Original articleAssessing the cellular and humoral immune response in Rhipicephalus sanguineus sensu lato (Acari: Ixodidae) infected with Leishmania infantum (Nicolle, 1908)
Introduction
Ticks are obligate ectoparasites that may parasitize domestic or wild animals and occasionally humans. They cause harm to the host via blood loss or transmission of a variety of pathogens, including bacteria, viruses and protozoa (Jongejan and Uilenberg, 2004; de la Fuente et al., 2008). The success of pathogen transmission by ticks is related to specific aspects of its biology, such as intracellular blood digestion by way of prevention of the action of digestive enzymes in the intestinal lumen. Moreover, the survival of ingested microorganisms seems to be associated with proteins present in the tick and maintenance of the redox balance in its intestine (Griesch et al., 2000; Sonenshine and Hynes, 2008). The redox balance is maintained by antioxidant enzymes such as catalase, glutathione peroxidase and thioredoxin, S-transferase glutathione and selenoproteins being expressed to destroy free radicals produced in the intestine (Anderson et al., 2008; Megy et al., 2012). Ticks have a well-developed innate immune system. However, despite their capacity to resist infection by various microorganisms, they remain important vectors of numerous causative agents of diseases, which can evade the tick’s immune system. Knowledge of these pathogen evasion mechanisms in ticks is still scant (Sonenshine and Hynes, 2008; Kopacek et al., 2010).
Most knowledge of the innate immunity of ticks is associated with cellular and humoral immune responses in the hemocoel tick. The innate immune response involves two main components. The first is cellular defense, in the form of hemocytes, which play an important role in the innate immune response of ticks when exposed to any pathogen or tissue damage. The hemocytes are also directly related to phagocytosis and encapsulation processes or lumps intended mainly for major pathogens such as protozoa and metazoa. The humoral response involves the production of antimicrobial peptides (AMPs), which act mainly against bacteria and fungi. The AMPs may be expressed by the hemocytes, body fat, the midgut, and in some cases other internal organs of the tick. Other peptides, not limited to antimicrobials, such as lysozyme, lectins, protease inhibitors, are also expressed in response to pathogen challenge (Griesch et al., 2000; Shi and Paskewitz, 2006; Sonenshine and Hynes, 2008).
Rhipicephalus sanguineus sensu lato (Acari: Ixodidae) is known as the brown dog tick and is an ectoparasite of domestic dogs found worldwide, mainly in tropical and temperate climates (Dantas-Torres, 2008). This tick species is the vector and host of pathogens such as the Rickettsia conorii bacterium and the Babesia vogeli and Hepatozoon canis protozoa (Parola et al., 2008; Baneth, 2011; Schnittger et al., 2012). However, little is known about the parasite-vector interaction between these pathogens and R. sanguineus s.l. Some studies have investigated the role of the possible relationship between R. sanguineus s.l. and Leishmania infantum (Coutinho et al., 2005; Paz et al., 2010; Dantas-Torres et al., 2010; Dantas-Torres and Otranto, 2011; Colombo et al., 2011; Solano-Gallego et al., 2012; de Morais et al., 2013). However, studies emphasize the presence of parasite DNA in ticks, necessitating further clarification regarding the vector-parasite interaction. The aim of this study was to evaluate aspects of the cellular and humoral innate immune response by evaluating hemocyte dynamics, phagocytosis, phenoloxidase activity and nitric oxide production in R. sanguineus s.l. challenged by L. infantum.
Section snippets
Tick and hemolymph collection
Ticks of the species R. sanguineus s.l., obtained from the Keizo Asami Immunopathology Laboratory - LIKA / UFPE, Recife strain, were used for the study. Rabbits, Oryctolagus cuniculus of New Zealand strain, were used to feed the colony. The R. sanguineus s.l. colony was maintained at 25 ± 3 °C and 85 ± 10% relative humidity (RH). The study used only adult females of R. sanguineus s.l. The hemolymph was collected by cutting the front legs of the coxal trochanter joint using siliconized glass
DNA quantify of L. infantum in R. sanguineus s.l. and hemocyte dynamics
The present study showed a statistically significant increase (p < 0.05) in the 18S ribosomal cDNA copies of L. infantum on days 1, 5 and 7 post-infection with L. infantum compared to time 0 post-infection (Fig. 1).
From the total and differential count of the five types of circulating hemocytes in the hemolymph of female R. sanguineus s.l. (plasmatocytes, granulocytes, spherulocytes, prohemocytes and adipohemocytes) the average total number of hemocytes in the R. sanguineus s.l. infected with
Discussion
A study by Prina et al. (2007) monitoring degradation of the DNA of Leishmania amazonensis, in vitro, in mouse macrophages, concluded that, after parasite death, DNA degraded in 1 h, with less than 1% of parasitic DNA remaining, disabling expression of their genes. In the present study, the presence of L. infantum DNA in R. sanguineus s.l. as well as a significant increase in copies of DNA was observed until the 7th day post-infection (dpi), suggesting that L. infantum remains viable in R.
Acknowledgements
The authors express their gratitude to the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) by fellowships. This work was developed in the Laboratory of Immunopathology Keizo Asami (LIKA)/UFPE and Institute Aggeu Magalhães (FIOCRUZ/PE).
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