14 March 2009 Discussion IMN is a relatively new pathology for L. vannamei, though it has already been described in several other crustacean species. Field clinical identification of IMN is an important preliminary step in the management and control of this disease during production, especially when laboratory diagnostic approaches are not available. Even though we have not shown histological or ultrastructural evidence of this viral disease, a prominent existing body of literature, including presentations of well documented and detailed histopathological and electron microscope slides concerning with IMN, can be found in Nash et al. (1987), Stentiford and Neil (2000), Lightner et al. (2004) and Tang et al. (2005). Both histopathologically and ultrastructurally, microscopic diagnosis is characterized by myofibrillar and sarcoplamic necrosis or fibrosis, with hemocytic infiltration during the chronic phase (Nash et al. 1987, Lightner et al. 2004). Shrimp with acute and chronic IMN showed lesions with coagulative muscle necrosis and coagulative–liquefactive necrosis, respectively Fig. 4. Unhealthy individuals showing pre-acute or grade 3 (indicated by a white arrow) and acute phase or grade 4 (indicated by arrow) during shrimp production. It is noted almost total necrosis of the abdominal segments (white color) and presence of red uropods during the acute phase. (Credit picture: ®Gustavo Dominguez). Fig. 5. A clear distinction of the IMN-acute phase (grade 4), when compared to a healthy Pacific white shrimp. (Credit picture: ®Gustavo Dominguez). (Lightner et al. 2004). The external appearance of infected individuals in advanced stages is similar to the morphological alterations previously found in penaeids, showing the myonecrosis in the distal abdominal segments, mainly from four to six (Rigdon and Baxter 1970, Lakshmim et al. 1978). Nash et al. (1987) pointed out that the myonecrosis is likely to predominate in distal segments inasmuch as this abdominal region presents the highest metabolic activity, most obviously in hypoxic conditions during hyperactive stress. The chronic advanced stage of myonecrosis and septic form of this disease is reached when the distal region of the abdomen turns red, becoming entirely necrotic and generally linked with shrimp mortality (Lightner 1993). Tang et al. (2005) reported that typical IMNV lesions on L. vannamei, injected with purified virions, were exhibited after six days, with a mortality of 20 percent. However, diagnosis of IMN based on only clinical signs and histopathological examinations is not sufficient to confirm the disease, so other methods, such as in situ hybridizations (ISH) to detect shrimp virus have been found to be effective for definitive IMN diagnosis (Tang et al. 2005). As evidenced by recent bioassays conducted in Brazil, the major route of exposure to this disease appears to be ingestion of contaminated food or infected tissues of shrimp when compared to fecal matter of birds fed with contaminated shrimp, horizontal infection using effluents from cultured contaminated shrimps and biomass of adult Artemia fed with contaminated shrimp extracts.3 From those preliminary experiments, the transmission of the IMN virus by direct consumption of contaminated tissues, 1.6 g of infected material caused a maximum mortality of 35 percent in a population of 25 juvenile shrimp ranging 0.2-0.5g (total biomass = 5-12.5 g) after 24 hr exposure under laboratory conditions (salinity = 35, temperature = 25-28° C; feeding was 5 percent of biomass/day with 35 percent protein; water was exchanged at 50 percent daily).3 Additionally, several environmental and intrinsic factors can promote the generation of IMN. For example, dramatic oscillations in salinity and temperature, hypoxia, overstocking, hyperactivity, poor handling and transfer techniques, collection by cast-net, direct solar radiation and low-quality feeds have been identified as stress factors associated with IMN (Rigdon and Baxter 1970, Lakshmim et al. 1978, Nash et al. 1987, Lightner 1993, Lightner 1988, Lightner et al. 2004). IMN induced by environmental stressors has been reported in marine shrimp, such as P. aztecus, P. japonicus and P. californiensis (Rigdon and Baxter 1970, Lakshmim et al. 1978, Momoyama and Matsuzato 1987). If predisposing environmental stressors are removed preior to development of the advanced grades of infections, IMN signs can be arrested (Rigdon and Baxter 1970, Lakshmim et al. 1978). Nash et al. (1987) reported that reduction of stocking density by transferring Asian freshwater shrimp postlarvae affected by IMN to different and lower density ponds apparently helps mitigate the mortality. Management health strategies, such as total cleaning and aseptic conditions associated with tanks and equipment, avoiding overcrowding and maintaining optimal aeration have been used successfully in prevent-
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