Epidemiological and clinical features of Leishmania (Viannia) braziliensis American cutaneous and mucocutaneous leishmaniasis in the State of Espírito Santo, Brazil

Between 1985 and 2000, epidemiological surveys of the American tegumentary leishmaniasis (ATL) were carried out in several rural and urban communities in Espírito Santo, Brazil. A total of 100 stocks of Leishmania (comprising isolates from both human and canine hosts with ATL) were identified by two methods of molecular characterization, using specific monoclonal antibodies and multilocus enzyme electrophoresis. Parasite isolates from 19 municipalities were found to belong to the same zymodeme and serodeme type as of the Leishmania (Viannia) braziliensis reference strain. In contrast, our genotyping studies have shown intra-specific variation among these parasites (comparisons of the variability of the internal transcribed spacers between the small and large subunits of the rRNA genes of the 22 stocks studiedrevealed at least 11 genotypes). Two main clusters of L. (V.) braziliensis genotypes were observed, representing parasites collected from different endemic regions in the state, where transmission reflects distinct eco-epidemiological features. Infection with this pathogen was associated with the characteristic disease forms, but neither the clinical outcome nor the response to treatment could be related to the genetic polymorphism of the isolates, as defined by using the proposed methodology.

Dominant character of the molecular marker of a Biomphalaria tenagophila strain (Mollusca: Planorbidae) resistant to Schistosoma mansoni

Biomphalaria tenagophila population from Taim (state of Rio Grande do Sul, Brazil) is totally resistant toSchistosoma mansoni, and presents a molecular marker of 350 bp by polymerase chain reaction and restriction fragment length polymorphism of the entire rDNA internal transcriber spacer. The scope of this work was to determine the heritage pattern of this marker. A series of cross-breedings between B. tenagophila from Taim (resistant) and B. tenagophila from Joinville, state of Santa Catarina (susceptible) was carried out, and their descendants F1 and F2 were submitted to this technique. It was possible to demonstrate that the specific fragment from Taim is endowed with dominant character, since the obtained segregation was typically mendelian.

The use of the polymerase chain reaction and restriction fragment length polymorphism technique associated with the classical morphology for characterization of Lymnaea columella, L. viatrix, and L. diaphana (Mollusca: Lymnaeidae)

The specific identification of Lymnaeid snails is based on a comparison of morphological characters of the shell, radula, renal and reproductive organs. However, the identification is complicated by dissection process, intra and interspecific similarity and variability of morphological characters. In the present study, polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP) techniques targeted to the first and second internal transcribed spacers (ITS1 and ITS2) rDNA and to the mitochondrial 16S ribosomal gene (16S rDNAmt) were used to differentiate the species Lymnaea columella, L. viatrix, and L. diaphana from some localities of Brazil, Argentina, and Uruguay as well as to verify whether the molecular results corroborates the classical morphological method.PCR-RFLP analysis of the ITS1, ITS2, and 16S using 12 restriction enzymes revealed characteristic patterns for L. columella and L. diaphana which were concordant with the classical morphology. On the other hand, for L. viatrix populations a number of 1 to 6 profiles were generated while morphology provided the species pattern results.

Molecular characterization of human Trypanosoma cruzi isolates from endemic areas in Panama

The present work provides information on Trypanosoma cruzi genotype circulating in endemic areas of Chagas disease in Panama. A total of 26 crude stocks of T. cruzi, isolated from the blood of persons with different clinical profiles of Chagas disease were collected and crio-conserved until used. Most of the stocks had been characterized by means of isoenzyme electrophoresis on cellulose acetate membranes. The clinical profiles of infected persons included 9 (34.6%) asymptomatic and 17 acute (65.4%) including 5 (19.2%) fatal cases, 2 under 5 years old and 3 adults. A multiplex-PCR assay based on the amplification of the non-transcribed spacer of the mini-exon gene was performed. All stocks of T. cruzi included in the study were found to correspond to Tc I group. This result supports the predominance of T. cruzi-I in the transmission cycles affecting the human population in the Republic of Panama.

Footprints of a trypanosomatid RNA world: pre-small subunit rRNA processing by spliced leader addition trans-splicing

The addition of a capped mini-exon [spliced leader (SL)] through trans-splicing is essential for the maturation of RNA polymerase (pol) II-transcribed polycistronic pre-mRNAs in all members of the Trypanosomatidae family. This process is an inter-molecular splicing reaction that follows the same basic rules of cis-splicing reactions. In this study, we demonstrated that mini-exons were added to precursor ribosomal RNA (pre-rRNA) are transcribed by RNA pol I, including the 5' external transcribed spacer (ETS) region. Additionally, we detected the SL-5'ETS molecule using three distinct methods and located the acceptor site between two known 5'ETS rRNA processing sites (A' and A1) in four different trypanosomatids. Moreover, we detected a polyadenylated 5'ETS upstream of the trans-splicing acceptor site, which also occurs in pre-mRNA trans-splicing. After treatment with an indirect trans-splicing inhibitor (sinefungin), we observed SL-5'ETS decay. However, treatment with 5-fluorouracil (a precursor of RNA synthesis that inhibits the degradation of pre-rRNA) led to the accumulation of SL-5'ETS, suggesting that the molecule may play a role in rRNA degradation. The detection of trans-splicing in these molecules may indicate broad RNA-joining properties, regardless of the polymerase used for transcription.

Comparative analyses of classical phenotypic method and ribosomal RNA gene sequencing for identification of medically relevant Candida species

As the distribution of Candida species and their susceptibility to antifungal agents have changed, a new means of accurately and rapidly identifying these species is necessary for the successful early resolution of infection and the subsequent reduction of morbidity and mortality. The current work aimed to evaluate ribosomal RNA gene sequencing for the identification of medically relevant Candida species in comparison with a standard phenotypic method. Eighteen reference strains (RSs), 69 phenotypically identified isolates and 20 inconclusively identified isolates were examined. Internal transcribed spaces (ITSs) and D1/D2 of the 26S ribosomal RNA gene regions were used as targets for sequencing. Additionally, the sequences of the ITS regions were used to establish evolutionary relationships. The sequencing of the ITS regions was successful for 88% (94/107) of the RS and isolates, whereas 100% of the remaining 12% (13/107) of the samples were successfully analysed by sequencing the D1/D2 region. Similarly, genotypic analysis identified all of the RS and isolates, including the 20 isolates that were not phenotypically identified. Phenotypic analysis, however, misidentified 10% (7/69) of the isolates. Phylogenetic analysis allowed the confirmation of the relationships between evolutionarily close species. Currently, the use of genotypic methods is necessary for the correct identification of Candida species.

Genetic diversity among isolates of stemphylium solani from cotton

The fungus Stemphylium solani causes leaf blight of tomato (Lycopersicon esculentum) in Brazil. In recent years, severe epidemics of a new leaf blight of cotton (Gossipium hyrsutum) caused by S. solani occurred in three major cotton-growing Brazilian states (PR, MT and GO). Molecular analysis was performed to assess the genetic diversity among the S. solani isolates from cotton, and to verify their relationship with representative S. solani isolates from tomato. Random amplified polymorphic DNA (RAPD) markers and internal transcribed spacers of ribosomal DNA (rDNA) were used to compare 33 monosporic isolates of S. solani (28 from cotton and five from tomato). An isolate of Alternaria macrospora from cotton was also used for comparison. RAPD analysis showed the presence of polymorphism between the genera and the species. The A. macrospora and the S. solani isolates from cotton and tomato were distinct from each other, and fell into separate groups. Variation by geographic region was observed for the tomato isolates but not for the cotton isolates. Amplifications of the ITS region using the primer pair ITS4/ITS5 resulted in a single PCR product of approximately 600 bp for all the isolates. Similarly, when amplified fragments were digested with eight restriction enzymes, identical banding patterns were observed for all the isolates. Hence, rDNA analysis revealed no inter-generic or intra-specific variation. The genetic difference observed between the cotton and the tomato isolates provides evidence that S. solani attacking cotton in Brazil belongs to a distinct genotype.

Intergenic and external transcribed spacers of ribosomal RNA genes in lizard-infecting Leishmania: molecular structure and phylogenetic relationship to mammal-infecting Leishmania in the subgenus Leishmania (Leishmania)

To establish the relationships of the lizard- and mammal-infecting Leishmania, we characterized the intergenic spacer region of ribosomal RNA genes from L. tarentolae and L. hoogstraali. The organization of these regions is similar to those of other eukaryotes. The intergenic spacer region was approximately 4 kb in L. tarentolae and 5.5 kb in L. hoogstraali. The size difference was due to a greater number of 63-bp repetitive elements in the latter species. This region also contained another element, repeated twice, that had an inverted octanucleotide with the potential to form a stem-loop structure that could be involved in transcription termination or processing events. The ribosomal RNA gene localization showed a distinct pattern with one chromosomal band (2.2 Mb) for L. tarentolae and two (1.5 and 1.3 Mb) for L. hoogstraali. The study also showed sequence differences in the external transcribed region that could be used to distinguish lizard Leishmania from the mammalian Leishmania. The intergenic spacer region structure features found among Leishmania species indicated that lizard and mammalian Leishmania are closely related and support the inclusion of lizard-infecting species into the subgenus Sauroleishmania proposed by Saf'janova in 1982.

Soil profile internal drainage for a central pivot fertigated coffee crop

Coffee cultivation via central-pivot fertigation can lead to fertilizer losses by soil profile internal drainage when water application is excessive and soils have low water retention and cation adsorption capacities. This study analyses the deep water losses from the top 1 m sandy soil layer of east Bahia, Brazil, cultivated with coffee at a high technology level (central-pivot fertigation), using above normal N fertilizer rates. The deep drainage (Q) estimation is made through the application of a climatologic water balance (CWB) program having as input direct measures of irrigation and rainfall, climatological data from weather stations, and measured soil water retention characteristics. The aim of the study is to contribute to the understanding of the hydric regime of coffee crops managed by central-pivot irrigation, analyzing three scenarios (Sc): i) rainfall only, ii) rainfall and irrigation full year, and iii) rainfall and irrigation dry season only. Annual Q values for the 2008/2009 agricultural year were: Sc i = 811.5 mm; Sc ii = 1010.5 mm; and Sc iii = 873.1 mm, so that the irrigation interruption in the wet season reduced Q by 15.7%, without the appearance of water deficit periods. Results show that the use of the CWB program is a convenient tool for the evaluation of Q under the cited conditions.

Internal consistency of the self-reporting questionnaire-20 in occupational groups

ABSTRACT OBJECTIVE To assess the internal consistency of the measurements of the Self-Reporting Questionnaire (SRQ-20) in different occupational groups. METHODS A validation study was conducted with data from four surveys with groups of workers, using similar methods. A total of 9,959 workers were studied. In all surveys, the common mental disorders were assessed via SRQ-20. The internal consistency considered the items belonging to dimensions extracted by tetrachoric factor analysis for each study. Item homogeneity assessment compared estimates of Cronbach’s alpha (KD-20), the alpha applied to a tetrachoric correlation matrix and stratified Cronbach’s alpha. RESULTS The SRQ-20 dimensions showed adequate values, considering the reference parameters. The internal consistency of the instrument items, assessed by stratified Cronbach’s alpha, was high (> 0.80) in the four studies. CONCLUSIONS The SRQ-20 showed good internal consistency in the professional categories evaluated. However, there is still a need for studies using alternative methods and additional information able to refine the accuracy of latent variable measurement instruments, as in the case of common mental disorders.

Simultaneous identification of Trypanosoma cruzi surface and internal antigens reactive to different immunoglobulin classes (radio-immunoblotting)

A radioactive Western-blotting technique was developed by which the reactivity of Immunoglobulins (Igs) from different classes to both membrane radiolabelled and internal parasite antigens is simultaneously identified. The method includes radioiodination of parasites, polypeptide fractionation by SDS-PAGE, Western-blot transfer and autoradiography of the immunoblots developed with anti-Igs conjugates labelled with enzymes. The analysis is then performed by the comparison of common bands on the autoradiograms and the respective substrate stained nitrocellulose blots. This technique was used to analyse T. cruzi trypomastigote surface labelled antigens reactive to IgM, IgA and IgG specific antibodies. A different pattern of reactivity with acute Chagas' disease patients sera was thus obtained.

The Portuguese language version of social phobia and Anxiety Inventory: analysis of items and internal consistency in a Brazilian sample of 1,014 undergraduate students

OBJECTIVE: Theoretical and empirical analysis of items and internal consistency of the Portuguese-language version of Social Phobia and Anxiety Inventory (SPAI-Portuguese). METHODS: Social phobia experts conducted a 45-item content analysis of the SPAI-Portuguese administered to a sample of 1,014 university students. Item discrimination was evaluated by Student's t test; interitem, mean and item-to-total correlations, by Pearson coefficient; reliability was estimated by Cronbach's alpha. RESULTS: There was 100% agreement among experts concerning the 45 items. On the SPAI-Portuguese 43 items were discriminative (p < 0.05). A few inter-item correlations between both subscales were below 0.2. The mean inter-item correlations were: 0.41 on social phobia subscale; 0.32 on agoraphobia subscale and 0.32 on the SPAI-Portuguese. Item-to-total correlations were all higher then 0.3 (p < 0.001). Cronbach's alphas were: 0.95 on the SPAI-Portuguese; 0.96 on social phobia subscale; 0.85 on agoraphobia subscale. CONCLUSION: The 45-item content analysis revealed appropriateness concerning the underlying construct of the SPAI-Portuguese (social phobia, agoraphobia) with good discriminative capacity on 43 items. The mean inter-item correlations and reliability coefficients demonstrated the SPAI-Portuguese and subscales internal consistency and multidimensionality. No item was suppressed in the SPAI-Portuguese but the authors suggest that a shortened SPAI, in its different versions, could be an even more useful tool for research settings in social phobia.

Chylothorax after myocardial revascularization with the left internal thoracic artery

A 38-year-old male underwent coronary artery bypass grafting (CABG). A saphenous vein graft was attached to the left marginal branch. The left internal thoracic artery was anastomosed to the left anterior descending artery (LAD). The early recovery was uneventful and the patient was discharged on the 5th postoperative day. After three months, he came back to the hospital complaining of weight loss, weakness, and dyspnea on mild exertion. Chest X-rays showed left pleural effusion. On physical examination, a decreased vesicular murmur was detected. After six days, the diagnosis of chylothorax was made after a milky fluid was detected in the plural cavity and total pulmonary expansion did not occur. On the next day, both anterior and posterior pleural drainage were performed by videothoracoscopy, and prolonged parenteral nutrition (PPN) was instituted for ten days. After seven days the patient was put on a low-fat diet for 8 days. The fluid accumulation ceased, the drains were removed and the patient was discharged with normal pulmonary expansion.

Application of receptive music therapy in internal medicine and cardiology

OBJECTIVE: To investigate the effects of receptive music therapy in clinical practice. METHODS: Receptive music therapy was individually applied via musical auditions, including five stages: musical stimulation, sensation, situation, reflection, and behavioral alteration. Following anamnesis and obtainment of consent, patients answered a first questionnaire on health risk evaluation (Q1), and after participating in 16 weekly music therapy sessions, answered a second one (Q2). RESULTS: Two men and 8 women, aged above 18 years, referred to us due to symptoms of stress, emotional suffering, and the need to change lifestyles (health risk behavior) were studied between August 1998 and December 1999. Comparison between answers to Q1 and Q2, showed a trend (P=0.059) for reduction of ingestion of cholesterol-rich foods and for increased prospects in life with a tendency towards improvement, and also of increased intake of fiber-rich food (55.6%), increased levels of personal satisfaction (44.5%), and decreased levels of stress (66.7%). CONCLUSION: The study demonstrated decreased stress levels and increased personal satisfaction, higher consumption of fiber-rich food, lower cholesterol intake, and a better perspective on life, suggesting that receptive music therapy may be applied in clinical practice as an auxiliary therapeutic intervention for the treatment of behavioral health risks.