BACKGROUND Trypanosoma cruzi circulates in sylvatic habitats, mainly through blood-feeding triatomines, although other routes also contribute to its dispersion. Sexual transmission of T. cruzi is an understudied topic, especially among wild mammals. Because of the difficulties inherent to field work, experimentally infected mice are frequently used to evaluate the transmission of T. cruzi. OBJECTIVE This study aimed to evaluate the sexual transmission of T. cruzi in acutely infected mice. METHODS Male and female mice in the acute phase of Chagas disease were mated with naïve partners. Then, parasitological tests, immunohistochemistry, serological assays, and polymerase chain reaction (PCR) assays were used to detect infection. FINDINGS Parasitological analysis showed trypomastigotes in the blood of 20% of the naïve mice after mating with infected partners. Serological assays detected anti-T. cruzi antibodies in all naïve females mated with infected males and in 60% of naïve males mated with infected females. PCR showed T. cruzi nDNA bands for all naïve mice mated with infected partners. The possibility of sexual transmission was also confirmed by visualisation of amastigotes in the testes. MAIN CONCLUSIONS Our results demonstrate that sexual transmission of T. cruzi is an ordinary event that may contribute to maintenance of the parasite's enzootic cycle.

BACKGROUND The Trypanosoma cruzi infection endemic in Latin America has now spread to several countries across four continents; this endemic involves triatomine vector-free protists. We hypothesised that the sexual transmission of T. cruzi contributes to the ongoing spread of Chagas disease. OBJECTIVES A short-term longitudinal study was conducted to evaluate this hypothesis. METHODS The study population comprised 109 subjects from four families, among whom 21 had been diagnosed with acute Chagas disease by direct parasitological analysis. Blood mononuclear cells and serum samples were obtained from each study subject once per year for three consecutive years. Enzyme-linked immunosorbent assay (ELISA) and indirect immunofluorescence serological examinations were used to detect specific T. cruzi antibodies. Polymerase chain reaction of T. cruzi DNA revealed 188-nucleotide bands, which hybridised to a specific radiolabelled probe and were confirmed by cloning and sequencing. RESULTS Three independent assessments at different time points revealed T. cruzi nuclear DNA footprints in 76% (83/109) of the study population with active infection. In contrast, the ELISA and indirect immunofluorescence assays detected the T. cruzi antibody in 28.4% (31/109) of the study samples. Moreover, the semen from 82.6% (19/23) of subjects people revealed harboured the 188- bp base pair T. cruzi footprint. Interestingly, the ejaculates of nuclear DNA-positive Chagas patient transmitted the T. cruzi upon peritoneal injection or infusion in the vagina of mice, and amastigotes were detected in the skeletal muscle, myocardium, vas deferens, and uterine tube. MAIN CONCLUSIONS T. cruzi infections can be transmitted from females or males to naïve mates through intercourse, and progeny showed discrepancies between the ratios of nuclear DNA footprints and specific antibody that can be explained by the tolerance attained during early embryo growth. Additional studies are needed to develop drugs to eradicate the infections. Additionally, the importance of a vigorous education, information, and communication program to prevent sexually transmitted Chagas disease in humans cannot be underemphasised.

En el cultivo de especies acuáticas, la nutrición adecuada es de importancia fundamental para el éxito de la actividad, especialmente durante las primeras etapas de la producción, período en que los animales son más susceptibles al mal manejo y a cambios ambientales repentinos. Objetivos: el propósito de este estudio fue evaluar la eficacia de la Spirulina platensis como suplemento alimenticio para el crecimiento y coloración de la tilapia roja. Métodos: las tilapias fueron cultivadas con alimento comercial (D1: control), alimento comercial + Spirulina platensis húmeda (D2); alimento comercial + Spirulina platensis seca (D3); alimento comercial + microalgas de agua dulce (D4). Resultados: en relación con el desempeño zootécnico, D2 mostró los mejores resultados. Las tasas de supervivencia de D1, D2 y D3 fueron mayores que D4 (p< 0.05). Los índices de masculinización y gastrointestinal fueron similares entre los tratamientos (p> 0.05). El índice hepato-somático fue similar entre D1 y D4 (p>0.05), pero menor que D2 y D3 (p<0.05). La intensidad del color amarillo en D2 fue moderada, mientras que en D3 fue débil (Abs 490nm). Los extractos de piel de peces que no consumieron S. platensis mostraron coloración muy débil. Los recuentos de microalgas acuáticas en D1 revelaron que la cianobacteria (género Microcystis) presentó 95% de dominancia. Conclusiones: los peces alimentados con S. platensis, húmeda o seca, presentaron mayor crecimiento y coloración que los animales expuestos a alimentos artificiales solos o combinados con microalgas de agua dulce. Las microalgas marinas demostraron ser eficaces en tilapia roja. Se sugiere realizar estudios sobre la relación costo-beneficio del cultivo y oferta de S. platensis para peces, y determinar las ventajas económicas que esta práctica puede traer a la piscicultura comercial.

Proper nutrition is critical for the production of aquatic species, especially during their early stages, when animals are more susceptible to mismanagement and sudden environmental changes. Objective: the purpose of this study was to evaluate the efficacy of Spirulina platensis as a nutritional supplement for growth and coloration of red tilapia. Methods: tilapia were fed with commercial feed (D1: control); commercial feed + wet Spirulina platensis (D2); commercial feed + dry Spirulina platensis (D3); commercial feed + freshwater microalgae (D4). Results: D2 showed the best growth performance results. Survival rates of D1, D2 and D3 were higher than D4 (p<0.05). Masculinization and gastrointestinal rates were similar between treatments (p>0.05). The hepatosomatic index was similar between D1 and D4 (p>0.05) but lower than D2 and D3 (p<0.05). Yellow color intensity in D2 was moderate, whereas D3 was weak (Abs. 490 nm). Skin extracts of fish not consuming S. platensis showed very weak staining. Aquatic algae counts in D1 revealed that cyanobacterium (Microcystis genus) presented 95% dominance. Conclusions: fish fed with wet or dr y S. platensis, showed higher growth and staining than animals exposed to commercial feed alone or combined with freshwater microalgae. Marine microalgae proved effective in red tilapia. It is suggested to study the cost-benefit of cultivation and supply of S. platensis to fish, and to determine the economic benefits that this practice can bring to commercial farming.

No cultivo de espécies aquáticas, uma nutrição adequada assume importância fundamental no sucesso da atividade, principalmente nas fases iniciais de criação, período em que os animais estão mais susceptíveis ao manejo errôneo e mudanças bruscas do ambiente. Objetivo: o Objetivo deste trabalho foi avaliar a eficácia de Spirulina platensis como suplemento alimentar no crescimento e coloração de tilápia vermelha. Métodos: as tilápias foram cultivadas com ração comercial (D1: controle), ração comercial + Spirulina platensis úmida (D2), ração comercial + Spirulina platensis seca (D3) e ração comercial + microalgas de água doce (D4). Resultados: em relação ao desempenho zootécnico, D2 obteve os melhores resultados. As taxas de sobrevivência alcançadas nos tratamentos D1, D2 e D3, foram superiores as alcançadas no tratamento D4 (p<0.05). Os índices de masculinização e gastrointestinais foram estatisticamente similares para todos os tratamentos (p<0.05). Os índices hepatossomáticos encontrados em D1 e D4 foram estatisticamente semelhantes (p<0.05), porém, ambos apresentaram valores abaixo dos encontrados em D2 e D3 (p<0.05). A intensidade da cor amarela de D2, após a extração de pigmentos das peles dos peixes, apresentou-se moderada, enquanto em D3 a intensidade foi fraca (Abs 490 nm). Os extratos das peles dos peixes que não consumiram S. platensis, apresentaram coloração muito fraca. As contagens das microalgas presentes na água verde de D1, realizadas através de microscopia óptica, revelou que a cianobactéria do gênero Microcystis apresentou 95% dominância. Conclusões: os peixes alimentados com S. platensis, úmida ou seca, obtiveram desempenho superior e coloração mais forte que os animais expostos somente ao alimento artificial ou combinado com microalgas de água doce. A microalga marinha demonstrou ser um suplemento alimentar eficiente para tilápia vermelha. Sugerem-se estudos da relação custo-benefício do cultivo e oferta de S. platensis para os peixes, desta forma elucidando as vantagens econômicas que esta prática poderá trazer para a tilapicultura comercial.

The horizontal transfer of Trypanosoma cruzi mitochondrial minicircle DNA to the genomes of naturally infected humans may play an important role in the pathogenesis of Chagas disease. Minicircle integrations within LINE-1 elements create the potential for foreign DNA mobility within the host genome via the machinery associated with this retrotransposon. Here we document integration of minicircle DNA fragments in clonal human macrophage cell lines and their mobilization over time. The movement of an integration event in a clonal transfected cell line was tracked at three months and three years post-infection. The minicircle sequence integrated into a LINE-1 retrotransposon; one such foreign fragment subsequently relocated to another genomic location in association with associated LINE-1 elements. The p15 locus was altered at three years as a direct effect of minicircle/LINE-1 acquisition, resulting in elimination of p15 mRNA. Here we show for the first time a molecular pathology stemming from mobilization of a kDNA/LINE-1 mutation. These genomic changes and detected transcript variations are consistent with our hypothesis that minicircle integration is a causal component of parasite-independent, autoimmune-driven lesions seen in the heart and other target tissues associated with Chagas disease.

Trypanosoma cruzi acute infections often go unperceived, but one third of chronically infected individuals die of Chagas disease, showing diverse manifestations affecting the heart, intestines, and nervous systems. A common denominator of pathology in Chagas disease is the minimal rejection unit, whereby parasite-free target host cells are destroyed by immune system mononuclear effectors cells infiltrates. Another key feature stemming from T. cruzi infection is the integration of kDNA minicircles into the vertebrate host genome; horizontal transfer of the parasite DNA can undergo vertical transmission to the progeny of mammals and birds. kDNA integration-induced mutations can enter multiple loci in diverse chromosomes, generating new genes, pseudo genes and knock-outs, and resulting in genomic shuffling and remodeling over time. As a result of the juxtaposition of kDNA insertions with host open reading frames, novel chimeric products may be generated. Germ line transmission of kDNA-mutations determined the appearance of lesions in birds that are indistinguishable from those seen in Chagas disease patients. The production of tissue lesions showing typical minimal rejection units in birds' refractory to T. cruzi infection is consistent with the hypothesis that autoimmunity, likely triggered by integration-induced phenotypic alterations, plays a major role in the pathogenesis of Chagas disease.

This work describes the development and functional testing of two episomes for stable transfection of Trypanosoma cruzi. pHygD contained the 5'- and 3'- flanking regions of the gene encoding the cathepsin B-like protease of T. cruzi as functional trans-splicing and polyadenylation signals for the hygR ORF. Evidence is presented to support extrachromosomal maintenance and organization as tandem repeats in transfected parasites. pPac was derived from pHygD by replacement of the entire hygR ORF with a purR coding region. The ability to modify pHygD and the availability of the complete DNA sequence make these plasmids useful tools for the genetic manipulation of T. cruzi.

The development of biotechnology in the last three decades has generated the feeling that the newest scientific achievements will deliver high standard quality of life through abundance of food and means for successfully combating diseases. Where the new biotechnologies give access to genetic information, there is a common belief that physiological and pathological processes result from subtle modifications of gene expression. Trustfully, modern genetics has produced genetic maps, physical maps and complete nucleotide sequences from 141 viruses, 51 organelles, two eubacteria, one archeon and one eukaryote (Saccharomices cerevisiae). In addition, during the Centennial Commemoration of the Oswaldo Cruz Institute the nearly complete human genome map was proudly announced, whereas the latest Brazilian key stone contribution to science was the publication of the Shillela fastidiosa genomic sequence highlythed on a Nature cover issue. There exists a belief among the populace that further scientific accomplishments will rapidly lead to new drugs and methodological approaches to cure genetic diseases and other incurable ailments. Yet, much evidence has been accumulated, showing that a large information gap exists between the knowledge of genome sequence and our knowledge of genome function. Now that many genome maps are available, people wish to know what are we going to do with them. Certainly, all these scientific accomplishments will shed light on many more secrets of life. Nevertheless, parsimony in the weekly announcements of promising scientific achievements is necessary. We also need many more creative experimental biologists to discover new, as yet un-envisaged biotechnological approaches, and the basic resource needed for carrying out mile stone research necessary for leading us to that "promised land"often proclaimed by the mass media.

Integration of kDNA sequences within the genome of the host cell shown by PCR amplification with primers to the conserved Trypanosoma cruzi kDNA minicircle sequence was confirmed by Southern hybridization with specific probes. The cells containing the integrated kDNA sequences were then perpetuated as transfected macrophage subclonal lines. The kDNA transfected macrophages expressed membrane antigens that were recognized by antibodies in a panel of sera from ten patients with chronic Chagas disease. These antigens barely expressed in the membrane of uninfected, control macrophage clonal lines were recognized neither by factors in the control, non-chagasic subjects nor in the chagasic sera. This finding suggests the presence of an autoimmune antibody in the chagasic sera that recognizes auto-antigens in the membrane of T. cruzi kDNA transfected macrophage subclonal lines.