epizootic ulcerative syndrome

Other Scientific Names

• mycotic granulamotoses
• red spot disease

International Common Names

• English: Aphanomyces invadans infection

English acronym

• EUS
• MG
• RSD

Epizootic ulcerative syndrome (EUS) is a disease of freshwater and brackishwater fish that has other synonyms such as mycotic granulomatosis (MG), red spot disease (RSD) and ulcerative mycosis (UM). More than 100 species of fish species with a wide geographic distribution have been reported to be affected by EUS. EUS outbreaks occur only when a number of causal factors combine.

The causative agent of this disease is the fungus, Aphanomyces invadans. There are a number of sufficient causes that lead to exposure of the dermis to fungal spore germination. In the early stages, red spots or small haemorrhagic lesions are found on the body surface. Intermediate stage lesions become small ulcers. Advanced stage lesions expand into large necrotic open ulcers resulting in death.

Positive diagnosis of EUS is made by the presence of mycotic granulomas in histological section and isolation of Aphanomyces invadans from infected fish. Quarantine and health certification practice for the movement of live fish between countries or regions are effective means of preventing the spread of EUS to new areas. For areas where EUS is endemic, prevention programmes should include eradication, exclusion, management, surveillance and treatment.

Outbreaks of EUS have continued to occur periodically in all target countries. The immunity of fish surviving EUS infections should be the subject of interest because records from many countries showed that the severity of EUS outbreak after the first few years tends to decrease (Mohan and Shankar, 1994; Bondad-Reantaso et al., 1992). The serious implications for the development of vaccines against EUS should be deliberated because there was a report on the strong reaction between the sera from immunized fish with the spores and mycelium of pathogenic strains of Aphanomyces and the extracts of pathogenic strains of Aphanomyces (Thompson et al., 1997).

Pathogenic Agents

Based on the investigation of the pathogenic organisms found from EUS-infected samples, a wide range of aetiologies have been attributed to EUS, such as bacteria, fungi, parasites and viruses.

Several species of parasitic protozoans (Chilodonella sp., Costia sp., Epistylis sp., Glossatella sp., Ichthyophthirius sp., Scyphidia sp., Trichodina spp.), myxosporeans (Henneguya sp. and Thelohania sp.), monogeneans and crustaceans (Lernaea sp.) have been recorded from diseased fish (Callinan et al., 1997; Reungprach et al., 1983). Nevertheless, there is no evidence to confirm that these parasites are the causative agent of the disease. Parasitic infections possibly induce stress in fish, thus predisposing them to infection (Subasinghe, 1993).

Aeromonas hydrophila, A. sobria, Pseudomonas spp. and Vibrio spp. are the species of bacteria isolated from the internal organs of EUS-infected fish (Burke and Rodgers, 1981; Llobrera and Gacutan, 1987; Tonguthai, 1985). Among these bacteria, A. hydrophila is the most commonly isolated species from the advanced stages of infected fish. It is usually not isolated from the early stage of the disease. On the other hand, A. hydrophila is known to be part of the normal microflora of fish and aquatic environments and is recognised as an opportunistic pathogen.

A number of birnaviruses, reoviruses and rhabdoviruses have been isolated from diseased snakehead and other susceptible species. These viruses most probably represent adventitious infections unrelated to EUS because of their heterogeneous nature, together with a low and non-consistent level of recovery from epizootics of disease. Therefore, the role of viruses in the aetiology of this disease remains unproven (Kanchanakarn, 1996; Roberts et al., 1994).

At least two species of fungi have been isolated from EUS-infected fish, namely, Achlya sp. which was isolated from the superficial area of the lesions (Pittchayangkula and Bodhalamik, 1983; Subasinghe et al., 1990) and Aphanomyces sp. which is normally isolated from the muscle area near the lesions (Egusa, 1992, Fraser et al., 1992; Hatai et al., 1994; Roberts et al., 1993). Recently, it has been shown that all Aphanomyces identified from EUS samples not only are the same species but also a single clonal lineage (Lilley et al., 1997). Therefore, it is confirmed that the fundamental aetiological agent of EUS is an oomycete fungus, Aphanomyces invaderis (Willoughby et al., 1995). However, the valid taxonomic name according to the International Code of Botanical Nomenclature (ICBN) of this species is A. invadans (Lilley et al., 1998). This specific pathogenic fungus showed the ability to invade the skin of EUS-infected fish (Kiryu et al., 2003). Isolation of this particular fungus is difficult as it is slow growing and most of the hyphae in the muscle are dead except for the tips that are penetrating deeper into the muscle tissue (Roberts et al., 1993)

Pathogenicity of the Fungus

After success in isolating the slow growing fungus, Aphanomyces invadans, from EUS-infected fish in various countries (Egusa, 1992; Fraser et al., 1992; Willoughby et al., 1995), pathogenicity studies were conducted. Intramuscular injection of spore suspensions into some susceptible fish species (barbs and snakehead) demonstrated a severe necrotising myogranulomatous condition at the injected site similar to the pathological feature of fish naturally infected with EUS. At the later stages, the fungus rapidly invaded the other internal organs, causing death to the host. The diameter of the hyphae of the fungus in lesions was always larger and the walls thicker than in cultured specimens (Roberts et al., 1993). Co-habitation transmission experiments with EUS-infected fish were successful (Balasuriya et al., 1990; Cruz-Lacierda and Shariff, 1995).The exposure of healthy Atlantic menhaden to suspension of zoospores of A. invadans demonstrated the germination and penetration of the germ tube through the epithelium layer of the experimental fish (Kiryu et al., 2003).

Histopathology

Lesions in all EUS-infected fish are similar, except that in snakehead chronic lesions can develop. This is because snakehead survive longer and the lesions are able to develop to a very advanced stage; other species die before this stage is reached.

Grossly the lesions are generally small red erosions on the body, head and fins. In the advanced stage, deep haemorrhaged ulcers spread through the whole body. Fungal mycelium is commonly found on top of the ulcerative lesions. The internal organs in the early stage of infection show only minimal inflammatory response. It is important to note that diseased fish with not too advanced lesions, kept in better water quality conditions, often recover and undergo a healing process leaving dark scars at the sites of infection.

Histopathological observation of early stage lesions reveal acute spongiosis with loss of epithelial cells. Degenerative changes are observed in the dermis, with hyperaemia, haemorrhages and inflammatory cells infiltrating between the fibres of the dermis. Inflammatory exudate and haemorrhages are also found in the hypodermis. Sarcolysis with haemorrhages and inflammatory exudate are obvious in the more advanced stage. The fungal hyphae are enclosed by a well developed epithelioid cell layer. Mycotic granulation of these non-septate fungal hyphae spreads through the infected muscle and other internal organs in the very late stages. In the advanced stage, there are no muscle fibres in the granuloma tissues. They are replaced by fibrosis, inflammatory cells and newly developing capillaries (Chinabut et al., 1995; Chinabut and Roberts, 1999). This distinct feature of the typical mycotic granulomas in the lesions of EUS-infected fish provides important histopathological information which can be used to distinguish EUS from other fish ulcerative diseases.

Cutaneous ulcerative disease, named epizootic ulcerative syndrome (EUS), has been a seasonal disease of freshwater and brackishwater fish in Asia, Australia and the United States of America since 1971. Earlier outbreaks of mycotic granulomatosis (MG) in Japan, red spot disease (RSD) in Australia and ulcerative mycosis (UM) in the United States of America are now known to be the same as EUS (Kiryu et al., 2003). EUS commonly occurs during periods of low temperature and after periods of heavy rainfall (Bondad-Reantaso et al., 1992). These conditions favour sporulation of the Aphanomyces pathogen (Lumanlan-Mayo et al., 1997) and low temperature has been shown to delay the inflammatory response of fish to fungal infection (Chinabut et al., 1995; Catap and Munday, 1998). It has been defined as "a seasonal epizootic condition of freshwater and estuarine warm water fish of complex infectious aetiology characterized by the presence of invasive Aphanomyces infection and necrotising ulcerative lesions typically leading to a granulomatous response" (Roberts et al., 1994). Recent studies suggest that a complex infectious aetiology is not necessarily involved in all cases of EUS (Callinan, 1997, Kiryu et al., 2003); and one species of invasive fungus, Aphanomyces invadans, is involved in the outbreaks. Region wide, over 100 species have been confirmed affected by histological diagnosis in Asia (Lilley et al., 1998), but some important cultured species, including Chinese carps, milkfish and tilapia, have been shown to be resistant.