Pathological and molecular characterization of avian malaria in captive Magellanic penguins (Spheniscus magellanicus) in South America.

Avian malaria is a mosquito-borne disease that affects multiple avian species and is caused by protozoans of the genus Plasmodium. An avian malaria infection caused by Plasmodium sp. in Magellanic penguins (Spheniscus magellanicus) with high mortality is described in a zoo in Southern Brazil. Clinically, three birds presented signs of inappetence, anorexia, pale mucosa, dyspnea, and opisthotonus, with death in a clinical course of 5-8 h. At the necropsy, all birds exhibited pale mucosa, marked splenomegaly and hepatomegaly, in addition to moderate leptomeningeal blood vessels ingurgitation in the brain. Microscopically, multiple exoerythrocytic meronts were observed in the cytoplasm of endothelial cells in the spleen, liver, heart, lungs, brain, kidneys, and pancreas. The spleen had a multifocal perivascular inflammatory infiltrate of lymphocytes, plasma cells, and macrophages, which also exhibited hemosiderosis and erythrophagocytosis. The liver had a multifocal periportal inflammatory infiltrate of lymphocytes, macrophages, and plasma cells, in addition to marked hemosiderosis in the hepatic sinusoids. Fragments of spleen, liver, brain, skeletal muscle, and lung were tested by the polymerase chain reaction technique for the detection of a fragment of the cytochrome B gene from haemosporidians, which resulted positive for Plasmodium spp. After sequencing, the samples were phylogenetically associated to Plasmodium sp. detected in Turdus albicollis (KU562808) in Brazil and matched to the lineage TURALB01 previously detected in T. albicollis. Avian malaria infections caused by Plasmodium sp. of lineage TURALB01 may occur in S. magellanicus with high mortality, and, thus, it is essential to detect and characterize the agent involved to obtain the differential diagnosis of the condition.

Different paths - the same virulence: experimental study on avian single and co-infections with Plasmodium relictum and Plasmodium elongatum.

Co-infections are prevalent worldwide, however, we are still struggling to understand interactions between different parasites and their impacts on host fitness. In the present experimental study we analysed the infection dynamics of two avian malarial parasites Plasmodium elongatum (genetic lineage pERIRUB01) and Plasmodium relictum (genetic lineage pSGS1) and their impacts on host health during single and co-infections. We reveal that P. elongatum intensity of parasitemia is enhanced by the presence of P. relictum during co-infection, while the parasitemia of P. relictum stays the same. This illustrates how development of a parasite (P. elongatum) which infects both mature and young (polychromatic) red blood cells (RBCs) is facilitated during co-infection with a parasite which specialises in adult RBCs only (P. relictum). The virulence of co-infections was similar to that of the more virulent parasite (P. elongatum). However, the profile of infection and the mechanisms that caused mortality were different. Birds infected with P. elongatum only start to die due to non-regenerative anaemia, when intensity of parasitemia is light and the number of polychromatic RBCs decrease dramatically. Meanwhile, co-infected birds start to die when the mean intensity of parasitemia reaches 10% and the number of polychromatic RBCs increases abnormally, reflecting regenerative anaemia. Our findings reveal that typically measured parameters of virulence (e.g., mortality rate, level of hematocrit) can be the same during single and co-infections, but the mechanisms responsible for the observed virulence can be different. This information serves a better understanding of the processes underpinning the interactions of co-infected parasite species.

Malaria parasites and related haemosporidians cause mortality in cranes: a study on the parasites diversity, prevalence and distribution in Beijing Zoo.

BACKGROUND: Malaria parasites and related haemosporidian parasites are widespread and may cause severe diseases in birds. These pathogens should be considered in projects aiming breeding of birds for purposes of sustained ex situ conservation. Cranes are the 'flagship species' for health assessment of wetland ecosystems, and the majority of species are endangered. Malaria parasites and other haemosporidians have been reported in cranes, but the host-parasite relationships remain insufficiently understood. Morbidity of cranes due to malaria has been reported in Beijing Zoo. This study report prevalence, diversity and distribution of malaria parasites and related haemosporidians in cranes in Beijing Zoo and suggest simple measures to protect vulnerable individuals. METHODS: In all, 123 cranes (62 adults and 61 juveniles) belonging to 10 species were examined using PCR-based testing and microscopic examination of blood samples collected in 2007-2014. All birds were maintained in open-air aviaries, except for 19 chicks that were raised in a greenhouse with the aim to protect them from bites of blood-sucking insects. Bayesian phylogenetic analysis was used to identify the closely related avian haemosporidian parasites. RESULTS: Species of Plasmodium (5 lineages), Haemoproteus (1) and Leucocytozoon (2) were reported. Malaria parasites predominated (83% of all reported infections). The overall prevalence of haemosporidians in juveniles was approximately seven-fold higher than in adults, indicating high susceptibility of chicks and local transmission. Juvenile and adult birds hosted different lineages of Plasmodium, indicating that chicks got infection from non-parent birds. Plasmodium relictum (pSGS1) was the most prevalent malaria parasite. Mortality was not reported in adults, but 53% of infected chicks died, with reports of co-infection with Plasmodium and Leucocytozoon species. All chicks maintained in the greenhouse were non-infected and survived. Species of Leucocytozoon were undetectable by commonly used PCR protocol, but readily visible in blood films. CONCLUSION: Crane chicks often die due to malaria and Leucocytozoon infections, which they likely gain from wild free-living birds in Beijing Zoo. Molecular diagnostics of crane Leucocytozoon parasites needs improvement. Because the reported infections are mainly chick diseases, the authors recommend maintaining of juvenile birds in vector-free facilities until the age of approximately 6 months before they are placed in open-air aviaries.

Cases of mortality in little penguins (Eudyptula minor) in New Zealand associated with avian malaria.

CASE HISTORY A little penguin (Eudyptula minor) of wild origin, in captivity at Wellington Zoo, became inappetent and lethargic in March 2013. Despite supportive care in the zoo's wildlife hospital, the bird died within 24 hours. CLINICAL FINDINGS Weight loss, dehydration, pale mucous membranes, weakness, increased respiratory effort and biliverdinuria were apparent on physical examination. Microscopic evaluation of blood smears revealed intra-erythrocytic stages of Plasmodium spp. and a regenerative reticulocytosis in the absence of anaemia. PATHOLOGICAL FINDINGS Post-mortem findings included reduced body condition, dehydration, pulmonary congestion and oedema, hepatomegaly, splenomegaly, hydropericardium and subcutaneous oedema. Histopathological findings included protozoal organisms in sections of lung, liver and spleen. A marked, diffuse, sub-acute interstitial histiocytic pneumonia was present. Accumulation of haemosiderin was noted in the Kupffer cells of the liver and in histiocytic-type cells in the spleen. MOLECULAR TESTING DNA was extracted from frozen portions of the liver. Nested PCR results and DNA sequencing confirmed infection of the deceased little penguin with Plasmodium (Huffia) elongatum lineage GRW06. DIAGNOSIS Avian malaria due to Plasmodium (Huffia) elongatum GRW06 RETROSPECTIVE INVESTIGATION A retrospective analysis of 294 little penguin cases in the Massey University post-mortem database revealed three other potential avian malaria cases. Analysis of archived tissues using a nested PCR for Plasmodium spp. followed by DNA sequencing revealed that a little penguin which died at Auckland Zoo was infected with P. elongatum GRW06 and two wild little penguins found dead on New Zealand beaches were infected with P. relictum SGS1 and Plasmodium. sp. lineage LINN1. Therefore, the overall frequency of deaths in little penguins associated with avian malaria was 4/295 (1.36%). CLINICAL RELEVANCE Our results suggest that avian malaria is associated with sporadic mortality in New Zealand's little penguins both in the wild and in captivity, but there is no evidence of mass mortality events due to Plasmodium spp. infection.

Diversity and distribution of avian malaria and related haemosporidian parasites in captive birds from a Brazilian megalopolis

The role of zoos in conservation programmes has increased significantly in last decades, and the health of captive animals is essential to guarantee success of such programmes. However, zoo birds suffer from parasitic infections, which often are caused by malaria parasites and related haemosporidians. Studies determining the occurrence and diversity of these parasites, aiming better understanding infection influence on fitness of captive birds, are limited. Methods: In 2011–2015, the prevalence and diversity of Plasmodium spp. and Haemoproteus spp. was examined in blood samples of 677 captive birds from the São Paulo Zoo, the largest zoo in Latin America. Molecular and microscopic diagnostic methods were used in parallel to detect and identify these infections. Results: The overall prevalence of haemosporidians was 12.6%. Parasites were mostly detected by the molecular diagnosis, indicating that many birds harbour subclinical or abortive infections. In this project, birds of 17 orders (almost half of all the orders currently accepted in taxonomy of birds), 29 families, and 122 species, were tested, detecting positive individuals in 27% of bird species. Birds from the Anatidae were the most prevalently infected (64.7% of all infected animals)...

Mortality and pathology in birds due to Plasmodium (Giovannolaia) homocircumflexum infection, with emphasis on the exoerythrocytic development of avian malaria parasites.

BACKGROUND: Species of avian malaria parasites (Plasmodium) are widespread, but their virulence has been insufficiently investigated, particularly in wild birds. During avian malaria, several cycles of tissue merogony occur, and many Plasmodium spp. produce secondary exoerythrocytic meronts (phanerozoites), which are induced by merozoites developing in erythrocytic meronts. Phanerozoites markedly damage organs, but remain insufficiently investigated in the majority of described Plasmodium spp. Avian malaria parasite Plasmodium (Giovannolaia) homocircumflexum (lineage pCOLL4) is virulent and produces phanerozoites in domestic canaries Serinus canaria, but its pathogenicity in wild birds remains unknown. The aim of this study was to investigate the pathology caused by this infection in species of common European birds. METHODS: One individual of Eurasian siskin Carduelis spinus, common crossbill Loxia curvirostra and common starling Sturnus vulgaris were exposed to P. homocircumflexum infection by intramuscular sub-inoculation of infected blood. The birds were maintained in captivity and parasitaemia was monitored until their death due to malaria. Brain, heart, lungs, liver, spleen, kidney, and a piece of breast muscle were examined using histology and chromogenic in situ hybridization (ISH) methods. RESULTS: All exposed birds developed malaria infection, survived the peak of parasitaemia, but suddenly died between 30 and 38 days post exposure when parasitaemia markedly decreased. Numerous phanerozoites were visible in histological sections of all organs and were particularly easily visualized after ISH processing. Blockage of brain capillaries with phanerozoites may have led to cerebral ischaemia, causing cerebral paralysis and is most likely the main reason of sudden death of all infected individuals. Inflammatory response was not visible around the brain, heart and muscle phanerozoites, and it was mild in parenchymal organs. The endothelial damage likely causes dysfunction and failure of parenchymal organs. CONCLUSION: Plasmodium homocircumflexum caused death of experimental passerine birds due to marked damage of organs by phanerozoites. Patterns of phanerozoites development and pathology were similar in all exposed birds. Mortality was reported when parasitaemia decreased or even turned into chronic stage, indicating that the light parasitaemia is not always indication of improved health during avian malaria. Application of traditional histological and ISH methods in parallel simplifies investigation of exoerythrocytic development and is recommended in avian malaria research.

Plasmodium spp. In a captive raptor collection of a Safaripark in northwest Italy.

Blood parasites infect all vertebrates (Clayton and Moore 1997). Avian malaria parasites (Plasmodium spp., Plasmodiidae) are cosmopolitan in their distribution and are responsible for severe diseases in domestic and wild birds.In September 2009, nine raptorial birds that either arrived recently or were maintained as permanent residents at the Safaripark Pombia (northwest Italy) showed loss of stamina, developing listlessness, anorexia and regurgitation. Within one month three animals died and were necropsied.Following the diagnosis of Plasmodium infection all other raptorial birds were treated: clinical improvement was observed in all birds, and blood smears made after one month resulted negative for parasites.

In situ hybridization and sequence analysis reveal an association of Plasmodium spp. with mortalities in wild passerine birds in Austria.

Native European passerine birds are frequently clinically inapparent carriers of haemosporidian parasites of the genus Plasmodium. Clinical disease and death are only exceptionally reported. In the present study, tissue samples of 233 wild passerine birds found dead in Eastern Austria were examined by in situ hybridization (ISH) and partial cytochrome B gene sequence analysis for the presence, abundance and taxonomic assignment of Plasmodium spp. In 34 cases (14.6%), ISH yielded a positive result with large numbers of developmental stages in different cell types of the spleen, liver, brain and lung. The abundance of the tissue stages, which was comparable to fatal cases of avian malaria in penguins, suggested a major contribution to the cause of death. Genetic analysis revealed infections with representatives of three different valid species of Plasmodium, Plasmodium elongatum, Plasmodium lutzi and Plasmodium vaughani. Genetically identical parasite lineages had been found in a previous study in penguins kept in the Vienna zoo, providing evidence for the role of wild birds as reservoir hosts. Further, this study provides evidence that several species of Plasmodium are able to abundantly proliferate in endemic wild birds ultimately resulting in mortalities.

Hand-rearing, release and survival of African penguin chicks abandoned before independence by moulting parents.

The African penguin Spheniscus demersus has an 'Endangered' conservation status and a decreasing population. Following abandonment, 841 African penguin chicks in 2006 and 481 in 2007 were admitted to SANCCOB (Southern African Foundation for the Conservation of Coastal Birds) for hand-rearing from colonies in the Western Cape, South Africa, after large numbers of breeding adults commenced moult with chicks still in the nest. Of those admitted, 91% and 73% respectively were released into the wild. There were veterinary concerns about avian malaria, airsacculitis and pneumonia, feather-loss and pododermatitis (bumblefoot). Post-release juvenile (0.32, s.e.  = 0.08) and adult (0.76, s.e.  = 0.10) survival rates were similar to African penguin chicks reared after oil spills and to recent survival rates recorded for naturally-reared birds. By December 2012, 12 birds had bred, six at their colony of origin, and the apparent recruitment rate was 0.11 (s.e.  = 0.03). Hand-rearing of abandoned penguin chicks is recommended as a conservation tool to limit mortality and to bolster the population at specific colonies. The feasibility of conservation translocations for the creation of new colonies for this species using hand-reared chicks warrants investigation. Any such programme would be predicated on adequate disease surveillance programmes established to minimise the risk of disease introduction to wild birds.

Food availability and competition do not modulate the costs of Plasmodium infection in dominant male canaries.

Understanding the different factors that may influence parasite virulence is of fundamental interest to ecologists and evolutionary biologists. It has recently been demonstrated that parasite virulence may occur partly through manipulation of host competitive ability. Differences in competitive ability associated with the social status (dominant or subordinate) of a host may determine the extent of this competition-mediated parasite virulence. We proposed that differences between subordinate and dominant birds in the physiological costs of infection may change depending on the level of competition in social groups. We observed flocks of domestic canaries to determine dominant or subordinate birds, and modified competition by providing restricted (high competition) or ad libitum food (low competition). Entire flocks were then infected with either the avian malaria parasite, Plasmodium relictum or a control. Contrary to our predictions we found that the level of competition had no effect on the outcome of infection for dominant or subordinate birds. We found that dominant birds appeared to suffer greater infection mediated morbidity in both dietary treatments, with a higher and more sustained reduction in haematocrit, and higher parasitaemia, than subordinates. Our results show that dominance status in birds can certainly alter parasite virulence, though the links between food availability, competition, nutrition and virulence are likely to be complex and multifaceted.

Identification of Plasmodium relictum causing mortality in penguins (Spheniscus magellanicus) from São Paulo Zoo, Brazil.

This study reports avian malaria caused by Plasmodium relictum in Magellanic Penguins (Spheniscus magellanicus) from São Paulo Zoo. The disease was highly infective among the birds and was clinically characterized by its acute course and high mortality. The penguins of São Paulo Zoo were housed for at least 2 years without malaria; however, they had always been maintained in an enclosure protected from mosquito exposure during the night period. When they presented pododermatitis, they were freed at night for a short period. São Paulo Zoo is located in one of the last forest remnants of the city, an area of original Atlantic forest. In the winter, the space destined for Zoo birds is shared with migratory species. Hence the possibility exists that the disease was transmitted to the penguins by mosquitoes that had previously bitten infected wild birds. Avian malaria parasites are transmitted mainly by mosquitoes of the genera Aedes and Culex, common vectors in the Atlantic forest. In this study, one Culex (Cux.) sp. was found, infected with P. relictum. There are diverse problems in housing distinct species of animals in captivity, principally when occupying the same enclosure, since it facilitates the transmission of diseases with indirect cycles, as is the case of Plasmodium spp., because certain species that cause discrete infections in some bird species can become a serious danger for others, especially penguins, which do not possess natural resistance. Thus, serious implications exist for periodically testing and administrating malaria therapy in captive penguins potentially exposed to mosquitoes during the night period, as well as other captive birds from São Paulo Zoo.

Global phylogeographic limits of Hawaii's avian malaria.

The introduction of avian malaria (Plasmodium relictum) to Hawaii has provided a model system for studying the influence of exotic disease on naive host populations. Little is known, however, about the origin or the genetic variation of Hawaii's malaria and traditional classification methods have confounded attempts to place the parasite within a global ecological and evolutionary context. Using fragments of the parasite mitochondrial gene cytochrome b and the nuclear gene dihydrofolate reductase-thymidylate synthase obtained from a global survey of greater than 13000 avian samples, we show that Hawaii's avian malaria, which can cause high mortality and is a major limiting factor for many species of native passerines, represents just one of the numerous lineages composing the morphological parasite species. The single parasite lineage detected in Hawaii exhibits a broad host distribution worldwide and is dominant on several other remote oceanic islands, including Bermuda and Moorea, French Polynesia. The rarity of this lineage in the continental New World and the restriction of closely related lineages to the Old World suggest limitations to the transmission of reproductively isolated parasite groups within the morphological species.

Interactive effects of turkeypox virus and Plasmodium hermani on Turkey poults.

Two experiments were conducted to examine the interactive effects of two disease agents of wild turkeys (Meleagris gallopavo), turkeypox virus and the malarial organism, Plasmodium hermani, on the health of turkey poults. Groups of domestic broad-breasted white turkey poults of 1 and 10 wk of age were infected with either turkeypox virus, P. hermani, both turkeypox virus and P. hermani, or were maintained as uninfected controls. The strains of turkeypox virus and P. hermani had been isolated from wild turkeys in southern Florida (USA). The goals of these experiments were two-fold and included both an examination of age differences in response to infections, and an examination of the effects of dual versus singular infections with the two agents. Both singular and concomitant infections of turkeypox virus and P. hermani were more detrimental to poults infected at 1 wk of age than to those infected at 10 wk, based on mortality, weight gain, and parasitemia. Dual infections of turkeypox virus and P. hermani were found to be slightly more harmful to 1-wk-old poults than were singular infections. No such interactive effects were noted in the poults infected at 10 wk of age.

Measuring the effects of an ever-changing environment on malaria control.

The effectiveness of malaria control measures depends not only on the potency of the control measures themselves but also upon the influence of variables associated with the environment. Environmental variables have the capacity either to enhance or to impair the desired outcome. An optimal outcome in the field, which is ultimately the real goal of vaccine research, will result from prior knowledge of both the potency of the control measures and the role of environmental variables. Here we describe both the potential effectiveness of control measures and the problems associated with testing in an area of endemicity. We placed canaries with different immunologic backgrounds (e.g., naïve to malaria infection, vaccinated naïve, and immune) directly into an area where avian malaria, Plasmodium relictum, is endemic. In our study setting, canaries that are naïve to malaria infection routinely suffer approximately 50% mortality during their first period of exposure to the disease. In comparison, birds vaccinated and boosted with a DNA vaccine plasmid encoding the circumsporozoite protein of P. relictum exhibited a moderate degree of protection against natural infection (P < 0.01). In the second year we followed the fate of all surviving birds with no further manipulation. The vaccinated birds from the first year were no longer statistically distinguishable for protection against malaria from cages of naïve birds. During this period, 36% of vaccinated birds died of malaria. We postulate that the vaccine-induced protective immune responses prevented the acquisition of natural immunity similar to that concurrently acquired by birds in a neighboring cage. These results indicate that dominant environmental parameters associated with malaria deaths can be addressed before their application to a less malleable human system.

Prevalência e variação dos estádios eritrocíticos do plasmodium (Novyella) juxtanucleare em Gallus gallus sob condições naturais, no período de um ano / Prevalence and variation of the erytrocytic forms of plasmodium (novyella) juxtanucleare in Gallus gallus about natural conditions, in the period of one year

The avian malaria caused by Plasmodium juxtanucleare in Gallus gallus, is a tipical plasmodiose from Brazilian gallinaceous. This disease can causes morbidy and mortality in its vertebrate hosts. This work was conducted at Boa Vista farm, Municipality of Santa Bárbara do Tugúrio, Minas Gerais, Brazil and the ours objectives were to evaluate the hight prevalence found in previous studies and to accompany the variation of the erytrocytic forms during one year. The bloods smears, dyed with Giemsa were examined in microscopy immersion. Twenty five half-breed fowls were accompany duting one year, monthly (from November/00 to May/01) and biweekly (from June/01 to October/01). The erytrocytic forms were registered and quantified by the observation of 100 microscopic fields. Was verified a prevalence of 100 por ciento by P. juxtanuclerare, but there wasn't statistics correlation between the increase of the erytrocytic forms during the year. The trophozoites were the more abundant form found in this studie.

Molecular characterization of malarial parasites in captive passerine birds.

Seven of 28 passerine birds that died in captivity were positive for malarial parasites by polymerase chain reaction targeting the mitochondrial cytochrome b (cytB) and apicoplast ribosomal RNA (rRNA) genes. Each bird was infected with a single parasite lineage having a unique genotype. Apicoplast rRNA sequences were present both in Haemoproteus spp. and Plasmodium spp. and had typically high adenosine + thymidine content. Phylogenies for cytB and apicoplast rRNA sequences were largely congruent and supported previous studies that suggest that Plasmodium-Haemoproteus spp. underwent synchronous speciation with their avian hosts, interrupted by sporadic episodes of host switching. Apicoplast phylogeny further indicated that Haemoproteus spp. are ancestral to Plasmodium spp. All the 7 infected passerine birds had histologic lesions of malaria, and malarial parasites may have contributed to the death of at least 4 animals. These findings provide new genetic data on passerine hematozoa, including initial sequences of apicoplast DNA, and emphasize the relevance of parasite prevalence, evolutionary relationships, and host switching to modern management and husbandry practices of captive birds.

Plasmodium Juxtanucleare associated with mortality in black-footed penguins (Spheniscus demersus) admitted to a rehabilitation center.

Five black-footed penguins (Spheniscus demersus) admitted to the Southern African Foundation for the Conservation of Coastal Birds, in Cape Town, South Africa, died from malaria infection. Evidence for malaria as the cause of death included antemortem clinical signs, parasitemia, splenomegaly, pulmonary edema, and the presence of histologically visible schizonts in the reticuloendothelial system. A portion of the malarial small subunit ribosomal ribonucleic acid gene was detected by polymerase chain reaction from postmortem blood samples from all the birds. A species-specific variable region of this gene was compared with the same region on genes from other known avian malarial organisms, establishing that Plasmodium juxtanucleare was involved.

The effect of Plasmodium gallinaceum on a challenge infection with Ascaridia galli in chickens.

The effect of a primary infection with the haemoparasite Plasmodium gallinaceum on the establishment of a challenge infection with the nematode Ascaridia galli in chickens was studied. Four groups were infected as follows. Group 1: inoculated intravenously with 10(6) P. gallinaceum-infected erythrocytes on day 0; group 2: orally infected with 500 embryonated A. galli eggs on day 10; group 3: infected with P. gallinaceum on day 0 and A. galli on day 10; and group 3: non-infected control birds. The results of this investigation demonstrates that a primary infection with P. gallinaceum in chickens alters the course of a subsequent infection with A. galli. Thus, an antagonistic effect was seen in which the malaria infection caused a significant reduction on the establishment of the nematode in concurrently infected animals.

Pathogenicity of avian malaria in experimentally-infected Hawaii Amakihi.

The introduction of avian malaria (Plasmodium relictum) and mosquitoes (Culex quinquefasciatus) to the Hawaiian Islands (USA) is believed to have played a major role in the decline and extinction of native Hawaiian honeycreepers (Drepanidinae). This introduced disease is thought to be one of the primary factors limiting recovery of honeycreepers at elevations below 1,200 m where native forest habitats are still relatively intact. One of the few remaining species of honeycreepers with a wide elevational distribution is the Hawaii Amakihi (Hernignathus virens). We measured morbidity and mortality in experimentally-infected Hawaii Amakihi that were captured in a high elevation, xeric habitat that is above the current range of the mosquito vector. Mortality among amakihi exposed to a single infective mosquito bite was 65% (13/20). All infected birds had significant declines in food consumption and a corresponding loss in body weight over the 60 day course of the experiment. Gross and microscopic lesions in birds that succumbed to malaria included enlargement and discoloration of the spleen and liver and parasitemias as high as 50% of circulating erythrocytes. Mortality in experimentally-infected amakihi was similar to that observed in Apapane (Himnatione sanguinea) and lower than that observed in Iiwi (Vestiaria coccinea) infected under similar conditions with the same parasite isolate. We conclude that the current elevational and geographic distribution of Hawaiian honeycreepers is determined by relative susceptibility to avian malaria.