Pesq. Vet. Bras. 35(5):462-465, maio 2014
Serosurvey of Leptospira interrogans, Brucella abortus and Chlamydophila abortus infection in free-ranging giant anteaters (Myrmecophaga tridactyla) from Brazil1
Flávia R. Miranda2*, Mariella Superina3, Fernanda Vinci4, Vanessa Hashimoto5, Julio Cesar Freitas4 and Eliana Reiko Matushima6
ABSTRACT.- Miranda F.R., Superina M., Vinci F., Hashimoto V., Freitas J.C. & Matushima E.R. 2015. Serosurvey of Leptospira interrogans, Brucella abortus and Chlamydophila abor- tus infection in free-ranging giant anteaters (Myrmecophaga tridactyla) from Brazil. Pesquisa Veterinária Brasileira 35(5):462-465. Projeto Tamanduá, Rua Vergueiro 3432, apto 171, Vila Mariana, São Paulo, SP 07600-000, Brazil. E-mail: A serological survey for antibodies against Leptospira interrogans, Brucella abortus, and Chlamydophila abortus was conducted in 21 clinically [email protected], free-ranging giant ant- eaters (Myrmecophaga tridactyla) from Parque Nacional das Emas (Goiás State, Brazil; n=6), Parque Nacional da Serra da Canastra (Minas Gerais State, Brazil; n=9), and RPPN SESC Pantanal (Mato Grosso State, Brazil; n=6) between July 2001 and September 2006. Sera were screened for antibodies against 22 serovars of Leptospira interrogans with a microscopic agglutination test. Twelve tested positive for L. interrogans serovars sentot (n=5 in PN Emas, n=2 in PN Serra da Canastra), butembo (n=2 in PN Serra da Canastra), autumnalis, bataviae, and shermani/icterohaemorrhagiae (n=1 each in SESC Pantanal). One adult female tested positive for B. abortus with the buffered plate antigen test. All sera were negative for C. abortus - port of pathogens that may interfere with the reproduction and population dynamics of free-ranging giant anteaters. using the complement fixation text. This is the first re INDEX TERMS: Anteaters, Myrmecophaga tridactyla, brucellosis, Brucella abortus, chla- mydiosis, Chlamydophila abortus, leptospirosis, Leptospira interrogans, Pilosa, serological survey, Vermilingua, wildlife, Xenarthra.
RESUMO.- [Inquérito sorológico de Leptospira inter- tamanduás-bandeira (Myrmecophaga tridactyla) de vida rogans, Brucella abortus e Chlamydophila abortus em livre do Parque Nacional das Emas (Goiás, Brasil, n=6), o tamanduás-bandeira (Myrmecophaga tridactyla) de Parque Nacional da Serra da Canastra (Minas Gerais, Bra- vida livre no Brasil.] Inquéritos sorológicos para detec- sil, n=9) e RPPN SESC Pantanal (Mato Grosso, Brasil, n=6) ção de anticorpos contra Leptospira interrogans, Brucella entre julho de 2001 e setembro de 2006. Os soros foram abortus, e Chlamydophila abortus foram realizados em 21 testados para anticorpos contra 22 sorotipos de Leptospi- ra interrogans com um teste de aglutinação microscópica. 1 Received on June 27, 2013. Doze animais foram considerados positivos para L. interro- Accepted for publication on December 17, 2014. gans sorovares sentot (n=5 em PN Emas, n=2 em PN Serra 2 Projeto Tamanduá, Rua Vergueiro 3432, apto 171, Vila Mariana, São da Canastra), butembo (n=2 em PN Serra da Canastra), au- Paulo, SP 07600-000, Brazil. *Corresponding author: tumnalis, bataviae e shermani/icterohaemorrhagiae (n=1 3 Laboratorio de Endocrinología de la Fauna Silvestre, IMBECU, CCT-CO- NICET Mendoza, Casilla de Correos 855, 5500 Mendoza,[email protected] Argentina. para cada sorovar em SESC Pantanal). Uma fêmea adulta 4 SHIN QI 7, conjunto 12, casa 18, Brasília, DF 031310-050, Brazil. testou positivo para B. abortus com o teste do antígeno 5 Departamento de Microbiologia, Centro de Ciências Biológicas, Univer- tamponado. Todos os soros se mostraram negativos para C. abortus 980, Brazil. Este é o primeiro relato de patógenos que podem interfe- sidade6 Departamento Estadual de de Londrina Patologia, (UEL), Faculdade Cx. Postal de Medicina6001, Londrina, Veterinária PR 86051- e Zoo- tecnia, Universidade de São Paulo (USP), Av. Prof. Dr. Orlando Marques de rir na dinâmica através reprodutiva do teste de populações fixação do complemento.de tamanduás Paiva 87, São Paulo, SP 05508-000, Brazil. em estado selvagem.
462 Serosurvey of Leptospira interrogans, Brucella abortus and Chlamydophila abortus infection in free-ranging giant anteaters 463
TERMOS DE INDEXAÇÃO: Tamanduás, Myrmecophaga tridactyla, Permits to capture and sample wild giant anteaters were brucelose, Brucella abortus, chlamydiose, Chlamydophila abortus, granted by the Brazilian Institute of Environment and Renewable leptospirose, Leptospira interrogans, Pilosa, inquérito sorológico, Natural Resources (IBAMA). Vermilingua, animais silvestres, Xenarthra. Sample analysis. All analyses were performed at the Departa- mento de Medicina Preventiva, Universidade Estadual de Londri- INTRODUCTION na, Brazil. A microscopic agglutination test (MAT) with live patho- gen was used to detect antibodies against Leptospira interrogans Diseases have been recognized as a growing problem in (Cole et al. 1973). The following reference serovars were used: biodiversity conservation (McCallum & Dobson 1995). Due australis, bratislava, autumnalis, butembo, bataviae, canicola, cas- to the paucity of information on pathogens affecting wild tellonis, copenhageni, cynopteri, fortbragg, grippotyphosa, hardjo, giant anteaters (Myrmecophaga tridactyla), it is currently hebdomadis, icterohaemorrhagiae, panama, pomona, pyrogenes, - sentot, shermani, tarassovi, whitcomb, and wolffi. Sera were con- M. tridactyla as Vulnerabledifficult to assessby the theIUCN role Red of List diseases of Threatened in the population Species (Mi de- Positive serum samples were serially diluted to determine the end randaclines that& Medri have 2010). led to Giantthe classification anteaters have of small litter si- pointsidered (Myers positive 1985). if agglutination If a sample showed was present cross-reactivity at dilutions between ≥1:10. two or more serovars of the same group, the one showing higher (Schauerte 2005). Pathogens that may interfere with their titers was considered positive. The Buffered Plate Antigen test (BPA, Angus & Barton 1984) zes, long gestations, and extended periods of parental care was used to screen sera for antibodies against B. abortus. A 0.03 ml suspension of deactivated B. abortus cells was homogenized andability bear to theproduce potential viable to offspringdrive this arecharismatic therefore species expected to with the same volume of serum. Samples showing macroscopic to have significant impacts on their population growth rate agglutination after four minutes were considered reactive. Leptospirosis, brucellosis, and chlamydiosis are impor- Sera were tested for antibodies against C. abortus using the tantextinction zoonoses (Boots with & Sasakiworldwide 2001). distribution that can lead C. abortus to reduced fertility and abortion (Acha & Szyfres 2003). In - Brazil, the causative agents of these diseases have been re- complementrimentale delle fixation Venezie, test Padova, (OIE 2008). Italy) Referencewas used as strain antigen. Titers ported in domestic (Tomich et al. 2007) and wild animals S26/3 (provided by Dr Carlo Turilli; Instituto Zooprofilattico Spe (Ito et al. 1998) sharing their habitat with giant anteaters. ≥1:32 were considered positive (OIE 2008). Contact between giant anteaters and infected animals may RESULTS lead to disease transmission, potentially affecting reproduc- tive rates and leading to population declines in M. tridactyla. males, 4 females), nine at PNSC (4 males, 5 females), and The objective of this study was to investigate the pre- A total of 21 giant anteaters were captured: six at PNE (2- sence of antibodies against Leptospira interrogans, Brucella dered clinically healthy. Twelve sera (57.14%) were posi- abortus, and Chlamydophila abortus in wild giant anteaters tivesix atfor SESC Leptospira (3 males, interrogans 3 females)., one All (4.76%) animals tested were positive consi inhabiting three protected areas of central Brazil. for B. abortus, and all were negative for antibodies against Chlamydophila abortus (Table 1). MATERIALS AND METHODS The large majority of anteaters with antibodies against Study area. Study animals were wild-caught in three diffe- L. interrogans had low agglutinating titers, but two animals from PNE had titers >1:100 for L. interrogans serovar sentot - (Fig.1). Determination of the more probable serovar was not zil,rent which localities protects of Brazil: 131,800 Parque ha of Nacional the Cerrado das Emas biome (PNE; in southwes 18˚16’S,- possible in one serum sample from SESC because co-aggluti- tern52˚53’W), Goiás locatedState (Scardua in the Brazilian et al. 2004); Central Parque Plateau Nacional of western da Serra Bra nation occurred and both serovars (shermani and icteroha- - emmorhagiae - Serovar sentot da Canastra (PNSC; 20˚00’-20˚30’S, 46˚15’-47˚00’W), which co tern Minas Gerais State (IBAMA 2005); and Reserva Particular do from two protected) were positiveareas. Antibodies at the same against maximum serovar dilution. vers approximately 200,000 ha of the Cerrado biome in southwes bu- tembo were restrictedwas to identified anteaters in from seven PNSC, giant and anteaters serovars locatedPatrimônio within Natural the Pantanal SESC Pantanal Biosphere (SESC; Reserve, 16–17˚S, in Mato56–57˚W), Grosso a autumnalis, bataviae, shermani, and icterohaemmorhagiae Stateprivately (Brandão owned et conservational. 2011). unit of approximately 88,000 ha were only found in animals from SESC (Fig.1). Sample collection. Line transects were performed in July 2001 and January to April 2003 (PNE); August 2003 (PNSC); Table 1. Number of seropositive giant anteaters and from July to September 2005 and June to September 2006 (Myrmecophaga tridactyla) per number of individuals (SESC). Anteaters were darted with a blowpipe using 10mg/kg tested for three pathogens at three protected areas of Brazil
a with 1mg/kg midazolam (Dormonid, Roche, São Paulo, Brazil) or Study site Disease agents ketamine chloride (Ketalar, Laboratorios Pfizer, São Paulo, Brazil) Leptospira Brucella Chlamydophila - interrogans abortus abortus ded,9.56mg/kg and age ketamine was determined chloride based with 1.6mg/kgon body mass xylazine and size.(Virbacyl, Mor- PNE 5/6 0/6 0/6 Virbac, São Paulo, Brazil). Sex and geographic location were recor PNSC 4/9 1/9 0/9 performed. Blood was collected into sterile test tubes by punctu- SESC 3/6 0/6 0/6 phometric measurements were taken and a clinical examination- Total 12/21 1/21 0/21 ner femoral vein. Serum was separated with a portable centrifuge, a PNE = Parque Nacional das Emas; PNSC = Parque Nacional da Serra da Ca- thenring the aliquoted external in jugular Eppendorf vein tubesor, less and frequently, stored in the liquid cephalic nitrogen. or in nastra; SESC = Reserva Particular do Patrimônio Natural SESC Pantanal.
Pesq. Vet. Bras. 35(5):462-465, maio 2014 464 Flávia R. Miranda et al.
eater. It is, however, also possible that the seropositive fe- male acquired the pathogen from a wild reservoir host. Gonçalves et al. (2009) noted that the presence of wild cer-
infected with B. abortus. Nevertheless, the role of cervids vids significantly increased the risk of cattle herds being prevalence of Brucella infection in cervids from the PNSC areaas reservoirs are lacking. requires confirmation because data on the None of the individuals from SESC and PNE was seropo- sitive for B. abortus. It should, however, be noted that only a few anteaters were tested in these areas. Prevalence of bovine brucellosis is very high in the vicinity of PNE (21% seropositive cattle herds, Rocha et al. 2009) and of SESC (37% seropositive herds, Negreiros et al. 2009), and cattle ranching occurs in private farms near both protected areas. Antibodies against B. abortus have also been found in pec- caries (Tayassu pecari) in the vicinity of PNE (Kashivakura Fig.1. Percentages of antibody against serovars of Leptospira in- et al. 2004) and in pampas deer (Ozotoceros bezoarticus) terrogans in free-ranging giant anteaters (Myrmecophaga tri- dactyla) from three protected areas of Brazil infrom PNE the and southern SESC. ItPantanal remains (Elisei to be etstudied al. 2010). whether Wild infecgiant- DISCUSSION tionanteaters with B.may abortus therefore leads also to clinical be exposed symptoms to this in giantpathogen ant- eaters. Considering that only one out of 21 assessed ant- the reproduction and population dynamics of free-ranging - giantThis is anteaters. the first report of pathogens that may interfere with sure related to high prevalences in cattle and other wildlife, Twelve of 21 tested giant anteaters were seroreactive to iteaters is also was possible seropositive that, similar in spite to of moose the elevated (Alces alcesrisk of, Forbes expo Leptospira interrogans, over 50% of which were positive for et al. 1996), Myrmecophaga tridactyla is highly susceptible serovar sentot. Epidemiological data on the occurrence of to the disease and dies soon after infection. this serovar in Brazil are scarce, as it is not always included None of the assessed giant anteaters was positive for as a reference strain (e.g., Tomich et al. 2007; Freitas et al. Chlamydophila abortus. This pathogen of worldwide distri- 2010). It is possible that the wide variety of serovars obser- bution is one of the most frequent causes of abortion in sheep ved in anteaters from SESC (Fig.1), as well as in domestic and goats (Longbottom & Coulter 2003). Data on its occur- animals and other wildlife from this area (Freitas et al. 2010; Jorge et al. 2011), is related to the area having favorable conditions for the survival of Leptospira in the environment, Sãorence Paulo in Brazilian State that wildlife recently are tested virtually positive non-existent, for Chlamydo with- - philathe notable spp. (Miranda, exception unpublished of a giant anteater data). Accordingkept in a zooto itsin dity levels (Acha & Szyfres 2003). Emmons et al. (2004) case history, the nulliparous female suffered spontaneous repeatedlysuch as seasonal observed flooding giant andanteaters high temperature wallowing in and mud humi and - bathing in shallow ponds that are used by other species as tom typical for infection with C. abortus in small ruminants watering place. This behavior may further increase the risk abortion in the final third of its first pregnancy, a symp- of anteaters becoming infected with L. interrogans. posure to this pathogen can interfere with reproduction in No clinical signs of leptospirosis were observed in any giant(Longbottom anteaters. & Coulter 2003). This case implies that ex To our knowledge, no epidemiological studies on C. wild animals rarely show symptoms or lesions attributable abortus have been performed in domestic or wild animals toof Leptospira the examined (Acha anteaters. & Szyfres This 2003) was. A not captive unexpected, giant ant- as within or near our study sites, which complicates the as- eater at Rio de Janeiro Zoo, however, showed clinical signs - due to L. interrogans serovar icterohaemmorhagiae infec- tion (Monteiro et al. 2003). identifysessment C. ofabortus the exposure risk of the giant anteaters stu A single giant anteater from PNSC tested positive for itsdied prevalence here. Only in few this laboratories country and, in especially, Brazil are at qualified our study to Brucella abortus. The adult female was clinically heal- sites (Silva et al. 2006)., which Additional may explain screenings the limited of giant data ant- on thy, but it is not known whether its fertility was affected eaters inhabiting areas where presence of C. abortus has important areas of cattle ranching, where roughly 6% needed to assess the importance of this pathogen in their ofby herds the pathogen. are infected PNSC with lies bovine within brucellosis one of Brazil’s (Gonçalves most reproductionbeen confirmed and in population domestic ordynamics. other wild animals will be vicinity of PNSC and on private properties within the na- Acknowledgments.- et al. 2009). Extensive cattle ranching occurs both in the tional park (IBAMA 2005). This situation bears the risk Roberto Aguilar for hisWe guidance. are thankful Financial to all support the persons was provided who helped by the us of close contact between domestic and wild animals, and with fieldwork and laboratory analyses . Special thanks are extended to cattle may have been the source of infection for this ant- Conservation Society. IUCN/SSC Anteater, Sloth and Armadillo Specialist Group and Wildlife
Pesq. Vet. Bras. 35(5):462-465, maio 2014 Serosurvey of Leptospira interrogans, Brucella abortus and Chlamydophila abortus infection in free-ranging giant anteaters 465
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