Transoceanic ships as a source of alien dinoflagellate invasions of inland freshwater ecosystems.

Ships' ballast water and sediments have long been linked to the global transport and expansion of invasive species and thus have become a hot research topic and administrative challenge in the past decades. The relevant concerns, however, have been mainly about the ocean-to-ocean invasion and sampling practices have been almost exclusively conducted onboard. We examined and compared the dinoflagellate cysts assemblages in 49 sediment samples collected from ballast tanks of international and domestic routes ships, washing basins associated with a ship-repair yard, Jiangyin Port (PS), and the nearby area of Yangtze River (YR) during 2017-2018. A total of 43 dinoflagellates were fully identified to species level by metabarcoding, single-cyst PCR-based sequencing, cyst germination and phylogenetic analyses, including 12 species never reported from waters of China, 14 HABs-causing, 9 toxic, and 10 not strictly marine species. Our metabarcoding and single-cyst sequencing also detected many OTUs and cysts of dinoflagellates that could not be fully identified, indicating ballast tank sediments being a risky repository of currently unrecognizable invasive species. Particularly important, 10 brackish and fresh water species of dinoflagellate cysts (such as Tyrannodinium edax) were detected from the transoceanic ships, indicating these species may function as alien species potentially invading the inland rivers and adjacent lakes if these ships conduct deballast and other practices in fresh waterbodies. Significantly higher numbers of reads and OTUs of dinoflagellates in the ballast tanks and washing basins than that in PS and YR indicate a risk of releasing cysts by ships and the associated ship-repair yards to the surrounding waters. Phylogenetic analyses revealed high intra-species genetic diversity for multiple cyst species from different ballast tanks. Our work provides novel insights into the risk of bio-invasion to fresh waters conveyed in ship's ballast tank sediments and washing basins of shipyards.

Pathological lesions associated with avian schistosomes (Digenea: Schistosomatidae) in the freshwater snail Chilina dombeiana (Gastropoda: Chilinidae) from Southern Chile.

Avian schistosomes inhabit the blood stream of domestic and wild birds with aquatic snails as their intermediate hosts. In the Neotropics there is an emerging effort to describe species from these hosts, including Chile, although the knowledge about their pathological consequences is mostly understudied. This study aimed to describe the pathological changes associated with the parasitism of a native schistosomatid restricted to the Southern Cone of Neotropics. To achieve this, a total of 401 Chilina dombeiana snails (Chilinidae) were collected in two locations from Southern Chile. All of them were disposed to cercarial release procedure for three consecutive days. Furcocercariae released were stained and characterized by microscopic evaluation. Then, all snails were dissected under stereomicroscope and preserved in 10 % buffered formalin until histopathological analysis was performed. Eight out 401 (P = 2 %) snails were found parasitized with avian schistosomes. The released furcocercariae were identified as Schistosomatidae gen. sp. Lineage II which was previously reported in the same host. The main pathological change was an atrophy of ovotestes and an absence or mild infiltration of hemocytes in the surrounding tissues. Besides, a co-infection with echinostomes was found which was associated with a moderate hemocyte infiltration, granuloma-like lesion, and a reduced presence of schistosome' sporocysts. The latter would suggest an antagonistic interaction between these two digeneans, as has been proposed in the Echinostoma spp.-Schistosoma mansoni model. Despite the above, the release of furcocercariae was present but reduced, in contrast with the non-release of echinocercariae. This interaction requires further attention. This study represents the first attempt to characterize the pathological consequences of parasitism by a native, yet undescribed, avian schistosome in an endemic snail. Future studies should consider experimental infections to understand the dynamics of single infections in other Chilina species, including inter- and intra-specific parasitism as previous studies have found, including this study.

Identification of a new gregarine parasite associated with mass mortality events of freshwater pearl mussels (Margaritifera margaritifera) in Sweden.

Freshwater bivalves play key ecological roles in lakes and rivers, largely contributing to healthy ecosystems. The freshwater pearl mussel, Margaritifera margaritifera, is found in Europe and on the East coast of North America. Once common in oxygenated streams, M. margaritifera is rapidly declining and consequently assessed as a threatened species worldwide. Deterioration of water quality has been considered the main factor for the mass mortality events affecting this species. Yet, the role of parasitic infections has not been investigated. Here, we report the discovery of three novel protist lineages found in Swedish populations of M. margaritifera belonging to one of the terrestrial groups of gregarines (Eugregarinorida, Apicomplexa). These lineages are closely related-but clearly separated-from the tadpole parasite Nematopsis temporariae. In one lineage, which is specifically associated with mortality events of M. margaritifera, we found cysts containing single vermiform zoites in the gills and other organs of diseased individuals using microscopy and in situ hybridization. This represents the first report of a parasitic infection in M. margaritifera that may be linked to the decline of this mussel species. We propose a tentative life cycle with the distribution of different developmental stages and potential exit from the host into the environment.

[Human parasites in surface water used for recreation in Salta, Argentina]. / Parásitos humanos en aguas superficiales de uso recreativo en Salta, Argentina.

Waterborne diseases can have different origins, micro-organisms such as bacteria and parasites being the most important ones. In this study, two recreational aquatic environments were studied in the province of Salta, Argentina. Water samples collected from three different locations, two from a creek and one from the outlet of a thermal complex, were monitored at four time points. Physicochemical and microbiological characterization of each point was conducted, as well as a search for parasites and amebae. Parasites were identified through optical microscopy observations and free-living amebae (FLA) were isolated by spiking in Petri dishes followed by subsequent molecular identification. Water samples from the outlet of the thermal complex showed different physicochemical characteristics from those of the creek. Bacterial indicators of contamination were detected at all points; however, the creek water had a significantly higher concentration of Pseudomonas sp. Sporadically, creek samples exhibited Ascaris spp. eggs, Giardia sp. cysts, and ancylostomid eggs. The presence of FLA was observed in all samples, 15 of which were isolated and identified as Acanthamoeba sp., mostly belonging to the T4 genotype. Parasite surveillance in recreational aquatic environments is an important complement to traditional microbial indicators for assessing water quality. The identified parasites represent a potential health risk for people using these environments.

New species of Polystomoides (Monogenoidea: Polystomatidae) parasitizing the urinary bladder of a freshwater turtle in Brazil.

Trachemys dorbigni is the most abundant freshwater turtle species in Rio Grande do Sul, southern Brazil. Chelonians are known to host a wide variety of pathogens, including viruses, bacteria, hemoparasites and helminths. Among these, nine genera of polystomatid flatworms (Monogenoidea; Polystomatidae) infect freshwater turtles: Apaloneotrema, Aussietrema, Fornixtrema, Manotrema, Pleurodirotrema, Polystomoidella, Polystomoides, Uropolystomoides and Uteropolystomoides. However, little is known about the biology of these parasites in the Neotropical Realm. Through investigative cystoscopy, specimens of Polystomatidae were located inside the urinary bladder of the host T. dorbigni. Retrieved specimens were fixed and stained whole mounts prepared for taxonomic identification. In the present paper, a new species of Polystomoides (Monogenoidea: Polystomatidae) parasitizing the urinary bladder of a freshwater turtle of the species T. dorbigni in Brazil is described. Polystomoides santamariensis n. sp. differs from the congeneric species on the length of the genital spines, which are longer. Given the enormous diversity of freshwater turtles around the world, it is likely that a large number of chelonian polystomatids are still unknown.

Geographic range, distribution patterns and interactions of Monogenea Van Beneden 1858, with species of native host freshwater fishes from Brazil / Distribuição geográfica, padrões de distribuição e interações de Monogenea Van Beneden 1858, com espécies de peixes hospedeiros nativos de água doce do Brasil

Abstract This paper investigated information on monogenean species using 312 scientific papers, to search for infection and geographic distribution patterns in native freshwater fish from Brazil. We used 1,698 samples of 296 fish species of 28 families distributed into Characiformes, Siluriformes, Cichliformes, Gymnotiformes, Perciformes, Mugiliformes, Osteoglossiformes and Clupeiformes, in addition to four hybrid fish. Among the hosts of the different orders and families, the greatest numbers of parasite-host associations were found for species of the families Serrasalmidae, Characidae, Loricariidae, Curimatidae and Anostomidae. The 578 species of monogeneans used in parasite-host interactions were distributed in 86 genera of six five families (Dactylogyridae, Gyrodactylidae, Diplectanidae, Microcotylidae, Ancylodiscoididae and Ancyrocephalidae), but with great predominance of Dactylogyridae species. There was variation in prevalence, intensity and abundance levels of monogeneans species among host fish species, as well as in infection sites that occurred predominantly in external organs. Positive correlations of prevalence, intensity and abundance with body length of hosts were observed. There was geographic distribution pattern of monogeneans limited mostly to two hydrographic basins those being the Amazon River and Paraná River. Just approximately 6% of potential monogeneans have been explored thus far, showing a clear need for further studies on this interesting group of parasites.

Resumo Este estudo investigou informações sobre espécies de monogenéticos, usando 312 artigos científicos para buscar padrões de infecção e distribuição geográfica em peixes nativos de água doce do Brasil. Foram utilizadas 1.698 amostras de 296 espécies de peixes de 28 famílias, distribuídas em Characiformes, Siluriformes, Cichliformes, Gymnotiformes, Perciformes, Mugiliformes, Osteoglossiformes e Clupeiformes, além de quatro peixes híbridos. Entre os hospedeiros das diferentes ordens e famílias, os maiores números de associações parasito-hospedeiro foram encontrados para espécies das famílias Serrasalmidae, Characidae, Loricariidae, Curimatidae e Anostomidae. As 578 espécies de monogenéticos, utilizadas nas interações parasito-hospedeiro, foram distribuídas em 86 gêneros de seis famílias (Dactylogyridae, Gyrodactylidae, Diplectanidae, Microcotylidae, Ancylodiscoididae e Ancyrocephalidae), mas com grande predominância de espécie de Dactylogyridae. Houve variação nos níveis de prevalência, intensidade e abundância das espécies de monogenéticos entre as espécies de peixes hospedeiros, bem como nos locais de infecção que ocorreram predominantemente em órgãos externos. Correlações positivas de prevalência, intensidade e abundância com o comprimento corporal dos hospedeiros foram observadas. Houve padrão de distribuição geográfica dos monogenéticos limitados principalmente a duas bacias hidrográficas, sendo elas o Rio Amazonas e Rio Paraná. Apenas aproximadamente 6% dos potenciais monogenéticos são conhecidos até agora, mostrando uma clara necessidade de mais estudos sobre esse interessante grupo de parasitos.

Taxonomy and molecular phylogeny of a new freshwater ciliate Frontonia apoacuminata sp. nov. (Protista, Ciliophora, Oligohymenophorea) from Qingdao, PR China.

The morphology and ciliature of a new freshwater ciliate, Frontonia apoacuminata sp. nov., isolated from an artificial pond in Qingdao, PR China, were investigated using live observation and silver staining methods. The main features separating F. apoacuminata sp. nov. from its congeners are as follows: a broad elliptical body that is slightly pointed at the posterior end, four ophryokineties, one or two spherical micronuclei of a 'compact' type, a dorsally positioned contractile vacuole, and peniculi 1-3 each with five kinetosome rows though the left-most two rows in peniculus 3 are extremely shortened (with only two or three kinetosomes each). Additionally, an improved diagnosis of F. acuminata is provided. Phylogenetic analyses based on the small subunit ribosomal RNA (SSU rRNA) gene show that F. apoacuminata sp. nov. clusters with F. atra, F. minuta, F. acuminata and F. terricola. These five species group with Disematostoma, Marituja and Stokesia rather than with other Frontonia species, causing polyphyly of the genus Frontonia.

Acanthamoeba species isolated from Philippine freshwater systems: epidemiological and molecular aspects.

Free-living amoeba (FLA) research in the Philippines is still in its infancy but has, by far, demonstrated the presence of potentially pathogenic species. Acanthamoeba may cause sight-threatening and central nervous system infections to humans, yet its epidemiologic distribution from local environmental sources is yet to be defined. The present study aimed to provide a baseline epidemiologic distribution of Acanthamoeba spp. in freshwater systems in the Philippines and establish potential pathogenicity of isolates through thermo-tolerance assay. A total of 63 water samples were collected from 13 freshwater systems all over the Philippine archipelago. The low-volume (50 ml) water samples were processed and cultured on non-nutrient agar lawned with Escherichia coli and observed for amoebic growth using light microscopy. Amoebic culture demonstrated 14.28% (9/63) positivity while further molecular testing of culture-positive plates using Acanthamoeba-specific primers demonstrated 100% (9/9) confirmation of Acanthamoeba species. Genotyping of Acanthamoeba isolates revealed T1, T3, T4, T5, T7, T11, and T15 genotypes. Thermo-tolerance assay demonstrated that T5 and T7 genotypes were potentially pathogenic strains. The evidence of environmental distribution of Acanthamoeba spp. in the freshwater systems in the Philippines and thermo-tolerance profile of isolates are significant aspects of amoeba study in public health and calls for initiatives in the dissemination of relevant information and the expansion of knowledge, awareness, and policies on pathogenic waterborne amoeba to mitigate, prevent, detect, and report cases of human infections.

Application of DNA barcoding and morphometric analysis in differentiating cystacanths of Polymorphus species (Acanthocephala: Polymorphidae) from central Alberta, Canada.

Acanthocephalans are multi-host endoparasites, many of which use freshwater amphipods as intermediate hosts for their larval stages (e.g., cystacanths) while adults live in the intestines of vertebrates, including waterfowl. In central Alberta, Canada, several co-occurring species of the acanthocephalan genus Polymorphus use the amphipod Gammarus lacustris Sars, 1863 as an intermediate host. We applied DNA barcoding and morphometric analysis to differentiate cystacanth larvae from G. lacustris sampled from 17 Albertan water bodies. We slide-mounted specimens and measured morphological traits relating to proboscis hooks. We sequenced the standard DNA barcoding region of the mitochondrial cytochrome c oxidase subunit I gene (COI). Morphometric analysis suggested that the acanthocephalans we collected belonged to four morphologically different groups that keyed to Polymorphus contortus (Bremser, 1821) Travassos, 1926; P. marilis Van Cleave, 1939; P. paradoxus Connel et Corner, 1957; and P. strumosoides (Lundström, 1942) Amin, 2013. Our Bayesian tree based on COI sequences generally corroborated the morphological results and supported that the specimens assigned to P. cf. contortus and P. cf. strumosoides belong to two distinct species. In contrast, the Bayesian tree showed that specimens of P. cf. marilis were nested as a cluster within the P. cf. paradoxus clade. Similarly, small pairwise genetic distance (< 2%) between specimens identified as P. cf. contortus and P. cf. strumosoides suggests that they are conspecific. Future studies should use morphology and sequence data from adult acanthocephalans to assess the taxonomic identity of the cystacanth-based Polymorphus taxa. Our study is the first to provide genetic information for the four Polymorphus taxa and emphasizes the importance of applying multiple approaches to differentiate parasite species.

Naturally Occurring Microbiota Associated with Mosquito Breeding Habitats and Potential Parasitic Species against Mosquito Larvae: A Study from Gampaha District, Sri Lanka.

A mosquito species has its own favourable requirements of abiotic and biotic characteristics including microbiota, in a breeding habitat. Some of the microbiota may cause parasitic or pathogenic effects to mosquito larvae such as species of viruses, parasitic bacteria, fungi, protists, entomopathogenic nematodes, and filamentous fungi. In Sri Lanka, there is a scarcity of information on microbiota associated with mosquito breeding habitats and their effect on mosquito larvae. Hence, the present study was conducted to determine microbiota species/taxa associated with a variety of mosquito breeding habitats in selected areas of the Gampaha District in Sri Lanka and the relationship, if any, the microbiota has with mosquito larva survival and breeding. Forty-five microbiota species belonging to 11 phyla were found from different mosquito breeding habitats with the highest percentage belonging to phylum Euglenozoa (27.89%). Species that belonged to the phylum Amoebozoa (1.22%) and Sarcodina (1.17%) had the lowest abundance, and each of its species richness was recorded as one. Philodina citrina followed by Monostyla bulla comprised 30.8% and 16.59%, respectively, of the total rotifer population. From the total microbiota, 25-50% existed as accidental while less than 25% rare, in the habitat type according to their abundance. Paddy fields had the highest species richness (17), evenness (23.52), Shannon-Weiner (66.64), and beta diversity (0.65) over 50% indicating high heterogeneity in microbiota composition among the habitats. Ciliated protists, namely, Vorticella microstoma, Zoothamnium spp., and Chilodinella sp., were identified as naturally occurring microbiota associated with Culex mosquito larvae that inhabited in paddy fields and associated irrigation canals. Only Vorticella microstoma caused a significant lethal effect on mosquito larvae. This study revealed that species of Cx. gelidus, Cx. pseudovishnui, Cx. tritaeniorhynchus, Cx. quinquefasciatus, and Cx. whitmorei served as hosts for V. microstoma where infectivity rate in Cx. tritaeniorhynchus reached 73.22. Chilodinella sp. selectively served as endoparasitic to Cx. gelidus larvae causing only 4.58% mortality, and invasive cysts of the pathogen were observed in the subcuticular layer of the host body. Even though Zoothamnium spp. were found on Cx. tritaeniorhynchus larvae, there was no lethal effect due to the attachment of the parasitic agent. The potential of these microbiotas in integrated vector controlling approaches in future perspectives is recommended.

Curcuma longa ethanol extract and Curcumin inhibit the growth of Acanthamoeba triangularis trophozoites and cysts isolated from water reservoirs at Walailak University, Thailand.

CURCUMA LONGA: (C. longa) rhizome extract has been traditionally used to treat many infections. Curcumin, a pure compound isolated from the plant, has been documented to possess a wide spectrum of pharmacological effects. The present study aimed to investigate the effects of Thai medicinal plant extracts including C. longa extract and Curcumin on Acanthamoeba triangularis, a causative agent of human Acanthamoeba keratitis. The parasite was isolated from the recreational reservoir at Walailak University, Thailand. The organism was identified as A. triangularis using morphology and 18S rDNA nucleotide sequences. The pathogen was tested for their susceptibility to ethanol extracts of Thai medicinal plants based on eye infection treatment. The ethanol C. longa extract showed the strongest anti-Acanthamoeba activity against both the trophozoites and cysts, followed by Coscinium fenestratum, Coccinia grandis, and Acmella oleracea extracts, respectively. After 24 h, 95% reduction of trophozoite viability was significantly decreased following the treatment with C. longa extract at 125 µg/mL, compared with the control (P < 0.05). The extract at 1,000 µg/mL inhibited 90% viability of Acanthamoeba cyst within 24 h, compared with the control. It was found that the cysts treated with C. longa extract at 500 µg/mL demonstrated abnormal shape after 24 h. The MIC values of C. longa extract and Curcumin against the trophozoites were 125 and 62.5 µg/mL, respectively. While the MICs of the extract and curcumin against the cysts were 500 and 1,000 µg/mL, respectively. The results suggested the potential medicinal benefits of C. longa extract and Curcumin as the alternative treatment of Acanthamoeba infections.

Detection of Acanthamoeba spp. in two major water reservoirs in the Philippines.

Water reservoirs are important manmade structures providing water security to deliver clean and safe water for drinking and other purposes to the community. Eighty water samples were collected from Magat and Ipo water reservoirs using purposive sampling between November 2018 and January 2019. Water samples were collected in sterile containers for testing. The samples were cultured in non-nutrient agar and lawned with Escherichia coli and incubated at 33 °C. Twelve out of the 80 (15%) water samples were positive for amoebic growth. Light and scanning electron microscopy (SEM) revealed double-walled cystic stages and were initially identified as Acanthamoeba spp. based on morphological characteristic in reference to Page's established criteria. Their extracted DNAs were used in polymerase chain reaction using JDP1 and JDP2 primers and confirmed the presence of Acanthamoeba DNA in agarose gel electrophoresis. Aligned sequences from PCR products were deposited in GenBank under accession numbers MK886460, MK909919, MK905437, MK910997, MK911021 and MK886514. The presence of potentially pathogenic Acanthamoeba spp. in water reservoirs is considered a potential risk for public health, requiring appropriate processing of water in treatment plants.

Decay of infectious adenovirus and coliphages in freshwater habitats is differentially affected by ambient sunlight and the presence of indigenous protozoa communities.

BACKGROUND: Sanitary quality of recreational waters worldwide is assessed using fecal indicator bacteria (FIB), such as Escherichia coli and enterococci. However, fate and transport characteristics of FIB in aquatic habitats can differ from those of viral pathogens which have been identified as main etiologic agents of recreational waterborne illness. Coliphages (bacteriophages infecting E. coli) are an attractive alternative to FIB because of their many morphological and structural similarities to viral pathogens. METHODS: In this in situ field study, we used a submersible aquatic mesocosm to compare decay characteristics of somatic and F+ coliphages to those of infectious human adenovirus 2 in a freshwater lake. In addition, we also evaluated the effect of ambient sunlight (and associated UV irradiation) and indigenous protozoan communities on decay of somatic and F+ coliphage, as well as infectious adenovirus. RESULTS: Our results show that decay of coliphages and adenovirus was similar (p = 0.0794), indicating that both of these bacteriophage groups are adequate surrogates for decay of human adenoviruses. Overall, after 8 days the greatest log10 reductions were observed when viruses were exposed to a combination of biotic and abiotic factors (2.92 ± 0.39, 4.48 ± 0.38, 3.40 ± 0.19 for somatic coliphages, F+ coliphages and adenovirus, respectively). Both, indigenous protozoa and ambient sunlight, were important contributors to decay of all three viruses, although the magnitude of that effect differed over time and across viral targets. CONCLUSIONS: While all viruses studied decayed significantly faster (p < 0.0001) when exposed to ambient sunlight, somatic coliphages were particularly susceptible to sunlight irradiation suggesting a potentially different mechanism of UV damage compared to F+ coliphages and adenoviruses. Presence of indigenous protozoan communities was also a significant contributor (p value range: 0.0016 to < 0.0001) to decay of coliphages and adenovirus suggesting that this rarely studied biotic factor is an important driver of viral reductions in freshwater aquatic habitats.

Genetic structure and geographical variation of Bithynia siamensis goniomphalos sensu lato (Gastropoda: Bithyniidae), the snail intermediate host of Opisthorchis viverrini sensu lato (Digenea: Opisthorchiidae) in the Lower Mekong Basin revealed by mitochondrial DNA sequences.

The freshwater snail Bithynia siamensis goniomphalos sensu lato is widely distributed in the Lower Mekong Basin where it acts as the first intermediate host of the liver fluke Opisthorchis viverrini, a group 1 carcinogen causing cholangiocarcinoma. This study explores the genetic structure and geographical variation of B. s. goniomphalos from eight previously studied catchments and eight new catchments. These catchments belong to five previously studied catchment systems and one new catchment system (Tonlesap) in the Lower Mekong Basin. Two new catchment systems, Prachin Buri and Bang Pakong from eastern and central Thailand, respectively, were also examined. We collected 289 specimens of B. s. goniomphalos from 15 previously studied localities and 18 new localities in Thailand, Lao PDR (People's Democratic Republic), and Cambodia. The mitochondrial cytochrome c oxidase subunit 1 and 16S ribosomal DNA sequences were used to determine genetic variation. Classification of haplotypes specified 100 at the cox1 locus and 15 at the rrnL locus. Comparison between 16 catchment populations found significant genetic differences (ФST) between all populations. The phylogenetic tree and haplotype network analyses classified B. s. goniomphalos into three evolutionary lineages (lineage I-III). Lineage I contained B. s. goniomphalos from the Mekong, Chi, Mun, Prachin Buri and Bang Pakong catchments in Thailand, including the Nam Ngum catchment in Lao PDR. Lineage II contained all specimens from the Tonlesap catchment, whereas lineage III contained specimens from the Mekong and Sea Bang Heang catchments in Thailand and Lao PDR, respectively. Interestingly, Bithynia siamensis siamensis was placed between lineages I and II of B. s. goniomphalos. This study supports the hypothesis that B. s. goniomphalos is a species complex containing at least three distinct evolutionary lineages in the Lower Mekong Basin, and that comprehensive molecular genetic analyses need to be conducted to further our understanding of the evolutionary and systematic relationships of these Bithynia snail taxa.

Redescription and supplementary molecular characteristics of Aspidogaster ijimai Kawamura, 1915 (Trematoda, Aspidogastrea, Aspidogastridae), a parasite of Cyprinus carpio Linnaeus, 1758 s. lato (Actinopterygii) and freshwater bivalves in East Asia.

A detailed morphological description and ribosomal DNA (rDNA) sequence molecular data for Aspidogaster ijimai from Japan are provided. Morphological analysis, including a description of the cirrus-sac, indicate the conspecificity of Japanese and continental East Asian A. ijimai specimens. Analysis of ITS1-5.8S-ITS2 rDNA sequences of Japanese, Chinese and Russian specimens confirmed the morphological results. Phylogenetic analysis using ITS rDNA sequences confirmed that A. ijimai is a sister species for Aspidogaster chongqingensis. Median-joining network analysis showed an initial molecular differentiation step of Russian specimens from group of Japanese-Chinese samples. Our 28S rDNA results based on maximum likelihood (ML) and Bayesian (BI) phylogenetic analyses indicated well-supported monophyly of the Aspidogaster conchicola + A. ijimai group, a finding that indicates that these species are congeneric. At the same time, our data demonstrated that the genus Lobatostoma is paraphyletic and the family Aspidogastridae is polyphyletic, results that confirm previous studies.

Low-cost internal controls for detection of Giardia cysts in water samples.

Giardia cysts stained with hot carbolfuchsin were used as internal controls in a concentration method for surface water samples. The morphological integrity of stained cysts and the stain's stability and intensity were tested with each of the chemical reagents used in the aluminum sulfate flocculation method. No alterations in morphology or color were noted. The stained cyst preparation has a low cost, high stability, and suitability for both light and immunofluorescent microscopy, making it affordable to researchers in low- and middle-income countries.

Comparison of pathogen-derived 'total risk' with indicator-based correlations for recreational (swimming) exposure.

Typical recreational water risk to swimmers is assessed using epidemiologically derived correlations by means of fecal indicator bacteria (FIB). It has been documented that concentrations of FIB do not necessarily correlate well with protozoa and viral pathogens, which pose an actual threat of illness and thus sometimes may not adequately assess the overall microbial risks from water resources. Many of the known pathogens have dose-response relationships; however, measuring water quality for all possible pathogens is impossible. In consideration of a typical freshwater receiving secondarily treated effluent, we investigated the level of consistency between the indicator-derived correlations and the sum of risks from six reference pathogens using a quantitative microbial risk assessment (QMRA) approach. Enterococci and E. coli were selected as the benchmark FIBs, and norovirus, human adenovirus (HAdV), Campylobacter jejuni, Salmonella enterica, Cryptosporidium spp., and Giardia spp. were selected as the reference pathogens. Microbial decay rates in freshwater and uncertainties in exposure relationships were considered in developing our analysis. Based on our exploratory assessment, the total risk was found within the range of risk estimated by the indicator organisms, with viral pathogens as dominant risk agents, followed by protozoan and bacterial pathogens. The risk evaluated in this study captured the likelihood of gastrointestinal illnesses only, and did not address the overall health risk potential of recreational waters with respect to other disease endpoints. Since other highly infectious pathogens like hepatitis A and Legionella spp. were not included in our analysis, these estimates should be interpreted with caution.

Snail transmitters of schistosomiasis and other mollusks of medical and economic importance at the simplício queda única hydroelectric plant, southeast Brazil

Abstract INTRODUCTION This paper presents the results of an extensive survey of freshwater mollusks in the Simplício Queda Única Hydroelectric Development area, Southeast Brazil. METHODS Mollusks were collected between 2008 and 2013. All specimens were examined for the presence of larval trematodes. RESULTS In total, 12,507 specimens classified into 16 genera were obtained. Known snail vectors of schistosomiasis and fascioliasis and exotic species were identified, and new records are reported. CONCLUSIONS No specimens parasitized by larval trematodes of medical interest were detected. However, the results reinforce the importance of surveillance in study areas vulnerable to the occurrence of schistosomiasis transmission.

Identification of a novel piscine Cryptosporidium genotype and Cryptosporidium parvum in cultured rainbow trout (Oncorhynchus mykiss).

This study reports for the first time the presence and molecular characterization of Cryptosporidium in farmed rainbow trout (Oncorhynchus mykiss Walbaum, 1792). A total of 360 fish, with no apparent clinical signs of disease, were collected and classified into groups according to their size. Cryptosporidium oocysts were detected by immunofluorescence microscopy in 33 specimens (9.2%), which were located in pyloric caeca samples (42.4%), intestinal scrapings (39.4%), or at both locations (18.2%). In the smallest (youngest) fish group, a higher percentage of positive samples were detected in the pyloric caeca relative to the intestinal location (58.8 vs. 17.6%; P = 0.01), including a cluster with more than 10 oocysts observed in the pyloric caeca of one specimen. PCR amplification and sequencing of fragments of SSU-rDNA and hsp70 genes identified a novel Cryptosporidium piscine genotype (genotype 9) in two specimens and Cryptosporidium parvum in seven fish, including the specimen in which the oocyst cluster was observed. Moreover, Cryptosporidium oocysts were detected in farm water samples (41.7 and 16.7% from influent and effluent, respectively). Although Giardia was not found in gastrointestinal samples, Giardia cysts were observed in 50.0 and 33.3% of the influent and effluent water samples, respectively. The results support the existence of natural infections by C. parvum in freshwater cultured fish, suggesting that the rainbow trout could shed infectious oocysts in aquatic environments and it may be a potential source of human infection when this edible fish is handled.

First molecular evidence of Thelohanellus wallagoi Sarkar, 1985 (Myxozoa) from economically important food fish, freshwater shark Wallago attu (Siluridae) in India.

The freshwater shark Wallago attu (Bl. and Schn.) is a frequent silurid in the River Ganga and one of the most commercially exploit fish in India. In a survey on its infection with myxosporeans, spore type belongs to Thelohanellus species was found in the gills, kidney and intestine respectively. Through morphological and molecular investigations, we identified the spore as Thelohanellus wallagoi Sarkar, 1985. They were pyriform in valvular view and slim in sutural view, and had one pyriform polar capsule with four to five turns. The spores measured 8.0 × 4.0 × 2.3 µm. T. wallagoi developed in small cysts in the gill lamellae, whereas cyst and scattered spores of T. wallagoi were also found in the kidney and intestine respectively. The 18S rDNA sequence of T. wallagoi isolates recovered from gills, kidney and intestine were found similar to each other and differed from any other Thelohanellus species available in GenBank and validated its status after 32 years of original description. Phylogenetic analysis signified that T. wallagoi was placed sister to Myxobolus species in the clade that indicated the polyphyletic nature of the genus Thelohanellus.