Safety Evaluation of a Compound Lonicera rupicola Hook.f.et & Thomson Injection

Haoyuan FENG Ling LI Yuhan LI Lu DAI Chaoxi CHEN

AbstractFrom 2015 to 2016, the hepatopancreatic lesion syndrome had a higher mortality rate, which caused great economic losses to farmers. In order to study the highefficiency diseasepreventing crab pond intercropping mode, an experiment was successfully conducted to intercrop Chinese sucker (Myxocyprinus asiaticus) in a crab pond in 2017. We optimized the traditional stocking mode, and mainly cultured river crabs when intercropping Chinese suckers. Meanwhile, the cultivation environment was strengthened, and the test pond was not affected by disease throughout the year. The results showed that the production benefits were river crab 115 620 yuan/hm2, Chinese sucker 44 280 yuan/hm2, and silver carp and bighead carp 5 418 yuan/hm2, with a total output of 165 318 yuan/hm2, and the outputinput ratio was 1∶1.72. The Chinese suckerintercropping river crab culture mode promoted the improvement of pond culture environment, effectively controlled the occurrence of hepatopancreatic lesion syndrome, and significantly improved the economic benefits of crab pond culture.

Key wordsEriocheir sinensis; Myxocyprinus asiaticus; Disease prevention model; Experimental study

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Received: July 26, 2018Accepted: October 21, 2018

Supported by Taizhou Science and Technology Support (Agriculture) Program (TN201707); Research Project of Jiangsu Agrianimal Husbandry Vocational College (NSF201614); Qing Lan Project for Talent Training in Colleges and Universities in Jiangsu Province; Taizhou Scientific Research Project for "311 Talent cultivation Engineering"; College Students Practice Innovation Training Program of Jiangsu Province ( 201812806022H).

Jiansheng YE (1981-), male, P. R. China, master, associate professor, devoted to research about economic aquaculture animal ecology and nutriology.

*Corresponding author.

River crab, with a scientific name of Eriocheir sinensis, is one of the most important freshwater economic crabs in China. It has strong adaptability to the environment, especially in Jiangsu, Anhui, Hubei, Shanghai and other areas along the Yangtze River. It is rich in nutrition and is fleshy. It is tender, delicious, suitable for people of all ages, and has a longstanding food culture. With the increasing demand for crab consumption at home and abroad, coupled with the continuous popularization of the concept of "Internet + Crab" online sales (purchasing), the culture area and culture scale have been expanding. Crab farming industry has also risen in areas such as Yunnan, Guizhou and Sichuan and achieved better economic benefit［1-4］, and the benefit of some pond have exceeded 300 000 yuan/hm2. Whether for the scale of river crab industry and brand building, or the integration of rural one, two, three industries and the construction of characteristic rural towns, crab industry has become Chinas worldfamous agricultural industry.

Chinese sucker (Myxocyprinus asiatius) belongs to Cypriniformes of Catostomidae. It grows fast and has delicious meat. It is kind of a traditional expensive economic fish, which has high crude protein content and moderate crude fat content, and is rich in nutrients including calcium, phosphorus and iron. The market value of Chinese sucker is extremely high, exceeding 120 yuan/kg, and the market development potential is huge. Raising Chinese sucker in river crab ponds can significantly improve comprehensive culture benefits.

At present, crab farming has weak market risk and resilience, and the breeding species is single. Its production potential has not been exerted. Most farmers only pay attention to consumers (river crabs) in production, but ignore decomposers and producers, resulting in seriouslyunbalanced ecological system［5-10］ and frequent disease. Especially in 2015-2016, the hepatopancreatic lesion syndrome of river crab in Jiangsu and surrounding provinces (there is no unified name, commonly known as "Shuibiezi" disease) had a high incidence and caused a high mortality rate, causing a large economic loss in some crab ponds. In the study, we optimized and innovated the traditional crabraising model, optimized the stocking mode and adopted a complete set of techniques. On the one hand, we promoted the improvement of pond ecological environment by intercropping Chinese sucker, and created a good environment for healthy growth of river crab. In species selection, silver carp was selected to improve the water quality, and promote the material circulation of the pond. On the other hand, the dissolved oxygen (DO) regulation was strengthened by adopting the composite aeration mode of "microporous aeration + impeller aeration" to accelerate the decomposition of organic matter in the pond, which reduced the "oxygen debt" at the bottom of the pool, reduced the use of chemicals, and reduced the metabolic load of the liver and pancreas on drugs and the environmental stress response.

Materials and Methods

Test location

The experiment was carried out at the Jiangsu Crab Breeding Base of Jiangsu Banqiao Aquatic Products Co., Ltd., in Liuge Development Zone, Yongfeng Town, Xinghua City. The experimental crab pond had an area of 3.73 hm2, and soil in the pond was clay. The pond was in eastwest direction, nearly square, with a slope ratio of 1∶2.5 and a water depth of 1.0-1.5 m. The drainage system of the crab pond was independent, with inlet and outlet ports disposed at the two ends of the pond. The irrigation and drainage were convenient. The water exchange channel outside the pool contained sufficient water, which was free of no pollution, and had good water quality.

Preparation of crab pond

Cleaning of crab pond

After the crab production in November, 2016, the crab pond water was first drained with a water pump to remove excess sludge from the bottom of the pond; lime water was evenly splashed all over the pond at 1 800 kg/hm2 to fully kill various pests and pathogens in the crab pond, followed by 30-50 d of exposure. Finally, in sunny days of the winter, weeds around the pond were removed, and holes and cracks in the pool wall were filled.

Preparation of crab pond

The test crab pond was surrounded by white calcium plastic boards, forming an antiescape wall with arcshaped corners, buried in the soil by 0.1 m, with a wall height of 0.6 m. The wall was slightly inclined to the inside of the pool which was smooth. The crab pond was excavated with a 0.6 m deep ring ditch (deep water area) accounting for 30% of the total area of the pond; and in the middle of the pond, a "crab island" (shallow water area) was built, accounting for 70% of the total pond area; and the ring ditch of the crab pond was equipped with a microporous aerator according to 2.25 kW/hm2, and the crab island in the middle of the crab pond was equipped with an impeller type aerator according to 3.75 kW/hm2.

Culture of water plants and input of aquatic snails

Culture of water plants

Before March 2017, 15 d after cleaning and preparation of the pond, the pond was filled with water with a depth of 40 cm. Elodea was transplanted on the shallow slope, forming a shape of the Chinese character with the meaning the same as "well"; and tape grass seeds were sown in the middle of the crab pond and protected by fence facilities to prevent crabs from entering the tape grass planting area too early, and the fence was dismantled when the grass grew to 10 cm.

After planting of aquatic plants, fertilization and water supply were well done in spring at a low temperature, to cultivate singlecell beneficial diatoms. Due to the low temperature, the diatom species and medium were used repeatedly in the ring ditch to keep the water color brown and the transparency of 30-40 cm. When the water temperature rose to 15℃, chlorella can be used regularly at an amount according to water fertility.

Input of aquatic snails

Fresh snails were placed before the end of March, 2017, and spread evenly. The snails were disinfected with 3‰-4‰ brine for 3-5 min before release. In order to prevent early outbreak of moss, the input of snails was changed from the original one time to three times, and the total amount of the snails was 6 000-7 500 kg/hm2. The first input was carried out before or after the Qingming period, at an amount of 750 kg/hm2; the second input time was from June to July, at an amount of about 3 000 kg/hm2; and the third input was in August, according to the situation of the crab pool at an amount of about 3 000 kg/hm2.

Stocking of breeds

Stocking of crab breed

In early March, at noon of a sunny day, large good crab breed was released to the temporary rearing region outside the fence. The crabs had a specification of 40-50 crabs/kg, and were disinfected with 3‰-4‰ saline before release and then released at a stocking density of 12 000 crabs/hm2.

Stocking of fingerlings

In order to give full play to the productivity of the crab pond, the benthic Chinese sucker in middle lower layer and the filterfeeding silver carp and bighead carp in the upper middle layer were stocked together with the river crabs to form a composite ecological body. Chinese sucker can feed on benthic animals, food residues, animal waste, etc., and silver carp and bighead carp can feed on phytoplankton, achieving the effects of improving water quality of the crab pond and controlling water fertilizer of the pond, reducing the growth of pathogens and parasites, reducing the probability of occurrence of crab disease, and improving overall benefit of aquaculture.

In April, 10-12 cm Chinese sucker fingerlings, with neat specifications, having strong body free of no disease and injury, were placed in the aquatic plant protection area in the fence, at a stocking density of 750 individuals/hm2. Silver carp and bighead carp were input the next day, with a stocking specification of 100 g/fingerlings and a stocking density of 600 fingerlings/hm2 (stocking ratio of silver carp to bighead carp at 3∶1). The Chinese sucker and silver carp and bighead carp were immersed in 3%-4% saline for 3-5 min before stocking. The specific stocking conditions are shown in Table 1.

Table 1Stocking conditions in unit area

Stocking speciesStocking specificationStocking timeStocking density

River crab40-50 crabs/kgMarch12 000 crabs/hm2

Chinese sucker10-12 cm/fingerlingsApril750 fingerlings/hm2

Silver carp and bighead carp100 g/fingerlingsApril600 fingerlings/hm2

Aquaculture management

Culture of aquatic plants

In the early culture period from April to May, the water level of the crab pond was controlled at 40-50 cm, fertilization was performed for multiple times to promote the growth of aquatic plants and inhibit the outbreak of moss (when a small amount of moss occurs, it is preferred to use sodium humate to sprinkle on the moss and inhibit growth of moss by shading, and not to use agrochemicals to kill the moss). When the temperature rose from the end of May to the beginning of June, as the water level gradually increased to higher than 60 cm, Elodea was controlled to 10 cm under water. During the hot season from July to August, the growth of Elodea was accelerated, and the tip of the grass was cut off in time to keep them growing 20 to 30 cm under the water surface; and the water level was deepened to 80 cm or more in time to reduce the water temperature. In late culture period, after September, floating, rotten and dead plants were removed in time, and substrate improvers such as potassium hydrogen persulfate tablets were regularly used to prevent water pollution.

Input of baits

Input of baits was mainly based on river crabs, which fed on both plant and animal baits among which animal baits were the main food. Chinese sucker and silver carp and bighead carp bait mainly fed on natural baits. In addition to the snails and water plants in the water body, the crabs were fed with crab pellets, iced fish, wheat and corn. March-May: Feeds containing 36%-40% of protein were mainly input, supplemented with iced fish; June-August: As the temperature rose, animal feeds were reduced to just maintain crab growth, and feeds with a protein content of 28%-30% were mainly input, supplemented with coarse grains such as wheat and corn. September-October: For late fattening, iced fish was mainly input, supplemented with feeds containing 36%-40% of protein.

In addition, the feeding amount of baits was adjusted according to the growth condition of crabs, the weather conditions and the intake condition. In the hot season from June to August, vitamin C and allicin were regularly added to the crab feeds to improve crab immunity and disease resistance, and heatclearing and detoxifying additives such as ginkgo lead extract were added to promote body metabolism and improve blood circulation.

Regulation of dissolved oxygen (DO)

River crabs and Chinese suckers have poor tolerance to hypoxia, so the complex aeration mode of "microporous aeration + impeller aeration" was adopted to ensure sufficient DO in the water. Water DO was measured periodically to ensure that the DO was in the range of 6-8 mg/L.

The aerator was started according to the DO condition in the crab pond and weather conditions. In April-May, the aerator was started at midnight of rainy days; in June-October, the machine was turned on for 2-3 h in the afternoon and for 2-3 h before sunrise, and in case of continuous rainy or lowpressure weather, the machine was started at 21:00-22:00 in the night until noon the next day; and in late culture period, the machine was operated frequently.

Daily management

The pond was observed regularly, to fully grasp the feeding, growth and activities of river crabs and Chinese suckers, and to handle special situations in time. Meanwhile, the antiescape and aeration facilities were checked, to find problems in time for maintenance.

Water was measured regularly. The water quality indicators included pH, DO, transparency, ammonia nitrogen, nitrite, hydrogen sulfide, water temperature, etc., and the growth condition of algae was examined with a microscope. In the early culture period, active microbial agents such as Bacillus subtilis, EM bacteria and chlorella were often used to improve water quality. Potassium hydrogen persulfate tablets were used regularly to improve the substrate in the middle and late culture period; and if the water quality deteriorated or changed abruptly, it should be regulated in time.

Diseases were prevented scientifically under the principle of "mainly preventing disease and focusing on prevention rather than treatment" and preventing and treating diseases early. Additives such as vitamin C, allicin, and ginkgo leaf extract were added to the feed regularly to improve the disease resistance of crabs.

Sales was conducted timely. According to market conditions, fishing and sales were conducted in time to ensure maximum benefits. Before the Mid-Autumn Festival, the price of crabs was higher, and the fished parts reached the marketing specification. Large crabs could be captured with cages and manually in the evening. They were divided into male and female ones, which were packaged for sale. The remaining crabs in the pool were gradually sold according to market conditions. At the end of the culture, remaining crabs and fish could be caught in dry pond.

Agricultural Biotechnology2019

Experimental Results

It could be seen from Table 2 that a total of 96 240 yuan/hm2 of breeds, snails, aquatic plants, water and electricity, feeds, iced fish, fishery drugs, pond rent and pond facilities were input. As shown in Table 3, it could be found that the benefits were river crab 115 620 yuan/hm2, Chinese sucker 44 280 yuan/hm2, and silver carp and bighead carp 5 418 yuan/hm2, with a total output of 165 318 yuan/hm2; and the contribution rates of the three cultured species to the increase of benefit per unit area were 69.94%, 26.78% and 3.28%, respectively. It could be seen from calculation that the net profit was 69 078 yuan/hm2, and the outputinput ratio was 1∶1.72.

Table 2Stocking conditions in unit area

Input itemsSpecificationUnit priceTotal released amountAverage input∥yuan/hm2

Crab40-50 crabs/kg120 yuan/kg240 kg28 800

Chinese sucker10-12 cm0.5 yuan/fingerlings750 fingerlings2 250

Silver carp and bighead carp100 g/fingerlings60 yuan/kg60 kg240

Snail7 500 kg/hm21.6 yuan/kg7 500 kg12 000

Aquatic plant1 500

Water and electricity4 500

Feed16 500

Fishery drug1 950

Iced fish9 750

Antiescape facility2 250

Aeration facility2 250

Pont rent14 250 yuan/hm214 250

Sum96 240

Table 3Benefit per unit area and contribution rate to increase of benefit per unit area

ItemsAverage specificationAverage priceyuan/kgAverage yieldkg/hm2Average benefityuan/hm2Contribution rate%

River crab♀124 g/crab♂163 g/crab82.01 410115 62069.94

Chinese sucker42 g/individual120.033944 28026.78

Silver carp and bighead carp1.82 kg/individual5.6967.55 4183.28

Sum165 318

The experimental results showed that under the Chinese suckerintercropping diseasepreventing crab culture mode, no disease occurred in the whole crab pond. It not only effectively avoided the occurrence of crab diseases, but also reduced the use of fisher drugs and exerted the utilization efficiency of the crab pond water body. It also greatly improved the economic benefits of farming.

Analysis and Discussion

Scientific selection of raised fish species

The benthic Chinese sucker lives in the lower middle layer of the water, mainly feeds on benthic invertebrates, underwater food residues, animal excrement, etc., and eats frequently, which is beneficial to the regulation of the culture substrate and the inhibition of growth of pathogens. Filterfeeding silver carp and bighead carp live in the upper middle layer of water, mainly feed on phytoplankton (such as diatoms, green algae and cyanobacteria) and zooplankton (such as protozoa, rotifers, cladocerans, copepods, etc.), and thus help to regulate water quality, promote nutrient conversion in ponds, control water body fertility and inhibit cyanobacteria. Through the innovation of the threedimensional stocking model of Chinese sucker, silver carp and bighead carp and river crab, Chinese sucker, silver carp and bighead carp and river crab together form a group of pond consumers, which can mutually benefit each other, thereby avoiding the risk of crab disease and improving selfpurification ability of the crab pond water. On the one hand, this culture mode obtained the greatest economic benefit, and improved the utilization rate of the crab pond water body, and on the other hand, it realized the minimum pollution that the crab pond environment can withstand, or even no pollution, and relieved the pressure of river crab production on water environment.

Optimization of aquaculture environment

Based on the living habits and feeding habits of Chinese sucker, it can be raised in river crab pond to remove excessive benthic animals, bottom residues, excreta and organic decay in the pond, reduce oxygen debt at the bottom of the pond, and reduce the use of disinfectants and chemicals such as substrate improvers, so as to regulate the culture environment and reduce the incidence of crab disease. The experiment results showed that the crab ponds produced 339 kg/hm2 of Chinese sucker, and the average benefit increased by 44 280 yuan/hm2, to which the contribution rate of Chinese sucker reached 26.78%. This condition greatly promotes the enthusiasm of the farmers, and the culture mode is an innovative breeding model that avoids the risk of "single fight" of river crab.

Adoption of composite aeration mode

In this study, we adopted the composite aeration mode of "microporous aeration + impeller aeration", which is obviously superior to microporous aeration or traditional impeller aerator. It achieved the effects of effectively promoting the exchange of water in the upper and lower layers of the crab pond and speeding up the decomposition of organic matter in the pond, greatly improving the substrate and water quality of the crab pond, reducing the living space of pathogens, and greatly reducing the occurrence of diseases and the use of fishery drugs. In 2017, DO in the test pond remained at a high level throughout the year, and the water quality indicators were normal throughout the whole process; and crabs and Chinese suckers grew well and suffered from no disease. The used fishery drugs in this study only cost 1 950 yuan/hm2, far lower than that in other crab ponds in the surrounding area.

Timely scientific fishing and sale

It is better to put the crabs into market before and after the MidAutumn Festival when the price of crabs is higher. According to market conditions and pond conditions, large river crabs were caught and sold; remaining crabs in the pond were selected for sale at a higher price around mid-November; and finally, all the animals were sold to ensure the highest farming benefit. Due to the low market supply of Chinese sucker, the market price has been high, and the average selling price has reached 120 yuan/kg. In 2017, crabs and Chinese suckers in the whole pond were basically harvested and sold at high prices. The two cultured species, river crab and Chinese sucker showed a total contribution rate to the average increase of benefit per unit area reaching 96.72%, and the final average net profit per unit area reached 69 078 yuan/hm2.

Therefore, the Chinese suckerintercropping diseasepreventing crab culture mode greatly reduced the incidence of crab diseases, especially the "Shuibiezi" disease of river crabs in the past two years, which greatly reduced the use of fishery drugs and improved the pond culture environment. The comprehensive benefits of crab pond farming have been improved, and the resistance of farmers to market risk has been improved (even if the market price of crabs is low, it can be compensated by the benefit of Chinese sucker), which has great promotion value to the promotion of sustainable and healthy development of crab industry, and is of great significance to the promotion of fishery transformation and upgrading.

References

［1］ HAO JUN, MA XZ, WANG YC, et al. The metazoam zooplankton community structure and dynamics of crab ecological farming ponds［J］. Journal of Shanghai Ocean University, 2015, 24(4): 523-531. (in Chinese)