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Saprolegnia Parasitica in Pseudochondrostoma Duriense (Coelho, 1985) (Cyprinidae)

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Saprolegnia Parasitica in Pseudochondrostoma Duriense (Coelho, 1985) (Cyprinidae) Bull. Eur. Ass. Fish Pathol., 36(2) 2016, 91 NOTE ȱȱȱȱȱȱȱ by Saprolegnia parasitica in Pseudochondrostoma duriense (Coelho, 1985) (Cyprinidae) ǯȱǯȱȬ Șǰȱ ǯȱǯȱȬ ǰȱǯȱ £¤£Ȭ Palacios, M. J. García-Iglesias and L. I. Pérez-Ordoyo Animal Health Department. Veterinary Faculty. University of àǯȱȱȱ£ǰȱȦǯȱŘŚŖŝŗȱàǯȱ Abstract ȱȱȱȱ¢ȱSaprolegnia parasiticaȱȱȱ¢ȱęȱPseudochondrostoma duriense ȱȱȱȱȱęȱǯȱP. durienseȱȱȱȱȱ ȱǯȱ ȱȱȱ ȱȱęȱȱȱȱȱȱǻǼǯȱȱȱȱȱȱ ȱǯȱȱ ȱȬȱȱ ȱȱȱȱǰȱȱȱ ȱȱȱȱȱęǰȱȱȱȱȱǯ Saprolegnia parasiticaȱȱȱȱȱSap- spread over their head, trunk, caudal peduncle rolegnia (Kingdom Chromista, Class Oomy- ȱęȱȱ ȱȱȱ¢ȱ¢ȱ ǼȱȱȱȬ ȱęǰȱ ȱ ǰȱ ȱȱę¢ȱ¢ȱSaprolegnia spp. Some ȱȱȱǯȱȱ ȱȱ ęȱȱȱȱȱȱȱȱȱȱ¢ȱ by Neish and Hughes (1980), Bruno et al. (2011) £ǰȱ ȱȱȱȱ¢ȱěǰȱȱ ȱ £ȱȱǯȱǻŘŖŗŚǼȱȱȱȱȱ ȱȱȱȱȱęȱǻȱ ęȱȱ ȱȱȱȱȱǯȱ ŗǰȱǼǯȱȱ ȱěȱęȱȱȱȱ- ȱȱ¢ȱȱȱęȱȱ ȱĚȱĚ¡ȱȱ¢ȱȱ£ȱ ȱ ȱȱȱȱȱPseudochon- Ĵȱǯ drostoma duriense (Coelho, 1985), the northern straight-mouth nase whose Spanish name is ȱěȱęȱǻ¡ȱǰȱȱǼȱ “boga del Duero”. were caught. Their average length was 17.9 ± 3.4 ȱȱ ȱşŗȱƹȱŚśǯŘȱǯȱȱȱȱȱ ȱȱȱ¢ȱȱŘŖŗŗȱȱȱȱ lesions were observed under the microscope ȱȱęȱȱȱȱȱǰȱ ȱȱ ȱȱĴȱǯȱ ȱ¢ȱȱȱȱǰȱ ȱȱĚ ȱ Others were cultured on glucose peptone agar ȱȱ¢ȱȱàȱȱȬȱǯȱ (GPA) with chloramphenicol (0.05 mg mL-1) at ȱęȱȱȱȱȱĴ¢ȱȱ ŘŖȱķȱȱȱȬȱǯȱSaproleg- * Corresponding author’s email: [email protected] 92, Bull. Eur. Ass. Fish Pathol., 36(2) 2016 A B Figure 1. ȱȱȱĴ¢ȱȱȱ¢ȱSaprolegnia parasitica on skin (head, trunk, caudal Ǽǰȱ¢ȱȱęȱȱPseudochondrostoma duriense (A, B). nia isolates were subcultured onto GPA with ęǯȱȱȱ ȱę¡ȱȱŗŖƖȱěȱ hemp seeds (Cannabis sativaǼȱ ȱ¢ȱ- ȱȱȱȱĜǰȱȱȱ ȱȱ£ȱȱȱȱȱȱ tissue sections were stained with haematoxy- ȱęȱȱȱȱ ǰȱȱŝȱ ȱȱǰȱȱȬěȱǻǼȱȱ ķȱȱȱŘŖȱķǰȱȱȱȱȱȱ Gomori methenamine silver (GMS). zoospores and sexual reproduction (oogonia) (Willoughby, 1978); and to observe the long ȱȱȱ cutaneous lesions hairs and indirect germination on the second- and signs described above, necropsy showed ¢ȱ¢ǰȱ¢ȱȱǰȱȱȱĴȱȱ no lesions in internal organs. Ovaries and ȱȱǻ¢ǰȱŗşŞśǼǯȱ ęȱȱ ȱ ȱ¢ȱȱȱȱęȱ ȱ Saprolegnia parasiticaȱ ȱȱȱȱȱ- ȱȱ ȱǯȱȱ¢ȱȱ phological characteristics: long hairs on the ȱȱȱȱȱȱȱ- secondary cysts, indirect (retracted) germination ȱĴȱȱȱ¢ȱȱ£- ȱȱȱŝȱķȱǻ ȱǼȱǻ- ȱȱSaprolegnia spp., and Saprolegnia loughby, 1985; Diéguez-Uribeondo et al., 2007). parasiticaȱǻ¡ȱȱŝȱķȱȱŘŖȱķǰȱ ȱȱ ȱȱȱ¢ǰȱȱȱ hairs and indirect germination on its second- spleen were grown on tryptone soy agar (TSA), ¢ȱ£ȱ¢Ǽȱ ȱȱȱȱ ȱȱȱ¢ȱȱȱȱ ȱȱȱȱȱȱ ǯȱȱ ȱȱȱŗŚȱęǯȱȱȱȱ ȱ ȱ¢ǰȱȱȱȱȱȱ ȱ ęȱ¢ȱ ȱȱ·¡ȱțȱ ȱ ȱȱǰȱ¢ȱ ȱ 2 System using the GN reagent card and then ȱȱȱȱęȱǻȱȱ ȱęȱ ȱȱȱ¡ȱ colony morphology). The seven selected iso- ȱȱȱȱ¢ȱ¢ȱȱŗŜȱȱ ǰȱȱȱȱȱęǰȱ ȱęȱȱ GyrB genes and using BLAST on GenBank. Shewanella putrefaciensȱǻŗȱęǼǰȱAeromonas media ǻŗȱęǼǰȱAeromonas sobriaȱǻŘȱęǼȱȱAeromonas A histopathological study was also carried out bestiariumȱǻŘȱęǼȱȱȱǰȱȱ on the skin and eyes with lesions, together with Aeromonas sobriaȱȱȱȱ¢ȱǻŗȱęǼǯȱ ȱ¢ǰȱǰȱȱȱȱȱȱ ȱ ȱęȱS. parasiticaȱ ȱȱȱȱ Bull. Eur. Ass. Fish Pathol., 36(2) 2016, 93 internal organs (kidney, spleen or liver) in bac- ȱȱȱȱȱȱ ȱǯȱ teriological cultures on TSA. P. durienseȱȱȱęȱȱȱȱȱ ęǰȱȱȱȱȬȱȱȱ ȱ Histopathological examination showed non- ȱǻȱȱǼȱȱȱȱȱ ȱȬȱȱ ȱȱȱȱ the rivers Duero and Miño (Douro and Minho) epidermis (Figure 2A). There was also marked ȱȱȱȱȱȱȱȱǰȱ ęȱ£ȱǰȱ¢ȱ£ǰȱ while the species P. polylepis and P. willkommii ¢ȱȱȱȱȱȱȱ ȱ¢ȱȱȱȱȱȱȱȱ ȱęȱȱȱȱȱǻȱŘǼǯȱ the Iberian Peninsula, respectively (FishBase, Additionally, there was ulcerative keratocon- ŘŖŗśDzȱ ǰȱŘŖŗśǼǯȱȱȱȱ- ȱȱȱǰȱ ȱȱȱȱ ȱěȱPseudochondrostoma duriense. anterior and posterior corneal epithelium and ȱȱȱęȱȱȱȱȱȱ ȱĚ¢ȱęȱȱȱǰȱ ȱȱȱȱǰȱ¢ȱ¢ǰȱ conjunctiva, iris, ciliary body and the anterior ȱěǯȱ¢ȱȱȱ¢ȱ ȱȱȱ¢ȱǻȱŘǼǯȱȱȱȱȱ ȱȱ ȱȱ¢ȱȱConfederación skin, muscle tissue and eyeball were associated ¤ęȱȱ yielded values within ȱȱȱȱ¢ȱ¢ȱȱ ȱȱǻǯȱǯǼǯȱȱęȱ ȱȱ with PAS and GMS staining (Figure 2A, B, C). ȱȱȱȱ ȱǰȱȱȱ No histopathological lesions or hyphae were having spawned and others being on the point seen in internal organs (kidney, liver, spleen ȱ ǯȱȱȱȱȱ- and intestines). ȱȱȱȱȱȱȱȱȱȱ ȱȱǰȱȱȱ ȱȱȱȱȱȱȱȱȱ ¢ȱȱȱȱȱȱȱȱȱȱ ¡ȱȱ ȱSaprolegnia spp. in ȱȱȱȱȱȱȱȱ ȱ ȱȱȱęǰȱȱ ȱȱǰȱȱ Ě ǯȱȱȱěȱȱȱȱ- ȱȱȱȱȱȱȱ taneous skeletal musculature, but not internal ȱȱȱȱȱ¢ȱȱȱ ǯȱ ȱȱS. parasiticaȱȱȱȱ ȱȱȱȱ ȱ¢ęȱ ȱ ȱęȱ ȱȱȱȱ¢ȱȱȱ degenerative changes. They have also noted hyphae or histopathological lesions, so that it ȱȱ¢ȱȱěȱȱȱǰȱ may have been due to S. parasitica being in the with any spreading to other internal tissues ȱȱȱȱȱȱȱ being rarer (Bly et al., 1992; Copland and Wil- ȱȱ¢ǯȱȱȱȱȱ loughby, 1982; Fregeneda-Grandes et al., 2001; internal organs belonged to various species Hatai and Hoshiai, 1992; Neish, 1977; Nolard- and were not associated with any lesions in the Tintigner, 1973; Srivastava et al., 1994; Xu and ȱȱȱȱęǰȱȱȱ¢ȱ ȱ Rogers, 1991). ȱȱȱȱȱ¢ȱȱ ȱ ȱS. parasitica. The genus Pseudochondrostoma (Cyprinidae) contains three species, P. duriense, P. polylepis ȱȱȱ ȱȱȱȱȱ and P. willkommii (Robalo et al., 2007), which are saprolegniosis in the wild brown trout Salmo 94, Bull. Eur. Ass. Fish Pathol., 36(2) 2016 A B C Figure 2. Saprolegniosis lesions on skin, ȱȱȱ¢ȱȱPseudochondrostoma duriense (GMS stain): (A) Skin: Saprolegnia parasitica hyphae in the dermis (ÎǼȱ ȱ ¢ȱ ęȱȱȱȱȱȱȱȱǻƇ) ȱ¢ȱǻȣǼǯȱǻǼȱȱȱDZȱ ¢¢ȱ£ȱ¢ȱęȱ£ȱȱȱ £ȱȱȱ¢ȱǻȣǼȱȱȱǻƅǼȱȱ ȱȱęDzȱ¢ȱȱS. parasitica associated with muscular lesion (Î). (C) Eye: Ulcerative keratitis ȱȱȱ£ȱ¢ȱȱȱȱ the anterior and posterior corneal epithelium (Ƈ) ȱĚ¢ȱȱęȱȱȱȱǻȣǼǰȱ ȱȱȱȱȱȱ¢ȱǻƅǼDzȱ¢ȱȱS. parasitica on the cornea (Î). Ĵ L. caused by Saprolegnia parasiticaȱǻ- ęȱȱ¢ȱȱP. willkommii and not ¢ȱęȱȱSaprolegnia sp. asexual or P. polylepis ȱȱ C. polylepis in the river Saprolegnia diclina Type 1) in the upper stretch Chanza (SIBIC, 2015). It would seem that the ȱȱȱȱȱȱȱȱȱȱ ȱȱȱȱęȱȱȱȱȱ ȱȱȱȱȱǻȬ ȱȱ ȱȱ¢ȱSaprolegnia parasiticaȱ- ¤£ȬÇ£ǰȱŗşŞŝDzȱȬȱ ȱȱ ȱȱȱPseudochondrostoma duriense al., 2000, 2009), but never in P. duriense. There (“boga del Duero”). ȱȱȱȱǰȱȱȱ ȱȱSaprolegnia was not isolated or iden- Acknowledgments ęǰȱȱȱȱȱChondrostoma pol- This work was supported through research ylepisȱȱȱȱȱȱ£ȱȱȱ ȱ ŘŖŗŖȬŗŞśŜŘȱȱȱȱMinis- ȱȱ ȱȱȱȱǻ £¤£ȱ terio de Ciencia e Innovación. C.G.P. received a de Canales et al., 2001). Currently C. polylepis ȱȱȱConsejería de Educaciónȱȱ ȱęȱȱPseudochondrostoma polylepis the Regional Government Junta de Castilla y ǻȱȱǯǰȱŘŖŖŝǼȱȱȱȱ ȱ LeónǰȱȬęȱ¢ȱȱȱȱǯȱ Bull. Eur. Ass. Fish Pathol., 36(2) 2016, 95 Aller-Gancedo JM (2000). Ultrastructural We wish to express our gratitude to the Servicio ¢ȱȱSaprolegnia secondary zoospore Territorial de Medio Ambiente and the Laboratorio ¢ȱȱȱȱ ȱ Regional de Sanidad Animal de Leónȱȱȱ- brown trout, ȱ Ĵ L., and river al Government Junta de Castilla and Leónȱȱȱ water indicates two distinct morphotypes amongst long-spined isolates. Journal of Fish ȱȱȱęȱȱȱ ȱȱȱ Diseases 23, 147-160. ȱȱȱŘȱ¢ǯ Ȭ ȱ ǰȱ ¤£Ȭ Díez M and Aller Gancedo JM (2001). 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