INTRODUCTION
Nowadays, Clonorchis sinensis is the most important helminth in aspects of the prevalence and clinical significance in the Republic of Korea (=Korea). The endemicity of this trematode infection, clonorchiasis, has maintained at relatively high levels in riverside areas [1–8]. Seo et al. [1] reported the prevalence of C. sinensis infection among the riverside residents in 7 major rivers, i.e., Nakdong-gang (River) (40.2%), Youngsan-gang (30.8%), Seomjin-gang (17.3%), Tamjin-gang (15.9%), Han-gang (15.7%), Gum-gang (12.0%), and Mangyeong-gang (8.0%). In 2008, Cho et al. [5] reported the egg positive rates of residents living in 4 major river basins, Nakdong-gang (17.1%), Seomjin-gang (11.2%), Youngsan-gang (5.5%), and Gum-gang (4.6%), located in southern parts of Korea. Recently, the prevalence of C. sinensis infection in adjacent residents of 5 major rivers, i.e., Nakdong-gang (11.7%), Seomjin-gang (9.9%), Gum-gang (6.5%), Youngsan-gang (3.1%), and Han-gang (1.0%), were reported by Jeong et al. [8]. By the nationwide survey on helminthic infections in Korea (2012), the prevalence of clonorchiasis was 1.9%, and 932,540 residents with this endemic disease were estimated to be infected, which is the highest value among the prevalences of helminthiasis in Korea [7].
On the other hand, freshwater fish, the source of infections, collected from various endemic riverside areas were examined by many Korean workers to estimate the endemicities of clonorchiasis. More than 48 fish species have been reported in Korea as the second intermediate hosts of C. sinensis [9–15]. Especially, in 2008, Kim et al. [12] widely surveyed 677 freshwater fish (21 species) from 34 localities to know the infection status with C. sinensis metacercariae (CsMc) in Korea. They examined fish from 2 sites, Imsil-gun (29 fish of 4 species), Jeollabuk-do, and Gokseong-gun (45 fish of 10 species), Jeollanam-do, as the fish from water systems of Seomjin-gang. In 2011, Cho et al. [14] investigated the infection status of CsMc in freshwater fish from 3 wide regions, which were tentatively divided by the latitude levels of the Korean peninsula. They also examined fish from 2 sites, Gokseong-gun (191 fish of 22 species) and Gurye-gun (68 fish of 14 species), Jeollanam-do, as the fish from Seomjin-gang. Cho et al. [15] surveyed on the prevalence of zoonotic trematode metacercariae in freshwater fish from Gangwon-do, Korea. Recently, Sohn et al. [16] investigated the infection status of trematode metacercariae, including C. sinensis, in freshwater fish from the water systems of Hantan-gang and Imjin-gang located in relatively northern regions of Korea. However, freshwater fish from the water systems of Seomjin-gang has not been widely and systematically examined. It is seriously needed to reveal the infection status of CsMc in fish from water systems of Seomjin-gang. Therefore, we performed the present study to investigate the prevalence of CsMc in freshwater fish from 7 local sites of Seomjin-gang.
MATERIALS AND METHODS
Fish collection
We collected total 1,604 freshwater fish from 7 local sites of Seomjin-gang, i.e., Osucheon (“cheon” means stream) (latitude: 38.43268; longitude: 127.4375) in Imsil-gun, Seomjin-gang (38.23047; 127.2179) in Sunchang-gun, Songdaecheon (38.23047; 127.2179) in Namwon-si, Jeollabuk-do, Seomjin-gang (38.23047; 127.2179) in Gokseong-gun, Seomjin-gang (37.94375; 127.07142) in Gurye-gun, Jeollanam-do, and Hoengcheon (38.0855; 127.07264) and Namsancheon (38.01408; 127.2088) in Hadong-gun, Gyeongsangnam-do, for 5 years (2012–2016) (Fig. 1).
Fish examined from upper reaches of Seomjin-gang
Total 236 freshwater fish in 11 species (no. of fish examined) were collected from Osucheon in Imsil-gun, Jeollabuk-do (2012 and 2013), which included Zacco platypus (80), Hemibarbus labeo (56), Squalidus chankaensis tsuchigae (35), Opsariichthys uncirostris amurensis (21), Squalidus japonicus coreanus (20), Microphysogobio jeoni (6), Acanthorhodeus gracilis (6), Pseudogobio esocinus (5), Carassius auratus (3), Rhodeus ocellatus (3), and Hemibarbus longirostris (1).
Total 310 freshwater fish in 29 species were collected from Seomjin-gang in Sunchang-gun, Jeollabuk-do (2014 and 2015), which included Pungtungia herzi (56), Z. platypus (51), P. esocinus (32), Coreoperca herzi (19), Zacco koreanus (15), O. uncirostris amurensis (14), C. auratus (14), Acheilognathus koreensis (12), Sarcocheilichthys variegatus wakiyae (12), S. japonicus coreanus (11), Acheilognathus rhombeus (10), Siniperca scherzeri (10), Odontobutis platycephala (9), Squalidus gracilis majimae (6), A. gracilis (6), Gnathopogon strigatus (5), H. longirostris (5), H. labeo (4), Coreoleuciscus splendidus (4), Plecoglossus altivelis (3), Acheilognathus yamatsutae (3), Abbottina springeri (3), Cyprinus carpio (2), Cobitis tetralineata (2), Acheilognathus lanceolatus (1), Acanthorhodeus macropterus (1), Hemiculter leucisculus (1), Cobitis lutheri (1), and Misgurnus anguillicaudatus (1).
Fish examined from middle reaches of Seomjin-gang
Total 196 freshwater fish in 14 species collected from Songdaecheon in Namwon-si, Jeollabuk-do were examined in 2012 and 2013. They included Z. koreanus (57), P. herzi (47), Z. platypus (46), Rhynchocypris oxycephalus (21), C. herzi (6), Microphysogobio koreensis (5), C. auratus (3), G. strigatus (2), S. gracilis majimae (2), Pseudorasbora parva (2), A. springeri (2), P. esocinus (1), Iksookimia longicorpus (1), and Lepomis macrochirus (1).
Total 289 freshwater fish in 15 species collected from Seomjin-gang in Gokseong-gun, Jeollanam-do were examined in 2015 and 2016. They included Z. koreanus (52), Z. platypus (43), C. splendidus (37), S. variegatus wakiyae (31), S. japonicus coreanus (27), P. herzi (22), H. longirostris (20), P. esocinus (16), M. koeensis (16), C. herzi (11), Acheilognathus majusculus (6), P. altivelis (3), H. labeo (2), O. uncirostris amurensis (2), and Ladislabia taczanowskii (1).
Fish examined from lower reaches of Seomjin-gang
Total 183 freshwater fish in 21 species were collected from Seomjin-gang in Gurye-gun, Jeollanam-do (2014), which included Z. platypus (24), P. herzi (21), Z. koreanus (14), S. japonicus coreanus (15), Sarcocheilichthys nigripinnis morii (14), O. uncirostris amurensis (13), A. rhombeus (11), C. herzi (10), H. labeo (10), S. gracilis majimae (8), M. koreensis (6), C. auratus (5), A. rivularis (5), A. gracilis (5), O. platycephala (4), P. parva (3), P. altivelis (3), A. lanceolatus (3), H. longirostris (1), and S. scherzeri (1).
Total 156 freshwater fish in 13 species were collected from Hoengcheon in Hadong-gun, Gyeongsangnam-do in 2014 and 2016, which included Z. koreanus (35), A. koreensis (27), Z. platypus (27), P. herzi (16), C. herzi (11), Zacco temminckii (10), H. longirostris (8), P. esocinus (8), A. springeri (7), C. splendidus (4), S. gracilis majimae (1), C. auratus (1), and O. platycephala (1).
Total 234 freshwater fish in 12 species were collected from Namsancheon in Hadong-gun, Gyeongsangnam-do in 2015 and 2016, which included Z. koreanus (57), P. esocinus (46), Z. platypus (41), P. herzi (35), C. auratus (16), H. longirostris (14), Z. temminckii (11), A. koreensis (7), S. gracilis majimae (3), A. gracilis (2), C. splendidus (1), and O. platycephala (1).
Examination methods
All collected fish with ice were transferred to the laboratory of the Department of Parasitology and Tropical Medicine, Gyeongsang National University School of Medicine, Jinju, Korea. After identification of fish species, they were individually ground with a mortar or a grinder. Each ground fish meat was mixed with artificial gastric juice, and the mixture was incubated at 36°C for 2 hr. The digested material was filtered with 1×1 mm of mesh, and washed with 0.85% saline until the supernatant became clear. The sediment was carefully examined under a stereomicroscope. Each species of zoonotic trematode metacercariae were separately collected by the general feature [1,2], and they were counted to get hold of infection rates (%) and densities (no. of metacercariae per infected fish) by fish species.
RESULTS
Infection status of fish from upper reaches of Seomjin-gang
The metacercariae of C. sinensis were detected in 102 (39.8%) out of 256 fish in 14 species from the upper reaches of Seomjin-gang, i.e., Osucheon in Imsil-gun, and Seomjin-gang in Sunchang-gun, Jeollabuk-do. Their average density was 9.0 per fish infected. Among 236 freshwater fish in 11 species collected from Osucheon, 33 (14.0%) in 6 species (54.6%), i.e., Z. platypus, S. chankaensis tsuchigae, O. uncirostris amurensis, M. jeoni, P. esocinus, and H. longirostris, were infected with CsMc. Total 69 (22.3%) freshwater fish in 9 species (31.0%), i.e., P. herzi, S. variegatus wakiyae, S. japonicus coreanus, A. rhombeus, S. gracilis majimae, H. longirostris, H. labeo, A. yamatsutae, and H. leucisculus, out of 310 fish in 29 species collected from Seomjin-gang in Sunchang-gun were infected with CsMc. Their infection status with CsMc by the fish species and surveyed areas was shown in Table 1.
Infection status of fish from middle reaches of Seomjin-gang
The metacercariae of C. sinensis were detected in 132 (48.0%) out of 275 fish in 12 species from the middle reaches of Seomjin-gang, i.e., Songdaecheon in Namwon-si, Jeollabuk-do, and Seomjin-gang in Gokseong-gun, Jeollanam-do. Their average density was 21.0 per infected fish. Among 196 freshwater fish in 14 species collected in Songdaecheon, 33 fish (16.8%) in 4 species (28.6%), i.e., P. herzi, M. koreensis, G. strigatus, and S. gracilis majimae, were infected with CsMc. Total 99 (34.3%) freshwater fish in 10 species (66.7%), i.e., Z. platypus, C. splendidus, S. variegatus wakiyae, S. japonicus coreanus, P. herzi, H. longirostris, M. koeensis, P. esocinus, A. majusculus, and L. taczanowskii, out of 289 fish in 15 species collected from Seomjin-gang in Gokseong-gun were infected with CsMc. The infection status with CsMc by the fish species and surveyed areas are shown in Table 2.
Infection status of fish from lower reaches of Seomjin-gang
The metacercariae of C. sinensis were detected in 77 (56.6%) out of 136 fish in 11 species from the lower reaches of Seomjin-gang, i.e., Seomjin-gang in Gurye-gun, Jeollanam-do, and Namsancheon in Hadong-gun, Gyeongsangnam-do. Their average density was 64.9 per infected fish. Among 183 freshwater fish in 21 species collected in Seomjin-gang in Gurye-gun, 74 fish (40.4%) in 11 species (52.4%), i.e., P. herzi, S. japonicus coreanus, S. nigripinnis morii, A. rhombeus, H. labeo, S. gracilis majimae, M. koreensis, A. rivularis, A. gracilis, P. parva, P. altivelis, and A. lanceolatus, were infected with CsMc. No metacercariae were detected in total 156 freshwater fish in 13 species collected from Hoengcheon in Hadong-gun, Gyeongsangnam-do. Total 339 C. sinensis metacercariae were found in only 3 P. herzi among 234 freshwater fish in 12 species collected from Namsancheon in Hadong-gun, Gyeongsangnam-do. Their infection status by the fish species and surveyed areas are shown in Table 3.
Infection status of Metorchis orientalis metacercariae in fish from Seomjin-gang
The metacercariae of M. orientalis were detected in fish from Seomjin-gang in Sunchang-gun, Songdaecheon in Namwon-si, Seomjin-gang in Gurye-gun, and Hoengcheon in Hadong-gun. Their infection status by the fish species and surveyed areas are shown in Table 4.
DISCUSSION
By the present study, it has been confirmed that CsMc are more or less prevalent in fish from Seomjin-gang. The total positive rate, 19.4%, and average metacercarial density, 27.9 per infected fish, were not so high (Table 5). However, the infection status was revealed with a certain trend in positive fish groups according to the surveyed localities tentatively divided; upper, middle, and lower reaches. The prevalences were 39.8%, 48.0%, and 56.6%, and metacercarial densities were 9.0, 21.0, and 64.9 per fish infected, respectively. Metacercarial densities in fish from upper to lower reaches, i.e., Osucheon in Imsil-gun, Seomjin-gang in Sunchang-gun, Songdaecheon in Namwon-si, Seomjin-gang in Gokseong-gun, and Seomjin-gang in Gurye-gun, were 4.4, 11.1, 16.7, 22.5, and 63.0 per infected fish, respectively. These findings suggested that the endemicity of CsMc is much higher in fish from lower reaches of Seomjin-gang.
The sampling of fish is one of the important factors in the successful metacercarial surveys to determine the epidemiology of trematode infections. Total 1,604 fish were collected from 7 localities in the water systems of Seomjin-gang in this study. The fish were tentatively grouped according to the surveyed areas, those of upper (546 fish: 34.0%), middle (485: 30.2%), and lower (573: 35.7%) reaches. The number of fish examined in each of these 3 reaches was relatively even. The pale chub (312: 19.5%), Z. platypus, was the most dominant fish species, and followed by the Korean chub (230: 14.3%), Z. koreanus, and the striped shinner (197: 12.3%), P. herzi, among the fish species examined in this study. The numbers of the pale chub examined in 3 reaches were 131 (upper: 42.0%), 89 (middle: 28.5%), and 92 (lower: 29.5%). Those of the Korean chub were 15 (6.5%), 109 (47.4%), and 106 (46.1%), and of the striped shinner were 56 (28.4%), 69 (35.0%), and 72 (36.6%). According to the above findings, the pale chub was collected more in the upper than in the middle and lower reaches, The Korean chub was collected more in the middle and lower reaches than in the upper reaches, and the striped shinner was relatively evenly collected in 3 reaches.
Total 48 fish species (in 34 genera, 7 families) have been reported as the second intermediate hosts of C. sinensis in Korea [9–15]. In the present study, CsMc were found in 24 fish species, i.e., A. rivularis, A. gracilis, A. lanceolatus, A. majuscules, A. rhombeus, A. yamatsutae, C. splendidus, G. strigatus, H. labeo, H. longirostris, H. leucisculus, L. taczanowskii, M. koreensis, M. jeoni, O. uncirostris, P. esocinus, P. herzi, P. parva, S. nigripinnis morii, S. variegates wakiyae, S. chankaensis tsuchigae, S. gracilis majimae, S. japonicus coreanus, and Z. platypus. They all had been listed as the second intermediate hosts of C. sinensis in Korea [9–15]. The striped shinner was used as an index fish species to compare the endemicities of C. sinensis infection in 3 reaches in the present study. However, the metacercarial positive rates (prevalence) in this fish species were 80.4%, 72.5%, and 30.6%, whereas the metacercarial densities were 15.2, 46.1, and 175.9 per infected fish in the upper, middle, and lower reaches, respectively (Table 5). In the distribution pattern of CsMc in another susceptible fish, P. parva, the prevalence is proportionally correlated with the metacercarial density in certain areas [17–20]. This reverse phenomenon is questionable, and it should be solved in the near future.
Kim et al. [12] detected CsMc in 3 fish species, P. parva, Acheilognathus intermedia and Odontobutis interrupta, from Imsil-gun, Jeollabuk-do and also in 3 fish species, P. parva, Zacco temminckii, and O. interrupta, from Gokseong-gun, Jeollanam-do in the examination of the fish from water systems of Seomjin-gang (total 74 fish of 11 species). Their infection rates and densities were not so high except for P. parva from Imsil-gun, Jeollabuk-do, when we compared them with our findings of Imsil-gun and Gokseong-gun. Fish from Seomjin-gang in Gokseong-gun (191 fish in 22 species) and Gurye-gun (68 fish in 14 species), Jeollanam-do were also examined in Cho et al. [14]. The prevalences were 35.3% and 66.1%, and densities were 59.4 and 37.6 per fish infected in fish from above 2 regions, respectively. However, the prevalences were 45.2% and 73.3%, and densities were 22.5 and 63.0 per fish infected in this study. From the above comparisons, it was confirmed that the metacercarial endemicity in fish from Gurye-gun is much higher in this study than in Cho et al. [14].
According to recent studies, CsMc were found in fish from the water system of Imjin-gang and Hantan-gang located in the northern part of Korea, although their prevalences and densities were very low [14–16]. However, CsMc were not detected in fish from the water systems of Han-gang in Gangwon-do [14,15]. In fish from Geum-gang, the prevalence and density of CsMc were also revealed in low levels [14]. They showed moderate levels in fish from Yeongsan-gang and Seomjin-gang in Jeollanam-do [14] like those in this study. However, fish from Tamjin-gang in Gangjin-gun, Jeollanam-do and Nakdong-gang in 3 localities of Gyeongsangbuk-do, and Sancheong-gun, Gyeongsangnam-do were highly infected with CsMc [14]. By the present study, it has been confirmed that CsMc are more or less prevalent in fish from some water systems of Seomjin-gang in Korea.
