Seasonal and Geographic Distribution of Cercarial Infection in Lymnaea gedrosiana (Pulmunata: Lymnaeidae) In North West Iran

Background

Trematodes are a diverse group of endoparasites which require molluscan and vertebrate animals as intermediate and definitive hosts in their life cycle. The present study was carried out to determine the diversity and geographic distribution of infection with trematodes''cercariae in the snail Lymnaea gedrosiana from north-west Iran.

Methods

A total number of 6759 Lymnaeidae snails were collected from 28 snail habitats; of these L. gedrosiana was the prevalent snail (74.37%) which examined for cercarial infection by shedding method.

Results

The overall infection rate was 8.03%. The most frequent trematodes cercariae in the snail were xiphidiocercariae (81.98%), furcocercariae (32.26%), echinostome cercariae (5.19%), and monostome cercariae (1.24%). The highest infection rate in L. gedrosiana (100%) was with echinostome cercariae from Golestaneh in autumn.

Conclusion

Due to the important role of pond snails in transmission of cercariae to fish as a source of zoonotic diseases, it is essential to estimate the distribution and abundance of the snails and the rate of their infection with different trematodes’ cercariae, and establish control programs in each region.     

Unravelling the riddle of Radix: DNA barcoding for species identification of freshwater snail intermediate hosts of zoonotic digeneans and estimating their inter-population evolutionary relationships

Radix spp. are intermediate host snails for digenean parasites of medical and veterinary importance. Within this genus, species differentiation using shell and internal organ morphology can result in erroneous species identification, causing problems when trying to understand the population biology of Radix. In the present study, DNA barcoding, using cox1 and ITS2 sequences, identified populations of Radix auricularia and Radix balthica from specimens originally morphologically identified as Radix peregra from the UK. Assessment of cox1 and ITS2 as species identification markers showed that, although both markers differentiated species, cox1 possessed greater molecular diversity and higher phylogenetic resolution. Cox1 also proved useful for gaining insights into the evolutionary relationships of Radix species populations. Phylogenetic analysis and haplotype networks of cox1 indicated that R. auricularia appeared to have invaded the UK several times; some haplotypes forming a distinct UK specific clade, whilst others are more akin to those found on mainland Europe. This was in contrast to relationships between R. balthica populations, which had low molecular diversity and no distinct UK specific haplotypes, suggesting recent and multiple invasions from mainland Europe. Molecular techniques therefore appear to be crucial for distinguishing Radix spp., particularly using cox1. This barcoding marker also enables the population biology of Radix spp. to be explored, and is invaluable for monitoring the epidemiology of fluke diseases especially in the light of emerging diseases and food security.     

Detection of Infection with Larval Stages of Ornithobilharzia turkestanicum using PCR in Field-Collected Snails of Lymnaea gedrosiana from Northwestern Iran

Background

Infection with Ornithobilharzia turkestanicum has been reported in a wide range of animals worldwide. This study was undertaken to assess the utility of polymerase chain reaction (PCR), for detecting the infection with O. turkestanicum larvae stages in Lymnaea gedrosiana.

Methods

A total of 6,759 Lymnaeidae snails were collected from six aquatic habitats in West Azarbaijan, northwest Iran. Of these, the snails of L. gedrosiana were identified. To detect infected L. gedrosiana with the larval stages of O. turkestanicum, they were subjected for cercarial shedding and molecular examinations. The genomic DNA was extracted and PCR was performed to specifically amplify a fragment of the nuclear 28SrRNA gene of O. turkestanicum.

Results

Of all collected snails, 5.4% (365/6,759) were the snails of L. gedrosiana. The cercarial shedding method revealed that 23.56% (86/365) of the snails were infected. The PCR patterns confirmed that 28.77% (105/365) snails of L. gedrosiana were infected with larval stages of O. turkestanicum. The infected snails were observed in five studied sites. The highest infection rate (66.66%, 20/30) was recorded in the snails of Ghargologh in the northern part. Only 35.24% (37/105) of the infected snails were from the plain areas, whereas the remaining existed in high altitudes.

Conclusion

It was concluded PCR method could be an efficient and fast method for uncovering the actual rate of infection with larval stages of O. turkestanicum in the snails of L. gedrosiana. This method can be also useful for the domestic animals and public health management programs in the country.     

Haemolymph Components of Infected & None Infected Lymnaea snails with Xiphidiocercariae

Background

In this study the haemolymph components of infected and none infected Lymnaea gedrosiana with xiphidiocercaria larvae was compared.

Methods

Five hundred Fifty Lymnaea snails were collected from Ilam and Mazandaran provinces, Iran, during 2008–2009. The snails were transported to the lab at Tehran University of Medical Sciences and their cercarial sheddings were studied. Haemolmyphs of snails were extracted and cells were counted using haemocytometer and cell-surface carbohydrate were recognized by conjugated lectin (Lentil). Haemolymph protein concentrations were measured by Bradford protein assay method and soluble protein compositions were determined on sodium dodecyl sulphate polyacrilamide gel electrophoresis (SDS-PAGE).

Results

From the 550 examined Lymnaea snails for cercariae, 27 snails were infected with xiphidiocercariae. Mean of haemolymph cells (haemocyte) number were obtained 93480±2.43 (cells/ml) for none infected snails (25 snail) and 124560±2800 (cells/ml) for infected snails (25 snail). Mannose carbohydrate was recognized on haemocyte of none infected and infected snails. Mean of protein concentration of haemolymph plasma was obtained as 1354±160 µg/ml (1.4 mg/ml) for none infected snails (25 snails) and 1802±138 µg/ml (1.8 mg/ml) for infected snail (25 snails). Comparing to none infected snails, the SDS-PAGE results of haemolymph plasma of infected snails, showed an extra protein band (70 kDa). The results showed a significant difference between the amounts and the kinds of proteins in haemolymph of infected and none infected snails.

Conclusion

This information might be useful to understand of parasite detection, adhesion, engulfment and antigen agglutination by snail.     

Development of a nest-PCR for detection of Fasciola hepatica DNA in the intermediate snail host,Radix cucunorica,and the prevalence in northwestern China

Fasciolosis, a foodborne zoonotic disease, caused by Fasciola species which is considered an important problem for human health and livestock husbandry development. Snails are intermediate hosts of F. hepatica, the epidemiological surveillance of snails can evaluate the transmission risk of this disease in human and livestock. In this study, we developed a nest-polymerase chain reaction (nest-PCR) to detect the DNA of F. hepatica in Radix cucunorica, a prevalent intermediate host of this parasite in northwestern China. The nest-PCR was used to amplify a 208 bp fragment of the second internal transcribed spacer (ITS-2) of F. hepatica with two pairs of primers. The method was able to detect up to 0.16 fg genomic DNA in a 25 μL PCR reaction system even effected with high concentrations of snail DNA, and no cross reaction was observed from the genomic DNA of Paramphistomum cervi, Clonorchis sinensis, Orientobilharzia turkestanicum, Metorchis orientalis, Dicrocoelium chinensis. To evaluate the transmission risk of this disease, 409 snail samples collected from different areas of Gansu province were used to detect and analyze the transmission risk of F. hepatica in this area. Of 409 snail samples, the overall prevalence is 43.76%. The prevalence was 92.75% in Gannan Tibetan Autonomous Prefecture, while no snail was positive for F. hepatica in Linxia Hui Autonomous Prefecture. The nest-PCR was firstly used to detect the infection of F. hepatica in snail. It is a novel, useful and convenient method with high sensitivity and specificity. This study is the first report about the epidemiological surveillance of snail infection by F. hepatica in northwestern China, which will help to evaluate the transmission risk of F. hepatica in northwestern China.     

A study on cercarial dermatitis in Khuzestan province, south western Iran

Background

Cercarial dermatitis' or swimmer's itch' is an itchy inflammatory response to the penetration of the skin by non-human schistosome parasites. In the hot season, (May to September) in Khuzestan province in the south west of Iran, swimming in canals and agriculture activities in swampy areas are common. This survey was made on people from villages north of Ahwaz city in south west Iran, to estimate cercarial dermatitis in this region.

Methods

2000 people were observed for clinical signs of cercarial dermatitis. Also 2000 Lymnaea gedrosiana snails were collected from agriculture canals and examined for animal schistosome cercariae during 1998–2000.

Results

From this survey 1.1% of people had pruritic maculopapular rash on their feet, hands or other parts of body. From the total of examined snails, 2.4% were found to be infected with bird schistosome cercariae including Trichobilharzia species.

Conclusion

Cercarial dermatitis could be a health problem in this area. This is the first report of cercarial dermatitis from this region of Iran.     

Exotic Small Mammals as Potential Reservoirs of Zoonotic Bartonella spp.

To evaluate the risk for emerging human infections caused by zoonotic Bartonella spp. from exotic small mammals, we investigated the prevalence of Bartonella spp. in 546 small mammals (28 species) that had been imported into Japan as pets from Asia, North America, Europe, and the Middle and Near East. We obtained 407 Bartonella isolates and characterized them by molecular phylogenetic analysis of the citrate synthase gene, gltA. The animals examined carried 4 zoonotic Bartonella spp. that cause human endocarditis and neuroretinitis and 6 novel Bartonella spp. at a high prevalence (26.0%, 142/546). We conclude that exotic small mammals potentially serve as reservoirs of several zoonotic Bartonella spp.     

Genetic diversity analysis of Blastocystis subtypes and their distribution among the domestic animals and pigeons in northwest of Iran

Blastocystis is a unicellular, anaerobic, eukaryotic protist, a common parasite found in the intestinal tracts of animals and humans. During the last few years, the host fecal DNA analysis by nucleic acid-based method has led to significant advances in Blastocystis diagnostics and enabled subtypes (STs). The zoonotic transmission of Blastocystis to humans is not well understood, therefore the present study was conducted to identify Blastocystis subtypes in Iran from different animal hosts from northwest of Iran. A total of 427 fresh fecal specimens were collected from cattle, sheep, poultry and pigeon (40,150,132,105 respectively). To detect the Blastocystis sp., each fecal specimen was examined microscopically. Total DNA from the samples that were positive for Blastocystis sp. was isolated, and the barcoding region of the small subunit of ribosomal rRNA (18S rRNA) was amplified and sequenced. Subsequently, sequence analyses, genetic diversity indices and evolutionary relationships of Blastocystis subtype populations were carried out. In total, 14.98% of the analyzed samples were positive for Blastocystis sp. and the subtypes detected were ST3,7,10 and 14. Among these, the ST10 was the main subtype that was found only in the cattle, sheep and poultry and the zoonotic subtype ST3 was present only from cattle. Our study shows the presence of Blastocystis subtypes in the sheep in north west of Iran and also demonstrated that the genetic approaches are crucial to understand the host specify of subtypes and the mode of infection. The study suggests that the genetic approaches will help us to understand the host specificity of subtypes and their role in infection if they are obtained from human and animals from the same geographical locations. Therefore, it is important to study the zoonotic aspects of this parasite with large number of samples from different groups of animals and from different geographical locations.     

A foreign invader or a reclusive native? DNA bar coding reveals a distinct European lineage of the zoonotic parasite Schistosoma turkestanicum (syn. Orientobilharzia turkestanicum ())

Natural foci of Schistosoma turkestanicum (syn. Orientobilharzia turkestanicum) has been identified in the Gemenc Forest regions of Hungary utilising red deer as the definitive host. In order to identify the origins of this parasite in Europe standard DNA bar coding techniques were employed to sequence fragments of the cytochrome oxidase 1 (cox1) and the nuclear ribosomal internal transcribed region (ITS) from 10 individual adult male worms. Phylogenetic reconstruction using maximum likelihood phylogenetic reconstruction and haplotype networks of the cox1 showed all the worms to be of a distinct unique Hungarian lineage although some ITS haplotypes were shared with worms from populations in China and Iran. Molecular clock analysis suggests an early divergence event around 270,000 years before present (YBP) between all S. turkestanicum populations giving rise to the Chinese, Iranian and Hungarian lineages. However, divergence of the sequences within the Hungarian population appears to have occurred approximately 63,000 YBP suggesting a long established population of S. turkestanicum in Europe. This suggests that the Hungarian population of S. turkestanicum has been native since the Ice Age and probably established itself during the last interglacial period as red deer moved into Europe from North Africa and the Middle East. This may also indicate that the parasite may have unknown populations established in several other countries in Eastern, Central and Southern Europe.     

Prevention and Control of Fish-borne Zoonotic Trematodes in Fish Nurseries, Vietnam

Worldwide, >18 million persons were infected with fish-borne zoonotic trematodes in 2002. To evaluate the effectiveness of interventions for reducing prevalence and intensity of fish-borne zoonotic trematode infections in juvenile fish, we compared transmission rates at nurseries in the Red River Delta, northern Vietnam. Rates were significantly lower for nurseries that reduced snail populations and trematode egg contamination in ponds than for nurseries that did not. These interventions can be used in the development of programs for sustained control of zoonotic trematodes in farmed fish.     

Electrophoretic studies of planorbid egg-proteins: The Bulinus africanus and B. forskalii species groups*

It is possible that physiological differences between populations of an intermediate snail host of schistosomes may be linked with differences in the pathogenicity of the parasite strains they harbour. With a view to differentiating between local populations of bulinid snails, a previous electrophoretic study of snail egg-proteins has been extended to cover all the recognized species of the Bulinus africanus group and most of those of the B. forskalii group.     

Helminth Infections of House Mouse (Mus musulus) and Wood Mouse (Apodemus sylvaticus) from the Suburban Areas of Hamadan City,Western Iran

Background

To determine the prevalence and intensity of helminths and their zoonotic importance in small rodents inhabiting in the suburban areas of Hamadan City, Iran.

Methods

The present survey was conducted on the helminth infections of two species of rodents Apodemus sylvaticus (n=60) and Mus musculus(n=72) in the suburban areas of Hamadan City during 2010-2012. Rodents were collected and examined for helminth in the different organs. The nematodes were collected in 5% formalin solution and cleared in lactophenol, cestodes and trematodes collected from intestine fixed in AFA solution and stained by acetocarmine, cleared in xylol for identification.

Results

Helminths found in A. sylvaticus and M. musculus and their prevalence for the first time in suburban areas of Hamadan City were as follows; In A. sylvaticus: Cysticercus fasciolaris(3.33%), Syphacia fredrici(26.67%), S. stroma(8.33%), Anoplocephalidae sp. (1.67%), Skrjabinotaenia lobata(5%), Plagiorchis muris(1.67%) and in M. musculus:Hymenolepis nana (16.67%), H.diminuta (5.55%), S. obvelata(30.56%), S. ohtarom (9.72%), Rodentolepis crassa (1.39%), C. fasciolaris (1.39%). Among 11 species in two rodents 4 species including S. obvelata, H. nana, H.diminuta,and P. muris have zoonotic importance. Statistically the relation between gender and their helminth infections was not significant in either M. musculus or A. sylvaticus (P>0.05).

Conclusion

This study reports 11 species of helminths and on the other hand 3 species were identified for the first time in Iran and 5 species of them have potential health importance for public health and cat.     

Distribution of the snail Biomphalaria glabrata, intermediate host of Schistosoma mansoni, within a St Lucian field habitat

A total of 6360 mud samples were obtained, in 62 collections made with an exhaustive sampling device, from banana drains on the West Indian island of St Lucia during fortnightly samplings over a 2½-year period. Analysis of counts of the snail Biomphalaria glabrata from these samples showed that this species had a contagious distribution. This finding is consistent with other evidence that banana drains form a rigorous habitat for B. glabrata. Its distribution was more contagious than that of Oncomelania quadrasi in certain Philippine habitats and several species of aquatic snail in various African irrigation canals. The exact transformation for normalizing the snail counts for standard statistical techniques was z = x^0.287 but the more convenient cube root transformation is probably adequate. However, if too few snails are collected (15 or fewer per 100 samples) or if the frequency distribution of snail counts is discontinuous, with too many widely separated high frequency counts, neither transformation will be entirely satisfactory.     

Prevalence and Genetic Characterization of Cryptosporidium Spp. In Diarrheic Children from Gonbad Kavoos City,Iran

Background: Cryptosporidium is an intestinal protozean parasite causing waterborne and foodborne outbreaks of diarrheal diseases. The present study was performed in order to find prevalence and subtypes of Cryptosporidium among children with diarrhea in Gonbad Kavoos City, Northern Iran. Methods: Diarrheic samples were collected from 547 children. The initial parasitological diagnosis was made based on detection of oocysts using the modified Ziehl-Neelsen acid-fast staining method. The positive microscopically samples were selected for sequence analysis of partial 60 kDa glycoprotein (gp60) gene. Results: Out of 547 collected samples, 27 (4.94%) were positive for Cryptosporidium oocysts. Fifteen from 27 positive samples successfully amplified in PCR. Sequences analysis of gp60 gene in 15 Cryptosporidium isolates revealed that all of them (100%) were C. parvum. The results showed three subtypes of IIa subtype family (7 cases) including IIaA16G2R1, IIaA17G1R1, IIaA22G3R1 and one subtype of IId subtype family (8 cases). The most common allele was IId A17G1d (53.3%). Conclusion: The predominance of zoonotic subtype families of C. parvum species (IIa, IId) in the present study is in concordance with previous studies in Iran and emphasizes the significance of zoonotic transmission of cryptosporidiosis in the country.Key Words: Cryptosporidium, Subtypes, Gp60 gene, Children, Iran     

Trichobilharzia anseri n. sp. (Schistosomatidae: Digenea), a new visceral species of avian schistosomes isolated from greylag goose (Anser anser L.) in Iceland and France

Parasitological investigations carried out on birds in Iceland and France highlight the presence of four species of avian schistosomes from greylag geese (Anser anser L.): the european nasal species Trichobilharzia regenti and three visceral species, among which an unknown species isolated from blood vessels of the large intestine and liver. Morphological and molecular analyzes of different parasite stages (eggs, adults) revealed new species of Trichobilharzia genus – Trichobilharzia anseri sp. nov. Studies on host–parasite relationship under natural conditions, showed that the life-cycle includes the snail Radix balthica (syn. R. peregra) as intermediate host. The cercariae, already isolated in Iceland from two ponds of the Reykjavik capital area – the Family park and Tjörnin Lake – are the same as those isolated in 1999 by Kolářová et al. during the first study on Icelandic parasitic agents of cercarial dermatitis.     

Patterns of adult tick parasitization of coexisting European (Erinaceus europaeus) and Algerian (Atelerix algirus) hedgehog populations in eastern Iberia

The availability of data regarding population abundance and dynamics of ticks in wild vertebrate populations is crucial to understand the host-tick relationship and to assess the risk for domestic animals and humans from possible zoonotic diseases. In this study we analyse several host-intrinsic and environmental factors affecting the tick population of two sympatric hedgehog species (the European and the Algerian hedgehog) in two differently anthropized habitats in eastern Spain. We captured 215 hedgehogs and sampled 356 adult ticks of four different species (Rhipicephalus sanguineus, R. turanicus, R. bursa and R. pusillus). We analysed by General Linear Mixed Models how the hosts’ species, sex, capture month and site and their interactions affected tick presence and tick load. Further, we carried out Spearman correlation tests to analyse how environmental temperature and precipitation affect tick presence and load on hedgehogs. We found that, in general, male hedgehogs were more infested than females. However, the effects of the different factors depended on the tick species, especially related to their endophilic or exophilic character. High values of general tick infestation can be explained by the coincidence of tick activity peaks and higher male host activity levels, especially within areas with higher habitat diversity and species richness. We also discuss how the potential immunosuppressive effect of testosterone could be affecting our results. Our results show that in highly anthropized environments hedgehogs potentially act as important mixing vessels for tick-borne zoonotic pathogens and that monitoring ticks in periurban wildlife is important to assess potential health risks for pets and humans.     

Experimental Life Cycle of Hypoderaeum conoideum (Block, 1872) Diez, 1909(Trematoda: Echinostomatidae) Parasite from the North of Iran

Background:

Human Echinostomiasis is an intestinal disease caused by the members of family Echinostomatidae parasites. The aim of present research was to identify echinostomatidae cercariae emitted by Lymnaea palustris snails from Mazandaran province in the north of Iran based on the morphological and morphometrical characteristics of the different stages of experimental parasite life cycle.

Methods:

Echinostomatidae cercariae were collected from L. palustris (Gastropoda: Lymnaeidae) of the north of Iran. To collect metacercaria, 50 healthy snails were infected with cercariae experimentally (50 cercariae for each). To obtain the adult stage, 9 laboratory animals (3 ducks, 2 rats, 2 mice and 2 quails) were fed with 60 metacercaria for each. To identify parasite, the different stages of worm were examined using light microscope and then the figures were draw under camera Lucida microscope and measures were determined.

Results:

Averagely, 15metacercaria were obtained from each snail that had been previously exposed with cercariae. Ducks presented worm eggs in feces after 10–15 days post-infection. Intestinal worms were collected and identified as Hypoderaeum conoideum on the bases of figures and measures of cephalic collar, the number of collar spine, suckers diameter ratio, testes arrangement, etc.

Conclusion:

H. conoideum cercariae and adult worm are described. This is the first report of the different stages of the experimental life cycle of this parasite in Iran.     

Further observations on Bulinus (Bulinus) truncatus rohlfsi (Clessin) in Ghana: Seasonal population fluctuations and biology

Bulinus (B.) truncatus rohlfsi is an important snail host of Schistosoma haematobium in Ghana and probably elsewhere in West Africa. Study of this snail in natural habitats in Northern Ghana has shown that the pronounced population fluctuations can be broadly related to the alternating wet and dry seasons and to any marked changes in the aquatic vegetation. An increase in snail density and reproductive activity begins during the rainy season, reaching a peak in the dry season. The onset of the contraction phase in the snail population is often abrupt, although it may be preceded by intense oviposition; during this phase there are but a few widely scattered snails and little reproductive activity with a low level of survival, particularly of juvenile snails. The factors which favour the survival of young specimens are clearly critical in the evolution of the snail population.     

Genetic similarity between Taenia solium cysticerci collected from the two distant endemic areas in North and North East India

Taenia solium taeniasis/cysticercosis is a major public health problem in developing countries. This study reports genotypic analysis of T. solium cysticerci collected from two different endemic areas of North (Chandigarh) and North East India (Dibrugarh) by the sequencing of mitochondrial cytochrome c oxidase subunit 1 (cox1) gene. The variation in cox1 sequences of samples collected from these two different geographical regions located at a distance of 2585 km was minimal. Alignment of the nucleotide sequences with different species of Taenia showed the similarity with Asian genotype of T. solium. Among 50 isolates, 6 variant nucleotide positions (0.37% of total length) were detected. These results suggest that population in these geographical areas are homogenous.     

Helminth Infections in Rattus ratus and Rattus norvigicus in Tehran,Iran

Background

The aim of this study was to determine intestinal and liver helminth infections in Rattus rodents in Tehran Iran.

Methods

Overall, 306 traps were put in 39 different regions in Tehran from 2009 to 2010. Rodents, including R. rattus and R. norvegicus were caught by live-traps. They become unconscious and the spinal cords were cut, afterwards the body was dissected and the stomach, small intestine, large intestine, liver, and cecum were studied separately. The dominant type and the prevalence rate of parasites in the rodents were determined based on the infected parts of their body.

Results

After recognition of the helminthes’ types, among the 120 total number of rodents, 39 belonged to males, while among the infected rodents, 57(47.5%) were female and 18(15%) were male. The prevalence of infection in Tehran was 62.5%. Seventy cases (58.33%) of helminth infections were observed in R. rattus and 5 cases (4.16%) were observed in R. norvegicus. The maximum prevalence (15.5%) was seen in the center and east part of Tehran, while the minimum (9.16%) was in the north part of the city. The helminthes types and the corresponding percentages were Hymenolepis nana fraterna (35.8%), Heterakis spumosa (17.5%), Hymenolepis diminuta (7.5%) and Capillaria annulosa (1.6%). The dominant rodent was Rattus rattus and among the identified helminthes, Hymenolepis diminuta and Hymenolepis nana fraterna are zoonotic ones.

Conclusion

The information presented here improves our understanding of the major parasitic infections that rodents harbor and can transmit to human and animal populations in Iran. To prevent infectivity of human, the hazard of the identified zoonotic species needs to be contemplated.