Occurrence of endo and hemoparasites in Sporophila caerulescens captured in the eastern region of the state of Acre , Brazil

Wild animals are hosts and reservoirs to many infectious agents, often unknown to the Scientific Community, which leads to serious health implications for natural and domestic environments. We conducted this research with the objective of diagnosing the occurrence of parasites of medical and veterinary interest in “coleirinhos” captured in the eastern region of the state of Acre, Brazil. We captured the specimens in August and September of 2017 using ornithological nets at the margins of Caeté river. We visually inspected the birds to evaluate the ectoparasites. Subsequently, we collected blood through a brachial vein puncture and rested the individuals in a cloth bag for 30 minutes to collect feces, after which we released them. Results of the visual inspection to detect and identify the ectoparasites were negative. Blood samples revealed the protozoan Haemoproteus sp., and the feces revealed cysts of Entamoeba histolytica and E. coli, oocysts of coccidia and eggs of Davaineidae. Coccidia was the most prevalent (69.2%) and abundant (34.15%) parasite reported. Parasites diagnosed in this study can promote the emergence of secondary infections in S. caerulescens individuals or other animals that are contaminated with these etiological agents, since, when migrating through different biomes of South America, other animal species can come into contact with these agents.


INTRODUCTION
The Amazon is the Brazilian biome with the highest number of bird species and endemism.In this biome, 92% of the bird species are resident, and only 8% migrate to the Brazilian territory (SICK, 1993).There are 708 confirmed bird species only in the region of Acre, of which 78 are migratory species.Many come through Acre for a brief period, in search for food, shelter, or rest (GUILHERME, 2016).
According to FREITAS et al. ( 2002) the wild animals are hosts and reservoirs of many infectious agents, often unknown to the Scientific Community, which leads to serious implications for the health of natural and domestic environments, especially when there is direct contact between these animals.These researches must be constant, given that, when concerning migratory birds, we can also evaluate the dissemination of parasites from distinct environments using a single species as study subject (COSTA et al., 2010).
A few researches have demonstrated the importance of migratory birds as a host of many infectious agents that can infect the human population.Among these is the research conducted by RODRIGUES et al. (2007) on the influenza virus on human population.The authors reported that migratory birds, especially species of the Anseriformes, Passeriformes and Charadriformes orders, are natural reservoirs for this antigen and, for most species, the virus does not cause clinical disorders, but can infect domestic animals and consequently, the human population.
Several species of parasites have already been described parasitizing several groups of resident and migratory birds in virtually all geographical regions (SANTOS-PREZOTO et al., 2004;FONSECA et al., 2005).Currently, attention is drawn to the genus Plasmodium, which causes avian malaria, which until 2005 was recorded in 13.5% of the world avifauna analyzed for hemoparasites (BRUM et al., 2016).
In addition to the lack of knowledge on the biology of these etiological agents transmitted by the wild animals, another aggravating factor is the proximity of the human population to these birds.Such proximity occurs through the need of using the natural resources for subsistence, many of which are used by the animals or even are the animals themselves, generating a direct or indirect contact with possible etiological agents.
In this research, we aimed to diagnose the occurrence of parasites of medical and veterinary interest on a species of "coleirinhos" (Sporophila caerulescens) captured in the eastern region of the state of Acre, Brazil.

Specimens of Sporophila caerulescens
were captured in the "Comunidade Cuidado" (S 09º08'50.9";W 069º01'22.3") located in the Cazumbá-Iracema Extractive Reserve, situated in the center-west region of the state of Acre, in the municipality of Sena Madureira.The reserve area is part of the West Corredor of the Amazon, one of the seven large ecological corridors proposed for Brazil.
We captured 18 individuals between August and September of 2017 using ornithological nets (Mistnets) placed in the morning (5:00 h -12:00 h) and afternoon (14:00 h -17:00 h), at the margins of Caeté river.After capturing the specimens, we identified and measured the individuals, then they were marked by the strategic feathers of the tail, for eventual recapture notification.A stool sample, when present and two blood samples were collected from each individual.Of the individuals recaptured, no new collection was performed.
We visually inspected the birds to evaluate the parasites.After inspection, we collected blood through brachial vein puncture on one of the wings after asepsis with cotton soaked in 70º alcohol (MATTER et al., 2010).Immediately after, we performed two blood smears from each individual in glass laminas fixed with methanol and stained using the Giemsa (Eosin-Methylene Blue) and Fast Panoptic (May-Grunwald-Giemsa) methods (DE CARLI, 2001).
For collecting the feces, we maintained the individuals in cloth bags containing absorbent paper within for 20 to 30 minutes.After this period, we released the birds and conditioned the feces, when any, in sterile plastic recipients, and sent them to the Wild Life Support Laboratory of the Federal University of Acre, where the parasitological analyzes were performed.
To prepare the laminas for microscopic analysis and quantifying the number of oocyts per defecation (OoPD), we used the fecal smear examination stained with Lugol solution by light microscopy, spontaneous sedimentation and centrifugation-flotation technique (DE CARLI, 2001).After sedimentation for 12 hours, part of the sediment was placed on glass slide, added lugol and examined under light microscopy (Olympus CX40; objectives: 10x, 40x, and 100x), being used a 40x increase for the photographs.For the coccidia research the modified formaldehyde concentration technique was used, followed by staining of the smears by the modified Ziehl-Neelsen technique (LENETTE et al., 1985).The stained smears were observed under immersion microscopy.The identification of the parasites was performed according to the characteristics described by FOREYT (2005).
We classified the infection intensity in (0) negative animal; (1) animal with a low infection between 1 and 100 OoPD; (2) animal with a moderate infection between 101 and 500 OoPD; and (3) animal with high infection of more than 500 OoPD (VASCONCELLOS et al., 2013).For the statistical analyzes of the data, prevalence and abundance were calculated, as defined by BUSH et at.(1997).

RESULTS
Thirteen stool samples were collected.We reported parasites in 92.3% (n=12) of the collected feces samples.The endoparasite identified were: cysts of Entamoeba histolytica and E. coli, oocysts of coccidia, and eggs of Davaineidae (Figure 1).
The visual inspection of the specimens to detect and identify ectoparasite was negative, and only 22.22% (4/18) of the individuals were positive for the presence of hemoparasites, identifying the protozoan Haemoproteus sp.
The presence of E. histolytica and E. coli concerns since they are parasites of public health interest and pathogens of human intestines.Human infection can occur by respiration when aspirating dust from areas contaminated by dry feces, or by ingesting dust or food contaminated with bird feces (SILVA & CAPUANO, 2008;COURA, 2015).
These birds are commonly reared in cages and the contact with the feces when cleaning the cages can represent another form for human infection  2009) reported the presence of coccidians parasitizing the species S. caerulescens and S. nigricollis when analyzing fecal samples of wild and exotic birds in Botocatu, SP.In addition to these records mortality caused by the coccidia has been described for the Sporophila species (VASCONCELOS et al., 2012).According to the authors two S. maximilliani individuals died in captivity due to the high infestation by coccidia.In "bicudos" (S. maximiliani) and "curiós" (S. angolensis), we found reports of outbreaks and mortality of young between two and three months of age caused by coccidia (CUBAS et al., 2014).
The infection caused by coccidia, manifest through diarrhea, apathy, egg drop decrease, and weight loss, which can lead to the death of the infected birds, especially the young ones (JÚNIOR et al., 2000;CUBAS et al., 2014).This is concerning to the researched specimens and other species that are infected with these parasites, given that the coccidia infect all classes of vertebrates, are easily transmitted with the host´s ingestion of sporulated oocyts (PAGE & HADDAD, 1995;FRIEND & FRANSON, 1999;LOPEZ et al., 2007;YABSLEY, 2008).
Among the blood parasites most common in birds, the genres Plasmodium and Haemoproteus are the most frequent, with more than 200 species of this hemoparasite being described based on morphology in 4,000 studies bird species (VALKIUNAS, 2005), results that corroborated those reported in this research.
According to BENSCH et al. (2000) the infections by hemoparasited lead to severe problems for the host, especially if native bird species are exposed for the first time.At the beginning of the 20 th century, the introduction of a new species of Plasmodium resulted in the extinction of birds native to Hawai, demonstrating the devastating effect of invasive disease (DASZAK et al., 2000;DOBSON & FOUFOPOULOS, 2001;HARVELL et al., 2000).
None of the evaluated animals presented clinical signs suggested any pathological change, and low values of infection indicators can be explained by the fact that parasitic infections do not generally manifest clinically under natural conditions given the balance between parasite and host populations.In addition, coccidian-infected birds develop a solid immunity that protects them against subsequent infections (GALHA et al., 2008).The main form of immunity involved in protection is cellular, mainly played by T cells in the nucleus of the intestinal group (GALT), since GALT has an immune function and triggering effector responses, but immunity does not expand cell invasion by schizonts, only development of the schizonts in the interior, delaying the infection (DIETERT et al., 1994;LILLEHOJ, 1999;GALHA et al., 2008).
However, according to DUARTE (1997) who gathered conditions that aid in controlling host resistance, such as malnutrition and intercurrent diseases, this balance can be altered in favor of the parasite, which can cause problems to the researched species or other species that occupy the same locations in which this species forages or pass during their migratory period due to dispersion and contamination.
According to FIGUEIREDO et al. (2010) parasitological studies using wild animals, especially migratory birds, as studied species, contribute to the knowledge on parasite fauna and aid in remedying questions on the different parasite species from distinct environments.These studies assisted in prophylactic measures that care for the sanity of these animals and the prevention of zoonoses present in distinct environments, and that can be disseminated by hosts often unknown to the scientific community.

CONCLUSION
Passeriform birds of the species S. caerulescens captured in the Extractive Reserve of Cazumbá-Iracema, State of Acre, presented parasitism by hemoparasite, intestinal helminths and protozoan, with a predominance of infection with coccidia.

BIOETHICS AND BIOSSECURITY COMMITTEE APPROVAL
This research was evaluated and approved by the Biodiversity Authorization and Information System -SISBIO nº 67436297 and Animal Use Ethics Committee of the Federal University of Acre nº 2107.009788/2016-01.

Table 1 -
Endoparasites present in the samples of faces of the Sporophila caerulescens captured in the Cazumbá-Iracema Extractive Reserve in Sena Madureira, Acre, Brazil.PS -positive samples, CS -collected samples, P -prevalence, and A -abundance.