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This study was carried out to assess the in vitro quality of canine semen frozen in an ultrafreezer at -152 degrees C and to evaluate the male-to-male variation of frozen semen in five male dogs of the Canarian Mastiff breed. Four ejaculates of each dog were processed individually (5% glycerol and 0.5% Equex) to reach a final concentration of 100 x 10(6) spermatozoa/ml. Then, two freezing techniques were tested to assess the seminal quality (sperm motility, live spermatozoa and abnormal sperm cell percentages) at 1, 30, 60, 120 and 360 days after freezing: (i) semen was frozen and stored in liquid nitrogen; (ii) semen was frozen and stored in the ultrafreezer at -152 degrees C. After freezing-thawing, both freezing protocols showed no significant differences in sperm motility and the percentages of live and abnormal spermatozoa. On the other hand, the microscopic characteristics of spermatozoa in fresh semen were practically similar among males; however, after the semen processing and freezing, significant differences were observed (p < 0.05) among males, especially as regards sperm motility. This inter-individual variability was detected in both freezing protocols, showing that the male-to-male variation in the seminal quality post-freezing was independent of the freezing technique used. The in vitro results obtained in the Canarian Mastiff breed confirmed that the use of ultra-freezers at -152 degrees C is a potential alternative to liquid nitrogen for storing canine semen for long periods of time.

This study investigated the in vitro and in vivo efficacy of trichlorfon against Dawestrema cycloancistrium, as well as its physiological effects on arapaima. Naturally parasitized arapaima gill arches were exposed in vitro to 100, 250, 500 and 750 mg/L of trichlorfon and a control group (only distilled water), in triplicate. Parasites were monitored, and mortality was used to determine the median effective concentration (EC 50 ). The 750 mg/L concentration demonstrated 100% in vitro efficacy against D. cycloancistrium after 60 min, while the intermediate (500 mg/L) and the lowest (100 and 250 mg/L) tested concentrations were completely efficient after 90 and 130 min, respectively. The EC 50-1h of trichlorfon for D. cycloancistrium was deter-

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Infection by the acantocephalan Neoechinorhynchus buttnerae is considered one of most important concerns for tambaqui fish (Colossoma macropomum ) production. Treatment strategies have been the focus of several in vivo studies; however, few studies have been undertaken on in vitro protocols for parasite maintenance. The aim of the present study was to develop the best in vitro culture condition for N. buttnerae to ensure its survival and adaptation out of the host to allow for the testing of substances to be used to control the parasite. To achieve this, parasites were collected from naturally infected fish and distributed in 6-well culture plates under the following treatments in triplicate: 0.9% NaCl, sterile tank water, L-15 Leibovitz culture medium, L-15 Leibovitz + agar 2% culture medium, RPMI 1640 culture medium, and RPMI 1640 + agar 2% culture medium. The plates containing the parasites were maintained at 24 °C, 28 °C, and 32 °C. The RPMI 1640 + agar 2% culture medium showed the best survival of 24 days at 24 °C. No body alterations such as swollen parasites, body deformation, dehydration and hardening were observed in the RPMI 1640 + 2% culture medium.

In the aquatic environment, fish are subject to different pollutants and environmental variations that favour stress on animals, making them vulnerable to pathogens. Fish parasites are excellent biological indicators of environmental quality, as their parasitic population may increase or decrease in response to changes in water parameters and/or the presence of pollutants. Ectoparasites are considered excellent indicators of environmental quality because they have a monoxenic life cycle with high reproductive rates that have been responsible for the immediate response to changes in aquatic biota. In turn, parasites with a heteroxenous life cycle are also responsive to environmental variation because, with the mortality of intermediate hosts, their population can be easily changed. However, studies with endoparasites and their relation with the environment are more numerous, as they are accumulators of pollutants, especially trace elements, either via fish or through stages of development outside the host that also contribute to the accumulation of metals. Moreover, Digenea, Acanthocephala, Nematoda and Cestoda parasites can compete for trace elements that are bioavailable in the environment, as fish with a greater abundance of parasites have a lower concentration of toxic metals in their tissues. Furthermore, they have a high affinity for trace elements that are toxic to human and fish health (arsenic, mercury, lead and cadmium). This review is a comprehensive overview of the use of fish parasites as environmental sentinels and describes their potential as bioindicators of effect and accumulation, and evaluates their application in fish farms environments.

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