解淀粉芽孢杆菌和胶红酵母复合菌对虹鳟生长性能及胃黏膜、肠黏膜菌群结构的影响

为研究饲料中添加不同水平的复合菌剂(解淀粉芽孢杆菌,Bacillus amyloliquefaciens V4和胶红酵母,Rhodotorula mucilaginosa)对虹鳟(Oncorhynchus mykiss)幼鱼生长及消化道黏膜微生物菌群结构的影响,选用体重为(205.1±4.82)g的虹鳟幼鱼360尾,随机分为4组(每组3个重复,每个重复30尾),分别投喂基础饲料(C0)和3种添加水平为5×10~6/5×10~7 CFU/g(T1),1.5×10~7/1.5×10~8 CFU/g(T3),2.5×10~7/2.5×10~8 CFU/g(T5)的复合菌剂(B.amyloliquefaciens V4/R.mucilaginosa),实验周期42 d。研究结果发现饲料中添加复合益生菌对虹鳟的生长及存活有一定的促进和提高,T1比例的复合益生菌能够显著提高虹鳟的增重率和特定生长率、显著降低饲料系数(P0.05),同时T1和T3比例添加显著降低虹鳟的死亡率(P0.05);对其消化道黏膜细菌群落16S rDNA进行聚合酶链式反应-变性梯度凝胶电泳(PCR-DGGE)指纹分析,结果表明:虹鳟胃黏膜和肠黏膜上微生物菌群种类存在差异;胃黏膜菌群DGGE图谱中分别检测到35.7±17.0(C0)、37.0±3.5(T1)、36.7±13.6(T3)、26.0±13.2(T5)条谱带,各处理组间谱带数目无显著差异(F=0.500,P=0.692),肠黏膜菌群DGGE图谱显示分别检测到23.3±5.8(C0)、22.3±3.2(T1)、16.7±8.0(T3)、24.7±7.4(T5)条谱带,各处理组间谱带数目也无显著差异(F=0.916,P=0.475);胃黏膜菌群多样性随着益生菌添加量增加,菌群多样性有升高趋势,但是在最高浓度组(T5)多样性降低,肠黏膜菌群多样性随着复合菌剂的添加,多样性指数持续降低,中浓度添加组(T3)多样性最低,但是随着添加浓度升高,呈现恢复和升高趋势(T5);基于所得PCR-DGGE指纹图谱中谱带丰度值数据的UPGMA聚类和PCA排序分析均显示胃黏膜微生物群落与肠黏膜微生物群落结构差异明显,大致分为两个不同的分支,胃黏膜和肠黏膜微生物菌群并没有按照不同处理组而有显著分化。以上结果表明解淀粉芽孢杆菌和胶红酵母复合添加能显著促进虹鳟的生长,提高存活率,外源益生菌添加对虹鳟消化道黏膜上优势菌群能产生一定影响,但并未对胃黏膜及肠黏膜菌群多样性产生显著影响,同时也未显著改变虹鳟肠黏膜微生物菌群结构,高比例添加降低消化道黏膜细菌数量及多样性风险。     

Effect of Bacillus cereus as a water or feed additive on the gut microbiota and immunological parameters of Nile tilapia

We evaluated the effects of Bacillus cereus, as an additive in water and feed, on the gut microbiota and immunological parameters of Nile tilapia (Oreochromis niloticus) fingerlings. Experiments were performed in tanks and net cages respectively. Experiment 1: Tilapia were housed in tanks for 42 days, and B. cereus was added to the water at 1.0 × 10⁴ cfu mL^?1 (Treatment 1) and 1.0 ×10⁵ cfu mL^?1 (Treatment 2) weekly. For the control, no probiotic was added. Experiment 2: Tilapia were housed in cages for 42 days, and the feed was supplemented with B. cereus at 1.0 × 10⁷ cfu g^?1 (Treatment 1) and 1.0 × 10⁸ cfu g^?1 (Treatment 2) weekly. For the control, no probiotic was added. Each treatment contained three replicates, with 50 male tilapias per replicate. The fish from the probiotic treatments in both tank and cage experiments had significantly higher serum lysozyme and peroxidase activities than the control. In the cage experiment, alkaline phosphatase and total superoxide dismutase activities in tilapia were significantly higher in probiotic treatments compared with the control. The results of polymerase chain reaction‐denaturing gradient gel electrophoresis showed that B. cereus supplementation in the feed and water affected the autochthonous gut bacteria community of tilapia and stimulated various potentially beneficial bacteria. Therefore, B. cereus, as a water or feed additive, could enhance the immune status and affect the gut microbiota of tilapia. Bacillus cereus was more effective as a feed supplement rather than a water additive for enhancing the immune status of tilapia.     

Two biomass preparation methods provide insights into studying microbial communities of intestinal mucosa in grass carp (Ctenopharyngodon idellus)

Animal digestive tract is habitat for a large number of autochthonous microbiota, which play central roles in multiple biological and physiological processes of the host. In this study, two different micro‐biomass preparation methods were employed to evaluate the diversity of intestinal mucosa‐associated microbiota in grass carp (Ctenopharyngodon idellus). Genomic DNAs were isolated either directly from intestinal mucosal samples (group A), or from micro‐biomass after microbial dissociation (group B). Community richness, diversity and evenness indices were all higher in group B, but differences were not statistically significant (P = 0.97, P = 0.33, P = 0.34 respectively). Furthermore, group B samples exhibited an increased ratio of bacterial DNA in comparison with group A samples, but the difference was also not statistically significant (P = 0.74). In addition, there were no statistically significant differences between the two groups (P > 0.05) at the taxonomic level. Our results support previous findings that there exists a great abundance of the intestinal mucosa‐adherent microbiota in the grass carp; among these, Proteobacteria, Firmicutes, Bacteroidetes, Actinobacteria, Spirochaetes and Fusobacteria were the most common phyla. Within these microbiota, Paenibacillus, Bacteroides, Bacillus and Cetobacterium genera comprise the majority of the community, implicating their functional importance (e.g. as probiotics) to their host. Our results contribute towards a better understanding of the intestinal microbial profile of grass carp. Both micro‐biomass preparation techniques proved to be feasible for studying mucosa‐adherent microbiota of grass carp; however, the second method (group B) provides a protocol that is somewhat more effective than the first method (group A).     

Impacts of diet on hindgut microbiota and short‐chain fatty acids in grass carp (Ctenopharyngodon idellus)

Diet is known to influence intestinal microbiota in fish, but the specifics of these impacts are still poorly understood. Different protein/fibre ratio diets may result in differing structures and activities of gut microbiota. We examined the hindgut microbiome of grass carp (Ctenopharyngodon idellus) fed three different diets: fish meal (FM, high protein – low fibre), Sudan grass (SG, high fibre – low protein) and compound feed (CF, intermediate). Microbial profiles of fish fed on FM were significantly different from profiles of fish fed CF and SG (F = 18.85, p < .01). Cetobacterium, known to be positively associated with protein digestion, was the dominant microbial group in FM samples (approximately 75.7%), while Lachnospiraceae and Erysipelotrichaceae, thought to be involved in fermentation of plant polysaccharides, were dominant in CF and SG samples (46.8% and 42.9% respectively). Network analyses indicated that the abundance of Lachnospiraceae and Erysipelotrichaceae was in a significantly positive correlation (r = .895, p = .001). Short‐chain fatty acid (SCFA) levels may indicate that the digestibility of diet by microbiota in the grass carp gut decreased from FM to SG (FM>CF>SG). Overall low SCFA levels indicate that hindgut fermentation probably provides a low proportion of energy requirements in grass carp.     

池塘养殖牙鲆肠道和环境菌群结构对益生菌制剂的响应

为研究益生菌制剂对池塘养殖牙鲆(Paralichthys olivaceus)肠道及环境菌群结构的调控效果,采用高通量测序技术和生物信息学分析手段构建牙鲆肠道、养殖水体、饵料和池塘底泥的16S rDNA基因测序文库,分析不同样品中菌群组成和多样性在益生菌制剂调控过程中的变化趋势.结果显示,添加益生菌制剂后,池塘底泥和牙...     

Role of gastrointestinal microbiota in fish

The gastrointestinal (GI) tract of an animal consists of a very complex and dynamic microbial ecosystem that is very important from a nutritional, physiological and pathological point of view. A wide range of microbes derived from the surrounding aquatic environment, soil/sediment and feed are found to colonize in the GI tract of fish. Among the microbial groups, bacteria (aerobic, facultative anaerobic and obligate aneraobic forms) are the principal colonizers in the GI tract of fish, and in some fish, yeasts are also reported. The common bacterial colonizers in the GI tract of freshwater and marine fish include Vibrio, Aeromonas, Flavobacterium, Plesiomonas, Pseudomonas, Enterobacteriaceae, Micrococcus, Acinetobacter, Clostridium, Fusarium and Bacteroides, which may vary from species to species as well as environmental conditions. Besides, several unknown bacteria belonging to Mycoplasma, Arthrobacter, Brochothrix, Jeotgailbacillus, Ochrobactrum, Psychrobacter and Sejongia species in the GI tract of different fish species have now been identified successfully using culture‐independent techniques. Gnotobiotic and conventional studies indicate the involvement of GI microbiota in fish nutrition, epithelial development, immunity as well as disease outbreak. This review also highlights the need for manipulating the gut microbiota with useful beneficial microbes through probiotic, prebiotic and synbiotic concepts for better fish health management.     

豆粕对乌鳢肠道菌群组成及微生物氨基酸代谢酶活性的影响

为研究豆粕替代鱼粉对乌鳢生长、肠道菌群组成和微生物相关氨基酸代谢酶活性的影响,分别用豆粕替代0、25%、50%和75%的鱼粉配制4组等氮等能的饲料饲喂初始体质量为(8.65±0.25)g的幼鱼(分别命名为G1、G2、G3和G4组),养殖实验在室内纤维玻璃钢桶中进行,实验周期为21 d。结果显示,各组间幼鱼成活率无显著性差异;G4组乌鳢幼鱼的生长性能显著低于G1和G2组,而G3组幼鱼生长性能与其余3组差异不显著。厚壁菌门在G1和G2组乌鳢肠道中的比例显著高于G3组,且在G4组中的比例最低;变形菌门在G1和G2组中的比例显著低于G3和G4组;G4组中乳球菌属、杆菌属、假单胞菌属、链球菌属、芽孢杆菌属和不动杆菌属的比例显著低于其余3组。另外,G4组幼鱼肠道微生物谷丙转氨酶、腺苷脱氨酶活性显著高于其余3组,而G1组乳酸脱氢酶的活性最高。研究表明,豆粕替代鱼粉除了对乌鳢幼鱼的生长产生影响外,也显著影响了肠道微生物组成及微生物相关氨基酸代谢酶的活性,本研究结果对于从微生态角度进一步理解豆粕在乌鳢配合饲料中的应用提供了理论依据和参考。     

船舶噪声声压级对大黄鱼幼鱼游泳、摄食行为及免疫生理指标的影响

通过录制大黄鱼(Larimichthys crocea)产卵场附近船舶航行时的噪声,并以此为刺激源,以大黄鱼幼鱼为实验对象,研究了船舶噪声声压级对大黄鱼幼鱼游泳、摄食行为及免疫生理指标的影响。研究发现,当噪声声压级<60 dB时,幼鱼趋避游泳行为不强烈;但随着声压级增大,开始呈现出不同强度的趋避行为,依次表现出:游泳速度加快、鱼与鱼之间及鱼与桶壁之间发生碰撞、瞬间反应无序、跳跃等行为;当声压级>200 dB时,刺激2 min后就出现了死亡个体。在120 dB和150 dB短期单次和多次刺激下,幼鱼血浆中的皮质醇、血糖、血红蛋白和乳酸这4个与应激相关的生理指标均显著上升,其中皮质醇、血糖和乳酸上升幅度尤为明显。另外,皮质醇单次刺激后即达到峰值,而多次刺激后反而较单次刺激有所下降;血糖、血红蛋白和乳酸则具有累加效益,多次刺激要高于单次刺激。在120 dB长期(30 d)刺激下,幼鱼生长明显减缓,血浆中部分免疫指标(免疫球蛋白M,干扰素-α,白介素-1β和肿瘤坏死因子-α)明显降低,肠道菌群也发生显著变化,突出表现为部分益生菌如芽孢杆菌、乳杆菌等相对丰度明显降低。摄食行为影响方面,研究发现幼鱼能准确识别噪声源的位置,并具有一定的记忆性。研究结果揭示了船舶噪声对大黄鱼幼鱼的危害,可为今后制定大黄鱼产卵场的保护措施提供数据支撑。     

Effects of a highly purified fucoidan from Undaria pinnatifida on growth performance and intestine health status of gibel carp Carassius auratus gibelio

A six‐week feeding trial was conducted to determine the effects of different concentrations of fucoidan (1 g/kg, 10 g/kg and 30 g/kg; w/w) from Undaria pinnatifida on gibel carp (Carassius auratus gibelio). Our results demonstrated that 30 g/kg fucoidan significantly increased (p < .05) growth performance, intestinal digestive enzyme activities, acid phosphatase activity and immunoglobulin M content. Histological examinations revealed that gibel carp receiving 30 g/kg fucoidan had significant higher abundance of mucin‐containing goblet cells in middle and distal intestine as compared with control treatment (p < .05). Intestinal microbiota analysis showed that 30 g/kg fucoidan supplementation significantly increased (p < .05) the abundance of Cetobacterium and Aeromonas, but lowered (p < .05) the prevalence of pathogenic bacteria Plesiomonas and a mucin‐degrading bacterium Mucinivorans. Furthermore, RNA‐seq and RT‐qPCR analysis indicated that 30 g/kg fucoidan caused significant changes (p < .05) in the expression of genes involved in immune regulation (such as interleukin‐8 and cyclooxygenase), signal transduction (such as phosphatidylinositol‐4,5‐bisphosphate 3‐kinase and protein kinase B) and nutrition utilization (maltase–glucoamylase and muscarinic acetylcholine receptor 3). Together, the current study shows that fucoidan supplementation could elevate the activity of intestinal digestive enzymes, modulate intestinal microbial communities and potentiate a higher state of immune readiness, which might consequently improve growth performance and intestine health status of gibel carp.