Occurrence of gastrointestinal and renal helminths in Zenaida auriculata ( Des Murs , 1847 ) trap-captured from Brazil

This study aimed to determine the prevalence of gastrointestinal and renal helminths from naturally infected Zenaida auriculata captured in Londrina, Paraná State. Two hundred and one Eared doves were trapped and the gastrointestinal and renal helminths were collected and identified according to morphological structures. One hundred and sixteen (57.71%) doves were parasitized by helminths with specific prevalences for Ornithostrongylus quadriradiatus in 50 doves (24.88%), Ascaridia columbae in 47 (23.38%), Paratanaisia bragai and P. confusa in 34 (16.92%), Tetrameres fissispina in 17 (8.46%), Synhimantus nasuta in 14 (6.47%), Brachylaima mazzantii in 4 (1.99%) and Raillietina allomyodes in 2 doves (1.00%). Seventy four/201 (37.00%) birds were infected with only one species, and 96/201 (48.00%) pigeons were infected with nematodes. The association between different classes of helminths occurred in 40/201 (20.00%) animals. The results showed statistically differences between the presence of nematode (p = 0.00001) and trematode species (p ≤ 0.05) in the doves, and there was an association between the local of capture and the presence of trematodes and A. columbae (p ≤ 0.05). This study is the first to report the infection of Z. auriculata from Brazil with O. quadriradiatus, A. columbae, T. fissispina, S. nasuta, R. allomyodes, P. bragai and P. confusa.


Introduction
Zenaida auriculata (Des Murs, 1847) (Aves: Columbiforme) is endemic to an area ranging from Antilles to Tierra del Fuego, passing through Brazil.This dove is popularly known as the eared dove (CÂNDIDO et al., 2008;SHIBATTA et al., 2009) and is found in fields, farms and urban areas (CÂNDIDO et al., 2008), where it can cause damage and potentially transmit pathogens (SHIBATTA et al., 2009).
Of the numerous health problems that can affect pigeons, parasite infections may be the most important (MARQUES et al., 2007).Little is known of the species of gastrointestinal parasites that infect these birds (MARQUES et al., 2007).
There are very few studies on the presence of gastrointestinal helminths in Z. auriculata; Travassos and Freitas (1942) documented gastrointestinal helminths of doves in Brazil's Northeast Railroad; Adriano et al. (2001) reported the presence of the trematode Brachylaima mazzantii in the small intestine of this dove at Junqueirópolis-SP.Additionally, Durette-Desset et al. (2000) described the presence of a new species of helminth, Ornithostrongylus volcani n. sp., in doves in Venezuela.The only study on the prevalence of helminths in Z. auriculata was conducted in Chile by González et al. (2004); these authors encountered Killigrewia delafondi (9.4%), Raillietina sp.(1.7%), Heterakis gallinarum (0.4%) and Echinostomum sp.(0.9%).
The aim of the present study was to determine the prevalence of gastrointestinal and renal helminths in naturally infected Z. auriculata captured in the city of Londrina, Paraná State.

Study area and host data collection
Londrina is located in the north of Paraná state, southern Brazil (23°08'47" and 23°55'46" S, 50°52'23" and 51°19'11" W) (IBGE, 2010).Two hundred and one doves (Z.auriculata) were captured using gauze-traps from January 2010 to June 2011.These animals were collected from three different areas: 140 birds from a soybean seed industry, 45 from the campus of State University of Londrina, and 16 from a dairy cattle farm.This work was approved by the Brazilian Institute of Environment (IBAMA) SISBIO N.

Helminth data collection
The animals were euthanized in a CO 2 chamber, and their gastrointestinal tracts were removed and opened from the esophagus to the cloaca.The esophagi, crops, proventriculi, gizzards, small intestines and large intestines were individualized and placed in Petri dishes with 0.9% saline solution.Each organ was cut lengthwise and scanned for helminths.The helminths were removed from the crop and proventriculus after a detailed examination.The gastrointestinal content was taken from the mucosa, washed and passed through sieves with apertures of 600 µm, 425 µm and 150 µm (PAVANELLI; FERNANDES, 1982) for the separation of the helminths.The kidneys were removed from the abdominal cavity and cut lengthwise, and the renal tubules were slightly compressed to obtain the helminths.

Permanent slides preparation and identification
To prepare permanent slides of the parasites, the helminths were refrigerated for 24 hours (PAVANELLI;FERNANDES, 1982).The nematodes were placed in 37 °C Railliet-Henry solution for fixation, while the cestodes and trematodes were pressed between glass slides and dipped in Railliet-Henry solution (TRAVASSOS, 1950).Subsequently, the nematodes were cleared in Hoyer solution and a slide and coverslip were prepared according to the method described by Sloss et al. (1999).The thin nematodes were stained with lactophenol cotton blue (TRAVASSOS, 1950) before the slide preparation.The cestodes and trematodes were washed for five hours in distilled water, stained with Carmine Mayer, then sallowed with acid alcohol and dehydrated in alcoholic series.After the dehydration, the cestodes and trematodes were cleared in Hoyer solution and placed between the slide and coverslip for observation by optic microscopy (PESSOA, 1967).Helminth specimens were identified following taxonomic keys of Travassos (1913Travassos ( , 1914Travassos ( , 1919Travassos ( , 1921)), Yamaguti (1958Yamaguti ( , 1959Yamaguti ( , 1961)), Travassos and Kohn (1966), Travassos et al. (1969), Vicente et al. (1995) and Saif et al. (2008).

Statistical analysis
The terms prevalence (P), abundance (A) and mean intensity of parasitism (MI) follow the definitions of Bush et al. (1997), and these parameters were evaluated in Microsoft Excel 2007 (Microsoft Corporation, Redmond, Washington, USA).Chi square (χ 2 ) test corrected by Yates was used to compare the variables (Epi Info Program, version 6.04).A P-value of ≤0.05 was considered significant.
Ornithostrongylus quadriradiatus was the most prevalent species and occurred in 24.88% of the eared doves; A. columbae was the second most prevalent species and was encountered in 23.38% of the doves.However, P. bragai + P. confusa had the highest mean intensity of parasitism (220.62).Table 1 presents the number of parasitized animals, the prevalence, the abundance, the mean intensity of parasitism and the range of parasitism of the species.There were significant differences when nematodes O. quadriradiatus, A. columbae, S. nasuta and T. fissispina were compared.Adittionally, when the presence of trematodes was studied (Table 1), the doves had 10.03 times-higher P. bragai + P. confusa than B. mazzantii (OR = 10.03,3.46<OR<39.51,p<0.05).
Of the 201 captured doves, 97/201 (48.26%) were male and 104/201 (51.74%) were female.A total of 49/97 (50.52%) of the males and 67/104 (64.42%) of the females had parasites.Both male and female doves presented each class of helminth in the gastrointestinal tract and renal tubules.However, 25 females had trematodes and only 13 males presented this class (Table 2).There were no differences between male and female doves considering the infection from Class Nematoda (p>0.10) and Class Trematoda (p>0.08).
Association between the parasites and the locality of trap capture is shown in Table 3.The parasites from Class Nematoda were taking all together and compared with place of trap capture, and no differences were observed (p>0.06),however, A. columbae was more frequent in dairy farms (p = 0.005).In Trematoda class the soybean industry had a higher prevalence than University (OR = 0.21, 0.04<OR<0.74,p = 0.01).This occur by the fact that P. bragai + P. confusa were found just in the industry (p = 0.00025), additionally, no trematodes were observed in dairy farms.
It was observed that among the positive birds, they had one to four different species of helminths in the gastrointestinal tract and renal tubules.The majority of infected Z. auriculata 74/201 (37.00%) were parasitized with one species of parasite.The Nematoda class was the most frequently encountered class of parasite, found in 96/201 (48.00%) doves, followed by the Trematoda class in 38/201 (19.00%) doves and the Cestoda class in 2/201 (1.00%) doves.

Discussion
A few studies on Z. auriculata helminths have been conducted worldwide.To the authors' knowledge, however, the data on prevalence, abundance, and mean intensity of parasitism of helminths in Z. auriculata have not previously been collected.Lee et al. (2004) worked with Z. macroura in the USA and they did not observe differences between local of capture, and sex of those birds compared with the presence of helminths.In a different study, with another species of bird, Z. asiatica, Glass et al. (2002) showed that helminths may be found equally in both sex.Those results are similar to what we observed in the present study.
The presence of A. columbae in doves from dairy farm was higher than other localities in our study, and this was observed previously (GLASS et al., 2002).In fact, the environment is more complex in farms and the presence of others intermediate hosts, which are important for such parasites, could explain this.However, we need further evaluation.
We observed a higher number of parasites of the Trematoda class in doves captured at the soybean industry; this finding could be related to a higher occurrence of intermediate hosts at this site, but other studies must be conducted to confirm this hypothesis.According to Freitas (1979), gastropod mollusks are intermediate hosts in the life cycles of cestodes and trematodes.Sick (1997) affirmed that Z. auriculata feeds on mollusks and diplopods during the reproduction season, suggesting that these birds require the ingestion of animal protein and calcium during this period.
The majority of the animals infected with helminths in this study had single infections (37.00%);only 21% of the birds had multiple infections.The most found mixed infection was between A. columbae and O quadriradiatus.Lee et al. (2004) also reported this helminth association in Z. macroura.

Conclusions
This study is the first to describe the occurrence of A. columbae, T. fissispina, S. nasuta, R. allomyodes, P. bragai and P. confusa in Z. auriculata from Brazil.More studies should be conducted to determine whether these doves represent sources of infection for other birds because none of the parasites observed in this study were species specific.
16428-1 and by the Ethics Committee of Animal Experiments of the State University of Londrina no.70/2008.

N
= number of doves, Mh = microhabitats, P = prevalence, A = abundance, MI = mean intensity of parasitism, SI -small intestine, Pr = proventriculus, RT = renal tubules, OR = Odds Ratio, CI = Confidence Interval, NC = not calculated, p-values, a, b = same letter are not significantly, however, different ones are.

Table 1 .
Prevalence, abundance, mean intensity of parasitism, the range of parasitism, the total of helminths collected, Odds ratio and p-value in Zenaida auriculata in Londrina, 2013.

Table 2 .
Prevalence, abundance, mean intensity of parasitism, the range of parasitism, the total of helminthes collected, Odds ratio and p-value in male and female Zenaida auriculata in Londrina, 2013.

Table 3 .
Outcome of association between the local of capture and helminths found in Zenaida auriculata, Londrina, 2013.= Odds Ratio, CI = Confidence Interval, NC = not calculated, p-values, 1 Fisher's exact test, a, b = same letter are not significantly, however, different ones are. OR