Dermocystidium sp. in the gills of farmed Oreochromis niloticus in Brazil

Abstract

Abstract: The genus Dermocystidium is very comprehensive in the host and site of infection, however this is the first report of the occurrence of Dermocystidium sp. in the gills of Nile tilapia. This study was carried out in a fish farming located in the state of Santa Catarina, Brazil. No mortalities were reported in the facility studied and the animals were clinically healthy. During the histopathological analysis of the gills, 8.33% of the fish presented spores of Dermocystidium sp. in the gill tissue. The spores reported herein had a mean length and width of 6.206 x 5.233 μm and a refractile body diameter of 1.965 μm and were studied by histopathology and Transmission Electron Microscopy. This study highlights the importance of a new branchial pathogen in farmed tilapia, as well as to its pathogenic potential, considering the outbreaks of mortalities associated with other fish species.

Key words
fish farming; dermatocystidiosis; Nile tilapia; histology; electron microscopy; spores


INTRODUCTION

The genus Dermocystidium comprises pathogens of the order Dermocistida, class Ichthyosporea (LangenmayerLANGENMAYER MC, LEWISCH M, GOTESMAN W, HOEDT M, SCHNEIDER M, EL-MATBOULI M and HERMANNS W. 2015. Cutaneous infection with Dermocystidium salmonis in cardinal tetra, Paracheirodon axelrodi (Schultz, 1956). J Fish Dis 38: 503-506. et al. 2015). Numerous different species of Dermocystidium have been described, infecting a variety of fish and producing gill infections, skin lesions and visceral diseases worldwide (FeistFEIST SW, LONGSHAW M, HURRELL RH and MANDER B. 2004. Observations of Dermocystidium sp. infections in bullheads, Cottus gobio L., from a river in southern England. J Fish Dis 27: 225-231. et al. 2004, ZhangZHANG Q and WANG Z. 2005. Dermocystidium sp. infection in cultured juvenile catfish Silurus meridionalis in China. Dis Aquatic Organ 65: 245-250. and Wang 2005).

Among the main species of fish of interest to aquaculture that have been affected by Dermocystidium spp., is the common carp with Dermocystidium koi (HoshinaHOSHINA T and SAHARA Y. 1950. A new species of the genus Dermocystidium, D. koi sp. nov., parasitic in Cyprinus carpio L. Bull Jap Soc Sci Fish 15: 825-829. and Sahara 1950), Dermocystidium cyprini (CervinkaCERVINKA S, VITOVEC J, LOM J, HOSKA J and KUB F. 1974. Dermocystidiosise- a gill disease of the carp due to Dermocystidium cyprini n. sp. J Fish Biol 6: 689-699. et al. 1974, LotmanLOTMAN K, PEKKARINEN M and KASESALU J. 2000. Morphological observations on the life cycle of Dermocystidium cyprini Cervinka and Lom, 1974, parasitic in carps (Cyprinus carpio). Acta Protozool 39: 125-134. et al. 2000), rainbow trout with Dermocystidium macrophagi (VanVAN DE MOER A, MANIER JF, BOUIX G and VUILLAUME A. 1987. An intracellular Dermocystidium associated with proliferative kidney disease (PKD) in rainbow trout (Salmo gairdneri R.). Bull Eur Assoc Fish Pathol 7: 76-77. de Moer et al. 1987), common perch with Dermocystidium percae (PekkarinenPEKKARINEN M and LOTMAN K. 2003. Occurrence and life cycles of Dermocystidium species (Mesomycetozoea) in the perch (Perca fluviatilis) and ruff (Gymnocephalus cernuus) (Pisces: Perciformes) in Finland and Estonia. J Nat Hist 37: 1155-1172. and Lotman 2003), several salmonids with Dermocystidium salmonis (OlsonOLSON RE, DUNGAN CF and HOLT RA. 1991. Water-borne transmission of Dermocystidium salmonis in the laboratory. Dis Aquatic Organ 12: 41-48. et al. 1991, Olson and Holt 1995OLSON RE and HOLT RA. 1995. The gill pathogen Dermocystidium salmonis in Oregon salmonids J Aquat Anim Health 7: 111-117.), kinguio, catfish and Nile tilapia with unidentified species of Dermocystidium spp. (Zhang and Wang 2005, MahmoudMAHMOUD MA, ALY SM, DIAB AS and JOHN G. 2009. The role of ornamental goldfish Carassius auratus in transfer of some viruses and ectoparasites to cultured fish in Egypt: comparative ultra-pathological studies. Afr J Aquat Sci 34: 111-121. et al. 2009, ShaheenSHAHEEN AA. 2000. Some studies on dermatocystidiosis among Oreochromis niloticus fish. Suez Canal Vet Med 3: 177-184. 2000). Still in tilapia, El-MansyEL-MANSY A. 2008. A New Finding of Dermocystidium - like Spores in the Gut of Cultured Oreochromis niloticus. Global Vet 2: 369-371. (2008) identified D. aegyptiacus in the intestinal epithelium of farmed O. niloticus in Egypt.

Gill infection by Dermocystidium is pathogenic and causes mortality in farmed salmonids (Olson et al. 1991), carps (Cervinka et al. 1974) and eels (WoottenWOOTTEN R and MCVICAR AH. 1982. Dermocystidium from cultured eels, Anguilla anguilla L., in Scotland. J Fish Dis 5: 215-222. and McVicar 1982, MolnárMOLNÁR K and SOVÉNYI JF. 1984. Dermocystidium anguillae infection in elvers cultured in Hungary. Aquacult Hung IV: 71- 78. and Sovenyi 1984). In Brazil, this pathogen was reported by EirasEIRAS JC and SILVA-SOUZA AT. 2000. Dermocystidium infection in Trichomycterus sp. (Osteichthyes, Trichomycteridae). Parasite 7: 323-326. and Silva-Souza (2000) in Trichomycterus sp., and more recently by FujimotoFUJIMOTO RY, COUTO MVS, SOUSA NC, DINIZ DG, MADI RR, MARTINS ML and EIRAS JC. 2017. Dermocystidium sp. infection in farmed hybrid fish Colossoma macropomum x Piaractus brachypomus in Brazil. J Fish Dis 00: 1-4. et al. (2017) in cultures of hybrid fish tambatinga; however, this is the first report of Dermocystidium sp. infecting gills of Nile tilapia Oreochromis niloticus (Linnaeus, 1758).

MATERIALS AND METHODS

The procedures adopted for this study were approved by the Committee on Ethics in the Use of Animals of Federal University of Santa Catarina - CEUA No PP00928. Sixty adult tilapia (average weight 480.9 ± 210.2 g and average length 28.1 ± 4.2 cm) were captured from 12 fish farms located in the state of Santa Catarina, southern Brazil. The study was conducted between May and December 2015. The fish were anesthetized with eugenol (75 mg L-1) and euthanized by rapid cerebral concussion. Then, the first right branchial arch was removed, divided and fixed in 10% buffered formalin and 2.5% glutaraldehyde.

The samples previously fixed in 10% buffered formalin were dehydrated in progressive graduation of alcohol, diaphanized in xylol and embedded in paraffin. Using a microtome PAT-MR10 (The Pathologist®, Brazil), samples were sectioned in 3 μm and stained with Harris haematoxylin and eosin (HH & E), mounted on permanent blades with Entellan® and analyzed by DIC (Differential Interference Contrast) microscope model Axio Imager A2 (Zeiss®, Germany).

For analysis in transmission microscopy, the gills were fixed in 2.5% glutaraldehyde in 0.1M sodium cacodylate buffer, pH 7.2 for 24 hours. Post-fixed with osmium tetroxide solution, dehydrated in increasing solution of ethanol and transferred to ethanol: spurr resin. Ultra thin (60 nm) sections were cut with a diamond blade and stained with uranyl acetate and lead citrate for microscopic observation (JEOL JEM-1011). The identification of the parasite was performed through morphological characteristics of the spores, such as refractile body, vacuoles and cytoplasm. To determine the approximate size of the spores, were performed by light microscopy at a magnification of 100x, the measurements of length (μm), width (μm) and refractile body diameter (μm) of 53 spores randomly found on the histological sections of the infected animals.

The spores (Figure 1) found in this study are spherical, with central refractile body and presented an average size of 6.206 x 5.233 μm (Table I). The size of the spores is similar to that reported by Wootten and McVicar (1982) who observed spheres of 4 to 7 μm spherical, measuring 25 cells, in gills of Anguilla anguilla. Based on the histological and ultrastructural findings, gill dermocystitis was diagnosed.

Figure 1
Photomicrographs of the gill tissue. a. interlamellar epithelial hyperplasia along the gill filaments (bar = 20 μm). b. Spores of Dermocystidium sp. between the secondary lamellae of gills of Nile tilapia, evidencing the hyaline cytoplasm (HH & E staining, 20 μm bar).

The presence of spores was observed through routine histopathological examination (Figure 1) and supported by Transmission Electron Microscopy (Figure 2).

Figure 2
Transmission Electron Microscopy of Dermocystidium spores showing the structures. V – vacuole; G – Golgi-complex; N – nucleus; M – mitochondria. (bar = 1 μm). b. Transmission Electron Microscopy of Dermocystidium spores showing the hyaline cytoplasm (bar = 2 μm).
TABLE I
Measurements (μm) of the spores of Dermocystidium sp. (n = 53) collected in the gills of Oreochromis niloticus,cultivated in Santa Catarina, southern Brazil. (Mean ± standard deviation).

According to BrunoBRUNO DW, NOWAK B and ELLIOTT DG. 2006. Guide to the identification of fish protozoan and metazoan parasites in stained tissue sections. Dis Aquatic Organ 70: 1-36. et al. (2006) Dermocystidium species that infect fish, are located in epithelial tissue of the skin, fins, gills or visceral organs. Infections usually appear as small white, round or oval nodules, or cysts in the affected tissue. In the histological sections, species attributed to the genus Dermocystidium are characterized by a spherical spore stage, with a large central vacuole or solid refractile body and the cytoplasm with the nucleus restricted to a narrow peripheral layer. In most species, the spores are relatively uniform in size (3 to 12 μm in size, depending on the species).

The histological sections of the present study showed gill alterations such as interlamellar epithelial hyperplasia (Figure 1a), secondary lamella epithelial hyperplasia and fusion of secondary lamellae. According to BrunoBRUNO DW. 2001. Dermocystidium sp., in Scottish Atlantic salmon, Salmo salar. Evidence for impact on fish in marine fish farms. Bull Eur AssoC Fish Pathol 21: 209-213. (2001) epithelial hyperplasia and the fusion of the gill lamellae are common alterations in the infection by Dermocystidium.

During collection, it was not possible to observe the fresh cysts in the gills and the diagnosis was made through histopathological analysis. Feist et al. (2004) report in their samples that several ruptured cysts were observed in the Cottus gobio epithelium, allowing the release of spores into the environment. The aforementioned authors believe that this is the usual mechanism of spore dispersion.

CONCLUSIONS

Based on the few morphological descriptions of Dermocystidium sp. found in the literature, it was not possible to identify the species. The authors report that it is difficult to compare the size of the spores, the shape and stage of development of the cysts, and the lack of sufficient data for the studied fish species (Feist et al. 2004).

Corroborating the results of the present study, Wootten and McVicar (1982) did not observe mortality that could be attributed directly to the infection by Dermocystidium sp. in farmed of Anguilla anguilla. Presumably, tilapia is a rustic fish, able to ensure adequate oxygen through the unaffected parts of the gills or skin. This contrasts with the severe disease outbreak and mortality in chinook salmon Oncorhynchus tshawytscha (Walbaum) and carp Cyprinus carpio (PauleyPAULEY GB. 1967. Prespawning adult salmon mortality associated with a fungus of the genus Dermocystidium. J Fish Res Board Can 24: 843-848. 1967, AllenALLEN RL, MEEKIN TK, PAULEY GB and FUJIHARA MP. 1968. Mortality among chinook salmon associated with the fungus Dermocystidium. J Fish Res Board Can 25: 2467-2475. et al. 1968, Cervinka et al. 1974) caused by Dermocystidium in the gills.

The occurrence of Dermocystidium sp. in the gills of Nile tilapia, recorded for the first time in asymptomatic fish, may be due to an infection detected at an early stage. Therefore, it is attentive to a possible emerging pathogen in farmed tilapia in Brazil. It is suggested the monitoring of fish farms to complete the identification of the agent, as well as epidemiological investigation and its pathogenicity in Nile tilapia.

ACKNOWLEGMENTS

The authors are grateful to the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) for financial support (CNPq 446072/2014-1) and research grant to ML Martins (CNPq 305869/2014-0), the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES-EMBRAPA n. 15/2014) for the award of the Master’s Scholarship to LD Steckert. To Luiz Rodrigo Motta Vicenti (Epagri), Ofélia Maria Campigotto (Gaspar fish farmers’ association), Susane Pahl-Klipp (Municipal Rural Development Foundation July 25) and Marcelo Tonial and Alexander Hilata (Nicolluzi) for assistance in collecting and to the fish farms of the state of Santa Catarina that donated the fish.

REFERENCES

  • ALLEN RL, MEEKIN TK, PAULEY GB and FUJIHARA MP. 1968. Mortality among chinook salmon associated with the fungus Dermocystidium. J Fish Res Board Can 25: 2467-2475.
  • BRUNO DW. 2001. Dermocystidium sp., in Scottish Atlantic salmon, Salmo salar. Evidence for impact on fish in marine fish farms. Bull Eur AssoC Fish Pathol 21: 209-213.
  • BRUNO DW, NOWAK B and ELLIOTT DG. 2006. Guide to the identification of fish protozoan and metazoan parasites in stained tissue sections. Dis Aquatic Organ 70: 1-36.
  • CERVINKA S, VITOVEC J, LOM J, HOSKA J and KUB F. 1974. Dermocystidiosise- a gill disease of the carp due to Dermocystidium cyprini n. sp. J Fish Biol 6: 689-699.
  • EIRAS JC and SILVA-SOUZA AT. 2000. Dermocystidium infection in Trichomycterus sp. (Osteichthyes, Trichomycteridae). Parasite 7: 323-326.
  • EL-MANSY A. 2008. A New Finding of Dermocystidium - like Spores in the Gut of Cultured Oreochromis niloticus. Global Vet 2: 369-371.
  • FEIST SW, LONGSHAW M, HURRELL RH and MANDER B. 2004. Observations of Dermocystidium sp. infections in bullheads, Cottus gobio L., from a river in southern England. J Fish Dis 27: 225-231.
  • FUJIMOTO RY, COUTO MVS, SOUSA NC, DINIZ DG, MADI RR, MARTINS ML and EIRAS JC. 2017. Dermocystidium sp. infection in farmed hybrid fish Colossoma macropomum x Piaractus brachypomus in Brazil. J Fish Dis 00: 1-4.
  • HOSHINA T and SAHARA Y. 1950. A new species of the genus Dermocystidium, D. koi sp. nov., parasitic in Cyprinus carpio L. Bull Jap Soc Sci Fish 15: 825-829.
  • LANGENMAYER MC, LEWISCH M, GOTESMAN W, HOEDT M, SCHNEIDER M, EL-MATBOULI M and HERMANNS W. 2015. Cutaneous infection with Dermocystidium salmonis in cardinal tetra, Paracheirodon axelrodi (Schultz, 1956). J Fish Dis 38: 503-506.
  • LOTMAN K, PEKKARINEN M and KASESALU J. 2000. Morphological observations on the life cycle of Dermocystidium cyprini Cervinka and Lom, 1974, parasitic in carps (Cyprinus carpio). Acta Protozool 39: 125-134.
  • MAHMOUD MA, ALY SM, DIAB AS and JOHN G. 2009. The role of ornamental goldfish Carassius auratus in transfer of some viruses and ectoparasites to cultured fish in Egypt: comparative ultra-pathological studies. Afr J Aquat Sci 34: 111-121.
  • MOLNÁR K and SOVÉNYI JF. 1984. Dermocystidium anguillae infection in elvers cultured in Hungary. Aquacult Hung IV: 71- 78.
  • OLSON RE, DUNGAN CF and HOLT RA. 1991. Water-borne transmission of Dermocystidium salmonis in the laboratory. Dis Aquatic Organ 12: 41-48.
  • OLSON RE and HOLT RA. 1995. The gill pathogen Dermocystidium salmonis in Oregon salmonids J Aquat Anim Health 7: 111-117.
  • PAULEY GB. 1967. Prespawning adult salmon mortality associated with a fungus of the genus Dermocystidium. J Fish Res Board Can 24: 843-848.
  • PEKKARINEN M and LOTMAN K. 2003. Occurrence and life cycles of Dermocystidium species (Mesomycetozoea) in the perch (Perca fluviatilis) and ruff (Gymnocephalus cernuus) (Pisces: Perciformes) in Finland and Estonia. J Nat Hist 37: 1155-1172.
  • SHAHEEN AA. 2000. Some studies on dermatocystidiosis among Oreochromis niloticus fish. Suez Canal Vet Med 3: 177-184.
  • VAN DE MOER A, MANIER JF, BOUIX G and VUILLAUME A. 1987. An intracellular Dermocystidium associated with proliferative kidney disease (PKD) in rainbow trout (Salmo gairdneri R.). Bull Eur Assoc Fish Pathol 7: 76-77.
  • WOOTTEN R and MCVICAR AH. 1982. Dermocystidium from cultured eels, Anguilla anguilla L., in Scotland. J Fish Dis 5: 215-222.
  • ZHANG Q and WANG Z. 2005. Dermocystidium sp. infection in cultured juvenile catfish Silurus meridionalis in China. Dis Aquatic Organ 65: 245-250.

Publication Dates

  • Publication in this collection
    23 Sept 2019
  • Date of issue
    2019

History

  • Received
    12 Sept 2018
  • Accepted
    22 Nov 2018
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