藤壶金星幼虫附着变态机制

藤壶属节肢动物门(Arthropoda)甲壳亚门(Crustacea)蔓足下纲(Cirripedia)围胸总目(Thoracica), 具备特殊的形态结构、生活史和种群生态特征,是最主要的海洋污损生物。其幼虫阶段通常经历6期无节幼体和1期不摄食的金星幼虫,从浮游的金星幼虫附着变态成固着的稚体是藤壶生活史中的一个关键环节。外界化学和生物因子中成体提取物、水溶性信息素、足迹、神经递质、激素、生物膜等均影响藤壶金星幼虫的附着变态;内在因子即金星幼虫的生理状态(能量储量和年龄)决定了其对外界因子的反应程度。概括了近年来藤壶附着变态生理机制和分子机制研究的进展,可为深入了解藤壶金星幼虫附着变态机制提供参考,也为开发新型、高效、环保的防污剂提供理论指导。     

Immunological studies on the settlement-inducing protein complex (SIPC) of the barnacle Balanus amphitrite and its possible involvement in larva-larva interactions.

Immunological investigation has revealed that a settlement-inducing protein complex (SIPC), which induces cypris settlement of the barnacle Balanus amphitrite, is synthesized during larval development and accumulates in the cypris larva. We previously purified the SIPC from adult B. amphitrite, which was active when bound to a substratum. The SIPC is a glycoprotein of high molecular mass, consisting of three major subunits of 76, 88 and 98 kDa with lentil lectin (LCA)-binding sugar chains. In the present study, we prepared antiserum against each LCA-binding subunit of SIPC, and performed immunoblot analyses. Immunoblotting of adult extracts showed that anti-76-kDa antibody reacted only with the 76-kDa protein, whereas anti-88-kDa and anti-98-kDa antibodies reacted with both the 88-kDa and the 98-kDa proteins. Immunoblotting of larval extracts indicated that reactivity of the 76-kDa protein to anti-76-kDa antiserum increased during larval development and cyprid extracts reacted strongly. Moreover, by using immunostaining we found that the SIPC was contained in ''footprints'' of cyprids, which have been shown to act as a settlement-inducing pheromone, and is secreted onto the antennular attachment discs. The results suggest that the SIPC (or SIPC-like proteins) is involved in both adult-larva and larva-larva interactions during settlement of the barnacle B. amphitrite.     

Smelly feet are not always a bad thing: the relationship between cyprid footprint protein and the barnacle settlement pheromone

A critical phase in the life cycle of sessile benthic marine invertebrates is locating a suitable substratum for settlement. For barnacles, it is the lecithotrophic cypris larva that makes this plankto-benthic transition. In exploring possible substrata for settlement, the cyprid leaves behind 'footprints' of a proteinaceous secretion that reportedly functions as a temporary adhesive, and also acts as a secondary cue in larval-larval interactions at settlement. Here, we show that two polyclonal antibodies raised against peptides localized at the N- and C-terminal regions of the adult settlement cue--the settlement-inducing protein complex (SIPC)--could both detect 'temporary adhesive' indicating that the SIPC is either a component of this secretion or that they are the same protein.     

Locating the barnacle settlement pheromone: spatial and ontogenetic expression of the settlement-inducing protein complex of Balanus amphitrite

Barnacles are prominent members of hard substratum benthic communities and their study has been important to advances in experimental ecology and contemporary ecological theory. Having recently characterized the cue to gregarious settlement of Balanus amphitrite, the settlement-inducing protein complex (SIPC), we use two polyclonal antibodies to examine the tissue distribution and ontogenetic expression of this glycoprotein. These antibodies were raised against two separate peptides located near the N- and C-termini of the SIPC and were used to detect the glycoprotein by western blotting and immunohistochemistry. By in situ hybridization we also show that the SIPC mRNA co-occurs with the expressed glycoprotein in the cuticles of both nauplius and cypris larval stages and the adult. In the larvae, the SIPC is expressed most strongly in the mouthparts and the hindgut of the stage 2 nauplius and in the thoracopods, antennules and bivalved carapace of the cyprid. In adult B. amphitrite, the expressed SIPC is present in protein extracts of the shell and in all organs that are lined by cuticular tissues. We suggest that the SIPC is produced by the epidermal cells that secrete the cuticle and discuss these observations with regard to earlier studies and the role of the SIPC as a contact pheromone.     

Discrimination at settlement in barnacles: Laboratory and field experiments on settlement behaviour in response to settlement‐inducing protein complexes

A major driving force to mechanistic studies of barnacle gregarious settlement is to contribute to an understanding of observed patterns of settlement in nature. In particular, how cyprids perceive adult conspecifics and how they discriminate between conspecific and allospecific barnacles are questions which have taxed researchers for nearly 50 years. The putative, active component of adult barnacles to which the cyprids respond has long been known to be a glycoprotein, referred to here as the settlement‐inducing protein complex (SIPC). The present study examines the discriminatory abilities of laboratory‐reared Balanus amphitrite and wild Semibalanus balanoides cyprids at settlement. Using a recently developed nitrocellulose membrane‐choice settlement assay, laboratory studies revealed that both species settled at a significantly higher rate on regions of membrane on which crude conspecific SIPC had been adsorbed compared to untreated regions. Moreover, when offered a choice between conspecific and allospecific SIPC, a trend to greater settlement on the conspecific regions was observed. The membrane assay was also evaluated in field trials using real‐time video footage of cyprid searching behaviour. Of 211 S. balanoides cyprids recorded during exploratory behaviour, only one settled. Exploratory behaviour was, however, clearly associated with regions of the membrane treated with either conspecific or allospecific (B. Amphitrite) SIPC compared to untreated regions. These results are generally in accord with previous reports on the discriminatory abilities of barnacle cyprids and suggest that the membrane assay may be usefully applied to field studies of settlement behaviour.     

Possible molecular mechanisms of species recognition by barnacle larvae inferred from multi-specific sequencing analysis of proteinaceous settlement-inducing pheromone

Gregarious settlement is essential for reproduction and survival of many barnacles. A glycoprotein, settlement-inducing protein complex (SIPC) has been recognized as a signal for settlement and it is expressed in both conspecific adults and larvae. Although the settlement-inducing activities of SIPC are species-specific, the molecular-based mechanism by which larvae distinguish conspecific SIPC from the SIPC of other species is still unknown. Here, the complete primary structure of the SIPC of Megabalanus coccopoma, as well as the partial structure of the SIPCs of Balanus improvisus, Megabalanus rosa, and Elminius modestus are reported. These SIPCs contain highly variable regions that possibly modulate the affinity for the receptor, resulting in the species specificity of SIPC. In addition, the distribution patterns of potential N-glycosylation sites were seen to be different among the various species. Differences in such post-translational modifications may contribute to the species specificity of SIPC.     

Possible molecular mechanisms of species recognition by barnacle larvae inferred from multi-specific sequencing analysis of proteinaceous settlement-inducing pheromone

Gregarious settlement is essential for reproduction and survival of many barnacles. A glycoprotein, settlement-inducing protein complex (SIPC) has been recognized as a signal for settlement and it is expressed in both conspecific adults and larvae. Although the settlement-inducing activities of SIPC are species-specific, the molecular-based mechanism by which larvae distinguish conspecific SIPC from the SIPC of other species is still unknown. Here, the complete primary structure of the SIPC of Megabalanus coccopoma, as well as the partial structure of the SIPCs of Balanus improvisus, Megabalanus rosa, and Elminius modestus are reported. These SIPCs contain highly variable regions that possibly modulate the affinity for the receptor, resulting in the species specificity of SIPC. In addition, the distribution patterns of potential N-glycosylation sites were seen to be different among the various species. Differences in such post-translational modifications may contribute to the species specificity of SIPC.     

Species specificity of barnacle settlement-inducing proteins

We previously isolated a larval settlement-inducing protein complex (SIPC) from adult extracts of the barnacle, Balanus amphitrite using a nitrocellulose membrane settlement assay. In the present study, we found that the extracts of other adult barnacles, Megabalanus rosa and Balanus eburneus, also induced the settlement of B. amphitrite cyprids although the inductive activity was slightly lower than that of conspecific extracts. Furthermore, we examined reactivity to anti-SIPC antibody in adult extracts from six species of Japanese barnacles other than B. amphitrite, brine shrimp and eight marine sessile organisms besides barnacles. The results showed that all barnacles examined contained SIPC-like proteins with slightly different molecular weight, while the other animals did not react to the antibody by immunoblot analysis. These findings suggest that species specificity in settlement-inducing proteins of barnacles is not so strict, but these proteins are characteristic to barnacle species.     

Pheromonal communication involved in courtship behavior in Diptera

Sex pheromones are known for many dipteran species and play an important role in courtship behavior, together with visual, tactile, acoustic and other factors. Pheromones for a number of dipterans have been recently identified. This survey covers a number of species in all the families that have been studied. The review discusses diverse courtship behaviors in Diptera, with a special focus on the sex pheromones involved. In the Nematocera suborder, pheromones are volatile components, which act at a distance. They are derived from short-chain alkanes with acetoxy groups (Cecidomyidae) or terpenes (Psychodidae). In the Cyclorrhapha, pheromones may be volatile, derived from alkanes (Tephritidae) or terpenes (Agromyzidae), or non-volatile, unsaturated or methyl-branched hydrocarbons, which act by contact (the other subgenera). The behavioral roles and regulation of these pheromones are described, and their importance in species recognition is discussed.     

Can the pheromones of predators modulate responses to herbivore‐induced plant volatiles?

Biological control of greenhouse pests has been successfully developed and applied. In greenhouse crops, several entomophagous species (predators and parasitoids) are used simultaneously in the crop cycle. One important aspect of these crops, which represent modified ecosystems, is the interactions among complexes of species, including plants, phytophagous insects, and predators. The chemical relationships (semiochemicals: pheromones and kairomones) among these species likely play an important role in greenhouse crops; however, few studies have focused on these relationships. The aim of this study was to analyse the importance of semiochemicals. Three groups of laboratory trials were conducted with two predatory species: Nabis pseudoferus and Nesidiocoris tenuis (Hemiptera: Nabidae and Miridae, respectively). The results of the first group of trials indicated that the adult females of both species were more attracted to herbivore‐induced plant volatiles (HIPVs) than they were to the control plants or plants with artificial damage. Based on the second group of trials, pheromones triggered an attraction in adult females of both species for conspecifics. Finally, based on the interactions of the adult females of the same species, pheromones changed or modulated the predatory responses to HIPVs. The implications of these results for the biological control of pest species in greenhouses are further discussed.     

The presence of GABA in gastropod mucus and its role in inducing larval settlement

Chemical substances that induce larval settlement have been the focus of many gastropod studies due to the importance of wild stock recruitment and production within aquaculture facilities. Gamma-aminobutyric acid (GABA), GABA analogs, and GABA-mimetics associated with certain crustose coralline algae (CCA), are known to induce larval settlement in commercial abalone (Haliotis) species, and other gastropods. Furthermore, mucus secreted from these gastropods has been shown to induce larval settlement, but the stimulatory components of mucus have not been thoroughly investigated. We now present data confirming that GABA is the settlement-inducing effector molecule contained within abalone mucus. To do this, we initially generated anti-GABA for use in immunoenzyme and immunofluorescent microscopy. Using these techniques GABA was identified in the nerves and epithelial cells of the foot, including mucus. Dried mucus samples subject to HPLC analysis revealed a mean concentration of 0.68 mM GABA after sample rehydration. The presence of GABA in these samples was confirmed by time-of-flight mass spectroscopy (TOF-MS). In addition, GABA was detected in the mucus of several abalone species and other gastropods by immunocytochemistry. Subsequent bioassays using both dry and fresh mucus strongly promoted induction of larval settlement.     

Purification and partial amino acid sequence analysis of the larval settlement-inducing pheromone from adult extracts of the barnacle,Balanus amphitrite (=Amphibalanus amphitrite)

A previously undescribed larval settlement-inducing protein was purified from adult extracts of the barnacle, Balanus amphitrite (=Amphibalanus amphitrite). Results of SDS-PAGE indicated that the relative molecular mass of the protein in reduced and denatured form is 31,600 ± 500 kDa, and that it is distinct from the Settlement Inducing Protein Complex (SIPC) which has previously been determined as a larval settlement-inducing pheromone. The N-terminal 33-residue sequence of the intact protein showed no similarity with previously reported proteins in the EMBL/Genbank/DDBJ databases. The purified protein at a concentration of 10 μg ml^?1 induced approximately four times more larval settlement than the control (filtered natural seawater). In addition, results of the assay using both 24-well polystyrene plates and agarose gels indicated that this protein is probably released into seawater and attracts cypris larvae. These results suggest that the purified protein is a waterborne type pheromone which induces settlement of larvae of B. amphitrite.     

Gregarious settlement in cypris larvae:the effects of cyprid age and assay duration

Gregarious behaviour of marine larvae is perhaps most clearly associated with finding a suitable habitat in a changeable or restricted environment, or with finding other conspecifics with which to mate. Prior work has shown that in settlement assays using cypris larvae of the barnacle Balanus amphitrite, gregarious interactions significantly affected the interpretation of experiments testing the activity of organic settlement promoters and inhibitors. Other studies have also shown effects of cyprid age and pheromone concentration on settlement behaviour. However, the effects of interactions between gregariousness and these two factors are not known. The aim of this study was to test the hypotheses that i) as cyprids age the effects of gregariousness become less apparent, and ii) as the duration of the experiment increases gregarious effects become more apparent, using cypris larvae of B. amphitrite and Balanus improvisus. Three age classes of cyprids were used at six densities in a fully factorial design. For B. improvisus cyprids significant gregarious effects occurred between 3 or more larvae, and although larval age and experiment duration had significant main effects, there were no interactions between these important factors and gregariousness. For B. amphitrite cyprids significant gregarious effects also occurred with 3 larvae per well, though this effect was strongly dependent upon experiment duration. B. amphitrite cyprid sensitivity to conspecific cues does not change with age, although increasing experiment duration and age interact to increase settlement. Differences between species may be due to different thresholds to conspecific larval cues, or B. improvisus cyprids release much more larval temporary adhesive during exploration.     

The role of pheromones in freshwater fishes

The study of fish pheromones is particularly relevant because of the conserved nature of chemoreception in vertebrates. However, most fish pheromone systems remain unstudied. All the major known pheromones of freshwater fish and their associated behaviours were reviewed. Importantly, those studies that have demonstrated the connection between behaviour and pheromones in freshwater fishes have resulted in a wide range of applications in management. For example, pheromones released by the sea lamprey Petromyzon marinus have a practical function in pheromone traps, showing how chemical communication can be used in the management of invasive species. Future research on fish pheromones should include olfactory systems in a wider range of species testing the possibility that a few distinct models could be applied to the all fishes. Progress in research on fish pheromones should include a closer collaboration with other research fields such as evolutionary biology to allow a better understanding of fish pheromones systems divergence and mate selection where correlation between phenotypic dominance and pheromone production is still largely ignored. Finally, the example of pheromone interaction between an invasive species topmouth gudgeon Pseudorasbora parva and a native endangered species sunbleak Leucaspius delineatus is provided to illustrate the concept of pheromone pollution that assists its establishment in a novel ecosystem.     

Physiology,endocrinology and chemical communication in aggressive behaviour of fishes

Fishes show remarkably diverse aggressive behaviour. Aggression is expressed to secure resources; adjusting aggression levels according to context is key to avoid negative consequences for fitness and survival. Nonetheless, despite its importance, the physiological basis of aggression in fishes is still poorly understood. Several reports suggest hormonal modulation of aggression, particularly by androgens, but contradictory studies have been published. Studies exploring the role of chemical communication in aggressive behaviour are also scant, and the pheromones involved remain to be unequivocally characterized. This is surprising as chemical communication is the most ancient form of information exchange and plays a variety of other roles in fishes. Furthermore, the study of chemical communication and aggression is relevant at the evolutionary, ecological and economic levels. A few pioneering studies support the hypothesis that aggressive behaviour, at least in some teleosts, is modulated by “dominance pheromones” that reflect the social status of the sender, but there is little information on the identity of the compounds involved. This review aims to provide a global view of aggressive behaviour in fishes and its underlying physiological mechanisms including the involvement of chemical communication, and discusses the potential use of dominance pheromones to improve fish welfare. Methodological considerations and future research directions are also outlined.     

白蚁踪迹信息素研究进展

近年来, 固相微萃取技术的使用显著促进了白蚁踪迹信息素研究的开展。目前, 已有77种白蚁的踪迹信息素得到研究, 常见化学成分为十二碳单烯醇、 十二碳二烯醇和十二碳三烯醇, 其次为新松柏烯。已经鉴定的踪迹信息素主要为单组分或双组分系统。白蚁踪迹信息素由腹板腺分泌, 除澳白蚁科的达氏澳白蚁Mastotermes darwiniensis具有3个腹板腺外, 现存的白蚁均具有1个腹板腺, 位于第4或第5腹节。所有腹板腺都具有类型Ⅰ和Ⅱ两类细胞, 原白蚁亚科（Termopsinae）、 齿白蚁科（Serritermitidae）、 鼻白蚁科（Rhinotermitidae）种类的腹板腺还具有类型Ⅲ细胞。踪迹信息素的生物合成还缺乏研究, 推测有甲羟戊酸、 脂肪醇和饱和表皮烃3种途径。白蚁踪迹信息素的简约性十分显著, 不同地理分布、 生物生态习性以及一些系统距离较远的种类具有相同和密切相关的踪迹信息素。对于许多种类, 相同的信息化合物具有踪迹信息素和配对性信息素双重功能。白蚁踪迹信息素种特异性和简约性的适应意义和进化机制需要进一步研究。     

Multiple web-borne pheromones in a spider Frontinella pyramitela (Araneae: Linyphiidae)

The bowl-and-doily spider, Frontinella pyramitela, is a common inhabitant of low vegetation throughout most of temperate North America. All instars build concave-upward, bowl-shaped, nonviscid webs supported above and below by meshworks of silk. Previous studies of this species have revealed that chemical(s) on the silk of adult females elicit both courtship behaviour and positive geotaxis from adult males that contact the silk. This study demonstrates (1) that two different contact pheromones are responsible for the dual action of the silk of adult females and (2) that the webs of different age and sex classes of bowl-and-doily spiders (including the webs of adult males) contain functionally different mixtures of the two pheromones.     

The role of pheromones and biostimulation in animal reproduction

It is now known that pheromonal communication plays an important role in mammalian behaviour and reproductive processes. Chemical communication with pheromones is one means of transmitting such information. In mammals, signalling and priming pheromones are thought to act either singly or in combination through olfaction, auditory, visual (sight) or tactile stimuli. Pheromones are air-borne chemical substances ("signals") released in the urine or feces of animals or secreted from cutaneous glands that are perceived by the olfactory system and that elicit both behavioural and endocrine responses in conspecifics.Extensive studies in insects, rodents, swine, sheep, goats and cattle have established the importance of pheromones in the strong influence exerted by the male on reproductive activity in the female. There is a pheromone produced by the queen honey bee, which has two functions: inhibition of queen rearing and suppression of oogenesis in workers and in addition attracts drones during nuptial flight. It has also been demonstrated that the urine of male mice, rats, feral species and other wild rodents contains a priming pheromone that is responsible for hastening puberty in the females. Pheromones in the wool, wax and urine of a ram are sufficient to stimulate ewes to ovulate, while the buck has a strong characteristic seasonal odor and a buck jar containing the odor of the buck can be used as an aid in the detection of oestrus in does. The mere presence of the boar at the time of insemination of the sow improves sperm transport and ovulation, while the presence of the vasectomised bull has been reported to hasten the onset of puberty in heifers and also early resumption of ovarian activity in cattle following parturition. The role of pheromones in bovine reproduction is not as clearly defined as in sheep, goats and swine.Pheromones and other allelomimetic cues can exert profound effects on reproductive activity via the hypothalamic system that generates pulses of gonadotropin-releasing hormone. Manipulations of these factors and other pathways linking environmental inputs to reproductive output can lead to developing the concept of "control systems technologies", aimed at controlling reproductive performance. The knowledge acquired on the effectiveness of biostimulation; the factor which conditions it and the biological mechanism which produces it in livestock species, allows its use as a breeding management tool. The understanding of the role of pheromones could be of potential economic importance in addressing some of the problems associated with livestock production in the tropics. The biostimulation technique offers a potentially useful and practical way to improve reproductive efficiency in livestock species in the tropics. The exact nature of the cues and the role of biostimulation in livestock species especially swine, sheep, goats and cattle in developing countries require more attention.