Chinese Journal of Animal Nutrition >

2025 , Vol. 37 >Issue 12: 8539 - 8550

DOI: https://doi.org/10.12418/CJAN2025.695

RESEARCH PAPER

Effects of Dietary Supplementation of Fermented Black Soldier Fly (Hermetia illucens L.) on Hepatic Histoarchitecture, Antioxidant Capacity and Intestinal Barrier Function of Channel Catfish (Ictalurus punctatus)

  • ZHENG Yushun , 1, 2 ,
  • CAI Yunchuan 3 ,
  • HUANG Yanhua 4 ,
  • WU Yamei 3 ,
  • PENG Kai 2 ,
  • HU Junru 2 ,
  • ZOU Jixing 1 ,
  • WANG Guoxia , 2, **
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  • 1 College of Marine Sciences, South China Agricultural University, Guangzhou 510642, China
  • 2 Guangdong Provincial Key Laboratory of Animal Breeding and Nutrition, Key Laboratory of Animal Nutrition and Feed Science in South China of Ministry of Agriculture and Rural Affairs, Institute of Animal Science, Guangdong Academy of Agricultural Sciences, Guangzhou 510640, China
  • 3 Agriculture Technology Extension Centre of Guangdong Province, Guangzhou 510520, China
  • 4 Guangzhou Fishtech Biotechnology Co., Ltd., Guangzhou 510640, China
** professor, E-mail:

* Contributed equally

Received date: 2025-04-10

  Online published: 2025-12-13

Fold

Abstract

This experiment aimed to investigate the effects of dietary supplementation of fermented black soldier fly (Hermetia illucens L.) on hepatic histoarchitecture, antioxidant indexes, and intestinal non-specific immune enzyme activities, antioxidant indexes, tight junction protein gene expression, and microbiota composition of channel catfish (Ictalurus punctatus). Based on a conventional diet formulation, four isonitrogenous and isolipidic experimental diets were prepared by adding 0 (G0), 2% (G2), 4% (G4), and 8% (G8) of fresh fermented black soldier fly larvae, respectively. A total of 480 channel catfish with an initial body weight of (2.75±0.01) g were selected and randomly divided into 4 groups (with 3 replicates per group and 40 fish per replicate). These groups were fed the corresponding four experimental diets for a 56-day feeding trial. The results showed as follows: 1) histological examination showed no pathological liver damage, with hepatocytes exhibiting regular arrangement, distinct nucleocytoplasmic boundaries, and no vacuolization or inflammatory infiltration was observed. 2) Compared with the control group (G0 group), hepatic total antioxidant capacity (T-AOC) and catalase (CAT) activity in the G2 and G4 groups were significantly increased (P<0.05), hepatic total superoxide dismutase (T-SOD) activity in the G4 group was significantly elevated (P<0.05), while hepatic protein carbonyl (PrC) content in the G2, G4 and G8 groups was significantly decreased (P<0.05). 3) Intestinal alkaline phosphatase (AKP) activity in the G2 group and acid phosphatase (ACP) activity in the G4 group were significantly higher than those in the control group (P<0.05); compared with the control group, intestinal CAT activity was significantly increased in the G2 group (P<0.05), while intestinal malondialdehyde (MDA) content was significantly reduced in the G2, G4, and G8 groups (P<0.05). 4) The relative expression levels of zonula occludens-1 (ZO-1) and claudin-2 (Claudin-2) genes in the G2 group were significantly upregulated compared with the control group (P<0.05). 5) The α and β diversity of intestinal microbiota in channel catfish among all groups showed no significant changes (P>0.05). In intestinal microbiota, G2 group showed increased Firmicutes relative abundance and G8 group displayed decreased Proteobacteria with elevated Fusobacteria (mainly Cetobacterium) relative abundance. In conclusion, dietary supplementation with 2% to 4% fresh fermented black soldier fly can improve hepatic antioxidant capacity, enhance intestinal mechanical barrier and immune barrier functions, and improve the intestinal microbiota structure of channel catfish.

Key words： Ictalurus punctatus; fermented black soldier fly; hepatic morphology; immune; antioxidant; intestinal microbiota structure

Cite this article

ZHENG Yushun , CAI Yunchuan , HUANG Yanhua , WU Yamei , PENG Kai , HU Junru , ZOU Jixing , WANG Guoxia . Effects of Dietary Supplementation of Fermented Black Soldier Fly (Hermetia illucens L.) on Hepatic Histoarchitecture, Antioxidant Capacity and Intestinal Barrier Function of Channel Catfish (Ictalurus punctatus)[J]. Chinese Journal of Animal Nutrition, 2025 , 37(12) : 8539 -8550 . DOI: 10.12418/CJAN2025.695

鱼类肝脏与肠道健康是其维持正常生长、发育与抗病能力的生物学基础。肝脏作为物质代谢和毒素转化的中枢器官,其组织结构的完整性直接影响营养物质的代谢周转、储存效率及内源性毒物的清除能力[1]。肠道除负责营养吸收外,更凭借机械屏障、免疫屏障与微生物屏障共同构成防御体系,在抵抗病原侵袭和维持机体稳态中发挥关键作用[2]。在集约化水产养殖条件下,饲料引起的营养代谢负荷易导致肝细胞脂肪蓄积、抗氧化系统功能紊乱以及肠道屏障损伤,进而引发生长性能下降和疾病易感性上升。因此,开发能够协同改善肝肠健康的新型饲料原料,已成为推动水产养殖业可持续发展的重要研究方向。
黑水虻(Hermetia illucens L.),隶属于双翅目水虻科扁角水虻属,广泛分布于全球热带和温带地区[3],其幼虫能有效转化多种有机废物为高营养价值的蛋白质和生物肥料。黑水虻富含优质蛋白质且氨基酸均衡,同时还含有多种维生素及矿物质[4],在水产饲料中被视为最有应用前景的饲料原料之一[5]。除作为优质蛋白质源外,黑水虻还含有甲壳素[6]、抗菌肽[7]和月桂酸[8]等多种生物活性物质,是一种具有良好应用前景的功能性饲料原料。本团队在前期已探究了黑水虻干幼虫粉、脱脂幼虫粉、虫油以及鲜虫浆等常用制品在水产饲料上的应用效果,作为饲料中蛋白质或者脂肪原料的部分替代物,黑水虻制品在大口黑鲈(Micropterus salmoides)[9-10]、花鲈(Lateolabrax japonicus)[11-12]、杂交鳢(Channa maculate♀×Channa argus ♂)[13-14]、黄颡鱼(Pelteobagrus fulvidraco)[15-18]、吉富罗非鱼(Oreochromis niloticus)[19]及凡纳滨对虾(Litopenaeus vannamei)[20-21]等水产动物上均具有较好的应用效果,并辅有降低血脂、减少肝脏脂肪沉积、促进肝脏和肠道发育、提高免疫能力、增强抗氧化功能以及改善肠道菌群结构等益处。作为饲料添加剂,饲料中添加黑水虻虫浆对大口黑鲈[22-23]、杂交鳢[24-25]有明显的促生长作用,并且表现出促进蛋白质和脂质沉积、增强免疫与抗氧化能力、增加肠道菌群多样性、改善肠道健康等作用。
采用微波或热风等干燥方法制备黑水虻干虫或虫粉成本较高且加工过程会损失生物活性物质[26],黑水虻鲜虫浆的运输与储存成本较高[27]。相比之下,将黑水虻幼虫匀浆及酵解处理制成发酵黑水虻,不仅可降低加工成本,且产物中还富含由黑水虻蛋白转化产生的抗氧化肽[28]及外壳几丁质转化产生的壳聚糖[29]等生物活性物质,理论上发酵黑水虻作用效果更优。然而,关于发酵黑水虻的应用研究较为匮乏,目前仅见大口黑鲈[30-31]、对虾[32]及杂交鳢[27]上有相关报道。
斑点叉尾鮰(Ictalurus punctatus)隶属于鲶形目鮰科,是我国重要的水产养殖品种,具有对环境适应性强、食性广、抗病抗逆能力强、肉质紧致、味道鲜美以及几乎没有肌间刺的优点,深受养殖者和消费者青睐。据统计,2023年我国斑点叉尾鮰养殖产量达44.1万t[33]。本团队在前期研究中发现,饲料中添加2%和4%的发酵黑水虻可提高斑点叉尾鮰的增重率,并且添加2%的发酵黑水虻还可改善脂质代谢并减轻肝脏负担,促进肠道发育并提升营养吸收效率,增强非特异性免疫力[34]。然而,上述有益效应背后的深层机制,尤其是其对肝脏抗氧化防御体系及肠道屏障功能的调控作用,仍有待深入阐明。本试验旨在系统探究发酵黑水虻对斑点叉尾鮰肝脏组织形态、抗氧化能力以及肠道免疫与抗氧化能力、紧密连接蛋白基因表达、菌群结构的影响,从肝脏与肠道健康的角度揭示发酵黑水虻的作用机理,为其作为一种功能性饲料原料在斑点叉尾鮰精准饲料配方中的应用提供理论依据,这对促进黑水虻资源的高值化利用和水产养殖业的减抗、健康养殖具有重要的实际应用价值。

1 材料与方法

1.1 试验材料

黑水虻酵素由广州飞禧特生物科技有限公司提供,由喂食餐厨垃圾的8日龄幼虫经粉碎磨浆,经酶解及发酵等工序加工精制而成,其风干物质中含有25.60%的粗蛋白质、9.61%的粗脂肪、6.73%的粗灰分、25.00%的总氨基酸、7.42%的肽、9.61%的酸溶蛋白及0.18%的几丁质。

1.2 试验设计

动物试验已由广东省农业科学院动物科学研究所实验动物伦理委员会批准,批准号:2023012。
以鱼粉、鸡肉粉、豆粕、双低菜籽粕和米糠粕为蛋白质源,豆油为脂肪源,高精面粉为碳水化合物源,并添加0(作为对照)、2%、4%和8%的发酵黑水虻鲜物质(对应的发酵黑水虻风干物质的添加量分别为0、10.4、20.8和41.6 g/kg),配制4种等脂等氮的试验饲料,并分别记为G0、G2、G4和G8。试验饲料组成及营养水平参见本团队前期发表的相关文献[34]。
选取外观健康、活力强健的斑点叉尾鮰鱼苗480尾,初始体质量为(2.75±0.01) g,随机分成4组(每组3个重复,每个重复40尾),对应饲喂试验饲料G0、G2、G4和G8,进行56 d的养殖试验。养殖试验在广东省农业科学院动物科学研究所白云试验基地循环系统玻璃纤维缸(直径80 cm,高70 cm,容积350 L)中进行,试验鱼的饲养管理参见本团队前期发表的相关文献[34]。

1.3 样品采集

养殖结束后,禁食24 h,每缸随机取2尾鱼,解剖取肝脏于10%福尔马林溶液中固定,备用;每缸随机取7尾鱼于冰上解剖,取肝脏及中肠,-80 ℃冰箱保存,用于检测抗氧化指标和非特异性免疫酶;每缸随机取2尾鱼于冰上解剖,取中肠置于-80 ℃冰箱保存,用于测定肠道紧密连接蛋白基因的表达情况;每缸随机取2尾鱼于冰上解剖,取后肠于-80 ℃冰箱保存,用于测定肠道菌群结构。

1.4 指标测定

1.4.1 肝脏组织结构

将在10%福尔马林溶液中固定的肝脏取出,制备成石蜡组织切片,经苏木精-伊红(HE)染色后用Pannoramic全景切片扫描仪扫描,观察肝脏组织形态与结构。

1.4.2 肝脏和肠道抗氧化指标

取肝脏和肠道分别匀浆并在4 ℃下以624.35×g离心15 min,收集上清液后进行抗氧化指标检测。总蛋白(TP)、蛋白质羰基(PrC)、丙二醛(MDA)含量,总抗氧化能力(T-AOC)及过氧化氢酶(CAT)、谷胱甘肽过氧化物酶(GSH-Px)、总超氧化物歧化酶(T-SOD)活性均使用商业试剂盒(南京建成生物工程研究所)按照说明书方法检测。

1.4.3 肠道非特异性免疫酶活性

取1.4.2中制备的肠道上清液,使用商业试剂盒(南京建成生物工程研究所)检测溶菌酶(LZM)含量及酸性磷酸酶(ACP)、碱性磷酸酶(AKP)活性。

1.4.4 肠道紧密连接蛋白基因

取-80 ℃冰箱保持的中肠样品,采用TRIzol法提取总RNA,并测定RNA的质量和完整性。将提取的RNA使用反转录试剂盒反转录合成cDNA。基于NCBI数据库获得斑点叉尾鮰闭锁小带蛋白-1(ZO-1)及闭合蛋白-2(Claudin-2)基因序列,以β-肌动蛋白(β-actin)为内参基因,在Primer-BLAST程序中设计引物,由生工生物工程(上海)股份有限公司合成引物,引物序列见表1。实时荧光定量PCR反应体系(20.0 μL):10 μL Taq Pro Universal SYBR qPCR Master Mix、1 μL cDNA、0.4 μL上游引物、0.4 μL下游引物、8.2 μL RNase free water;反应程序:95 ℃ 30 s;95 ℃ 8 s,56 ℃ 30 s,40个循环。采用2-ΔΔCt法计算目的基因的相对表达量。
表1 肠道紧密连接蛋白相关基因引物序列

Table 1 Primer sequences of genes related to intestinal tight junction proteins

基因
Genes		 上游引物序列
Forward primer sequences (5'—3')		 下游引物序列
Reverse primer sequences (5'—3')
闭锁小带蛋白-1 ZO-1 TACCAAACCGTGGATACAAACC CTTCTATGGGTGGAGGAGGC
闭合蛋白-2 Claudin-2 CAAATTTGCAGGGCAGCAGT AAGAAGACAATCTATATTCGCCCA

1.4.5 肠道菌群

委托北京诺禾致源科技股份有限公司利用Illumina MiSeq测序平台,对后肠组织进行DNA提取和PCR扩增,通过16S rRNA高通量测序,对肠道菌群进行分类鉴定和多样性分析。

1.5 数据统计与分析

采用SPSS 26.0软件对试验数据进行单因素方差分析(one-way ANOVA),并采用Duncan氏法分析组间差异显著性,差异显著水平为P<0.05。试验数据分析结果均采用平均值±标准误表示。

2 结果与分析

2.1 肝脏组织结构

由图1可见,各组肝脏组织未见坏死,肝细胞排列均较为整齐,细胞界限较为明显,细胞核居中清晰可见,均无明显空泡化出现,无明显炎症细胞浸润。
图1 饲料中添加发酵黑水虻对斑点叉尾鮰肝脏组织结构的影响

黑色箭头为肝细胞 。

Fig.1 Effects of dietary supplementation of fermented black soldier fly on hepatic histoarchitecture of channel catfish (40×)

The black arrow indicates hepatocytes.

2.2 肝脏抗氧化指标

由表2可知,相较于对照组(G0组),G2和G4组肝脏T-AOC和CAT活性显著升高(P<0.05),G4组肝脏T-SOD活性显著升高(P<0.05),G2、G4及G8组肝脏PrC含量显著降低(P<0.05);各组间肝脏GSH-Px活性和MDA含量无显著差异(P>0.05)。
表2 饲料中添加发酵黑水虻对斑点叉尾鮰肝脏抗氧化指标的影响

Table 2 Effects of dietary supplementation of fermented black soldier fly on hepatic antioxidant indexes of channel catfish

项目
Items		 组别Groups
G0 G2 G4 G8
总抗氧化能力T-AOC/(U/mg prot) 0.58±0.07b 1.02±0.04a 0.88±0.05a 0.57±0.08b
过氧化氢酶CAT/(U/mg prot) 49.87±1.58c 76.91±5.33a 64.60±3.29b 45.29±3.44c
谷胱甘肽过氧化物酶GSH-Px/(U/mg prot) 468.75±17.89 491.66±24.33 487.87±12.33 505.67±31.69
总超氧化物歧化酶T-SOD/(U/mg prot) 43.14±0.52b 46.82±3.84ab 53.53±2.33a 49.73±3.32ab
蛋白质羰基PrC/(nmol/mg prot) 6.38±0.11a 3.44±0.75b 2.46±0.33bc 1.58±0.35c
丙二醛MDA/(nmol/mg prot) 2.98±0.25 3.39±0.18 3.44±0.23 3.86±0.58

同行数据肩标无字母或相同字母表示差异不显著(P>0.05),不同小写字母表示差异显著(P<0.05)。下表同。

In the same row, values with no letter or the same letter superscripts mean no significant difference (P>0.05), while with different small letter superscripts mean significant difference (P<0.05). The same as below.

2.3 肠道免疫与抗氧化指标

由表3可知,与对照组相比,G2组肠道AKP活性与G4组肠道ACP活性显著增加(P<0.05),且G2组肠道CAT活性显著增加(P<0.05),同时G2、G4和G8组肠道MDA含量显著降低(P<0.05);各组间肠道LZM含量及T-AOC活性无显著差异(P>0.05)。
表3 饲料中添加发酵黑水虻对斑点叉尾鮰肠道免疫与抗氧化指标的影响

Table 3 Effects of dietary supplementation of fermented black soldier fly on intestinal immune and antioxidant indexes of channel catfish

项目
Items		 组别Groups
G0 G2 G4 G8
溶菌酶LZM/(μg/mg prot) 54.24±5.82 41.21±4.01 45.98±7.80 44.74±5.64
碱性磷酸酶AKP/(金氏单位/g prot) 81.51±7.95b 110.03±4.26a 95.80±2.67ab 95.36±7.26ab
酸性磷酸酶ACP/(金氏单位/g prot) 13.90±0.37b 17.85±0.89ab 20.37±1.73a 15.50±1.41ab
过氧化氢酶CAT/(U/mg prot) 37.80±1.50b 46.58±1.45a 35.81±1.18b 40.51±3.78ab
总抗氧化能力T-AOC/(U/mg prot) 1.82±0.06 1.86±0.16 1.68±0.13 1.70±0.12
丙二醛MDA/(nmol/mg prot) 6.31±0.54a 4.48±0.32b 3.87±0.44b 3.72±0.64b

2.4 肠道紧密连接蛋白基因

由图2可知,与对照组相比,G2组肠道ZO-1和Claudin-2基因的相对表达量显著上升(P<0.05)。
图2 饲料中添加发酵黑水虻对斑点叉尾鮰肠道紧密连接蛋白基因表达的影响

数据柱标注不同小写字母表示差异显著(P<0.05)。

Fig.2 Effects of dietary supplementation of fermented black soldier fly on expression of genes related to intestinal tight junction proteins of channel catfish

Data columns with different letters mean significant difference (P<0.05).

2.5 肠道菌群

2.5.1 肠道菌群多样性

如图3所示,饲料中添加不同水平发酵黑水虻对斑点叉尾鮰肠道菌群(属水平)的α多样性指数如Shannon(P=0.181)、Simpson(P=0.104)、Chao1(P=0.510)、Dominance指数(P=0.104)均无显著影响。
图3 属水平下斑点叉尾鮰肠道菌群的α多样性

Fig.3 α diversity of intestinal microbiota at genus level of channel catfish

主坐标分析(PCoA)(图4)显示,各样本间聚集及区分不明显,表明各组之间肠道菌群的β多样性相似。
图4 属水平下斑点叉尾鮰肠道菌群的β多样性(主坐标分析)

Fig.4 β diversity of intestinal microbiota at genus level of channel catfish (PCoA)

2.5.2 肠道菌群组成

如图5-A所示,在门水平上,斑点叉尾鮰肠道中的主要菌门有变形菌门(Proteobacteria)、梭杆菌门(Fusobacteriota)、厚壁菌门(Firmicutes)、放线菌门(Actinobacteriota)、拟杆菌门(Bacteroidota)、蓝细菌门(Cyanobacteria)等,其中G0和G2组的优势菌门为变形菌门和梭杆菌门,G4组的优势菌门为变形菌门和放线菌门,G8组的优势菌门虽也为变形菌门和梭杆菌门,但变形菌门的相对丰度下降,梭杆菌门的相对丰度增加;厚壁菌门的相对丰度随发酵黑水虻添加量的增加呈先增加后下降趋势。如图5-B所示,在属水平上,G0、G2和G8组斑点叉尾鮰肠道中优势菌属均为鲸杆菌属(Cetobacterium)和邻单胞菌属(Plesiomonas),G4组的优势菌属则为不动杆菌属(Acinetobacter);此外,G8组鲸杆菌属的相对丰度与其他组相比明显升高。
图5 饲料中添加发酵黑水虻对斑点叉尾鮰肠道菌群在门(A)和属(B)水平上组成的影响

Fig.5 Effects of dietary supplementation of fermented black soldier fly on intestinal microbiota composition at phylum (A) and genus (B) levels of channel catfish

3 讨论

3.1 饲料中添加发酵黑水虻对斑点叉尾鮰肝脏健康的影响

鱼类肝脏结构的完整性是其生理功能的基础,营养代谢异常引发的病理改变常伴随氧化应激[35]。研究表明,黑水虻外骨骼高几丁质含量是诱导肝脏损伤的关键限制因子。本团队前期发现,添加9.6%的脱脂黑水虻虫粉可致鲈鱼肝脏空泡化[11],添加2%~8%黑水虻鲜虫浆可导致大口黑鲈肝细胞液泡变性及炎症浸润[23]。上述研究结果均显示了黑水虻中几丁质的负面影响。高剂量的几丁质通过干扰脂质代谢[36]以及激活炎症通路[37]损伤肝脏。本试验发现,黑水虻虫浆经发酵处理制成发酵黑水虻后,适宜的添加量(2%~8%鲜物质)下未对斑点叉尾鮰的肝脏产生损伤。该结果与在巴沙鱼(Pangasianodon hypophthalmus)[38]及大口黑鲈[30]的研究中得出的发酵黑水虻维持肝脏健康的结论一致,其关键机制在于发酵过程可将几丁质降解为小分子壳聚糖。
本试验进一步研究发现,发酵黑水虻不仅可缓解斑点叉尾鮰的肝脏损伤,还可通过调节氧化-抗氧化平衡系统提高鱼体健康水平。氧化应激本质是机体氧化-抗氧化系统失衡[39],其中线粒体膜超极化促进的活性氧(ROS)过度生成[40],可攻击生物膜多不饱和脂肪酸引发以MDA为标志物的脂质过氧化[41],同时导致蛋白质氧化形成羰基化产物。本研究中,与未添加发酵黑水虻的对照组相比,添加不同水平发酵黑水虻的3个试验组斑点叉尾鮰的肝脏PrC含量显著降低,说明在斑点叉尾鮰饲料中添加发酵黑水虻可降低鱼体氧化损伤,减少蛋白质氧化产物的产生,这与在大口黑鲈上得出的饲料中添加发酵黑水虻降低氧化损伤的研究结论[30]一致。T-AOC反映的是机体的整体抗氧化能力,超氧化物歧化酶(SOD)和CAT通过消除自由基提供对氧化损伤的保护,是关键抗氧化酶[42]。本研究中,饲料中添加2%~4%发酵黑水虻鲜物质可显著提高斑点叉尾鮰肝脏T-AOC和CAT活性,添加4%发酵黑水虻鲜物质还可显著提高肝脏T-SOD活性,说明适宜添加量的发酵黑水虻可通过增强自由基清除效能改善机体抗氧化功能。这一结论在不同的鱼种也得到了验证:在大口黑鲈[30-31]和杂交鳢[27]饲料中添加适量发酵黑水虻均可增强鱼体的抗氧化能力;当巴沙鱼饲料中发酵黑水虻虫粉替代鱼粉量超过25%时,其抗氧化能力增强[38]。减少氧化产物及增强抗氧化能力这种双重保护机制可能与黑水虻虫浆经发酵后的产物特性密切相关:一方面,黑水虻虫浆中几丁质的降解极大消除了促氧化因素;另一方面,黑水虻虫浆中富含的月桂酸可能通过激活核因子E2相关因子2(Nrf2)/抗氧化反应元件(ARE)信号通路上调抗氧化酶表达,协同增强机体的抗氧化防御能力[43]。

3.2 饲料中添加发酵黑水虻对斑点叉尾鮰肠道屏障功能的影响

肠道是鱼类进行营养物质消化吸收的场所,其屏障系统更承担着抵御病原体入侵及维持内环境稳态的重要功能[44]。本试验探究了发酵黑水虻对斑点叉尾鮰肠道多重屏障的协同调控机制。
在机械屏障层面,肠道紧密连接由闭合蛋白(Claudins)和闭锁小带蛋白(ZOs)等紧密连接蛋白构成,通过连接肠上皮细胞形成选择性屏障,其完整性破坏将导致病原体入侵并诱发炎症反应[45]。本研究结果显示,饲料中添加2%发酵黑水虻鲜物质可显著上调斑点叉尾鮰肠道ZO-1和Claudin-2基因的相对表达量,表明该添加量下发酵黑水虻可提高斑点叉尾鮰肠道紧密连接蛋白的完整性,降低肠道通透性,从而强化机械屏障功能。这与吴佳瑄[46]报道的黄粉虫粉改善大口黑鲈肠道屏障功能的结果相一致。发酵处理可使黑水虻外骨骼中的几丁质降解为生物活性更高的壳聚糖,后者可通过激活细胞外信号调节激酶1/2(ERK1/2)信号通路促进紧密连接蛋白合成[47],这可能是发酵黑水虻改善肠道屏障完整性的重要分子基础。
在免疫屏障层面,LZM通过水解细菌肽聚糖层破坏细胞壁完整性,介导广谱抗菌作用[46]。AKP和ACP参与非特异性免疫反应,在免疫系统中起重要作用[48-50]。本试验中,饲料中添加2%和4%发酵黑水虻分别显著提高斑点叉尾鮰肠道AKP及ACP活性,表明发酵黑水虻可通过增强非特异性免疫酶活性改善肠道免疫功能。发酵黑水虻中高含量的月桂酸可能通过直接提高AKP和ACP活性以及调节TLR/NF-κB等炎症信号通路来增强肠道免疫功能[51]。此外,本试验中添加发酵黑水虻能显著改善斑点叉尾鮰肠道的抗氧化状态,具体表现为G2组CAT活性显著升高以及各添加组MDA含量显著降低。发酵黑水虻对肠道抗氧化状态的改善与其对免疫功能的增强密切相关,一方面,CAT活性的提升和MDA含量的下降,直接清除了过量的ROS,减轻了氧化应激对肠道上皮细胞的损伤,这不仅保护了细胞膜的完整性,也避免了过量ROS破坏紧密连接蛋白,从而为前述ZO-1和Claudin-2基因表达的上调创造了有利的微环境,巩固了机械屏障;另一方面,氧化应激的缓解间接稳定了免疫屏障,MDA等脂质过氧化产物本身即可激活炎症通路,其含量的降低有助于抑制过度的炎症反应。同时,一个氧化还原稳定的肠道环境确保了AKP及ACP等非特异性免疫酶能够高效行使功能。
在微生物屏障方面,饲料中添加发酵黑水虻虽未显著改变斑点叉尾鮰肠道菌群的α多样性和β多样性,但引发了具有功能特异性的菌群结构变化。饲喂添加2%发酵黑水虻鲜物质饲料的斑点叉尾鮰肠道菌群中,擅长分解复杂碳水化合物并产生如丁酸和乙酸等短链脂肪酸的厚壁菌门的相对丰度增加,这些短链脂肪酸可为肠上皮细胞供能并提升营养吸收效率[52],这与本团队前期观察到的该组增重率提高相呼应[34];饲喂添加8%发酵黑水虻鲜物质饲料的斑点叉尾鮰肠道菌群中,梭杆菌门尤其是鲸杆菌属的相对丰度异常升高。梭杆菌门可利用发酵产物进行增殖,并通过维生素B12合成和短链脂肪酸代谢等途径直接促进宿主生长并抑制致病菌[53-54],此类功能变化无需依赖整体菌群多样性的改变。在大口黑鲈饲料中添加发酵黑水虻后也观察到类似的梭杆菌门优势转变[30]。此外,含有多种条件致病菌的变形菌门的相对丰度降低,这可能与发酵产物中几丁质衍生物及有机酸等成分的抗菌作用有关,进一步优化了肠道微生态环境。

4 结论

综上所述,在斑点叉尾鮰饲料中添加2%~4%的发酵黑水虻鲜物质能维持肝脏健康,提高肝脏抗氧化能力,增强肠道机械屏障、免疫屏障功能以及抗氧化能力,改善肠道菌群结构。

References

Publishing order | Descend order by publishing year | Descend order by cited within
[1]
MA X C, LU Q S, HAN G L, et al. Effects of different carbohydrate levels on glycolipid metabolism and liver health of channel catfish (Ictalurus punctatus)[J]. Aquaculture Reports, 2025, 43:102921.

DOI

[2]
ARAGÃO C, GONÇALVES A T, COSTAS B, et al. Alternative proteins for fish diets:implications beyond growth[J]. Animals, 2022, 12(9):1211.

DOI

[3]
SINGH A, KUMARI K. An inclusive approach for organic waste treatment and valorisation using black soldier fly larvae:a review[J]. Journal of Environmental Management, 2019, 251:109569.

DOI

[4]
LILAND N S, ARAUJO P, XU X X, et al. A Meta-analysis on the nutritional value of insects in aquafeeds[J]. Journal of Insects as Food and Feed, 2021, 7(5):743-759.

DOI

[5]
VAN HUIS A. Edible insects:future prospects for food and feed security[M]. Rome: Food and Agriculture Organization of the United Nations, 2013.

[6]
ELKADAOUI S, AZZI M, DESBRIERES J, et al. Valorization of Hermetia illucens breeding rejects by chitins and chitosans production. influence of processes and life cycle on their physicochemical characteristics[J]. International Journal of Biological Macromolecules, 2024, 266(Pt 2):131314.

DOI

[7]
ZHANG Z L, MENG Y Q, LI J J, et al. Effects of antimicrobial peptides from dietary Hermetia illucens larvae on the growth,immunity,gene expression,intestinal microbiota and resistance to Aeromonas hydrophila of juvenile red claw crayfish (Cherax quadricarinatus)[J]. Fish & Shellfish Immunology, 2024, 147:109437.

[8]
HURTADO-RIBEIRA R, SILVAN J M, FORNARI T, et al. Modulation of the lipolysis and subsequent antibacterial activity of the fat from black soldier fly (Hermetia illucens) by the combined selection of slaughtering,drying and defatting methods of the larvae[J]. Innovative Food Science and Emerging Technologies, 2023, 90:103510.

DOI

[9]
彭凯, 萧鸿发, 莫文艳, 等. 黑水虻幼虫粉替代鱼粉对加州鲈生长性能、形体指标、体成分及营养物质沉积率的影响[J]. 动物营养学报, 2021, 33(11):6340-6348.

DOI

PENG K, XIAO H F, MO W Y, et al. Effects of replacing fish meal with black soldier fly larvae meal on growth performance,physical indexes,body composition and nutrient retention rates of Micropterus salmoides[J]. Chinese Journal of Animal Nutrition, 2021, 33(11):6340-6348. (in Chinese)

[10]
彭凯, 萧鸿发, 莫文艳, 等. 饲料中添加黑水虻幼虫粉对加州鲈血清代谢物、抗氧化与免疫指标及肠道组织结构的影响[J]. 动物营养学报, 2021, 33(12):6964-6972.

DOI

PENG K, XIAO H F, MO W Y, et al. Effects of dietary black soldier fly larvae meal on serum metabolites,antioxidant and immune indexes and intestinal histological appearance of Micropterus salmoides[J]. Chinese Journal of Animal Nutrition, 2021, 33(12):6964-6972. (in Chinese)

[11]
WANG G X, PENG K, HU J R, et al. Evaluation of defatted black soldier fly (Hermetia illucens L.) larvae meal as an alternative protein ingredient for juvenile Japanese seabass (Lateolabrax japonicus) diets[J]. Aquaculture, 2019, 507:144-154.

DOI

[12]
胡俊茹, 王国霞, 莫文艳, 等. 黑水虻幼虫粉替代鱼粉对鲈鱼幼鱼生长性能、体组成、血浆生化指标和组织结构的影响[J]. 动物营养学报, 2018, 30(2):613-623.

HU J R, WANG G X, MO W Y, et al. Effects of fish meal replacement by black soldier fly (Hermetia illucens L.) larvae meal on growth performance, body composition,plasma biochemical indexes and tissue structure of juvenile Lateolabrax japonicas[J]. Chinese Journal of Animal Nutrition, 2018, 30(2):613-623. (in Chinese)

[13]
王国霞, 莫文艳, 范怡杰, 等. 黑水虻对杂交鳢生长、肌肉组成和血清指标的影响[J]. 水产科学, 2019, 38(5):603-609.

WANG G X, MO W Y, FAN Y J, et al. Effect of black soldier fly larvae on growth performance,muscular composition and serum indices of hybrid snakehead Channa argus×C.maculata[J]. Fisheries Science, 2019, 38(5):603-609. (in Chinese)

[14]
谢雨桐, 黄文庆, 李诗洋, 等. 黑水虻虫粉替代鱼粉对杂交鳢血清免疫抗氧化、肠道功能和抗病力的影响[J]. 水产学报, 2024, 48(11):163-173.

XIE Y T, HUANG W Q, LI S Y, et al. Effects of replacing fish meal with black soldier fly larvae meal on serum immune antioxidant indices,intestinal function and disease resistance of hybrid snakehead (Channa maculata♀×Channa argus ♂)[J]. Journal of Fisheries of China, 2024, 48(11):163-173. (in Chinese)

[15]
王国霞, 陈冰, 孙育平, 等. 脱脂亮斑扁角水虻幼虫粉替代鱼粉对黄颡鱼幼鱼生长性能、营养素沉积率、血清生化指标和消化酶活性的影响[J]. 水产学报, 2020, 44(6):987-998.

WANG G X, CHEN B, SUN Y P, et al. Effects of replacing fish meal with defatted black soldier fly (Hermetia illucens) larvae meal on growth performance,nutrient retention,serum biochemical parameters and digestive enzymes activity of juvenile Pelteobagrus fulvidraco[J]. Journal of Fisheries of China, 2020, 44(6):987-998. (in Chinese)

[16]
WANG G, ZHAO H, HU J, et al. Effects of substitution of fish meal with black soldier fly (Hermetia illucens) larvae meal,in yellow catfish (Pelteobagrus fulvidraco) diets[J]. Israeli Journal of Aquaculture-Bamidgeh, 2017, 69(1):1382.

[17]
胡俊茹, 易昌金, 王国霞, 等. 黑水虻虫油替代豆油对黄颡鱼幼鱼生长、血清生化指标和肝脏脂滴面积的影响[J]. 水生生物学报, 2020, 44(4):717-727.

HU J R, YI C J, WANG G X, et al. Effects of dietary soybean oil replaced with black soldier fly larvae oil on growth performance,plasma biochemical indexes and liver lipid droplets of juvenile yellow catfish[J]. Acta Hydrobiologica Sinica, 2020, 44(4):717-727. (in Chinese)

[18]
陈晓瑛, 胡俊茹, 王国霞, 等. 黑水虻幼虫粉替代鱼粉对黄颡鱼幼鱼生长性能、肌肉品质及血清生化指标的影响[J]. 动物营养学报, 2019, 31(6):2788-2799.

CHEN X Y, HU J R, WANG G X, et al. Effects of fish meal replacement by black soldier fly (Hermetia illucens) larvae meal on growth performance,serum biochemical indices and meat quality of juvenile yellow catfish (Pelteobagrus fulvidraco)[J]. Chinese Journal of Animal Nutrition, 2019, 31(6):2788-2799. (in Chinese)

[19]
胡俊茹, 王绥涛, 邱世殿, 等. 黑水虻幼虫培养基替代豆粕对吉富罗非鱼生长、体组成和血清生化指标的影响[J]. 淡水渔业, 2016, 46(6):98-103.

HU J R, WANG S T, QIU S D, et al. Effect of replacing soybean meal with black soldier fly (Hermetia illucens) larvae culture meal on growth performance,body composition and serum parameters for GIFT tilapia Oreochromis niloticus[J]. Freshwater Fisheries, 2016, 46(6):98-103. (in Chinese)

[20]
WANG G X, PENG K, HU J R, et al. Evaluation of defatted Hermetia illucens larvae meal for Litopenaeus vannamei:effects on growth performance,nutrition retention,antioxidant and immune response,digestive enzyme activity and hepatic morphology[J]. Aquaculture Nutrition, 2021, 27(4):986-997.

DOI

[21]
胡俊茹, 王国霞, 黄文庆, 等. 黑水虻幼虫粉替代鱼粉对凡纳滨对虾幼虾生长性能、体组成、血清生化指标和抗氧化能力的影响[J]. 动物营养学报, 2019, 31(11):5293-5300.

HU J R, WANG G X, HUANG W Q, et al. Effects of fish meal replacement by black soldier fly (Hermetia illucens) larvae meal on growth performance,body composition,serum biochemical indexes and antioxidant ability of juvenile Litopenaeus vannamei[J]. Chinese Journal of Animal Nutrition, 2019, 31(11):5293-5300. (in Chinese)

[22]
PENG K, MO W, XIAO H, et al. Dietary black soldier fly pulp affects growth,antioxidant and immune capacity of Micropterus salmoides[J]. Journal of Insects as Food and Feed, 2022, 8(11):1197-1203.

DOI

[23]
PENG K, MO W Y, XIAO H F, et al. Effects of black soldier fly pulp on growth performance,histomorphology and lipid metabolism gene expression of Micropterus salmoides[J]. Aquaculture Reports, 2021, 20:100737.

DOI

[24]
谢雨桐, 李诗洋, 彭凯, 等. 黑水虻虫浆对杂交鳢生长性能、营养物质沉积率和血清生化指标的影响[J]. 动物营养学报, 2023, 35(6):3878-3887.

DOI

XIE Y T, LI S Y, PENG K, et al. Effects of black soldier fly larvae pulp on growth performance,nutrient deposition rates and serum biochemical indices of hybrid snakehead[J]. Chinese Journal of Animal Nutrition, 2023, 35(6):3878-3887. (in Chinese)

[25]
谢雨桐, 朱喜锋, 李诗洋, 等. 黑水虻虫浆对杂交鳢血清免疫、抗氧化指标和肠道形态、菌群及抗病力的影响[J]. 动物营养学报, 2023, 35(8):5297-5308.

DOI

XIE Y T, ZHU X F, LI S Y, et al. Effects of black soldier fly larvae pulp on serum immune and antioxidant indices,intestinal morphology,and disease resistance of hybrid snakehead[J]. Chinese Journal of Animal Nutrition, 2023, 35(8):5297-5308. (in Chinese)

[26]
王斌, 邹仕庚, 彭运智, 等. 黑水虻在畜禽饲料中的应用研究进展[J]. 中国畜牧杂志, 2021, 57(6):8-15.

WANG B, ZOU S G, PENG Y Z, et al. Research progress on application of black soldier fly (Hermetia illucens) in animal feed[J]. Chinese Journal of Animal Science, 2021, 57(6):8-15. (in Chinese)

[27]
朱喜锋, 黄文庆, 李国立, 等. 发酵黑水虻对杂交鳢生长性能、体成分及抗氧化和免疫能力的影响[J]. 中国畜牧兽医, 2024, 51(9):3807-3816.

DOI

ZHU X F, HUANG W Q, LI G L, et al. Effects of fermented black soldier fly (Hermetia illucens L.) on growth performance,body composition,antioxidant and immune ability of hybrid snakehead[J]. China Animal Husbandry & Veterinary Medicine, 2024, 51(9):3807-3816. (in Chinese)

[28]
ZHU D, HUANG X W, TU F, et al. Preparation,antioxidant activity evaluation,and identification of antioxidant peptide from black soldier fly (Hermetia illucens L.) larvae[J]. Journal of Food Biochemistry, 2020, 44(5):e13186.

[29]
KACZMAREK M B, STRUSZCZYK-SWITA K, LI X K, et al. Enzymatic modifications of chitin,chitosan,and chitooligosaccharides[J]. Frontiers in Bioengineering and Biotechnology, 2019, 7:243.

DOI

[30]
XU F M, HOU S W, WANG G X, et al. Effects of zymolytic black soldier fly (Hermetia illucens) pulp as dietary supplementation in largemouth bass (Micropterus salmoides)[J]. Aquaculture Reports, 2021, 21:100823.

DOI

[31]
黄文庆, 王梦华, 李国立, 等. 发酵黑水虻对大口黑鲈生长性能、体成分、抗氧化指标以及肠道组织结构的影响[J]. 饲料工业, 2024, 45(12):20-28.

HUANG W Q, WANG M H, LI G L, et al. Effects of fermented black soldier fly larvae (Hermetia illucens) on growth performance,body composition,antioxidant indexes and intestinal structure of Micropterus salmoides[J]. Feed Industry, 2024, 45(12):20-28. (in Chinese)

[32]
王梦华, 周萌, 黄文庆, 等. 饲料中添加发酵黑水虻对凡纳滨对虾生长性能、体成分和消化性能的影响[J]. 饲料工业, 2024, 45(12):28-37.

WANG M H, ZHOU M, HUANG W Q, et al. Effects of adding fermented Hermetia illucens L.in diets on growth performance,body composition and digestive performance of Litopenaeus vannamei[J]. Feed Industry, 2024, 45(12):28-37. (in Chinese)

[33]
中华人民共和国农业农村部渔业渔政管理局, 全国水产技术推广总站, 中国水产学会. 2024中国渔业统计年鉴[M]. 北京: 中国农业出版社, 2024.

Fishery Administration Bureau of the Ministry of Agriculture and Rural Affairs of the People’s Republic of China,National Aquaculture Technology Promotion Station,Chinese Fisheries Society. 2024 China fishery statistical yearbook[M]. Beijing: China Agriculture Press,2024. (in Chinese)

[34]
郑雨顺, 陈桂琼, 黄文庆, 等. 发酵黑水虻对斑点叉尾鮰生长、肝脏生化指标、消化吸收和免疫力的影响[J]. 动物营养学报, 2025, 37(3):1940-1953.

DOI

ZHENG Y S, CHEN G Q, HUANG W Q, et al. Effects of fermented black soldier fly on growth,liver biochemical indices,digestion and absorption and immunity of channel catfish (Ictalurus punctatus)[J]. Chinese Journal of Animal Nutrition, 2025, 37(3):1940-1953. (in Chinese)

[35]
SHI X, LUO Z, CHEN G H, et al. Replacement of fishmeal by a mixture of soybean meal and chlorella meal in practical diets for juvenile crucian carp,Carassius auratus[J]. Journal of the World Aquaculture Society, 2017, 48(5):770-781.

DOI

[36]
LI S L, JI H, ZHANG B X, et al. Defatted black soldier fly (Hermetia illucens) larvae meal in diets for juvenile Jian carp (Cyprinus carpio var. Jian):growth performance,antioxidant enzyme activities,digestive enzyme activities,intestine and hepatopancreas histological structure[J]. Aquaculture, 2017, 477:62-70.

DOI

[37]
ZARANTONIELLO M, BRUNI L, RANDAZZO B, et al. Partial dietary inclusion of Hermetia illucens (black soldier fly) full-fat prepupae in zebrafish feed:biometric,histological,biochemical,and molecular implications[J]. Zebrafish, 2018, 15(5):519-532.

DOI

[38]
ARDRA M, PRADHAN C, DAS S, et al. The effect of fishmeal replacement with organic acid fermented black soldier fly (Hermetia illucens) larvae meal on growth,nutrient utilization,metabolic enzyme activity,antioxidant status and immunity in pangasius (Pangasianodon hypophthalmus)[J]. Aquaculture, 2024, 591:741114.

DOI

[39]
AW T Y. Cellular redox:a modulator of intestinal epithelial cell proliferation[J]. Physiology, 2003, 18(5):201-204.

DOI

[40]
YAO W X, ZHANG C Y, MAO H X, et al. Effects of dietary defatted black soldier fly (Hermetia illucens) larvae meal substituting fish meal on growth,antioxidative capacity,immunity,intestinal histology and microbiota of juvenile Chinese mitten crab (Eriocheir sinensis)[J]. Aquaculture Reports, 2024, 38:102302.

DOI

[41]
YAO W X, LI X Q, ZHANG C Y, et al. Effects of dietary synbiotics supplementation methods on growth,intestinal health,non-specific immunity and disease resistance of Pacific white shrimp,Litopenaeus vannamei[J]. Fish & Shellfish Immunology, 2021, 112:46-55.

[42]
KERRIDGE A, LAPPIN-SCOTT H, STEVENS J R. Antibacterial properties of larval secretions of the blowfly,Lucilia sericata[J]. Medical and Veterinary Entomology, 2005, 19(3):333-337.

DOI

[43]
孙彩云, 董宏标, 王文豪, 等. 月桂酸单甘油酯对花鲈脂质代谢的影响[J]. 南方水产科学, 2021, 17(1):67-75.

SUN C Y, DONG H B, WANG W H, et al. Effects of glycerol monolaurate on lipid metabolism of Lateolabrax maculatus[J]. South China Fisheries Science, 2021, 17(1):67-75. (in Chinese)

[44]
ASSIMAKOPOULOS S F, TRIANTOS C, MAROULIS I, et al. The role of the gut barrier function in health and disease[J]. Gastroenterology Research, 2018, 11(4):261-263.

DOI PMID

[45]
GUAN P Y, YU H, WANG S N, et al. Dietary rutin alleviated the damage by cold stress on inflammation reaction,tight junction protein and intestinal microbial flora in the mice intestine[J]. The Journal of Nutritional Biochemistry, 2024, 130:109658.

DOI

[46]
吴佳瑄. 黄粉虫粉在大口黑鲈“优鲈3号”配合饲料中的应用研究[D]. 硕士学位论文. 重庆: 西南大学, 2023.

WU J X. Application of Tenebrio molitor meal in compound diet of Micropterus salmoides[D]. Master’s Thesis. Chongqing: Southwest University, 2023. (in Chinese)

[47]
LI Y, WU L Y, YONG Y H, et al. Enhancing gut barrier integrity:upregulation of tight junction proteins by chitosan oligosaccharide through the ERK1/2 signaling pathway[J]. Nutrition, 2024, 124:112428.

DOI

[48]
OKSANEN E. Lysozyme revisited[J]. Structure, 2025, 33(1):6-7.

DOI PMID

[49]
LALLÈS J P. Biology,environmental and nutritional modulation of skin mucus alkaline phosphatase in fish:a review[J]. Fish & Shellfish Immunology, 2019, 89:179-186.

[50]
LI J, WU Z B, ZHANG Z, et al. Effects of potential probiotic Bacillus velezensis K2 on growth,immunity and resistance to Vibrio harveyi infection of hybrid grouper (Epinephelus lanceolatus ♂×E. fuscoguttatus ♀)[J]. Fish & Shellfish Immunology, 2019, 93:1047-1055.

[51]
LIN H X, TAN B P, YANG Q H. The effect of glycerol monolaurate on intestinal health and disease resistance in cage-farmed juvenile pompano Trachinotus ovatus[J]. Aquaculture Nutrition, 2023, 2023:8580240.

[52]
KONONOVA S V, ZINCHENKO D V, MURANOVA T A, et al. Intestinal microbiota of salmonids and its changes upon introduction of soy proteins to fish feed[J]. Aquaculture International, 2019, 27(2):475-496.

DOI

[53]
TSUCHIYA C, SAKATA T, SUGITA H. Novel ecological niche of Cetobacterium somerae,an anaerobic bacterium in the intestinal tracts of freshwater fish[J]. Letters in Applied Microbiology, 2008, 46(1):43-48.

[54]
MA C, CHEN C X, JIA L, et al. Comparison of the intestinal microbiota composition and function in healthy and diseased Yunlong grouper[J]. AMB Express, 2019, 9(1):187.

DOI

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