OBJECTIVE: To evaluate the performance of recombinase-aided amplification (RAA) assay in detection of Clonorchis sinensis metacercariae in freshwater fish samples, so as to provide insights into standardization and field application of this assay. METHODS: Wild freshwater fish samples were collected in the rivers of administrative villages where C. sinensis-infected residents lived in Jiangyan District, Xinghua County and Taixing County of Taizhou City, Jiangsu Province from June to September 2022. Genomic DNA was extracted from six freshwater fish specimens (5 g each) containing 0, 1, 2, 4, 8 and 16 C. sinensis metacercariae for fluorescent RAA assay, and the diagnostic sensitivity was evaluated. Fluorescent RAA assay was performed with genomic DNA from C. sinensis, Metorchis orientalis, Haplorchis pumilio and Centrocestus formosanus metacercariae as templates to evaluate its cross-reactions. In addition, the detection of fluorescent RAA assay and direct compression method for C. sinensis metacercariae was compared in field-collected freshwater fish samples. RESULTS: Positive amplification was found in fresh-water fish specimens containing different numbers of C. sinensis metacercariae, and fluorescent RAA assay was effective to detect one C. sinensis metacercaria in 5 g freshwater fish specimens within 20 min. Fluorescent RAA assay tested negative for DNA from M. orientalis, H. pumilio and C. formosanus metacercariae. Fluorescent RAA assay and direct compression method showed 5.36% (93/1 735) and 2.88% (50/1 735) detection rates for C. sinensis metacercariae in 1 735 field-collected freshwater fish samples, with a statistically significant difference seen (χ² = 478.150, P < 0.001). There was a significant difference in the detection of C. sinensis metacercariae in different species of freshwater fish by both the direct compression method (χ² = 11.20, P < 0.05) and fluorescent RAA assay (χ² = 20.26, P < 0.001), and the detection of C. sinensis metacercariae was higher in Pseudorasbora parva than in other fish species by both the direct compression method and fluorescent RAA assay (both P values < 0.05). CONCLUSIONS Fluorescent RAA assay has a high sensitivity for detection of C. sinensis metacercariae in freshwater fish samples, and has no cross-reactions with M. orientalis, H. pumilio or C. formosanus metacercariae. Fluorescent RAA assay shows a higher accuracy for detection of C. sinensis infections in field-collected freshwater fish than the direct compression method.
Animals ; Clonorchis sinensis/genetics* ; Metacercariae/genetics* ; Recombinases ; Fresh Water ; Fishes ; DNA ; Fish Diseases/diagnosis*

Ecdysis is a common phenomenon that happens throughout the life phase of the giant freshwater prawn Macrobrachium rosenbergii. It is vital to better understand the correlation between cannibalism and biochemical compound that exists during the moulting process. The objective of the present study was to determine the amino acid profile released by M. rosenbergii during the ecdysis process that promotes cannibalism. To accomplish this, changes in amino acid levels (total amino acid (TAA) and free amino acid (FAA)) of tissue muscle, exoskeleton, and sample water of culture medium from the moulting (E-stage) and non-moulting (C-stage) prawns were analysed using high-performance liquid chromatography (HPLC). Comparison study revealed that among the TAA compounds, proline and sarcosine of tissues from moulting prawn were found at the highest levels. The level of FAA from water that contains moulting prawns (E-stage) was dominated by tryptophan and proline. Significant values obtained in the present study suggested that these amino acid compounds act as a chemical cue to promote cannibalism in M. rosenbergii during ecdysis. The knowledge of compositions and compounds that were released during the moulting process should be helpful for better understanding of the mechanism and chemical cues that play roles on triggering cannibalism, and also for future dietary manipulation to improve feeding efficiencies and feeding management, which indirectly impacts productivity and profitability.
Amino Acids/chemistry* ; Animals ; Aquaculture ; Cannibalism ; Chromatography, High Pressure Liquid ; Feeding Behavior ; Fresh Water ; Malaysia ; Molting ; Palaemonidae/physiology* ; Proline/chemistry* ; Tryptophan/chemistry*

Freshwater snail is an important biological group in aquatic ecosystem and an intermediate host of many parasites. Intestinal flora plays an important role in animal energy metabolism and resistance to pathogens. We analyzed the intestinal microbiota diversity of Radix auricularia (RA) and Planorbella trivolvis (PL) by 16S rRNA high-throughput sequencing. At the phylum level, RA had 23 phyla, including Proteobacteria (33.63%), Cyanobacteria (15.33%), Chloroflexi (13.95%), and Actinomycetes (12.99%). PL had 13 phyla, including Proteobacteria (54.88%), Bacteroidetes (28.49%), and Actinomycetes (7.65%). At the genus level, there were 445 genera in RA, including Pleurocapsa, Thiodictyon, Leptotrichia, and Nocardioides. There were 238 genera in PL, including Cloacibacterium, OM60NOR5_clade, Pseudomonas, and Rhodobacter. Ninety-three genera were the common core flora of the two snail species (all the samples were present), and 27 genera had an abundance greater than 0.5%. The structure of intestinal microbiota was significantly different between the two groups (P=0.027). We performed the functional prediction of intestinal microbiota using Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt), and the results show that the KEGG functional composition of the intestinal flora of the two snails was similar, and the abundance of the amino acid metabolism, carbohydrate metabolism and membrane transport were large. In summary, the intestinal microbiota of the two snails was high in diversity and significantly different, but there were a large number of common core flora.
Animals ; Auricularia ; Ecosystem ; Fresh Water ; Gastrointestinal Microbiome/genetics* ; High-Throughput Nucleotide Sequencing ; Phylogeny ; RNA, Ribosomal, 16S/genetics* ; Snails

A 46-year old man visited our outpatient clinic with complaint of foreign body sensation in throat after consuming raw freshwater fish 5 days ago. Laryngoscopic examination revealed a motile worm attached on posterior pharyngeal wall. The worm was removed using biopsy forceps under transnasal endoscopy and evidently identified as Clinostomum complanatum after microscopic examination. Patient’s subjective foreign body sensation of throat and hyperemia of laryngeal mucosa remained for approximately 2 weeks post-removal, which were eventually resolved after administration of non-steroidal anti-inflammatory drug and anti-refluxant drug for 2 weeks. Treatment was ended at three weeks since the first visit. C. complanatum infections in humans are rare, and only four cases have been reported in Korea. Symptoms resembling pharyngitis or laryngitis occurs by consumption of raw, infected freshwater fish and treatment is done by mechanically removing the parasite.
Ambulatory Care Facilities ; Biopsy ; Endoscopy ; Foreign Bodies ; Fresh Water ; Humans ; Hyperemia ; Korea ; Laryngeal Mucosa ; Laryngitis ; Parasites ; Pharyngitis ; Pharynx ; Sensation ; Surgical Instruments

The infection status with Clonorchis sinensis metacercariae (CsMc) was examined in freshwater fishes from Yangcheon (a branch of Gyeongho-gang), which is located in Sancheong-gun, Gyeongsangnam-do, the Republic of Korea. Total 2,201 fishes in 26 species were examined by the artificial digestion method through 7 years. CsMc were detected in 1,171 (53.2%) fishes in 21 spp. (80.8%) and their density was 85 per fish infected. Total 532 (99.6%) out of 534 Pungtungia herzi (index fish) examined were infected with 147 CsMc per fish infected. Metacercarial densities in this fish were highest in 2015 (179 CsMc), followed by 2012 (168), 2013 (152), 2016 (145), 2014 (114), and 2017 (89) respectively. In the gobioninid fish group, i.e., P. herzi, Sarcocheilichthys spp., Squalidus spp., Pseudogobio esocinus, Hemibarbus longirostris, and Hemibarbus labeo, 841 (92.7%) fishes were infected with 117 CsMc per fish infected. Total 250 (54.7%) acheilognathinid fish (bitterlings), Acheilognathus spp. and Acanthorhodeus spp. were infected with 5.8 CsMc. In the rasborinid fish (chubs) group, i.e., Zacco platypus, Zacco temminckii, and Zacco koreanus, 77 (13.7%) out of 563 fish examined were infected with 2.4 CsMc in average. The susceptibility indices of CsMc were 49.09 in the overall positive fish group, 104.15 in the gobioninid group, 3.17 in the acheilognathinid group and 0.35 in the rasborinid fish group respectively. Only 1 CsMc was detected in 3 fish species, Coreoperca herzi, Channa argus, and Lepomis macrochirus, respectively. Conclusively, it was confirmed that CsMc are moderately prevalent in fishes from Yangcheon in Sancheon-gun, Gyeongsangnam-do, Korea.
Clonorchis sinensis ; Cyprinidae ; Digestion ; Fishes ; Fresh Water ; Gyeongsangnam-do ; Korea ; Metacercariae ; Methods ; Platypus ; Republic of Korea

Primary amebic encephalitis (PAM) is a devastating central nervous system infection caused by Naegleria fowleri, a free-living amoeba, which can survive in soil and warm fresh water. Here, a 43-year-old healthy male was exposed to warm freshwater 5 days before the symptom onset. He rapidly developed severe cerebral edema before the diagnosis of PAM and was treated with intravenous conventional amphotericin B while died of terminal cerebral hernia finally. Comparing the patients with PAM who has similar clinical symptoms to those with other common types of meningoencephalitis, this infection is probably curable if treated early and aggressively. PAM should be considered in the differential diagnosis of purulent meningoencephalitis, especially in patients with recent freshwater-related activities during the hot season.
Adult ; Amoeba ; Amphotericin B ; Brain Edema ; Central Nervous System Infections ; Central Nervous System Protozoal Infections ; Diagnosis ; Diagnosis, Differential ; Encephalitis ; Encephalocele ; Fresh Water ; Humans ; Male ; Meningoencephalitis ; Naegleria fowleri ; Seasons ; Soil

The infection status of digenetic trematode metacercariae (DTM) was investigated in fishes from 2 representative visiting sites of migratory birds in Gyeongsangnam-do, the Republic of Korea (Korea). A totaly 220 freshwater fishes (7 species) were collected from Junam-jeosuji (reservoir), and 127 fishes (7 species) were also collected from Woopo-neup (swamp) in June and October 2017. As the control group, total 312 fish (22 spp.) from Yangcheon in Sancheong-gun, Gyeongsangnam-do were also collected in June and October 2017. All fishes collected in 3 sites were examined with the artificial digestion method. In the fishes from Junam-jeosuji, more than 4 species, i.e., Clonorchis sinensis, Echinostoma spp., Diplostomum spp. and Cyathocotyle orientalis, of DTM were detected and their endemicy was very low, 0.70. More than 6 species, i.e., C. sinensis, Echinostoma spp., Metorchis orientalis, Clinostomum complanatum, Diplostomum spp. and C. orientalis, of DTM were found in the fishes from Woopo-neup, and their endemicy was low, 5.16. In the fishes from Yangcheon, more than 8 species, i.e., C. sinensis, Metagonimus spp., Centrocestus armatus, C. complanatum, C. orientalis, M. orientalis, Echinostoma spp., and Diplostomum spp., of DTM were detected, and their endemicity was relatively high, 95.48. The percentages of avian trematode metacercariae (ATM) were 99.6% and 94.7% in fishes from Junam-jeosuji and Woopo-neup whereas it was 74.1% in the control site, Yangcheon. The above findings suggested that migratory birds partly affect in endemicity of DTM in the fish in the 2 visiting sites in Gyeongsangnam-do, Korea.
Birds ; Clonorchis sinensis ; Digestion ; Echinostoma ; Fishes ; Fresh Water ; Gyeongsangnam-do ; Heterophyidae ; Korea ; Metacercariae ; Methods ; Republic of Korea

Freshwater snails, as the first intermediate hosts of trematodes, can cause health hazards in animals and humans. Recently, the World Health Organization has included Iran in a list of 6 countries known to have serious problems with fascioliasis. In addition, cercarial dermatitis is a job-related disease that is seen often in paddy workers, agricultural labourers, and fishermen in Iran, particularly in Mazandaran Province. Many studies have been conducted in Iran to survey larval trematodes in freshwater snails. However, to the best of our knowledge, no comprehensive data exist regarding infections in gastropods. Therefore, the aim of the present study was to estimate the types and prevalence of cercarial infections in snails in Iran. Electronic English-language and Persian-language databases were searched to identify 24 published articles reporting the prevalence of trematode infections in snails (9 species from 6 families) in various provinces of Iran. In total, 4.4% of gastropods were infected with the larval stages of trematodes. According to the studies reviewed in this meta-analysis‚ the highest infection prevalence was found in Radix auricularia (9.9%). Twelve larval species of trematodes were identified, and the highest prevalence of cercariae was found for Echinostomatidae cercariae (4.3%). Among the provinces explored, West Azerbaijan had the highest prevalence of infected snails (16.9%). The presence of trematodes in snails could pose a serious health problem in Iran. Thus, further studies are necessary to characterize these infections in other provinces.
Animals ; Azerbaijan ; Cercaria ; Dermatitis ; Echinostomatidae ; Farmers ; Fascioliasis ; Fresh Water ; Gastropoda ; Humans ; Iran ; Prevalence ; Snails ; Trematode Infections ; World Health Organization

Freshwater snails, as the first intermediate hosts of trematodes, can cause health hazards in animals and humans. Recently, the World Health Organization has included Iran in a list of 6 countries known to have serious problems with fascioliasis. In addition, cercarial dermatitis is a job-related disease that is seen often in paddy workers, agricultural labourers, and fishermen in Iran, particularly in Mazandaran Province. Many studies have been conducted in Iran to survey larval trematodes in freshwater snails. However, to the best of our knowledge, no comprehensive data exist regarding infections in gastropods. Therefore, the aim of the present study was to estimate the types and prevalence of cercarial infections in snails in Iran. Electronic English-language and Persian-language databases were searched to identify 24 published articles reporting the prevalence of trematode infections in snails (9 species from 6 families) in various provinces of Iran. In total, 4.4% of gastropods were infected with the larval stages of trematodes. According to the studies reviewed in this meta-analysis‚ the highest infection prevalence was found in Radix auricularia (9.9%). Twelve larval species of trematodes were identified, and the highest prevalence of cercariae was found for Echinostomatidae cercariae (4.3%). Among the provinces explored, West Azerbaijan had the highest prevalence of infected snails (16.9%). The presence of trematodes in snails could pose a serious health problem in Iran. Thus, further studies are necessary to characterize these infections in other provinces.
Animals ; Azerbaijan ; Cercaria ; Dermatitis ; Echinostomatidae ; Farmers ; Fascioliasis ; Fresh Water ; Gastropoda ; Humans ; Iran ; Prevalence ; Snails ; Trematode Infections ; World Health Organization

There are several diagnostic findings required for confirming a postmortem diagnosis of drowning. However, postmortem diagnosis of drowning remains challenging for forensic pathologists. In previous reports, several biochemical tests using various body fluids have been studied for their potential use in the postmortem diagnosis of drowning. In this study, the concentration of sodium and chloride was tested in various postmortem body fluids (vitreous humor, sphenoid sinus fluid, pleural fluid, cerebrospinal fluid, etc.) and their results were interpreted for their potential use in postmortem diagnosis of drowning. We examined 67 autopsy cases (freshwater drowning, 12 cases; seawater drowning, 16 cases; control group, 39 cases). The sodium and chloride concentration in the vitreous humor, sphenoid sinus fluid, and pleural fluid significantly correlated with each other. Furthermore, the concentrations of sodium, chloride, and the sum of the concentrations of the two in the various postmortem body fluids were significantly different in the three groups, when compared with each other (generally the concentration being the highest in the seawater drowning group, followed by the control group and the freshwater drowning group). Biochemical tests using various postmortem body fluids may serve as useful indicators for the postmortem diagnosis of drowning and for the differential diagnosis between freshwater and seawater drowning.
Autopsy ; Biochemistry ; Body Fluids ; Cerebrospinal Fluid ; Diagnosis ; Diagnosis, Differential ; Drowning ; Fresh Water ; Seawater ; Sodium ; Sphenoid Sinus ; Vitreous Body