e Rese tur arc ul h c & a u D q e A v e f l o o Zadeh et al., J Aquac Res Development 2014, 5:7 l p a m n Journal of Aquaculture r e u n o t DOI: 10.4172/2155-9546.1000289 J ISSN: 2155-9546 Research & Development Review Article OpenOpen Access Access The Detection of Ichthyophonus hoferi in Naturally Infected Fresh Water Ornamental Fishes Mahsa JafariZadeh1*, Rahim Peyghan2 and Shadi Eftekhar Manavi2 1Fisheries Department, Islamic Azad University, Ahvaz Branch, Ahvaz-Iran, Iran 2Veterinary Faculty, Shahid Chamran University of Ahvaz, Ahvaz, Iran Abstract Ichthyophoniasis is one of the most important systematic infections among different species of fishes that caused by Ichthyophonus hoferi. In the present study, we have reported this parasite from two species of ornamental fish, black tetra (Gymnocorymbus ternetzi) and tiger barb (Pentius tetrazona) in Ahvaz-Iran. Examined fishes had marked signs such as abnormal swimming, lethargy, swelling abdominal and low rate mortality. In this study, the two phases of life cycle of I. hoferi involving active and passive detected. The obvious internal sign was white cysts and nodules, which embedded in infected spleens. The cysts were full of schizonts that were surrounded by collagen fibers and many eosinophilic cells. Plasmodium spherical bodies with variable sizes were detected by microscopic examination of wet mount squash from the infected organs. In addition, histopathology studies showed that there were many granulation tissues surrounded by multilayer connective tissues in the infected tissues. Tissue samples were also isolated and put in to Minimum Essential Medium (MEM) to detect the germination of Ichthyophonus hoferi for distinguish Ichthyophoniasis from Mycobacterial infections. Keywords: Ichthyophonus hoferi; Ornamental fishes; Black tetra; (Gymnocorymbus ternetzi) and swollen abdominal of one tiger barb Tiger barb; Histopathology; Nodules; Spleen (Pentius tetrazona). This aquarium, involved 35 adult fish from both species. This facility was filled with closed systems water, tap water, Introduction of Ahvaz city. As a random seven fishes from both species, five black Ichthyophonus hoferi was first identified in cultured brown and tetra and two tiger barb were collected. After anesthesia, the fishes were brook trout in Germany by von Hofer in 1893. The protozoan nature killed, and total length and total weight of the fishes were measured of I. hoferi described and named by Plehn and Mulsow in 1911 as a and recorded. Tiger barbs had average length of 5.2 ± 1.2 cm and fungus. Ultimately this parasite classified into Mesomycetozoea [1-3]. average weight of 1.5 ± 0.5 g. The black tetras average length was 5.3 ± Signs are species-related and depend on the condition of individual 0.8 cm and average weight was 2.5 ± 0.3 g. All organs were examined fish.Ichthyophonus hoferi causes a systematic granulomatous infection by routine clinical methods. Within examination the pericardial cavity, in their hosts. I. hoferi has produced infections in many different many visible white nodules were seen in spleens of the studied fishes. In species of freshwater, estuarine, marine teleosts and adapt to a wide addition, one specimen had some nodules in heart organ. range of environmental conditions [4]. Ichthyophonus sp. has been Microscopic examination reported from many temperate and some tropical waters. Couch in 1985 detected two cases of fishes, sea catfish, and spot, infected by I. Wet and dry smear were prepared from spleen tissues and the dry hoferi in Gulf coast. For the first time Ichthyophonus sp. is reported in smears were stained with Giemsa staining. two cultured marine fish including Mugil capito and Lia salines from Tissue culture Spain [5]. This disease has records from over 80 species of both marine and freshwater fishes and results into mass mortalities and economic Samples from infected tissues were taken and incubated in MEM losses [6]. This parasite detected in several commercial fish [7]. In medium (Eagle’s minimum essential medium supplemented with 10% the case of aquarium fish, Ichthyophonus hoferi reported in sumatra fetal bovine serum, 100 IU/ml penicillin, and 100 mg/ml streptomycin). barb (Systomus tetrazona) and black tetra (Gymnocorymbus ternetzi) Cultures were incubated in 25°C and the tissue cultures were examined by Reichenbach-Klinke, H in 1954 and 1955. Ozturk et al. in 2010 every day under microscope. Eventually after 2 weeks, the tissue dry screened this pathogen in some organs of goldfish (Carassius auratus). smear was prepared and stained by Giemsa method. Discus fish (Symphysodon.spp) is the new host record of ornamental fish that infected by I. hoferi [8]. I. hoferi reported in herring in the northwest Atlantic (up to 25% infection) with associated reductions in the population, up to 10% infection in U.K waters with mortality, and *Corresponding author: JafariZadeh M, Fisheries Department, Islamic Azad mackerel up to 69% infection and haddock up to 80% infection [9]. University, Ahvaz Branch, Ahvaz-Iran, Iran, Tel: 989163128034; E-mail: There are many reports that I. hoferi causes mortality in post-smolts [email protected] of Atlantic salmon and Chinook salmon [10,11]. The purpose of the Received October 12, 2014; Accepted November 07, 2014; Published November present study is the description of the morphology and pathology of I. 12, 2014 hoferi as found in two species of ornamental fish in Iran. Citation: Zadeh MJ, Peyghan R, Manavi SE (2014) The Detection of Ichthyophonus hoferi in Naturally Infected Fresh Water Ornamental Fishes. J Aquac Res Case Presentation Development 5: 289. doi:10.4172/2155-9546.1000289 In a disease case, from a local aquarium fish propagation center, Copyright: © 2014 Zadeh MJ, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted the culture facility with the size of 40 cm in length and 30 cm in width use, distribution, and reproduction in any medium, provided the original author and was regarded because of swimming disorders of one the black tetra source are credited. J Aquac Res Development ISSN: 2155-9546 JARD, an open access journal Volume 5 • Issue 7 • 1000289 Citation: Zadeh MJ, Peyghan R, Manavi SE (2014) The Detection of Ichthyophonus hoferi in Naturally Infected Fresh Water Ornamental Fishes. J Aquac Res Development 5: 289. doi:10.4172/2155-9546.1000289 Page 2 of 5 Histopathological sampling and examination Samples from heart, liver, intestine, gonads and spleen were cut and fixed in 10% phosphate-buffered formalin for histological examination. By using standard techniques, samples were processed and then sections stained either in haematoxylin-eosin [12] and Periodic Acid-Schiff. Micrometric measurement was carried out using Sa-Iran camera microscope and Axio-vision software. Results Infection to Ichthyophonus hoferi was found in internal organs of all examined aquarium fishes including five black tetra and two tiger barb. During gross examination, white nodules were found on spleen organs of both fish species. In one specimen (tiger barb), some nodules were also seen in heart organ. However, no external sign of Figure 2: A. Encapsulated schizonts. B. Un-encapsulated schizonts, inside well-defined host cellular granulomas in naturally infected tiger barb, spleen, infection was observed on the body surface of naturally infected fishes. x40 magnification. The morphology and developmental stage of life style of I. hoferi were similar in both specimens. Schizonts of Ichthyophonus hoferi were the most commonly characteristic of squashed infected organs. The size of the schizonts, include the wall varied from 54 µm to 182 µm in all of specimens (Average 85.10). Differences between schizonts size indicate that, the schizonts of black tetras were bigger compare to the tiger barbs. Microscopic examination of wet mount squash of infected organs were represented, thick walled spherical bodies (resting schizonts) (Figure 1). The subdivision of the contents of each of the spherical bodies results to produce small uninucleate stages. The reason for this incident is rotational movement in the wall of spherical body (Woo and Btuno 2006). By studying the histopathological sections encapsulated and un-encapsulated schizont inside well-defined host cellular granulomas were observed. In some cases encapsulated schizont consist of 3-5 fibrous layers or fibrotic capsule that related to the passive phase and, some nodules had thin fibrotic layer, which represented active phase (Figure 2). In addition, the Hydropic degeneration observed in the infected spleen. Detected black spots (melanin reaction) around schizonts had different sizes from 6.77 µm to 2.49 µm (average 4.57 µm) in infected spleen. After incubation in Minimum Essential Medium, Figure 3: A. Plasmodium. B. Existing a collapsed Ichthyophonus hoferi the germination of the infective agent and formation of new budding schizonts with two plasmodia. Culture media (MEM). Black tetra, spleen, yeast of plasmodium, non-septate germination tubes and terminating x400 magnification. club-shaped cells from Ichthyophonus schizonts, were seen. (Figures 3-5). Then the two plasmodia free within the tissue and migrating away black tetras showed the black spots or melano-macrophage centers. from an empty schizonts (Figure 3). Histopathologic examination of Although, these melanized centers were not seen in all examined tiger the lesion of spleen showed the presence of characteristics granuloma. barbs.