基于转录组学解析铜驯化缓解高温胁迫对大黄鱼的氧化损伤

为探讨铜驯化对高温胁迫下大黄鱼(Larimichthys crocea)氧化损伤和基因表达水平的影响,将体质量为(48.92±3.62) g的大黄鱼暴露在铜离子浓度为0和10μg·L^-1的水体中14 d,再暴露在温度为32℃的水体中24 h。结果显示,高温胁迫显著增加了大黄鱼肝脏中活性氧(ROS)和脂质过氧化物(LPO)含量(P<0.05)。铜驯化对常温下ROS和LPO含量不产生影响,而显著降低了高温胁迫下大黄鱼ROS和LPO含量(P<0.05),表明铜驯化缓解了高温胁迫对大黄鱼的氧化损伤。从高温胁迫组vs对照组、铜驯化组vs对照组和(铜驯化+高温胁迫)组vs高温胁迫组中分别筛选到828个、2 311个和1 084个差异基因。GO和KEGG分析发现,差异基因主要富集在与TCA循环、氧化磷酸化、谷胱甘肽代谢、内质网蛋白加工、吞噬体和MAPK信号通路等相关功能上。聚类分析表明,高温胁迫下调了TCA循环、氧化磷酸化、谷胱甘肽代谢、吞噬体和MAPK信号等通路中的大部分基因表达,上调了内质网蛋白加工通路中的大部分基因表达,而铜驯化则对高温胁迫下大黄鱼的这些基因表...     

基于转录组解析铜驯化对低温胁迫下大黄鱼氧化损伤的影响

为探讨铜驯化对低温胁迫下大黄鱼(Larimichthys crocea)氧化损伤和基因表达水平的影响, 本研究将体重为 (48.92±3.62) g 的大黄鱼暴露在铜浓度为 0 和 10 μg/L 的水体中 14 d, 再暴露在温度为 8 ℃的水体中 24 h。结果显示, 低温胁迫显著增加了活性氧(ROS)和脂质过氧化物(LPO)含量。尽管铜驯化对 ROS 和 LPO 含量不产生影响, 但铜驯化显著增加了低温胁迫下大黄鱼 ROS 和 LPO 含量, 表明铜驯化加剧了低温胁迫对大黄鱼的氧化损伤。从铜驯化相对对照组、低温胁迫相对对照组和铜驯化+低温胁迫相对低温胁迫中分别筛选到 2288 个、1425 个和 1382 个差异基因。GO 和 KEGG 分析发现差异基因主要富集在与脂肪酸代谢、糖类有氧代谢、谷胱甘肽代谢、内质网应激、自噬和凋亡等相关的通路中。聚类分析表明, 低温胁迫上调了不饱和脂肪酸合成、内质网应激、自噬和凋亡等相关通路中的大部分基因表达, 而铜驯化则对低温胁迫下大黄鱼的这些基因表达调控产生了拮抗效应, 表明铜驯化通过抑制不饱和脂肪酸合成、内质网应激、自噬和凋亡来降低大黄鱼的低温胁迫耐受性。研究结果为深入研究铜污染物对大黄鱼低温胁迫耐受性的影响及其分子机制提供科学依据。     

盐度驯化改善大黄鱼盐度胁迫耐受性的作用机制

为探讨盐度驯化改善大黄鱼(Larimichthys crocea)盐度胁迫耐受性的作用机制,将大黄鱼暴露在盐度为25或20的水体中7 d,再暴露在盐度为12或40的水体中24 h,分为对照组(25)、低盐组(25+12)、高盐组(25+40)、低盐驯化组(20)、低盐驯化+低盐组(20+12)和低盐驯化+高盐组(20+40) 6个处理组,分别记为C组、CL组、CH组、A组、AL组、AH组。结果显示, CL vs C中大黄鱼肝脏的丙二醛(MDA)和脂质过氧化物(LPO)含量显著上升,超氧化物歧化酶(SOD)和溶菌酶(LZM)活性显著上升,过氧化氢酶(CAT)活性显著降低(P<0.05),碱性磷酸酶(AKP)的活性变化不显著(P>0.05)。CH vs C中MDA和LPO含量显著上升, SOD和LZM的活性显著上升, CAT和AKP显著降低(P<0.05)。A vs C中MDA和LPO含量显著上升(P<0.05), SOD、LZM和AKP的活性未发生显著变化(P>0.05), CAT活性显著降低(P<0.05)。AL vs CL与AH vs CH中MDA...     

铜驯化改善大黄鱼低温耐受性的作用机制

为了探讨铜驯化改善大黄鱼(Larimichthyscrocea)低温耐受性的作用机制,将大黄鱼幼鱼放入含Cu20μg/L的水体中驯化7 d,然后暴露在10℃的水体中6 h和24 h,检测其抗氧化能力、能量代谢和非特异性免疫力指标的变化。结果表明,低温胁迫增加了大黄鱼的活性氧(ROS)含量,加重了肝细胞畸形,表明低温胁迫对机体产生了氧化损伤。与低温胁迫相比,铜驯化+低温胁迫提高了谷胱甘肽过氧化物酶、过氧化氢酶、ATP合成酶、琥珀酸脱氢酶、碱性磷酸酶和溶菌酶的活性,增加了谷胱甘肽和ATP含量,降低了ROS、乳酸含量和肝细胞空泡率,表明铜驯化通过改善大黄鱼的抗氧化能力、能量代谢效率和非特异性免疫力来缓解低温胁迫对机体的损伤。结论认为,铜驯化通过产生适应性反应来提高大黄鱼的低温胁迫耐受性,表现了毒物兴奋效应作用。     

低盐胁迫下金乌贼转录组学分析

为探讨低盐胁迫对金乌贼影响的分子机制,通过转录组测序技术,测定了正常盐度(30)培养和低盐胁迫(15)6 h的孵出30 d、体质量(1.3±0.3)g金乌贼幼体的转录组数据。测序共获得87326026条序列,经过质量剪切和从头拼接得到575171条转录本和513053条Unigenes。分别在NR、Swiss-Prot、KEGG、String和Pfam数据库对Unigenes进行功能注释,共获得62485条注释结果。Unigenes包含数目较多的KEGG通路有嘌呤、嘧啶和碳代谢,PI3K-AKT、cAMP和Rap1信号通路,内吞作用,RNA转运,局灶性黏附,赖氨酸降解和泛素介导的蛋白水解等。低盐胁迫产生1923条差异表达基因,GO功能富集分析显示,一些可能与低盐胁迫相关的生物学过程如α-氨基酸、羧酸、氧乙酸、有机酸和RNA等代谢过程得到了显著富集。GO可视化分析发现,低盐胁迫对金属离子、阴离子及核苷酸结合,α-氨基酸代谢和水解酶活性等过程影响显著。KEGG通路富集分析显示,低盐胁迫6 h后差异表达基因主要富集到雌激素和心肌细胞肾上腺素信号通路,类固醇生物合成,抗原加工与表达,脂肪和蛋白质消化与吸收,甘油脂质、花生四烯酸和酪氨酸代谢等信号通路上。本研究中获得的通路及基因信息可为今后开展金乌贼低盐胁迫生理机制的探讨、分子标记的挖掘和关键基因的克隆等提供技术支撑。     

低盐胁迫下坛紫菜叶状体的转录组分析

为探索坛紫菜叶状体响应低盐胁迫的分子生理学机制,本研究采用第二代高通量测序技术,分析比较在正常盐度(26,对照组)和低盐度(3,胁迫组)下培养不同时间(3、6和9 h)的坛紫菜叶状体转录组数据。结果显示,测序数据经de novo组装,获得了33 872条unigenes,平均长度为612 bp;与对照组相比,3个胁迫组分别产生了1108、1638和1881条差异性表达基因。GO功能富集分析显示,一些可能与低盐胁迫相关的重要生物过程得到了显著富集,如单分子有机物代谢过程,单糖的合成代谢和分解过程,糖质新生,有机物分解代谢过程等。KEGG代谢通路富集分析发现,低盐胁迫不同时间富集到的代谢通路存在很大差异,低盐胁迫3 h响应的差异性表达基因富集到3个KEGG代谢通路,其中2个与氨基酸合成相关,1个与光合作用相关;而低盐胁迫6和9 h的差异性表达基因则大多富集到与糖类功能相关的代谢通路。荧光定量PCR(q RT-PCR)验证结果显示,所选取的8个差异性表达基因的表达趋势与高通量测序结果相一致。上述结果说明,坛紫菜叶状体在受到低盐胁迫时,应激反应具有时间特异性,早期主要通过合成或者降解蛋白质来抵御低盐环境,随着胁迫时间延长,细胞通过改变可溶性物质的含量、减慢能量代谢等途径以抵御低盐逆境。     

低盐养殖大黄鱼肌肉营养成分分析

为研究低盐度养殖条件对大黄鱼肌肉主要营养成分的影响,分别在盐度2、4、6、8、10、24(对照组)的水体中饲养大黄鱼幼鱼,6个月后,检测大黄鱼肌肉营养成分,并采用氨基酸评分(AAS)、化学评分(CS)和必需氨基酸指数(EAAI)来评定肌肉蛋白质的氨基酸营养价值。结果显示:大黄鱼肌肉中的水分含量与盐度呈负相关性,盐度2组(S_2)肌肉中水分含量最高,达(77.4±0.7)%,显著高于对照组(P0.05);各组间粗蛋白、粗脂肪和粗灰分的含量差异不显著(P0.05);盐度6组(S_6)氨基酸总量(TAA)、必需氨基酸含量(EAA)、呈味氨基酸含量(DAA)分别为(11.4±3.4、)%、(4.3±0.1)%和(4.5±1.5)%,均为最高(P0.05),且该组EAAI最大(73.88);盐度8组(S_8)多不饱和脂肪酸含量(∑PUFA)和必需脂肪酸含量(∑EFA)分别为(39.1±3.0)%和(26.8±1.6)%,显著大于其他组(P0.05);各低盐养殖组饱和脂肪酸总含量(∑SFA)和单不饱和脂肪酸总含量(∑MUFA)显著高于对照组(S_(24))(P0.05);各组二十碳五烯酸(EPA)+二十二碳六烯酸(DHA)的含量在(9.8±0.4)%～(13.0±0.2)%。试验表明,大黄鱼在盐度6～8的水体中养殖,肌肉营养较佳。结果将为大黄鱼低盐工厂化养殖和抗低盐新品种选育提供理论基础。     

慢性盐度胁迫下金钱鱼卵巢转录组分析

为了阐明盐度胁迫对金钱鱼(Scatophagus argus)代谢和生殖的影响,采用RNA-seq技术对低盐组(5)、对照组(25)和高盐组(35)处理40 d后的2龄性成熟金钱鱼卵巢进行转录组分析。结果发现,金钱鱼卵巢转录组测序获得raw reads共398681318,clean reads共396910398。从低盐胁迫相对对照组(5 vs 25)和高盐胁迫相对对照组(35 vs 25)中分别筛选到373个和874个差异表达基因(DEGs)。与对照组相比,低盐胁迫后氨基酸代谢相关基因(sds、bhmt)和脂肪酸代谢相关基因(pnpla2)显著下调;高盐胁迫后pgr、cyp17a1和ers1等生殖相关基因显著下调。KEGG通路富集分析发现,低盐胁迫组相对对照组显著富集与代谢相关的通路为半胱氨酸和甲硫氨酸代谢,缬氨酸、亮氨酸和异亮氨酸生物合成代谢,脂肪酸生物合成,脂肪酸代谢,皮质醇的合成和分泌,醛固酮的合成和分泌,脂肪细胞脂解的调节等;高盐胁迫组相对对照组中显著富集的与生殖相关的通路为雌激素信号传导通路。这些结果表明,氨基酸和脂肪酸的代谢调控可能在金钱鱼卵巢的低渗透压调节中起重要作用...     

低盐胁迫对黄姑鱼(Nibea albiflora)幼鱼鳃离子调节、呼吸代谢酶和皮质醇的影响

挑选平均体重为(5.0±1.4)g,平均全长为(8.3±0.8)cm的黄姑鱼(Nibea albiflora)幼鱼,进行低盐度胁迫试验,以23盐度组为对照组,设置9和16两个盐度胁迫组,在0、1、3、7 d进行取样。通过检测鳃Na~+/K~+-ATP酶(NKA)、Ca~(2+)-ATP酶、H~+-ATP酶、乳酸脱氢酶(LDH)、琥珀酸脱氢酶(SDH)活力以及血清皮质醇含量,研究低盐度对黄姑鱼离子调节和呼吸代谢的影响。结果显示,鳃NKA活力在盐度胁迫后出现显著增强(P0.05);Ca~(2+)-ATP酶活力略有上升后即出现下降的变化,而H~+-ATP酶活力呈现上升后恢复的变化。血清皮质醇含量在低盐度胁迫后呈显著的波动变化(P0.05),9和16盐度组的皮质醇水平交替上升。鳃LDH活力在低盐度胁迫后显著增强(P0.05),且9盐度组增强程度大于16盐度组。鳃SDH活力先减弱然后增强,9盐度组在7 d时增强十分显著(P0.05)。试验中,仅9盐度组出现实验鱼死亡两尾的情况,16及23盐度组黄姑鱼进食和活动均正常,9盐度组略差。研究表明,盐度降低可显著影响黄姑鱼幼鱼离子调节能力及呼吸代谢功能。盐度胁迫程度超过黄姑鱼适应能力与机体储备的承受范围会对机体造成伤害。     

低盐胁迫对黄姑鱼(Nibea albiflora)幼鱼鳃离子调节、呼吸代谢酶和皮质醇的影响

挑选平均体重为(5.0±1.4) g,平均全长为(8.3±0.8) cm的黄姑鱼(Nibea albiflora)幼鱼,进行低盐度胁迫试验,以23盐度组为对照组,设置9和16两个盐度胁迫组,在0、1、3、7 d进行取样。通过检测鳃Na+/K+-ATP酶(NKA)、Ca2+-ATP酶、H+-ATP酶、乳酸脱氢酶(LDH)、琥珀酸脱氢酶(SDH)活力以及血清皮质醇含量,研究低盐度对黄姑鱼离子调节和呼吸代谢的影响。结果显示,鳃NKA活力在盐度胁迫后出现显著增强(P<0.05);Ca2+-ATP酶活力略有上升后即出现下降的变化,而H+-ATP酶活力呈现上升后恢复的变化。血清皮质醇含量在低盐度胁迫后呈显著的波动变化(P<0.05),9和16盐度组的皮质醇水平交替上升。鳃LDH活力在低盐度胁迫后显著增强(P<0.05),且9盐度组增强程度大于16盐度组。鳃SDH活力先减弱然后增强,9盐度组在7 d时增强十分显著(P<0.05)。试验中,仅9盐度组出现实验鱼死亡两尾的情况,16及23盐度组黄姑鱼进食和活动均正常,9盐度组略差。研究表明,盐度降低可显著影响黄姑鱼幼鱼离子调节能力及呼吸代谢功能。盐度胁迫程度超过黄姑鱼适应能力与机体储备的承受范围会对机体造成伤害。     

Effects of fasting on the meat quality and antioxidant defenses of market-size farmed large yellow croaker (Pseudosciaena crocea)

The study was conducted to investigate fasting effect on flesh composition and antioxidant defenses of market-size large yellow croaker (Pseudosciaena crocea). Two hundred fish (main initial weight 380 g) were divided into two groups (control and fasted) and reared in 6 cages. After two weeks of adaptation, group I fasted for 28 days; group II was fed normally as a control. In 3, 7, 14, 21 and 28 days, 6 fish per group were processed for proximate flesh composition, liver antioxidant enzyme activities and malondialdehyden flesh content analyses. In fasted fish, the reduction of lipid content in muscle occurred after day 3, and, compared to controls, the content of protein decreased from day 21, the activities of liver antioxidative enzymes superoxide dismutase (SOD) and glutathione peroxidase (GPX) increased from day 3, and flesh malondialdehyde (MDA) levels increased from day 21. Muscle lipid reduction shows that the fasting technique is effective in end product improvement of large yellow croaker. However, considering flesh protein loss and the subsequent oxidative stress, the fasting technique should be used with precautions.     

An outbreak of visceral white nodules disease caused by Pseudomonas plecoglossicida at a water temperature of 12°C in cultured large yellow croaker (Larimichthys crocea) in China

Visceral white nodules disease (VWND) caused by Pseudomonas plecoglossicida is a common disease in cage-farmed large yellow croaker (Larimichthys crocea) in China. VWND usually occurred at water temperature of 16–19℃, resulting in high mortality in farmed large yellow croaker. Now, P. plecoglossicida as its pathogen has been considered nonpathogenic at 7–12℃. During February 2019, an infectious disease outbreak was observed in cage-farmed large yellow croaker at a water temperature of 12℃ in Ningde, China. This disease is characterized by white granulomatous lesions in internal organs of the diseased fish, which was similar with the symptoms of the VWND in large yellow croaker. Then, we isolated a bacterial strain named PQLYC4 from visceral lesions of the diseased fish. The experimental infection studies demonstrated that the strain PQLYC4 was the pathogen of the disease, which was further identified as P. plecoglossicida by the analysis of morphology, 16s rRNA gene homology and average nucleotide identity based on the whole genome sequence. Our results revealed that P. plecoglossicida strain PQLYC4 could cause the outbreak of the VWND at 12℃, a water temperature lower than that reported previously, thus providing new knowledges of prevalence and prevention of the VWND in large yellow croaker.     

Visceral granulomas in farmed large yellow croaker,Larimichthys crocea (Richardson), caused by a bacterial pathogen,Pseudomonas plecoglossicida

An enzootic disease characterized by granulomas in internal organs occurred in cage‐farmed large yellow croaker, Larimichthys crocea (Richardson), in April and November 2010, in Ningbo, Zhejiang Province. One bacterial strain, named XSDHY‐P, was isolated from the diseased fish and identified by biochemical characterization, fatty acid methyl ester (FAME) analysis and multilocus sequence analysis (MLSA). According to the results obtained from the biochemical tests, FAME analysis and phylogenetic analysis derived from 16S ribosomal RNA, gyrB, oprF, oprI, oprL and rpoD gene sequencing, the bacterial isolate, XSDHY‐P, was identified as Pseudomonas plecoglossicida. Moreover, lethal dose, 50% trials were carried out to demonstrate the virulence of XSDHY‐P in large yellow croaker when administered at 2.13 × 10⁵ colony‐forming units per fish. Visceral granulomas were found in the experimentally infected fish as well as in the naturally infected fish, indicating that P. plecoglossicida is another bacterial pathogen that causes granulomatosis in L. crocea.     

大黄鱼过氧化氢酶基因的克隆及其对鳗弧菌感染的响应

为探究过氧化氢酶(catalase,CAT)对大黄鱼机体的保护作用,实验基于大黄鱼基因组序列数据库克隆获得CAT基因1584 bp的完整开放阅读框(GenBank登陆号:KKF-14425.1),该序列编码527个氨基酸残基,包含与其他动物高度保守的酶活性中心序列FDRERIPERVVHAKGA、亚铁血红素结合信号序列RLFSYPDTH、3个催化位点残基His75、Asn148和Tyr358,以及12个NADPH结合位点等,理论分子量为59.98 ku,等电点为8.37。多序列比对显示,大黄鱼CAT氨基酸序列与其他鱼类具有较高的一致性,与同属于石首鱼科的军曹鱼和条石鲷同源性高达94%,在进化树中也聚类于同一进化分支,说明该序列为CAT家族成员。实时荧光定量PCR检测显示,大黄鱼CAT基因在所检测的7种组织(肝脏、脾脏、脑、肾、肌肉、鳃、肠)中均有表达,但在肝脏中表达水平最高(为肌肉中的6.68倍)。鳗弧菌感染后,大黄鱼肝组织中CAT基因的表达随着时间的推移而变化明显,感染后12 h,达到最高(7.48倍),随后逐渐下降,到72 h已基本恢复到原始水平,注射PBS的对照组,CAT基因表达只略有上调,说明病原菌侵染可能引起鱼体产生大量活性氧自由基及H2O2,CAT基因则可以清除体内过量的活性氧,进而防止它们对细胞造成损伤。     

中国主要海洋底层鱼类生物学研究的回顾与展望

回顾了中国海洋主要底层经济鱼类资源生物学的研究历史和科研成果,并重点叙述了带鱼、小黄鱼、大黄鱼和绿鳍马面鲀等重要经济鱼类资源生物学研究概况.概述的内容包括地理种群及其产卵群体的鉴别和划分、生活史型的演变、种群和群体结构、种群数量变动、年龄组成和生长特性、摄食习性、性成熟周期、性腺成熟指数(GSI)、产卵群体生殖力、产卵场、索饵场、越冬场及洄游路线、资源量和渔获量、各种作业渔船的调整及其捕捞力量的限制措施等,并展望了中国海洋底层经济鱼类资源生物学研究的前景.     

Effects of antimicrobial peptide APSH‐07 on the growth performance,anti‐oxidation responses,stress resistance and intestine microbiota in large yellow croaker Larimichthys crocea

A 65‐day feeding trial was conducted to investigate the effects of dietary antimicrobial peptide APSH‐07 on the growth performance, anti‐oxidation responses, stress resistance and intestine microbiota of large yellow croaker Larimichthys crocea. Four isonitrogenous and isolipidic diets were formulated with 0, 30, 60 and 90 mg/kg of APSH‐07, respectively. The results showed that the fish fed with 90 mg/kg of dietary APSH‐07 had the significantly highest specific growth rate (p < .05). The activities of catalase, superoxide dismutase, the total anti‐oxidative capacity and lysozyme had the significantly highest values in liver of fish fed with 90 mg/kg of dietary APSH‐07 (p < .05). Serum glucose increased significantly in fish fed diets without APSH‐07 supplementation after temperature stress, while serum cortisol increased significantly in those after trawl stress (p < .05). Fish fed with 90 mg/kg of dietary APSH‐07 showed significantly higher operational taxonomic units, ACE estimator and phylogenetic diversity whole tree in intestine microbiota compared to fish fed without APSH‐07 (p < .05). In conclusion, under the present experimental condition, 90 mg/kg of antimicrobial peptide APSH‐07 supplementation in diet had the better growth performance, higher anti‐oxidation and stress resistance capacity, and a potentially more beneficial intestine microbial community of large yellow croaker.     

Effects of salinity on the physiological responses of the large yellow croaker Pseudosciaena crocea under indoor culture conditions

The physiological responses to different environmental salinities were assessed in juveniles of large yellow croaker Pseudosciaena crocea. This species shows a good capacity to adapt to comparatively low environmental salinities by evaluating some physiological responses, i.e. superoxide dismutase (SOD), acid phosphatase (ACP), alkaline phosphatase (AKP) and lysozyme (LZM) in the liver, spleen, gill and kidney respectively. Growth and survival at salinity 5‰, 10‰ and 25‰ were better than those at salinity 15‰ and 20‰. No significant differences in ACP, AKP, SOD and LZM in the liver were observed among different salinity treatments; SOD, AKP and LZM in the spleen among different treatments only showed significant differences at the beginning or the 2nd week; in the gill, no significant difference of AKP and LZM were observed during the whole experiment, SOD among different treatments showed significant difference at the beginning and the 8th week, and ACP only showed significant difference at the end of the experiment; in the kidney, significant differences in ACP, AKP and SOD among different salinities were merely observed at the end of experiment, and LZM showed significant difference among different treatments at the 2nd week. Overall, some slight stress responses were observed, but few significant differences were observed between low salinity and normal salinity, especially the growth and physiological functions were not influenced by low salinity, i.e. 5‰ and 10‰. We conclude that juvenile large yellow croaker is tolerant to low salinity and shows a potential for low salinity culture.