Considerations about the recommendations of the Commission on the Limits of the Continental Shelf on the Amazon fan

In 2004, Brazil submitted to the Commission on the Limits of the Continental Shelf (CLCS) a Submission for the outer limit of the Brazilian continental shelf for its extension beyond the limits of 200 nautical miles. In 2007, the CLCS presented its recommendations, however it did not recommend four areas proposed by Brazil, the Amazon Fan among them. The objective of this study is to present the main legal and technical aspects of the controversy about the Amazon Fan, in order to evaluate some alternatives for a future submission, new or revised.

Optimization of the Sybr Green real time PCR for the detection of Human Herpes Virus type 6 (HHV-6)

HHV-6 is the etiological agent of Exanthem subitum which is considered the sixth most frequent disease in infancy. In immuno-compromised hosts, reactivation of latent HHV-6 infection may cause severe acute disease. We developed a Sybr Green Real Time PCR for HHV-6 and compared the results with nested conventional PCR. A 214 pb PCR derived fragment was cloned using pGEM-T easy from Promega system. Subsequently, serial dilutions were made in a pool of negative leucocytes from 10-6 ng/µL (equivalent to 2465.8 molecules/µL) to 10-9 (equivalent to 2.46 molecules/µL). Dilutions of the plasmid were amplified by Sybr Green Real Time PCR, using primers HHV3 (5' TTG TGC GGG TCC GTT CCC ATC ATA 3)'and HHV4 (5' TCG GGA TAG AAA AAC CTA ATC CCT 3') and by conventional nested PCR using primers HHV1 (outer): 5'CAA TGC TTT TCT AGC CGC CTC TTC 3'; HHV2 (outer): 5' ACA TCT ATA ATT TTA GAC GAT CCC 3'; HHV3 (inner) and HHV4 (inner) 3'. The detection threshold was determined by plasmid serial dilutions. Threshold for Sybr Green real time PCR was 24.6 molecules/µL and for the nested PCR was 2.46 molecules/µL. We chose the Real Time PCR for diagnosing and quantifying HHV-6 DNA from samples using the new Sybr Green chemistry due to its sensitivity and lower risk of contamination.

Delimitation of kala-azar risk areas in the district of Vaishali in Bihar (India) using a geo-environmental approach

Remote sensing and geographical information technologies were used to discriminate areas of high and low risk for contracting kala-azar or visceral leishmaniasis. Satellite data were digitally processed to generate maps of land cover and spectral indices, such as the normalised difference vegetation index and wetness index. To map estimated vector abundance and indoor climate data, local polynomial interpolations were used based on the weightage values. Attribute layers were prepared based on illiteracy and the unemployed proportion of the population and associated with village boundaries. Pearson's correlation coefficient was used to estimate the relationship between environmental variables and disease incidence across the study area. The cell values for each input raster in the analysis were assigned values from the evaluation scale. Simple weighting/ratings based on the degree of favourable conditions for kala-azar transmission were used for all the variables, leading to geo-environmental risk model. Variables such as, land use/land cover, vegetation conditions, surface dampness, the indoor climate, illiteracy rates and the size of the unemployed population were considered for inclusion in the geo-environmental kala-azar risk model. The risk model was stratified into areas of "risk"and "non-risk"for the disease, based on calculation of risk indices. The described approach constitutes a promising tool for microlevel kala-azar surveillance and aids in directing control efforts.

Species diversity of sandflies (Diptera: Psychodidae) during different seasons and in different environments in the district of Taquaruçú, state of Tocantins, Brazil

Phlebotomine sandflies are the vectors for the protozoan parasites that cause leishmaniasis. The present study investigated the species composition of sandfly fauna in the rural district of Taquaruçú, municipality of Palmas, state of Tocantins, Brazil and compared the diversity of species among intradomicile, peridomicile and forest environments during the dry and rainy seasons. Sandflies were collected using CDC light traps over the course of three months during the dry and rainy seasons. A total of 767 specimens were captured, belonging to different 32 species. The most abundant species were Micropygomyia goiana (Martins, Falcão & Silva), Sciopemyia sordellii (Shannon & Del Ponte), Evandromyia carmelinoi (Ryan Fraiha, Lainson & Shaw), Evandromyia termitophila (Martins, Falcão & Silva), Nyssomyia whitmani (Antunes & Coutinho) and Lutzomyia longipalpis (Lutz & Neiva). The highest species diversity (30) and the greatest percentage of specimens (78.3%) were obtained during the rainy season. During the dry season, the species richness and abundance were greater in domestic environments. However, during the rainy season, the forest displayed the highest species richness and the domestic environment exhibited the greatest species abundance. Several important vector species are reported in this study.

The performance of semi-quantitative differential PCR is similar to that of real-time PCR for the detection of the MYCN gene in neuroblastomas

Amplification of the MYCN gene in neuroblastomas is a potent biological marker of highly aggressive tumors, which are invariably fatal unless sound clinical management is applied. To determine the usefulness of semi-quantitative differential PCR (SQ-PCR) for accurate quantification of MYCN gene copy number, we evaluated the analytical performance of this method by comparing the results obtained with it for 101 tumor samples of neuroblastoma to that obtained by absolute and relative real-time PCR. Similar results were obtained for 100 (99%) samples, no significant difference was detected between the median log10 MYCN copy number (1.53 by SQ-PCR versus 1.55 by absolute real-time PCR), and the results of the two assays correlated closely (r = 0.8, Pearson correlation; P < 0.001). In the comparison of SQ-PCR and relative real-time PCR, SQ-PCR versus relative real-time PCR concordant results were found in 100 (99%) samples, no significant difference was found in median log10 MYCN copy number (1.53 by SQ-PCR versus 1.27 by relative real-time PCR), and the results of the two assays correlated closely (r = 0.8, Pearson correlation; P < 0.001). These findings indicate that the performance of SQ-PCR was comparable to that of real-time PCR for the amplification and quantification of MYCN copy number. Thus, SQ-PCR can be reliably used as an alternative assay in laboratories without facilities for real-time PCR.