First report of Urosporidium sp., a haplosporidian hyperparasite infecting digenean trematode Parvatrema duboisi in Manila clam, Ruditapes philippinarum on the west coast of Korea.

In this study, we first report on the occurrence of Urosporidium sp., a haplosporidian hyperparasite infecting the trematode, Parvatrema duboisi, which parasitizes Manila clams, Ruditapes philippinarum on the west and south coasts of Korea. The larval P. duboisi infected by the sporocyst stage of Urosporidium sp. demonstrated numerous small yellowish spores in their tissues. The heavily infected metacercariae exhibited degenerate bodies and the larvae were often motionless. Clams heavily infected by the metacercariae of P. duboisi also displayed abnormal golden spots on the mantle tissue. In histology, different life stages of Urosporidium sp. could be identified, including the uni-nucleate, plasmodial, sporogonic stages and the acid fast mature spores released from the cyst. In scanning electron microscopy (SEM), the mature spore exhibited a semi-circular rim around the apical end and the orifice was covered internally with a flap. Loop-like filaments ornamentation was also identified from Urosporidium sp. in SEM, suggesting that Urosporidium sp. found in this study is a new member in the genus. Prevalence of Urosporidium sp.-infected trematodes in this study ranged from 2.5% to 24.0% in April 2010 and the infection was observed from 8 sampling sites out of the 26 sites surveyed on the west and south coasts.

Electrochemical genosensors for the detection of Bonamia parasite. Selection of single strand-DNA (ssDNA) probes by simulation of the secondary structure folding.

A post-PCR nucleic acid work by comparing experimental data, from electrochemical genosensors, and bioinformatics data, derived from the simulation of the secondary structure folding and prediction of hybridisation reaction, was carried out in order to rationalize the selection of ssDNA probes for the detection of two Bonamia species, B. exitiosa and B. ostreae, parasites of Ostrea edulis. Six ssDNA probes (from 11 to 25 bases in length, 2 thiolated and 4 biotinylated) were selected within different regions of B. ostreae and B. exitiosa PCR amplicons (300 and 304 bases, respectively) with the aim to discriminate between these parasite species. ssDNA amplicons and probes were analyzed separately using the "Mfold Web Server" simulating the secondary structure folding behaviour. The hybridisation of amplicon-probe was predicted by means of "Dinamelt Web Server". The results were evaluated considering the number of hydrogen bonds broken and formed in the simulated folding and hybridisation process, variance in gaps for each sequence and number of available bases. In the experimental part, thermally denatured PCR products were captured at the sensor interface via sandwich hybridisation with surface-tethered probes (thiolated probes) and biotinylated signalling probes. A convergence between analytical signals and simulated results was observed, indicating the possibility to use bioinformatic data for ssDNA probes selection to be incorporated in genosensors.