Parasite Immunology, 2008, 30, 515–524 DOI: 10.1111/j.1365-3024.2008.01051.x SBOIplRaecInkGniwcINeclhAaPLnugbAeliRsﬂhnTiIvnCigscLeErtadSllaeishmaniamsismation and structural changes of splenic lymphoid tissue in visceral leishmaniasis: A study on naturally infected dogs C. C. SANTANA,1 J. VASSALLO,2 L. A. R. DE FREITAS,1 G. G. S. OLIVEIRA,1 L. C. PONTES-DE-CARVALHO1 & W. L. C. DOS-SANTOS1 1Centro de Pesquisas Gonçalo Moniz, Fundação Oswaldo Cruz, Fundação Oswaldo Cruz, Candeal, Salvador, Bahia, Brazil, 2Faculdade de Ciências Médicas, Universidade de Campinas, Cidade Universitária ‘Zeferino Vaz’, Campinas, São Paulo, Brazil SUMMARY The aim of this study was to identify splenic immuno-inﬂammatory patterns associated with natural infection by Leishmania chagasi. Spleen samples were obtained from 72 stray dogs from an endemic area of visceral leishmaniasis. The animals were grouped into four categories as follows: (i) potentially resistant to visceral leishmaniasis, with a positive leishmanin skin test result, and negative splenic culture for Leishmania parasites (ii) potentially susceptible to visceral leishmaniasis, with a negative leishmanin skin test and positive splenic culture for Leishmania (iii) infected with undeﬁned susceptibility status, with a positive leishmanin skin test and positive splenic culture for Leishmania, and (iv) noninfected, with a negative leishmanin skin test, negative splenic culture for Leishmania, and negative serology for anti-Leishmania antibodies. Histopathological analyses showed that there was a higher frequency of perisplenitis (18/25, P < 0·0001), granuloma (7/25, P = 0·0102), structural disorganization (14/25, P < 0·0001), and atrophy of the lymphoid follicles (20/25, P = 0·0036) and of the marginal zone (15/25, P = 0·0025) in the potentially susceptible group than in the other groups. The data presented here show changes in the white pulp of the spleen that are associated with naturally acquired visceral leishmaniasis. Keywords canine visceral leishmaniasis, Leishmania chagasi, Leishmania infantum, lymphoid tissue, Spleen Correspondence: WLC Dos-Santos Washington LC dos-Santos, Centro de Pesquisas Gonçalo Moniz-FIOCRUZ, Rua Waldemar Falcão 121, Candeal, 40·296–710, Salvador, Bahia, Brazil (e-mail: wluis@bahia.ﬁocruz.br). Re-use of this article is permitted in accordance with the Creative Commons Deed, Attribution 2·5, which does not permit commercial exploitation. Received: 10 March 2008 Accepted for publication: 28 May 2008 © 2008 The Authors Journal compilation © 2008 Blackwell Publishing Ltd INTRODUCTION Visceral leishmaniasis is used to describe two clinically, parasitologically and epidemiologically different diseases: antroponotic visceral leishmaniasis, caused by Leishmania donovani, and zoonotic visceral leishmaniasis (VL), caused by L. chagasi/L. infantum. The latter disease is endemic in Mediterranean countries and the American continent, and its causal agents are transmitted to mammalian hosts by sand ﬂies of the Phlebotomus and Lutzomyia genera (1). Leishmania chagasi/L. infantum infection does not always lead to VL (2). Most infected humans (2,3) and at least a proportion of infected dogs (4,5) remain asymptomatic or develop a mild disease, which is spontaneously cured. Occasionally, dogs and humans develop a severe form of VL (2,6) that is usually lethal if left untreated. In this report, we use the term VL to refer to this severe form of the disease. In both humans and dogs, the disease proceeds with emaciation, enlargement of the liver and spleen, fever, anaemia, and an increased predisposition to bacterial infection (2,7–10). In dogs, such signs of disease are further accompanied by a variety of skin and ocular lesions (9). Much that is known about the immune response to Leishmania parasites, and even the immune response in general has come from studies in murine models of leishmaniases (11–13). Most of these models, however, are intrinsically compromised by the artiﬁcial nature of the infective inocula. Instead of consisting only of naturally infective, metacyclic promastigotes, these inocula usually contain (i) either relatively large number of complement- ﬁxing, noninfective procyclic promastigotes (usually constitut- ing more than 70% of the inoculum) (14) or (ii) amastigotes, which are not the usual infectious form of the parasite. Canine VL, arising from natural infection by L. chagasi, is a disease worthy not only of study in its own right for veterinary and epidemiological reasons (15), but also for consideration as an attractive animal model for the human disease, because 515