Histoplasma capsulatum isolated from Didelphis albiventris (Marsupialia: Didelphidae) in the State of Minas Gerais, Brazil

We report the isolation of Histoplasma capsulatum from a culture of the viscera of Didelphis albiventris, one of the marsupial species found in Brazil. To our knowledge, this is the first report of the isolation of this fungus from this mammalian species. This finding confirms the ubiquitous presence of H. capsulatum in nature.     

Mapping of the distribution of Trypanosoma cruzi infection among small wild mammals in a conservation unit and its surroundings (Northeast-Brazil)

Maps are a useful tool that permits correlation of landscapes with hotspots of parasite transmission. Here, they were used as a tool for geovisualization to evaluate variables involved in the transmission of Trypanosoma cruzi among small wild mammals in an area endemic for Chagas disease, the "Serra da Capivara" National Park (PARNA) and its surroundings in Piauí State, Northeast Brazil. The implementation of a Geographical Information System (GIS) allowed the observation that a previously noted aggregated distribution of Triatoma sordida and Triatoma brasiliensis, T. cruzi prevalence and infection pattern of small wild mammals was directly or indirectly influenced by the local relief and human action. Small mammalian species diversity was higher in mesic refugia inside the park and in its buffer zone and lower in the disturbed area by anthropic activities. Didelphis albiventris was more abundant in the areas affected by human action. Thrichomys laurentius demonstrated to be an eclectic species and a competent reservoir of T. cruzi, being infected in all study areas. Small wild mammals infected with the TCII genotype of T. cruzi were localized only in the buffer zone of PARNA while TCI infected specimens were found in both areas, inside the PARNA and its buffer zone. The impact of biodiversity loss on the transmission cycle of T. cruzi in the wild environment was discussed.     

Experimental transmission of Sarcocystis speeri Dubey and Lindsay, 1999 from the South American opossum (Didelphis albiventris) to the North American opossum (Didelphis virginiana)

Sarcocystis speeri Dubey and Lindsay, 1999 from the South American opossum Didelphis albiventris was successfully transmitted to the North American opossum Didelphis virginiana. Sporocysts from a naturally infected D. albiventris from Argentina were fed to 2 gamma-interferon knockout (KO) mice. The mice were killed 64 and 71 days after sporocyst feeding (DAF). Muscles containing sarcocysts from the KO mouse killed 71 DAF were fed to a captive D. virginiana; this opossum shed sporocysts 11 days after ingesting sarcocysts. Sporocysts from D. virginiana were fed to 9 KO mice and 4 budgerigars (Melopsittacus undulatus). Schizonts, sarcocysts, or both of S. speeri were found in tissues of all 7 KO mice killed 29-85 DAF; 2 mice died 39 and 48 DAF were not necropsied. Sarcocystis stages were not found in tissues of the 4 budgerigars fed S. speeri sporocysts and killed 35 DAE These results indicate that S. speeri is distinct from Sarcocystis falcatula and Sarcocystis neurona, and that S. speeri is present in both D. albiventris and D. virginiana.     

Nippostrongylus brasiliensis-induced depression of heterologous immune responses: effects of the histamine H2 receptor antagonist, cimetidine

Both mucosal and systemic immune responses are depressed in mice infected with the nematode Nippostrongylus brasiliensis and this is correlated with a striking increase in the numbers of histamine-containing, mucosal-associated mast cells. As suppressor T cells bear histamine H2 receptors, we have used the H2 antagonist, cimetidine, to test whether histamine mediates N. brasiliensis-induced immunodepression. In uninfected mice, mitogenic responses to PHA and Con A were increased by treatment with cimetidine; in some experiments this treatment also increased antibody responses to a T-dependent antigen (TNP-BGG). By contrast, these T- and B-cell responses were markedly depressed in mice infected with N. brasiliensis, and cimetidine treatment did not alter this parasite-induced immunodepression. These results imply that although histamine can modulate immune responses in uninfected animals, it is not a major component of the immunoregulatory pathway induced by infection with N. brasiliensis.     

Molecular characterization of an opossum Didelphis albiventris (Marsupialia: Didelphidae) population in an urban fragment of the Brazilian Atlantic Rainforest and support to species barcode identification

We made a molecular study of 40 opossums, Didelphis albiventris, from an urban fragment of the Atlantic Rainforest in southeastern Brazil, analyzing a 653-bp sequence of cytochrome c oxidase, subunit I. We found three close connected haplotypes, with low nucleotide diversity and a haplotype diversity of 59.1% and confirmed sympatry between D. albiventris and D. aurita in this region. The clear phylogenetic separation shows the appropriateness of DNA barcode identification methodology for effectively discriminating between these opossum species.     

Intestinal distribution of worms and host ingesta in Nippostrongylus brasiliensis.

The gastrointestinal nematode Nippostrongylus brasiliensis is thought to feed on host ingesta, and it is generally thought that the presence of ingesta determines the distribution of this parasite within the host intestine. However, these assertions have not been supported by direct evidence. The purpose of this study was to test the hypothesis that N. brasiliensis worms are preferentially found in regions of the host small intestine containing ingesta. The relationship between worm and ingesta distribution was investigated using mice infected with N. brasiliensis and killed on day 8 postinfection at 0130, 0730, 1330, or 1930 hr. There was an inverse relationship between worm and ingesta distributions, and the worms were distributed significantly more anteriad in the intestine than host ingesta, at all times during the 24 hr. To determine what the worms fed on, host ingesta, tissue, and blood were differentially labeled with the fluorescent dyes rhodamine B and Fluoresbrite. The results of this study suggest that N. brasiliensis feeds on the host's intestinal wall, and that habitat distribution of this parasite within the small intestine is not directly related to the presence of luminal ingesta.     

Paracoccidioides brasiliensis induces secretion of IL-6 and IL-8 by lung epithelial cells. Modulation of host cytokine levels by fungal proteases

Paracoccidioides brasiliensis is a pathogenic, dimorphic fungus that causes paracoccidioidomycosis, a systemic human mycosis that is highly prevalent in Latin America. In this study, we demonstrated that P. brasiliensis yeasts induced interleukin (IL)-8 and IL-6 secretion by human lung epithelial A549 cells. However, tumor necrosis factor-α and interferon-γ were undetectable in these cultures. Moreover, P. brasiliensis yeasts induced activation of p38 mitogen-activated protein kinase (MAPK), c-Jun NH(2)-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK) 1/2 in A549 cells, and IL-8 and IL-6 secretion promoted by this fungus was dependent on activation of p38 MAPK and ERK 1/2. In addition, IL-8 and IL-6 levels were significantly higher in culture supernatants of A549 cells that were incubated with formaldehyde-fixed P. brasiliensis compared to cultures of cells that were infected with live yeasts. Our results indicate that the observed cytokine level differences were due to protease expression, in live yeasts, that degraded these cytokines. Degradation of human recombinant IL-8 and IL-6 by live P. brasiliensis was inhibited by AEBSF and aprotinin, suggesting that these proteases belong to a family of serine proteases. This is the first report showing that P. brasiliensis may modulate host inflammation by expressing proteases that degrade proinflammatory cytokines.     

Involvement of extracellular matrix proteins in the course of experimental paracoccidioidomycosis

We aimed at determining involvement of extracellular matrix proteins (ECMp) and an ECM-binding adhesin (32-kDa protein) from Paracoccidioides brasiliensis, in the course of experimental paracoccidioidomycosis. BALB/c mice were infected with P. brasiliensis conidia previously incubated with soluble laminin, fibronectin and fibrinogen or a mAb against the fungal adhesin. Inflammatory response, chitin levels and cytokine production at different postinfection periods were determined. Chitin was significantly decreased in lungs of mice infected with ECMp-treated conidia when compared with controls at week 8, especially with laminin and fibrinogen. Contrariwise, when animals were infected with mAb-treated conidia no differences in chitin content were found. The observed inflammatory reaction in lungs was equivalent in all cases. IFN-gamma increased significantly in lungs from mice infected with soluble ECMp - (at day 4 and week 12) or mAb-treated conidia (at week 12) when compared with animals infected with untreated conidia. Significant increased levels of tumour necrosis factor-alpha were observed at 8 weeks in animals infected with ECMp-treated conidia while no differences were observed during the remaining periods. These findings point toward an inhibitory effect of ECMp on P. brasiliensis conidia infectivity and suggest that these proteins may interfere with conidia initial adhesion to host tissues probably modulating the immune response in paracoccidioidomycosis.     

Suppression of extraintestinal and intestinal Nippostrongylus brasiliensis-induced eosinophilia by Eimeria nieschulzi

Eosinophil responses in extraintestinal and intestinal tissues were examined in August and Sprague-Dawley rats infected with Nippostrongylus brasiliensis or Eimeria nieschulzi (or both), and in uninfected controls to test the hypothesis that E. nieschulzi suppresses the systemic N. brasiliensis-induced eosinophil response. Caudal vein blood, femoral bone marrow, bronchoalveolar lavage fluid, peritoneal lavage fluid, and duodenal and jejunal samples were collected on day 8 postinfection (PI) with E. nieschulzi, on day 16 PI of the N. brasiliensis infection, when these days coincided in the concurrently infected rats, and from uninfected controls. Differential white blood cell counts were made from blood smears and cytocentrifuged preparations, and duodenal and jejunal eosinophils per villus crypt unit were quantified. Eimeria nieschulzi significantly reduced N. brasiliensis-induced eosinophil levels in peripheral blood, lavage fluids, and duodenal and jejunal tissues in both rat strains. August and Sprague-Dawley rats monospecifically infected with N. brasiliensis and concurrently with both parasites demonstrated elevated eosinopoiesis compared with uninfected controls and rats infected with only E. nieschulzi; however, despite this, concurrently infected rats had a significantly greater level of eosinopoiesis than those infected with only the nematode. In addition, E. nieschulzi induced elevated neutrophil levels in both monospecifically and concurrently infected rats in all extraintestinal tissues examined in both rat strains, whereas lymphocyte counts decreased concomitantly. This study suggests that the intestinal coccidian E. nieschulzi has the ability to modulate the systemic inflammatory response to N. brasiliensis and that this is not a rat strain-specific phenomenon.     

Suppression of Nippostrongylus brasiliensis (Nematoda)-induced lysophospholipase activity and peripheral eosinophilia by Eimeria nieschulzi (Apicomplexa)

Interspecific interactions between Nippostrongylus brasiliensis and Eimeria nieschulzi were studied by measuring fecal lysophospholipase (LYPH) activity and relative numbers of peripheral eosinophils in rats singly or concurrently infected with one or both parasite species. Three groups of 10 rats each were inoculated with 2 X 10(3) N. brasiliensis L3 larvae and/or 5 X 10(5) E. nieschulzi sporulated oocysts. Groups 1 and 2 were infected with E. nieschulzi or N. brasiliensis, respectively. Group 3 rats were infected first with N. brasiliensis, followed on day 8 postinoculation (PI) with E. nieschulzi. Each rat served as its own control. Results revealed LYPH levels rose steadily in Group 2 rats, reaching significant peaks on days 10 and 12 PI before decreasing to control levels. Lysophospholipase activity in Groups 1 and 3, however, did not differ from control values. Group 2 rats also demonstrated peripheral eosinophilia, with peak values occurring on days 10, 12, 14, and 16 PI, while rats in Groups 1 and 3 exhibited no eosinophilia. These results demonstrate that E. nieschulzi suppressed intestinal LYPH activity and relative peripheral eosinophilia and demonstrate that a host's immune response to a single parasite may be significantly altered when a second parasite species is present.     

Physiological responses of rats to primary infection with Nippostrongylus brasiliensis

The physiological responses of well-nourished rats to primary infection with the intestinal nematode Nippostrongylus brasiliensis were examined. Infected rats fed ad libitum were compared with uninfected control rats fed ad lib. and with uninfected rats which were pair-fed to the infected rats. Following infection with N. brasiliensis rat food intake was reduced from day 2 post infection (pi) and there were two periods of minimal food intake (days 2 to 3 and 8 to 9 pi). The water intake of infected rats was only reduced on days 2, 3 and 9 pi and not to the same extent as food intake. Muscle catabolism in infected rats was more severe than could be explained on the basis of their food intake reduction. The rectal temperature and rate of oxygen consumption per g body-weight of rats was not significantly altered by the infection. Host responses to N. brasiliensis are compared with those seen in microbial infections and some of them are found to be considerably different.     

In vitro cultivation of schizonts of Sarcocystis speeri Dubey and Lindsay, 1999

Schizonts of Sarcocystis speeri Dubey and Lindsay, 1999 were cultured in vitro in bovine monocyte and equine kidney cell cultures inoculated with infected tissues of nude and gamma-interferon knockout mice fed sporocysts from opossums, Didelphis albiventris. At least 1 asexual cycle was completed in 3 days. In vitro-grown merozoites were structurally and antigenically distinct from those of Sarcocystis neurona and Sarcocystis falcatula. Culture-derived merozoites of S. speeri were not infective to budgerigars (Melopsittacus undulatus).     

Phenotypic and functional characterization of NK cells in human immune response against the dimorphic fungus Paracoccidioides brasiliensis

Besides their role in fighting viral infection and tumor resistance, recent studies have shown that NK cells also participate in the immune response against other infectious diseases. The aim of this study was to characterize the possible role of NK cells in the immune response against Paracoccidioides brasiliensis. Purified NK cells from paracoccidioidomycosis patients and healthy individuals were incubated with P. brasiliensis yeast cells or P. brasiliensis-infected monocytes, with or without the addition of recombinant IL-15. We found that NK cells from paracoccidioidomycosis patients exhibit a lower cytotoxic response compared with healthy individuals. NK cells are able directly to recognize and kill P. brasiliensis yeast cells, and this activity seems to be granule-dependent but perforin-independent, whereas the cytotoxicity against P. brasiliensis-infected monocytes is perforin-dependent. These results indicate that NK cells participate actively in the immune response against the P. brasiliensis infection either by directly destroying yeast cells or by recognizing and killing infected cells. Granulysin is the possible mediator of the cytotoxic effect, as the reduced cytotoxic activity against the yeast cells detected in patients with paracoccidioidomycosis is accompanied by a significantly lower frequency of CD56(+)granulysin(+) cells compared with that in healthy controls. Furthermore, we show that NK cells released granulysin in cultures after being stimulated by P. brasiliensis, and this molecule is able to kill the yeast cells in a dose-dependent manner. Another important finding is that stimulated NK cells are able to produce proinflammatory cytokines (IFN-γ and TNF-α) supporting their immunomodulatory role in the infection.     

Responses of inbred mouse strains to infection with intestinal nematodes

Comparisons were made of the immune and inflammatory responses of four strains of inbred mice to infection with the intestinal nematodes Trichinella spiralis and Nippostrongylus brasiliensis to determine whether genetically determined 'high responsiveness' to infection, seen most clearly in intestinal responses, is independent of the parasite concerned and necessarily correlated with protection. The time course of infection was followed by counting adult worms at intervals after infection. Mucosal mast cells and Paneth cell numbers were determined as indices of the intestinal inflammatory response. Levels of IgG2a and IgG1 antibodies and of the cytokines IFN-gamma and IL-5 released from in vitro-stimulated mesenteric node lymphocytes were measured to assess type 1 and type 2 responses. NIH and CBA mice were the most resistant to T. spiralis and N. brasiliensis respectively, resistance in each case being correlated with the most intense intestinal inflammatory responses. C57BL/10 (B10) and B10.BR were the least resistant to T. spiralis, but were as resistant as CBA to N. brasiliensis, despite their intestinal inflammatory responses to both parasites being much lower than the other two strains. Mice infected with T. spiralis made the expected switch from a type 1 (IFN-gamma) to a type 2 (IL-5) response between days 2 and 8, and there were no significant differences in levels of these cytokines between the strains. In contrast, when infected with N. brasiliensis, CBA showed an IFN-gamma response at day 4, all strains switching to IL-5 by day 8 and NIH mice releasing the greatest amount of IL-5. The results indicate that the "high responder" phenotype to intestinal nematode infection is in part determined by host characteristics, but is also determined by the parasite concerned--seen most clearly by the differences between NIH and CBA when infected with T. spiralis and N. brasiliensis. The fact that "low responder" B10 background mice were more resistant to N. brasiliensis than "high responder" NIH implies that each parasite elicits a particular pattern of protective host responses, rather than parasites being differentially susceptible to the same response profile.     

Arthroderma benhamiae infection in the Central African hedgehog,Erinaceus albiventris,and a report of a human case

The occurrence of ringworm caused by Arthroderma benhamiae Ajello & Cheng is reported in Central African hedgehogs (Erinaceus albiventris Wagner), caught near Nairobi, Kenya. Of the 45 animals examined, 10 were positive on culture, including a litter of 4 young. Six infected animals were without lesions, and 2 littermates showed scaly areas similar to those described in E. europaeus L. caused by Trichophyton erinacei (Smith & Marples) Padhye & Carmichael. No correlation with mite infestation or mange lesions was evident. Ringworm-like lesions were found which were repeatedly negative on culture. A human infection by A. benhamiae was contracted from the hedgehogs.     

Comparison between human and armadillo Paracoccidioides brasiliensis isolates by random amplified polymorphic DNA analysis

Sixty-three Paracoccidioides brasiliensis isolates obtained from three nine-banded armadillos ( Dasypus novemcinctus), one Amazonian armadillo's and 19 clinical isolates were compared by random amplified polymorphic DNA analysis with the primer OPG-19. The isolates were divided into three major clusters, I, II and III. Coincidences between human and armadillo isolates were observed in clusters I and II. Cluster III consisted only of armadillos' isolates. The results suggested that (I) humans may acquire P. brasiliensis infection by contact with armadillo's environment, (II) there may be P. brasiliensis genotypes peculiar to the animal, and (III) individual armadillos may be infected with P. brasiliensis cells with different genotypes. This revised version was published online in June 2006 with corrections to the Cover Date.     

Invasion of epithelial mammalian cells by Paracoccidioides brasiliensis leads to cytoskeletal rearrangement and apoptosis of the host cell

Paracoccidioides brasiliensis (Pb) yeast cells can enter mammalian cells and probably manipulate the host cell environment to favor their own growth and survival. We studied the uptake of strain Pb 18 into A549 lung and Vero epithelial cells, with an emphasis on the repercussions in the cytoskeleton and the apoptosis of host cells. Cytoskeleton components of the host cells, such as actin and tubulin, were involved in the P. brasiliensis invasion process. Cytochalasin D and colchicine treatment substantially reduced invasion, indicating the functional participation of microfilaments (MFs) and microtubules (MTs) in this mechanism. Cytokeratin could also play a role in the P. brasiliensis interaction with the host. Gp43 was recognized by anti-actin and anti-cytokeratin antibodies, but not by anti-tubulin. The apoptosis induced by this fungus in infected epithelial cells was demonstrated by various techniques: TUNEL, DNA fragmentation and Bak and Bcl-2 immunocytochemical expression. DNA fragmentation was observed in infected cells but not in uninfected ones, by both TUNEL and gel electrophoresis methods. Moreover, Bcl-2 and Bak did not show any differences until 24 h after infection of cells, suggesting a competitive mechanism that allows persistence of infection. Overexpression of Bak was observed after 48 h, indicating the loss of competition between death and survival signals. In conclusion, the mechanisms of invasion of host cells, persistence within them, and the subsequent induction of apoptosis of such cells may explain the efficient dissemination of P. brasiliensis.     

The epidemiologic importance of Triatoma brasiliensis as a chagas disease vector in Brazil: a revision of domiciliary captures during 1993-1999

To clarify the epidemiologic importance of Triatoma brasiliensis, the most important Chagas disease vector in the Northeastern of Brazil, capture data related to this species, its distribution, capture index, and percentages of natural infection by Trypanosoma cruzi were examined in 12 different Brazilian states. The Brazilian National Health Foundation collected these data from 1993 to 1999, a period during which a total of 1,591,280 triatomines (21 species) were captured in domiciles within the geographic range of T. brasiliensis. Of this total, 422,965 (26.6%) were T. brasiliensis, 99.8% of which were collected in six states, and 54% in only one state (Ceará). The percentage of bugs infected with T. cruzi varied significantly among states, ranging from 0% (Goiás, Maranh?o, Sergipe, and Tocantins) to more than 3% (Alagoas, Minas Gerais, and Rio Grande do Norte) with an average of 1.3%. This latter value represents a dramatic reduction in the natural infection percentages since 1983 (6.7%) suggesting that, despite the impossibility of eradicating this native species, the control measures have significantly reduced the risk of transmission. However, the wide geographic distribution of T. brasiliensis, its high incidence observed in some states, and its variable percentages of natural infection by T. cruzi indicate the need for sustained entomological surveillance and continuous control measures against this vector.     

Model of in vitro granulomatous hypersensitivity in human paracoccidioidomycosis

With the purpose of studying the immunological components of granulomatous hypersensitivity in patients infecteded with Paracoccidioides brasiliensis, we used the model of in vitro granuloma formation developed for schistosomiasis studies, that correlates with in vivo granulomatous reactivity occurring around eggs trapped in organs of infected donors. In this case, granuloma formation can be determined examining cellular reactivity manifested as multiple cell layers surrounding antigen-conjugated polyacrilamide beads. Our results showed that peripheral blood mononuclear cells (PBMC) obtained from acute treated and chronic paracoccidioidomycosis patients proliferate and generate in vitro granulomas in response to P. brasiliensis antigens (PbAg). In contrast, no proliferation or granuloma formation were observed when PBMC from acute non-treated patients were used. These studies demonstrate the feasibility of investigating granulomatous hypersensitivity in P. brasiliensis-infected patients by using an in vitro granuloma model. This revised version was published online in June 2006 with corrections to the Cover Date.     

Natural infection of the anal glands of the opossum (Didelphis albiventris) by Trypanosoma cruzi in the municipality of Bambuí--MG, Brazil

Out of 87 opossums, Didelphis albiventris, captured in the Bambuí area (Minas Gerais State), 32 (36.7%) were found infected by Trypanosoma cruzi; the rates varied according to whether the specimens originated from sylvan, rural peridomiciliar or urban surroundings, being 34.9, 81.8 and 7.7 respectively. From 20 of the infected opossums the anal glands were repeatedly examined and found positive in only one (5%) specimen (GA 9), with 7 positive examinations out of 17 performed through an 18-months periods. Material from these glands produced patent parasitemia in opossums and sub-patent infections in mice. Isolates from the opossum GA 9, obtained through xenodiagnoses and hemocultures and from cultures of the infected anal glands fitted into zymodeme Z1.