Orthogonal test design for optimization of the extraction of polysaccharides from Phascolosoma esulenta and evaluation of its immunity activity

Yield of polysaccharides from Phascolosoma esulenta obtained by phosphate buffer extraction through an orthogonal experiment (L₉(3)⁴) were investigated to get the best extraction conditions. The results showed that extraction temperature, ratio of phosphate buffer to raw material, extraction time, and ratio of trypsinase to raw material were the main four variables that influenced the yields of extracts. The highest yield was obtained when extraction temperature, ratio of phosphate buffer to raw material, extraction time and ratio of trypsinase to raw material were 40 °C, 2, 5.5 h and 1.6, respectively. The immunity-stimulating method showed that polysaccharides from P. esulenta could significantly raise liver, spleen and thymus index of mice and enhance Con A-stimulated mouse spleen cells proliferation. These results indicate that polysaccharides from P. esulenta had significantly higher immunity-stimulating activities.     

弓形革囊星虫组织结构和超微结构观察

弓形革囊星虫(Phascolosoma arcuatum)是近年兴起的水产养殖品种,目前已形成一定养殖规模,但是其生物学特性相关的研究只有零星报道。本文主要采用石蜡切片苏木精-伊红(H.E)染色和扫描电镜方法对弓形革囊星虫的整体形态和各主要器官的结构进行研究。结果表明,弓形革囊星虫主要由吻部和躯干部构成,躯干部有乳突分布;吻部由触手和项器构成,伸长可达躯干部的两倍,具有感光性。其消化系统由口、食道、肠、肛门四个部分构成,不同部分的肌肉组织存在明显差异;收吻肌一端与吻相连,另一端分成4支,分别连于背部两侧(背收吻肌)和腹部两侧(腹收吻肌);食道贴于收吻肌,后连接肠。肠分为肠下回环与肠上回环,肠上回环与肠下回环缠绕盘旋,肠上回环后接直肠。直肠具有盲囊,后连接肛门;两条肾管分布在腹侧。     

可口革囊星虫肾管纤毛的分布及结构特征

为了解可口革囊星虫(Phascolosoma esculenta)肾管纤毛的结构特点及其功能,采用显微及亚显微技术观察研究了可口革囊星虫肾管纤毛的分布位置及形态结构特征。结果表明,肾管外膜多纤毛细胞表面簇生纤毛、内部柱状上皮细胞与立方上皮细胞游离面着生分散的纤毛,肾口内面也着生纤毛。纤毛结构由纤毛干、过渡区、基体及其纤毛小根组成;纤毛干由"9+2"结构的轴丝外被纤毛膜构成;纤毛干与基体之间为过渡区,中央微管终止于此,外周双联微管通过过渡区和基体的外周轴丝相连;基体呈圆筒状,为"9+0"结构;纤毛小根分长根和短根,均为基体发出的由微细原纤维组成的圆锥形结构,具间隔70 nm的明显横纹。肾管纤毛可能在促进体腔液流动、提高肾管对体腔液的过滤作用以及引导成熟精卵进入肾管等方面发挥作用。     

An examination of the Phascolosotna subgenera Antillesoma, Rueppellisoma and Satonus (Sipuncula)

Three of the subgenera in the sipunculan genus Phascolosoma are reviewed, based on examination of type material. The presumed difference between the subgenera Antillesoma and Rueppellisoma (number of retractor muscles) is shown to be invalid and the taxa are therefore synonymous. The 15 species are either considered junior synonyms of P. antillarum (eight), transferred to other species or genera (six), or considered incertae sedis (one). Phascolosoma antillarum is redescribed. Of the eight species in the subgenus Satonus, only P. pectinatum remains valid: the others are considered either to belong to other taxa (four) or to be incertae sedis (three).     

可口革囊星虫体腔中卵细胞大小组成的周年变化

在浙江温岭沿海逐月采样,显微观察了可口革囊星虫体腔中卵细胞大小组成的周年变化。3—5月,卵径30~90μm的卵细胞占大部分。6月,卵径70~130μm的卵细胞占多数。7月,大部分卵细胞卵径达100μm以上,其中130~150μm的卵细胞约占1/3。8月,卵细胞卵径均在110μm以上,其中130~150μm的卵细胞约占2/3。9月,仍多数卵细胞卵径在110μm以上,130~150μm的卵细胞约占1/3。10月,以110~140μm的卵细胞占多,40μm以下的卵细胞较少,大卵细胞开始退化。11月至翌年2月,随着大型卵细胞的逐渐退化,小型及中型卵细胞逐渐增多,2月仍可见少量未退化的大型卵细胞。分析认为,可口革囊星虫在浙江温岭的繁殖期主要在7—9月。     

可口革囊星虫对Cd2+和Hg2+的富集及其影响

采用原子吸收、原子荧光等分析检测技术,探讨了Cd2+、Hg2+在可口革囊星虫中的富集规律及其对生长与主要营养成分的影响.结果表明:在试验设定的胁迫浓度内,可口革囊星虫体壁肌肉对Cd2+、Hg2+的富集均随胁迫时间的延长而增加,最终达到饱和浓度;环境中Cd2+、Hg2+浓度越高,富集速度越快,达到饱和的时间越短,饱和浓度也越高.可口革囊星虫体质量增长随着重金属胁迫浓度的升高而减慢,且联合胁迫的影响程度大于单一胁迫.体壁肌肉蛋白质含量随重金属胁迫浓度的增加而升高,Cd2+、Hg2+分别在0.05和0.02 mg·L-1胁迫浓度下达到最高,然后开始降低.联合胁迫也呈同样的规律,且影响程度更大.体壁肌肉脂肪含量随重金属胁迫浓度的增加而降低,联合胁迫下降低程度更大.     

Production of recombinant protein and polyclonal mouse antiserum for ferritin from Sipuncula Phascolosoma esculenta

The iron storage protein, ferritin, plays a key role in iron metabolism, but its regulation and functions in many invertebrate species are still largely unknown. In our previous work, an inducible ferritin cDNA from Phascolosoma esculenta with a full-length of 1017 bp has been cloned. In this follow-up study, the deducted ferritin protein sequence was predicted to be a polypeptide of 175 amino acids with a molecular mass of 20.1955 kDa and an isoelectric point of 5.08. The cDNA sequence of P. esculenta ferritin was constructed into pET system expression system and efficiently expressed in E. coli BL21 under IPTG induction. The recombinant ferritin was detected as a 24 kDa protein by SDS-PAGE. After purification directly from the gel, the recombinant ferritin was used to immunize mice and the anti-serum was prepared. The antibody displayed a strong immunological reactivity and specificity when used in Western-blot analysis. For the first time, our work provided a set of molecular tools essential for the further studies of ferritin protein functions in P. esculenta.     

可口革囊星虫肾管肌组织的结构特征

采用显微及亚显微技术观察了可1：7革囊星虫肾管肌组织的结构特征。肾管肌组织位于柱状上皮层下,由纵肌及环肌组成。肌细胞（肌纤维）呈长梭形,核位于细胞边缘并明显突向细胞外基质中,核周围有较多线粒体及少量内质网。肌纤维表面有许多囊状或指状突起的肌质囊,内含肌浆、光面内质网、线粒体及糖原颗粒。肌质囊之间的肌膜内面具膜相关电子致密斑。肌纤维内含粗、细两种肌丝,细肌丝围绕在粗肌丝周围,在肌丝之间分布有糖原颗粒、线粒体及胞质致密体。线粒体及糖原为肌纤维的代谢提供能量,肌组织的收缩对促进肾管的过滤排泄及繁殖时配子进入肾管可能起重要作用。     

可口革囊星虫胚胎与幼体的发育

解剖经降温8-10℃刺激30-40h后的可口革囊星虫(Phascolosomaesculenta)亲体取得精、卵,用人工授精法获得的受精卵进行胚胎和幼体发育的研究。结果表明:可口革囊星虫的卵为圆形或椭圆形,卵的大小为120μm×142μm-125μm×148μm。精子由头部、中段和尾部组成,全长约为32-40μm。在海水盐度23.8ppt、pH8.1、温度15℃的条件下,精、卵不能受精;在海水盐度23.8ppt、pH8.1、温度20℃的条件下,精卵能受精,但胚胎发育到原肠胚期停止;在海水盐度23.8ppt、pH8.1条件下,25℃、30℃和35℃的不同水温,精卵均能受精,受精卵发育到初期海球幼虫分别需要72h0min、47h08min和42h53min。在海水盐度23.8-25.5ppt,pH8.1-8.2,温度28-30℃条件下,从初期海球幼虫发育到稚虫需要15-20d。     

可口革囊星虫消化道的形态及组织学结构

对可口革囊星虫(Phascolosoma esculenta)消化道各部分结构进行了形态学和组织学观察,表明可口革囊星虫消化道由翻吻、咽、食道、肠、直肠和肛门组成。翻吻收缩性很强;肠道极长,沿纺锤肌螺旋缠绕成肠索。消化道管壁由内向外分为粘膜层、粘膜下层、肌层和外膜。粘膜上皮主要由柱状细胞组成。除肠外,消化道上皮细胞均有发达的纤毛,肠上皮细胞主要具发达的微绒毛;粘膜下层及肌层的发达程度因消化道的部位不同而异。