image/svg+xmlISSN Versión impresa 2218-6425ISSN Versión Electrónica 1995-1043Neotropical Helminthology, 2022, 16(1), ene-jun:-56.49ORIGINAL ARTICLE / ARTÍCULO ORIGINAL1Instituto de Investigaciones de la Amazonía Peruana (IIAP). Laboratorio de Parasitologia y Sanidad Acuícola. Carretera Iquitos-Nauta Km 4.5, Iquitos, Loreto-Peru.2Amazon Research Center for Ornamental Fishes - ARCOF, Asentamiento Humano 31 de mayo, Mz. 22 Lt. 2, Iquitos, Loreto, Perú.*Corresponding author: germantiss1106@gmail.comGermán Augusto Murrieta-Morey: https://orcid.org/0000-0001-6244-2654Carlos Alfredo Tuesta-Rojas: https://orcid.org/0000-0002-1961-3574Angela María Echevarria-Matos: https://orcid.org/0000-0003-0296-0921Carlos Tobías Chuquipiondo-Guardia: https://orcid.org/0000-0002-7979-20251*1Germán Augusto Murrieta-Morey; Carlos Alfredo Tuesta-Rojas; 22Angela María Echevarria-Matos & Carlos Tobías Chuquipiondo-GuardiaABSTRACTKey words: Aquaculture – cichlids – Clinostomum – ornamental fish – piscivorous birdThe ornamental fish trade is one of the most important economic activities for thousands of people in the Amazon region. Among fish that are exported from Peru, Apistogramma sp., Cichlasoma amazonarum (Kullander, 1983) and Pterophyllum scalare (Schultze, 1823) are highly demanded in the ornamental fish trade activities. To date, in Peru, studies concerning parasites that affect aquaculture activities are scarce. The present study is first to report the occurrence of Clinostomum sp. parasitizing three species of cichlids in the Peruvian Amazonia. Samples of Apistogramma sp., C. amazonarum and P. scalare were collected in a fish farming pond of the “Amazon Research Center for Ornamental Fishes (ARCOF) in Iquitos, Peru. Analyzed samples revealed the presence of Clinostomum sp. parasitizing the fins of the fish. As some species of cichlids are cultivated in the Peruvian Amazon for exportation as ornamental fish, the presence of this trematodecould compromise the health and appearance of the fish and may cause mortalities, either, due to susceptibility to predation by piscivorous birds or due to injuries caused by the infection of parasites. Neotropical Helminthology49doi:10.24039/rnh20221611381METACERCARIAE OF CLINOSTOMUM SP.(TREMATODA: CLINOSTOMIDAE) INFESTING ORNAMENTAL CICHLIDS CULTIVATED IN THE PERUVIAN AMAZONIAMETACERCARIA DE CLINOSTOMUMSP. (TREMATODA: CLINOSTOMIDAE) INFESTANDO A CÍCLIDOS CULTIVADOS ORNAMENTALES EN LA AMAZONÍA PERUANAEste artículo es publicado por la revista Neotropical Helminthology de la Facultad de Ciencias Naturales y Matemática, Universidad Nacional Federico Villarreal, Lima, Perú auspiciado por la Asociación Peruana de Helmintología e Invertebrados Afines (APHIA). Este es un artículo de acceso abierto, distribuido bajo los términos de la licencia Creative Commons Atribución 4.0 Internacional (CC BY 4.0) [https:// creativecommons.org/licenses/by/4.0/deed.es] que permite el uso, distribución y reproducción en cualquier medio, siempre que la obra original sea debidamente citada de su fuente original.DDDD
image/svg+xmlThe ornamental fish trade is one of the most important economic activities for thousands of people in the Amazon region (Moreau & Coomes, 2007). Nearly all Amazon fish exported from the natural environment are obtained by local fishermen, who transfer the fish to agents who, later on, transfer them to exporters so as to reach fish stores worldwide. It has been estimated that in this transport chain, 73% of the fish die, due to several causes (Olivier, 2001).Cichlids are one of the major ornamental fish families, due to the large number of species and the demand for these fish for ornamental purposes. This demand is explained by the variety of shapes, extravagant colors and special behavior of the different species that are included in this family (Kullander & Silfvergrip, 1991; Garcia-Dávila et al., 2021). According to Garcia-Dávila et al.(2021), in the Peruvian Amazonia, 37 species of cichlids are commercialized as ornamental fish. Between these species, Apistogramma sp., Cichlasoma amazonarum (Kullander, 1983) and Pterophyllum scalare (Schultze, 1823)are highly demanded for ornamental fish importers. In the region of Loreto, Peru, most of the cichlids used in the ornamental trade are captured from natural environments while others are breed under captivity in artificial ponds (Garcia et al.,2021). El comercio de peces ornamentales es una de las actividades económicas más importantes para miles de personas en la región amazónica. Entre los peces que se exportan desde Perú, Apistogrammasp., Cichlasomaamazonarum(Kullander, 1983) y Pterophyllumscalare(Schultze, 1823) son muy demandados en las actividades comerciales de peces ornamentales. A la fecha, en el Perú, los estudios referentes a los parásitos que afectan las actividades acuícolas son escasos, es así que el presente estudio registra por primera vez en la Amazonía peruana la ocurrencia de Clinostomumsp. parasitando tres especies de cíclidos. Se recolectaron muestras de Apistogrammasp., C. amazonarumy P. scalareen un estanque de piscicultura del “Centro de Investigación de Peces Ornamentales de la Amazonía (ARCOF) en Iquitos, Perú. Las muestras analizadas revelaron la presencia de Clinostomumsp. parasitando las aletas de los peces. Como algunas especies de cíclidos se cultivan en la Amazonía peruana para su exportación como peces ornamentales, la presencia de este trematodo podría comprometer la salud de los peces y causar mortalidades, ya sea por susceptibilidad a la depredación por aves piscívoras o por lesiones causadas por la infección de parásitos.50Neotropical Helminthology, 2022, 16(1), ene-junINTRODUCTIONParasites are abundant in tropical climates, and are one of the major causes of death and disposal of ornamental fish, thus representing large economic losses. Because of the small size of these fish, they are easily affected by small numbers of parasites (Takemoto et al., 2004).Metacercariae of Clinostomum spp.have the ability to infect muscles, fins and internal organs of fish. This parasite is usually found on the host body wall below the integumentary tissue, causing characteristic nodular swelling that can be easily recognize during physical examination (Simsek et al., 2018). In the life cycle of these parasites, the first intermediate hosts are species of freshwater snails which have been infected from eggs deposited in the water by definitive host fish-eating birds (Liao, 1993). Fish species act as second intermediate hosts, manifesting the pathology known as “yellow-grub disease” (Ramos-Alves et al., 2001).Thus, the present study registers for the first time in the Peruvian Amazonia the occurrence of Clinostomum sp. parasitizing ornamental cichlids collected from an artificial fish pond. In a fish farming pond of the “Amazon Research Center for Ornamental Fishes (ARCOF) in Iquitos, RESUMENPalabras claves: Acuicultura – Ave piscívora – cíclidos – ClinostomumPez ornamentalMurrieta-Morey et al.MATERIAL AND METHODS
image/svg+xmlOOPeru (03 46.433' S, 073 15.688'W), specimens of “Apistograma tapiche” dwarf cichlid Apistogramma sp. with 4.30 ± 0.88 cm standard length and 3.50 ± 0.68 g of total weight; “bujurqui” Amazon cichlid C. amazonarumwith 4.28 ± 0.84 cm standard length and 3.60 ± 1.03 g of total weight and “pez ángel” wild Angelfish P. scalare with 7.20 ± 0.57 cm standard length and 5.80 ± 0.83 g of total weight (were observed with signs of erratic swimming: fish swam very close to the surface, showing difficulty in their movement). Specimens were collected and placed in plastic bags with water and oxygen for transportation to the “Laboratorio de Parasitología y Sanidad Acuícola” of the “Instituto de Investigaciones de la Amazonía Peruana” (IIAP), Loreto, Peru. The surface of the body and fins were analyzed by using a stereomicroscope. Parasites observed in the fins were collected with dissecting needles and placed in Petri dishes for counting the number of individuals collected per infested fish. For fixation of the parasites, alcohol-formalin-acetic acid (AFA) was used for 24 h and then conserved in 70% ethanol. For taxonomic identification, it was necessary to stain the parasites with Langeron's alcoholic Carmin. Finally, stained parasites were observed under stereoscope and microscope. Parasitological indices (Prevalence, mean intensity and mean abundance) followed Bush et al.(1997). Parasites were deposited in the parasitological collection of the “Laboratorio de Parasitología y Sanidad Acuícola” from IIAP.Ethical disclosures This study was developed in accordance with the principles adopted by IIAP, and authorization from Ethics Committee of the IIAP was carried out.51Neotropical Helminthology, 2022, 16(1), ene-junAnalyzed fish revealed the presence of metacercariae of Clinostomum sp. (Figs. 1 and 2) on the fins of infested fish species. The main characteristics observed in the parasites were: Body linguiform with slight strangulation at the level of ventral sucker. Oral sucker located in the center of a circular dilation of the body. Pharynx present with bifurcation of the intestinal cecum extending to the posterior extremity of the body. Large intestinal cecum with irregular contour; pre-equatorial acetabulum; testes located between the end of the second third and the upper region of the last third of the body, anterior and posterior testis with lobular triangular shape. Intratesticular and post-equatorial ovary. Intercaecal, pre-testicular and post-acetabular uterus. Vitelline consisting of small, post-acetabular, intra and extracaecal follicles.According to site of infestation, for Apistogramma sp., 47% of the metacercariae were found on the dorsal fin; 39% in the anal fin; 10% on the caudal fin; 02% on the ventral fin and 02% on the pectoral fin. For C. amazonarum, 82% of the parasites were found on the dorsal fin; 11% on the pectoral fin; 05% on the caudal fin and 02% on the ventral fin. For P. scalare, 55% of the parasites were found on the dorsal fin; 20% on the caudal fin; 18% on the ventral fin and 07% on the pectoral fin (Fig. 3). Parasitological indices of Clinostomum sp.reported in the three-host species are showed in Table 1.RESULTSTable 1. Parasitological indices of Clinostomum sp.reported in three species of cichlids. AF = analyzed fish, PF = parasitized fish, P% = prevalence, N = number of C. marginatum recorded in each host species, mI = mean intensity of infestation, mA = mean abundance of infestation.Species of fichlidsAFPFP%NmImAApistogramma sp.20168022213.87± 2.411.1± 1.8Cichlasoma amazonarum331957.57552.89± 0.621.66± 0.44Pterophyllum scalare 5480448.8± 1.911± 2.5Infestation of Clinostomum sp.
image/svg+xmlInfections of Clinostomum spp.have been reported in different fish species. In Brazil: Semaprochilodus insignis (Jardine, 1841); Cichla ocellaris Bloch & Schneider, 1801; Crenicichla sp. (Castelo, 1984; Thatcher, 1991); Hoplosternum littorale (Hancock, 1828)(Sao Clemente et al., 1998); P. scalare (Ramos-Alves et al., 2001); Synbranchus marmoratus Bloch, 1795(Isaac et al., 2004); Geophagus brasiliensis (Quoy & Gaimard, 1824) (Paraguassú et al.,2005); Pygocentrus nattereri Kner, 1858 (Morais et al., 2011); Colossoma macropomum (Cuvier, 1816)(Murrieta-Morey & Malta, 2016);52DISCUSSIONNeotropical Helminthology, 2022, 16(1), ene-junPterygoplichthys pardalis (Castelnau, 1855) (De Souza et al., 2020a); Gymnotus coropinae Hoedeman, 1962, Gymnorhamphycthys petiti Géry & Vu, 1964, Brachyhypopomus brevirostris(Steindachner 1868) and Microsternarchus bilineatusFernández-Yépez, 1968 (De Souza et al.,2020b). In the present study, Apistogramma sp., C. amazonarum and P. scalare are cited for the first time as hosts of Clinostomum sp. metacercariae and therefore, we constitute the first study in the Peruvian Amazon related to infestations of trematodes in fish species cultivated.The fact that the dorsal and anal fins were the most common locations of the parasites is probably due to low movements of those fins, therefore Figure 1.Metacercariae of Clinostomum sp. A. Ventral view showing the complete body. B. Anterior part of the body. C. Posterior part of the body. os = oral sucker, f = pharynx, ic = intestinal caecum, a = acetabulum, u = uterus, at = anterior testes, o = ovary, pt = posterior testes. Scale bar: A = 2 mm, B, C = 300 µm.Murrieta-Morey et al.
image/svg+xml53Neotropical Helminthology, 2022, 16(1), ene-junFigure 2. Esquematic design of metacercariae of Clinostomum sp. A. Ventral view showing the complete body. B. Anterior part of the body. C. Posterior part of the body. os = oral sucker, ph = pharynx, ic = intestinal caecum, a = acetabulum, u = uterus, at = anterior testes, o = ovary, pt = posterior testes. Scale bar: A = 2 mm, B, C = 300 µm.facilitating the contact with the invasive cercariae. On the other hand, the low value found for the infection in the caudal fin, as compared with the other fins, is likely to be due to the more intense movements of the caudal fin; therefore, making difficult the penetration by the cercariae into the body of Apistogramma sp. Similar report are present by Eiras et al.(1999) where infected loricarids present more metacercariae of C. marginatum attached into dorsal and anal fins compared with the caudal fin that presented low levels of infection. Metacercariae of trematodes may change the behavior of their second intermediate fish host, enhancing the chance to be eaten by the final host (Szidat, 1969; Simsaket al., 2018). As an adaptive strategy, Clinostomum spp. are able to infect the Infestation of Clinostomum sp.
image/svg+xml54Neotropical Helminthology, 2022, 16(1), ene-junFigure 3.Metacercariae of Clinostomum sp.parasitizing the fins of ornamental cichlids. A. Lateral view of Pterophyllum scalare showing metacercariae in dorsal and pectoral fins. B. Pectoral fin of P. scalare with metacercariae. C. Lateral view of complete body of P. scalarewith metacercariae on dorsal and pectoral fins. D. Dorsal fin of Cichlassoma amazonarum with trematodes. E. Anal fin of Apistogramma sp. with encysted metacercariae. F. Dorsal fin of Apistogramma sp. with encysted metacercariae Scale bar A, C, D = 5 mm; B, E, F = 3 mm.Murrieta-Morey et al.
image/svg+xmlBush, A O, Lafferty, K D, Lotz, J M. & Shostak W A. 1997. Parasitology meets ecology on its own terms: Margolis et al.revisited. The Journal of Parasitology, vol. 83, pp. 575–583.Castelo, F P. 1984. Ocorrência de Clinostomum marginatumRudolphi, 1819 “Yellow-Spot Disease” em filé de jaraqui (Semaprochilodus insignisShomburgk, 1814).Acta Amazonica, vol. 13, pp. 325-326.De Souza, D C, De Sousa, L F, Coelho, T A. & Corrêa L L. 2020a. Host–parasite interaction between trematode, Clinostomum marginatum(Clinostomidae) and armoured catfish, Pterygoplichthys pardalis(Loricariidae) from Brazilian Amazon. Annals of Parasitology, vol. 66, pp. 243-249.De Souza, D C, Nogueira, A. & Corrêa L L. 2020b. Parasitism by Clinostomum marginatum(Clinostomidae) in neotropical electric fish (Gymnotiformes) in the Brazilian Amazon.Annals of parasitology, vol. 66, pp. 101-106.Eiras, J, Dias, M L, Pavanelli, G C. & Machado M H. 1999. Histological studies on the effects of Clinostomum marginatum(Digenea, Clinostomidae) in its second intermediate host Loricariichthys platymetopon(Osteichthyes, Loricariidae) of the upper Paraná River, Brazil. Acta Scientiarum. Biological Sciences, vol. 21, pp. 237-241.García-Dávila, C, Estivals, G, Mejia, J, Flores, M, Angulo, C, Sánchez, H, Nolorbe, C, Chuquipiondo, C, Castro-Ruiz, D, García, A, Ortega, H, Pinedo, L, Oliveira, C, Römer, U, Mariac, C, Duponchelle, F, Renno, J F. 2021. Peces Ornamentales de la Amazonía Peruana. Instituto de Investigaciones de la Amazonia Peruana (IIAP). 503 p.Isaac, A, Guidelli, G M. & De França J G. 2004. Composição e estrutura das infracomunidades endoparasitárias de Gymnotusspp. (Pisces: Gymnotidae) do rio Baía, Mato Grosso do Sul, Brasil. Acta Scientiarum, Biological Sciences, vol. 26, pp. 453-462. Kullander, S. & Silfvergrip A. 1991. Review of thefins of the fish, causing difficulty for swimming and therefore be more susceptible to predation by piscivorous birds (Eiras et al., 1999; Simsak et al., 2018). Additionally, in infected fish, metacecariae induce to present slight degeneration of the muscle fibers and the destruction of the fat cells of the fins, making the movements and the ability to response to stimulus slower than in non-infected specimens (Eiras et al., 1999). In the present study, the location of the metacercaria on the fins of cichlids may be a strategy to induce to erratic swimming, susceptibility to swimming closer to the surface of the water and to cause other disturbs for running away from their predators.According to Lo et al. (1982), when a large amount of cercariae of Clinostomum spp. penetrate the fish body at the same time they may cause irritation and lesions to fish tissue, which may induce mortality of small fish. Furthermore, specimens may become weak and grew more slowly than healthy fish, dying during transportation (Lo et al., 1982). As “Apistograma tapiche” Apistogramma sp., C. amazonarum and P. scalareare cultivated in the Peruvian Amazon for exportation as ornamental fish, the presence of C. marginatum could compromise the health of the fish and to cause mortalities, either due to susceptibility to predation by piscivorous birds or due to injuries caused by the infection of parasites. In that way, to prevent also economic losses, prophylactic measures should be taken into consideration: avoid the presence of aquatic snails and piscivorous birds near the breeding ponds.Molecular studies will be necessary to be able to identify at the species level the metacercariae of Clinostomumcollected from the analyzed cichlids. This, in order to guarantee the correct taxonomic identification based not only on morphological characters, but also based on the genetic sequence of the parasites.55Neotropical Helminthology, 2022, 16(1), ene-junThe authors thank the Amazon Research Center for Ornamental Fishes (ARCOF), Perú, for providing the samples used in the present study.ACKNOWLEDGEMENTSBIBLIOGRAPHIC REFERENCESInfestation of Clinostomum sp.
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