脱脂蚕蛹替代日粮中鱼粉对建鲤肠道菌群的影响

利用免培养的16S rDNA梯度凝胶电泳技术(DGGE)研究脱脂蚕蛹替代鱼粉对建鲤(Cyprinus carpio var. jian)肠道菌群的影响。6个饲养组分别饲喂6种等氮等能饲料, 分别为含10%鱼粉的基础饲料、3种替代饲料(脱脂蚕蛹替代鱼粉蛋白, 替代水平分别为25%、50%和75%)及2组分别在50%和75%替代基础上分别再添加0.6%和0.7%晶体赖氨酸的饲料 (分别记为DSP0、DSP25、DSP50、DSP50-Lys、DSP75、DSP75-Lys)。饲养结束后采集肠道样品, 分析建立指纹图谱, DSP0、DSP25、DSP50、DSP50-Lys、DSP75、DSP75-Lys组分别产生了19、15、11、12、8和12条可鉴别条带, 各组均存在优势种群条带。DSP0和其他各组(DSP25、DSP50、DSP50-Lys、DSP75、DSP75-Lys)的相似性指数为50.7%、50.6%、69.0%、42.4%、66.7%。对PCR-DGGE指纹图谱主要条带进一步回收、克隆和测序, 共得到19条序列, 将得到的序列在NCBI数据库中进行同源分析, 发现建鲤肠道菌群归为芽孢杆菌纲、螺旋体纲、β-变形菌纲、梭菌纲、放线菌纲、γ-变形菌纲、蓝藻纲、拟杆菌纲、梭杆菌纲等, 大部分为不可培养细菌。研究发现, 脱脂蚕蛹替代鱼粉对建鲤肠道菌群组成产生显著影响, 随着脱脂蚕蛹替代水平的提高, 肠道菌群多样性下降, 细菌种类从19种锐减至8种, 而饲料中补充赖氨酸可减弱这种影响, 本研究可为蚕蛹在鲤配合饲料中的开发应用提供参考资料。

Effect of substituting dietary fish meal with defatted silkworm pupae on the intestinal bacterial community structure of Jian carp, Cyprinus carpio var. jian

Our objective was to evaluate the effects of fish meal (FM) replacement with defatted silkworm pupae (DSP) on the intestinal bacterial community structure in Jian carp (Cyprinus carpio var. jian) using culture-independent 16S rDNA PCR-DGGE fingerprinting. Five isonitrogenous (36.8% crude protein) and isoenergetic (5.5% crude lipid) diets were formulated as follows: one basal diet with 10% FM (DSP0), three diets with 25%, 50%, and 75% FM replacement (DSP25, DSP50, DSP75), and two FM replacement diets (DSP50 and DSP 75) that were supplemented with 0.7% crystalline lysine (DSP50-Lys, DSP75-Lys). A total of 324 juvenile Jian carp (mean body weight: 15.30±2.97 g) were randomly assigned to one of 6 groups (3 replicates per group, 18 fish per replicate). A PCR-DGGE fingerprint analysis revealed that DSP0, DSP25, DSP50, DSP50-Lys, DSP75, DSP75-Lys had 19, 15, 11, 12, 8, and 12 bands, respectively. The dominant bacteria were represented by 1-2 bands. The similarity coefficient between the DGGE fingerprint of DSP0 bacteria and the other groups was 50.7%, 50.6%, 69.0%, 42.4%, and 66.7%, respectively. The major bands were recovered, cloned, and sequenced, yielding 19 sequences. The sequences were analyzed by comparison with the NCBI database. The bacterial community consisted primarily of Bacilli, Spirochaetes, Beta proteobacteria, Clostridia, Actinbacteria, Gammaproteobacterial, Cyanobacteria, Bacteroidetes, and Fusobacterial, and included 15 sequences that were uncultured. In conclusion, replacement of graded dietary fish meal protein with defatted silkworm pupae meal had a significant effect on the intestinal bacterial community structure in Jian carp. However, the addition of lysine reduced this effect. Our results provide a foundation for further development of alternative carp feeds.