Canine bartonellosis: serological and molecular prevalence in Brazil and evidence of co-infection with Bartonella henselae and Bartonella vinsonii subsp. berkhoffii Pedro Paulo Vissotto de Paiva Diniz, Ricardo Maggi, Denise Schwartz, María Cadenas, Julie Bradley, Barbara Hegarty, Edward Breitschwerdt To cite this version: Pedro Paulo Vissotto de Paiva Diniz, Ricardo Maggi, Denise Schwartz, María Cadenas, Julie Bradley, et al.. Canine bartonellosis: serological and molecular prevalence in Brazil and evidence of co-infection with Bartonella henselae and Bartonella vinsonii subsp. berkhoffii. Veterinary Research, BioMed Central, 2007, 38 (5), pp.697-710. 10.1051/vetres:2007023. hal-00902882 HAL Id: hal-00902882 https://hal.archives-ouvertes.fr/hal-00902882 Submitted on 1 Jan 2007 HAL is a multi-disciplinary open access L’archive ouverte pluridisciplinaire HAL, est archive for the deposit and dissemination of sci- destinée au dépôt et à la diffusion de documents entific research documents, whether they are pub- scientifiques de niveau recherche, publiés ou non, lished or not. The documents may come from émanant des établissements d’enseignement et de teaching and research institutions in France or recherche français ou étrangers, des laboratoires abroad, or from public or private research centers. publics ou privés. Vet. Res. 38 (2007) 697–710 Available online at: c INRA, EDP Sciences, 2007 www.vetres.org DOI: 10.1051/vetres:2007023 Original article Canine bartonellosis: serological and molecular prevalence in Brazil and evidence of co-infection with Bartonella henselae and Bartonella vinsonii subsp. berkhoffii Pedro Paulo Vissotto De Paiva Da, Ricardo Guillermo Ma, Denise Saretta Sb, María Belén Ca, Julie Meredith Ba,BarbaraHa, Edward Bealmear Ba* a Intracellular Pathogens Research Laboratory, Center for Comparative Medicine and Translational Research, Department of Clinical Sciences, College of Veterinary Medicine, North Carolina State University, 4700 Hillsborough Street, Raleigh, NC, 27606, USA b School of Veterinary Medicine and Animal Science, São Paulo State University (FMVZ – UNESP), Botucatu, São Paulo, Brazil (Received 17 November 2006; accepted 4 April 2007) Abstract – The purpose of this study was to determine the serological and molecular prevalence of Bartonella spp. infection in a sick dog population from Brazil. At the São Paulo State University Veterinary Teaching Hospital in Botucatu, 198 consecutive dogs with clinicopathological abnor- malities consistent with tick-borne infections were sampled. Antibodies to Bartonella henselae and Bartonella vinsonii subsp. berkhoffii were detected in 2.0% (4/197) and 1.5% (3/197) of the dogs, respectively. Using 16S-23S rRNA intergenic transcribed spacer (ITS) primers, Bartonella DNA was amplified from only 1/198 blood samples. Bartonella seroreactive and/or PCR positive blood samples (n = 8) were inoculated into a liquid pre-enrichment growth medium (BAPGM) and sub- sequently sub-inoculated onto BAPGM/blood-agar plates. PCR targeting the ITS region, pap31 and rpoB genes amplified B. henselae from the blood and/or isolates of the PCR positive dog (ITS: DQ346666; pap31 gene: DQ351240; rpoB: EF196806). B. henselae and B. vinsonii subsp. berkhof- fii (pap31: DQ906160; rpoB: EF196805) co-infection was found in one of the B. vinsonii subsp. berkhoffii seroreactive dogs. We conclude that dogs in this study population were infrequently ex- posed to or infected with a Bartonella species. The B. henselae and B. vinsonii subsp. berkhoffii strains identified in this study are genetically similar to strains isolated from septicemic cats, dogs, coyotes and human beings from other parts of the world. To our knowledge, these isolates provide the first Brazilian DNA sequences from these Bartonella species and the first evidence of Bartonella co-infection in dogs. dogs / Bartonella infections / heart disease / culture / Brazil 1. INTRODUCTION teremia in mammals and are transmit- ted by animal bites and scratches or by Bartonella are fastidious hemotropic many different vectors including sand- bacteria, which cause long-lasting bac- flies, body lice, fleas and potentially ticks. The genus Bartonella contains nu- * Corresponding author: [email protected] merous recently described species, many Article available at http://www.vetres.org or http://dx.doi.org/10.1051/vetres:2007023 698 P.P.V.P. Diniz et al. of which are new and emerging human more prevalent than was previously recog- pathogens [8, 12, 16, 24]. nized. In dogs, infection with five Bartonella The objective of this study was to deter- species has been described: B. clarridgeiae mine the serological and molecular preva- and B. vinsonii subsp. berkhoffii (here- lence of Bartonella infection in a subset of after B. v. berkhoffii) DNA was detected the sick dog population examined at a Vet- in aortic valves of six dogs with vegeta- erinary Teaching Hospital in southeastern tive endocarditis, based upon independent Brazil. Successful culture and molecular studies [9,14,32]; B. elizabethae DNA was characterization of B. henselae and B. v. detected in a dog with weight loss and berkhoffii from two clinically ill dogs is sudden death [37]; B. henselae DNA was also reported, including the first evidence sequenced from a dog with peliosis hepatis of simultaneous infection with these two and from a dog with granulomatous hep- organisms in the same host. atitis [22]; and B. washoensis was isolated from a dog with mitral valve endocardi- tis [15]. A potentially new species closely 2. MATERIALS AND METHODS related to B. clarridgeiae (proposed name 2.1. Study population B. rochalimaea) has been isolated from three dogs and from 22 gray foxes in One hundred and ninety-eight EDTA- 1 California [32] . anticoagulated blood and serum samples To date, no studies have reported sero- from dogs with historical, clinical and/or logical or molecular evidence of Bar- laboratory findings consistent with tick- tonella infection in dogs from Brazil or borne diseases were prospectively se- other South American countries. With the lected at a Veterinary Teaching Hospi- notable exception of B. bacilliformis, there tal at the College of Veterinary Medicine are only a limited number of reports and Animal Husbandry (FMVZ – Un- describing human infection with other esp) in Botucatu, southeast Brazil (latitude Bartonella species in South America. In 22◦ 53 09 S, longitude 48◦ 26 42 W) Brazil, B. quintana was detected by PCR between October 2002 and November in an infant [3] and two fatal cases of 2003. Dogs included in this study had endocarditis were associated with high B. at least three of the following clini- henselae antibody titers [41]. In contrast to cal or laboratory criteria: presence of humans, B. henselae seroreactivity was re- tick infestation at the time of examina- 2 ported in 46% of 102 cats suggesting that tion, bleeding disorders (epistaxis, me- exposure to this organism in Brazil may be lena, ecchymosis, bleeding gums), neu- rological signs, inflammatory ocular dis- 1 Henn J.B., Koehler J.E., Gabriel M., Kasten ease (uveitis, chorioretinitis, hyphema), R.W., Brown R.N., Papageorgiou S., Chomel fever (rectal temperature > 39.4 ◦C), ane- B.B., Zoonotic Bartonella spp. in domestic mia (PCV < 35%), leukopenia (WBC dogs and gray foxes from California, in: Proc. < /µ 20th Meeting of the American Society for Rick- 6 000 cells L), thrombocytopenia < /µ ettsiology and 5th International Conference (platelets 150 000 cells L) and hyper- > / on Bartonella as Emerging Pathogens, Pacific proteinemia (total protein 7.8 g dL). Grove, 2006, pp. 70. Dogs treated with tetracycline or imi- 2 Slhessarenko N., Camargo M.C.G.O., docarb dipropionate during the previous D’Auria S.R.N., Soroprevalência de Bartonella 30 days were excluded from the study pop- henselae em gatos do municipio de São Paulo, ulation. This study was approved by the Rev. Soc. Bras. Med. Trop. (1996) 29:104 Medical Ethical and Animal Care Com- (Abstract). mittee of São Paulo State University. In Bartonella species in Brazilian dogs 699 addition, each owner authorized blood col- blood pressure was normal; however, lection and enrollment of the dog into the echocardiography identified an increased study. left ventricle wall (14.5 mm) thickness From the selected dogs, two had resulting in reduction of left ventricular Bartonella species detected by culture chamber size (33 mm) during diastole. All (techniques described below). Dog #1 other infectious disease diagnostic tests was a 6.3 kg 2-year-old male Poodle described above for Dog #1 were also that was examined because of anorexia, negative for Dog #2. bleeding gums, cutaneous petechiae, and ecchymoses. The dog was febrile (39.7 ◦C) and infested with ticks. Hematological 2.2. Serology-based assays abnormalities included severe anemia The microimmunofluorescence test (PCV = 14%; normal 39%), leukope- (IFA) for the detection of anti-B. v. nia (2 692/µL; normal 6 000/µL), berkhoffii (isolate 93-CO-1, ATCC thrombocytopenia (2 500/µL; normal #51672) and anti-B. henselae (strain 190 000/µL), and hypoproteinemia Houston-1, ATCC #49882) antibodies (4.6 g/dL; normal 6.1 g/dL). Cardiovas- in canine sera was performed as de- cular abnormalities included hypotension scribed [30]. The starting dilution was (systolic blood pressure = 75 mmHg; at 1:16 and the cut-off for seroreactivity normal 90 mmHg), myocardial dysfunc- was defined at 1:64. Endpoint titers were tion (fractional shortening = 24%; normal determined as the last dilution at which 30%) and increased serum cTnI concen-