Effectiveness of the Air Stripping in Two Salmonid Fish, Rainbow Trout (Oncorhynchus Mykiss) and Brown Trout (Salmo Trutta Morpha fario)

Summary

The principal aim of this study is to standardize and test the pneumatic method (air stripping) of collecting eggs in rainbow trout and brown trout. This method allows effective and simple collection of the eggs without the necessity of fish abdomen massage.

Abstract

Egg collection is one of the most crucial procedures during fish reproduction in salmonid hatcheries. Classic methods involve the use of hand massage on fish abdomens to expel the eggs. An alternative method uses the pressure of gas injected into the body cavity, which causes the subsequent release of the eggs. This method is believed to have less negative effects on both the welfare and egg quality of the broodstocks. Herein, we compare the results of air and hand stripping methods with respect to one-year survival and egg quantity and quality in two salmonid fish, rainbow trout (Oncorhynchus mykiss) and brown trout (Salmo trutta morpha fario). Our results indicate that air stripping yielded a better quality of eggs and higher one-year survival rate in rainbow trout. In addition, air stripping resulted in lower mortality rate than the group subjected to hand stripping (25% vs. 35%). The pH and hatching rate of the hand stripped group was lower than those of the air stripped group. In the case of brown trout, the quality of eggs obtained by both hand and air-stripping methods was similar; however, the one-year losses in fish were higher in air stripped group (15% compared to 0% in hand stripped fish). Although the advantages of air stripping method over hand stripping in terms of egg quality might not be observed in all salmonid species, the air-stripping procedure might be a promising option to be adopted in hatcheries as it ensures a high level of reproducibility and efficiency.

Introduction

The rainbow trout (Oncorhynchus mykiss) and brown trout (Salmo trutta morpha fario) belong to the Salmonidae family of Salmoniformes¹. The production of these two species in global aquaculture has gained rapid growth, owing to their commercial and recreational importance. In Poland, production of salmonid fish stands at around 20 000 tonnes, with rainbow trout being the dominant species. On the other hand, the brown trout represents a major source of freshwater fish resources in Europe because of its commercial value in aquaculture and importance for angling. In many aquatic ecosystems, the existence of brown trout populations is threatened. As a result, artificial reproduction has been applied to restock local populations of this species².

Rainbow trout and brown trout attain sexual maturity usually at the age of three, with males maturing a year earlier than females¹. Under artificial conditions, eggs and sperm of rainbow trout and brown trout are usually collected by gentle abdominal massage. In practice, on a small scale, a hand-stripping method of collecting eggs is cost-effective. However, on a large scale, the hand-stripping method may be labor-intensive and exhausting. As a result, this technique can result in broken eggs due to the weariness of the hatchery workers. Breakage of eggs often leads to a decrease in the pH of ovarian fluid and the release of egg yolk, both of which negatively affect the fertilization of eggs³. Furthermore, this traditional method of egg collection (repeatedly pressing of the abdomen) may also result in broodstock mortality caused by skin infection associated with the deterioration of the protective barrier of mucus.

Pneumatic method of egg collection was first used in Australia in 1957⁴. The method is based on the injection of gases (air, nitrogen or oxygen) into the body cavity to expel eggs using gas pressure. This fast and easy technique has been successfully applied in salmonid fish without any negative side effects on the fish⁴. Recently, this method was used in wild species (Northern pike, Esox lucius) to collect mature eggs in artificial conditions⁵. It was shown that air stripped eggs had higher fertilization and hatching rates in comparison with hand stripped eggs, regardless of the solution used for fertilization (Woynarovich solution, Billard buffer or hatchery water)⁵.

The principal aim of this study was to test the pneumatic method (air stripping) of collecting eggs in rainbow trout and brown trout in comparison with traditional hand stripping method. Efficiencies of both methods (volume of obtained eggs and time of collection, quality of gametes (ovarian fluid pH), as well as hatching rate and post-spawning mortality) were compared.

Protocol

Procedures were carried out in accordance with the Local Committee on the Ethics of Animal Experiments in Olsztyn, Poland.

1.Equipment Preparation

1. Use a syringe and needle for injection of the gas into the fish body cavity.
2. Set the gas pressure at 0.5 bar, and maintain this pressure to be below 1 bar throughout.
3. Control the air flow. For the salmonid fish, 1.5 L/min is the most efficient speed of gas exhaustion. Maintain the optimum speed because higher speed might not obtain all the eggs while a slower speed might unnecessarily prolong the procedure.
4. Connect the syringe with a screwed needle (0.8-1.2 mm diameter) to avoid needle detachment caused by the gas pressure.

2. Air Spawning

1. Anesthetize the fish with 0.2% Propiscin. Place the fish into a water bath with the 0.2% Propiscin and wait until the fish becomes unresponsive.
2. Carefully dry out before spawning by wiping with a dry cotton cloth.
3. Place the fish on the slant bed about 35-40°, on her side tail side down. With a gentle push of fish abdomen, check the egg flow from the genital tract. If the eggs are not present, the fish should not be spawned.
4. Place a container covered with mesh below the fish papilla to collect the eggs.
5. Quickly insert the needle below the abdominal fins.
6. Freely remove the eggs by the gas until only singular eggs are removed from the fish body.
7. Gently massage the abdomen to remove excess gas from the fish body.
8. After spawning, gently transfer the fish to a tank with water flow. Fish released to tank after air spawning might initially swim on the surface of the water. After several minutes the fish releases the remaining gas while swimming and starts swimming normally.
   NOTE: The obtained eggs are very clean and free from blood or feces contamination.

Results

Mature males and females of rainbow trout (age 2+, 1700 ± 328 g) and brown trout (age 3+, 1900 ± 435 g) were obtained from the Salmonid Fish Hatchery (Rutki-Żukowo, Inland Fishery Institute in Olsztyn, Poland). The males and females were placed in separate tanks with a volume of about 5 m³. The water temperature was 12±1 °C. Before manipulation, fish were anesthetized with 0.2% Propiscin. The fish were divided into two groups. Part of them underwent a traditional m...

Discussion

The pneumatic method of egg collection from fish, although more time-consuming than the traditional method (hand stripping), can secure a high quality of matured oocytes (this work). This is related to the low risk of mechanical disruption of the eggs while applying this procedure. High pH of the ovary fluid, as well as an increase in the fertilization rate, attest to the usefulness of the pneumatic method. Overall, the air stripping method of egg collection in fish reproduction could be a promising option in terms of st...

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Glycine pure P.A.	AVANTOR	527560117	sperm activating buffer
Trizma base	Sigma Aldrich	T1503	sperm activating buffer
Sodium chloride Bioreagent	Sigma Aldrich	S5886	sperm activating buffer
Calcium chloride anhydrous	Sigma Aldrich	C4901	sperm activating buffer
Propiscin (0.2% etomidate solution)	IRS Olsztyn, Poland	not indicated
compressor	Thomas  Sheboigan WI USA	DT/SR 070800001882
reduction valve	Camozzi	cRJUS U7J
air hose	ZEC T.P.U.	SH.98
syrgine	EFD	7012118
Air spawning stage	Biopasz, Poland	PNEU001
Orion Ross Ultra electrode	Thermo Scientific, Waltham, MA, USA	8102BNUWP