Sexual dimorphism in neuronal projections from the antennae of silk moths (Bombyx mori,Antheraea polyphemus) and the gypsy moth (Lymantria dispar)

Summary The antennal lobe of both sexes of the silk moth Bombyx mori contains 55–60 ventrally located antennal glomeruli; in addition, that of the male contains a dorsal macroglomerular complex (MGC). A group of identifiable glomeruli consisting of two lateral large glomeruli (LLG) and four medial small glomeruli (MSG) is present in both sexes, but the LLG are greatly enlarged in the female. A MGC is also present in the male gypsy moth Lymantria dispar and male giant silk moth Antheraea polyphemus. The MGC in all of these species is organized into 3–4 distinct levels of glomeruli. Antennal sensory fibers were stained by cobalt backfills in B. mori, A. polyphemus, and L. dispar. Most fibers stained from cut long hairs (sensilla trichodea) projected to MGC in males and LLG in both sexes of B. mori. The distribution of fibers in the MGC of B. mori was topographically biased in that a majority of fibers from anterior branches projected medially in MGC while most fibers from posterior branches projected laterally or anteriorly. Terminal arborizations of single fibers were each restricted to a single glomerular level of the MGC. Fibers projecting to the posterior antennal center were frequently stained in cut-hair and control preparations, apparently by uptake of cobalt through intact sensilla on flagellar branches.     

Structure of isolated sensilla and sensory neurons in vitro: Observations on developing silkmoth antennae

Summary By combined enzymatic and mechanical treatment, it was possible to dissociate the sensory epithelium of developing antennae of male Antheraea polyphemus and A. pernyi silkmoths from the stage of separation of the antennal branches up to the early stages of cuticle deposition. Large numbers of entire developing trichoid sensilla were isolated. These are characterized by a large trichogen cell with a long apical, hair-forming process and a large nucleus. A cluster of 2–3 sensory neurons, enclosed by the thecogen cell, is situated in the basal region. The dendrites run past the nucleus of the trichogen cell into the apical process from which they protrude laterally. The nuclei of the tormogen and a 4^th enveloping cell can be distinguished near the base of the prospective hair. After further dissociation, only the neuron clusters remain, still enclosed by their thecogen cell and often attached to the antennal branch nerve via their axons. It is finally possible to disrupt the thecogen cells and the axons, leaving the sensory neurons with inner dendritic segments and axon stumps. The majority of these neurons can be expected to be olfactory.     

Utilization of cuticular components ofAntheraea mylitta Drury larvae byPenicillium citrinum Thom.

The major cuticular components of Indian tasar silkworm,Antheraea mylitta Drury, were sequentially extracted and estimated to ascertain preferential utilization of these components for growth by the entomopathogenic fungusPenicillium citrinum Thom. Proteins which constituted 61.64% dry weight of cuticule were found to play a key role in the growth ofP. citrinum whereas lipids (7.15%) and chitin (30.02%) were least involved. Also, this study suggests absence of any mycocidal substance in the cuticle ofA. mylitta.     

密码子优化后的柞蚕溶菌酶在酵母中的表达及活性测定

【目的】提高柞蚕溶菌酶在毕赤酵母中的表达量,对柞蚕溶菌酶活性进行测定。【方法】根据毕赤酵母密码子偏爱性对柞蚕溶菌酶基因进行优化,并在目的基因3′端加上6个组氨酸标签,优化后的基因克隆至表达p PIC9K载体中,并通过电转化的方法导入毕赤酵母GS115中。利用不同浓度梯度的G418进行高拷贝转化子的筛选,经甲醇诱导实现分泌表达。通过硫酸铵盐析、镍柱亲和层析等工艺分离纯化柞蚕溶菌酶,利用琼脂扩散方法测定柞蚕溶菌酶对溶壁微球菌、金黄色葡萄球菌、枯草芽孢杆菌、大肠杆菌、苍白杆菌及过氧化醋杆菌的抑菌作用,利用比浊法测定酶活大小。【结果】密码子优化后的柞蚕溶菌酶在诱导温度25°C、甲醇量0.75%和p H 7.0条件下实现了最高表达,表达量达到了2.4 g/L,并且纯化后的柞蚕溶菌酶酶活达到23 970 U/mg。【结论】密码子优化后的柞蚕溶菌酶在毕赤酵母中实现了高效表达,纯化后的柞蚕溶菌酶对溶壁微球菌、金黄色葡萄球菌、枯草杆菌、大肠杆菌、苍白杆菌及过氧化醋杆菌均有抑菌作用。     

Lectin-Binding Sites Involved in Paramecium primaurelia Mating Pair Formation

ABSTRACT. Mating pair formation in Paramecium primaurelia was shown to be inhibited by incubating mating-competent mating type (mt) I and mt U cells with Limulus polyphemus agglutinin (LPA) or wheat germ agglutinin (WGA). Preincubation of LPA and WGA with their specific binding-monosaccharides, N-acetylneuraminic acid (NeuAc) and N-acetylglucosamine (GlcNAc), respectively, prevented the lectin effect on pair formation. Addition either of NeuAc or GlcNAc resulted in a reversal of cell pairing inhibition by LPA or WGA, respectively. Both NeuAc and GlcNAc monosaccharides inhibited pair formation when their concentration exceeded a threshold value. The pattern of the relative distribution of NeuAc and GlcNAc molecules on the cell surface was analyzed using fluorescence resonance energy transfer techniques combined with imaging systems. Mt I1 cells showed a high lectin-binding site density localized just on the surface region engaged in conjugation. The pattern of mt I cells, consisting of a quite homogeneous lectin-binding site density spread on the cell surface, was also common to nonmating-competent cells and to immature cells. These findings suggest that in P. primaurelia pair formation involves both NeuAc and GlcNAc residues and that the development of mating-competence is related to a modification in NeuAc and GlcNAc relative distribution on the cell surface of mt 11 cells.     

柞蚕感染微孢子虫后血淋巴免疫应答蛋白质的分离与鉴定

了解柞蚕Antheraea pernyi感染微孢子虫初期血淋巴内免疫系统及刺激应答相关蛋白质种类, 本研究以柞蚕5龄雌幼虫的起蚕（结束4眠, 刚完成蜕皮的幼虫）添食柞蚕微孢子虫Nosema pernyi为材料, 对感染后血淋巴利用SDS-PAGE进行分离后, 利用LC-MS/MS质谱技术和蛋白质组学分析对差异蛋白质条带进行鉴定。结果显示： 感染微孢子虫144 h后, 血淋巴中分子量约为44 kD （AP44）和28 kD （AP28）的蛋白质条带表达量增高。质谱分析AP28和AP44蛋白质条带样品, 共鉴定117个不重复蛋白质, 其中2个样品共有蛋白质12个, AP28独有蛋白质52个, AP44独有蛋白质53个。对质谱数据利用COG数据库进行搜寻鉴定, 显示AP28和AP44的鉴定蛋白质中涉及柞蚕免疫系统及刺激应答生物过程的蛋白质共有29个, 其中AP28中包括热激蛋白、 泛素样蛋白、 泛素结合酶E2、 保幼激素环氧水解酶、 微管结合蛋白、 溶菌酶、 ADP-核糖基化因子、 防御蛋白、 肽聚糖识别蛋白等15个, AP44中包括DRK、 酚氧化酶原、 类免疫球蛋白等10个; 二者共有热激蛋白hsp21.4、 酚氧化酶原、 抗菌肽等4个。本研究结果可以为今后研究柞蚕对微孢子虫的免疫应答及防御机制提供参考。     

Gopher Tortoise Demographic Responses to a Novel Disturbance Regime

The long-term viability of gopher tortoise (Gopherus polyphemus) populations is jeopardized by increased urbanization and habitat degradation owing to fire suppression. Because the species' remaining natural habitats in the southeastern United States exist within a mosaic of anthropogenic land uses, it is important to understand demographic responses to contrasting land uses and habitat management regimes. We examined differences in demographic parameters among fire-suppressed sandhill, restored sandhill, and former sandhill (i.e., ruderal) land use-land cover (LULC) types at Archbold Biological Station in south-central Florida, USA. Using Program MARK, we estimated population size, and sex-specific and LULC-specific survivorship based on 6 years of mark-recapture data. We also analyzed individual growth trajectories and clutch sizes to determine whether growth rates or reproductive output differed among LULC types. Tortoises in an open, ruderal field occurred at a higher density (7.79/ha) than in adjacent restored (1.43/ha) or fire-suppressed (0.40/ha) sandhill. Despite this higher density, both adult survivorship and body size were significantly higher in the ruderal field. Furthermore, the larger female body size in the ruderal field likely contributed to increased annual survivorship and slightly larger average clutch sizes. We did not detect offsetting negative demographic effects; in particular, we did not find significant biological or statistical differences in body condition, asymptotic body size, or growth rate among the 3 LULC types. Our results suggest that anthropogenic, grass-dominated land-cover types may be important components of the habitat mosaic currently available to this at-risk species. © 2019 The Wildlife Society.     

Species-specific pheromonal compounds induce distinct conformational changes of pheromone binding protein subtypes from Antheraea polyphemus

We have investigated the structural features of three pheromone binding protein (PBP) subtypes from Antheraea polyphemus and monitored possible changes induced upon interaction with the Antheraea pheromonal compounds 4E,9Z-14:Ac [(E4,Z9)-tetradecadienyl-1-acetate], 6E,11Z-16:Ac [(E6,Z11)-hexadecadienyl-1-acetate], and 6E,11Z-16:Al [(E6,Z11)-hexadecadienal]. Circular dichroism and second derivative UV-difference spectroscopy data demonstrate that the structure of subtype PBP1 significantly changes upon binding of 4E,9Z-14:Ac. The related 6E,11Z-16:Ac was less effective and 6E,11Z-16:Al showed only a small effect. In contrast, in subtype PBP2 pronounced structural changes were only induced by the 6E,11Z-16:Al, and the subtype PBP3 did not show any considerable changes in response to the pheromonal compounds. The UV-spectroscopic data suggest that histidine residues are likely to be involved in the ligand-induced structural changes of the proteins, and this notion was confirmed by site-directed mutagenesis experiments. These results demonstrate that appropriate ligands induce structural changes in PBPs and provide evidence for ligand specificity of these proteins. Electronic Publication     

Molecular Cloning of a Putative Cyclic Nucleotide-Gated Ion Channel cDNA from Limulus polyphemus

Abstract : Cyclic nucleotide-gated channels have been proposed to mediate the electrical response to light in the ventral photoreceptor cells of the horseshoe crab, Limulus polyphemus . However, a cyclic nucleotide-gated channel has not been identified from Limulus . We have cloned a putative full-length cyclic nucleotide-gated channel cDNA by screening cDNA libraries constructed from Limulus brain using a probe developed from Limulus ventral eye nerves. The putative full-length cDNA was derived from two overlapping partial cDNA clones. The open reading frame encodes 905 amino acids ; the sequence shows 44% identity to that of the α subunit of the bovine rod cyclic GMP-gated channel over the region containing the transmembrane domains and the cyclic nucleotide binding domain. This Limulus channel has a novel C-terminal region of ~200 amino acids, containing three putative Src homology domain 3 binding motifs and a putative coiled-coil domain. The possibility that this cloned channel is the same as that detected previously in excised patches from the photoreceptive membrane of Limulus ventral photoreceptors is discussed in terms of its sequence and its expression in the ventral eye nerves.     

Infection and development of Nosema bombycis (Microsporida: Protozoa) in a cell line of Antheraea eucalypti

Spores of Nosema bombycis derived from diseased insects were highly purified by Urografin density gradient centrifugation. Antheraea eucalypti cells were inoculated with the purified spores primed with 0.1 n KOH solution to start a continuous propagation of N. bombycis in cell culture. The first increase in the number of infected A. eucalypti cells was observed at 48 hr postinoculation, and it was caused by the secondary infective forms of N. bombycis. The secondary infective forms were produced during the course of sporoblast differentiation. The parasites in cell cultures divided synchronously until 36 hr postinoculation. Mature spores were observed initially 6 days postinoculation at 27°C. The infected cultures were subcultured extensively for more than 1 year with the addition of healthy A. eucalypti cells.