Axial vertebrae deformities, such as scoliosis, lordosis, kyphosis, and platyspondyly can occur in teleost fish^[1]. While rarely seen in wild fish, they are commonly observed in captive stocks^[2]. Biotic factors linked to bone deformities include nutrition, genetics, rearing density, handling, rapid growth rates, parasites, and infections^[3-8]. Abiotic factors include light intensity, water chemistry, water quality, water quantity, water velocity, and radiation^[1,9-11].

Water temperature is the most influential abiotic factor on skeletal development in teleost fish, with temperature fluctuations potentially hampering skeletal formation during egg incubation^[12,13]. In some salmonids rearing temperatures at high and low extremes can produce high gene expression levels that may ultimately affect skeletal formation^[6,14-16].

Substantial bone deformities have been observed in brook trout Salvelinus fontinalis eggs spawned at Crystal Lakes Trout Hatchery in Fortine, Montana, USA. Elevated egg incubation temperature was hypothesized to be a potential contributor to the relatively high incidence of bone deformity. Thus, the objective of this study was to examine the impact of incubation water temperature on the incidence of vertebral deformity in brook trout eggs.

Methods

Brook trout eggs were obtained from Crystal Lakes Trout Hatchery on October 31, 2018. After fertilization, eggs were water-hardened in lake water (4.4°C) for 30 min, rinsed, and transported in lake water to Giant Springs State Fish Hatchery, Great Falls, Montana, USA. Upon arrival at the hatchery, the eggs received a static treatment in a 100 mg/L active iodine solution for 10 min. They were then tempered to ambient incubation water temperature of 12.2°C at a rate of 1°C/min.

After tempering, the eggs were rinsed and divided into six, 100-egg aliquots for incubation in vertically stacked incubation trays (Marisource, Burlington, Washington, USA) receiving 18.5 L/min spring water (322 mg/L total hardness as CaCO₃; 176 mg/L Alkalinity as CaCO₃; 7.52 pH). Two incubation water temperature treatments were used (N=3): 9.4°C and 12.2°C. Water was chilled using Current’s Prime water chiller (model number, Vista, California, USA). To prevent saprolegnia infestations, eggs received flow-through treatments of 500 mg/L hydrogen peroxide every-other-day (Peroxiade, Chemical Montana, Helena, Montana).

Survival to the eyed stage of egg development was calculated based on the number of eggs alive on November 23, 2018 (484 daily temperature units) for eggs incubated at 12.2°C and November 28, 2018 (476 daily temperature units) for eggs incubated at 9.4°. After egg hatch, the number of hatched fry (alevins) was recorded. In addition, the number of alevins with skeletal deformities was noted. The following formulas were used:

Survival to Eye (%) = 100 x (number of eyed-eggs / initial number of eggs).

Survival to Hatch (%) = 100 x (number of alevins/ initial number of eggs).

Deformities (%) = 100 x (number of deformities/initial number of eggs).

Data Analysis

Data was analyzed using t-tests. Percentage data was arcsine square root transformed to stabilize the variances^[17]. Significance was pre-determined at P < 0.05.

Results

Skeletal deformity was significantly less in eggs incubated at 9.4°C compared to those incubated at 12.2°C. There were no significant differences in survival to eye-up or hatch between the incubation temperatures.

Table 1: Mean (± SE) percent spinal deformity and egg survival of brook trout eggs incubated at one of two temperatures (N=3).

Discussion

Incubation water temperature impacts metabolism and subsequent physiological development, dictating the rate of development in teleost fish^[18]. During incubation, eggs can be subjected to ambient rearing temperature variations which can impact individual physiological traits^[20-22]. The relatively lower bone deformity rates in the eggs incubated at 9.4°C in the current study supports the findings of^[20,21].

Temperature thresholds are species specific, with brook trout temperatures ranging from 1.7°C to 15.6°C. Wargelius et al^[21].found that warmer temperature changes at specific developmental periods can cause deformities of the spine in Atlantic Salmon Salmo salar by effecting sclerotomal cell patterning. Similarly, Titzsimmons and Perutz (2006) found that bone deformity prevalence and severity increased with warmer incubation temperatures in Atlantic cod Gadus morhua. Conversely, Okamura et al^[23] observed an opposite reaction in Japanese eel Anguilla japonica, where bone deformity rates decreased with increasing water temperatures. These results suggest that bone deformity rates could be minimized by finding the optimal incubation temperature for each unique fish species.

The results of this study show the benefit to incubating brook trout eggs in relatively cooler water. Although chilling incubation water may be cost prohibitive, it would increase the number of surviving progeny, maximizing their potential genetic contributions to the population. In other words, chilling water may maximize genetic diversity by reducing the number of malformed fish^[24]. Chilling eggs will also slow development and increase incubation times, potentially reducing the amount of time the fish will be on feed^[25]. Unless growth within a specific time period is the most important consideration, incubating brook trout eggs in cooler temperatures is recommended.

References

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