Fasciolopsis Research Articles

Screening and identification of DNA aptamers toward Schistosoma japonicum eggs via SELEX.

Schistosomiasis is a major parasitic disease caused by blood flukes of the genus Schistosoma. Several million people all over the world are estimated to suffer from severe morbidity as a consequence of schistosomiasis. The worm’s eggs, which cause the symptoms of schistosomiasis, are generally used to diagnose the disease. In this study, we employed egg-based systematic evolution of ligands by exponential enrichment (egg-SELEX) and identified a panel of ssDNA aptamers specifically binding to eggs derived from S. japonicum. Among these, two aptamers LC6 and LC15 exhibited strong binding to and specific recognition of S. japonicum eggs, but not eggs from Fasciolopsis buski, Enterobius, Ascaris or Clonorchis sinensis. Furthermore, tissue imaging results revealed that LC15 could recognize S. japonicum eggs laid in liver tissues with a detection ratio of 80.5%. Collectively, therefore, we obtained useful aptamers specifically recognizing S. japonicum eggs, which will facilitate the development of an effective tool for both schistosomiasis diagnosis and drug delivery.

Intestinal and liver flukes of birds of prey (Accipitriformes, Falconiformes, Strigiformes) from Slovakia: uniform or diverse compound?

During 2012-2014 up to 286 birds of the orders Falconiformes (5 species), Accipitriformes (11 species), and Strigiformes (7 species) were examined for trematodes and this represents the first detailed study in Slovakia. A total of 12 trematode species belonging to the families Diplostomidae, Cyathocotylidae, Strigeidae, and Opisthorchiidae were identified. Rare infections were found in falcons where only two species (40%) and three of 85 examined birds (3.5%) were infected with a low range of two to four worms of generalists Strigea falconis or Plagiorchis elegans. Contrary to that, ten accipitriformes species (90.9%) and 63 of 156 bird individuals (40.4%) were infected with nine flukes: Conodiplostomum perlatum, Conodiplostomum spathula, Neodiplostomum attenuatum, Neodiplostomum spathoides, Parastrigea flexilis, Strigea falconis, Strigea vandenbrokae, Paracoenogonimus ovatus, and Metorchis bilis. S. falconis and N. attenuatum were the most frequent, occurring in parallel in eight and four bird species, in numbers up to 575 and 224. The intensity of infection with other fluke species was low ranging from one to 13 worms. Three owl (Strigiformes) representatives (42.9%) were exclusive hosts for Neodiplostomum canaliculatum and Strigea strigis, and the proportion of positive and dissected individual birds was 10:45 (22.2%). Both trematodes occurred in two or three owl species. In conclusion, apparent dissimilarity of trematode load of three unrelated lines of falcons, eagles, and owls was revealed. The present study extends our knowledge on the composition of the trematode fauna in Slovakia as all species except S. falconis and P. elegans that represent new host and species records in Slovakia.

Complete Mitochondrial Genome of Echinostoma hortense (Digenea: Echinostomatidae).

Echinostoma hortense (Digenea: Echinostomatidae) is one of the intestinal flukes with medical importance in humans. However, the mitochondrial (mt) genome of this fluke has not been known yet. The present study has determined the complete mt genome sequences of E. hortense and assessed the phylogenetic relationships with other digenean species for which the complete mt genome sequences are available in GenBank using concatenated amino acid sequences inferred from 12 protein-coding genes. The mt genome of E. hortense contained 12 protein-coding genes, 22 transfer RNA genes, 2 ribosomal RNA genes, and 1 non-coding region. The length of the mt genome of E. hortense was 14,994 bp, which was somewhat smaller than those of other trematode species. Phylogenetic analyses based on concatenated nucleotide sequence datasets for all 12 protein-coding genes using maximum parsimony (MP) method showed that E. hortense and Hypoderaeum conoideum gathered together, and they were closer to each other than to Fasciolidae and other echinostomatid trematodes. The availability of the complete mt genome sequences of E. hortense provides important genetic markers for diagnostics, population genetics, and evolutionary studies of digeneans.

Identification and characterization of microRNAs in the zoonotic fluke Fasciolopsis buski.

Fasciolopsis buski is a food-borne zoonotic parasite which is transmitted by aquatic plants, with pigs and humans as the definitive hosts. The objective of the present study was to characterize the microRNA (miRNA) profiles of this parasite by Solexa deep sequencing and bioinformatic analysis. Approximately 12 million high-quality reads were obtained from adult F. buski. A total of 286 miRNA candidates were found and 24 miRNA candidates were conserved miRNAs in the miRBase database. Three novel miRNAs were identified and confirmed by stem-loop reverse transcriptase polymerase chain reaction (RT-PCR). The miRNAs found in the present study belong to 13 families whose members showed high bias. The guanine (G) was the dominant nucleotide at the beginning and middle of the conserved miRNAs, particularly at the positions of 2nd, 6th, and 13th. To our knowledge, this is the first report of the miRNA profiles of F. buski, which would lay a foundation for further functional studies of miRNAs of F. buski.

Complete mitochondrial genome annotation of the giant intestinal fluke, Fasciolopsis buski (Indian isolate) as revealed by ion torrent and illuminanext-generation sequencing

The complete mitogenome sequences of the intestinal fluke Fasciolopsis buski are presented for the first time in this study. It is 14,119 bp long and is thus the shortest trematode mitochondrial genome sequenced to date. The F. buski mtDNA genome has a close resemblance with F. hepatica and has a similar gene order tallying with that of other trematodes. The overall base composition of F. buski mitogenome is 17.89% for A, 9.16% for C, 27.59% for G and 45.36% for T, and has a GC content of 36.75%. The assembled mitogenome (GenBank accession number KX449331) consists of 12 protein-coding genes (PCGs), 22 transfer RNAs and two ribosomal RNA genes. The mtDNA for the intestinal fluke reported herein would help investigate Fasciolidae taxonomy and systematics with the aid of mtDNA NGS data.

Clonorchis sinensis and Opisthorchis spp. in Vietnam: current status and prospects.

Clonorchis sinensis and Opisthorchis viverrini are clinically important small liver flukes because of their known association with development of cholangiocarcinoma. In Vietnam, high prevalence of C. sinensis infection in humans was previously reported in northern provinces, and O. viverrini infection has been detected in several central provinces. However, diagnosis of C. sinensis and O. viverrini infections in the past was merely based on faecal egg examination. This method alone can lead to misidentification at the species level because of morphological similarity between the eggs of these liver flukes and minute intestinal trematodes of the family Heterophyidae. In fact, recent surveys in Vietnam revealed that infection with several minute intestinal flukes, such as Haplorchis pumilio and H. taichui, are much more common than infection with C. sinensis or O. viverrini, and they often co-infect humans. Thus, previously reported prevalence of small liver fluke infection in Vietnam was likely over-estimated due to mis identification of parasites in copro-parasitological examinations. In addition, there is some confusion about identification of cercariae, metacercariae and also adults of C. sinensis and O. viverrini in intermediate and definitive hosts. The aim of this review is, therefore, to draw realistic pictures of the past and present scientific reports on the epidemiology and biology of C. sinensis and Opisthorchis spp. infection in Vietnam.

Fasciolopsis buski Vomited Out by a Child; The First Case Reported from Nepal

Live adult worms of Fasciolopsis buski are rarely seen in humans except in autopsy. Only a few such cases have been reported in the world literature. We reported a case of fasciolopsiasis in a child of age 14 months who coughed out the live adult Fasciolopsis buski after administration of antihelminthic drug. The patient was a resident of Terai (Far Western) region of Nepal and had history of travelling to India. This is the first case of fasciolopsis reported from Nepal.Nepal Journal of Biotechnology. Dec. 2015 Vol. 3, No. 1: 66-67

Food-borne zoonotic trematodes: devious old flukes in an urbanized, global village

Food-borne trematodes constitute a family of lung, liver and intestinal flukes that have parasitized mankind and domesticated animals since the distant past. In the modern global village, food-borne trematodes present a daunting challenge of infection with severe morbidity, despite being highly neglected as causes of disease. Urbanization is the population shift from rural to urban centers, as a necessity for industrialization and development. It is currently the hallmark of rapidly developing low and middle income countries and usually characterized by both poor planning and sustainability. In many communities, the population shift observed is mainly due to rural-urban migration. With urban industrialization and globalization however, immigration plays a significant role as well. The most significant effect is seen in rapid population growth, beyond the capacity of available resources. Aside from localized businesses, cuisine and cultural centers are the first and fastest growing establishments in newly emerging urban centers. In the low and middle income countries, water and food safety are most affected in this environment. The opportunity for introduction and transmission of imported water and food-borne trematode infections is particularly high in coastal cities or those emerging around major natural and man-made water bodies. A typical outcome of globalization, urbanization and immigration is a rise in the prevalence and spread of food-borne trematodiases.

A molecular phylogeny of Asian species of the genus Metagonimus (Digenea)—small intestinal flukes—based on representative Japanese populations

Metagonimus Katsurada, 1912 is a genus of small intestinal parasites. The genus comprises eight species, primarily from far-eastern Asia, with two exceptions reported from Europe. Metagonimus yokogawai, the most widespread species, is the main agent responsible for the intestinal disease, metagonimiasis, in Japan and some other East Asian countries. On the basis of the ratio of the size of the ventral and oral suckers, Metagonimus has traditionally been morphologically divided into two groups; however, the genus has not been extensively studied using molecular data. To reveal phylogenetic relationships within Metagonimus based on molecular data, we analyzed six of the seven species present in Asia using samples collected in central Japan. Maximum likelihood and Bayesian analyses of a combined 28S ribosomal DNA (rDNA), internal transcribed spacer 2 (ITS2), and mitochondrial cox1 gene sequence dataset separated the six species into two well-supported clades. One clade comprised M. yokogawai, M. takahashii, M. miyatai, and M. hakubaensis, whereas the other consisted of M. otsurui and M. katsuradai. Genetic distances calculated from 28S rDNA and ITS2 nucleotide sequences and a comparison of the predicted amino acid sequences of cox1 gene suggested that M. otsurui and M. katsuradai may have diverged recently. None of the four main morphological characters used to delimit species of Metagonimus (i.e., sucker ratio, positions of the uterus and testes, and distribution of vitelline follicles) was consistent with the distribution of species in the molecular tree.

Zoonotic helminths parasites in the digestive tract of feral dogs and cats in Guangxi, China.

BackgroundIn Guangxi, a province of southern China, an important number of dogs and cats roam freely in rural settings, and the presence of these animals in proximity of people may represent a risk of parasitic zoonoses. The objective of the present study was to investigate the presence and identify gastrointestinal helminths in feral carnivores in Guangxi province. Therefore, post mortem examination was performed in 40 dogs and in 39 cats.ResultsThe Gastrointestinal helminths were found in all the necropsied dogs and in 37 out of 39 cats. Fifteen species were identified including 7 trematodes, 3 cestodes and 5 nematodes. Most of them may be responsible for zoonotic infections.ConclusionsMajor zoonotic gastrointestinal helminths, including liver and intestinal flukes, Toxocara spp., and Ancylostoma spp., are present in feral dogs and cats in Guangxi, and may represent a significant risk for public health.

Molecular characterization of Stictodora tridactyla (Trematoda: Heterophyidae) from Kuwait Bay using rDNA ITS and mtCO1.

Stictodora tridactyla is an intestinal fluke in the family Heterophyidae that parasitizes shorebirds and mammals, including humans. Its metacercarial cyst stage was reported in the Arabian killifish, Aphanius dispar, at Kuwait Bay. In the present study, Cerithidea cingulata was found to serve as the first intermediate host of S. tridactyla. In order to establish the snail-fish link in the life cycle of S. tridactyla, complete sequences of ribosomal DNA internal transcribed spacer region 1 and 2 (rDNA ITS1 and ITS2) and partial sequence of cytochrome oxidase subunit 1 were obtained for metacercarial cysts isolated from the fish A. dispar and rediae isolated from the snail C. cingulata. Sequence alignment demonstrated that these larval stages belong to the same heterophyid species, S. tridactyla. Phylogenetic analysis based on rDNA ITS1, ITS2, and mtCO1 confirmed the position of S. tridactyla within the Heterophyidae and found it to cluster with Haplorchis spp. The present study represents the first molecular study correlating the larval stages of S. tridactyla using rDNA ITS1, ITS2, and mtCO1 and examining the phylogenetic relationships of S. tridactyla with different heterophyid species.

Fasciolopsiasis: Endemic focus of a neglected parasitic disease in Bihar

Purpose: To describe a newly discovered, previously unreported endemic focus of fasciolopsiasis in the Phulwaria village, under tehsil Sugauli, East Champaran, Bihar. Materials and Methods: A study was conducted at village Phulwaria, following diagnosis of fasciolopsiasis in three children from the village. A total of 120 individuals, including all the children and adults who gave history of recent passage of red fleshy masses in their stool, were included in the study. The cases of fasciolopsiasis were treated with Praziquantel 25 mg/kg, three doses a day. Risk factors for the transmission of the parasite in the village were also studied. Results: Questionnaire revealed majority of the population suffering from abdominal discomfort and passage of red fleshy masses in stool. These fleshy masses were identified as Fasciolopsis buski. One hundred and eighteen individuals were presumably considered as cases of the parasitic infection. After treatment with Praziquantel, all of them passed the parasite in their stool for the next 2–3 days. On investigating, it was observed that all the conditions required for effective continuation of the life cycle of the parasite were present in this village. Conclusion: This study draws attention to a new endemic focus of fasciolopsiasis in Bihar, with a very high prevalence due to poverty, the lack of awareness about the parasite in villagers as well as ignorance among local medical practitioners. There is an urgent need for mass campaign around the region for its effective control.

Endoscopic extraction of Fasciolopsis buski presenting as acute upper GI bleeding in a child

Endoscopic extraction of Fasciolopsis buski presenting as acute upper GI bleeding in a child

Neglected tropical diseases among the Association of Southeast Asian Nations (ASEAN): overview and update.

The ten member states of the Association of Southeast Asian Nations (ASEAN) constitute an economic powerhouse, yet these countries also harbor a mostly hidden burden of poverty and neglected tropical diseases (NTDs). Almost 200 million people live in extreme poverty in ASEAN countries, mostly in the low or lower middle-income countries of Indonesia, the Philippines, Myanmar, Viet Nam, and Cambodia, and many of them are affected by at least one NTD. However, NTDs are prevalent even among upper middle-income ASEAN countries such as Malaysia and Thailand, especially among the indigenous populations. The three major intestinal helminth infections are the most common NTDs; each helminthiasis is associated with approximately 100 million infections in the region. In addition, more than 10 million people suffer from either liver or intestinal fluke infections, as well as schistosomiasis and lymphatic filariasis (LF). Intestinal protozoan infections are widespread, while leishmaniasis has emerged in Thailand, and zoonotic malaria (Plasmodium knowlesi infection) causes severe morbidity in Malaysia. Melioidosis has emerged as an important bacterial NTD, as have selected rickettsial infections, and leptospirosis. Leprosy, yaws, and trachoma are still endemic in focal areas. Almost 70 million cases of dengue fever occur annually in ASEAN countries, such that this arboviral infection is now one of the most common and economically important NTDs in the region. A number of other arboviral and zoonotic viral infections have also emerged, including Japanese encephalitis; tick-borne viral infections; Nipah virus, a zoonosis present in fruit bats; and enterovirus 71 infection. There are urgent needs to expand surveillance activities in ASEAN countries, as well as to ensure mass drug administration is provided to populations at risk for intestinal helminth and fluke infections, LF, trachoma, and yaws. An ASEAN Network for Drugs, Diagnostics, Vaccines, and Traditional Medicines Innovation provides a policy framework for the development of new control and elimination tools. Together with prominent research institutions and universities, the World Health Organization (WHO), and its regional offices, these organizations could implement important public health improvements through NTD control and elimination in the coming decade.

Occurrence and seasonality of internal parasite infection in elephants, Loxodonta africana, in the Okavango Delta, Botswana.

It is known from studies in a wide range of wild and domestic animals, including elephants, that parasites can affect growth, reproduction and health. A total of 458 faecal samples from wild elephants were analysed using a combination of flotation and sedimentation methods. Coccidian oocysts (prevalence 51%), and nematode (77%) and trematode (24%) eggs were found. Species were not identified, though trematode egg morphology was consistent with that of the intestinal fluke Protofasciola robusta. The following factors were found to have a significant effect on parasite infection: month, year, sex, age, and group size and composition. There was some evidence of peak transmission of coccidia and nematodes during the rainy season, confirmed for coccidia in a parallel study of seven sympatric domesticated elephants over a three month period. Nematode eggs were more common in larger groups and nematode egg counts were significantly higher in elephants living in maternal groups (mean 1116 eggs per gram, standard deviation, sd 685) than in all-male groups (529, sd 468). Fluke egg prevalence increased with increasing elephant age. Preservation of samples in formalin progressively decreased the probability of detecting all types of parasite over a storage time of 1-15 months. Possible reasons for associations between other factors and infection levels are discussed.

A survey for potentially zoonotic gastrointestinal parasites of dogs and pigs in Cambodia.

There is little information available on parasites of zoonotic significance in Cambodia. In 2011, in an effort to obtain data on potentially zoonotic gastrointestinal parasites in domestic animals, 50 dogs and 30 pigs residing in 38 households located in Ang Svay Check village, Takeo province, Cambodia were examined for parasites from faecal samples. The samples were processed using the formalin-ethyl acetate concentration technique (FECT). Hookworms were the most common zoonotic parasite found in dogs (80.0%) followed by Echinostomes (18.0%). While, in pigs, Fasciolopsis buski was the most common zoonotic parasite (30.0%) followed by Ascaris suum (13.3%). This study provides baseline data on gastrointestinal parasites in dogs and pigs from Cambodia and underscores the importance of domestic animals as reservoir hosts for human parasites for Cambodian veterinary and public health agencies. Follow-up studies are required to further taxonomically characterize these dog and pig parasites and to determine their role in human parasites in this community.

Health Status of Immigrant Children and Environmental Survey of Child Daycare Centers in Samut Sakhon Province, Thailand.

Samut Sakhon is a Thai province popular among immigrants attracted to work in factories and the Thai food industry, especially people from Myanmar. Poor personal-hygiene behaviors, crowded accommodation and limited sanitation, result in health problems among immigrant workers. Various infectious diseases among this group are seen and managed by Samut Sakhon General Hospital. The impact of intestinal parasitic infections on public health is well known; they can spread from infected immigrant areas to uninfected areas via close contact and fecal-oral transmission from contaminated food and water. This study aimed to determine the prevalence of intestinal parasitic infections among 372 immigrant children at 8 child-daycare centers during their parents' work time, by physical examination, fecal examination, and examination of the environment around the centers. Physical examinations were generally unremarkable, except that head-lice and fingernail examinations were positive in two cases (0.8%). The results showed intestinal parasitic infections to be highly prevalent, at 71.0%. These infections comprised both helminths and protozoa: Trichuris trichiura (50.8%), Enterobius vermicularis (25.2%), Ascaris lumbricoides (15.3%), hookworm (11.6%), Giardia lamblia (10.2%), Endolimax nana (3.5%), Entamoeba coli (2.7%), and Blastocystis hominis (0.5%). The environmental survey found a small number of houseflies near the accommodation to be positive for helminthic eggs (0.2%), including A. lumbricoides, E. vermicularis, hookworms, Taenia spp., and minute intestinal flukes. Regarding the high prevalence of intestinal parasitic infections among children, it has been conjectured whether they were infected, along with their parents, during their daily lives before or after settling in Thailand. Intestinal parasites among immigrant children may involve a significant epidemiological impact, since immigrant children can serve as carriers and transmitters of disease.

Neglected Tropical Diseases: Trematodiases—The Indian Scenario

Neglected tropical diseases (NTDs) represent the most common infections that mainly affect the poorest of the poor people living in tropical and subtropical countries. Food-borne trematodiases (FBTs) constitute an important group of NTDs caused by digenetic trematodes, which include the lung, liver and intestinal flukes. At least 56 million people are estimated to suffer globally with one or more FBTs. Food-borne trematodes are endemic in several parts of the world, particularly Southeast Asia including the Indian subcontinent. In the context of India, the tropical climate along with the traditional food practices of various native societies provide a congenial transmission environment for many of these trematodes. In the present review, the spectrum of FBTs reportedly occurring in the country and having a zoonotic potential is discussed in brief.

Prevalence of intestinal helminths among inhabitants of Cambodia (2006-2011).

In order to investigate the status of intestinal helminthic infections in Cambodia, epidemiological surveys were carried out on a national scale, including 19 provinces. A total of 32,201 fecal samples were collected from schoolchildren and adults between 2006 and 2011 and examined once by the Kato-Katz thick smear technique. The overall egg positive rate of intestinal helminths was 26.2%. The prevalence of hookworms was the highest (9.6%), followed by that of Opisthorchis viverrini/minute intestinal flukes (Ov/MIF) (5.7%), Ascaris lumbricoides (4.6%), and Trichuris trichiura (4.1%). Other types of parasites detected were Enterobius vermicularis (1.1%), Taenia spp. (0.4%), and Hymenolepis spp. (0.2%). The northwestern regions such as the Siem Reap, Oddar Meanchey, and Banteay Meanchey Provinces showed higher prevalences (17.4-22.3%) of hookworms than the other localities. The southwestern areas, including Koh Kong and Preah Sihanouk Provinces showed higher prevalences of A. lumbricoides (17.5-19.2%) and T. trichiura (6.1-21.0%). Meanwhile, the central and southern areas, in particular, Takeo and Kampong Cham Provinces, showed high prevalences of Ov/MIF (23.8-24.0%). The results indicate that a considerably high prevalence of intestinal helminths has been revealed in Cambodia, and thus sustained national parasite control projects are necessary to reduce morbidity due to parasitic infections in Cambodia.

Specific diagnosis of Opisthorchis viverrini using loop-mediated isothermal amplification (LAMP) targeting parasite microsatellites

Opisthorchis viverrini and other food-borne trematode infections are major health problems in Thailand, the Lao People's Democratic Republic, Vietnam and Cambodia. Differential diagnosis of O. viverrini based on the microscopic observation of parasite eggs is difficult in areas where Clonorchis sinensis and minute intestinal flukes coexist. Recently, loop-mediated isothermal amplification (LAMP) has been widely used for detection and identification of trematode for its simple method that is useful in low-resource or field settings. We have reported ITS1-LAMP assay to detect O. viverrini infection from human feces. The sensitivity and specificity of the test was 100% and 61.5%. The sensitivity of the test appeared to be higher than microscopic egg examination; however non-specific amplification from other parasites could not be ruled out. We therefore targeted microsatellites of O. viverrini that is a species specific sequence. By using hydroxyl naphthol blue (HNB)-LAMP, O. viverrini microsatellite 6 (OVMS6) could specifically amplify DNA from O. viverrini genome, but not other parasites such as C. sinensis, Opisthorchis felineus, Centrocestus caninus, Haplorchis taichui, Fasciola gigantica and Haplorchoodes sp. The detection limit of the test is 1ng genomic DNA, which was 1000 times lower than the ITS1-LAMP, but targeting microstellites showed more specific detection of O. viverrini. In addition, the colorimetric LAMP assay was simple and effective; this makes it potentially applicable for point-of-care diagnosis.