Chinese Herbal Compound Screening and Its Pharmacodynamic Efficacy Evaluation Against Shrimp Postlarva Bacterial Vitrified Syndrome (BVS) Induced by Vibrio alginolyticus

Abstract

Penaeus vannamei is an important breeding shrimp species in China. Moreover, the quality of the larvae is the cornerstone for the sustainable development of the shrimp industry. In recent years, shrimp postlarva bacterial vitrified syndrome (BVS) has caused huge economic losses in the shrimp postlarva breeding industry, with epidemiological characteristics of rapid onset and high mortality, occurring at 2–3 days post-hatch. The diseased postlarva symptoms include emaciation, decreased activity, and an empty intestinal tract and stomach. The hepatopancreas shows atrophy, blurring of contour, paleness, and even vitrified syndrome. The causative agents of the disease are Vibrio alginolyticus and Vibrio parahaemolyticus. Furthermore, antibiotics are often used to treat BVS; however, frequent and inappropriate use of antibiotics can lead to bacterial resistance and drug residues. With the advantages of antibacterial activity, immunity regulation, low toxicity, few side effects, and lack of drug resistance and drug residues, Chinese herbal medicines play an important role in aquaculture. There are currently no effective drugs available to prevent and treat BVS. It is urgent and important that effective Chinese herbal compounds are developed for BVS prevention and treatment. In this study, first, an antibacterial test was carried out in vitro by the Oxford cup and double dilution methods. Overall, 50 types of Chinese herbal medicines were screened, and those with a good bacteriostasis effect on V. alginolyticus and V. parahaemolyticus were identified. Then, a pathological model of BVS was constructed with different concentrations of V. alginolyticus. The efficacies of the Chinese herb compounds were evaluated based on the mortality, histopathology and ultrastructural pathology characteristics of the postlarvae. The bacterial inhibition test showed that the inhibition zone of V. alginolyticus could be > 12 mm with the application of one of six types of Chinese herbs, including Schisandrae chinensis fructus, Verbenae herba, and Granati pericarpium. The inhibition zone of V. parahaemolyticus could be > 13 mm with the application of one of five types of Chinese herbs, including Chebulae fructus, S. chinensis fructus, Moutan cortex and Sanguisorbae radix. Then, 11 types of antibacterial and two types of immunological drugs were selected for further analysis. Moreover, the MIC and MBC of G. chinensis, C. fructus, Caryophylli flos, and S. chinensis fructus against V. alginolyticus and V. parahaemolyticus were ≤ 12.5 mg/mL and ≤ 50 mg/mL, respectively. Then, three types of compounds were used for in vivo pharmacodynamics tests by combining Chinese antibacterial medicine with Chinese immunological medicine (Cyperi rhizoma and Gardeniae fructus), which were named prescription 1 (C. fructus 30 g, S. chinensis fructus 20 g, C. rhizoma 20 g), prescription 2 (C. fructus 40 g, G. pericarpium 30 g, C. flos 30 g), and prescription 3 (C. fructus 40 g, G. chinensis 30 g, G. fructus 30 g), respectively. Florfenicol was used as the positive drug control, and V. alginolyticus infection without treatment was used as the positive control. The BVS model indicated that V. alginolyticus caused the same vitrification symptoms in shrimp postlarvae as a natural infection. The LD50 of shrimp postlarvae was 2.82 × 105 CFU/mL after 72 h of infection with V. alginolyticus. The pathological model of BVS was constructed by immersing shrimp postlarvae with 5 × 104 CFU/mL V. alginolyticus. Based on this, the infected shrimp postlarvae were treated with drugs for 7 days. After this period, the mortality rates of each group from low to high were: blank control 0.05). Thus, prescription 1 can significantly reduce the mortality of BVS-positive shrimp postlarvae. Histopathology observation showed that after 7 days of administration, the hepatopancreas and intestinal epithelium of larvae in the prescription 1 group were more intact, with more hepatic tubules, less exfoliation and necrosis of the epithelial cells, and more abundant and well-arranged intestinal epithelial cells. Ultrastructural pathology observation showed that after 7 days of administration, the cell membrane and nucleus of the liver tubule epithelial cells of the diseased shrimp postlarvae in the prescription 1 group were normal, and the mitochondria and endoplasmic reticulum were abundant. Histopathology observation showed that the hepatopancreas and intestinal tract lesions of the shrimp postlarvae in the prescription 1 treatment were significantly less than those in the other groups.In summary, Chinese herbal compounds (C. fructus 30 g, S. chinensis fructus 20 g, C. rhizoma 20 g) had the best effect for preventing and treating shrimp postlarva BVS caused by V. alginolyticus. The results of this study lay a scientific foundation for the development of specialized herbal compounds for the prevention and treatment of bacterial infections and contribute to the green and high-quality development of the shrimp industry.

Keywords

• penaeus vannamei
• bacterial vitrified syndrome (bvs)
• pathological model
• chinese herbal compound
• pharmacodynamics