Subconjunctival human dirofilariasis by Dirofilaria repens in the Mediterranean Basin.

Purpose: Human dirofilariasis cases have increased in Europe for the last few decades. We document an autochthonous case of dirofilariasis infection in the subconjunctival space of a 60-year-old Spanish man. Observations: The patient presented with pain in his right eye, which was diagnosed as epi-scleritis. In spite of the treatment administered, no improvement was reported. An external exam showed conjunctival congestion on the temporal part of the right eye bulbar conjunctiva. The anterior chamber showed no flare or cells in both eyes and dilated fundus exam was normal. Nevertheless, a long whitish vermiform mobile mass was detected under the conjunctiva in the temporal part of the right eye. The worm was surgically removed and diagnosed morphologically and also molecularly as Dirofilaria repens. This species uses dogs and wild canids as definitive hosts and principal reservoirs, while mosquito species are transmitters. Humans act as accidental hosts, and clinical manifestations depend on the location of the worm in the organs and tissues. Conclusions and importance: The correct diagnosis and control of subcutaneous/ocular dirofilariasis by D. repens require several professionals under the One Health approach to deal with this zoonotic disease, which poses a serious public health problem, at least in the Mediterranean Basin.

Soil-transmitted Helminth Infections in Schoolchildren of Laguna de Perlas (Nicaragua).

Soil-transmitted helminth (STH) prevalence, intensity, polyparasitism and co-infections in 425 children from 3 schools of Laguna de Perlas (Nicaragua) were investigated. Single stool samples were analysed by the formalin-ether method and the Kato-Katz. A total of 402 (94.6%) children were infected. Trichuris trichiura 308 (72.4%), Ascaris lumbricoides 115 (27.1%) and Hookworms 54 (12.7%) were the most prevalent STHs. Polyparasitism (322; 75.8%) with two species was most prevalent (109; 25.6%). T. trichiura with A. lumbricoides (19.3%) and T. trichiura with Hookworm (6.8%) were the most common combinations. Positive associations were observed between T. trichiura and A. lumbricoides and T. trichiura and Hookworm (p = 0.0001). Highest Hookworm intensities appeared when three STH co-infections occurred. Moderate- heavy STH intensities appear in up to 42.1% in trichuriasis, 57.5% in ascariasis and 11.1% in Hookworm infections. Integrated control interventions covering children need to be implemented mainly in a rural environment.

An update on human echinostomiasis.

Echinostomiasis, caused by trematodes belonging to the family Echinostomatidae, is an important intestinal foodborne parasitic disease. Humans become infected after ingestion of raw or insufficiently cooked molluscs, fish, crustaceans and amphibians, thus, understanding eating habits is essential to determine the distribution of the disease. Despite the public health impact of echinostomiasis, it has been neglected for years. Traditionally, echinostomiasis has been considered as a minor disease confined to low-income areas, mainly in Asia. However, the geographical boundaries and the population at risk are currently expanding and changing in relation to factors such as growing international markets, improved transportation systems, new eating habits in developed countries and demographic changes. These factors make it necessary to improve our understanding of intestinal trematode infections. Herein, we review the main features of human echinostomiasis in relation to their biology, epidemiology, host-parasite relationships, pathogenicity, clinical aspects, diagnosis, treatment and control.

Partial resistance to homologous challenge infections of the digenean Echinostoma caproni in ICR mice.

In the present paper, we analyse the effect of a primary infection of ICR mice with Echinostoma caproni (Trematoda: Echinostomatidae) on the generation of resistance against homologous challenge infections. In ICR mice, E. caproni induces chronic infections concomitantly with strong responses characterized by the development of T-helper 1 (Th1)-type local immune responses with elevated levels of local interferon-gamma (IFN-γ) and inflammatory and antibody responses. Here, the effect of the response generated against a primary infection with E. caproni in the generation of resistance against subsequent homologous infections was analysed. For this purpose, ICR mice were challenged with metacercariae of E. caproni and the results obtained showed that primary infection induces partial resistance against subsequent homologous infections in ICR mice. This resistance was expressed as a reduced rate of infection, worm recovery and worm size, indicating that primary infection induces changes in the host, making a hostile environment for the development of the parasite.

Echinostoma caproni (Trematoda): differential in vivo mucin expression and glycosylation in high- and low-compatible hosts.

Enhanced mucus production and release appears to be a common mechanism for the clearance of intestinal helminths, and this expulsion is normally mediated by Th2-type immune responses. To investigate the factors determining the expulsion of intestinal helminths, we have analysed in vivo expression of mucin genes at the site of infection in two host species displaying different compatibility with Echinostoma caproni (Trematoda). Surprisingly, a general down-regulation on mucin mRNA expression was detected in low-compatible hosts (rats) coinciding with the development of Th2/Th17 responses and the early rejection of the worms from the intestinal lumen. This suggests the existence of a mechanism by which the parasites can modulate the mucus barrier to favour their survival. In highly compatible hosts (mice), some mucin genes were found to be up-regulated throughout the infection, probably, to protect the intestinal epithelium against the infection-induced inflammation developed in this host species. Moreover, infection-induced changes on mucin glycans were also studied by lectin histochemistry. Similar alterations were detected in the ileum of infected mice and rats, except with SNA lectin, indicating that sylated mucins might play an important role in determining the evolution of the infection in each host species.

Haematological changes in the laboratory rat Rattus norvegicus infected with Echinostoma caproni (Trematoda: Echinostomatidae).

To study possible indirect effects of the infection with intestinal helminths, 12 Rattus norvegicus (Wistar) were each experimentally exposed to 100 metacercariae of Echinostoma caproni, and blood samples were taken weekly up to 4 weeks post-exposure for comparison with control rats. Values of haematocrit (HCT), red blood cells (RBC), platelets (PLT), white blood cells (WBC), haemoglobin (HGB) and haematimatrix indices, and mean corpuscular haemoglobin concentrations (MCHC) were determined. In addition, leucocyte counts, including lymphocytes, neutrophils, monocytes, eosinophils and basophils were analysed. These parameters, including the leucocyte counts, showed no significant differences, except for MCHC at 4 weeks post-exposure. The present results indicate that in rats infected with E. caproni, although eosinophilia did not significantly increase, a significant reduction in MCHC was associated with an increase in the number of RBC.

Prevalence and risk factors related to intestinal parasites among children in Department of Rio San Juan, Nicaragua.

BACKGROUND: The present study was carried out to assess the prevalence of intestinal parasite infections among children in Department of Rio San Juan (Nicaragua), to explore the extent of polyparasitism, and to identify the risk factors that might favour transmission of these parasites. METHODS: A coprological study of single stool specimen, collected at random from 382 children (167 boys and 215 girls) aged 2-15 years from two municipalities, and a questionnaire, concerning demographic and epidemiological data on sanitary and behavioural habits, was filled in to identify risk factors. RESULTS: About 93% of the children presented infection. Blastocystis hominis, the most frequently protozoa, and Trichuris trichiura, Ascaris lumbricoides and hookworm, the most prevalent helminths. Polyparasitism (85.4%[326/382]), with the highest percentages detected in females (p=0.001), in the 6-11 year age group (p<0.001) and having a rural background (p<0.001). A firm relationship between any parasite and age (OR=1.2, p=0.036) and rural background (OR=0.3, p=0.027) was detected. Living in rural areas and drinking river water were also positively associated with each of the STH species detected, and walking barefoot was also associated with hookworm infection. CONCLUSIONS: Government efforts should be focused on controlling the risk factors associated with these enteroparasites, with health education programmes in rural areas of Nicaragua.

Human infection by a "fish tapeworm", Diphyllobothrium latum, in a non-endemic country.

We document a case of locally acquired "broad- or fish-tapeworm" infection caused by Diphyllobothrium latum in a 27-year-old Spanish man, confirmed by molecular analysis (COI gene). The patient had naturally expelled a worm of 110 cm in length, but the physical examination did not yield any remarkable findings, and the patient did not suffer from any particular symptoms. Laboratory test results were normal except for a remarkable increase in the red blood cell count, and an evident decrease in the mean corpuscular volume and mean corpuscular hemoglobin. Vitamin B12 and folic acid values were in the normal range without signs of anemia. It was suggested that these anomalies in erythrocyte formation might not be related to the parasite, and analysis of the patient's anamnestic data revealed that the infection could only have been caused by the ingestion of imported fish, although no light could be shed on the specific source of infection. From a public health viewpoint, this human case of fish-borne zoonosis is exemplary, suggesting that not only is control of fish and fish product quality essential, but also increased awareness of the general population with regard to changes in culinary habits.

The influence of adult worm age and definitive host on the transmission of Echinostoma caproni: egg hatchability and miracidial infectivity.

We analysed the influence of different ages of Echinostoma caproni adult worms reared in different definitive host-species on egg hatchability and miracidial infectivity. Eggs obtained from 2-, 4- and 6-week-old adult worms collected from rats, hamsters and mice, hatched miracidia that were used to infect laboratory-reared specimens of Biomphalaria glabrata. Highest egg hatchability was obtained with eggs derived from hamsters and mice. Egg hatchability did not seem to be affected by the age of E. caproni adult worms. Miracidial infectivity was not affected by the definitive host species. However, miracidial infectivity was affected by the age of adult worms when they developed in the mice intestine. The long period of miracidial infectivity in hamsters revealed this definitive host as the most suitable for experimental development of the life cycle and parasite transmission.

Current status of food-borne trematode infections.

Food-borne trematodiases constitute an important group of the most neglected tropical diseases, not only in terms of research funding, but also in the public media. The Trematoda class contains a great number of species that infect humans and are recognized as the causative agents of disease. The biological cycle, geographical distribution, and epidemiology of most of these trematode species have been well characterized. Traditionally, these infections were limited, for the most part, in populations living in low-income countries, particularly in Southeast Asia, and were associated with poverty. However, the geographical limits and the population at risk are currently expanding and changing in relation to factors such as growing international markets, improved transportation systems, and demographic changes. The diagnosis of these diseases is based on parasitological techniques and only a limited number of drugs are currently available for treatment, most of which are unspecific. Therefore, in-depth studies are urgently needed in order to clarify the current epidemiology of these helminth infections and to identify new and specific targets for both effective diagnosis and treatment. In this review, we describe the biology, medical and epidemiological features, and current treatment and diagnostic tools of the main groups of flukes and the corresponding diseases.

Th17 responses in Echinostoma caproni infections in hosts of high and low compatibility.

In order to investigate the factors determining the expulsion of intestinal helminths, we have analyzed the in vivo expression of IL-17, TGF-ß and IL-23 in several tissues of two host species displaying different compatibility with Echinostoma caproni (Trematoda). We did not observe upregulation of these cytokines in any of the tissues of the high compatible host (mice). In contrast, the responses in the host of low compatibility (rats) with the parasite were markedly different. Significant increases in the expression of IL-17 and TGF-ß were observed in the Peyer's patches and the intestine from the 2 to 8 weeks post-infection. The expression of IL-23 was upregulated from 2 to 4 weeks post-infection in the spleen, Peyer's patches and the intestine. Considering together our results with those published previously the development of chronic infections appears to be related with the development of local Th1 responses, whereas the early rejection of the worms is mediated by the development a biased Th17/Th2 phenotype. The Th17 response generated in rats may facilitate the worm expulsion via the suppression of the inflammatory Th1 responses and the increase in intestinal contractility.

Prevalence and molecular characterization of Cryptosporidium in schoolchildren from department of Rio San Juan (Nicaragua).

A cross-sectional study of cryptosporidiosis was carried out in Nicaragua. The prevalence of Cryptosporidium infection was determined in 272 (110 boys and 162 girls) schoolchildren, aged between 4 to 15 years from department of Rio San Juan. The total percentage obtained for Cryptosporidium (35.7%) was one of the highest reported so far. Cryptosporidium appeared in 94.8% of multiparasitism cases. No significant statistical differences were detected in the cryptosporidiosis prevalence between loose/watery (22.2%) and soft/formed (36.7%) stool samples. No significant age and sex differences were observed. This is the first report to identify Cryptosporidium in Nicaragua at species level, providing a preliminary molecular characterization of all positive samples, such as Cryptosporidium parvum (genotype 2). The high prevalence of C. parvum suggests that animals may be potential sources of infection for human cryptosporidiosis, although C. parvum infections may have originated from humans themselves. The human health problem caused by Cryptosporidium in this region may be related to the poor human hygiene/sanitation and contamination of the environment, food, or water supplies. Continuous exposure to the parasite could have been protective against development of symptoms in the children examined.

Helminth parasites in faecal samples from the endangered Iberian lynx (Lynx pardinus).

The Iberian lynx is the most endangered felid in the world. Enteropathogens may threaten its survival, and therefore we analysed faecal samples from 66 different individuals (37 males and 29 females), the largest population representation studied to date. The samples were obtained from November 2005 to October 2008 in the two areas where the Iberian lynx survives: Sierra Morena and Doñana (Andalusia, southern Spain). A total of 56.1% samples were parasitized with at least 6 species of helminths, including two cestodes (Hymenolepis spp. and Taenia spp.) and four Nematodes (Ancylostoma spp., Toxocara spp., Toxascaris leonina, and Capillaria sp.). In this work, the presence of Hymenolepis is reported for the first time in Lynx pardinus. The relevance of our findings is discussed focussed on the conservation of this endangered felid.

Proteomic analysis of Strongyloides stercoralis L3 larvae.

Strongyloidiasis can be perpetuated by autoinfection with the filariform larvae L3, causing asymptomatic chronic infections and creating a population of carriers, affecting not only developing countries. So far, very little is known about the proteins that interact with the human host, and few proteins from the infective Strongyloides stercoralis L3 have been characterized. Here, we report results obtained from a proteomic analysis of the proteins from S. stercoralis L3 larvae obtained from patients. Since the genome of S. stercoralis is not yet available, we used proteomic analysis to identify 26 different proteins, 13 of them released by short digestion with trypsin, which could represent surface-associated proteins. The present work extends our knowledge of host-parasite interactions by identifying proteins that could be of interest in the development of diagnostic tools, vaccines, or treatments for a neglected disease like strongyloidiasis.

Sympatric and allopatric experimental infections of the planorbid snail Gyraulus chinensis with miracidia of Euparyphium albuferensis (Trematoda: Echinostomatidae).

An experimental infection with echinostomatid miracidia in sympatric or 'local' vs. allopatric or 'away' snail combinations, as a model to examine parasite compatibility, was carried out. We employed Euparyphium albuferensis miracidia to infect Gyraulus chinensis snails, from three different natural parks: Albufera (Valencia, Spain); the Ebro Delta (Tarragona, Spain) and Coto de Doñana (Huelva, Spain). Insignificant differences between the three snail strains were noted for the infection rate and the rhythm of daily cercarial production. However, a significantly higher total cercarial production per snail, patent period and life span were observed in local snails. The different infection characteristics in the three G. chinensis strains considered reveal that E. albuferensis miracidia demonstrate local adaptation.

Protozoa in respiratory pathology: a review.

Among the micro-organisms that may affect the respiratory apparatus are the protozoa. The diseases they may give rise to constitute a relatively uncommon group of respiratory ailments with, in the majority of cases, an underlying clinical situation corresponding to states of suppressed immunity (AIDS, transplants, malign haemopathies, corticotherapy, etc.). Other factors, such as visits to endemic areas and immigration, also have to be taken into account. In view of the probable increase in the number of cases and the appearance of new emerging diseases, it is the intention of the present work to review the publications available, in different fields of medicine, that refer to the principal kinds of protozoa (Entamoeba, Acanthamoeba, Balamuthia, Leishmania, Trypanosoma, Trichomonas, Lophomonas, Cryptosporidium, Cyclospora, Toxoplasma, Plasmodium, Babesia, Encephalitozoon, Enterocytozoon and Balantidium) and, at the same time, detail and comment on the latest findings on this subject.

Molecular cloning and characterization of Echinostoma caproni heat shock protein-70 and differential expression in the parasite derived from low- and high-compatible hosts.

We cloned and expressed Echinostoma caproni HSP70 in Escherichia coli. This molecule presents an open reading frame (ORF) of 655 amino acids, and a theoretical molecular weight of 71 kDa. E. caproni HSP70 protein showed a high homology to other helminth molecules, major differences being located in the C-terminal region of the molecule, with a hydrophobic portion. Studies of protein and messenger RNA (mRNA) expression revealed a distinct pattern, depending on the host (low- or high-compatible). Specific polyclonal antisera raised against the recombinant protein expressed in Escherichia coli demonstrated its selective presence in excretory/secretory products (ESP) of adult parasites obtained from high-compatible hosts. Immunological studies showed clearly the association of HSP70 with the parasite surface and other structures, including eggs.

Identification of antigenic proteins from Echinostoma caproni (Trematoda) recognized by mouse immunoglobulins M, A and G using an immunoproteomic approach.

Antigenic proteins of Echinostoma caproni (Trematoda) against mouse IgM, IgA, IgG, IgG1 and IgG2a were investigated by immunoproteomics. Excretory/secretory products (ESP) of E. caproni separated by two-dimensional (2D) gel electrophoresis were transferred to nitrocellulose membranes and probed with the different mouse immunoglobulin classes. A total of four proteins (enolase, 70 kDa heat-shock protein (HSP-70), actin and aldolase) were accurately identified. Enolase was recognized in eight different spots of which seven of them were detected in the expected molecular weight and were recognized by IgA, IgG or IgG and IgG1. Another spot identified as enolase at 72 kDa was only recognized by IgM. Digestion with N-glycosidase F of the 72 kDa band rendered a polypeptide with an apparent molecular weight similar to that expected for enolase recognized by Western immunoblotting using anti-enolase antibodies. This suggests that glycosylated forms of enolase may be involved in the early thymus-independent responses against E. caproni. Early IgM responses were also generated by actin and the HSP-70 which suggests that these proteins are exposed early to the host and may be of importance in the parasite establishment. The IgA responses also appear to be mediated by the HSP-70 and aldolase which could be related with the close contact of these proteins with the host mucosal surface after secretion.

Leucine aminopeptidase is an immunodominant antigen of Fasciola hepatica excretory and secretory products in human infections.

The liver fluke Fasciola hepatica parasitizes humans and ruminant livestock worldwide, and it is now being considered a reemerging zoonotic disease, especially in areas in which it is endemic, such as South America. This study investigates the immune response to excretory and secretory products produced by F. hepatica in a group of patients from the Peruvian Altiplano, where the disease is highly endemic. Using a proteomic approach and immunoblotting techniques, we have identified the enzymes leucine aminopeptidase (LAP) and phosphoenolpyruvate carboxykinase as immunodominant antigens recognized by sera from fasciolosis patients. An indirect enzyme-linked immunosorbent assay using recombinant LAP as the antigen was developed to check sera from individuals of this region. Our results demonstrate that LAP produces a specific and strong reaction, suggesting its potential use in the serologic diagnosis of F. hepatica infections in humans.

Infectivity of Echinostoma friedi miracidia to different snail species under experimental conditions.

The infectivity of Echinostoma friedi (Trematoda: Echinostomatidae) miracidia was studied experimentally in a range of laboratory-reared snails that coexist in the same natural locality, namely Radix peregra, Lymnaea fuscus, L. truncatula (Lymnaeidae), Gyraulus chinensis, Helisoma duryi (Planorbidae) and Physella acuta (Physidae), and snails from different geographical origins acting naturally or experimentally as intermediate hosts of Schistosoma spp., namely Planorbarius metidjensis (from Málaga, Spain), Biomphalaria glabrata (Guadeloupe), B. alexandrina (Egypt) (Planorbidae), Bulinus cernicus (Mauritius), B. globosus (Zambia), B. natalensis (South Africa) and B. truncatus (Niger) (Bulinidae). Six species of snails were found to be susceptible, with the rate of infection ranging from 0 to 36.7%. The highest infection was detected in R. peregra. The low host specificity of E. friedi might have an epidemiological significance as a requisite for a recent establishment in a new geographical area.