Survey
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
Program Agenda First Annual CL Davis Symposium on Diagnostic Pathology of Diseases of Aerial, Terrestrial and Aquatic Wildlife April 9-11, 2008 Covance Laboratory Learning Center, Madison, WI Ocean Health & Disease: Coral Reefs & Sea Turtles Thierry M. Work USGS-National Wildlife Health Center-Honolulu Field Station PO Box 50167, Honolulu, HI 96850 Tel: 808 792-9520; Fax: 808 792-9596; [email protected] Web: www.nwhc.usgs.gov/hfs/Homepage.htm Slide Topic 1 Title 2 Outline 3 4 Photo of turtle Life cycle 5 Photo of boat strike, fishing line 6 FP photo 7 Map Comments Investigation of disease in tropical marine ecosystems poses particular challenges. You can only stay underwater for limited time (limited observation), when animals die, they get eaten (no diagnostic specimens), and if specimens are found, the heat will decompose them rapidly. Adding to the complexity is that we have limited knowledge about the basic physiologic and anatomic processes for many marine organisms, and elucidating etiology of diseases is often stymied by lack of laboratory reagents and inability to culture suspect agents in the laboratory. This session will cover some of what we know about major diseases of reef organisms including marine turtles, fish, and invertebrates. There are 7 species of sea turtles worldwide. One challenge of working with sea turtles is that a portion of life cycle remains a mystery. Where do hatchlings go when they go out to sea? Obvious causes of mortality in turtles include boat strikes characterized by linear parallel lesions on carapace and fishing line entanglement and ingestion, which can cause imbrication of intestines and ulceration of intestinal mucosa. By far the most important disease of sea turtles is fibropapillomatosis (FP) manifested as sessile to pedunculated smooth to rugose growths arising from the skin FP was first documented in Florida in the 1930s but since has been found wherever marine turtles are found (tropical oceans). The disease has been documented mainly in green, loggerhead, and olive ridley turtles. 8 FP photo 9 Oral tumors 10 Internal tumors: heart, bone, lung 11 Internal tumors: kidney, heart 12 Locations 13 Graph 14 Histogram 15 Pie chart 16 Histo Skin 17 Lung tumor 18 Kidney tumor 19 Tumor intestines/bone 20 Heart tumor 21 Pie chart/graph Tumors can reach very large sizes and impede movement of limbs and cause loss of vision. FP has been most intensively studied in Florida and Hawaii. Prevalence in Florida continues to remain stable at ~20-50% whereas in Hawaii, the disease peaked at ~60% but has been steadily declining for the past 5 years for unknown reasons Interestingly, oral tumors are a characteristic of FP in Hawaii that is not found in Florida. Ca. 50% of turtles with oral tumors also have secondary pulmonary infections. Turtles with FP also have internal tumors in various organs characterized by firm white to grey wellcircumscribed masses. In some cases, these masses are filled with clear fluid, and in other cases, larger tumors can have cores of friable debris. In Hawaii, tumors are most often seen on the eyes followed by the mouth, flippers, and seams and scutes. There also seems to be a predilection for FP to occur more in the front than the rear of animals. In most cases, for internal tumors, only one organ is affected For both HI and Florida, the most commonly affected organs for internal tumors include the lungs, heart, and kidney. Interestingly, in contrast to FL, liver tumors in HI are rare. Skin tumors are characterized by acanthotic orthokeratotic epidermis with prominent rete pegs overlying a collagen matrix with numerous pleomorphic plump hapahazardly arranged fibroblasts. In some cases, keratin cysts are present within connective tissue matrix, Internal tumors are variations of connective tissue tumors. In some cases like the lung, tumors are well circumscribed. In other cases, tumors infiltrate adjoining tissues causing atrophy (e.g. note atrophied renal tubules). In other cases, tumors can be invasive into bone causing reabsorption. Depending on staining characteristics, tumors have been classified as fibromas, myxofibromas… …or in the case of heart, fibrosarcomas of low-grade malignancy. In general, tumors have appearance of slow growth with few mitotic figures. Severity of disease in turtles is graded from 0 to 3. Most stranded turtles are TS 2 and 3, and area of tumors 22 Map 23 Flukes in artery 24 Egg diagram 25 Chart 26 Chart of SCL 27 Possible causes of FP 28 Okadaic acid 29 Immunology 30 Flukes/viruses 31 Viruses 32 Tumor diagram 33 Vectors 34 Tree increases with age In Main HI islands, FP is found in all islands but rarely on west coast of Hawaii for unknown reasons. 100% of stranded turtles also have infections with vascular trematodes. Interestingly, turtles from pelagic stage have little to no infection suggesting these parasites are acquired once turtles recruit to near shore habitat. The life cycle is unknown, but it is suspected that it is similar to schistosomiasis with mollusk intermediate host. In HI, turtles are infected by 4 species of trematodes of which one is probably endemic (not found in other oceans). The endemic one can be differentiated from 4 other by egg morphology Eggs in tissue can be quantified in standardized manner to get an idea of severity of infection. Distribution of eggs burden in tissues suggests that endemic parasite makes a small component of trematode community in turtles. It also appears that trematodes have a negative effect on body condition of sea turtles independent of FP status Several potential causes of FP have been postulated including UV radiation, flukes, viruses, marine toxins, and immunosuppression. Experimentally, OA has caused tumors in mice. This toxin is produced by Prorocentrum lima, a dinoflagellate. Although turtles are exposed to OA, the toxin can be found in both tumored and non-tumored turtles. Immunology studies on turtles with FP suggests that those with severe disease are immunosuppressed but that immunosuppression is not necessarily a prequel to disease. Experimental trials in Florida using captive green turtles revealed that FP could be reproduced using cell free filterable agents. This along with histopathology showing intranuclear inclusions in some tumors suggested a viral cause Subsequent work in New York and Florida revealed that a herpes virus is associated with tumors. Incidentally, turtles also are infected with an endogenous retrovirus. Using PCR, herpes viral DNA can be found in tumors but not non-tumored tissue. There also is a gradient with more viral DNA found in superficial tumors Viral DNA has been found in a variety of epibionts associated with turtles. Phylogeny places the turtle herpes virus in the 35 Diagram 36 37 Questions? Brick scale soldierfish 38 39 40 41 42 43 44 45 46 alphaherpes category Unfortunately, because the virus cannot be cultured, Koch’s postulates remain to be fulfilled. While quite a bit of information exists on diseases of aquarium fish, there is very little data on diseases of wild coral reef fish. Existing literature reviews (Panek 2005) focus mainly on the Western Atlantic, and major causes of mortality in reef fish there include blooms of toxic algae, bacterial infections, and, more rarely, tumors. Butterfly fish and map In contrast to the Atlantic, tumors in reef fish appear relatively more common in the Pacific ocean. In Hawaii, Okihiro (1998) documented chromatophoromas in two species of butterfly fish (multiband and milletseed). These tumors were found in fish from Southwest Maui and Western Lanai Depth gradient During extensive surveys in the early 1980s, Okihiro found that prevalence of tumors ranged from 5-25% with decreasing prevalence of tumors as depth increased and increasing prevalence of tumors over time Fish in wild with A recent follow up study done in 2005 revealed that tumors tumors continue to exist in multiband butterfly fish. Dead multiband In multiband butterflyfish, these tumors are characterized butterflyfish with by raised sessile grey to iridescent nodular firm masses tumors with a homogenous texture on cut surface sometimes associated with hemorrhage on margins. Tumors appear most often distributed laterally. Live milletseed with In milletseed butterflyfish, tumors are sessile, raised tumors nodular dark masses usually appearing to initially arise from the first dorsal spine. Dead milletseed with Larger tumors are often ulcerated. tumors Histopath of On histology, tumors are characterized by mixed milletseed tumor populations of chromatophores round and spindle cells mixed with a connective tissue matrix effacing epidermal architecture and infiltrating underlying skeletal muscle with occasional nidi of necrosis. Recent surveys in 2005 at the same sites surveyed by Okihiro revealed a much smaller prevalence of tumors (<5%). Kole live Another skin lesion that has been seen in reef fish in the Pacific are pigment anomalies in goldring surgeonfish. These are typically distributed laterally at the base of the tail but more severe cases can encompass the entire body. Kole dead Fish with this pigment anomaly have been found on Maui, Oahu, and the northwestern Hawaiian islands. To date, no efforts have been made to quantify prevalence, 47 Kole histo 48 Oahu Map 49 Fish photos 50 Pie charts 51 Micrograph 52 Histogram 53 Diagram of island 54 Graph 55 Goatfish photos 56 Multiband liver, gill, heart however, it appears to be fairly low. Early lesions manifest as hyperplasia of epidermis. Advanced lesions consists of masses of round to spindle shaped cells mixed with chromatophores effacing skin architecture and infiltrating underlying skeletal muscle. Etiology for both types of tumors (butterflyfish, surgeonfish) is unknown. Monitoring sewer outfalls for liver tumors in fish is required by EPA for Hawaii. This has presented an opportunity do survey wild fish for disease agents (in addition to looking at tumors). Each year, three species of fish are collected by City and County of Honolulu (blue lined snapper, brick scale soldierfish, big eyed scad). Some of these fish are necropsied to detect presence of liver tumors. Surveys of fish revealed that prevalence of histozoic parasites in blue lined snappers was considerably higher than for other two species of fish. This can probably be partly explainable by life history patterns (more parasites in bottom feeders). The two most commonly encountered parasites in snappers were a coccidian and an epitheliocystis-like organism (ELO) in the spleen and kidney. Molecular work on coccidium at OSU reveals it to be Goussia sp. ELO stains positive with Giemsa and Gimenez. Prevalence of infection with coccidium increases with age of fish, however, prevalence of ELO decreases with increasing age of fish. Fish seem to mount little to no inflammatory response to coccidia, however, a chronic inflammatory response is seen with ELO infections. Tropical islands are prone to suffer from invasive species (Plants and animals) because a lot of the native biota lost adaptive life history traits to defend against predators. For microorganisms in HI, this has been exemplified by severe impact of imported avian malaria on native forest birds. Blue lined snappers were introduced from Marquesas to HI in 1950s, and this brought up question as to whether they were sharing parasites with native fish with which they school. Surveys over time indicate that prevalence of Goussia and ELO in snappers remains fairly constant Question then arose about possibility that snappers, which school with native goatfish, may be sharing parasites with these species. Turns out that, as in many other wild fish, multiband goatfish are infected at low levels with coccidia in liver, 57 Multiband liver, spleen, caudal kidney 58 Orange goatfish spleen, gill, heart 59 Orange goatfish spleen, gill 60 Yellow tail goatfish spleen and muscle Yellow stripe goatfish spleen, muscle, liver 61 62 Graph summary 63 Coral reef scene 64 Cnidarian diagram 65 Anatomy 66 Black Band monogean trematodes in gills, intermediate stage of parasites in heart and epitheliocystis-like organisms in gills. However, multiband goatfish also are infected, albeit at low prevalence, with ELO and coccidia in spleen and kidney. As in multiband, orange goatfish also have infections with trematodes (eggs, in various organs) and also have occasional infections with ciliates in gills (trichodinid ciliates). However, >90% of orange goatfish infected with coccidia in spleen. Also see occasional monogean trematodes in gills. Ca. 30% of yellow tail goatfish have coccidia in spleen. Occasional microsporidia are in skeletal muscle. Ca. 50% of yellow stripe goatfish have coccidia in spleen with occasional microsporidia in muscle and coccidia in liver as well. So, it appears that at least for coccidia, native goatfish share parasite that is morphologically similar to that of snappers. The big question remains as to whether these parasites have detectable effects on fish health Corals are and indispensable component of reefs providing nurseries for a variety of fish and invertebrates and laying the foundation for the diversity of organisms seen in tropical marine ecosystems. Lose your corals, you lose your ecosystem. Corals belong to the cnidaria (animals with stinging cells). Corals are colonial sessile organisms that, along with algae, make up a majority of the biomass of reefs. They are unique in that they live with symbiotic algae (zooxanthellae). Anatomically, they are simple organisms with polyps consisting of tentacles surrounding a mouth and a digestive system (gastrovascular canal) that is connected between polyps. At the cellular level, corals consist of three cell layers: the calicoblastic epithelium that secretes the skeleton, the gastrodermis (cells that digest and also harbor zooxanthellae), the mesoglea (a connective tissue matrix), and the epidermis (containing supporting cells and stinging cells or nematocysts). Coral disease have had the most severe impact in the Western Atlantic causing massive loss of corals. Of these, black band disease and vibrio associated bleaching are the best characterized. The typical lesion of black 67 Map of black band 68 Bleaching 69 Bleaching diagram 70 Vibrio associated bleaching Vibrio bleaching Diagram 71 72 Aspergillosis 73 Aspergillus diagram 74 Aspergillus map 75 Dust storm 76 Porites Trematodes 77 Putative life cycle 78 White syndrome 79 White syndrome diagram band is circular loss of tissues revealing intact white skeleton separated from intact tissue by a black band. This black band consists of a consortium of cyanobacteria and vibrios. In the field, the presence of disease appears to progress more rapidly with increasing water temperatures. Black band was first documented in Central American and has been found throughout the Caribbean and the Pacific. Bleaching is the loss of pigmentation of corals revealing the underlying white skeleton The causes of bleaching are due to exit of zooxanthellae from the gastrodermis leading to loss of tissue pigments. Can be associated with temperature In Israel, researchers have characterized a form of bleaching that is due to infection with Vibrio In this scenario, the vibrio infects the coral and enters a viable but non-culturable state. The bacterium produces a toxins that kills zooxanthellae and induces bleaching. In the 1980s, researchers noted sea fans with lesions associated with fungal hyphae. Subsequent investigations led to discovery of Aspergillus sydowii as a cause of these lesions Field research shows that prevalence of seafan lesions increase with size of sea fan and with increasing depth suggesting that water motion may play a role Seafan aspergillosis was initially found in central America and Venezuela but has since been documented in multiple islands throughout Caribbean Research by USGS suggests that dust blown from Africa may help spread coral disease (Aspergillus spore have been found in the dust). In the Pacific, certain species of coral are infected with the intermediate stage of a fish trematode. These lesions are characterized by small pink to tan raised nodules Research in the Pacific has shown that butterfly fish feed on these lesions preferentially and serve as the definitive host for the trematode. How corals get infected remains a mystery. This is a catch all term for acute tissue loss in corals revealing white skeleton. White syndrome are often lethal to coral colonies and are deemed responsible for coral reef degradation in W. Atlantic. The etiology of most of these syndromes is unknown. In the Caribbean, WS has resulted in major shifts in reef structure with extermination of the dominant coral, 80 White syndrome map 81 Diagram of disease investigation 82 Pie charts 83 Map 84 85 Photodocument Pathology diagram 86 87 88 89 Discoloration photo GA photo TL photo Discoloration Histo 90 Tissue loss histo 91 GA Histo 92 93 Pie charts Known causes 94 Mixed causes 95 White syndrome Johnston Atoll 96 WS Johnston Atoll 97 Time line slide 98 Acknowledgements Acropora palmate and its replacement with encrusting corals and algae WS was first documented in Caribbean and has since been found throughout the Western Atlantic. One of the problems with coral disease research in corals has been a lack of standardization of nomenclature of lesions which has led to a lot of confusion When you look at coral disease literature, not much is produced regarding morphology, yet morphology is a critical part of case definitions of disease Accordingly, in the pacific, emphasis has been on developing standardized nomenclature of gross lesions in corals and follow up with histopath Lesions in corals are broadly characterized as tissue loss, skeletal growth anomalies, and discoloration As expected, histology of each of these lesions reveal different manifestations. For example, bleached corals manifest loss of zooxanthellae Tissue loss can be due to infection with ciliates, fungi, algae or can be uncomplicated Growth anomalies manifest as proliferation of polyp basal body wall. So far, no evidence of true neoplasia in classical sense. Summary of gross vs histo findings in corals In some cases, cause of lesion is evident such as barnacles, predation, or mucus sheathing In other cases, gross lesions induced by predators (starfish) can be similar to those of unknown etiology (white syndromes). Also a challenge is that determining prevalence of colonies with lesions can be challenging, particularly for certain morphologies of corals. What we do know is that coral diseases can kill colonies outright. Example photos of a reef taken in 2001 and 2006 reveal >90% loss of live coral. Coral disease is way behind other animals in level of knowledge. Lots to learn about in the future. References: Herbst LH (1994) Fibropapillomatosis of marine turtles. Annual Review of Fish Diseases 4: 389-425 Okihiro MS (1988) Chromatophoromas in two species of Hawaiian butterflyfish, Chaetodon multicinctus and C. miliaris. Veterinary Pathology 25: 422-431 Panek F (2005) Epizootics and disease of coral reef fish in the tropical Western Atlantic and gulf of Mexico. Reviews in Fisheries Science 13: 1-21 Work T, Aeby G (2006) Systematically describing gross lesions in corals. Diseases of Aquatic Organisms 70: 155-160