Abstract
From December 2002 through January 2003 a large-scale seabird mortality was recorded off the shores of the Falkland (Malvinas) Islands, in the southwestern Atlantic Ocean. Affected species were gentoo (Pygoscelis papua), rockhopper (Eudyptes chrysocome), and Magellanic penguins (Spheniscus magellanicus), as well as albatrosses, petrels, and prions (Procellariidae). Best estimates suggest that 100,000–200,000 seabirds may have died during this event. Clinical signs in affected birds led us to investigate for the presence of toxins. Results showed the presence of neosaxitoxin (NeoSTX) and gonyautoxin 4 (GTX4), which are components of the paralytic shellfish poisons. Histopathology of affected birds showed minor microscopic changes. Infectious disease serology on albatross and gentoos (affected and apparently healthy animals), included paramyxovirus (Newcastle’s, type 2, and type 3), avian adenovirus, avian reovirus, avian influenza virus, avian laryngotracheitis virus, infectious bronchitis virus, infectious bursal disease, avian encephalomyelitis, Salmonella pullorum, Chlamydophila psittaci, and Marek’s disease. Results were negative for all of the above diseases except avian adenovirus. This is the first report of seabird mass mortality in the Falklands to affect such a wide range of species simultaneously and the first report of PSP affecting/killing seabirds in the South Atlantic. Because of the increasing occurrence of toxic algal blooms in this area, we encourage the establishment of a regular monitoring system and response team to investigate wildlife mortalities as they are reported. An organized, joint multiagency effort, with governmental support is key for successful investigation and preventive action.
Introduction
From December 2002 through January 2003 a large-scale seabird mortality was recorded off the shores of the Falkland (Malvinas) Islands, in the southwestern Atlantic Ocean (51 45 S, 59 00 W). Affected animals were mostly adult penguins, including gentoo (Pygoscelis papua), rockhopper (Eudyptes chrysocome) and Magellanic penguins (Spheniscus magellanicus).
However, several other species were reported ill and/or dying, such as albatrosses, petrels, and prions (Procellariidae). Mortalities occurred at several seabird colonies located on different islands within the archipelago, mostly in islands located to the northwest and south. In some areas, breeding pairs were down to 10% of the usual size of the specific colony and in others, entire breeding sites were abandoned even though eggs had been laid.
Methods
In order to evaluate possible causes of death, an array of samples was collected from healthy, clinically ill, and dead seabirds. Blood samples were collected from clinically “normal” black-browed albatross (Diomedea melanophris) (n=26), rockhopper (n=55), Magellanic (n=11), and gentoo penguins (n=68) and from 9 gentoo penguins in bad physical condition or showing severe clinical signs. After blood collection, four of the severely affected gentoo penguins were euthanatized and complete necropsies were performed. Additionally, tissue samples were collected from freshly dead animals that were found on the beach, four Magellanic and six gentoo penguins. Blood samples were centrifuged, and harvested plasma was stored in liquid nitrogen. Tissue samples were collected for histopathology and stored in 10% buffered formalin. A duplicate set of tissues (liver, spleen, kidneys, GI tract, and brain) and gastrointestinal contents when present, were frozen in liquid nitrogen.
Analyses for biotoxins were conducted at the Instituto de Fomento Pesquero (IFOP, Punta Arenas, Chile). Samples were analyzed by high-performance liquid chromatography (HPLC) on a Shimadzu spectrofluorometric analyzer in search of red tide toxins from the paralytic shellfish poisoning group (PSP). Histopathology and bacteriology analyses were performed at the National Veterinary Institute in Uppsala, Sweden.
Results and Discussion
Results showed the presence of neosaxitoxin (NeoSTX) and gonyautoxin 4 (GTX4), which are components of the PSPs. Five of 11 individual penguins submitted tested positive (45%). Of these positive individuals, one or more tissue samples had high levels of toxins (levels above those considered unsafe for humans: 80 µg STXeq 100-1 g shellfish meat). Additionally, low or trace levels of other PSP toxins were detected, particularly GTX1, GTX2, GTX3, and STX. Tissues from which high toxin levels were recovered were intestine or intestinal content (80% of tested samples were positive), followed by liver (45%), and stomach content or stomach wall (33%). No toxins of this group were found in fat tissue or aqueous humor of the eye.
Positive animals included two gentoo penguins collected after death (livers positive to NeoSTX). Both of these animals and a third gentoo also had low levels of STX and GTX2 and GTX3. On the other hand, three of the four ill gentoos euthanatized were positive to NeoSTX and GTX4, and had trace levels of STX, GTX1, GTX2, and GTX3. In these animals, tissues with detectable toxins were intestinal content, liver, kidney, and stomach content in two of them, and liver, brain, spleen, intestine, and intestinal content in the other. Assuming that dead birds had died from toxin ingestion, samples collected from euthanatized or dying animals proved to be better for toxin isolation, compared to those from animals found dead, even if they were fresh.
Histopathology of birds found ill (n=4) or dead (n=6, 5 gentoos and 1 Magellanic) did not show microscopic changes indicative of acute intoxication. For those birds which were euthanatized, there were no gross necropsy findings of significance, most were in good body condition and no parasites were found in their gastrointestinal tract. On histology, most birds showed older inflammatory reactions in bile ducts with periportal infiltrates of eosinophils and heterophils and some other leukocytes. Areas of fibrosis were found in one bird. Hemosiderosis was present in the liver of seven penguins, but no hepatosplenomegaly was observed. Liver cells and parenchyma appeared normal with no signs of necrosis or reaction. No inclusion bodies (neither IC nor ICP) were observed. Older inflammatory reactions were found in intestines with degeneration of crypts and epithelium, and infiltration of WBC. No lesions were present in the brains, meninges, hearts, kidneys, or lungs (except for congestion in the lungs). Spleens were found to be active, some with hyperplasia, but no necrosis. No signs of adenovirus (inclusion bodies) or plasmodium infection were detected. Bacteriology was performed on lung, kidney, brain, muscle, and heart tissue of one Magellanic and one gentoo penguin with negative results.
Paralytic shellfish poisoning (PSP) is caused by several dinoflagellates including Alexandrium spp., Gymnodinium catenatum, Pyrodinium bahamense that produce toxins known as saxitoxins and derivatives such as neosaxitoxin and gonyautoxins I–IV. PSP results from ingestion of shellfish, crustaceans, fish, and other organisms that have accumulated those potent toxins as a result of consuming dinoflagellates.9 PSP manifestations in humans are acute paresthesias and other neurologic manifestations which may progress rapidly to respiratory distress, muscular paralysis, and death.13 It is believed that birds are more susceptible to PSPs than other warm-blooded animals. For seabirds, symptoms such as loss of equilibrium, incoordination, convulsions, paralysis, vomiting, abnormal green-brown feces, and congestion of organs including lung have been previously described.13
Organisms of genus Alexandrium have been reported in Argentina (A. tamarense)2,8,9 and Chile (A. catenella)5. Geographically, positive animals in our investigation were collected at different locations in the archipelago which lies on the Argentina continental shelf. Gentoo penguins are opportunistic feeders, and around the Falklands are known to take fish (56% of diet, mainly from the cod family Patagonotothen sp., but also Micromesistius australis and other species), crustaceans (36%, Munida gregaria), and squid (11% Loligo gahi, Gonatus antarcticus, Moroteuthis ingens).3,10 PSP toxins have been detected in sardines (plankton feeders) but also in higher trophic level feeders such as mackerel (Scomber japonicus)1,8 and anchovy (Engraulis anchoita)9 in the South Atlantic. Montoya et al. (1997) report a mackerel mortality event off the shores of Buenos Aires province in 1993, probably associated with penguin and other seabird deaths. It has been suggested that herbivorous zooplankton is the main biologic PSP vector for these fish,8,9 and that mackerel could have the ability of accumulating toxins in their liver. However, no information on toxin detections on the above mentioned gentoo prey items has been reported.
This is the first report of seabird mass mortality in the Falklands (Malvinas) to affect such a wide range of species simultaneously. Producing a realistic figure for the number of birds that died as a result of this event is very hard to do at the islands’ scale. In the few colonies of gentoo penguins that were again monitored in November 2003, after the event, numbers were down to one third, while colonies not affected in the north east of the islands were either stable or increasing. This leads to a rough estimate of a minimum of 30,000–50,000 gentoo penguins missing. Estimating the number of rockhopper penguins is more complicated as fewer colonies that were affected were monitored the following season. Some affected colonies had their numbers drop by one third only, while the main colony of Steeple Jason, lost 60,000 pairs out of 89,000 counted in the 2000 census. Again, a rough estimate of birds actually dying from poisoning, island wide, might be between 30,000–70,000 for rockhopper penguins. Census of albatross after the event was only conducted on Steeple Jason where numbers dropped from 158,000 in 2000 to 112,000 in November 2003. Albatrosses also suffer from mortality associated with fishing activities, so not all the 45,000 missing pairs might be due to intoxication and a reasonable estimate would be between 10,000 and 30,000 albatrosses may have died from poisoning. Due to lack of effective census, no numbers can be produced for Magellanic penguins and burrowing petrels, but considering the number of reports of affected birds, numbers could be in the tens of thousands (Huin, unpublished data). Even though seabird numbers suggest that up to date these events have had a very low frequency of occurrence, it seems like they have been getting more important in the last decade, reflecting global features of harmful algal blooms (HABs), exhibiting a higher frequency, a higher geographic cover, and a higher intensity.
In addition, serology for infectious diseases was performed on the euthanatized penguins and on a sample of apparently healthy animals at various locations with evidence of mortalities. Serology included paramyxovirus (Newcastle’s, type 2, and type 3), avian adenovirus, avian reovirus, avian influenza virus, avian laryngotracheitis virus, infectious bronchitis virus, infectious bursal disease, avian encephalomyelitis, Salmonella pullorum, Chlamydophila psittaci, and Marek’s disease. The albatross and gentoos were antibody negative for all of the above diseases except avian adenovirus. Results for Magellanic and rockhoppers are still pending. For the gentoos tested, of the ones showing neurologic symptoms 7 of 9 tested positive (78%), and of the ones that appeared clinically normal 15 of 27 tested positive (55%).
There are no published references for adenovirus causing disease in penguins or seabirds. Nonetheless, avian adenoviruses are distributed worldwide, and many avian species are known to be susceptible. This agent causes disease in Galliformes and has also been reported as a possible pathogen in pigeons, raptors, Psittaciformes, and waterfowl.4 Positive antibody titers have been found by Karesh et al. (1999) in rockhopper penguins, in southern giant petrels (Macronectes giganteus),15 and in other seabirds such as Magellanic penguins, imperial cormorants (Phalacrocorax albiventer) and rock shags (P. magellanicus) from Argentina (Uhart et al., unpublished data). However, the test used for avian adenovirus antibody detection presents moderate specificity and has not been validated for these species, and could result in false positives.6,12
An unexpected result was the relative absence of evidence of exposure to the other infectious diseases in the birds sampled. We have found positive titers to many of these in seabirds in Argentina and Peru using the exact same sampling and test methods7 (Karesh, Uhart unpublished data). A possible explanation would be that other migratory birds do not come in contact with the albatross or the gentoo colonies which, due to their relative isolation, have not been exposed to infectious diseases that are endemic in populations elsewhere. The lack of positive antibody titers also suggests that these birds may suffer heavily if or when these diseases are introduced to the colonies. Good sanitation practices to prevent contamination or cross-contamination among colonies and islands would be prudent to prevent the spread of infectious agents introduced by tourists/visitors, researchers, landowners, or officials.
This is the first report of PSP affecting/killing seabirds in the South Atlantic. Though it is possible that events such as the one reported here have occurred before8,11,14 in this area, it has not been possible to identify specific toxins in tissues of affected/dead animals. The spatial and temporal distribution of toxic dinoflagellate blooms seems to have expanded over the last 2 decades in the Argentinian Sea.2 Therefore, we encourage the establishment of a regular monitoring system of plankton in coastal waters as well as an immediate response team to investigate wildlife mortalities as they are reported. The experience gained has shown that there must be an organized joint multiagency effort with governmental support for successful investigation and preventive action.
Literature Cited
1. Carreto, J.I., R. Akselman, M.O. Carignan, A.D. Cucchi Colleoni and M. Pajaro. 1993. Presencia de veneno paralizante de moluscos en higado de caballa de la region costera bonaerense. INIDEP Doc Cient. 2:53–59.
2. Carreto, J.I., N. Montoya, A.D. Cucchi Colleoni and R. Akselman. 1998. Alexandrium tamarense blooms and shellfish toxicity in the Argentine Sea: a retrospective view. In: Harmful Algae (B. Reguera, J. Blanco, M.L. Fernández and T. Wyatt, eds.), (Xunta de Galicia and IOC-UNESCO), 131–134.
3. Clausen, AP and K. Pütz. 2002. Recent trends in diet composition and productivity of gentoo, Magellanic and rockhopper penguins in the Falkland Islands. Aquatic Conservation: marine and freshwater ecosystems. 12:51–61.
4. Gerlach, H. 1994. Viruses. In: Ritchie, B.W., G.J. Harrison, and L.R. Harrison (eds.). Avian Medicine: Principles and Application. Wingers Publishing. Lake Worth, Florida. 862–948.
5. Guzmán, L.H. Pacheco, G. Pizarro and C. Alarcón. 2002. Alexandrium catenella y Veneno Paralizante de los Mariscos en Chile. (M.E. Sar, M. Ferrario y B. Reguera, eds.). Monografías del Instituto Español de Oceanografía. 235–256.
6. Hietala, S.K. and I.A. Gardner. 1999. Validity of using diagnostic tests that are approved for use in domestic animals for nondomestic species. In: Fowler and Miller (eds.). Zoo and Wild animal medicine. Current Therapy, 4. WB Saunders, Philadelphia, Pennsylvania. 55–58.
7. Karesh, W.B., M.M. Uhart, E. Frere, P. Gandini, W.E. Braselton, H. Puche and R.A. Cook. 1999. Health evaluation of free-ranging rockhopper penguins (Eudyptes chrysocome) in Argentina. Journal of Zoo and Wildlife Medicine. 30(1):25–31.
8. Montoya, N.G., R. Akselman, M. Pajaro, R.G. Perrota and J.I. Carreto. 1997. Mortandad de caballa (Scomber japonicus) en la plataforma bonaerense (Mar Argentino) asociada a un florecimiento del dinoflagelado toxico Alexandrium tamarense. Revista de Investigacion y Desarrollo Pesquero. 11:145–152.
9. Montoya, N.G., M.I. Reyero, R. Akselman, J.M. Franco and J.I. Carreto. 1998. Paralytic shellfish toxins in the anchovy Engraulis anchoita from the Argentinian coast. Xunta de Galicia and Intergovernmental Oceanographic Commission of UNESCO. 72–73.
10. Pütz, K., R.J Inghma, J.G. Smith and J.P. Croxall. 2001. Population trends, breeding success and diet composition of gentoo (Pygoscelis papua), Magellanic (Spheniscus magellanicus) and rockhopper (Eudyptes chrysocome) penguins in the Falkland Islands. A review. Polar Biology. 24:793–807.
11. Quintana, F., M.M. Uhart and P. Garcia Borboroglu. 2001. Causa de mortandad masiva de aves marinas en las costas de Chubut. Unpublished report, 18 January 2001.
12. Ritchie, B.W., K.S. Latimer, C.R. Gregory, D. Pesti, V. Burnley and P.D. Lukert. 2001. Diagnostic testing for infectious diseases. In: Proceedings from Annual Conference of the Association of Avian Veterinarians. 359–367.
13. Shunway S.E., S.M. Allen and P.D. Boersma. 2003. Marine birds and harmful algal blooms: sporadic victims or under-reported events? Harmful algae. 2:1–17.
14. Uhart, M., F. Quintana, E. Frere, P. Gandini, W. Karesh and R. Cook. 2002. Patagonia seabirds as indicators of marine ecosystem health. In: Proceedings from 51st Annual Wildlife Disease Association Conference. 39.
15. Uhart, M., F. Quintana, W. Karesh and E. Braselton. 2003. Baseline hematology, biochemistries and disease serology of the southern giant petrel in Patagonia, Argentina. Journal of Wildlife Diseases. 39(2):359–365.