冷藏大黄鱼SSO希瓦氏菌致腐能力差异机制初探

为探讨特定腐败菌(SSO)希瓦氏菌(Shewanella)致腐能力的差异机制,采用生化和16S rDNA鉴定冷藏大黄鱼货架期终点的产H2S菌,在灭菌鱼汁和无菌鱼块筛选4株致腐差异的希瓦氏菌,扩增氧化三甲胺(TMAO)还原酶基因及分析其表达量,并预测其蛋白质的理化性质。结果显示,22株产H2S菌均为希瓦氏菌属,其中 S.baltica占54.5%,为S. putrefaciens占40.9%,S. hafniensis占4.5%。希瓦氏菌在灭菌鱼汁中致腐能力存在显著差异,其中S. balticaXH2和XH8的缺点评分和TVB-N最高,S. putrefaciensXH14和XH17菌最低。接种无菌鱼块的4株希瓦氏菌中XH2的样品在72h出现腐败气味,48h细菌高于107cfu/g,产生较高的TMA、TVB-N、尸胺和腐胺和,XH8菌次之,XH14和XH17菌最慢。四株希瓦氏菌都扩增出2490bp的torA基因,且torA基因的表达量与致腐能力密切相关,S.balticaXH2最高。预测的TorA蛋白中S.balticaXH2的分子量和不稳定指数最大,理论等电点和总平均疏水性最小。可见,S. baltica XH2为冷藏大黄鱼的SSO,其强致腐能力与torA基因高表达量和TorA蛋白理化性质有关。研究为阐明希瓦氏菌致腐机制奠定了良好的基础。     

水蛭来源荧光假单胞菌的分离鉴定及系统发育树分析

为调查山东某水蛭养殖场病情暴发原因,从该场患病水蛭中分离得到优势菌株SZ-1,对其进行生理生化特性及分子生物学分析。形态学、生理生化分析显示,菌株SZ-1为革兰氏阴性菌,杆状,两端钝圆,有鞭毛;Biolog GenIII分析表明,该菌株生长代谢利用情况与假单胞菌属吻合,与荧光假单胞菌B特征相似。基于16S rRNA基因序列分析同源性,该菌株16S rRNA基因序列与荧光假单胞菌(Pseudomonas fluorescens)相聚类,聚合置信度可达99%。综合生物学及分子特征,判定SZ-1为荧光假单胞菌。     

一株河鲈源致病性虫草菌Cordyceps confragosa CHL02菌株的全基因组测序及比较基因组分析

致病性虫草菌Cordyceps confragosa CHL02菌株是从患病河鲈(Perca fluviavilis)鱼体分离鉴定的一株昆虫致病菌,其无性阶段蜡蚧轮枝菌(Lecanicillium lecanii)广泛用于农业中昆虫防治。本研究基于Illumina PE150测序平台进行CHL02菌株的全基因组测序,对测序数据进行组装和组分分析,进行基因预测与功能注释,预测次级代谢产物合成基因簇,并进行病原宿主互作以及比较基因组分析。测序结果显示,CHL02基因组大小为36.17 Mb,GC含量为53.09%;预测包含8093个编码基因、1618个转座因子(TEs)、4572个串联重复序列及114个tRNA;共注释7724个基因,其中,1985个基因获得KOG注释,GO聚类分析中,2687个基因参与代谢过程,预测到22个次级代谢产物合成基因簇,1162个基因参与病原宿主互作机制中。基因聚类分析和系统发育树均显示,CHL02菌株与参考菌株昆虫源粗糙虫草菌(C. confragosa) RCEF 1005具有较高的同源性。本研究首次报道了河鲈源致病性虫草菌C. confragosa CHL02菌株的全基因组序列并分析其基本特征,与参考菌株进行比较基因组分析,为后续深入开展该病菌侵染河鲈的作用机制等相关研究奠定理论基础。     

一株河鲈源致病性虫草菌Cordyceps confragosa CHL02菌株的全基因组测序及比较基因组分析

致病性虫草菌Cordyceps confragosa CHL02菌株是从患病河鲈(Perca fluviavilis)鱼体分离鉴定的一株昆虫致病菌,其无性阶段蜡蚧轮枝菌(Lecanicillium lecanii)广泛用于农业中昆虫防治。本研究基于Illumina PE150测序平台进行CHL02菌株的全基因组测序,对测序数据进行组装和组分分析,进行基因预测与功能注释,预测次级代谢产物合成基因簇,并进行病原宿主互作以及比较基因组分析。测序结果显示,CHL02基因组大小为36.17 Mb,GC含量为53.09%;预测包含8093个编码基因、1618个转座因子(TEs)、4572个串联重复序列及114个t RNA;共注释7724个基因,其中,1985个基因获得KOG注释,GO聚类分析中,2687个基因参与代谢过程,预测到22个次级代谢产物合成基因簇,1162个基因参与病原宿主互作机制中。基因聚类分析和系统发育树均显示,CHL02菌株与参考菌株昆虫源粗糙虫草菌(C. confragosa) RCEF 1005具有较高的同源性。本研究首次报道了河鲈源致病性虫草菌C. confragosa CHL02菌株的全基因组序列并分析其基本特征,与参考菌株进行比较基因组分析,为后续深入开展该病菌侵染河鲈的作用机制等相关研究奠定理论基础。     

大黄鱼源气单胞菌的分离鉴定及致腐表型

为鉴别大黄鱼腐败菌气单胞菌的致腐能力,采用生理生化和分子方法(16S r RNA)鉴定冷藏大黄鱼货架期终点不产H2S的菌株。分析菌株在体外培养条件下的生长、胞外酶活性、三甲胺(TMA)和生物胺形成的腐败表型,将菌株接种在灭菌鱼汁中测定其致腐能力。结果显示,分离株AE03和AE04在0~37°C及0~60的盐度下生长良好,在4和25°C条件下AE03菌株的生长速率显著高于AE04菌株。2株细菌不产H2S,能还原氧化三甲胺(TMAO),分解尿素和液化明胶,氧化酶、赖氨酸、鸟氨酸脱羧酶、精氨酸水解酶等都呈阳性,能利用葡萄糖。结合16S r RNA鉴定发现分离株AE03为杀鲑气单胞菌,AE04为软体动物气单胞菌。在体外培养中,2种气单胞菌均能产生蛋白酶、形成TMA和生物被膜,其中AE03菌株生成各指标的能力高于AE04菌株,同时AE03菌株还具有脂酶活性,产生生物胺,其中尸胺含量较高。气单胞菌AE03在灭菌鱼汁中产生的挥发性盐基氮(TVB-N)显著高于AE04。通过紫色杆菌CV026和根癌农杆菌A136 2种报告菌检测发现,2株气单胞菌是高丝氨酸内酯(AHLs)产生菌,其中AE03菌株产生的AHLs含量显著高于AE04菌株。研究表明,杀鲑气单胞菌AE03为冷藏大黄鱼的强致腐菌,且能分泌高含量的AHLs,研究为从群体感应角度阐明气单胞菌的致腐机理奠定了良好的基础。     

河蟹细菌病病原分离与鉴定

1999～2004年对安徽省巢湖市境内8个乡镇的1300 hm2池塘、稻田养殖的中华绒螯蟹细菌性疾病进行了研究。结果表明:中华绒螯蟹细菌性疾病主要出现“颤抖”、“黑鳃”、“肝腺糊化”、“腹水”和“烂肢或腐壳”等5种常见病症;从患病蟹体内分离、鉴定出嗜水气单胞菌、粪产碱菌、少动假单胞菌、反硝化产碱菌、产吲哚黄杆菌、产碱假单胞菌和苏云金芽孢杆菌等7株病原菌。     

鲤鱼竖鳞病病原的研究

从本院试验场患竖鳞病鲤鱼鳞囊、肝、肾中分离出病原菌3株,对其中2株进行分类鉴定,分离自病鱼鳞囊的菌株为鲶爱德华氏菌(Edzuardsiella ictaluri ),分离自病鱼肾脏的菌株为荧光假单胞菌(Pseudomonas fluorescens).药敏试验结果表明:卡那霉素、氧哌嗪青霉紊对三株菌均有较强抑制作用.     

罗非鱼冷藏过程细菌种群的变化

对养殖尼罗罗非鱼(Oreochromis niloticus)0～10℃贮藏过程中感官、化学、微生物品质和细菌种群消长研究表明,初始样品挥发性盐基氮为(8.08±0.29)mg/(100g),菌落总数(CFU/g)的对数值为4.79±0.60;细菌种群复杂,种类繁多,分离到570株菌,其中80.5%为革兰氏阴性菌,13.6%为革兰氏阳性菌,优势菌是假单胞菌(Pseudomonas spp.)(36.5%)、肠杆菌科细菌(Enterobacteriaceae)(14.2%)和气单胞菌(Aeromonas spp.)(15.3%),同时检出了一定比例的不动杆菌和其他细菌。冷藏过程中肠杆菌科细菌、气单胞菌等生长受到抑制,细菌菌相组成逐渐变得单一,适应低温环境下革兰氏阴性菌比例不断增加,0℃、5℃、10℃贮藏至货架期终点时,阴性菌的比例分别达到86.4%、88.9%、100.0%;假单胞菌增殖显著,比例分别达到80.7%、68.1%、60.0%,可确认其为0～10℃贮藏罗非鱼的优势腐败菌。假单胞菌的种类变化趋势显示,优势顺序分别为荧光假单胞菌(P.uorescens)、恶臭假单胞菌(P.putida)、铜绿假单胞菌(P.aeruginosa)、产碱假单胞菌(P.alcaligenes),0℃、5℃、10℃贮藏至货架期终点时,假单胞菌中荧光假单胞菌分别占52.1%、53.1%、59.3%,可确认其为最优势种群。     

健康养殖南美白对虾肠道细菌的抗菌活性

用双层琼脂扩散法检测从健康养殖南美白对虾肠道内分离出的111株菌对8株病原菌的抗菌活性。结果表明:111株菌对白假丝酵母菌,非01群霍乱弧菌,嗜水气单胞菌指示菌无抗菌活性,48.6%的菌株对余下的5株指示菌有抗菌活性,其中气单胞菌属和发光杆菌属对荧光假单胞菌与金黄色葡萄球菌的抑制性最强。各菌属中以气单胞菌属,发光杆菌属的抗菌谱最广,弧菌属、肠杆菌科、黄单胞菌属、芽孢杆菌属、小球菌属次之。土壤杆菌属、棒状杆菌属、黄杆菌属、产碱杆菌属、假单胞菌属和色杆菌属则没有抗菌活性。     

采用Illumina MiSeq测序技术比较日本鲭和大黄鱼冷藏期间的腐败特性

为比较日本鲭和大黄鱼肌肉中微生物和代谢功能的变化及其与鱼肉腐败特性之间的关系,本研究检测了2种鱼在冷藏过程中的理化指标和菌落总数的变化,利用Illumina Miseq测序技术分析细菌群落变化,并利用皮尔森相关性分析检验微生物与鱼肉腐败及组胺产生相关性,结合功能预测分析细菌群落组成与代谢功能之间的关系。结果显示,冷藏期间日本鲭和大黄鱼的pH、挥发性盐基氮、组胺、菌落总数等均呈上升趋势,且日本鲭上升较快;冷藏末期2种鱼TVB-N值和组胺含量分别达到76.34、59.98和59.92、3.11 mg/100 g;日本鲭肌肉中细菌丰富度和多样性先增加后减少,大黄鱼则整体呈下降趋势;2种鱼肌肉中的优势腐败菌均为希瓦氏菌属;日本鲭体内与TVB-N产生相关的菌共12种,其中10种与组胺产生具有显著相关性;大黄鱼体内与TVB-N产生相关的菌共7种,但未检测出与组胺产生具有相关性的细菌;冷藏过程中氨基酸代谢和碳水化合物代谢为最主要的代谢通路,日本鲭样品组氨酸、精氨酸、脯氨酸等氨基酸代谢相关基因和丁酸丁酯代谢、丙酸酯代谢及丙酮酸代谢丰度均显著高于同一时期的大黄鱼,本实验从微生物代谢水平解释了日本鲭比大黄鱼更易腐败的原因,为不同水产品腐败特性的研究提供新思路。     

Desalted Cod Spoilage Flora Inhibition by Citric Acid and Potassium Sorbate Combination

ABSTRACT

Desalted cod spoilage isolates were tested in culture broth for growth inhibition by citric acid and potassium sorbate. Preservative effect was also evaluated in refrigeratedcod juice and desalted cod spiked with Shewanella putrefaciens and Pseudomonas fluorescens orputida. Combination of 0.15% citric acid and 0.1% potassium sorbate inhibited growth of approximately 60% of isolates, including the former two strains. Total inhibition was observed for S. putrefaciens and partial inhibition was noticed for P. fluorescens or putida, in both treated juice and desalted cod. Preservatives delayed growth of halotolerant bacteria in cod. Citric acid plus potassium sorbate addition lowered the pH to extend desalted cod shelf life.     

瓦氏黄颡鱼全基因组微卫星的分布特征及其定位的初步研究

文章探究了瓦氏黄颡鱼(Pelteobagrus vachelli)全基因组微卫星的分布特征及其规律,旨在为相关功能性微卫星分子标记的筛选提供依据.利用MISA(MIcroSAtellite identification tool)软件对其全基因组微卫星进行筛查和分析,并对外显子中含有微卫星的基因进行了GO(Gene O...     

Comparative genomic insights into the taxonomy of Edwardsiella tarda isolated from different hosts: Marine,freshwater and migratory fish

Edwardsiella tarda, a Gram‐negative member of the family Enterobacteriaceae, has been isolated from many animal species worldwide, especially fish species. Its broad host range indicates the diversity in taxonomy, which attracted the attention of many researchers. Here, we added genome of E. tarda strain isolated from freshwater fish to comparative genomics study for the first time. We sequenced and assembled the genome of E. tarda ASE201307 which was isolated from freshwater Asian swamp eel. ASE201307 genome contained a single circular chromosome of 3.68M with G+C 57.09% content. Comparative genomics including SNP calling, synteny block, Core/Pan genes analysis and phylogeny analysis was conducted among ASE201307 and other Edwardsiella strains isolated from different fish species. Results of SNP analysis and synteny block demonstrated the close relative of ASE201307, FL95.01 and DT which were all isolated from freshwater fish. In further analysis heat map of dispensable genes and phylogenetic tree, all E. tarda strains were divided into two groups. One was isolated from freshwater fish and the other was isolated from marine/migratory fish. Based on all studies above, we proposed the living environment of hosts as a new taxonomic character and divided E. tarda isolated from diseased fish into freshwater group and marine/migratory group.     

Assessment of the Antibacterial Effect of Chitosan Coated with Heracleum persicum Oil on Rainbow Trout Quality

Today, due to the rapid spoilage of fish, the use of natural preservatives is a priority over those of synthetic varieties. Also, the natural antimicrobial effects of essential oils can help to increase shelf life. There are very few studies concerning the use of essential oils in this regard. In the present study, the antibacterial effect of chitosan coated with Heracleum persicum oil was investigated on the quality of rainbow trout. The control and the coated fish samples were analyzed periodically by generalized estimating equation (GEE) for total viable count (TVC), psychrophilic bacteria, total volatile base nitrogen (TVB-N), polyunsaturated fatty acids (PUFA), texture, taste, odor, and overall acceptance parameters. The results showed that by increasing the amount of chitosan, TVC, psychrophilic bacteria, and TVB-N decreased while PUFA, texture, taste, odor, and overall acceptance parameters increased.     

鲤和草鱼IL17受体基因家族的全基因组识别、起源进化及表达分析

为了研究鲤科鱼类最具代表性的二个物种—鲤和草鱼IL17受体基因家族的起源进化,实验采用比较基因组学和生物信息学的方法,分别在鲤和草鱼基因组数据库进行序列比对和注释,然后对得到的基因进行结构域和系统发育学分析,最后在12个不同组织中进行基因表达分析研究。结果显示,在鲤和草鱼中分别注释得到9个和5个IL17受体基因家族成员。系统发育分析显示,该基因家族不存在鱼类特有的基因,在硬骨鱼类中具有一定的保守性。比较基因组学结果显示,与四足动物相比,大多数硬骨鱼类中IL17受体基因没有明显增多。鲤与草鱼等其他硬骨鱼类相比,除IL17RB以外,其余IL17受体基因家族成员均加倍。不同组织的表达分析结果显示全基因组复制后不同基因拷贝的功能发生了分化。研究表明,虽然硬骨鱼经历第三轮全基因组复制,但是由于复制发生时间久远,大多数基因已经发生改变或退化,进而在基因组中丢失。而鲤第四轮基因组复制时间发生在820万年前,复制发生时间较近,故复制后的基因基本得以保留。但是对于一些具有特殊功能的高度保守基因(例如IL17RB),也会发生在极短时间内出现丢失现象。鲤和草鱼健康组织的表达谱分析结果同样表明,鲤IL17受体基因的不同拷贝之间已经发生了快速进化及亚功能化,并且这种现象在鲤四倍体基因组中普遍存在。     

Dietary self-selection in sharpsnout seabream (Diplodus puntazzo) fed paired macronutrient feeds and challenged with protein dilution

Macronutrient selection and responses to dietary protein dilution were investigated in 95-g sharpsnout seabream, an omnivorous fish species of interest for Mediterranean aquaculture. Five groups of 18 sharpsnout seabream were distributed among circular 450-l tanks and allowed to self-feed from 3 feeders each of which distributed feeds composed of different pairs of macronutrients: PC (75% protein, 25% carbohydrate), PF (75% protein, 25% fat) or FC (50% fat, 50% digestible carbohydrate). Regardless of self-feeders position, the fish composed a diet containing 63% protein, 19% fat and 18% carbohydrate. When the protein-containing diets (PC and PF) were diluted 50% with cellulose, the fish increased their food intake to compensate for the energy dilution (170 kJ/kg BW/day vs. 168 kJ/kg BW/day after dilution), and dietary composition (63% protein, 16% fat and 21% carbohydrate) did not change significantly. These results demonstrate that sharpsnout seabream can select from incomplete diets to compose a balanced diet, and the fish are also able to compensate for a dietary dilution to regulate both energy intake and the relative proportions of macronutrients.