Isolation and characterization of a new agglutinin from the red marine alga Hypnea cervicornis J. Agardh.

The biochemical characterization of a new lectin (Hypnea cervicornis agglutinin or HCA) isolated from the Brazilian red alga H. cervicornis is reported. The haemagglutinating activity of the lectin was only inhibited by the glycoprotein porcine stomach mucin at a minimum inhibitory concentration of 19 microg x mL(-1). No haemagglutination inhibition was detected after the addition of simple sugars. The MALDI-TOF molecular masses of native and reduced and carbamidomethylated HCA were, respectively, 9196.6 Da and 9988.2 Da, indicating that the primary structure of the protein is crosslinked by 7 disulfide bonds. This unusual structural feature among lectins, along with its N-terminal sequence and amino-acid composition, clearly shows that HCA belongs to a protein family distinct from the isolectins Hypnin A1 and A2 isolated from the related Japanese alga Hypnea japonica. On the other hand, HCA displayed a high degree of similarity to the agglutinin from the Brazilian species Hypnea musciformis. Our data indicate the occurrence of structural diversity among lectins of closely related species living in distant ecosystems, i.e., the Pacific coast of Japan and the Atlantic coast of Brazil, and support the hypothesis that the lectin content (lectinome) might serve as a biomarker for taxonomical purposes.

Megabladder in experimental Chagas disease: pathological features of the bladder wall.

Mega-organs, primarily in the digestive tract, are well known to occur in chronic Chagas disease. Acute experimental infection with Trypanosoma cruzi results in parasitism of a wide range of cells, tissues, and organs, including the urinary bladder. Infection of BALB/c mice with 100,000 bloodstream forms of the Y strain of T. cruzi induced acute infection with intense parasitism of all layers of the urinary bladder. Parasites were found in the mucosa, lamina propria, muscular, adventitial connective, and fat tissue. Desquamate epithelial cells with amastigotes in the bladder lumen were also found. After 60 days of infection, mice inoculated with 50 bloodstream forms developed dilated, thin-walled bladders that had inflammatory infiltrates and foci of fibrosis replacing areas of damaged muscular layer. These lesions result from direct damage to the muscle fibers by the T. cruzi, leading to myosites, muscle damage, and scarring. Direct damage of paraganglia cells secondary to parasitism, leading to dilatation, damage of muscle fibers, and scarring with replacement of muscular tissue with connective tissue, should also be considered as a cause of functional disturbance of the urinary bladder.

Clinical analysis and parasite genetic diversity in human immunodeficiency virus/Chagas' disease coinfections in Brazil.

To evaluate the possible role of parasitemia on Chagas' disease reactivation in Chagas' disease/human immunodeficiency virus (HIV) coinfection cases and the impact of HIV coinfection on Trypanosoma cruzi genetic diversity, 71 patients with Chagas' disease (34 HIV+ and 37 HIV-) were surveyed. Moreover, 92 T. cruzi stocks from 47 chronic chagasic patients (29 HIV+ and 18 HIV-) were isolated and analyzed by multilocus enzyme electrophoresis and a random amplified polymorphic DNA procedure. High parasitemia appeared to play a major role in cases of Chagas' disease reactivation. In HIV+ patients, the genetic diversity and population structure (clonality) of T. cruzi was similar to that previously observed in HIV- patients, which indicates that immunodepression does not modify drastically genotype repartition of the parasite. There was no apparent association between given T. cruzi genotypes and specific clinical forms of Chagas' disease/HIV associations.

Life cycle of Trypanosoma cruzi (Y strain) in mice.

Since 1958, we have studied experimental Chagas' disease (CD) by subcutaneous inoculation of 1,000 blood forms of Trypanosoma cruzi (Y strain) in Balb/C. mice. Evolution of parasitemia remained constant, beginning on the 5th and 6th day of the disease, increasing progressively, achieving a maximum on about the 30th day. After another month, only a few forms were present, and they disappeared from the circulation after the third month, as determined from direct examination of slides and the use of a Neubauer Counting Chamber. These events coincided with the appearance of amastigote nests in the tissues (especially the cardiac ones), starting the first week, and following the Gauss parasitemia curve, but they were not in parallel until the chronic stage. In 1997, we began to note the following changes: Parasites appeared in the circulation during the first week and disappeared starting on the 7th day, and there was a coincident absence of the amastigote nests in the tissues. A careful study verified that young forms in the evolutionary cycle of T. cruzi (epi + amastigotes) began to appear alongside the trypomastigotes in the circulation on the 5th and 7th post-inoculation day. At the same time, rounded, oval, and spindle shapes were seen circulating through the capillaries and sinusoids of the tissues, principally of the hematopoietic organs. Stasis occurs because the diameter of the circulating parasites is greater than the vessels, and this makes them more visible. Examination of the sternal bone marrow revealed young cells with elongated forms and others truncated in the shape of a "C" occupying the internal surface of the blood cells that had empty central portions (erythrocytes?). We hypothesize that there could be a loss of virulence or mutation of the Y strain of Trypanosoma cruzi.

Reactivation of Chagas' disease in a human immunodeficiency virus-infected patient leading to severe heart disease with a late positive direct microscopic examination of the blood.

We report a human immunodeficiency virus (HIV)-infected man with chronic Chagas' disease who developed a congestive heart failure that could not be clinically controlled. Endomyocardial biopsy revealed severe myocarditis and the xenodiagnosis result was positive, but Trypanosoma cruzi by direct microscopic examination of the blood was found only four months after the symptoms had started. Treatment with benznidazole was effective in reducing parasitemia, stabilizing the clinical status, and controlling tissue damage related to the parasite. Although the finding of T. cruzi trypomastigotes by direct microscopic examination of the blood has been considered the mark of Chagas' reactivation in immunocompromised patients with chronic disease, in this case it was a late finding.

Experimental toxoplasmosis, and immunosuppression (parasite evaluation and histological alterations)

With the purpose of studying immunosuppressant drug action on parasites, the effect of Cyclosporine and cortisone in experimental toxoplasma evaluation, a highly virulent strain of Toxoplasma gondii (RH strain) was used. The animals were divided in two groups: control and immunosuppressed. Two animals from each group were sacrificed daily. Evaluation was based on parasitemia and parasitism of peritoneal exsudate and presence of the parasite in heart, lung, liver, spleen, small bowel, brain, cerebellum, spinal cord and eyes. Parasite infestation in peritoneal exsudate was 10 times greater than parasitemia. Red spots were observed after the second day; after the fourth day, focal necrosis with softening and fluidification of brain tissue spattered throughout the nervous tissue with enhanced vascularization. Capillaries obstructed by Toxoplasmas gondii were found, causing infarction and necrosis due to toxic or enzymatic substances. Images suggestive of cells an "owl-eye", Cytomegalovirus-like were found, with T. gondii, both in the brain, as well as in the tear glands and small bowel. Blood stream parasite infestation was observed, no histopathological lesions were found in the remaining tissue, possibly due to lack of time, for all animals died within a week's observation. We conclude that immunosuppression has worsened the outcome, anticipating the disease in 24 hours.

Hypovolemic hemorrhagic shock (an experimental study).

With the objective of studying the natural defense mechanisms against traumatic hemorrhages, 20 mice were studies. Some of the large vessels were examined histologically after fixing with formaldehyde, in cases of bleeding, bleeding + hemodilution and controls. In the hemo-diluted group, important alterations in the distribution of the components of the blood stream were confirmed and in one case, there was extravasation of liquid poor in cellular elements. The results show that hemodilution is an unfavorable condition for the natural defense mechanisms against hemorrhage. The repercussion of these experimental results on clinical cases of traumatic hemorrhage is speculated.

[Evaluation of the therapeutic activity of itraconazole in chronic infections, experimental and human, by Trypanosoma cruzi]. / Avaliação da atividade terapêutica do itraconazol nas infecções crônicas, experimental e humana, pelo Trypanosoma cruzi.

Other authors have demonstrated that itraconazole has antiparasitic activity against Trypanosoma cruzi both in vitro and in animal acute infection. Because of these observations, we decided to evaluate the chronic phase of this protozoal disease, since it is the most important form under a clinical and assistential point of view. We studied an infected mouse model as well as human cases of Chagas' disease. One hundred mg/kg/day by gastric tube, and 100 or 200 mg/day orally were given, respectively, during three months, without showing any beneficial effect, at least with the adopted methods.

[Xenodiagnosis in Chagas disease: influence of the triatominae sex]. / O xenodiagnóstico na doença de chagas: influência do sexo dos triatomíneos.

We evaluated, comparatively, the performance of female and male Triatoma infestans in the 5th instar of development, in xenodiagnosis for Chagas' disease. Xenodiagnosis were done in 40 patients with chronic infectans. For each diagnosis 20 nymphs female and 20 nymphs males were used, which were weighed immediately before and after the meal. Intestinal contents were examined about 20, 30, 60 and 90 days after the xenodiagnosis application. The females and males ingested the mean of 230 and 210 mg of blood, respectively. The results revealed positivity of 15 (37.5%) out of 40 xenodiagnosis, but any of the positive tests show 100% of the nymphs, female and/or male infected by T. cruzi. The females ingested significantly more blood than the males; this is coincidental with the higher female positivation to T. cruzi here observed. Our results suggest that the female nymphs seem able to increase the sensibility of the xenodiagnosis for chronic patients with Chagas' disease being necessary further studies are necessary to confirm this hypothesis.