Studies On The Development Of The Digestive System In Bullfrog Tadpoles

The digestive system of Rana catesbeiana tadpole stage is not fully developed,the digestive capacity is limited,and the lack of special compound feed and immature seedling breeding technology leads to the high mortality rate and low metamorphosis rate of Rana catesbeiana tadpole,which seriously restricts the healthy development of Rana catesbeiana industry.Through exploring bullfrog tadpoles each developmental stage structure of digestive organs morphology,digestive enzyme activity(lipase,amylase,trypsin,pepsin,alkaline phosphatase)and thyroid hormone(T3,T4),growth hormone(GH),insulin-like growth factor-I(IGF-I)the change of the content,and analyze the abnormal tadpoles intestinal flora changes,With development change rule of bullfrog tadpoles digestive system,rich bullfrog(tadpoles)the basic biology of early development stage,for the development of tadpoles special compound feed and provide theoretical basis for suitable feeding strategies.Research shows that:1 changes of digestive system structure of Bullfrog tadpoleThe structural changes in the digestive organs of bullfrog tadpoles during growth and development are closely related to the change in food habits.At 3 days of age,the digestive organs of bullfrog tadpoles are not differentiated and the yolk sac provides energy for the body,which is the stage of endogenous nutrition;at 5 days of age,the eyes and mouth are differentiated and feeding begins,the yolk sac shrinks in size and the stomach,intestine and hepatopancreas begin to differentiate,which is the stage of mixed nutrition;at 6 days of age,the yolk sac is exhausted and the stage of exogenous nutrition is entered;at 37 days of age,a small number of cup-shaped cells can be seen in the stomach,and thereafter,the stomach gland,gastric rim,intestinal gland,intestinal rim and other structures can be seen.At 72 days of age(stage 43-45),the histological structures of the stomach and intestine(mucosal layer,submucosal layer,muscular layer and plasma membrane)are clearly layered,the gastric glands are well developed,the intestine has a large number of cup cells,the hepatocytes are increased,and the hepatocyte cords and hepatic blood sinuses are evident;at 74 days of age(stage 46),the tadpole is completely metamorphosed,the stomach wall is thickened,the gastric glands and gastric villi are increased,the intestinal villi are dense,and the intestinal mucosal layer has a large number of cup cells.The liver lobules are very inconspicuous.The hepatic lobules are very inconspicuous,the hepatocytes are densely packed,the hepatocyte cords are closely arranged,and the digestive system is gradually improving in terms of structure and function.2 Digestive physiological characteristics of the developing digestive system of bullfrog tadpoles(1)Changes in the main digestive enzymesAll five major digestive enzymes could be detected in bullfrog tadpoles at 5 days of age,and the soluble protein content showed a general trend of first decreasing and then increasing;from 5 to 74 days of age,the amylase activity showed a general trend of first decreasing and then increasing and then decreasing,and the specific activity showed a trend of first increasing and then decreasing,among which,the activity was higher from 5 to 58 days of age and the specific activity was lower,and from 70 to 72 days of age,they all decreased significantly(P < 0.05).The activity and specific activity of lipase showed a general trend of increasing and then decreasing,with the highest activity 14.31 U/g prot at 37 days of age and the highest specific activity 8.34 U/g prot at 70 days of age;the activity and specific activity of pepsin were low from 5 to 46 days of age and significantly increased from 60 to 74 days of age(P < 0.05);the activity of trypsin showed a significant trend of increasing from 22 to 46 days of age(P < 0.05).Trypsin activity showed a significant upward trend from 22 to 46 days of age(P < 0.05),with the highest 3759.44 U/mg prot at 46 days of age,while specific activity fluctuated upward from 22 to 70 days of age(P < 0.05),with a peak 4.82 U/mg prot at 70 days of age;alkaline phosphatase activity and specific activity both showed an upward and then downward trend from 5 to 28 days of age,with a significant increase in activity from70 to 74 days of age(P < 0.05),while the specific activity decreased and then increased(P < 0.05).(2)Changes in the content of major endocrine hormonesAll four hormones were detected in bullfrog tadpoles at 5 days of hatching age.During the growth and development of tadpoles,T4 content was the highest,followed by T3 content and GH content was the lowest,and T3,T4 and IGF-I content all peaked at 72 days of age(stage 43-45)and then showed a significant decreasing trend(P <0.05),while GH content showed an increasing trend after 72 days of age(P < 0.05).The levels of T3,T4 and IGF-I hormones increased significantly during the peak of metamorphosis in bullfrog tadpoles and decreased after metamorphosis was completed(P < 0.05),while GH levels changed in the opposite direction.3 Analysis of intestinal flora diversity in bullfrog tadpoles during metamorphosisThe results showed that at the phylum level,Fusobacteriota,Firmicutes,Actinobacteria,Bacteroidetes and Proteobacteria were the main intestinal flora.At the genus level,the main genera are Cetobacterium,Actinomyces and Bacteroides.The phylum Cetobacterium is the main dominant species in the metamorphic stage of tadpoles,with relative abundances ranging from 28.5% to 56.1%.A total of 106 ASVs were recorded at 37,46,58 and 70 days of age as the host developed.at the KEGG level 2,the main functions of the bullfrog tadpole metamorphosis gut flora were associated with transport and catabolism,age,endocrine and metabolic diseases,the nervous system and the endocrine system.