Cryptic species of Paracoccidioides brasiliensis: impact on paracoccidioidomycosis immunodiagnosis

We aimed to evaluate whether the occurrence of cryptic species of Paracoccidioides brasiliensis, S1, PS2, PS3 and Paracoccidioides lutzii, has implications in the immunodiagnosis of paracoccidioidomycosis (PCM). Small quantities of the antigen gp43 were found in culture filtrates of P. lutzii strains and this molecule appeared to be more variable within P. lutzii because the synonymous-nonsynonymous mutation rate was lower, indicating an evolutionary process different from that of the remaining genotypes. The production of gp43 also varied between isolates belonging to the same species, indicating that speciation events are important, but not sufficient to fully explain the diversity in the production of this antigen. The culture filtrate antigen AgEpm83, which was obtained from a PS3 isolate, showed large quantities of gp43 and reactivity by immunodiffusion assays, similar to the standard antigen (AgB-339) from an S1 isolate. Furthermore, AgEpm83 was capable of serologically differentiating five serum samples from patients from the Botucatu and Jundiaí regions. These patients had confirmed PCM but, were non-reactive to the standard antigen, thus demonstrating an alternative for serological diagnosis in regions in which S1 and PS2 occur. We also emphasise that it is not advisable to use a single antigen preparation to diagnose PCM, a disease that is caused by highly diverse pathogens.

Revalidation and redescription of Triatoma brasiliensis macromelasoma Galvão, 1956 and an identification key for the Triatoma brasiliensis complex (Hemiptera: Reduviidae: Triatominae)

Triatoma brasiliensis macromelasoma is revalidated based on the results of previous multidisciplinary studies on the Triatoma brasiliensis complex, consisting of crossing experiments and morphological, biological, ecological and molecular analyses. These taxonomic tools showed the closest relationship between T. b. macromelasoma and Triatoma brasiliensis brasiliensis. T. b. macromelasoma is redescribed based on specimens collected in the type locality and specimens from a F1 colony. The complex now comprises T. b. brasiliensis, T. b. macromelasoma, Triatoma melanica, Triatoma juazeirensis and Triatoma sherlocki. An identification key for all members of the complex is presented. This detailed comparative study of the morphological features of T. b. macromelasoma and the remaining members of the complex corroborates results from multidisciplinary analyses, suggesting that the subspecific status is applicable. This subspecies can be distinguished by the following combination of features: a pronotum with 1+1 narrow brownish-yellow stripes on the submedian carinae, not attaining its apex, hemelytra with membrane cells darkened on the central portion and legs with an incomplete brownish-yellow ring on the apical half of the femora. Because the T. brasiliensis complex is of distinct epidemiological importance throughout its geographic distribution, a precise identification of its five members is important for monitoring and controlling actions against Chagas disease transmission.

Phospholipase gene expression during Paracoccidioides brasiliensis morphological transition and infection

Phospholipase is an important virulence factor for pathogenic fungi. In this study, we demonstrate the following: (i) the Paracoccidioides brasiliensis pld gene is preferentially expressed in mycelium cells, (ii) the plb1 gene is mostly up-regulated by infection after 6 h of co-infection of MH-S cells or during BALB/c mice lung infection, (iii) during lung infection, plb1, plc and pld gene expression are significantly increased 6-48 h post-infection compared to 56 days after infection, strongly suggesting that phospholipases play a role in the early events of infection, but not during the chronic stages of pulmonary infection by P. brasiliensis.

Domestic, peridomestic and wild hosts in the transmission of Trypanosoma cruzi in the Caatinga area colonised by Triatoma brasiliensis

The role played by different mammal species in the maintenance of Trypanosoma cruzi is not constant and varies in time and place. This study aimed to characterise the importance of domestic, wild and peridomestic hosts in the transmission of T. cruzi in Tauá, state of Ceará, Caatinga area, Brazil, with an emphasis on those environments colonised by Triatoma brasiliensis. Direct parasitological examinations were performed on insects and mammals, serologic tests were performed on household and outdoor mammals and multiplex polymerase chain reaction was used on wild mammals. Cytochrome b was used as a food source for wild insects. The serum prevalence in dogs was 38% (20/53), while in pigs it was 6% (2/34). The percentages of the most abundantly infected wild animals were as follows: Thrichomys laurentius 74% (83/112) and Kerodon rupestris 10% (11/112). Of the 749 triatomines collected in the household research, 49.3% (369/749) were positive for T. brasiliensis, while 6.8% were infected with T. cruzi (25/369). In captured animals, T. brasiliensis shares a natural environment with T. laurentius, K. rupestris, Didelphis albiventris, Monodelphis domestica, Galea spixii, Wiedomys pyrrhorhinos, Conepatus semistriatus and Mus musculus. In animals identified via their food source, T. brasiliensis shares a natural environment with G. spixii, K. rupestris, Capra hircus, Gallus gallus, Tropidurus oreadicus and Tupinambis merianae. The high prevalence of T. cruzi in household and peridomiciliar animals reinforces the narrow relationship between the enzootic cycle and humans in environments with T. brasiliensis and characterises it as ubiquitous.

Thermal preferences and limits of Triatoma brasiliensis in its natural environment - Field observations while host searching

The goal of this work was to explore the thermal relationship between foraging Triatoma brasiliensis and its natural habitat during the hottest season in the state of Ceará, Brazil. The thermal profiles were determined using infrared analysis. Although the daily temperature of rock surfaces varied in a wide range, T. brasiliensisselected to walk through areas with temperatures between 31.7-40.5ºC. The temperature of T. brasiliensisbody surface ranged from 32.8-34.4ºC, being higher in legs than the abdomen. A strong relationship was found between the temperature of the insect and the temperature of rock crevices where they were hidden (r: 0.96, p < 0.05). The species was active at full sunlight being a clear example of how the light-dark rhythm may be altered, even under predation risk. Our results strongly suggest a thermal borderline for T. brasiliensisforaging activity near 40ºC. The simultaneous determination of insect body and rock temperatures here presented are the only obtained in natural habitats for this or other triatomines.

Antifungal activity of extracts from Atacama Desert fungi againstParacoccidioides brasiliensis and identification ofAspergillus felis as a promising source of natural bioactive compounds

Fungi of the genus Paracoccidioides are responsible for paracoccidioidomycosis. The occurrence of drug toxicity and relapse in this disease justify the development of new antifungal agents. Compounds extracted from fungal extract have showing antifungal activity. Extracts of 78 fungi isolated from rocks of the Atacama Desert were tested in a microdilution assay against Paracoccidioides brasiliensis Pb18. Approximately 18% (5) of the extracts showed minimum inhibitory concentration (MIC) values≤ 125.0 µg/mL. Among these, extract from the fungus UFMGCB 8030 demonstrated the best results, with an MIC of 15.6 µg/mL. This isolate was identified as Aspergillus felis (by macro and micromorphologies, and internal transcribed spacer, β-tubulin, and ribosomal polymerase II gene analyses) and was grown in five different culture media and extracted with various solvents to optimise its antifungal activity. Potato dextrose agar culture and dichloromethane extraction resulted in an MIC of 1.9 µg/mL against P. brasiliensis and did not show cytotoxicity at the concentrations tested in normal mammalian cell (Vero). This extract was subjected to bioassay-guided fractionation using analytical C18RP-high-performance liquid chromatography (HPLC) and an antifungal assay using P. brasiliensis. Analysis of the active fractions by HPLC-high resolution mass spectrometry allowed us to identify the antifungal agents present in the A. felis extracts cytochalasins. These results reveal the potential of A. felis as a producer of bioactive compounds with antifungal activity.

Breeding sites of Neotropical Drosophilidae (Diptera): IV. living and fallen flowers of Sessea brasiliensis and Cestrum spp. (Solanaceae)

The flowering phenologies of Sessea brasiliensis and three species of Cestrum (C. amictum, C. schlechtendalii and C. sendtnerianum), all belonging to the Solanaceae, were monitored in a fragment of the montane Atlantic rain forest within São Paulo city from July 1993 through June 1995. A total of 11,021 flies belonging to five species of the flavopilosa group of Drosophila (D. cestri, D. cordeiroi, D. hollisae, D. incompta and D. mariaehelenae), plus 1,244 flies belonging to four species of Zygothrica (Z. dispar, Z. vittimaculosa, Z. vittinubila and Z. aff. vittipunctata) emerged from 33,955 sampled flowers. Cestrum amictum, C. sendtnerianum and Sessea brasiliensis are recorded for the first time as host plants for the species of the Neotropical flavopilosa group ofDrosophila. Additionally, at least ten less abundant taxa belonging also to the Drosophilidae, plus 1,073 microhymenopteran parasitoids (Braconidae, Eucoilidae, and Pteromalidae), emerged from the sampled flowers of S. brasiliensis e Cestrum spp. The present study raised from nine to eleven the number of species of Cestrum previously recorded as breeding sites for these flies. A comprehensive list of species of Solanaceae known to host six species of Drosophila and four species of Zygothrica in southeastern and southern Brazil is included.

Mosquitoes (Diptera, Culicidae) inhabiting foliar tanks of Guzmania brasiliensis Ule (Bromeliaceae) in central Amazonia, Brazil

Mosquitoes in the family Culicidae develop their immature forms in different environments depending on various circumstances. This study represents the first contribution to the ecological knowledge of culicids living in the bromeliad G. brasiliensis. The objectives of the study were to compare the Culicidae fauna of bromeliads in two seasonal periods, analyzing their abundance, richness and the influence of pH, conductivity and water volume, on the populations. Sampling was done at Reserva Adolpho Ducke, Manaus, Amazonas, Brazil between 2003 and 2005. Six samplings were done in the rainy season and six in the dry season. On each sampling day, six terrestrial and six epiphytic bromeliads, totaling 144 sampling units, were retrieved. A total of 444 immature mosquitoes were collected, distributed in the following species: Culex (Microculex) stonei Lane & Whitman, Culex (Microculex) chryselatus Dyar & Knab, Culex (Microculex) sp., Wyeomyia (Hystatomyia) autocratica Dyar & Knab and Wyeomyia (Hystatomyia) splendida Bonne-Wepster & Bonne. The relationship between abundance of Culicidae and sampling period was not significant, although abundance was higher in the dry period. There were significant correlations between water volume and the abundance (p = 0.003) and richness (p = 0.001) of culicids. The pH was positively related to the occurrence of the species Wy. autocratica (p= 0.010) and Wy. splendida (p= 0.006), demonstrating the influence of this factor in the structuring of bromeliad-inhabiting communities.

Propriedades físicas de latossolo e argissolo em função de práticas de manejo aplicadas na entrelinha da cultura da seringueira (Hevea brasiliensis)

Avaliou-se a influência das práticas de manejo (gradagem, Pueraria phaseoloides e roçadeira) nas entrelinhas da cultura da seringueira (Hevea brasiliensis), plantada em 1992, sobre a densidade do solo e a macro e microporosidade de solos do Planalto Paulista. Foram retiradas amostras nos anos de 1998 e 1999, após seis anos consecutivos de manejo, nas profundidades de 0-10, 10-20, 20-30 e 30-40 cm do Latossolo Vermelho distrófico textura argilosa A moderado caulinítico hipoférrico relevo plano e Argissolo Vermelho-Amarelo distrófico abrúptico, Tb, A moderado textura areia/média fase floresta tropical subperenifólia e relevo suave ondulado, localizados, respectivamente, nos municípios de Jaboticabal e Tabapuã (SP). Para cada profundidade, foram retiradas oito amostras por tratamento, por experimento, em cada ano, totalizando 394 amostras. Os atributos físicos do solo (densidade do solo e macroporosidade) foram avaliados de acordo com as práticas de manejo aplicadas. Dentre os sistemas de manejos aplicados na entrelinha da seringueira, a roçadeira foi o que provocou maior compactação do solo em todas as profundidades, evidenciada pelos elevados valores de densidade do solo e reduzida macroporosidade, sendo este efeito mais pronunciado nas camadas superficiais dos solos.

Desenvolvimento in vitro de Agaricus brasiliensis em meios suplementados com diferentes farelos

O cogumelo Agaricus brasiliensis normalmente é cultivado em meios à base de batata ou composto orgânico. O objetivo deste trabalho foi avaliar o desenvolvimento in vitro da linhagem ABL 97/11 de A. brasiliensis, cultivada em meio à base de composto suplementado com diferentes concentrações de farelos de milho, trigo, arroz e soja, à temperatura constante de 28ºC. Avaliou-se, diariamente, o diâmetro da colônia e obteve-se, aos seis dias de cultivo, a massa miceliana. A adição de farelos de soja ou arroz não favorece o desenvolvimento in vitro de A. brasiliensis. O meio de cultura suplementado com 20% de farelo de trigo apresenta as maiores massa miceliana e velocidade de crescimento, comparado aos meios suplementados com outros farelos, na mesma concentração. Na concentração de 10% de farelo, o milho promove a maior velocidade de crescimento do cogumelo.

Trânsito gastrintestinal de dieta seca em Salminus brasiliensis

O objetivo deste trabalho foi avaliar tempo de passagem do alimento e seqüência de repleção e depleção dos órgãos no trato gastrintestinal de Salminus brasiliensis. Cem peixes (38,8±7,6 g) foram mantidos em 20 gaiolas (60 L), a 24,8±0,2ºC e alimentados com ração contendo 1% de Cr2O3. A cada 60 min a partir da primeira alimentação foram sacrificados cinco peixes e foi verificada a localização da digesta no trato gastrintestinal. A partir da segunda hora depois da alimentação, observou-se redução gradativa do conteúdo estomacal e subseqüente presença de digesta no intestino. Esse padrão se mantém até a 16ª hora; uma e duas horas depois, estômagos e intestinos, respectivamente, encontram-se completamente vazios.

Apparent digestibility of ingredients in diets for Salminus brasiliensis

The objective of this work was to determine the nutritional value of different protein sources for "dourado" (Salminus brasiliensis). Thirty juveniles per group (33.51±1.4 g) were hand fed on a reference diet (70%) added of tested ingredients (30%) and chromium oxide III (0.1%). Apparent digestibility coefficients of the gross energy (ADC GE), crude protein (ADC CP) and amino acids of the tested ingredients were evaluated. Corn gluten meal yielded the best results for ADC GE and ADC CP (95.7 and 96.9%, respectively) amongst plant ingredients. Spray-dried blood meal yielded the best values of ADC GE and ADC CP amongst animal ingredients (94.1 and 96.3%, respectively). Wheat bran yielded poorest ADCs coefficients (77 for ADC GE and 88.2% for ADC CP).

Biology and parasitic potential of Doryctobracon areolatuson Anastrepha fraterculus larvae

The objective of this work was to study the biology and parasitic potential of Doryctobracon areolatus on larvae of Anastrepha fraterculus. The egg-adult period, the sex ratio, the longevity, the pupal viability, and the parasitism rate of D. areolatus were determined in laboratory, using A. fraterculus as host. The parasitoid development from oviposition to adult emergence required 25.00±1.70 days, the sex ratio was 0.62±0.09, and the mean longevity was 16.36±3.62 days for males and 10.24±1.71 days for females. The mean parasitism rate was 53.50±8.93%, varying from 41.60 to 68.60%, which shows the potential of this parasitoid for biological control of A. fraterculus.

Lysine and arginine requirements of Salminus brasiliensis

The objective of this work was to determine the dietary lysine (DL) and dietary arginine (DA) requirements of dourado (Salminus brasiliensis), through dose-response trials using the amino acid profiles of whole carcasses as a reference. Two experiments were carried out in a completely randomized design (n=4). In the first experiment, groups of 12 feed-conditioned dourado juveniles (11.4±0.2 g) were stocked in 60 L cages placed in 300 L plastic indoor tanks in a closed circulation system. Fish were fed for 60 days on diets containing 1.0, 1.5, 2.0, 2.5, 3.0, or 3.5 % dietary lysine. In the second experiment, dourado juveniles (27.0±0.8 g) were fed for 60 days on semipurified diets containing arginine at 1.0, 1.5, 2.0, 2.5 or 3.0%, in similar conditions to those of the first experiment. Optimal DL requirements, as determined by broken-line analysis method for final weight, weight gain and specific growth rate, were 2.15% DL or 5% lysine in dietary protein, and 1.48% DA or 3.43% arginine in dietary protein. The best feed conversion ratio is attained with 2.5% DL or 5.8% lysine in dietary protein and 1.4% DA or 3.25% arginine in dietary protein.

Estaquia do porta-enxerto de videira '43-43' (V. vinifera x V. rotundifolia) resistente à Eurhizococchus brasiliensis

O experimento foi realizado com o objetivo de avaliar a propagação vegetativa por estaquia do porta-enxerto de videira '43-43'. Ramos herbáceos coletados em janeiro e semilenhosos em março que foram separados em dois lotes: um de estacas retiradas do ápice e outro da base dos ramos. As estacas foram então imersas por 10 segundos em soluções contendo diferentes concentrações de ácido indolilbutírico (0; 1.000; 1.500; 2.000; 2.500 ou 3.000 mg.L-1), totalizando 12 tratamentos para cada época de enraizamento. Posteriormente, as estacas foram plantadas em vasos de polipropileno com areia e mantidas em casa de vegetação sob sistema de nebulização. Sessenta dias após o plantio das estacas, para cada época, foram avaliadas as seguintes variáveis: porcentagem de estacas enraizadas, comprimento médio de raízes, massa fresca de raízes e porcentagem de estacas brotadas. Estacas herbáceas e semilenhosas apresentaram alta porcentagem de enraizamento (66,0% e 68,0%, respectivamente) mesmo quando não tratadas com o fitorregulador. Estacas apicais apresentaram maior comprimento e massa fresca de raízes. A aplicação de AIB reduziu a porcentagem de enraizamento e de brotação de estacas herbáceas e semilenhosas. Estacas semilenhosas, quando tratadas com o AIB a 1.000 mgL-1, apresentaram maior comprimento e massa fresca de raízes.