Dead fries were observed for yellowfin bream among the three treatments and the control on the 3rd day; thereafter, the survival rates of all treatments gradually decreased with time. The survival rates of MI, TC and the control group were not significantly different in the whole period (p < 0.05), and the two marking methods did not raise the death rate compared with the control. However, the survival rate of MI was lower than that of the above three treatments throughout the whole period (p < 0.05) (Fig. 1).
The mark retention rate of marking skin by spraying fluorescent pigment (MS) can reach above 80% in salmon19, while it is under 60% in this study, which perhaps can be accounted for by inadequate spray gun air pressure19,36. But this method is not friendly to fish, therefore, this study decided not to use this method. The ineffective MI in this experiment may come from many unsuitable processes, such as solution concentration, immersion time, or the dye used. Similar low mark retention rates were observed in earlier studies16,37. Most likely, the TC is unsuitable for mass Tagging fry. Although the survival rate was the lowest, the mark retention rate was the highest among all treatments in this study. This study suggested that the immersion time of MI may be so long that the physiological function of the fry may be damaged38,39. In other words, MI can be a good and suitable marking method under a shorter immersion time for yellowfin bream.
For yellowfin bream, the survival rates of all treatments and the control were above 80%, except for MI of 0.2% treatment and the control on the 14th day of the whole period. The survival rates of MI of 0.2% treatment, 0.25% and the control were not significantly different except MI between the 2nd and 12th days (p > 0.05), so the two marking methods did not increase the death rate compared with the control. That of MI of 0.75% treatment was the highest, and TC was the lowest on the MI of 0.2% treatment 14th day (p < 0.05) (Fig. 2).
The mark retention rate of MI of 0.75% treatment was the highest, MI of 0.5% treatment was next, and both MI of 0.2% treatment and the control were lowest in the entire period (p < 0.05). Among them, MI of 0.25% treatment and 0.2% were not significantly different from the 2nd to 6th days (p > 0.05), while the former was higher than the latter between the 8th and 14th days (p < 0.05).
The high mark retention rates of MI of 0.75% treatment and 0.5% (above 80%) indicate that both methods were effective for the entire period. The medium initial mark retention rate of MI of 0.75% treatment, which survival rates dropped to near 0% during the experimental period, indicates that the method was unsuitable for mass marking even though it is inexpensive and easy to complete16 (Fig. 2).
The survival rates of MI were above 95% and higher than those of the other treatments and the control throughout the whole period (p < 0.05). This study suggests that tetracycline is an antibiotic that could enhance the fish resistance and increase survival rates.
The mark retention rate of TC was the highest among all the treatments and stayed at almost 100.0% in the whole period, that of MI was quickly down to 0% from the 3rd to the 15th day (Fig. 3). It is obvious that the marking method of TC was effective, and MI could be used but not well enough.
The mark retention rate of MS is above 80% and similar to earlier studies of salmon19,18. This similarity of high mark retention rates could come from yellowfin bream, which, like salmon, belong to small-scale species that are under the same spray gun air pressure. The survival rate of MI of 0.75% treatment was the lowest among the three treatments and the control in the whole period (p < 0.05). This study suggests that the immersion time may be so long that the physiological function of the fry may be impeded or influenced by yellowfin bream. The low mark retention rate of MD is similar to earlier studies16,37. Most likely, MI of 0.75% treatment is unsuitable for mass marking of fry in this study.
No dead fries were observed in either double marking or the control from the 1st to the 7th day (p > 0.05); thereafter, the survival rates of double marking were kept above 80% and higher than that of the control on the 9th and 15th days (p < 0.05) (Fig. 3). This study concludes that double marking by combining MI and TC would not increase the death rate of the samples compared with the control. The reason may come from the use of tetracycline, which can enhance the fry resistance.
Both tag and mark retention rates of TC and MI were both 100%, while the marker retention rate of the control period was 0% (Fig. 3). The TC and MI markers of all fry can be detected at the same time. In other words, the double marking method combining TC and MI is effective and is suitable for yellowfin bream fry.
All the MI of 0.2% treatment, 0.25%, and 0.5% survival rates of yellowfin bream were higher. The reasons can be attributed to improved techniques, including the immersion time of MI. The TC, and MI survival rates of yellowfin bream is relatively high. The reasons can be attributed to technical improvements, including the reduction of the MI soaking time from 120 minutes to 30 minutes. The conclusion of this study is that the soaking time of MI may have an optimal value; otherwise, too short a time will result in low mark retention of fry, and too long will affect the osmotic adjustment of fry38.
The retention of almost 100% of MI markers in yellowfin bream indicates that this method is suitable for fry. The double marking method applied to yellowfin bream in this study includes two fluorescent marking and tagging methods, which can be applied to fry population marking at the same time. In other words, the fluorescent dual marking method was used for the first time in fry marking research. In the future, when marked fry are released to increase fishery resources, even if the marking is lost, they can still be found and restored. In addition, the loss rate of each mark can be estimated4,40.