Molecular method for the characterization of Coxiella burnetii from clinical and environmental samples: variability of genotypes in Spain

[EN] Background. Coxiella burnetii is a highly clonal microorganism which is difficult to culture, requiring BSL3 conditions for its propagation. This leads to a scarce availability of isolates worldwide. On the other hand, published methods of characterization have delineated up to 8 different genomic groups and 36 genotypes. However, all these methodologies, with the exception of one that exhibited limited discriminatory power (3 genotypes), rely on performing between 10 and 20 PCR amplifications or sequencing long fragments of DNA, which make their direct application to clinical samples impracticable and leads to a scarce accessibility of data on the circulation of C. burnetii genotypes. Results: To assess the variability of this organism in Spain, we have developed a novel method that consists of a multiplex (8 targets) PCR and hybridization with specific probes that reproduce the previous classification of this organism into 8 genomic groups, and up to 16 genotypes. It allows for a direct characterization from clinical and environmental samples in a single run, which will help in the study of the different genotypes circulating in wild and domestic cycles as well as from sporadic human cases and outbreaks. The method has been validated with reference isolates. A high variability of C. burnetii has been found in Spain among 90 samples tested, detecting 10 different genotypes, being those adaA negative associated with acute Q fever cases presenting as fever of intermediate duration with liver involvement and with chronic cases. Genotypes infecting humans are also found in sheep, goats, rats, wild boar and ticks, and the only genotype found in cattle has never been found among our clinical samples. Conclusions: This newly developed methodology has permitted to demonstrate that C. burnetii is highly variable in Spain. With the data presented here, cattle seem not to participate in the transmission of C. burnetii to humans in the samples studied, while sheep, goats, wild boar, rats and ticks share genotypes with the human population.

Direct estimation of the backward flow

[EN] The aim of this work is to propose a new method for estimating the backward flow directly from the optical flow. We assume that the optical flow has already been computed and we need to estimate the inverse mapping. This mapping is not bijective due to the presence of occlusions and disocclusions, therefore it is not possible to estimate the inverse function in the whole domain. Values in these regions has to be guessed from the available information. We propose an accurate algorithm to calculate the backward flow uniquely from the optical flow, using a simple relation. Occlusions are filled by selecting the maximum motion and disocclusions are filled with two different strategies: a min-fill strategy, which fills each disoccluded region with the minimum value around the region; and a restricted min-fill approach that selects the minimum value in a close neighborhood. In the experimental results, we show the accuracy of the method and compare the results using these two strategies.