Polymorphism evidence in Plasmodium (Haemamoeba) lutzi Lucena, 1939 (Apicomplexa, Haemosporida) isolated from Brazilian wild birds.

Plasmodium parasites can infect great variety of bird species around the world inflicting the so called avian malaria, an illness that could be fatal in some cases and consequently, should be monitored and widely included into conservation programs. The aim of this study was to characterize two lineages of Plasmodium (Haemamoeba) lutzi found in some birds in the Atlantic Forest of Minas Gerais - Brazil, that were morphologically identified after blood smears analyses under light microscopy and molecularly by sequencing the mitochondrial cytochrome b gene (cyt b). Besides these two lineages could be clearly morphologically identified as P.(H.) lutzi, some variations in comparison with its original description were noticed: absence of meronts and gametocytes (early and fully grown) in polychromatic erythrocytes, the larger size of pigment granules in meronts and gametocytes, and the presence of small vacuoles between pigment accumulation in fully grow macrogametocytes. Moreover, a certain degree of genetic intraspecific diversity was also observed across the lineages of P. (H.) lutzi, indicating the existence of polymorphisms within this taxon, which is uncommon in Haemosporida. These results allow discussion about species boundaries within avian hemosporidians and highlight the importance of multidisciplinary approaches for a more efficient species identification and characterization.

Molecular and Morphological Characterization of a Brazilian Lineage of Plasmodium ( Novyella) Unalis in Turdus Spp. (Passeriformes) of the Atlantic Forest, with Remarks on New Hosts and High Genetic Variation.

Plasmodium spp. are haemosporidian protozoans that alternate their live cycles between bloodsucking Culicidae dipterans and vertebrate hosts (mammals, reptiles, and birds). In birds, these parasites are the causative agents of the so-called avian malaria, a disease associated with considerable declines and extinctions in the avifauna in different geographical regions. In this work, we applied a multidisciplinary approach, light microscopy and cytochrome oxidase b (cyt b) gene sequence analysis, for characterization of Plasmodium spp. found in association with wild birds of the genus Turdus, collected in Atlantic forest fragments of southeastern Brazil. From the total 90 analyzed birds, 58 (47 Turdus rufiventris, 9 Turdus leucomelas, 1 Turdus albicollis, and 1 Turdus flavipes) were positively infected with Plasmodium unalis, a haemosporidian that was previously detected in Turdus fuscater in Colombia and in penguins in Brazil, but has never been found in association with these Turdus species of this present work. Moreover, all 7 new sequences of P. unalis cyt b gene clustered into a monophyletic clade with previously characterized P. unalis sequences with a mean genetic divergence of 1.6% and with a maximum divergence of 3.1%, indicating for a high degree of intraspecific polymorphism within this parasitic species. Together, our data highlight the existence a high degree of intraspecific variation within P. unalis and highlight the importance of integrative taxonomy to an accurate identification and characterization of avian haemosporidian parasites.

Multidisciplinary re-description of Plasmodium (Novyella) paranucleophilum in Brazilian wild birds of the Atlantic Forest kept in captivity.

Haemosporidian blood parasites of the Plasmodium genus are the causative agents of avian malaria in many parts of the world. Despite the great diversity of Brazilian avifauna, few studies have been conducted to examine the haemosporidians of wild birds found in the Brazilian Atlantic Forest, especially those kept in captivity. This study aimed to re-examine and further characterize the South American avian parasite Plasmodium paranucleophilum using a multidisciplinary approach. Blood samples were collected from 68 captive birds representing 15 species found in the Atlantic Forest of southeastern Brazil. Morphometric and morphological characterization was performed, in addition to PCR and sequencing of the mitochondrial cytochrome b gene and subsequent phylogenetic analysis. The overall prevalence of P. paranucleophilum infection in the study was 13.23% (n = 9), with a mean parasitemia of 0.58%. We observed the highest parasitemia of 3.88% in Rupornis magnirostris. In our phylogenetic analysis, P. paranucleophilum and P lasmodium nucleophilum formed distinct, highly supported clades, with a mean genetic divergence of 2.48%. This study provides new morphological and molecular data, expanding our knowledge of the haemosporidians of wild birds in Brazil and highlighting the need for further investigation. The true depth of diversity in Brazilian avian haemosporidians remains largely unknown, and given the enormous variety of vectors and avian species, there may be many more species of these blood parasites yet to be described.

Prevalence of Haemoproteus spp. (Apicomplexa: Haemoproteidae) in tortoises in Brazil and its molecular phylogeny.

Captive terrestrial tortoises of the species Chelonoidis carbonaria (n = 17) and Chelonoidis denticulata (n = 37) in the state of Minas Gerais, southeastern Brazil, were examined for hematozoans by using a combination of microscopic and molecular methods. Microscopic examination revealed young intra-erythrocytic forms in blood smears from both species of tortoises. The results of PCR, sequencing, and phylogenetic analysis indicated that these parasites belonged to the Haemoproteus spp., whose observed prevalence was 17.6 % in C. carbonaria and 13.5 % in C. denticulata. Phylogenetic analysis indicated that these sequences formed a clade that was grouped with other sequences of Haemoproteus spp. parasites in birds, separate from the clade formed by Haemoproteus spp. of reptiles. This study expands the information regarding the occurrence and distribution of hemosporidia in turtles and is the first study of blood parasites in C. carbonaria.

Molecular Characterization and Biochemical and Histopathological Aspects of the Parasitism of Haemoproteus spp. in Southern Caracaras (Caracara plancus).

Haemoproteid species have a wide global distribution, and they have been described in falcon species in several parts of the world. However, few studies in South America have focused on these birds. Haemoproteus spp. infections have been reported as the causative agents of serious histopathological changes, which can lead to the death of the host. Thus, this study aimed to molecularly and phylogenetically characterize Haemoproteus spp. in Caracara plancus, to characterize aspects of parasitism through clinical analysis and biochemical parameters, and to describe the histopathology of infection. To examine these aspects, 5 southern caracaras were examined clinically, and blood samples were collected. Blood smears were subsequently utilized in parasitemia calculations, PCR amplification, and serum biochemical investigations. Histological sections of the liver, kidneys, spleen, and heart were examined to check for possible histopathological changes. The birds showed clinical signs such as pallor and prostration that are consistent with Haemoproteus spp. infection. Moreover, the examination of the blood smears revealed 0.07% parasitemia in young gametocytes only. The PCR and sequencing results confirmed that the parasites belonged to Haemoproteus spp. The activity of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) enzymes, albumin, total serum proteins, and enzymatic urea were first described in C. plancus and serve as reference for future studies of bird species parasitized by Haemoproteus spp. Histopathology results showed signs of injury that were consistent with haemosporidian infection in the tissues of the analyzed organs. The present study is preliminary, and additional studies of Haemoproteus spp. infections in other bird species are needed to better understand the relationship between parasites and hosts, because despite the low parasitemia recorded, biochemical and histopathological changes in various organs were observed.

Plasmodium spp. and Haemoproteus spp. infection in birds of the Brazilian Atlantic Forest detected by microscopy and polymerase chain reaction / Infecção por Plasmodium spp. e Haemoproteus spp. em aves da Mata Atlântica brasileira detectada por microscopia e reação em cadeia da polimerase

In recent years haemosporidian infection by protozoa of the genus Plasmodium and Haemoproteus, has been considered one of the most important factors related to the extinction and/or population decline of several species of birds worldwide. In Brazil, despite the large avian biodiversity, few studies have been designed to detect this infection, especially among wild birds in captivity. Thus, the objective of this study was to analyze the prevalence of Plasmodium spp. and Haemoproteus spp. infection in wild birds in captivity in the Atlantic Forest of southeastern Brazil using microscopy and the polymerase chain reaction. Blood samples of 119 different species of birds kept in captivity at IBAMA during the period of July 2011 to July 2012 were collected. The parasite density was determined based only on readings of blood smears by light microscopy. The mean prevalence of Plasmodium spp. and Haemoproteus spp. infection obtained through the microscopic examination of blood smears and PCR were similar (83.19% and 81.3%, respectively), with Caracara plancus and Saltator similis being the most parasitized. The mean parasitemia determined by the microscopic counting of evolutionary forms of Plasmodium spp. and Haemoproteus spp. was 1.51%. The results obtained from this study reinforce the importance of the handling of captive birds, especially when they will be reintroduced into the wild...

Nos últimos anos infecção por protozoários hemosporídeos dos gêneros Plasmodium e Haemoproteus, tem sido considerada um dos fatores mais importantes relacionados com a extinção e / ou declínio da população de várias espécies de aves em todo o mundo. No Brasil, apesar da grande biodiversidade aviária, poucos estudos foram desenvolvidos para detectar a infecção, especialmente entre as aves silvestres mantidas em cativeiro. Assim, o objetivo deste estudo foi analisar a prevalência de infecção por Plasmodium spp. e Haemoproteus spp. em aves silvestres em cativeiro na Mata Atlântica do sudeste do Brasil, utilizando microscopia convencional e reação em cadeia da polimerase. Amostras de sangue de 119 aves mantidas em cativeiro no Ibama durante o período de julho de 2011 a julho de 2012, foram coletadas. A densidade parasitária foi determinada com base apenas em leituras de esfregaços de sangue por microscopia fotônica. A prevalência média de infecção por Plasmodium spp. e Haemoproteus spp. obtida por exame microscópico de esfregaços sanguíneos e PCR foi semelhante (83,19% e 81,3%, respectivamente), com Caracara plancus e Saltator similis sendo as espécies mais parasitadas. A parasitemia média determinada pela contagem microscópica de formas evolutivas de Plasmodium spp. e Haemoproteus spp. foi de 1,51%. Os resultados obtidos neste estudo reforçam a importância do manejo de aves em cativeiro, especialmente quando serão reintroduzidas na natureza...