Limited studies provide direct evidence of Clonorchis sinensis adults in the early stage of gallbladder stone formation. Our current research systematically studied 33 gallbladder stones resembling adult worms and shed light on the definite connection of C. sinensis infection with concomitant cholelithiasis. A total of 33 gallbladder stones resembling adult C. sinensis worms were systematically analysed. Fourier transform infrared spectroscopy, scanning electron microscopy and X-ray energy spectrometry were used to analyse the composition and microstructure. Meanwhile, a histopathological examination of the stone was carried out. The 33 gallbladder stones resembling adult C. sinensis worms included nine calcium carbonate (CaCO3) stones, 12 bilirubinate stones and 12 mixed stones. Clonorchis sinensis eggs were found in 30 cases, including all CaCO3 and mixed stones. Parasite tissues were detected in 12 cases, which were mainly CaCO3 stones or bilirubinate–CaCO3 mixed stones. The outer layer of stones was wrapped with 12.88% calcium salt, as revealed by X-ray energy spectrometry, while surprisingly, many C. sinensis eggs were found in the inner part of these stones. Based on our current findings, we concluded that calcification and packaging occurred after C. sinensis adult entrance into the gallbladder, subsequently leading to the early formation of CaCO3 or bilirubinate–CaCO3 mixed gallbladder stones. This discovery highlights definite evidence for C. sinensis infection causing gallbladder stones.

Blue sharks, Prionace glauca, are cosmopolitan, extremely vagile sharks and the species among elasmobranchs for which most surveys containing tapeworm community data are available worldwide. In this study we report on the tapeworm fauna of three samples of blue sharks (n = 37) from two new regions (one sample from Galicia, north-east Atlantic, and two from Valencia, western Mediterranean), and compared it with previous studies, assessing the relative role of the ecological and evolutionary factors in structuring local tapeworm assemblages. Nine cestode taxa were identified, of which four included adult specimens, that is, Platybothrium auriculatum, Prosobothrium armigerum, Anthobothrium caseyi and Molicola horridus. The abundance of these species, and Brillouin's diversity index, differed significantly among samples without a clear geographical signal. A comparison with six previous surveys revealed that tapeworm assemblages were composed of the same ‘core’ taxa, with mean species richness typically ranging from two to four species. Global records of adult tapeworms in blue sharks included: 15 taxa identified at species level, of which only eight (generalist trypanorhynchs) were shared with other sympatric host species; five mostly with other carcharhinids; and three with large lamnid sharks sharing the blue sharks’ habitat. The composition of tapeworm communities of blue sharks is thus highly constrained by strong host specificity, with composition and abundance varying across localities depending on idiosyncratic environmental conditions.

In Serbia, in most cases, small family outbreaks of trichinellosis occur due to the consumption of untested infected meat from domestic pigs that are raised and slaughtered in the backyards of small individual farms. The aim of this study is to present data regarding 24 patients involved in two outbreaks that occurred in two neighbouring districts in Serbia during a closely related period of time in August 2014. The source of infection in the first outbreak was undercooked pork, while raw pork sausages caused the second outbreak. Meat samples and sausages were Trichinella spp. positive by artificial digestion. With the aim to obtain a second opinion and validate the serological findings discovered at the Public Health Institute Nis, all samples were sent to the National Reference Laboratory for Trichinellosis, INEP. Serodiagnosis showed that 21 persons were positive (87.5%) and three (12.5%) were negative for anti-Trichinella antibodies, while 15 patients fulfilled the trichinellosis case definition. Western blot analysis (using an epitope unique for the muscle larvae stage of the Trichinella genus) confirmed the diagnosis of trichinellosis in five patients. Six patients also had specific antibodies against Toxocara canis (T. canis). Due to the fact that in endemic foci in Serbia there is the presence of Trichinella spiralis and T. canis and that these two infections could be asymptomatic, we consider that trichinellosis cases were irrefutably proven. The dilemma about the existence of co-infection with T. canis remained open due to the lack of clinical findings.

A parasitological survey of terrestrial slugs and snails was conducted at popular dog walking locations across the city of Nottingham, with the intensions of finding gastropods infected with parasites of medical (or veterinary) importance such as lungworm (metastrongyloid nematodes) and trematodes. A total of 800 gastropods were collected from 16 sites over a 225 km2 area. The extracted nematodes and trematodes were identified by molecular barcoding. Of the 800 gastropods collected, 227 were infected (172 had nematode infections, 37 had trematode infections and 18 had both nematode and trematode infections). Of the nematode infected gastropods genotyped, seven species were identified, Agfa flexilis, Angiostoma gandavense, Angiostoma margaretae, Cosmocerca longicauda, Phasmarhabditis hermaphrodita, Phasmarhabditis neopapillosa and an unknown Cosmocercidae species. Of the trematode infected gastropods genotyped, four species were identified, Brachylaima arcuate, Brachylaima fuscata, Brachylaima mesostoma and an unknown Plagiorchioidea species. No lungworm species were found within the city of Nottingham. To our knowledge, this study represents the first survey of gastropod-associated nematodes and trematodes in the East midlands of the United Kingdom.

As part of surveillance of snail-borne trematodiasis in Knowsley Safari (KS), Prescot, United Kingdom, a collection was made in July 2021 of various planorbid (n = 173) and lymnaeid (n = 218) snails. These were taken from 15 purposely selected freshwater habitats. In the laboratory emergent trematode cercariae, often from single snails, were identified by morphology with a sub-set, of those most accessible, later characterized by cytochrome oxidase subunit 1 (cox1) DNA barcoding. Two schistosomatid cercariae were of special note in the context of human cercarial dermatitis (HCD), Bilharziella polonica emergent from Planorbarius corneus and Trichobilharzia spp. emergent from Ampullacaena balthica. The former schistosomatid was last reported in the United Kingdom over 50 years ago. From cox1 analyses, the latter likely consisted of two taxa, Trichobilharzia anseri, a first report in the United Kingdom, and a hitherto unnamed genetic lineage having some affiliation with Trichobilharzia longicauda. The chronobiology of emergent cercariae from P. corneus was assessed, with the vertical swimming rate of B. polonica measured. We provide a brief risk appraisal of HCD for public activities typically undertaken within KS educational and recreational programmes.

Calodium hepaticum is a zoonotic nematode with a worldwide distribution. Although the host range of C. hepaticum includes a wide spectrum of mammals (including humans), this parasite is predominantly associated with the families Muridae and Cricetidae. Several Sigmodontinae species from Argentina were found to be infected by C. hepaticum, with a high prevalence in Akodon azarae. The present study focuses on C. hepaticum eggs from natural infection of three species of sigmodontine rodents from Argentina. Eggs were genetically characterized (intergenic 18S rRNA region). The objectives of this work are: (i) to propose a new analytical methodology; and (ii) to morphologically characterize C. hepaticum eggs, from three Sigmodontinae species (A. azarae, Calomys callidus and Oligoryzomys flavescens). Analyses were made by the Computer Image Analysis System based on the new standardized measurements and geometric morphometric tools. The resulting factor maps clearly illustrate global size differences in the parasite eggs from the three Sigmodontinae species analysed. The degree of similarity between egg populations was assessed through pairwise Mahalanobis distances, showing that the largest distances were detected between parasite eggs from C. callidus and O. flavescens. Herein, the phenotypical plasticity of C. hepaticum eggs is shown. Significant positive correlations were obtained between each egg parasite principal component 1 and rodent corporal characteristics: weight; liver weight; rodent length; and rodent body condition. The usefulness of the geometric morphometric analysis in studies of the relationship between C. hepaticum and its host must be highlighted. The high prevalence observed in A. azarae, associated with the wide size range of the parasite eggs evidenced by principal component analysis, suggests A. azarae to be the Sigmodontinae host species that plays the most important role as reservoir host for C. hepaticum in the New World.

The type species of the genus Epacrolaimus, Epacrolaimus declinatoaculeatus, is studied from the re-examination of type material of Aporcelaimus vorax, its junior synonym, and the observation of several Iberian populations and a few Iranian specimens. Morphologically, it displays a recognizable morphological pattern characterized by, among other features, the incurved nature of its odontostyle aperture, presence of perioral liplets or lobes, lip region 24–31 μm wide, odontostyle 21–25 μm long and comparatively anterior location of S2N pharyngeal gland nuclei. Nevertheless, variations in some morphological traits (vagina shape and tail shape) and in several morphometrics (body length, uterus length, vulva position, tail length and spicule length) are also noted. Sequences of D2–D3 domains of the 28S rDNA, 18S rDNA and COI mtDNA were obtained from several Iberian populations. Their analyses, in particular those from D2–D3 sequences, revealed the existence of a highly supported clade ((Epacrolaimus + Sectonema) + Metaporcelaimus), with a closer relationship between Epacrolaimus and Palaearctic populations of Sectonema, whereas the remaining aporcelaimid genera occupied placements in other clades. These results are discussed, with especial emphasis on the intricate separation of Epacrolaimus and Sectonema, which display significantly different protruding stomatal structure in spite of their close evolutionary relationship as derived from molecular trees.

Trichinellosis is an important worldwide foodborne zoonosis. The gold standard test to detect Trichinella spp. larvae in muscle samples of animals intended for human consumption is the artificial digestion method. Handling and dispensing of conventional pepsin powder present significant safety risks for analysts. The use of pepsin powder that is resistant to aerosolization should alleviate these safety concerns. The aim of this study was to compare the efficacy of an aerosol-resistant pepsin powder to conventional pepsin powder in the artificial digestion method. Proficiency samples of pork diaphragm containing specific numbers of viable Trichinella spiralis larvae were tested in two laboratories. The results revealed that aerosol-resistant pepsin was simple, effective and convenient to use, and showed good solubility and larval recovery that met the requirements of the European Union regulation EU 2015/1375. Overall, the efficacy of the aerosol-resistant pepsin was comparable to the conventional pepsin and safer for analysts.

The parasite biodiversity of mouse opossums in Brazil remains incompletely explored. We describe a new species of Subulura (Ascaridida: Subuluroidea) from the large intestine of the white-bellied woolly mouse opossum, Marmosa constantiae, based on the results of light and scanning electron microscopy (SEM). We also partially sequenced the mitochondrial cytochrome c oxidase I (MT-CO1) gene of the new species, using molecular phylogenetic analyses to determine its relationships within the Subuluroidea superfamily. As molecular data on subuluroid species are extremely limited, few inferences could be drawn from our phylogenies. Our SEM observations showed the detailed morphology of the cephalic extremity, precloacal pseudo-sucker, caudal papillae, phasmids and vulva. Subulura eliseae sp. n. differs from the other four Subulura parasites species of marsupials by the number of caudal papillae and the structure dimensions, and size of the spicule. Moreover, S. eliseae sp. n. has ten pairs of caudal papillae, which is unique compared to other species. We present morphometric and molecular data on this new species, contributing to future studies on subuluroids.

Steinernema populi n. sp. was recovered by baiting from beneath poplar trees in China. Morphological and molecular features provided evidence for placing the new species into the Kushidai clade. The new species is characterized by the following morphological features: third-stage infective juveniles (IJ) with a body length of 1095 (973–1172) μm, a distance from the anterior end to excretory pore of 77 (70–86) μm and a tail length of 64 (55–72) μm. The Body length/Tail length (c) ratio and Anterior end to Excretory pore/ Tail length × 100 (E%) of S. populi n. sp. are substantially greater than those of all other ‘Feltiae–Kushidai–Monticolum’ group members. The first-generation males can be recognized by a spicule length of 66 (57–77) μm and a gubernaculum length of 46 (38–60) μm. The new species is further characterized by sequences of the internal transcribed spacer and partial 28S regions of the ribosomal DNA. Phylogenetic analyses show that Steinernema akhursti and Steinernema kushidai are the closest relatives to S. populi n. sp.

The influence of climate on parasite distribution has been demonstrated in different regions worldwide. Despite its small size, Gran Canaria (Canary Islands, Spain) constitutes a ‘biodiversity laboratory’ due to the huge climatic differences between municipalities. Feral cats may represent a threat to biodiversity due to their predatory behaviour. In addition, they may be a source of pathogens zoonotic to humans. To study the climatic/seasonal influence and prevalence of feral cat parasites throughout the island, a total of 290 stool samples from 29 feral cat colonies were analysed following standard concentration protocols (sodium chloride, formol-ether and zinc sulphate). In total, 13 feline parasitic taxa were found, with the most common species being Ancylostoma spp., which, together with Toxocara spp., Toxoplasma gondii and Giardia spp., are considered a concern for human health. Nematodes were the most common parasites in all areas. Nematodes and protozoans were significantly more prevalent in temperate mild (75.0% and 30.0%) than in dry desert areas (29.3% and 18.7%). In contrast, cestodes were significantly more prevalent in dry desert than in temperate mild areas (26.0% and 13.3%). Only protozoans exhibited statistically significant seasonal patterns, mostly in the wet season. Data reported in this study endorse the usage of small and diverse islands such as Gran Canaria to study the climatic influence on parasitic communities in wild/feral animals. Cat colonies require better management to reduce their threat to endemic wildlife, domestic animals and public health, being invasive species that harbour zoonotic parasites.

Digenetic trematodes are important parasites of humans and animals. They have complex life cycles and typically infect a gastropod as the first intermediate host. Bithynia siamensis goniomphalos, the first intermediate host of the liver fluke, Opisthorchis viverrini, harbours a wide variety of other trematode species. Morphological details of cercariae of 20 trematode taxa from B. s. goniomphalos, collected mainly in Thailand from 2009 to 2014, were provided in an earlier paper. Correct identification to the species or genus level based on morphology of these cercariae is generally not possible. Therefore, we used molecular data to improve identification and to investigate the diversity of the species of trematodes infecting B. s. goniomphalos. We were successful in extracting, amplifying and sequencing portions of the 28S ribosomal RNA (rRNA) gene for 19 of these 20 types of cercaria, and the internal transcribed spacer 2 region for 18 types. BLAST searches in GenBank and phylogenetic trees inferred from the 28S rRNA sequences identified members of at least nine superfamilies and 12 families. Only a few cercariae could be assigned confidently to genus or species on the basis of the sequence data. Matching sequence data from named adult trematodes will be required for definitive identification. There is clearly a great diversity of trematode species utilizing B. s. goniomphalos in Thailand.

The alveolar hydatid disease, also known as alveolar echinococcosis, of humans is certainly one of the most dangerous zoonoses worldwide. The disease is caused by Echinococcus multilocularis – the fox tapeworm. Red foxes (Vulpes vulpes) are currently counted as the most important carriers (reservoirs) of E. multilocularis in the Northern Hemisphere. The possible routes of infection of E. multilocularis to humans are complex and still require research. Until now, it has been unknown whether E. multilocularis eggs can be moved by wind at all. This analysis shows, based on calculations, that E. multilocularis eggs can be transported by wind. Using a mathematical model, flight distances depending on wind speed and take-off heights are calculated for dense and less dense (coniferous) forest areas. The results – differentiated for seasons and as overall average – are based on mean values of wind speeds which were measured over a ten-year period in an experimental forest stand in the Solling (Germany). Due to their rate of descent, wind-related spreading of E. multilocularis eggs is possible. The average flight distance covered by E. multilocularis eggs in forest areas, depending on their starting altitude and wind speed, is between approximately 1.3 m and approximately 17 m. From the mathematical point of view, the wind factor can definitely be seen as one of the multiple vectors associated with environmental contamination by E. multilocularis eggs. Consequently, the possible wind-borne spread of E. multilocularis eggs poses an infection risk to humans that should be considered and requires further research.

During nematode surveys in natural vegetation in Sierra Mágina, Jaén province, southern Spain, a Longidorus species closely resembling Longidorus carpetanensis was found, but application of integrative taxonomic approaches clearly demonstrated that it is a new species described herein as Longidorus maginicus n. sp. The new species is amphimictic, characterized by a moderately long body (4.2–5.2 mm); lip region anteriorly flattened, slightly separated from the rest of body by a depression, 9.0–11.0 μm wide and 3.5–6.0 μm high; amphidial fovea not lobed; relatively short odontostyle (61.0–70.5 μm); guiding ring located 23.5–27.0 μm from anterior end; vulva located at 42.0%–51.3% of body length; female tail 39.0–61.0 μm long, conoid, dorsally convex with rounded terminus (c′ = 1.3–2.1), with two or three pairs of caudal pores; and males common (1:2 ratio males:females), with moderately long spicules (39.0–48.5 μm) and 1 + 6–9 ventromedian supplements and three juvenile developmental stages. According to the polytomous key, codes for the new species are (codes in parentheses are exceptions): A2-B1-C2-D2-E1-F2(3)-G2-H5(4)-I2-J1-K6. The results of molecular analysis of D2–D3 28S, internal transcribed spacer region, partial 18S rDNA, and cytochrome oxidase c subunit 1 (coxI) gene sequences further characterized the new species status, and separated it from L. carpetanensis and other related species.

Trichinellosis in humans is most often caused by the parasite Trichinella spiralis. The clinical course of the disease is diverse and the symptoms can vary from mild to severe. Symptoms usually disappear within a few months, but encysted larvae in the muscles can cause myalgia and weakness that last for years. However, the existence of chronic trichinellosis as a disease is still debatable. This study presents the results obtained at the National Reference Laboratory for Trichinellosis – INEP, Serbia. The study was conducted to assess the immunoserological, biochemical and symptomatic disease parameters of twelve patients who acquired trichinellosis 13 and 18 years ago, respectively. They were involved in two T. spiralis outbreaks in Serbia, at the village of Kumane and the city of Belgrade (nine and three patients, respectively). Results indicated the presence of specific anti-Trichinella antibodies in 83% of the total number of patients. However, while the humoral immune response to Trichinella lasted for more than a decade reaching almost two decades after the acute infection phase (7/9 and 3/3 respectively, in two outbreaks), persistence of chronic muscular pain, as the most prolonged symptom of trichinellosis, could be found in the majority of patients from the Kumane outbreak (7/9). As a consequence, these patients suffered from limitations in daily living activities for the same period of time. The results presented in this paper are our contribution to the view that trichinellosis as a chronic disease with symptoms exists and may be related to the severity of the disease in the acute phase.

Two populations of the species Diastolaimus grossus have been obtained from bark of trees in Bosnia and Herzegovina and the Czech Republic. The species is described and characterized in detail using morphological techniques (light and electron scanning microscopy) and morphometrical (Gower General Similarity coefficient of morphological characters) and molecular analyses, including phylogenetic analysis of all related and already sequenced species of the family Chambersiellidae. Morphological and molecular analyses, based on 18S and 28S ribosomal DNA sequences, show that the family Chambersiellidae is polyphyletic, being the subfamily Chambersiellinae related with Cephalobomorpha and Tylenchomorpha, and the subfamily Macrolaiminae is located into Panagrolaimomorpha. The genus Diastolaimus, previously belonging to Macrolaiminae, is transferred to Chambersiellinae. Diastolaimus mexicanus is proposed as a junior synonym of D. grossus.

To investigate methods for in vitro assessment of anthelmintic efficacy against the chicken nematode Ascaridia galli this study firstly evaluated sample preparation methods including recovery of eggs from excreta using different flotation fluids and induced larval hatching by the deshelling–centrifugation method and the glass-bead method with or without bile. It then evaluated two in vitro assays, the in-ovo larval development assay (LDA) and larval migration inhibition assay (LMIA), for anthelmintic efficacy testing against A. galli using fresh eggs and artificially hatched larvae, respectively. Four anthelmintics, thiabendazole (TBZ), fenbendazole (FBZ), levamisole (LEV) and piperazine (PIP) were employed using an A. galli isolate of known susceptibility. The results suggested that the LDA and LMIA could successfully be used to generate concentration response curves for the tested drugs. The LDA provided EC50 values for inhibition of egg embryonation of 0.084 and 0.071 μg/ml for TBZ and FBZ, respectively. In the LMIA, the values of effective concentration (EC50) of TBZ, FBZ, LEV and PIP were 105.9, 6.32, 349.9 and 6.78 × 107 nM, respectively. For such in vitro studies, a saturated sugar solution showed high egg recovery efficiency (67.8%) and yielded eggs of the highest morphological quality (98.1%) and subsequent developmental ability (93.3%). The larval hatching assays evaluated did not differ in hatching efficiency but the deshelling–centrifugation method yielded larvae that had slightly better survival rates. For final standardization of these tests and establishment of EC50 reference values, tests using isolates of A. galli of defined resistance status need to be performed.

The heterogeneous landscape of Nicaragua harbours a large diversity of freshwater fishes. The great Nicaraguan lakes, Managua and Nicaragua, and several adjacent crater lakes harbour numerous endemic fish species. However, information about their parasite fauna is still fragmentary. Here, we surveyed the great Nicaraguan lakes and four crater lakes and provide data for 17 metazoan parasite taxa infecting seven fish host species. We also gathered all the published records from the literature on the parasites reported from Nicaraguan freshwater fishes, as well as those for Costa Rica and Panama to discuss the region of Lower Central America as a whole. With this information we built a parasite–host and a host–parasite checklist. With data from near 50% of the native and endemic freshwater fishes in Nicaragua, the parasite fauna comprises 101 taxa in 51 fish species allocated in 11 families. Cichlids are the most diverse group of fishes in this region and have been the most extensively surveyed for their metazoan parasites. Helminths are the best-represented groups of metazoan parasites, with 42 trematodes, five cestodes, 24 monogeneans, two acanthocephalans, 20 nematodes and one hirudinean. Additionally, freshwater fishes are parasitized by copepods, branchiurans and oribatid mites. Even though the inventory is not yet complete, the patterns of diversity uncovered revealed promising information about the origin, biogeography and evolutionary history of the Nicaraguan freshwater fish parasite fauna. More studies are necessary to complete our knowledge about the diversity, host association and distribution of metazoan parasites in Nicaragua and other Central American countries.

For decades, it has remained unclear how the Asian swim bladder nematode Anguillicola crassus was able to supplant the previously stable population of its relative from New Zealand Anguillicola novaezelandiae in the Lake Bracciano, Italy. Previously, researchers have hypothesized that A. crassus possesses an ecological advantage due to a more efficient life cycle in combination with a pattern of unidirectional hybridization between A. novaezelandiae females and A. crassus males. The present study focuses on the viability of hybrid offspring and their allelic pattern, particularly in developed adult stages of the hybrid F1 generation. While the percentages of hybrid individuals from A. novaezelandiae mothers and A. crassus fathers increased from egg to adult stages, it was more distinct in egg stages of A. crassus females and A. novaezelandiae males, but did not occur in adult F1 individuals at all. Therefore, we corroborate the hypothesis of unidirectional hybridization by differentiating between egg and adult stages, and suggest this as another explanatory factor for the extinction of A. novaezelandiae in Lake Bracciano in Italy and the predominance of A. crassus.

Regardless of the highly efficient anthelmintics available and the control measures taken by dog owners and veterinarians, gastrointestinal parasites, especially zoonotic helminths, are still abundant in dogs and pose a health risk to humans. Free-ranging dogs in rural areas can be an important source of helminth infection. The aims of the present work were to collect scats of rural dogs, determine the environmental contamination caused by helminth infections among rural dogs of Western Estonia, analyse how diet affects helminth infection rate and compare the findings to a previous study focusing on dog helminths in urban areas of Estonia. To differentiate the scats of dogs from other sympatric canids, a genetic method was applied. Of 328 samples, genetic analysis identified 84 scats belonging to dogs, of which 87.0% were infected with helminths. A high proportion of rural dog scats harboured eggs of Taeniidae (65.5%), followed by Trichuris spp./Eucoleus spp. (15.5%), Uncinaria stenocephala (14.7%) and Toxocara canis (4.3%). Coinfections occurred in 34.5% of the samples, being the most common between Taeniidae and U. stenocephala (41.4%). The intensity model indicated higher helminth infection rate in rural dogs preying on rodents and game. In comparison to urban dogs, rural dogs were nine times more likely to be infected with intestinal parasites. These results emphasize the need to implement measures to reduce helminth infections in dogs living in rural areas of Western Estonia. Among a complex of measures to be taken, we suggest that it is also important to diagnose which gastrointestinal parasite species infect dogs to determine specific anthelmintic treatment against these parasites.