SciELO - Scientific Electronic Library Online

 
vol.47 issue6Dengue in Brazil and Colombia: a study of knowledge, attitudes, and practicesSeroprevalence and risk factors associated with visceral leishmaniasis in dogs in Jaciara, State of Mato Grosso author indexsubject indexarticles search
Home Pagealphabetic serial listing  

Services on Demand

Journal

Article

Indicators

Related links

Share


Revista da Sociedade Brasileira de Medicina Tropical

Print version ISSN 0037-8682

Rev. Soc. Bras. Med. Trop. vol.47 no.6 Uberaba Nov./Dec. 2014

http://dx.doi.org/10.1590/0037-8682-0042-2014 

Short Communications

Outbreak of canine visceral leishmaniasis in Barra Mansa, State of Rio de Janeiro

Cintia Xavier de Mello [1]  

Fabiano Borges Figueiredo [2]  

Artur Augusto Velho Mendes Júnior [2]  

Marina Carvalho Furtado [2]  

Luciana Freitas Campos de Miranda [1]  

Maria de Fátima Madeira [1]  

[1]Laboratório de Vigilância em Leishmanioses, Instituto de Pesquisa Clínica Evandro Chagas, Fundação Oswaldo Cruz, Rio de Janeiro, RJ

[2]Laboratório de Pesquisa Clínica em Dermatozoonoses, Instituto de Pesquisa Clínica Evandro Chagas, Fundação Oswaldo Cruz, Rio de Janeiro, RJ

ABSTRACT

Introduction

In Brazil, visceral leishmaniasis (VL) has spread to various regions. This study reports canine cases of VL in Barra Mansa, where human VL cases were recently reported.

Methods

Using the human index case, a canine survey was performed by dual-path platform immunochromatography and enzyme-linked immunosorbent assay. Seropositive animals were euthanized. Cultures were collected to detect Leishmania parasites.

Results

Serological tests detected 141 canine VL cases, and Leishmania chagasi were isolated from 82.2% animals.

Conclusions

Leishmania chagasi is in circulation in Barra Mansa. This study broadens information on the parasite's distribution in the State of Rio de Janeiro.

Key words: Barra Mansa; Canine survey; Canine visceral leishmaniasis

Visceral leishmaniasis (VL) is of serious public health importance, and affects approximately 1-2 million people annually around the world1. Autochthonous transmission has been recorded in nearly all Brazilian states. The domestic dog is an important reservoir of infection, therefore constituting a primary target for disease control2.

Human cases are rare in the State of Rio de Janeiro, although canine cases have often been recorded over the years in municipalities such as Angra dos Reis, Mangaratiba3, Maricá4, and Volta Redonda5, and in different urban areas of the capital3,6. Canine VL cases historically precede spatial and temporal human cases7; therefore, the geographical expansion of canine cases in the State of Rio de Janeiro has put epidemiological surveillance authorities on alert8.

Barra Mansa is a municipality in the Southern region of the state where nine human cases of VL were recently reported9. Confirmation of these cases triggered a series of actions in Barra Mansa, in particular, the initiation of a canine survey directed at tracking the occurrence and extent of VL among domestic dogs. In this paper, our focus was to report the initial results of the canine survey in Barra Mansa during the years 2011-2013 and to identify Leishmania species in seroreactive animals.

Barra Mansa is a Brazilian municipality in the southern State of Rio de Janeiro. It is located at latitude 22°32′39″ S, longitude 44°10′17″ W, altitude 381m, and has an area of 547,226km2. The average annual temperature is 28°C with rainy summers and a relative humidity of 77%. The population is approximately 177,000 inhabitants. Barra Mansa borders the municipalities of Valença, Quatis, Rio Claro, Piraí, Bananal, Barra do Piraí, Resende, and Porto Real, and forms a conurbation with the Cities of Volta Redonda and Pinheral10.

The canine survey included domestic dogs living within a radius of approximately 100 meters of recorded humans VL cases. The animals were evaluated by a dual-path platform immunochromatographic test [DPP rapid test; Bio Manguinhos/Fundação Oswaldo Cruz (FIOCRUZ), Rio de Janeiro, Brazil], and DPP-positive animals were confirmed by an enzyme-linked immunosorbent assay [Ensaio imunoenzimático para diagnóstico da leishmaniose visceral canina (EIE-LVC); Bio Manguinhos/FIOCRUZ/Rio de Janeiro, Brazil] at the Central Laboratory Noel Nutels [Laboratório Central (LACEN), Rio de Janeiro, Brazil], as recommended by the Ministry of Health8. All animals confirmed as seropositive by enzyme-linked immunosorbent assay (ELISA) were collected and sent for euthanasia at the Laboratório de Pesquisa Clínica em Dermatozoonoses/FIOCRUZ, Rio de Janeiro, Brazil. Euthanasia followed the technical norms defined by Resolution no 714/02 from the Federal Council of Veterinary Medicine and by the screening-culling procedure adopted by the Brazilian Ministry of Health in its Program for Visceral Leishmaniasis Control2.

Before euthanasia, the animals were sedated with ketamine (10mg/kg) mixed with acepromazine (0.2mg/kg) and clinically classified as asymptomatic (i.e., dogs without clinical signs of VL upon physical examination), oligosymptomatic (i.e., dogs with 1-3 clinical signs), and symptomatic (i.e., dogs that had >3 clinical signs of VL such as emaciation, skin lesions, lethargy, lymph node enlargement, or signs of hepatomegaly or splenomegaly). Euthanasia was performed with an overdose of sodium thiopental 5%. Tissue samples (from liver, spleen, and intact skin fragments) and bone marrow and lymph node aspirates were collected and processed by culture for the isolation of parasites using the protocol described by Madeira et al.3. Tissue samples were first placed in sterile saline solution with antibiotics and an antifungal, and later transferred to a biphasic culture medium (Novy-MacNeal-Nicolle medium + Schneider medium supplemented with 10% fetal bovine serum) and incubated at 27°C ± 1°C, whereas the material obtained by puncture was immediately seeded in test tubes containing the same culture medium. Cultures were examined weekly for 30 days. When positive, they were reserved for etiological characterization by isoenzyme electrophoresis (i.e., multilocus enzyme electrophoresis) which employed five enzymatic systems: malic enzyme (ME, EC1.1.1.40), nucleosidase (NH, EC3.2.2.1), glucose-6-phosphate dehydrogenase (G6PDH, EC1.1.1.49), glucose phosphate isomerase (GPI, EC5.3.1.9), and 6-phosphogluconate dehydrogenase (6PGDH, EC1.1.1.43). Leishmania braziliensis (MHOM/BR/75/M2903) and Leishmania chagasi (MHOM/BR/74/PP75) were the reference samples in all experiments.

During the study period, 3,103 dogs were evaluated. Of these, 141 (4.5%) dogs were seropositive and hence euthanized. Promastigote forms were isolated from different sites in 116 (82.2%) euthanized animals (Table 1). Twenty-five euthanized dogs were culture-negative. Among these, 15 dogs were asymptomatic, nine dogs were oligosymptomatic, and one dog was symptomatic.

TABLE 1 Results of parasitological cultures using different clinical specimens from euthanized seropositive dogs. 

Clinical specimen Percentage (number of isolates/number of dogs) (CI:95%)
Lymph node fragment 73.8% (48/65; 84.5-63.0)
Spleen fragment 69.5% (98/141; 77.0-61.9)
Skin fragment 58.5% (38/65; 70.4-46.5)
Bone marrow aspirates 56.7% (80/141; 60.8-52.5)
Lymph node aspirates 52.3% (33/63; 64.6-40.0)

CI95%: 95% confidence interval.

At least one sample of the isolated organisms from each animal was processed by multilocus enzyme electrophoresis. Leishmania chagasi was confirmed in 100% (n = 116) of dogs.

The expansion of VL has been reported in various regions of Brazil, and the results of this study corroborate this. In 1977, in the State of Rio de Janeiro, the first human case of VL occurred in the municipality of Rio de Janeiro. From this case, the protocol of the canine survey was officially established in 1980 in this state6. At that time and in subsequent years, this survey was performed only in certain areas of the City of Rio de Janeiro. However, it has recently been extended to other regions of the state such as Barra Mansa because of the discovery of human and/or canine cases in these regions. The displacement of infected animals and the high adaptability of the vector have been the main reasons for the dispersal of VL in Brazil, and the State of Rio de Janeiro is no exception to this rule. Barra Mansa is bordered by the municipalities of Resende and Volta Redonda where VL cases have also recently been described5.

In this respect, a point that should be emphasized is the need for epidemiological surveillance in regions with favorable conditions for the establishment of VL. In Barra Mansa, the presence of L. chagasi has been confirmed, which places the region on alert.

Canine seropositivity may vary, depending on the epidemiological characteristics of the region. In Belo Horizonte, is approximately 7% which is an area of intense transmission11. In Barra Mansa, we reported a prevalence of 4.5%. It is interesting that by 2009 canine leishmaniasis seropositivity had already been reported at levels that reached 10% in Barra Mansa12. At that time, L. chagasi was not detected in the dogs studied; however, one dog was infected with Leishmania braziliensis. This suggests that canine visceral leishmaniasis (CVL) was perhaps already spreading silently in Barra Mansa, and led to an explosion in the number of cases such as those reported in this paper.

The limitation of serological testing is a major argument against euthanasia of seropositive dogs11. In our study, L. chagasi was confirmed by culture in 82% of the 141 euthanized seropositive dogs. This speaks in favor of the routine - DPP followed by enzyme-linked immunosorbent assay that is currently employed for the diagnosis of CVL in Brazil8. In 25 seropositive dogs, parasitological confirmation was not possible. This may be explained by the clinical condition of the animal at the time of euthanasia because 15 (60%) dogs were classified as asymptomatic animals. Studies have shown that the chance of parasite isolation is less in asymptomatic dogs than in symptomatic animals13.

Another point that should be emphasized is the need for laboratory confirmation, primarily in areas with no record of VL. Human cases have been reported in Barra Mansa; however, this is the first time that L. chagasi has been shown in VL cases in this region.

The culture is the standard reference tool for the diagnosis of leishmaniasis; however, this approach is not used in epidemiologic studies14,15.

The isolation of L. chagasi from the assessed dogs reinforces the involvement of this animal in the transmission cycle of VL in Barra Mansa. Despite reports of L. braziliensis in this region, this study did not detect it in any of the evaluated dogs. In the present study we evaluated a variety of clinical specimen, and the highest rates of positivity were in lymph node fragments, whereas interestingly the lowest rates were in lymph node aspirates. We believe this result may be because of variations in the volume of material collected by puncture. In recent years, leishmaniasis has been classified, along with other arthropod-borne diseases, as an emerging and re-emerging disease with a spread that can be associated with numerous factors. Barra Mansa, which was previously an area without recorded cases, now has an explosion of VL canine cases. The sequence of events demonstrated in this study suggests that CVL was possibly already initiated and spreading silently. The canine survey continues to be conducted in Barra Mansa, along with health education activities to professionals from different sectors that are aimed at containing the spread of VL in this region.

CONFLICT OF INTEREST

The authors declare that there is no conflict of interest.

FINANCIAL SUPPORT

This study was supported by the municipal prefecture of Barra Mansa and the Evandro Chagas National Institute of Infectology (Instituto Nacional de Infectologia Evandro Chagas)/Fundação Oswaldo Cruz (FIOCRUZ). Maria de Fátima Madeira and Fabiano Borges Figueiredo hold a grant from Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) for their productivity in the research.

ACKNOWLEDGMENTS

The authors would like to thank Evandro Chagas National Institute of Infectology (Instituto Nacional de Infectologia Evandro Chagas)/Fundação Oswaldo Cruz (FIOCRUZ) for the infrastructure to realize the research.

REFERENCES

. World Health Organization. Research Priorities for Chagas Disease, Human African Trypanosomiasis and Leishmaniasis. Geneva: World Health Organization; 2012. [ Links ]

. Ministério da Saúde. Manual de Vigilância da Leishmaniose Visceral Americana. Ministério da Saúde: Brasília; 2006. [ Links ]

. Madeira MF, Schubach AO, Schubach TMP, Pereira AS, Figueiredo FB, Baptista C, et al. Post mortem parasitological evaluation of dogs seroreactive for Leishmania from Rio de Janeiro, Brazil. Vet Parasitol 2006; 138:366-370. [ Links ]

. Paula CC, Figueiredo FB, Menezes RC, Mouta-Confort E, Bogio A, Madeira MF. Canine visceral leishmaniasis in Maricá, State of Rio de Janeiro: first report of an autochthonous case. Rev Soc Bras Med Trop 2009; 42:77-78. [ Links ]

. Campos MP, Silva DA, Madeira MF, Velho Jr AA, Figueiredo FB. First autochthonous case of canine visceral leishmaniasis in Volta Redonda, Rio de Janeiro, Brazil. Rev Bras Parasitol Vet 2013; 22:424-426. [ Links ]

. Marzochi MCA, Fagundes A, Andrade MV, Souza MB, Madeira MF, Mouta-Confort E, et al. Visceral leishmaniasis in Rio de Janeiro, Brazil: eco-epidemiological aspects and control. Rev Soc Bras Med Trop 2009; 42:570-580. [ Links ]

. Brazil RP. The dispersion of Lutzomyia longipalpis in urban áreas. Rev Soc Bras Med Trop 2013; 45:120-121. [ Links ]

. Coordenação geral de doenças transmissíveis. Coordenação geral de laboratórios de saúde publica. Nota técnica n° 01/2011. Rio de Janeiro; 2011. [ Links ]

. Pimentel MIF, Alves ELM, Silva MHFF, Moza PG, Mello CX, Schubach AO, et al. High lethality of visceral leishmaniasis in Barra Mansa, a new area of transmission in the state of Rio de Janeiro, Brazil. Rev Soc Bras Med Trop 2014; 47:521-523. [ Links ]

. Instituto Brasileiro de Geografia e Estatística (IBGE). Cidades@ [Internet]. Rio de Janeiro: IBGE. [Cited 2013 September 23]. Available at: http://www.ibge.gov.br/cidadesat/painel/painel.php?codmun=330040/. [ Links ]

. Romero GA, Boelaert M. Control of visceral leishmaniasis in Latin America a systematic review. PLoS Negl Trop Dis 2010; e584. [ Links ]

. Figueiredo FB, Bonna IC, Nascimento LD, Costa T, Baptista C, Pacheco TM, et al. Serological evaluation for detection of anti-Leishmania antibodies in dogs and cats in the district of Santa Rita de Cássia, municipality of Barra Mansa, State of Rio de Janeiro. Rev Soc Bras Med Trop 2009; 42:141-145. [ Links ]

. Almeida AB, Sousa VR, Gasparetto ND, Silva GF, Figueiredo FB, Dutra V, et al. Canine visceral leishmaniasis: diagnostic approaches based on polymerase chain reaction employing different biological samples. Diagn Microbiol Infect Dis 2013; 76:321-324. [ Links ]

. Madeira MF, Figueiredo FB, Pinto AG, Nascimento LD, Furtado M, Mouta-Confort E, et al. Parasitological diagnosis of canine visceral leishmaniasis: is intact skin a good target? Vet Sci 2009; 87:260-262. [ Links ]

. Almeida AB, Sousa VR, Sorte EC, Figueiredo FB, Paula DA, Pimentel MF, et al. Use of parasitological culture to detect Leishmania (Leishmania) chagasi in naturally infected dogs. Vector Borne Zoonotic Dis 2011; 11:1555-1560. [ Links ]

Received: February 22, 2014; Accepted: May 29, 2014

Address to: Dra. Maria de Fátima Madeira. Lab Vigileish/IPEC/FIOCRUZ. Av. Brasil 4365, Manguinhos, 21040-360 Rio de Janeiro, RJ, Brasil. Phone/Fax: 55 21 3865-9541. e-mail: fatima.madeira@ipec.fiocruz.br

Creative Commons License This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License, which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.