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The relationships between host phylogenetics, functional traits and parasites in wildlife remain poorly understood in the Neotropics, especially in habitats with marked seasonal variation. Here, we examined the effect of seasonality and host functional traits on the prevalence of avian haemosporidians (Plasmodium and Haemoproteus) in the Brazilian Caatinga, a seasonally dry tropical forest. 933 birds were evaluated for haemosporidian infections. We found a high parasitism prevalence (51.2%), which was correlated with phylogenetic relatedness among avian species. Prevalence varied drastically among the 20 well-sampled species, ranging from 0 to 70%. Seasonality was the main factor associated with infections, but how this abiotic condition influenced parasite prevalence varied according to the host-parasite system. Plasmodium prevalence increased during the rainy season and, after excluding the large sample size of Columbiformes (n = 462/933), Plasmodium infection rate was maintained high in the wet season and showed a negative association with host body mass. No association was found between non-Columbiform bird prevalence and seasonality or body mass when evaluating both Plasmodium and Haemoproteus or only Haemoproteus infections. Parasite community was composed of 32 lineages including 7 new lineages. We evidenced that even dry domains can harbour a high prevalence and diversity of vector-borne parasites and pointed out seasonality as a ruling factor.

We report, for the first time, the presence of ungulate malaria parasites in South America. We conducted PCR-based surveys of blood samples of multiple deer species and water buffalo from Brazil and detected Plasmodium sequences from pampas deer (Ozotoceros bezoarticus) samples. Phylogenic analysis revealed that the obtained sequences are closely related to the Plasmodium odocoilei clade 2 sequence from North American white-tailed deer (Odocoileus virginianus). Nucleotide differences suggest that malaria parasites in South American pampas deer and North American P. odocoilei clade 2 branched more recently than the Great American Interchange. Keywords: Malaria, Pampas deer, South America, Plasmodium odocoilei, Brazil

Abstract Background Haemosporidian parasites of the genera Plasmodium and Haemoproteus can have detrimental effects on individual birds and populations. Despite recent investigations into the distribution and richness of these parasites and their vertebrate hosts, little is known about their dipteran vectors. The Neotropics has the highest diversity of mosquitoes in the world, but few studies have tried to identify vectors in this area, hampering the understanding of the ecology of avian malaria in the highly diverse Neotropical environments. Methods Shannon traps and active collection were used to capture 27,110 mosquitoes in a Seasonally Dry Tropical Forest in southeastern Brazil, a highly endangered ecosystem. Results We screened 17,619 mosquito abdomens from 12 different species and several unidentified specimens of Culex, grouped into 1,913 pools, for the presence of haemosporidians. Two pools (out of 459) of the mosquito Mansonia titillans and one pool (out of 29) of Mansonia pseudotitillans were positive for Plasmodium parasites, with the detection of a new parasite lineage in the former species. Detected Plasmodium lineages were distributed in three different clades within the phylogenetic tree revealing that Mansonia mosquitoes are potential vectors of genetically distant parasites. Two pools of Culex spp. (out of 43) were positive for Plasmodium gallinaceum and closely related lineages. We found a higher abundance of these putative vectors in pasture areas, but they were also distributed in areas at intermediate and late successional stages. One pool of the mosquito Psorophora discrucians (out of 173) was positive for Haemoproteus. Conclusions The occurrence of different Plasmodium lineages in Mansonia mosquitoes indicates that this genus encompasses potential vectors of avian malaria parasites in Brazil, even though we did not find positive thoraces among the samples tested. Additional evidence is required to assign the role of Mansonia mosquitoes in avian malaria transmission and further studies will add information about evolutionary and ecological aspects of avian haemosporidia and untangle the diversity of their vectors in Brazil.

Blood parasites are potential threats to the health of penguins and to their conservation and management. Little penguins Eudyptula minor are native to Australia and New Zealand, and are susceptible to piroplasmids (Babesia), hemosporidians (Haemoproteus, Leucocytozoon, Plasmodium) and kinetoplastids (Trypanosoma). We studied a total of 263 wild little penguins at 20 sites along the Australian southeastern coast, in addition to 16 captive-bred little penguins. Babesia sp. was identified in seven wild little penguins, with positive individuals recorded in New South Wales, Victoria and Tasmania. True prevalence was estimated between 3.4% and 4.5%. Only round forms of the parasite were observed, and gene sequencing confirmed the identity of the parasite and demonstrated it is closely related to Babesia poelea from boobies (Sula spp.) and B. uriae from murres (Uria aalge). None of the Babesia-positive penguins presented signs of disease, confirming earlier suggestions that chronic infections by these parasites are not substantially problematic to otherwise healthy little penguins. We searched also for kinetoplastids, and despite targeted sampling of little penguins near the location where Trypanosoma eudyptulae was originally reported, this parasite was not detected.

Babesia spp. are tick-borne parasites, and 16 avian-infecting species have been described to date, including one species (Babesia peircei) that infects penguins. Considering the results of a recent study reporting Babesia sp. in penguins on Deception Island, South Shetland Islands, we re-examined the samples obtained in a previous investigation on the occurrence of blood parasites in adult Adélie (Pygoscelis adeliae), chinstrap (Pygoscelis antarcticus) and gentoo penguins (Pygoscelis papua) on King George and Elephant islands, South Shetland Islands. Notwithstanding a comprehensive re-examination of the blood smears, Babesia sp. was not detected. When we employed two nested PCR tests targeting the 18S rRNA gene of Babesia, a considerable proportion of the samples produced positive results; however, gene sequencing revealed these were due to cross-amplification of non-target organisms. We therefore did not detect Babesia sp. infection in penguins on King George and Elephant islands. Additional studies will be valuable to clarify the distribution and epidemiology of tick-borne pathogens in sub-Antarctic and Antarctic seabirds.

From an ecological point of view, parasites are considered important selective forces that influence all aspects of a host's life. The purpose of this study was to assess the health state of Turdus leucomelas Vieillot, 1818 (Turdidae: Passeriformes) inhabiting a Brazilian Cerrado in the breeding and molting seasons through a hematological parameter analysis and an evaluation of infection by blood parasites. The birds were collected with mist-nets, ringed and blood sampled to assess hematological parameters (hematocrit, hemoglobin concentration and white blood cells) and blood parasites. We detected blood parasites through optical microscopy and subsequently used PCR (amplification of the 18sRNA gene) to verify the presence of Plasmodium spp. (avian malaria). During the breeding season, the hemoglobin level and CHGM percentage were greater. Global leukocyte count was positively related to hemoglobin level, CHGM percentage and body weight. Of the total 31 T. leucomelas individuals examined, 18 presented Plasmodium parasites (58% of prevalence). There was a significant relationship between the presence of Plasmodium spp. and decreased CHGM. These results suggest a connection between the health parameters of wild birds and the physiological stress associated with the breeding season.

Background The integration of diagnostic methods holds promise for advancing the surveillance of malaria transmission in both endemic and non-endemic regions. Serological assays emerge as valuable tools to identify and delimit malaria transmission, serving as a complementary method to rapid diagnostic tests (RDT) and thick smear microscopy. Here, we evaluate the potential of antibodies directed against peptides encompassing the entire amino acid sequence of the PvMSP-1 Sal-I strain as viable serological biomarkers for P. vivax exposure. Methods We screened peptides encompassing the complete amino acid sequence of the Plasmodium vivax Merozoite Surface Protein 1 (PvMSP-1) Sal-I strain as potential biomarkers for P. vivax exposure. Here, immunodominant peptides specifically recognized by antibodies from individuals infected with P. vivax were identified using the SPOT-synthesis technique followed by immunoblotting. Two 15-mer peptides were selected based on their higher and specific reactivity in immunoblotting assays. Subsequently, peptides p70 and p314 were synthesized in soluble form using SPPS (Solid Phase Peptide Synthesis) and tested by ELISA (IgG, and subclasses). Results This study unveils the presence of IgG antibodies against the peptide p314 in most P. vivax-infected individuals from the Brazilian Amazon region. In silico B-cell epitope prediction further supports the utilization of p314 as a potential biomarker for evaluating malaria transmission, strengthened by its amino acid sequence being part of a conserved block of PvMSP-1. Indeed, compared to patients infected with P. falciparum and uninfected individuals never exposed to malaria, P. vivax-infected patients have a notably higher recognition of p314 by IgG1 and IgG3.

South America has different biomes with a high richness of wild bird species and Diptera vectors, representing an ideal place to study the influence of habitat on vector-borne parasites. In order to better understand how different types of habitats do or do not influence the prevalence of haemosporidians, we performed a new analysis of two published datasets comprising wild birds from the Brazilian Savanna (Cerrado) as well as wild birds from the Venezuelan Arid Zone. We investigated the prevalence and genetic diversity of haemosporidian parasites belonging to two genera: Plasmodium and Haemoproteus. We evaluated data from 676 wild birds from the Cerrado and observed an overall prevalence of 49%, whereas, in the Venezuelan Arid Zone, we analyzed data from 527 birds and found a similar overall prevalence of 43%. We recovered 44 lineages, finding Plasmodium parasites more prevalent in the Cerrado (15 Plasmodium and 12 Haemoproteus lineages) and Haemoproteus in the Venezuelan Arid Zone (seven Plasmodium and 10 Haemoproteus lineages). No difference was observed on parasite richness between the two biomes. We observed seven out of 44 haemosporidian lineages that are shared between these two distinct South American biomes. This pattern of parasite composition and prevalence may be a consequence of multiple factors, such as host diversity and particular environmental conditions, especially precipitation that modulate the vector’s dynamics. The relationship of blood parasites with the community of hosts in large and distinct ecosystems can provide more information about what factors are responsible for the variation in the prevalence and diversity of these parasites in an environment.

Avian haemosporidians are parasites with great capacity to spread to new environments and new hosts, being considered a good model to host-parasite interactions studies. Here, we examine avian haemosporidian parasites in a protected area covered by Restinga vegetation in northeastern Brazil, to test the hypothesis that haemosporidian prevalence is related to individual-level traits (age and breeding season), species-specific traits (diet, foraging strata, period of activity, species body weight, migratory status, and nest shape), and climate factors (temperature and rainfall). We screened DNA from 1,466 birds of 70 species captured monthly from April 2013 to March 2015. We detected an overall prevalence (Plasmodium/Haemoproteus infection) of 22% (44 host species) and parasite’s lineages were identified by mitochondrial cyt b gene. Our results showed that migration can be an important factor predicting the prevalence of Haemoproteus (Parahaemoproteus), but not Plasmodium, in hosts. Besides, the temperature, but not rainfall, seems to predict the prevalence of Plasmodium in this bird community. Neither individual-level traits analyzed nor the other species-specific traits tested were related to the probability of a bird becoming infected by haemosporidians. Our results point the importance of conducting local studies in particular environments to understand the degree of generality of factors impacting parasite prevalence in bird communities. Despite our attempts to find patterns of infection in this bird community, we should be aware that an avian haemosporidian community organization is highly complex and this complexity can be attributed to an intricate net of factors, some of which were not observed in this study and should be evaluated in future studies. We evidence the importance of looking to host-parasite relationships in a more close scale, to assure that some effects may not be obfuscated by differences in host life-history.

Three clones isolated from the Y strain of Trypanosoma cruzi - YP1, YP2 and YP3 - were adapted to in vitro cultivation in VERO cells. The recovery of the parasites from the Y strain and clone YP3 was similar after 24 hr of contact with cells (3.2% and 2.7%, respectively) and much lower than the recovery of clones YP1 and YP2 (56.7% and 60.0% of inoculum, respectively). After five days incubation, the ratio Trypomastigotes/Amastigotes released into the supernatants was about 90/10 for clone YP1, YP3 and Y strain, and 50/50 for clone YP2. After nine days, the ratio was 62/38 for clone YP1, 97/3 for clone YP3, 81/19 for Y strain and 50/50 for clone YP2. The susceptibility of tissue culture derived trypomastigotes (TCT) to lysis in the presence of chronic chagasic human sera and human complement was assessed using Complement Mediated Lysis reaction (CML). Trypomastigotes from clone YP2 were consistently less susceptible to CML (% lysis less than 20), than parasites from the other clones and Y strain. Parasites of clone YP3 had susceptibility to CML comparable to that of the Y strain (about 70%), while TCT of clone YP1 had intermediary susceptibility (40%).

Habitat modification may change vertebrate and vector-borne disease distributions. However, natural forest regeneration through secondary succession may mitigate these effects. Here we tested the hypothesis that secondary succession influences the distribution of birds and their haemosporidian parasites (genera Plasmodium and Haemoproteus) in a seasonally dry tropical forest, a globally threatened ecosystem, in Brazil. Moreover, we assessed seasonal fluctuations in parasite prevalence and distribution. We sampled birds in four different successional stages at the peak and end of the rainy season, as well as in the middle and at the end of the dry season. A non-metric multidimensional scaling analysis revealed that bird communities in the pasture (i.e., highly modified) areas were different from those in the early, intermediate, and late successional areas (secondary forests). Among 461 individual birds, haemosporidian prevalence was higher in pasture areas than in the more advanced successional stages, but parasite communities were homogeneous across these areas. Parasite prevalence was higher in pasture-specialists birds (resilient species) than in forest-specialists species, suggesting that pasture-specialists may increase infection risk for co-occurring hosts. We found an increase in prevalence between the middle and end of the dry season, a period associated with the beginning of the breeding season (early spring) in southeastern Brazil. We also found effects of seasonality in the relative prevalence of specific parasite lineages. Our results show that natural forest recovery through secondary succession in SDTFs is associated with compositional differences in avian communities, and that advanced successional stages are associated with lower prevalence of avian haemosporidian parasites.

Effect of Aedes fluviatilis saliva on the development of Plasmodium gallinaceum experimental infection in Gallus (gallus) domesticus was studied in distinct aspects. Chickens subcutaneously infected with sporozoites in the presence of the mosquito salivary gland homogenates (SGH) showed higher levels of parasitaemia when compared to those ones that received only the sporozoites. However, the parasitaemia levels were lower among chickens previously immunized by SGH or non-infected mosquito bites compared to the controls, which did not receive saliva. High levels of anti-saliva antibodies were observed in those immunized chickens. Moreover, 53 and 102 kDa saliva proteins were recognized by sera from immunized chickens. After the sporozoite challenge, the chickens also showed significant levels of anti-sporozoite antibodies. However, the ability to generate anti-sporozoites antibodies was not correlated to the saliva immunization. Our results suggest that mosquito saliva components enhance P. gallinaceum parasite development in naive chickens. However, the prior exposure of chickens to salivary components controls the parasitemia levels in infected individuals.

Avian malaria is a mosquito-borne disease caused by Plasmodium spp. Avian plasmodia are recognized conservation-threatening pathogens due to their potential to cause severe epizootics when introduced to bird populations with which they did not co-evolve. Penguins are considered particularly susceptible, as outbreaks in captive populations will often lead to high morbidity and rapid mortality. We used a multidisciplinary approach to investigate an outbreak of avian malaria in 28 Magellanic penguins (Spheniscus magellanicus) at a rehabilitation center during summer 2009 in Florianópolis, Brazil. Hemosporidian infections were identified by microscopic and molecular characterization in 64% (18/28) of the penguins, including Plasmodium (Haemamoeba) tejerai, Plasmodium (Huffia) elongatum, a Plasmodium (Haemamoeba) sp. lineage closely related to Plasmodium cathemerium, and a Haemoproteus (Parahaemoproteus) sp. lineage closely related to Haemoproteus syrnii. P. tejerai played a predominant role in the studied outbreak and was identified in 72% (13/18) of the hemosporidian-infected penguins, and in 89% (8/9) of the penguins that died, suggesting that this is a highly pathogenic parasite for penguins; a detailed description of tissue meronts and lesions is provided. Mixed infections were identified in three penguins, and involved P. elongatum and either P. tejerai or P. (Haemamoeba) sp. that were compatible with P. tejerai but could not be confirmed. In total, 32% (9/28) penguins died over the course of 16 days despite oral treatment with chloroquine followed by sulfadiazine-trimethoprim. Hemosporidian infections were considered likely to have occurred during rehabilitation, probably from mosquitoes infected while feeding on local native birds, whereas penguin-mosquito-penguin transmission may have played a role in later stages of the outbreak. Considering the seasonality of the infection, rehabilitation centers would benefit from narrowing their efforts to prevent avian malaria outbreaks to the penguins that are maintained throughout summer.

Habitat alteration can disrupt host-parasite interactions and lead to the emergence of new diseases in wild populations. The cerrado habitat of Brazil is being fragmented and degraded rapidly by agriculture and urbanization. We screened 676 wild birds from three habitats (intact cerrado, disturbed cerrado and transition area Amazonian rainforest-cerrado) for the presence of haemosporidian parasites (Plasmodium and Haemoproteus) to determine whether different habitats were associated with differences in the prevalence and diversity of infectious diseases in natural populations. Twenty one mitochondrial lineages, including 11 from Plasmodium and 10 from Haemoproteus were identified. Neither prevalence nor diversity of infections by Plasmodium spp. or Haemoproteus spp. differed significantly among the three habitats. However, 15 of the parasite lineages had not been previously described and might be restricted to these habitats or to the region. Six haemosporidian lineages previously known from other regions, particularly the Caribbean Basin, comprised 50-80% of the infections in each of the samples, indicating a regional relationship between parasite distribution and abundance.