Severity Classification of Clinical Signs and Defining the Moribund State as an Experimental Endpoint for Acute Toxicity Testing Using Japanese Medaka (Oryzias latipes).

Use of the moribund state as an endpoint for acute toxicity testing in fish is complicated by the lack of consensus about the sublethal signs that define the state or that can predict imminent death. Defining the moribund state in fish requires determination of the association between each observable sublethal sign and transition to imminent death. To establish a criterion for defining the moribund state in Japanese medaka (Oryzias latipes), we determined death/clinical-sign ratios, defined as the proportion of fish with each clinical sign that transitioned to imminent death, for 13 clinical signs observed in individually housed fish exposed to one of two typical toxicants, three pharmaceuticals, two pesticides, and one metal. Also, the time from onset of each clinical sign to transition to imminent death (referred to as the survival time) was determined by continuous monitoring. Three of the observed clinical signs-immobility, lethargy, and immobility at the surface-were found to be indicators of the moribund state, with each of these signs having a death/clinical-sign ratio of 1.0. Evaluation of the survival time after onset of the other 10 signs enabled determination of whether timely euthanasia would be appropriate, thereby providing a means of reducing the suffering of laboratory fish in the period before death. Environ Toxicol Chem 2022;41:1089-1095. © 2022 SETAC.

Effects of oxygenated polycyclic aromatic hydrocarbons on the early life stages of Japanese medaka.

Oxygenated polycyclic aromatic hydrocarbons (oxyPAHs) are directly discharged into the atmosphere with exhaust gas from diesel engine automobiles and industry and are also generated by photo-oxidation and/or microbial metabolism of parent polycyclic aromatic hydrocarbons (PAHs). They are widely distributed in environment, especially around urban areas. We evaluated the effects of exposure to oxyPAHs as acenaphthenequinone (ANQ), 7,12-benz(a)anthracenequinone (BAQ), 1,4-naphthoquinone (NAQ), and 9,10-phenanethrenequinone (PHQ) in Japanese medaka (Oryzias latipes) embryos. ANQ (>2720 µg/L) caused caving in the yolk sac, and BAQ (>22.4 µg/L) caused tanning of the oil droplet. Additionally, NAQ (>314 µg/L) and PHQ (>734 µg/L) stopped development of medaka embryos beginning 2 days after the start of exposure. The hatched larvae from embryos exposed to ANQ, BAQ, NAQ, or PHQ at 518, 9.86, 36.5, or 80.7 µg/L, respectively, exhibited incomplete development of the cephalic region, incomplete palate, unabsorbed and hypertrophied yolk sac, tubular heart, altered axial curvature, and poor swimming ability. These symptoms were similar to those observed in blue sac disease, which is caused by exposure to PAHs.

Toxic evaluations of sediments in Tokyo Bay, Japan, using Japanese medaka embryos.

Toxic risks of sediments collected from seven sites in Tokyo Bay were evaluated using Japanese medaka embryos. Those sediments with slight pore water were placed in grass petri dishes without overlying water. The most remarkable effect in the field sediment was to cause hatching delay in embryos, and the longest time until hatching took was 12.5 ± 1.6 days post-fertilization (dpf), although that in control group was 10.1 ± 0.7 dpf. A significant delay in hatching was observed at four sites. Because total carbon concentrations were relatively high in sediments at three of these four sites, several chemicals were expected to be residues in these sites and could cause their delay. Although extreme mortality was not observed at all sites, sediments collected from the site close to Kawasaki city induced 10 % mortality. Polycyclic aromatic hydrocarbon (PAH) concentrations were remarkably high at this site compared with other sites, and thus PAH toxicities could be causing the mortality. Concentration of heavy metals such as cadmium, copper, lead, and zinc in sediments were also determined, but no clear relationship was found between toxicities to embryos and the distribution of their concentrations.