The sperm motility pattern in ecotoxicological tests. The CRYO-Ecotest as a case study.

Changes in environmental stressors inevitably lead to an increasing need for innovative and more flexible monitoring tools. The aim of this work has been the characterization of the motility pattern of the cryopreserved sea bream semen after exposure to a dumpsite leachate sample, for the identification of the best representative parameters to be used as endpoints in an ecotoxicological bioassay. Sperm motility has been evaluated either by visual and by computer-assisted analysis; parameters concerning motility on activation and those describing it in the times after activation (duration parameters) have been assessed, discerning them in terms of sensitivity, reliability and methodology of assessment by means of multivariate analyses. The EC50 values of the evaluated endpoints ranged between 2.3 and 4.5ml/L, except for the total motile percentage (aTM, 7.0ml/L), which proved to be the less sensitive among all the tested parameters. According to the multivariate analyses, a difference in sensitivity among "activation" endpoints in respect of "duration" ones can be inferred; on the contrary, endpoints seem to be equally informative either describing total motile sperm or the rapid sub-population, as well as the assessment methodology seems to be not discriminating. In conclusion, the CRYO-Ecotest is a multi-endpoint bioassay that can be considered a promising innovative ecotoxicological tool, characterized by a high plasticity, as its endpoints can be easy tailored each time according to the different needs of the environmental quality assessment programs.

Ecotoxicological effects of sediments from Mar Piccolo, South Italy: toxicity testing with organisms from different trophic levels.

The Mar Piccolo of Taranto (Ionian Sea, Southern Italy) is a semi-enclosed and strongly polluted basin. For decades, it has been subjected to different anthropogenic impacts. These stressors caused severe sediments contamination with high concentration of different pollutants (PAHs, PCB, heavy metals). In order to assess the current status of sediments contamination, an ecotoxicological investigation combined with chemical analysis (heavy metals, PAH, and PCB) has been performed. In order to derive ecologically relevant conclusions, a multiorganisms and multiend-points approach has been applied, exposing organisms from different trophic levels to elutriate and whole sediment. The battery of bioassays consists of a microalgal growth inhibition test (Dunaliella tertiolecta), acute and sublethal assays (end-points: mortality, immobilization and swimming speed alteration) on crustaceans larvae and juveniles, and rotifers (Amphibalanus amphitrite, Artemia salina, Corophium insidiosum and Brachionus plicatilis), and embryotoxicity test on echinoderms (Paracentrotus lividus). Considering the high levels of sediment contamination highlighted from chemical analysis, an unexpected very low toxic effect was observed, even considering the sublethal end-point (larval swimming speed alteration). The results of this study suggest a very complex contaminants dynamic in the Mar Piccolo sediments that, despite a strong level of contamination, seems to not affect in a proportional manner the biological compartment.

Development of a multiplex PCR for the simultaneous amplification and genotyping of glycoprotein N among human cytomegalovirus strains.

Genomic variation among human cytomegalovirus (HCMV) strains is probably involved in HCMV-induced pathogenesis. The envelope glycoprotein N (gN) showed extensive genetic polymorphism as HCMV isolates have been clustered into four distinct gN variants (gN-1, gN-2, gN-3, gN-4) whose distribution has been analyzed worldwide using different methodological approaches (PCR-RFLP, PCR-Cloning, PCR-Sequencing). This paper describes a new method for concurrent detection of gN genotypes among HCMV strains using a multiplex gN-variants specific PCR plus visualization on agarose gel, avoiding subsequent steps such as cloning, restriction or sequencing. This novel approach will reduce costs and shorten the detection time of gN polymorphisms among HCMV clinical isolates.