巨型艾美耳球虫偏菱形样蛋白EmRP免疫原性分析

在前期研究中,从巨型艾美耳球虫(Eimeria maxima)cDNA文库中筛选到了免疫保护性抗原偏菱形样蛋白EmRP,本研究为揭示该抗原的免疫保护机理,进一步检测了其免疫原性。以E.maxima卵囊cDNA为模板,用RT-PCR技术扩增EmRP,并构建真核表达质粒pVAX1-EmRP,将其经腿部肌肉注射2周龄的雏鸡,3周龄加强免疫。分别于首次免疫后和加强免疫后1周,采集血清,用ELISA方法检测血清特异性IgG水平,用荧光定量PCR方法(qPCR)检测细胞因子水平,用流式细胞术检测CD4^+T淋巴细胞和CD8^+T淋巴细胞的比例,分析EmRP诱导的免疫反应。序列分析表明,EmRP含有偏菱形样蛋白超家族成员保守区域,为非跨膜蛋白,分子量约为28.4 kD。真核表达质粒pVAX1-EmRP免疫鸡后,与对照组相比,鸡体IFN-γ、IL-2、IL-10和IL-17等细胞因子转录水平显著提升;CD8^+和CD4^+ T细胞比例显著提高,而血清特异性IgG水平与对照组差异不显著。结果表明,EmRP诱导的免疫保护作用主要是由其诱导的细胞免疫反应实现的,可以作为研制E.maxima新型疫苗的候选抗原。     

Pectin purified from pomelo (Citrus maxima) peel as a natural hatching agent for fish embryos

Pectin is a biodegradable polysaccharide, and it has been recently applied as a gene delivery, drug delivery, wound healing and tissue engineering agent. In this study, pectin was extracted from pomelo (Citrus maxima) peel and characterized. The extraction recovery of pectin form pomelo peel was 14.5%, and it had 72.56% degree of esterification, 1,245.56 equivalent weight, 7.82% methoxyl and 68.27% anhydrouronic acid contents. Use of pomelo pectin as a hatching enhancing agent for fish embryos and its effect on hatching enzyme 1 (ZHE1) was investigated. The pectin‐exposed zebrafish embryos (100 µg/ml) showed significantly (p < .05) higher hatching rate (96.6%) compared with untreated (control) embryos (66.6%) at 60 hpf. The mRNA expression of ZHE1 was also significantly (p < .05) elevated up to 55.6‐fold in pectin‐exposed embryos at 24 hpf. In situ hybridization results revealed remarkably strong expression of ZHE1 in pectin‐exposed embryos compared with the control group. In addition, considerably larger size of the hatching gland was observed in pectin‐exposed larvae than that of the unexposed larvae group. These results clearly indicate that pectin isolated from pomelo peel has an ability to enhance the hatching process of zebrafish embryos via upregulation of ZHE1.     

Thermal preference and tolerance of green abalone Haliotis fulgens (Philippi, 1845) and pink abalone Haliotis corrugata (Gray, 1828)

The thermoregulatory behaviour of green abalone Haliotis fulgens and pink abalone H. corrugata was investigated. Haliotis fulgens juveniles ranging in wet weight from 3.0 to 3.3 g and from 28.7 to 30.5 mm shell length and of H. corrugata 2.0 g and 25.7 mm in shell length were exposed to 19°C for 30 days in a flow‐through water system. Temperature preference was determined in a horizontal thermal gradient and was found to be 25.4°C for green abalone and 25.0°C for pink abalone. Displacement velocity was 4.3 cm h⁻¹ for H. fulgens and 12.8 cm h⁻¹ for H. corrugata. The optimum temperature for growth calculated for both abalone species was 24.6 and 24.5°C respectively. The critical thermal maxima (CTMax) of H. fulgens and H. corrugata were determined as a measure of thermal tolerance. Abalones were subjected to increasing water temperatures at a rate of 1°C on 30 min until they detached from the substrate. The CTMax at 50% were 33.6 and 32.0°C for green and pink abalone respectively. The results are discussed in relation to site selection and commercial rearing.     

大珠母贝人工苗水泥池中间培育技术的初步研究

为了解决大珠母贝幼贝海区养殖大规模死亡问题,开展了贝苗水池中间培育技术研究。本实验共用3批人工繁殖的2～3mm贝苗,经6个月的水泥池（335m^2）养殖,平均壳长分别增长至35．8mm、27．7mm、23．4mm,平均壳高分别增长至30．1mm、24．9mm、20．1mm,成活率分别为29．5％、17．4％、17．9％。该试验表明,大珠母贝种苗池塘中间培育生长速度快,成活率高,有望克服海区养殖大规模死亡问题。     

柚皮粗提物对松梢螟幼虫生物活性影响研究

通过测定柚皮粗提物对松梢螟幼虫的拒食活性、生长发育影响和毒杀活性,研究其对松梢螟幼虫的室内毒力作用.结果显示:100 mg/mL的柚皮粗提物生物毒性最强,处理24、48 h后的松梢螟幼虫的拒食率分别达到了99.28％ 、94.23％,拒食中浓度AFC50分别为7.61、9.28 mg/mL;对松梢螟四龄幼虫的生长抑制率在处理后24、48、96 h分别为96.32％ 、95.44％和93.19％,LC50分别为9.56、13.52、8.38 mg/mL;三龄幼虫、五龄幼虫的校正死亡率分别为90.00％和80.00％,与对照药剂噻虫啉的效果处于同一显著水平.柚皮粗提物对三龄幼虫毒杀作用效果优于五龄幼虫,LD50分别为18.46、25.99 mg/mL.说明柚皮粗提物对松梢螟幼虫生物活性有一定影响.     

大杯伞柄多糖与菌丝体粗多糖的免疫识别方法初探

采用热水(80℃)提取和75%乙醇沉淀、Sepharose CL-4B凝胶过滤及DEAE Sepharose CL-4B阴离子交换等方法分离纯化大杯伞(Clitocybe maxima)柄粗多糖,获得伞柄多糖纯品,制备多糖抗体;采用热水法提取大杯伞菌丝体粗多糖。用碳二亚胺法将纯化伞柄多糖与牛血清白蛋白进行共价化学偶联,并将偶联产物(拟糖蛋白)免疫兔子以制备抗血清,以酶联免疫法检测抗体滴度。依据该抗体与来自相同菌株的菌丝体粗多糖免疫反应差异对这两种多糖进行免疫识别,结果表明:经过亲和层析纯化后,第六次免疫抗血清对伞柄多糖的抗体滴度可达64K,而且对菌丝体粗多糖反应呈阴性,显示该抗体可识别这两种多糖之间的结构差异。     

Colouring ornamental fish (Cyprinus carpio and Carassius auratus) with microalgal biomass

Koi carp and goldfish value increases with intensity of skin colour, which is an important quality criterion. Fish cannot fully synthesize their own carotenoid colourings and these must therefore be included in their diet. Two trials were undertaken to investigate skin colour enhancement in ornamental species (i.e. three chromatic varieties of koi carp (Cyprinus carpio), namely Kawari (red), Showa (black and red) and Bekko (black and white) and goldfish (Carassius auratus)) by feeding a dietary carotenoid supplement of freshwater microalgal biomass [Chlorella vulgaris, Haematococcus pluvialis, and also the cyanobacterium Arthrospira maxima (Spirulina)], using a diet containing synthetic astaxanthin and a control diet with no colouring added for comparison. In the first trial, five homogeneous duplicate groups of 25 juvenile koi carp (C. carpio) (initial mean body weight 24.6 ± 0.7 g) were fed, for 10 weeks, one of the four diets containing 80 mg colouring/kg diet. In the second trial, this procedure was repeated for five homogeneous duplicate groups of 25 goldfish (C. auratus) (initial mean body weight of 0.9 ± 0.1 g). Initial and final samples of skin along the dorsal fin were withdrawn, from five fish per group, for subsequent analysis of total carotenoid content (spectrophotometric analysis), and red hue (colorimetric analysis, CIE (1976) L* a* b* colour system). Growth and feed efficiency were not significantly different between groups administered by the various dietary treatments. In both trials, dietary carotenoid supplementation increased total skin carotenoid content. The more efficient colouring for koi carps was found to be C. vulgaris biomass, providing both maximum total carotenoid deposition and red hue for the three chromatic koi carp varieties studied, and particularly for the kawari variety. For goldfish the best colouring obtained, as ascertained by total carotenoid content, was also achieved using C. vulgaris biomass, and red hue was maximum when using H. pluvialis biomass.     

Excavating sponges that are destructive to farmed pearl oysters in Western and Northern Australia

Species of the family Clionaidae (Porifera: Demospongiae) that excavate shell of the silver‐lip pearl oyster, Pinctada maxima, in north Western and Northern Australia are described. Two species belong to the genus Cliona and one species to the closely related genus Pione. Cliona orientalis has only recently been reported from Australia in living and dead coral on the Great Barrier Reef, and this is the first report of this species from north Western and Northern Australia. Cliona dissimilis is reported from Australia for the first time. Pione velans was first described from Shark Bay, Western Australia and this study extends its distribution from Albany, south Western Australia to Port Bremer in the Northern Territory. The most common species found was C. dissimilis. Pione velans and C. orientalis were also present, although the latter species was rare. Cliona dissimilis and P. velans were found to be sexually reproductive in some shells with specimens of C. dissimilis with oocytes in May and P. velans in September 1999. Egg development was synchronous, indicating that the sponges were oviparous and would broadcast gametes in a spawning event.     

Growth of cultured mussels (Perna canaliculus Gmelin 1791) at a deep-water chlorophyll maximum layer

We tested the hypothesis that yield of the mussel Perna canaliculus (Gmelin 1791) could be increased by placing culture ropes deep into the water column to take advantage of deep‐water chlorophyll a maxima. The study site, in Pelorus Sound, New Zealand, showed periods of thermal stratification of the water column, causing a high concentration of phytoplankton in deeper water, at the thermocline. Mussels were grown simultaneously at 5 and 17 m, for up to 96 days. Mussel growth and condition index data showed no significant differences between the two depth treatments, indicating that in this system there may not be substantial mussel productivity benefits from lowering mussel farms to the deep‐water chlorophyll maximum layer.