Protozoal and epitheliocystis-like infections in the introduced

More documents

Recommendations

Info

60 Dis Aquat Org 57: 59–66, 2003 that at least one nematode species was introduced into Hawaii with the importation of taape. Okihiro (1988) found a high prevalence of skin tumors on native butterfly fish (Chaetodon multicinctus and C. miliaris) in Maui, and suspected contaminants as a possible cause, based on the distribution of affected fish. Kent & Heidel (2001) found that pen-raised opakaka Pristipomoides filamentosus suffered mainly from bacterial infections of swim bladder and hemorrhage and emphysema of the eyes, although various protozoan and metazoan parasites were noted at low infection levels. As part of a larger study, taape caught near Oahu were surveyed for systemic parasites to provide baseline information on their health status. MATERIALS AND METHODS During 2001 and 2002, fish were collected using the line and hook method at depths ranging from 15 to 70 m throughout Southern Oahu. Fish were anesthetized with MS-222 in seawater, and bled from the caudal tail vein using sterile 3 cc syringes and 1 × 38 mm needles. Blood smears were made immediately, air dried, and fixed in absolute methanol. Fish were then humanely euthanized with an overdose of MS-222 in seawater. Necropsies consisted of measuring total and fork length (0.5 cm) with a ruler, weight (0.1 g) with an electric scale, and a complete external and internal exam. Spleen, liver, cranial and caudal kidneys, swim bladder, brain, heart, skeletal muscle, gill, and gonad were stored in 10% neutral buffered formalin. Tissues were processed for histology by paraffin embedding, sectioning at 5 µm, and staining with hematoxylin and eosin. Tissues were examined microscopically for presence of lesions and signs of infectious organisms. Giemsa was used to identify protista, and Gimenez, and Gram stains were used to identify bacteria (Prophet et al. 1992). Protista included microsporidians and coccidia, bacteria included epitheliocystis-like organisms, and metazoans included myxozoa, trematodes or their eggs, and migratory tracts associated with helminth. For electron microscopy, tissues were fixed in Trump’s fixative (McDowel & Trump 1976), rinsed in 0.1 M Sorenson’s phosphate buffer, and post fixed in 2% osmium tetroxide. Epoxy embedded tissues were cut into 1 µm thick toluidine blue-stained sections. Ultrathin sections were stained with uranyl acetate, post stained with lead citrate and examined with a Zeiss EM 109 electron microscope. Blood smears were stained with Wright’s Giemsa and examined for the presence of hemoparasites. Data were tested for normality and equal variance. We used linear regression to evaluate the relationship between weight and fork length, the t-test to evaluate the difference in weight between males and females, and ANOVA to compare weights of parasitized and parasite-free fish (Daniel 1987). For all comparisons, α = 0.05. RESULTS A total of 120 taape (78 males, 42 females) were examined. Mean weight (mean ± SD) of males 241.7 ± 59.5 was significantly (t = 6.2, df = 118, p < 0.001) greater than that of females 178.7 ± 40.2. Grossly, no lesions were seen other than trauma associated with capture. There was a significant relationship between fork length and weight (F = 1535, df = 119, R 2 = 0.93, p< 0.0001). With microscopy, protista were the most common organisms, which infected 56 (47%) fish. Of these, infection was in the spleen (42 fish), kidney (4 fish), or both organs (10 fish). Protista were in well-defined multicellular aggregates (Fig. 1A,B), and were sometimes heavily infiltrated with melanized macrophages. Some appeared banana shaped with an eosinophilic cytoplasm, while others appeared to have multiple nuclei. They stained weakly positive with Giemsa and were not associated with tissue necrosis. On electron microscopy, protista were intracytoplasmic within monocytes and appeared to displace the host nucleus (Fig. 1C). The organisms were sausage-shaped, and sometimes were grouped within a membrane. Individual organisms had a distinct nucleus, conoid, rhoptries, amylopectin granules and micronemes (Fig. 1D). No hemoparasites were seen on smears of peripheral blood. Bacteria compatible in morphology with the epitheliocystis-like organism were seen in 26 (22%) fish. Of these, infection was in the kidney (23 fish), spleen (2 fish), or both organs (1 fish). On light microscopy, the epitheliocystis-like organism consisted of well-defined spherical aggregates of basophilic organisms that stained negative with Gram stain and positive with Gimenez and Giemsa. In putative early infections, organisms were amorphous and homogenous and were accompanied by a mild mononuclear response or no inflammation (Fig. 2A). In later infections, particularly in the kidney, epitheliocystis-like organisms became infiltrated with clumps of eosinophilic material and were surrounded by a prominent capsule of collagen mixed with fibroblasts (Fig. 2B). On electron microscopy, epitheliocystis-like organisms had a thick capsule (Fig. 2C) surrounding a granular matrix containing mitochondria in various stages of degeneration (Fig. 2D). Deeper into the organism, a thin inner membrane (Fig. 2E) surrounded variably sized aggregates of membrane-bound spherical structures, ranging in diameter from 1 to 2 µm. These structures contained
Work et al.: Pathology of bluestripe snapper in Hawaii 61 Fig. 1. Lutjanus kasmira. (A) Coccidia in spleen. Note lack of inflammatory response. Scale bar = 50 µm. (B) Close-up of coccidia in (A) (arrows). Scale bar = 20 µm. (C) Coccidia within macrophage (arrow). Note displacement of host nucleus (n). Scale bar = 1µm. (D) Coccidia (probable merozoite). Scale bar = 1 µm. n = nucleus, a = amylopectin granules, r = rhoptries, c = conoid, m = microneme, g = dense granule
Page 1: DISEASES OF AQUATIC ORGANISMS Vol.
Page 5 and 6: Work et al.: Pathology of bluestrip
Page 7 and 8: Work et al.: Pathology of bluestrip

Protozoal and epitheliocystis-like infections in the introduced

You also want an ePaper? Increase the reach of your titles

Delete template?

Save as template?