ISSN (rint) 0023-4001 ISSN (Online) 1738-0006 Korean J arasitol Vol. 50, No. 4: 317-325, December 2012 http://dx.doi.org/10.3347/kjp.2012.50.4.317 Foodborne rematode Metacercariae in Fish from Northern Vietnam and heir Adults Recovered from Experimental Hamsters Jong-Yil Chai 1, Nguyen Van De 2 and Woon-Mok Sohn 3, * 1 Department of arasitology and ropical Medicine, Seoul National University College of Medicine, and Institute of Endemic Diseases, Seoul National University Medical Research Center, Seoul 110-799, Korea; 2 Department of arasitology, Hanoi Medical University, Hanoi, Vietnam; 3 Department of arasitology and Institute of Health Sciences, Gyeongsang National University School of Medicine, Jinju 660-751, Korea Abstract: he prevalence of foodborne trematode (FB) metacercariae was investigated in fish from 2 localities of northern Vietnam in 2004-2005. Freshwater fish (9 species) were collected from local markets in Hanoi City (n= 76) and Nam Dinh rovince (n=79), and were examined for FB metacercariae using the artificial digestion technique. Adult flukes were obtained from hamsters experimentally infected with the metacercariae at day 8 post-infection. hree (Haplorchis pumilio, Centrocestus formosanus, and rocerovum varium) and 6 (Haplorchis taichui, H. pumilio, C. formosanus,. varium, Stellantchasmus falcatus, and Heterophyopsis continua) species of FB metacercariae were detected in the 2 regions, respectively. Overall, among the positive fish species, H. pumilio metacercariae were detected in 104 (80.0%) of 130 fish examined (metacercarial density per infected fish; 64.2). C. formosanus metacercariae were found in 37 (40.2%) of 92 fish (metacercarial density; 14.7).. varium metacercariae were detected in 19 (63.3%) of 30 fish (Anabas testudineus and Mugil cephalus) (metacercarial density; 247.7). S. falcatus metacercariae were found in all 10 M. cephalus examined (metacercarial density; 84.4). H. continua metacercariae (2 in number) were detected in 1 fish of Coilia lindmani. Morphologic characteristics of the FB metacercariae and their experimentally obtained adults were described. he results have demonstrated that various FB species are prevalent in northen parts of Vietnam. Key words: Haplorchis taichui, Haplorchis pumilio, Centrocestus formosanus, rocerovum varium, Stellantchasmus falcatus, Heterophyopsis continua, foodborne trematode, metacercaria, fish, Vietnam INRODUCION Received 15 July 2012, revised 12 August 2012, accepted 29 August 2012. * Corresponding author (wmsohn@gnu.ac.kr) 2012, Korean Society for arasitology and ropical Medicine his is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. Foodborne trematode (FB) infections are an important public health concern in various Asian countries, including Lao DR, Vietnam, Cambodia, hailand, the hilippines, aiwan, China, and the Republic of Korea. Human FB infections are caused by habitual consumption of raw fish containing infective larvae (metacercariae) [1-3]. Studies on FB metacercarial infections have revealed that some species of freshwater and brackish water fish play important roles as the source of human infections in endemic areas [4-7]. It is known that FB are not only pathogenic to the human host provoking remarkable morbidities but also harmful to fish hosts causing a serious economic loss in the aquaculture fish industry [1]. Surveys on metacercarial infection in second intermediate hosts, in combination with surveys on adult fluke infections in humans, are essential to understand the epidemiology and host-parasite relationships of FB infections in particular geographical areas. However, fecal examinations are not usually suitable to determine the exact infection status in humans, since the specific identification of eggs is very difficult in cases of mixed infections with small-sized trematode eggs, in particular, the liver fluke, heterophyids, gymnophallids, and lecithodendriid flukes [8-10]. herefore, investigation of metacercarial infections in second intermediate hosts available in the area around can provide valuble information on the trematode epidemiology. Hanoi City and Nam Dinh rovince are located in the northern part of Vietnam. hrough adult fluke recovery by Dung et al. [11], it has been known that many residents in Nam Dinh rovince are infected with FB, such as Clonorchis sinensis, heterophyids (Haplorchis pumilio, Haplorchis taichui, Haplorchis yok 317
318 Korean J arasitol Vol. 50, No. 4: 317-325, December 2012 ogawai, and Stellantchasmus falcatus), and Fasciolopsis buski [11]. After that survey, metacercatrial infections were investigated in fish intermediate hosts caught in several local areas in Vietnam [12-17]. In particular, han et al. [17] surveyed on FB metacercarial infection in freshwater fish from small-scale farms (family-based household fish farm) in Nam Dinh rovince. However, those studies did not provide quantitative details of FB metacercarial infections in fish hosts, mainly focusing on qualitative aspects of commercially important cultured fish species. herefore, in the present study, we intended to reveal the infection status (both qualitative and quantitative aspects) of FB metacercariae in wild fish caught from 2 localities of northern Vietnam (Hanoi and Nam Dinh rovince). In addition, we described the morphologic characteristics of FB metacercariae and their adults obtained from experimentally infected hamsters. MAERIALS AND MEHODS In Hanoi and Nam Dinh rovince (Fig. 1), 76 and 79 fish (9 N species each), respectively, were collected from local markets in November 2004 to April 2005. All collected fish were transferred with ice to the laboratory of Department of arasitology, Gyeongsang National University School of Medicine, Jinju, Korea. heir length and weight were measured and the species were identified with the aid of FishBase site in internet (able 1) [18]. Individual fish was ground with a mortar with pestle, or a grinder, and the ground fish meat was mixed with artificial gastric juice. he mixture was incubated at 36 C for 2 hr. he digested material was filtered through a 1 1 mm mesh, and washed with 0.85% saline untill the supernatant became clear. he sediment was carefully examined using a stereomicroscope. he metacercariae were classified by their general features and were identified at the species level based on detailed morphologies and demensions using a light microscope. Identified metacercariae were experimentally infected to hamsters to obtain adult worms. At day 8 after infection, the hamsters were killed by cervical dislocation, and their small intestines were isolated and longitudinally opened with a scissors in a beaker containing 0.85% saline. Adult flukes were recovered from the sediment of the intestinal contents which were diluted and washed with 0.85% saline. Recovered worms were fixed with 10% formalin under a cover glass pressure, stained with Semichon s acetocarmine, and observed using a light microscope equipped with a micrometer. Hanoi Nam Dinh rovince RESULS FB metacercariae in fish from Hanoi In fish from Hanoi, 3 species of metacercariae, H. pumilio, Centrocestus formosanus, and rocerovum varium were recovered. H. pumilio metacercariae were detected in 7 of 9 fish species examined. Among them, Ctenopharyngodon idella was most heavily infected (able 2). C. formosanus metacercariae were found in 4 fish species, and Anabas testudineus revealed a relatively high infection rate and high metacercarial density (able 3).. varium metacercariae were detected in only 1 fish species, A. testudineus. he infection rate of this fish with. varium metacercariae was 100%, and the average metacercarial density was 466 per fish (able 4). Fig. 1. Surveyed areas ( ) (Hanoi City and Nam Dinh rovince), located in northern Vietnam. FB metacercariae in fish from Nam Dinh rovince In fish from Nam Dinh rovince, 6 species of metacercariae, H. taichui, H. pumilio, C. formosanus,. varium, S. falcatus, and Heterophyopsis continua, were recovered. H. taichui metacercari-
Chai et al.: FB metacercariae in fish and their adults from Vietnam 319 able 1. Fish collected from local markets in Hanoi City and Nam Dinh rovince, Vietnam Species of fish No. of fish examined Length (cm) Weight (g) Range Average Range Average Fish from Hanoi a Cirrhinus molitorella 18 6.0-8.2 7.1 3.4-10.8 7.4 Hypophthalmichthys molitrix 10 6.0-6.8 6.4 2.4-3.9 3.3 Carassius auratus 10 5.0-7.0 6.1 3.3-10.6 6.5 Anabas testudineus 10 5.0-6.6 5.8 4.1-8.9 6.5 Albulichthys albuloides 10 5.2-7.5 6.1 2.6-7.9 4.6 arabramis pekinensis 7 7.0-8.6 7.6 7.0-14.2 9.0 untius semifasciolatus 5 4.0-4.7 4.3 1.5-3.4 2.2 Barbodes balleroides 3 4.5-5.2 4.7 2.1-3.6 2.7 Ctenopharyngodon idella 3 7.3-11.4 9.2 6.2-25.3 14.9 Fish from Nam Dinh b Carassius auratus 21 4.6-8.2 5.8 2.6-12.6 5.4 Ctenopharyngodon idella 11 5.7-10.0 7.8 4.0-16.2 9.1 Mugil cephalus 10 7.4-11.0 9.4 7.8-19.3 14.0 Anabas testudineus 10 5.5-8.3 6.8 5.6-16.7 10.4 untius semifasciolatus 10 4.5-5.8 5.0 2.7-5.5 3.7 Rasbora aurotaenia 5 8.9-9.8 9.5 6.0-11.3 9.2 Coilia lindmani 5 16.2-17.6 17.0 21.0-31.0 25.5 richogaster trichopterus 5 6.4-7.1 6.8 9.5-12.4 10.9 Acheilognathus barbatulus 2 4.8-5.9 5.4 3.4-4.8 4.1 a 76 fish and b 79 fish in total (9 species each) were examined. able 2. Haplorchis pumilio metacercarial infection in fish from local markets in Hanoi City and Nam Dinh rovince, Vietnam Species of fish No. of fish examined No. (%) of fish infected No. of metacercariae detected otal Range Average Fish from Hanoi Cirrhinus molitorella 18 18 (100) 431 11-50 23.9 Hypophthalmichthys molitrix 10 9 (90.0) 92 3-22 10.2 Carassius auratus 10 3 (30.0) 5 1-2 1.7 Albulichthys albuloides 10 5 (50.0) 10 1-6 2.0 arabramis pekinensis 7 6 (85.7) 46 2-14 7.7 Ctenopharyngodon idella 3 3 (100) 5,217 1,360-2,118 1,739 Barbodes balleroides 3 1 (33.3) 1-1.0 Subtotal 61 45 (73.8) 5,802 1-2,118 128.9 Fish from Nam Dinh Carassius auratus 21 20 (95.2) 188 1-26 9.4 Anabas testudineus 10 10 (100) 465 1-218 46.5 Ctenopharyngodon idella 11 9 (81.8) 191 1-43 21.2 untius semifasciolatus 10 8 (80.0) 27 1-8 3.4 Rasbora aurotaenia 5 4 (80.0) 14 2-8 3.5 Coilia lindmani 5 3 (60.0) 11 1-9 3.7 richogaster trichopterus 5 4 (80.0) 17 3-6 4.3 Acheilognathus barbatulus 2 1 (50.0) 35-35.0 Subtotal 69 59 (85.5) 878 1-218 14.9 otal 130 104 (80.0) 6,680 1-2,118 64.2 ae (6 in total number) were detected in 3 (60%) of 5 Rasbora aurotaenia fish. H. pumilio metacercariae were detected in 8 of 9 fish species examined. Among them, A. testudineus revealed relatively higher prevalence (able 2). C. formosanus metacer-
320 Korean J arasitol Vol. 50, No. 4: 317-325, December 2012 able 3. Centrocestus formosanus metacercarial infection in fish from local markets in Hanoi City and Nam Dinh rovince, Vietnam Species of fish No. of fish examined No. (%) of fish infected No. of metacercariae detected otal Range Average Fish from Hanoi Cirrhinus molitorella 18 9 (50.0) 36 1-9 4.0 Anabas testudineus 10 9 (90.0) 403 7-194 44.8 arabramis pekinensis 7 7 (100) 69 1-22 9.9 Barbodes balleroides 3 1 (33.3) 4-4.0 Subtotal 38 26 (68.4) 512 1-194 19.7 Fish from Nam Dinh Carassius auratus 21 1 (4.8) 1-1.0 Anabas testudineus 10 4 (40.0) 9 1-4 2.3 Ctenopharyngodon idella 11 4 (36.4) 20 1-10 5.0 untius semifasciolatus 10 1 (10.0) 1-1.0 Acheilognathus barbatulus 2 1 (50.0) 2-2.0 Subtotal 54 11 (20.4) 33 1-10 3.0 otal 92 37 (40.2) 545 1-194 14.7 able 4. rocerovum varium metacercarial infection in fish from local markets in Hanoi City and Nam Dinh rovince, Vietnam Species of fish No. of fish examined No. (%) of fish infected No. of metacercariae detected otal Range Average Fish from Hanoi Anabas testudineus 10 10 (100) 4,655 25-1,928 465.5 Fish from Nam Dinh Anabas testudineus 10 3 (30.0) 37 5-21 13.5 Mugil cephalus 10 6 (60.0) 14 1-5 2.3 Subtotal 20 9 (45.0) 51 1-21 5.7 otal 30 19 (63.3) 4,706 1-1,928 247.7 cariae were recovered in 5 fish species, and their infection rate and densities were relatively low (able 3).. varium metacercariae were detected in 2 fish species (climbing perch A. testudineus and striped mullet Mugil cephalus). S. falcatus metacercariae were detected in all 10 M. cephalus, and the average metacercarial density was 84.4. Only 2 H. continua metacercariae were recovered in a Lindman s grenadier anchovy, Coilia lindmani. Morphology of metacercariae (All measurement unit is µm) H. taichui metacercariae (n=5) were elliptical, 190-220 (av. 205) 160-190 (175) in size, had a baseball glove-shaped ventrogenital sac with 11-18 rodlets and an O-shaped excretory bladder occupying the large portion of the posterior body. hese metacercariae were identical with those from Laotian and Chinese fish [5,6]. H. pumilio metacercariae (n=20) were elliptical, 160-195 (179) 145-175 (159) in size, had 36-42 deer horn-like minute spines arranged in 1-2 rows around the ventrogenital complex, and an O-shaped excretory bladder occupying the large portion of the posterior body. hey were identical with those from Chinese fish [6]. C. formosanus metacercariae (n=20) were elliptical, 155-190 (169) 123-150 (138) in size, had 32 circumoral spines around the oral sucker arranged in 2 rows, and a X-shaped excretory bladder occupying the greater portion of the posterior body. hey were identical with those from Laotian fish [5].. varium metacercariae (n=20) were elliptical, 165-208 (187) 115-163 (147) in size, had yellowish-brown pigment granules scattering in the area near the intestinal bifurcation, a pair of eyespots lateral to the pharynx, a submedian ventral sucker, a thick-walled bulb-like expulser, and a D-shaped (half moonshaped) excretory bladder with grouped granules (Fig. 2A). S. falcatus metacercariae (n=10) were elliptical, 255-330 (297) 225-250 (232) in size, had yellowish-brown pigment granules scattered all over the body, a submedian ventral sucker, a thick-walled bulb-like expulser, and a V-shaped excretory bladder (Fig. 2B). H. continua metacercariae (n= 2) were nearly round with relatively thick cyst wall, 440-475 (458) 435-470 (453) in size, had brownish pigment scattered all over the body, a ventral sucker located median, a genital sucker located just behind the ventral sucker, and a Y-shaped excretory bladder (Fig. 2C). Morphology of adult flukes H. taichui (n=2; Fig. 3A): Body small, pear-shaped, 580-730
Chai et al.: FB metacercariae in fish and their adults from Vietnam 321 E EB EB A B C Fig. 2. FB metacercariae detected in fish hosts from Hanoi and Nam Dinh rovince, Vietnam. A). varium metacercaria; elliptical and 187 147 μm in average size, having yellowish-brown pigment granules, a pair of eyespots, a muscular oral sucker () and a submedian ventral sucker (), a thick-walled expulser, and a D-shaped excretory bladder (EB) with grouped granules. Scale bar = 50 μm. B) S. falcatus metacercaria; elliptical and 297 232 μm in average size, having yellowish-brown pigment granules, a muscular oral sucker (), and a submedian ventral sucker (), a thick-walled expulser (E), and a V-shaped excretory bladder (EB). Scale bar = 50 μm. C) H. continua metacercaria; round with relatively thick cyst wall and 458 453 μm in average size, having brownish pigments scattered all over the body, a muscular oral sucker () and a ventral sucker () located median, a genital sucker located just behind the ventral sucker, and a Y-shaped excretory bladder (EB). Scale bar = 100 μm. (av. 655) long and 300-310 (305) wide, with the greatest width at middle, the ovarian level. Oral sucker subterminal, 45 by 60 in size. harynx subglobular or elliptical, 35-38 (36) by 28-30 (29). Esophagus short, 75-95 (85) in length. Ventrogenital sac small with 11-18 rodlets, baseball glove-shaped, 88-100 (94) by 70-80 (75). Seminal vesicle saccular, bipartite, 113-125 (119) by 73-75 (74). Ovary spherical or subspherical, 85-88 (86) by 71-74 (73), dextral to midline. Seminal receptacle ellipsoidal, 80-88 (84) by 50-53 (51), lying at the right side of the ovary. estis single, globular or subglobular, 160-193 (176) by 120165 (143), lying in the posterior 1/4 of the body. Uterus with eggs occupying from the anterior 1/3 to the posterior end (most of the hind-body). Vitellaria follicular, distributing in post-ovarian fields. Eggs small, yellow, and 24-27 (26) by 12-14 (13). H. pumilio (n= 15, Fig. 3B): Body small, pear-shaped, 415550 (496) long and 195-245 (217) wide, with the greatest width at middle, the ovarian level. Oral sucker subterminal, 43-50 (46) by 50-58 (54). harynx subglobular or elliptical, 25-33 (29) by 20-30 (25). Esophagus short, 40-83 (57) in length. Ven trogenital sac small with 36-42 deer horn-like minute spines, 65-80 (74) by 50-75 (57). Seminal vesicle saccular, 28-100 (60) by 18-75 (45). Ovary spherical or subspherical, 55-70 (62) by 30-68 (52), slightly dextral to midline. Seminal receptacle elliptical, 38-85 (51) by 25-58 (38), lying at the right side of the ovary. estis single, globular or subglobular, 65-103 (82) by 75-105 (85), lying at the posterior 1/4 of the body. Uterus with eggs occupying from the anterior 1/3 to the posterior end (most of the hind-body). Vitellaria follicular, distributing in post-ovarian fields. Eggs small, yellow, and 28-31 (30) by 15-18 (16).. varium (n= 20, Fig. 3C): Body small, pear-shaped, 355-475 (434) long and 190-250 (223) wide, with the greatest width at posterior 1/3 of the body. Oral sucker subterminal, 35-43 (38) by 38-48 (42). harynx subglobular or elliptical, 23-30 (27) by 18-23 (20). Esophagus short, 33-75 (55) in length. Ventral sucker very small, 15-30 (25) by 18-33 (27), embedded in the ventrogenital sac. Expulsor long and thick-walled, 88-138 (115) by 25-35 (29). Ovary spherical or subspherical, 45-75 (59) by 38-70 (48), slightly dextral to midline. Seminal receptacle saccular, 45-83 (63) by 33-55 (44), lying at the right side of the testis. estis single, globular or subglobular, 105-150 (127) by 100-180 (127), situated at the middle of the hind-body. Uterus with eggs occupying from the anterior 1/3 to the posterior end (most of the hind-body). Vitellaria follicular, distributing from the posterior border of the ovary to the posterior extremity. Eggs small, yellow, and 25-28 (26) by 11-15 (13). S. falcatus (n= 15, Fig. 3D): Body small, 400-650 (481) long and 200-300 (239) wide. Oral sucker subterminal, 30-43 (38) by 45-55 (49). harynx subglobular, 23-30 (27) by 20-38 (24).
322 Korean J arasitol Vol. 50, No. 4: 317-325, December 2012 VGS VGS SR SV O SV SR E O O A B C E SV O O D E Fig. 3. Adult heterophyid flukes recovered from experimental hamsters at day 8 post-infection. All scale bars = 100 μm. A) H. taichui adult. Body small, 655 305 μm in average size, having a muscular oral sucker () and pharynx (), a ventrogenital sac (VGS) arm ed with 11-18 rodlets, a saccular seminal vesicle (SV) and seminal receptacle (SR), a spherical ovary (O), single globular testis (), and follicular vitellaria distributed in post-ovarian fields. B) H. pumilio adult. Body small, 496 217 μm in average size, having a muscular oral sucker () and pharynx (), a ventrogenital sac (VGS) equipped with 36-42 deer horn-like minute spines, a saccular seminal vesicle (SV) and seminal receptacle (SR), a spherical ovary (O), single globular testis (), and follicular vitellaria distributed in post-ovarian fields. C). varium adult. Body small, 434 223 μm in average size, having a muscular oral sucker () and a pharynx (), a small ventral sucker and a long and thick-walled expulsor (E), a spherical ovary (O), single globular testis (), and follicular vitellaria distributing in post-ovarian fields. D) S. falcatus adult. Body small, 481 239 μm in average size, having a muscular oral sucker () and phar ynx (), a small ventral sucker () and a long and thick-walled expulsor (E), a spherical ovary (O), 2 globular testes (), and follicular vitellaria distributed in post-ovarian fields. E) C. formosanus adult (left, unstained; right, acetocarmine-stained). Body very small, 367 207 μm in average size, having an oral suc ker () armed with 32 circumoral spines (arrow marks), a muscular pharynx (), a welldeveloped ventral sucker (), a spherical ovary (O), 2 globular testes (), and follicular vitellaria distributed along extracecal margins from the pharyngeal level to the posterior end.
Chai et al.: FB metacercariae in fish and their adults from Vietnam 323 Esophagus slender, 45-118 (72) in length. Ventral sucker small, 25-30 (27) by 28-30 (29). Expulsor long and thick-walled, 75-100 (87) by 30-38 (33). Seminal vesicle saccate, 63-70 (66) by 33-40 (37). Ovary spherical, 63-100 (79) by 43-75 (62). estes paired, ovoid or globular, slightly oblique and widely separated; right 83-180 (111) by 45-93 (62); left 83-188 (108) by 30-88 (55). Uterus with eggs occupying from the anterior 1/3 to the posterior end (most of the hind-body). Vitellaria follicular, distributing in the post-ovarian fields. Eggs small, yellow, and 20-23 (21) by 11-13 (12). C. formosanus (n=15, Fig. 3E): Body very small, 320-470 (367) long and 170-235 (207) wide. Oral sucker subterminal, 35-43 (40) by 38-50 (42), armed with about 32 circumoral spines. repharynx very short, 10-25 (17) long. harynx globular, 30-43 (35) by 20-28 (23). Esophagus very short. Ventral sucker round or elliptical, 33-43 (37) by 45-53 (51). Ovary elliptical, 50-75 (59) by 30-53 (39), dextral to midline. Seminal receptacle large and saccular, 50-75 (63) by 40-55 (49). wo testes ellipsoidal, side by side near the posterior end; right 60-88 (72) by 33-50 (41), left 55-88 (66) by 30-43 (36). Vitellaria follicular, distributing along the extracecal margins from the pharyngeal level to the posterior end. Eggs small, yellow, and 29-33 (30) by 15-18 (17). DISCUSSION he 6 species of FB metacercariae (H. taichui, H. pumilio, C. formosanus,. varium, S. falcatus, and H. continua) detected in this study were all minute intestinal flukes and members of the Heterophyidae. Among them, H. pumilio was the dominant species. It is of note that Dung et al. [11] reported a high prevalence (64.9%) of small trematode eggs and also high prevalence for soil-transmitted helminths among residents of 2 communes in Nam Dinh rovince, northern Vietnam, in April 2005. hey recovered adult flukes of 6 trematode species (Clonorchis sinensis, H. pumilio, H. taichui, H. yokogawai, S. falcatus, and Fasciolopsis buski) from 33 peoples who revealed over 1,000 eggs per gram of feces (EG) for small trematode eggs after praziquantel treatment and purgation. Among the 6 trematode species, H. pumilio was recovered in all 33 (100%) residents and the worm load averaged 416 per person [11]. Accordingly, it is presumed that more than 8 species of FB, including H. pumilio, may be distributed in the northern part of Vietnam, although we could not find C. sinensis and H. yokogawai metacercariae. Recently, studies on FB metacercarial infections in Vietnam have been performed popularly. hu et al. [12] surveyed on metacercarial infections in cultured catfish and snakeheads, and also in several wild fish species from An Giang rovince, a major fish production area in the Mekong Delta of Vietnam in 2005-2006. hien et al. [13] examined 13 major cultured fish species from ien Giang rovince and Can ho City, located in central areas of the Mekong Delta of Vietnam, in 2005-2006. Hop et al. [14] examined FB metacercarial infections in wastewater-fed aquaculture fish in northern Vietnam [14]. Chi et al. [15] performed a cross-sectional survey of FB metacercariae in farmed fish in Nghe An rovince, about 300 km south of Hanoi, in 2005 [15]. Vo et al. [16] surveyed 2 species of groupers (Epinephelus coioides and Epinepheles bleekeri), and mullet (M. cephalus) in a central coastal area of Vietnam (Khanh Hoa rovince) in 2008 [16]. han et al. [17] surveyed on the infection status of FB metacercariae in freshwater fish from smallscale farms (family-based household fish farm) in Nam Dinh rovince in 2010 [17]. hese studies focused mainly on metacercarial infections in commercially important fish species and in qualitative aspects. Studies on quantitative aspects of FB metacercarial infections in each species of fish and thereby the suitability and susceptability of fish hosts for each FB species have not been available. In the present study, H. pumilio metacercariae were detected in 104 (80.0%) of 130 fish examined, and the metacercarial density was 64.2 per fish infected. he metacercarial density was higher in fish from Hanoi (av. 129 metacercariae) than those from Nam Dinh rovince (av. 15 metacercariae). he most heavily infected fish with H. pumilio metacercariae was the grass carp (Ctenopharyngodon idella) caught from Hanoi. he high susceptibility of this fish species was also shown in the study of han et al. [17] which was performed in Nam Dinh rovince [17]. A similar finding was shown in Guangxi Zhuang Autonomous Region located in the southern part of China, slightly north of our surveyed area [6]. In this area of China, 18 fish species (Carassius auratus, Acheilognathus tonkinensis, Hemibarbus maculatus, Hypophthalmichthys molitrix, Hemiculter leucisculus, C. idella, oxabramis houdemeri, Microphysogobio fukiensis, seudohemiculter dispar, Opsariichthys bidens, Squalidus argentatus, Metzia lineata, Cyprinus carpio, untius semifasciolatus, Saurogobio dabryi, Culter recurviceps, Chanodichthys dabryi, and seudorasbora parva) were listed as hosts for H. pumilio [6]. On the other hand, in Nam Dinh rovince of Vietnam, han et al. [17] listed 17 fish species (Labeo rohita, H. molitrix, Cirrhinus
324 Korean J arasitol Vol. 50, No. 4: 317-325, December 2012 mrigala, C. idella, C. auratus, Squaliobarbus curriculus, iaractus brachypomum, C. carpio, Cirrhinus molitorella, Hypophthalmichthys nobilis, Barbonymus gonionotus, H. leucisculus, Anabas testudineus, Oreochromis niloticus, Clarias batrachus, Channa orientalis, and Notopterus notopterus) for the host for H. pumilio. In the present study, 7 more fish species (Acheilognathus barbatulus, Albulichthys albuloides, Barbodes balleroides, Coilia lindmani, arabramis pekinensis, Rasbora aurotaenia, and richogaster trichopterus) have been newly added as the host for H. pumilio. C. formosanus metacercariae were detected in 8 fish species (A. barbatulus, A. testudineus, B. balleroides, C. auratus, C. molitorella, C. idella,. pekinensis, and. semifasciolatus) in the present study. han et al. [17] detected the same species of metacercariae in 10 fish species (L. rohita, H. molitrix, C. mrigala, C. idella, S. curriculus,. brachypomum, C. carpio, C. batrachus, C. orientalis, and A. testudineus) in Nam Dinh rovince [17]. In China, Sohn et al. [6] reported 10 fish species (M. fukiensis, A. tonkinensis, S. argentatus, C. carpio, H. molitrix, A. rivularis, H. leucisculus, M. lineata, S. dabryi and. parva) as the fish hosts for C. formosanus [6]. herefore, it appears that more than 24 fish species play the role of second intermediate hosts for C. formosanus in Vietnam and China. It has been known that 2 species of liver flukes (Clonorchis sinensis and Opisthorchis viverrini) distribute in Vietnam. C. sinensis is distributed in the northern part, whereas O. viverrini is found in the southern part. However, the metacercariae of these liver flukes were only rarely detected in fish from Vietnam. In An Giang rovince, a southern part, only 19 metacercariae of O. viverrini were detected in 1.9% of 108 fish examined [12]. hien et al. [13] could not find any liver fluke metacercariae in all 13 important cultured fish species from ien Giang rovince and Can ho City, in the Mekong Delta of Vietnam [13]. No liver fluke metacercariae were detected in a cross-sectional survey of FB metacercariae in farmed fish from Nghe An rovince [15]. Meanwhile, only a small number of C. sinensis metacercariae were found in 1 of 1,185 silver carps (H. molitrix) examined in small-scale farms in Nam Dinh rovince. In the present study, we could not find any C. sinensis metacercariae in fish from the same study area. By contrast, Dung et al. [11] reported a high positive rate (51.5%) of C. sinensis adult worms (their worm load was not so high; 1-18 per individual) from 33 residents with over 1,000 EG for small trematode eggs in Nam Dinh rovince [11]. his discrepancy between the metacercarial infection in fish hosts and the adult fluke infections in humans should be clarified in the near future. In the present study,. varium metacercariae were detected in 2 fish species (A. testudineus and M. cephalus), although these metacercariae were previously recorded in 9 fish species (L. rohita, H. molitrix, C. mrigala, C. idella, S. curriculus,. brachypomum, C. batrachus, B. gonionotus, and A. testudineus) in Nam Dinh rovince [17]. Vo et al. [16] also detected. varium metacercariae together with 2 other heterophyid metacercariae (H. continua and. summa) in 2 species of groupers and mullet from Khanh Hoa rovince in Vietnam. On the other hand, S. falcatus and H. continua metacercariae were found in M. cephalus and C. lindmani, respectively, in the present study. Although morphologic characteristics of FB metacercariae had previously been reported, we redescribed some of their characteristic features to provide a useful aid for epidemiologic studies in Southeast Asian countries. Among the FB metacercariae detected in our study, 3 species (H. taichui, H. pumilio, and C. formosanus) were morphologically identical with those from China and Lao DR [5,6]. On the other hand, the metacercariae of. varium (187 147 µm) were smaller than those (210 180) reported in a freshwater fish (Oryzias melastigma) from Visakhapatnam, India [19]. However, our. varium metacercariae were almost identical with those found in groupers from Nha rang district in Khanh Hoa rovince, Vietnam [16]. With regard to S. falcatus metacercariae, they were previously detected in cultured giant gouramy (Osphronemus gourami) from ien Giang rovince and Can ho City [13], and also in the common carp (C. carpio) and grass carp (C. idella) from Nghe An rovince, Vietnam [15]. However, no morphologic descriptions were available on S. falcatus metacercariae in Vietnam. H. continua metacercariae (458 453 µm) detected in a C. lindmani fish in the present study were similar in shape with but larger than those (380 in diameter) recovered in mullets from Khanh Hoa rovince [16]. In the Republic of Korea, it is agreed that H. continua metacercariae detected in various fish species (Laterolabrax japonicus, Acanthogobius flavimanus, Clupanodon punctatus, lecoglossus altivelis, Conger myriaster, Boleophthalmus pectinirostris, and Scartelaos sp.) show wide size ranges by fish hosts examined [20]. Among the FB species distributed in Vietnam, 2 species of liver flukes (C. sinensis and O. viverrini) are highly important in clinical and pathological aspects, although their metacercariae were not detected in this study. Being similar in general shape, the liver fluke metacercariae can be distinguished from those of heterophyid flukes by the presence of a large well-developed ventral sucker which is nearly equal in size with the oral suck-
Chai et al.: FB metacercariae in fish and their adults from Vietnam 325 er. he ventral suckers of H. taichui, H. pumilio, C. formosanus,. varium, and S. falcatus are characteristically smaller than their oral suckers. Nevertheless, when these metacercariae were mixed together, it is not easy to distinguish them unless each metacercaria is subjected to a detailed observation under a light microscope. he experimentally obtained adults of the 5 species of FB were morphologically compatible with those previously reported. Dung et al. [11] only briefly mentioned on the adult worm morphologies of H. pumilio, H. taichui, H. yokogawai, and S. falcatus after recovery in residents of Nam Dinh rovince, Viet nam. he size of H. taichui (756 421) and H. pumilio (632 291) adults obtained from residents were slightly larger than those from our study, whereas the adult of S. falcatus (468 298) from residents was smaller in size than our specimens [11]. 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