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Pollution-free loach seed breeding methods
**Foreword**
The standard "Pollution-free Loach" (DB33/561-2005) is divided into three parts:
- Part 1: Technical Specifications for Seed Breeding
- Part 2: Aquaculture Technical Specifications
- Part 3: Quality and Safety Requirements
This section corresponds to the first part of DB33/561-2005. It was proposed and managed by the Zhejiang Ocean and Fisheries Bureau. The section was drafted by the Zhejiang Provincial Institute of Freshwater Fisheries and the Xinchang County Fisheries Technology Promotion Station. The main contributors include Zhou Zhiming, Zhang Yuming, Hu Tingjian, and Huang Xiaohong.
**Introduction**
Loach is a highly nutritious fish with a protein content of up to 21% and notable medicinal benefits, making it a popular choice as a health food. However, due to overfishing and habitat degradation, natural loach populations have declined significantly. Artificial breeding has become the primary source for market supply, and aquaculture areas have expanded rapidly both within and outside the province. To ensure safe and sustainable loach production, this standard was developed to regulate breeding environments, promote aquaculture techniques, and guarantee product quality and safety.
**Pollution-free Loach: Technical Specifications for Seed Breeding**
**1 Scope**
This section outlines the environmental requirements, broodstock standards, breeding methods, and seedling cultivation techniques for loach (*Misgurnus anguillicaudatus*). It applies specifically to the breeding of loach seedlings.
**2 Normative References**
The following documents are referenced in this section and form its basis. For dated references, only the specified version applies. For undated references, the latest edition is applicable.
- GB 11607: Fishery Water Quality Standard
- GB/T 18407.4: Environmental Requirements for the Production of Pollution-free Aquatic Products
- NY 5051: Water Quality for Harmless Freshwater Aquaculture
**3 Environmental Requirements**
3.1 The production environment must meet the requirements of GB/T 18407.4.
3.2 The water source should be clean, abundant, and free from pollution. The water quality must comply with GB 11607.
3.3 Aquaculture ponds should be well-constructed, preferably cement or tri-soil ponds. Cement ponds should have a 15–20 cm layer of loam at the bottom. Soil ponds must not have sandy substrates. The pond walls should be made of cement or plastic, with a height of 100 cm. Inlet and outlet openings should be covered with wire mesh or nylon screens (0.15 mm mesh size, 20 mesh).
**4 Broodstock Requirements**
4.1 Broodstock should be collected from natural spawning areas or selected from the farm’s own stock. Individuals with clear growth advantages and no inbreeding should be chosen.
4.2 Broodstock should be 2–3 years old, with females weighing 40–100 g and males 20–50 g. They should exhibit typical germplasm characteristics, be healthy, active, and free from disease or injury.
4.3 Mature females typically have swollen abdomens, wide pectoral fins, and a rounded anal fin. Males have smaller abdomens, narrow pectoral fins, and visible testes. When pressed, milky white milt should be released. White spots on the abdomen indicate unsuitable individuals.
**5 Broodstock Breeding**
5.1 Stocking density should be 6–10 groups per square meter, with a 1:1 to 2 male-to-female ratio.
5.2 Feeding should occur twice daily in autumn, using animal-based feeds like minced meat, fish meal, and viscera, supplemented with plant-based feeds such as wheat bran and rice bran. Feed amounts should be 3–5% of body weight. During winter and early spring, feeding is suspended, and fermented pig manure is added. From late March, high-protein feed is introduced at 5–10% of body weight.
5.3 Water depth should be maintained at 40 cm or more, with regular water changes to keep the water clear and cool, with a transparency of 30 cm.
**6 Seed Breeding**
6.1 Induction
6.1.1 Spawning season begins in late April to mid-May when water temperature exceeds 22°C.
6.1.2 Oxytocin (HCG) is used at 500–700 IU per female, with half the dose for males.
6.1.3 Injection is administered into the back muscle or dorsal fin, avoiding the abdominal midline. A No. 4 needle is used, with 1–1.5 cm exposed. The injection angle should be 30°, and the depth should be 0.2 cm.
6.2 Spawning
6.2.1 Optimal water temperature for spawning is 22–28°C, with 25°C being ideal. At 22°C, eggs are laid after 14 hours; at 25–28°C, after 10 hours.
6.2.2 Fertilization
6.2.2.1 After natural fertilization, eggs are placed in cages with mesh or brown paper inducers. Each cage holds 50–80 broodstock pairs (1:2 to 3 ratio). Eggs are then moved to an incubator.
6.2.2.2 Artificial insemination involves gently squeezing the female's abdomen and collecting sperm from the male. Eggs and semen are mixed in a porcelain bowl with Galinger’s solution. After rinsing with 0.6–0.7% saline, fertilized eggs are placed in sterilized nests for incubation. Direct sunlight should be avoided.
6.3 Incubation
6.3.1 Fertilized eggs can be incubated in indoor or outdoor microflow water. Eggs are placed in bamboo baskets and submerged in water.
6.3.2 Dissolved oxygen in incubation water should be at least 6 mg/L, meeting NY 5051 standards.
6.3.3 Incubation density should be 150–200 × 10ⴠparticles per cubic meter.
6.3.4 Hatching time varies with temperature: 40–44 hours at 22–25°C, and 24–28 hours at 25–28°C. Three days after hatching, fry change color from black to light yellow and are ready for seedling cultivation.
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