Development of an indirect immunofluorescence technique for the diagnosis of toxoplasmosis in bottlenose dolphins.

The diagnosis of toxoplasmosis is often complicated by the lack of specific clinical symptoms or postmortem features, in humans and other animals. The only diagnostic test described so far for the serological diagnosis of Toxoplasma gondii in marine mammals is the modified agglutination test (Dubey et al., Am J Vet Res 48(8):1239-1243, 1987). The development of more sensible and specific immunological techniques requires specific antibodies, which are currently unavailable in the scientific market. Indirect immunofluorescence (IIF) is one of the most widely used methods for the diagnosis of toxoplasmosis in humans (Auer et al., Parasitol Res 12:965-970, 2000). In order to develop and apply this technique to the bottlenose dolphin (Tursiops truncatus), immunoglobulins were firstly purified using ion-exchange chromatography. The purified immunoglobulins were then injected in New Zealand rabbits in order to obtain polyclonal antibodies. These antisera were validated by the IIF technique, using as controls serum samples of dolphins infected by Toxoplasma. The results were visualized using antirabbit IgG labeled with fluorescein. This newly developed and specific serological assay was then tested with the dolphin collection of Loro Parque, Tenerife, Spain (group I), and L'Oceanogràfic of Valencia, Spain (group II). The obtained results in this study showed that none of the dolphins from group 1 were infected by T. gondii and two animals were positive in group 2. Furthermore, we conclude that this study has produced antibodies with high specificity against dolphin immunoglobulins and an IIF method which may be used as immunological diagnostic tools, especially for the serological diagnosis of toxoplasmosis.

A DEVH-box RNA Helicase from Leishmania braziliensis is Associated to mRNA Cytoplasmic Granules.

RNA helicases are ubiquitous enzymes that participate in almost all aspects of RNA processing, including RNA and RNA-protein complex remodelling. In trypanosomatids, which post-transcriptionally regulate gene expression, the formation of different kinds of ribonucleoprotein granules under stress conditions modulates the parasite's RNA metabolism. This paper describes the isolation of a putative DEVH-box RNA helicase produced by promastigotes of Leishmania braziliensis. Using a Cy3-labelled dT30 oligo, FISH showed the localization of this protein to mRNA granules under starvation stress conditions. The central region of the protein was shown to be responsible for this behaviour.

Estimating statistical uncertainty of Monte Carlo efficiency-gain in the context of a correlated sampling Monte Carlo code for brachytherapy treatment planning with non-normal dose distribution.

Correlated sampling Monte Carlo methods can shorten computing times in brachytherapy treatment planning. Monte Carlo efficiency is typically estimated via efficiency gain, defined as the reduction in computing time by correlated sampling relative to conventional Monte Carlo methods when equal statistical uncertainties have been achieved. The determination of the efficiency gain uncertainty arising from random effects, however, is not a straightforward task specially when the error distribution is non-normal. The purpose of this study is to evaluate the applicability of the F distribution and standardized uncertainty propagation methods (widely used in metrology to estimate uncertainty of physical measurements) for predicting confidence intervals about efficiency gain estimates derived from single Monte Carlo runs using fixed-collision correlated sampling in a simplified brachytherapy geometry. A bootstrap based algorithm was used to simulate the probability distribution of the efficiency gain estimates and the shortest 95% confidence interval was estimated from this distribution. It was found that the corresponding relative uncertainty was as large as 37% for this particular problem. The uncertainty propagation framework predicted confidence intervals reasonably well; however its main disadvantage was that uncertainties of input quantities had to be calculated in a separate run via a Monte Carlo method. The F distribution noticeably underestimated the confidence interval. These discrepancies were influenced by several photons with large statistical weights which made extremely large contributions to the scored absorbed dose difference. The mechanism of acquiring high statistical weights in the fixed-collision correlated sampling method was explained and a mitigation strategy was proposed.