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The expression of trans-sialidase and sialidase activities in the kinetoplastid protozoa was explored as a potential marker to discriminate between the morphologically indistinguishable flagellates isolated from human, insects and vertebrate reservoir hosts. By virtue of the differences observed in the ratios of these enzyme activities, a collection of 52 species and strains comprising the major taxa of these parasites could be separated into four expression types. Type-I parasites express comparable levels of both trans-sialidase and sialidase activities (Endotrypanum species and Trypanosoma lewisi). Type-I1 parasites express predominantly trans-sialidase activity (Trypanosoma cruzi and Trypanosoma conorhini).Type-I11 parasites express sialidase activity exclusively (Trypanosoma rangeli and Trypanosoma leeuwenhoeki). Type-IV parasites do not express either activity (Leishmania species and Trypanoplasma borreli). The measurement of trans-sialidase and sialidase activities thus permits the differentiation of parasites frequently found in the same insect vectors that are difficult to distinguish, such as T. cruzi and 7: rangeli, or in the same sylvatic vertebrate and invertebrate hosts, such as Leishmania and Endotrypanum.Trans-sialidases are a class of recently described cell-surface-exposed trans-glycosidases that differ from the Golgiassociated sialyltransferases in that they catalyze the transfer of N-acylneuraminate (sialic acid) residues from substrates other than cytidine monophosphate (CMP)-sialic acid to glycoproteins, glycolipids and polysaccharides. Like all transglycosidases, the trans-sialidases also exhibit hydrolase (i.e. sialidaseheuraminidase) activity in the absence of suitable acceptors.Trans-sialidase activity was discovered in the protozoan parasite Tqpanosoma cruzi (order Kinetoplastida, family Trypanosomatidae) [ 1 -51, the agent of Chagas' disease in the American continent, and shortly after in their African counterparts Trypanosoma brucei ssp. [6, 71, the causative organisms of sleeping sickness in humans and of nagana in domestic mammals. Trans-sialidases are, however, not restricted to trypanosomes as they also occur in the non-pathogenic trypanosomatids of the genus Endotrypanum [8], and in the pathogenic fungus Pneumocystis carinii (L. Trimble, N. Pavia & M. E. A. Pereira, unpublished results). Although unequivocal evidence for the specific biological roles of the trans-sialidases in these parasite systems is lacking, the T. cruzi trans-sialidase is believed to be involved in the process Currently, it is difficult to discriminate between pathogenic and non-pathogenic trypanosomatids. Striking similarities at the morphological, molecular and biological levels exist between many trypanosomatids isolated from sylvatic insects and/or vertebrate reservoir hosts that make the identification of the medically important parasites demanding (reviewed in [15, 161). Moreover, the geographical distributions of the different vectors and vertebrate reservoirs overlap. For example, the neotropical tre...

Epidemiological studies have been conducted to better understand the dynamics of American Cutaneous Leishmaniasis (ACL) in the Amazon region where distinct species of Leishmania circulate. In endemic areas, the optimal diagnosis must be made in the earlier clinical presentation to avoid the complications of chronic disease. The scarcity of financial support, laboratory infrastructure and trained persons are the major obstacles in this reality. This paper describes the result of performing different diagnostic methods for ACL in Amazonas State between the years 2010 and 2011. The tests used were the intradermal skin test (Montenegro's skin test), ELISA (Enzyme-Linked Immunosorbent Assay), direct examination, culture isolation and identification of Leishmania species. A total of 38 suspected human cases of ACL were diagnosed by different methods, of which 71.0% (n = 27) were positive by direct examination, 75.6% (n = 28) had positivity in the culture isolates and, of these, 54.0% (n = 19) had infection with Leishmania (Viannia) guyanensis. The positivity of the intradermal skin test with the leishmanin solution was observed in 77.0% of cases analyzed and the serology with detection of IgG and IgM showed the presence of antibodies in 100% of exams realized results, showing variation in the titles of antibodies. The success of Leishmaniasis treatment depends on an effective and early diagnosis. Parasitological diagnosis is highly specific, but sensitivity is subject to variation because the tissue distribution of parasites generally is not homogeneous and depends on the specie of parasite. Moreover, parasitological tests require invasive procedures and depend on restrictive conditions for the collection of biological sample, which limit their use in large-scale for epidemiological studies. ELISA has been the most widely used serological method for the diagnosis of Visceral Leishmaniasis (VL) as it is easy to perform and has a low cost. However, flaws in specificity are observed in the diagnosis of cutaneous leishmaniasis. Actually the diagnosis needs to be done as an associated methods depending on the question to be solved.

The authors discuss in this paper the role of inflammatory, anti-inflammatory, and regulatory cytokines in patients infected with different species of Leishmania in Amazonas State, Brazil. A comparative analysis was made of serum concentrations of these cytokines in the peripheral blood of 33 patients infected with cutaneous leishmaniasis. The isolates were identified as Leishmania guyanensis, L. naiffi, and L. amazonensis. Most (64%) of the patients were male ranging in age from 18 to 58 years. Protein expression profiles of IL-2, IL-4, IL-6, IL-10, IFN-γ, TNF-α, and IL-17 cytokines were shown to vary significantly between infected and noninfected (control group) individuals and according to the Leishmania species. Infection caused by L. guyanensis accounted for 73% of the cases and patients with this parasite also showed higher concentrations of IL-2, IFN-γ, IL-4, and IL-17 when compared to infection by L. amazonensis. Patients with infection caused by L. naiffi showed higher concentration of the cytokines analyzed when compared to uninfected patients; however, there was no statistically significant difference with the other species analyzed.

In this study, we have analysed enzyme polymorphism among a group of protozoan parasites of the genus Endotrypanum (Kinetoplastida: Trypanosomatidae). Seventeen stocks of Endotrypanum spp. isolated from sloths (Choloepus didactylus and C. juruanus) in the Amazon Region of Brazil were analysed by enzyme electrophoresis, and their electromorphic profiles were compared with reference strains reported previously. The 16 enzymic loci were analysed, and the strains were classified into zymodemes, each representing parasites with unique enzyme profiles. Each zymodeme was considered as an elementary taxon, and using numerical analyses (cladistic, agglomerative hierarchical and ordination techniques) the genus was shown to be monophyletic and the 12 zymodemes characterized could be divided into 3 groups (A, B, C). The heterogeneous population (which may represent a complex of parasite species or strains variants) showed, however, no correlation with the origin (i.e. host species involved or geographic area of isolation) of Endotrypanum stocks. Eight isolates of Endotrypanum sp. from Rondônia State (Brazil) and a parasite strain from Panama were clustered together into a zymodeme, which was phenetically closely related to the E. monterogeii from Costa Rica. The data indicate that E. schaudinni is a species complex.

Key words: Endotrypanum -Protozoa -Kinetoplastida -Trypanosomatidae -molecular taxonomy -enzyme activities -monoclonal antibodies -enzyme electrophoresis -DNA analyses -mammalian reservoirssandflies vectorsBiological characteristics of Endotrypanum spp. -Parasitic protozoa of the genus Endotrypanum are unique among the Kinetoplastida in that they infect erythrocytes of their mammalian host. Infection with Endotrypanum appears to be restricted to edentates, principally of forest-dwelling two-toed sloths of the genus Choloepus, but rarer infections with these flagellates seems to occur in three-toed sloths (genus Bradypus) (Fig. 1). As shown in Fig. 2, inside the erythocyte the Endotrypanum assumes an epimastigote or trypomastigote form, while in the sandfly or during in vitro culture the parasite assumes promastigote morphology (Shaw 1992). However, the complete life cycle of Endotrypanum spp. has not been reproduced in experimental studies. Other developmental stages of the parasite may occur within the mammalian host (Deane 1961).In nature, Endotrypanum parasites are probably transmitted by the bite of infected phlebotomine sandflies (Diptera: Psychodidae). Arias et al. (Rogers et al. 1988). Further evidence for the development of Endotrypanum in phlebotomines was obtained by feeding several laboratory-reared sandfly species on infected sloths (Shaw 1964, 1969, 1981, Christensen & Herrer 1977, 1979. In these studies, infections developing within the insect gut (Shaw 1964(Shaw , 1969 were similar to those found in wild caught sandflies (Johnson et al. 1963, Arias et al. 1985. Moreover, we have reported (Franco et al. 1997b) the developmental biology of Brazilian strains of Endotrypanum for three sandfly species. Development of Endotrypanum varied for each parasitehost species association. After feeding on culture forms of E. schaudinni, significantly more Lu. shannoni (100%, 9/9) became infected than did Lu. longipalpis (62.3%, 33/53) or Phlebotomus papatasi (27.3%, 15/55). The greatest number of infections were in the midgut and hindgut from 6 to 16 days after feeding, but flagellates also were present in the Malpighian tubules. Moreover, distinct development patterns in the sandfly gut were obtained when the Callejon Lu. longipalpis colony was fed on cultures of other Endotrypanum strains.

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