocystis rileyi; and S. neurona. Nuclear 18S rDNA sequences were generated for H. heydorni, H. hammondi, C. suis, C. canis, C. felis, C. rivolta, C. cf. ohioensis, S. neurona, and S. rileyi. Aligned, concatenated 18S rDNA and COI sequences were Bayesian analysed using partitioned nucleotide substitution models [HKY + I + G for 18S; GTR + I + G codon (code = metmt) for COI]. Phylogenetic hypotheses supported a monophyletic Sarcocystidae and its subfamilie with two major clades within the Toxoplasmatinae: (1) a monophyletic clade of Cystoisospora spp. with Nephroisospora eptesici; and (2) a clade of Toxoplasma, Neospora and Hammondia. Within the latter, Hammondia was shown to be paraphyletic; H. heydorni and H. triffittae were monophyletic with N. caninum [canine definitive hosts (DHs)], whereas H. hammondi was monophyletic with T. gondii (feline DHs). A new genus is erected to resolve the paraphyly of the genus Hammondia confirmed using mt COI and combined 18S/COI sequence datasets.

The protozoan parasite Leishmania is endemic in large parts of the world which causes leishmaniasis. Its visceral form is fatal if not treated and is caused mostly by Leishmania donovani, Leishmania infantum and Leishmania chagasi. Given the difficulties linked to vector (sandfly) control and the lack of an effective vaccine, the control of leishmaniasis relies mostly on chemotherapy. Unfortunately, the prevalence of parasites becoming resistant to the first-line drug pentavalent antimony (SbV) is increasing worldwide. Few alternative drugs are available that includes amphotericin B, pentamidine and miltefosine (oral). Already, decreases in efficacy, resistance and toxicity have been noted against these drugs. Dry antileishmanial pipeline further indicates the slow pace of drug discovery in this field where resistance as a major barrier. Full understanding of the genetic and molecular basis of the parasite is lagging. Since leishmaniasis is a neglected disease and occurs predominantly in the developing world largely, therefore, it is unaddressed. The pharma industry argues that development of the new drug is too costly and risky to invest in low return neglected diseases is very high. Research is also needed to identify new and effective drug targets. The lack of drug research and development for neglected diseases will require some new strategies. We have discussed the various cause of slow pace of antileishmanial drug discovery in this review to pay attention of researchers and also take the public and private initiative to make the process fast for new antileishmanial drug development.

Leishmania virulence and disease development critically depends on the ability of Leishmania promastigotes to infect, differentiate into amastigote forms and replicate inside mammalian host macrophages. Understanding changes associated with amastigote differentiation in axenic culture conditions is a key to identifying virulence factors. Here we compared efficiency of the conventional pH–temperature-dependent shift method to induce amastigote differentiation with the recently identified trigger for differentiation mediated by mitochondrial reactive oxygen species (ROS). Using two different visceral leishmaniasis species, L. infantum and. L. donovani, we show that ROS-generating methods such as iron deprivation or exposure to sub-lethal concentrations of hydrogen peroxide (H2O2) or menadione are significantly more effective in promoting promastigote–amastigote differentiation than the low pH–high temperature shift, leading to higher survival rates, morphological changes and gene expression patterns characteristic of the amastigote stage. Notably, both H2O2 and menadione-mediated differentiation did not require upregulation of the mitochondrial electron transport chain-associated protein p27, suggesting that treatment with oxidants bypasses the necessity to upregulate mitochondrial activity, a precondition for mROS generation. Our findings confirm that ROS-induced differentiation occurs in multiple Leishmania species, including the medically important visceralizing species, and provide mechanistic rationale for earlier reports demonstrating markedly increased virulence of L. infantum promastigotes pre-treated with oxidative reagents.

Tick-borne apicomplexan haemoparasites infect wild and domestic animals, but studies on their distribution among free-living animals are comparatively fewer. Tissues from 241 wild carnivores of eight Mustelidae, two Canidae, one Viverridae and one Felidae species were collected in Northern Spain, and analysed by real-time PCR targeting the 18S rRNA gene and sequencing. Babesia vulpes (formerly known as Theileria annae) was the only piroplasm detected in red foxes (Vulpes vulpes). Badgers (Meles meles) were shown to harbour two novel Babesia sp. sequence types (A and B) that only shared 96.7% homology between them and were closely related (ca. 97–98%) to, but distinct from B. vulpes and other babesia from carnivores. Analysis of PCR-derived sequences also revealed the presence of Cystoisospora sp. and Hepatozoon sp. in badgers and showed that wild cats (Felis silvestris catus) were infected with Cytauxzoon sp. Forty-two per cent of the animals subjected to a detailed external examination were parasitized by ixodid ticks, being Ixodes hexagonus and Ixodes ricinus the most abundant species. This study provided novel data on the different haemoparasites that can infect European wild carnivores and showed that they can be hosts for a range of haemoparasites and pose a potential risk for transmission to domestic animals.

The Nigerian national control programme (NCP) for schistosomiasis does not have a specific action plan for female genital schistosomiasis (FGS), mainly due to gaps in epidemiological and clinical surveillance. To address this, we conducted a pilot investigation for FGS in girls (>5 and ≤15 years) and adult women (≥16 years) in four rural communities near Abeokuta, Ogun State. Urine samples were collected from 317 participants [children (n = 187), adults (n = 130)] and examined for ova of Schistosoma haematobium by microscopy. A total of 149 participants (47·0%) had egg-patent urogenital schistosomiasis [children (64·7%), adults (21·6%)], reported blood in urine was common (44·5%) as well as sign-post symptoms of FGS. To verify FGS directly, 20 adult women each underwent a gynaecological investigation by colposcopy with observed lesions classified according to the WHO FGS pocket atlas. Fourteen (70·0%) women presented with FGS as grainy-sandy patches (n = 10), homogenous yellow sandy patches (n = 6), nabothian cysts and rubbery papules (n = 1). Our study confirms FGS in Ogun State and calls for further appraisals of this disease in other areas where urogenital schistosomiasis is endemic. The Nigerian NCP should be encouraged to develop an appropriate response to FGS not only to detect it, but also to prevent it.

Cryptosporidium is a well-known cause of neonatal enteritis in cattle worldwide. Cattle are commonly infected with four different species of Cryptosporidium but only one of these, Cryptosporidium parvum, is associated with clinical disease. Identification of species in cases of calf scour can give an indication if Cryptosporidium is the causative agent or not. In addition, C. parvum is a zoonotic species and so has implications for human health, for this reason it is important to identify the species of Cryptosporidium infecting cattle particularly where a farm is implicated in an outbreak of cryptosporidiosis in humans. Here a multiplex PCR test, which can identify the four common cattle-adapted Cryptosporidium species, including C. parvum, has been developed. This test allows quick and accurate detection of Cryptosporidium species in cattle fecal samples including mixed infections, which could be missed by the more common method of sequencing the same gene.

The current mainstay for control of the four major helminth diseases in humans (lymphatic filariasis, onchocerciasis, soil-transmitted helminthiases and schistosomiasis) is with preventive chemotherapy by mass administration of key anthelminthics. Following the London Declaration on Neglected Tropical Diseases in 2012, a roadmap for the elimination and control of these helminthiases by 2020 has been devised. With expected declines in prevalence and intensity of these infections, there is urgent need for implementing more sensitive, high-throughput and cost-effective diagnostic tools. Currently available diagnostic approaches for surveying, monitoring and evaluating helminth control programmes are based on microscopical observation of eggs/larvae, and/or detection of antibodies or parasite antigens in stool, urine or blood; all relatively low-throughput and of limited sensitivity and specificity. Newly proposed approaches for helminthiases diagnosis include the nucleic acid-based methods of (multiplex) real-time polymerase chain reaction assays, loop-mediated isothermal amplification and recombinase polymerase amplification. However, as well as sensitivity/specificity evaluation, their comparison to current ‘gold standard’ diagnostics and future application in individual-/community-based diagnosis, or in xenomonitoring requires consideration of relative costs, agreement of standard methods and strategic interpretation of resulting data before control/elimination programmes might best utilize molecular diagnostics to inform decision making. We review current nucleic-acid-based molecular diagnostic methods and highlight the needs and future research required to refine these tools for monitoring and evaluation of control and elimination programmes for four major human helminthiases.

Sixteen intestinal helminth species were recovered from 271 necropsied stray dogs during a survey undertaken in Raoued and Soukra, two northeastern rural regions of Tunisia. Recorded parasites included trematodes (Brachylaemus sp., Phagicola italica, Heterophyes heterophyes), cestodes (Echinococcus granulosus, Dipylidium caninum, Diplopylidium noelleri, Mesocestoides lineatus, Mesocestoides litteratus, Taenia hydatigena, Taenia pisiformis, Taenia multiceps), nematodes (Toxocara canis, Ancylostoma caninum, Uncinaria stenocephala, Trichuris vulpis) and one acanthocephalan Macracanthorhynchus hirudinaceus. This is the first record of Brachylaemus sp., P. italica, H. heterophyes, D. noelleri and M. hirudinaceus in dogs from Tunisia. Echinococcus granulosus was found in 5·16% of dogs with a higher intensity of infection recorded in younger animals (303 worms/infected dog). Molecular analysis confirmed E. granulosus sensu stricto as the cause of canine echinococcosis. This epidemiological study investigating the status of intestinal helminths of dogs recorded a prevalence of 98·89% and a mean intensity of 87·62 worms per infected dog and confirmed the infection of 95·14% of dogs with helminths of potential zoonotic concern. Results of this study emphasize the need to interrupt parasite transmission using preventive approaches in zoonoses control programmes mainly against cystic echinococcosis, as well as reducing transmission to other animals by regular dog dosing treatments and proper management of dog populations.

The African buffalo (Syncerus caffer), a mammal species whose population is declining, can play a role as a reservoir or carrier of a wide number of arthropod-borne pathogens. Translocation procedures have been used as an alternative approach for species conservation. However, the veterinary aspects of this sort of procedures are extremely important to minimize the impact on animal health. In order to detect Bartonella and haemoplasmas, two important group of bacterial that have an impact in both human and animal health, EDTA whole-blood samples were screened for the presence of these bacterial pathogens by molecular techniques. As a result, a molecular occurrence of 4.1 and 15.4% for Bartonella spp. and haemoplasmas, respectively, was reported among 97 wild buffaloes sampled during a translocation procedure from Marromeu to Gorongosa Reserve, Mozambique. Additionally, phylogenetic analyses of the obtained sequences were conducted. At least, three bovine-associated pathogens, namely B. bovis, M. wenyonii and ‘Candidatus M. haemobos’, as well as a probably new Bartonella genotype/species were detected in S. caffer. Further studies are needed in order to determine whether these bacterial species may cause impact in buffaloes and other sympatric ruminant species living in the release site.

A cross-sectional survey was carried out in primary schools to determine prevalence, intensity and spatial co-distribution of Schistosomiasis and soil transmitted helminths (STH) infections in Ogun State, Nigeria. A total of 2148 pupils from 42 schools were examined for Schistosoma and STH infections from urine and fresh fecal samples respectively. Ethyl ether concentration method prepared in sodium acetate – acetic acid – formalin ether was used to concentrate parasites’ ova before microscopic examination. The overall prevalence of schistosomiasis and STH infections were 4.0% (95% CI = 3.21–4.92) and 34.64% (95% CI = 32.62–36.69) respectively. Schistosoma haematobium and Ascaris lumbricoides were the most prevalent across the study area among the Schistosoma and STH species respectively. Overall, intensity of infection was higher in males than in females for all Schistosoma and STH infections, but with no significant difference (P > 0.05), except for Trichuris trichiura (χ2 = 6.490, P < 0.05). Infection intensity was significantly inversely correlated (χ2 = 12.953, P < 0.05) with an increase in age group. Co-distribution of Schistosoma and STH infections occurred in 15 (35.7%) out of 42 schools, and only 30 children (1.4%) had co-infection of Schistosoma and STH. This study provides information on the prevalence and spatial risk of schistosomiasis and STH in Ogun State. This will serve as decision-support tool for Ogun State programme managers to help facilitate integration of schistosomiasis and STH control.

In the present study, the potential activity of two 5-nitroindazole derivatives previously proposed as suitable antichagasic prototypes was further evaluated on diverse Trypanosoma cruzi strains belonging to two discrete typing units (DTUs) frequently associated with human infection (i.e. DTUs TcII and TcVI). The trypanocidal profile that both 2-benzyl-1-propyl (22) and 2-benzyl-1-butyl (24) derivatives achieved on Tulahuen amastigotes (IC50 = 3·56 ± 0·99 and 6·31 ± 1·04 µ m, respectively) correlates with that of formerly obtained on CL Brener, corroborating an outstanding activity on DTU TcVI parasites. Moreover, a sequential screening on extracellular and intracellular stages of T. cruzi Y (DTU TcII) demonstrated also the effectiveness of 22 and 24 over this strain on a similar range of activity (IC50 epimastigotes = 3·55 ± 0·47 and 7·92 ± 1·63 µ m, IC50 amastigotes = 2·80 ± 0·46 and 9·02 ± 5·26 µ m, respectively). These results, supported by a lack of toxicity registered over either L929 fibroblasts or primary cultures of cardiomyocytes, confirm that 5-nitroindazolinones 22 and 24 display great selectivity on both drug-sensitive (CL and Tulahuen) and drug-moderately resistant (Y) T. cruzi strains, and therefore, represent an important outcome in the research of Chagas disease chemotherapy.

Feline leishmaniasis has been reported in various parts of the world in recent years, occurring mainly in areas where zoonotic visceral leishmaniasis (VL) is endemic. The purpose of this study was to assess the occurrence of natural infection by Leishmania spp. in domestic cats (Felis catus) in the municipality of Teresina, Piauí state, Brazil, an endemic area of VL in Brazil. The prevalence of infection by Leishmania spp. in the population under study was 4% (3/83) in the enzyme-linked immunosorbent assay (ELISA) and 4% (3/83) by smear observation and isolation in a culture medium, using popliteal lymphnode sample. Only one of the three infected cats was positive for ELISA, also being positive for feline immunodeficiency virus. In the haematologic parameters, normocytic normochromic anaemia was the most common change, being present in the three infected animals. In the biochemical measurements also were observed alterations in infected animals. The Leishmania spp. strains isolated from the culture medium were characterized as L. infantum. The presence of L. infantum infection in cats in the city of Teresina, an area endemic for VL, suggests the possible involvement of these animals in the epidemiological chain of L. infantum in high-transmission areas.

Blastocystis sp. is a protozoan commonly found in human and animal stool samples. Several pathogenic and zoonotic aspects of this organism are still unknown. The aim of the present study was to investigate Blastocystis subtypes (STs) in samples from patients of the Hospital das Clínicas of the Faculdade de Medicina at the Universidade de São Paulo (HC-FMUSP), Brazil. Blastocystis sp.-positive stool samples diagnosed at the Section of Parasitology of the Central Laboratory (HC-FMUSP) were used for DNA isolation. Polymerase chain reaction (PCR) was performed using specific primers targeting the small-subunit rRNA gene. Direct DNA sequencing of the PCR products was performed and the DNA sequences were then aligned and compared with other sequences obtained from the GenBank database. Phylogenetic analysis was used to identify STs and determine the phylogenetic relationships between the sequences. Four STs were identified: ST1 (22·5%), ST2 (12·5%), ST3 (60%) and ST6 (5%). In conclusion, ST3 was the most prevalent ST among the human isolates followed by ST1. The present study is one of the few providing STs data from the human population in South America. Determining ST prevalence in human samples may contribute to the monitoring of Blastocystis sp. infection transmission in endemic regions.

Host manipulation is a common strategy of parasites employed to increase their fitness by changing the phenotype of their hosts. Whether host manipulation might be affected by environmental factors such as resource availability, has received little attention. We experimentally infected laboratory-bred copepods with the cestode Schistocephalus solidus, submitted infected and uninfected copepods to either a high or a low food treatment, and measured their behaviour. Infection reduced host activity and speed in both feeding treatments. However, the difference between the infected and uninfected copepods was smaller under low food conditions, because uninfected, but not infected, copepods moved slower under these conditions. We suggest that these differences are mediated by the physical condition of copepods rather than changes in how strongly the parasite manipulated host behaviour. Additionally, we measured three fitness-relevant traits (growth, development and infection rate in the next host) of the parasite to identify potential trade-offs with host manipulation. The largest parasites in copepods appeared the least manipulative, i.e. their hosts showed the smallest behavioural alterations, but this may again reflect variation in copepod condition, rather than life history trade-offs between parasite growth and host manipulation. Our results point to the possibility that parasite transmission depends on environmental conditions.

Visceral leishmaniasis is an important neglected parasitic disease that is generally caused by Leishmania infantum, Leishmania donovani and Leishmania chagasi. However, several causative species of cutaneous leishmaniasis (CL) causes an interstitial form of leishmaniasis which known viscerotropic leishmaniasis. The aim of this paper is a systematic review of the cases of viscerotropic leishmaniasis to present the main causative agents, clinical manifestations, treatment and outcomes of the cases. An electronic search (any date to August 2017) without language restrictions was performed using Medline, PubMed, Scopus and Google Scholar. The searches identified 19 articles with total 30 case reports. Of them, old world Leishmania species was reported from 23 (76.7%) cases, including 20 cases of L. tropica and three cases of L. major; whereas new world Leishmania species were reported in seven (23.4%) cases. The infection was more prevalent in male (24/30, 80%) than female (5/30, 16.7%) patients. Co-morbidity/co-infection was observed in 13 out of 30 cases (43.4%), which the most of them was HIV/AIDS (10 out of 13 cases, 76.9%). The results suggested that viscerotropic leishmaniasis should be more attended in the endemic countries of CL and in immunocompromised patients in order to exact discrimination from other endemic infectious diseases.

The main goal of this work consisted in cloning, purifying and characterizing a protein phosphatase 2C (PP2C) from promastigotes of Leishmania major. The gene was cloned and amplified by PCR using specific oligonucleotides and the recombinant protein was purified by affinity chromatography. The peak with maximal protein concentration was analysed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and revealed a protein of 44·9 kDa with PP2C activity. This activity was dependent on divalent cations (Mg+2 and Mn+2) and was optimal at pH of 8·5, using phosphothreonine as the substrate. Sanguinarine inhibited the activity of the recombinant LmPP2C, while protein tyrosine phosphatase inhibitors had no effect. The recombinant LmPP2C was used to generate polyclonal antibodies. These antibodies recognized a protein of 44·9 kDa in different Leishmania species; the LmPP2C was localized in the flagellar pocket and the flagellum of promastigotes.

Oxyspirura petrowi is a heteroxenous parasitic nematode that has been reported in high prevalences from birds in the Order Galliformes experiencing population declines in the USA. There is a paucity of information regarding the natural history O. petrowi, including the life cycle and effects of infection on wild bird populations. In order to study the life cycle of this parasite, we collected plains lubber grasshoppers (Brachystola magna) from a field location in Mitchell County, Texas. We found third-stage larvae (L3) in 37.9% (66/174) B. magna. We determined that they were O. petrowi through morphological comparison of L3 from experimentally infected Acheta domesticus and by sequence analysis. Then, we showed that B. magna are a potential intermediate hosts for O. petrowi infections in northern bobwhites (Colinus virginianus) in a laboratory setting by experimental infection. We first detected shedding of eggs in feces using a fecal float technique 52 days post infection. In addition, we recovered 87 O. petrowi from experimentally infected northern bobwhites. Although we detected shedding in feces, recovery of eggs was low (>5 eggs/g). Future work is needed to understand shedding routes and shedding patterns of northern bobwhites infected with O. petrowi.

Populations of blue-footed boobies in the Galápagos Islands have declined by at least 50% compared with numbers recorded during the 1960s. Recently, concern has been expressed about potential adverse effects of introduced blood parasites; in particular, malarial parasites (i.e. Plasmodium sp.), which are frequently pathogenic and can cause mortality and morbidity in birds, and blood parasites of the genus Haemoproteus, which can reduce fitness. We screened blue-footed boobies at six breeding colonies within Galápagos for both genera of parasite. At two sites, we also investigated whether there was any relationship between infection status and physiological condition or breeding success. We found a high prevalence of Haemoproteus spp. in adults and chicks using PCR, although none was found on blood smears. We found no evidence of Plasmodium. Effects of parasitism on the condition of birds were limited. Also, there was no evidence that infection with Haemoproteus adversely affected breeding success or chick growth. Our results suggest that poor success of breeding birds was not strongly associated with Haemoproteus infection. However, regular colony monitoring and examination of dead birds are necessary to assess whether acute Haemoproteus infection might be responsible for some mortality, particularly of chicks.

On 9 November 2015, a juvenile male fin whale of 11·60 m length was observed on the bulb of a merchant vessel in the Channel Terneuzen – Ghent (The Netherlands – Belgium). A severe parasitosis was present in the right heart ventricle and caudal caval vein. Parasites were identified as Crassicauda boopis based on macroscopic and microscopic observations. The sequence of the 18S rRNA gene obtained from the parasite samples was 100% similar to the sequence of the 18S rRNA gene from Crassicauda magna available on GenBank. While adults of C. boopis and C. magna are morphologically distinct and found at different locations in the body, the molecular analysis of the 18S rRNA gene seems insufficient for reliable species identification. Although numerous C. boopis were found, the cause of death was identified as due to the collision with the ship, as suggested by the presence of a large haematoma, and the absence of evidence of renal failure. The young age of this whale and the absence of severe chronic reaction may suggest that the infestation was not yet at an advanced chronic stage.

Haemosporidian parasites are widespread in avian species and modulate their ecology, behaviour, life-history and fitness. The prevalence of these vector-transmitted parasites varies with host intrinsic and extrinsic factors, such as host resistance and behaviour, and habitat-related characteristics. In this study, we evaluated the prevalence of avian haemosporidians in great tit Parus major populations inhabiting two areas with different degrees of exposure to aerial emissions from pulp mills, to assess if this type of pollution impacted parasite prevalence. We also compared the physiological condition of infected and uninfected individuals. Haemosporidian infection prevalence was high (58%), varied seasonally, but was not associated with air pollution exposure. Fledged birds presented higher infection rates than nestlings and infected fledged birds had higher levels of blood glutathione peroxidase activity. These results allow us to infer that infection by blood parasites may activate antioxidant defences, possibly to protect the organism from the negative oxidative stress side-effects of immune activation against parasites. Because oxidative stress is one of the mechanisms responsible for ageing and senescence and may affect fitness, the relationship between parasitism and oxidative stress markers should be further investigated through studies that include experimental manipulation of infection in model organisms.