Toxic Effects of Methylene Blue on the Growth, Reproduction and Physiology of Daphnia magna

Abstract

Methylene blue (MB) is a disinfectant used in aquaculture to prevent and treat fish diseases. However, the release of MB can pose a risk to the receiving water bodies. Zooplankton are the most sensitive organisms among aquatic life. Hence, this study examined the acute and chronic toxic effects of MB on zooplankton using Daphnia magna (D. magna) as a test organism to provide basic data for risk assessment. The results show that 48 h-EC₅₀ and 24 h-LC₅₀ were 61.5 ± 2.3 and 149.0 ± 2.2 μg/L, respectively. Chronic exposure to MB affected the heart rate, beat frequency of the thoracic limbs, and reproductive ability of D. magna at environmental concentrations higher than 4.7 μg/L. The cumulative molts, time to production of the first brood, and total number of living offspring were affected at different MB concentrations, while "abortions" were observed in high-exposure groups. The activity of superoxide dismutase was increased, while glutathione S-transferase activity was stimulated at low concentrations and inhibited at high concentrations. In addition, the malondialdehyde content increased with increasing concentrations of MB. Our findings demonstrate the impact of MB on the reproduction and growth of freshwater species, as well as their physiological responses. These results have implications for establishing guidelines on the use of MB in aquaculture and setting discharge standards.

Keywords: antimicrobial dyes; life table parameters; oxidative damage; zooplankton.

Figure 1

Trend of immobilization rate fitted by Hill function in acute toxicity tests. Means and standard errors are shown.

Figure 2

Trend of mortality rate fitted by Hill function in acute toxicity tests. Means and standard errors are shown.

Figure 3

Damage to D. magna caused by MB in acute toxicity tests: (A) swollen and whitish carapace; (B) blue intestines; (C) swollen carapace and blue intestines; (D) carapace stuck with blue substance; (E) swollen carapace and partial loss of thoracic limb; (F) blue substance sticking to second antenna; (G) body turned blue throughout, with swollen carapace and holes; (H) swollen carapace and blue substance sticking to thoracic limb; (I) blue deposits in intestines and thoracic limbs.

Figure 4

Effect of MB on (a) heart rate and (b) thoracic limb activity of D. magna (mean ± SE, n = 10); significant differences (p < 0.05) between different treatment groups and the control group are indicated by different letters.

Figure 5

Effects of MB on (a) heart rate and (b) thoracic limb activity of D. magna after fitting with the Log3P1 curve. Means and standard errors are shown.

Figure 6

Effect of MB on (a) body length and (b) morphology of D. magna in each group at 21 days (mean ± SE, n = 10); significant differences (p < 0.05) between different treatment groups and the control group are indicated by different letters.

Figure 7

Influences of MB on (a) number of molts, (b) time to production of first brood, (c) number of first brood, (d) total number of living offspring, (e) total number of broods, and (f) total number of living offspring per brood (mean ± SE; n = 10). Significant differences (p < 0.05) among groups are indicated by different letters.

Figure 8

Cumulative number of living offspring over a 21-day period. Means and standard errors are shown.

Figure 9

(a) Control group of D. magna and aborted eggs in groups exposed to (b) 15 and (c) 26.7 μg/L observed under a stereomicroscope (3×).

Figure 10

Intrinsic rate of population increase of D. magna (mean ± SE; n = 10).

Figure 11

Effects of MB on (a) SOD activity, (b) GST activity, and (c) MDA content of D. magna (mean ± SE; n = 3; * p < 0.05 between the different treatment groups and the control group; ** p < 0.01 between the different treatment groups and the control group).