甜菜夜蛾核型多角体病毒在宿主种群中的垂直传播研究

以两种浓度的病毒液感染甜菜夜蛾4龄、5龄幼虫,以研究甜菜夜蛾核型多角体病毒从亲代到子代垂直传递的方式和效率。实验结果表明,该病毒可通过母体介导(卵内、卵表)和父体介导等3种方式传递给子代,造成子代感染。幼虫经口感染后母体介导传递实验表明,病毒垂直传播可引起20%～48%的子代幼虫病死,并能使子代成虫产卵量下降;母体介导传播实验中,以5%福尔马林进行卵表面消毒证实仍有18.8%的幼虫感染病毒死亡,表明了该病毒可经卵内传播,并同时存在着附卵传播;母体介导传递实验还表明病毒对宿主有弱化作用,亲代免于病死的蛹重量减轻,羽化所得雌成虫的产卵量下降,产卵期延长。以交互配对将感染与未感染的雌雄成虫彼此交配,表明雄成虫精子也能垂直传递病毒(父体介导),引起20％～40%子代幼虫感病死亡。成虫经口感染,对所产卵块表面消毒后,仍导致28.6%的子代幼虫死于病毒感染,也表明存在卵内传播。     

Ultrastructure of fat body cells of the velvetbean caterpillar infected with a nuclear polyhedrosis virus

During a study of the ultrastructure of a nuclear polyhedrosis virus of the velvetbean caterpillar, Anticarsia gemmatalis, various types of nuclear and cytoplasmic inclusions were found in fat body tissue heavily infected with the virus. Virogenic stroma was present in the nuclei of most infected cells. Bundles of fibrous material were observed in the nuclei and cytoplasm of cells containing polyhedral bodies. Other nuclear inclusions included concentric multilayered material, vacuoles, and membrane structures.     

自同安钮夜蛾幼虫分离的一种核型多角体病毒

对在林间导致同安钮夜蛾Anua indiscriminate Moore幼虫死亡的一种病原物进行了分离和鉴定,并对该病原物进行了室内毒力测定。结果表明：该病原物为核型多角体病毒, 定名为同安钮夜蛾核型多角体病毒(AiNPV)。其对同安钮夜蛾幼虫的致死浓度LC₅₀和LC₉₀分别为3.4×10⁴和3.8×10⁷ PIB/mL。     

Reproduction-flight relationship in the beet armyworm, Spodoptera exigua (Lepidoptera: Noctuidae)

To evaluate the relationship between reproduction and migratory flight, we examined ovaries, tethered flight, and egg laying in the beet armyworm, Spodoptera exigua (Hübner). Ovarian development in female S. exigua was classified into five stages in relation to the postemergence age of the moths. Oviposition peaked at ovarian stages III and IV. Mating accelerated ovarian development in this species, and thus the mated females developed faster than the virgin moths in ovaries. In virgin adults, flight capacity increased from the first day to the third day after emergence and remained at a high level until the moths were 7 d old. In contrast, the mated moths reached the maximum flight capacity in 2 d after emergence and maintained a high level until the age of 7 d. However, mating status could not influence the flight capacity of the beet armyworm, although there were general differences in flight capacity between virgin and mated adults at the same age. Moreover, significant correlations between the number of eggs laid and the flight capacity were not performed by the mated moths. These results showed that mating status and ovarian development did not seem to influence flight capacity in the beet armyworm, which was not in agreement with the definitions of the oogenesis-flight syndrome observed in many other migratory insects.     

Concentrating Autographa californica nuclear polyhedrosis virus and recombinant alkaline phosphatase from insect cells using a temperature-sensitive hydrogel

Recombinant alkaline phosphatase expressed in insect cells was concentrated by a factor of one and half times at a separation efficiency of 54.2% using hydrogel ultrafiltration. Enzyme concentration was confirmed by SDS-PAGE as well as by spectrophotometric measurement. Wild and recombinant Autographa californica nuclear polyhedrosis viruses (AcNPV) were concentrated 1.4 and 1.6 times of the feed solution at 48.5 and 60.0% separation efficiency, respectively. Hydrogel ultrafiltration appears to be an attractive alternative for the concentration of AcNPV and recombinant proteins from insect cells. This revised version was published online in June 2006 with corrections to the Cover Date.     

甜菜夜蛾交配行为和能力

在(27±1)℃,光周期L14∶D10的条件下对甜菜夜蛾I>Spodoptera exigua的交配行为及能力进行了研究。结果表明：成虫在羽化当晚即可进行交配,交配率以羽化后头三个晚上的较高(>82%),但从第4天起则显著下降。成虫一天中的交配时间出现于23:30～05:30之间,交配高峰出现在01:30～02:30和03:00～04:00 之间, 其中以第1高峰的发生频率较高。成虫交配持续时间从22～191 min不等,但以30～60 min的为多(40.8%, n=97), 60～90 min的次之(19.4%),超过180 min的较少(10.2 %)。另外,交配持续时间与蛾龄紧密相关。蛾龄越大,交配持续的时间越长,且差异显著。雄蛾一生的交配能力由1～11次不等,但受性比的影响显著：在性比为1∶1的条件下,雄蛾平均交配次数仅为3.0 次,而在2♀∶1♂至5♀∶1♂时,则增加到5.1～6.0 次。雌蛾交配比例及次数受性比的影响也很大：没有交配的雌蛾比例从1∶1时的8.3%增加到5♀∶1♂时的32%,仅交配一次的比例从16.7%增加到38.7%,而交配≥5 次的比例则从 25%下降到0。最后,对这些结果在甜菜夜蛾防治中应用的可能性进行了讨论。     

Dissolution of Autographa californica nuclear polyhedrosis virus polyhedra by the digestive fluid of Trichoplusia ni (Lepidoptera:Noctuidae) larvae

The dissolution of polyhedra of Autographa californica nuclear polyhedrosis virus by digestive fluid collected from 5th stage Trichoplusia ni larvae was studied in vitro. Observations were made at timed intervals using phase contrast microscopy, and scanning and transmission electron microscopy. Dissolution occurred rapidly and in a detectable sequence. Under phase contrast, most polyhedra lost their refringence by 0.5 min. The polyhedra became rounded in appearance with small protuberances on the surface and Brownian movement was observed within. After 1 min, the envelope of most polyhedra had ruptured, releasing the enclosed virions. The protuberances were also observed under the scanning electron microscope after digestion for 0.5 min. Many shell fragments devoid of internal contents were seen after more lengthy digestion. Internal structural changes were revealed by electron microscopy. After 1 min of exposure, polyhedra were observed in all stages of dissolution. By 3 min, only virions, scattered about in heterogeneous material, could be distinguished.     

Transmission of nuclear polyhedrosis virus in laboratory population of the armyworm, Mythimna (Pseudaletia) separata

Armyworm, Mythimna (Pseudaletia) separata, a serious pest of 12 agricultural crops, was artificially infected with its nuclear polyhedrosis virus to study the modes of virus transmission among the laboratory population. Virus transmitted by transovum and transovarial modes passed through polyhedra-fed moths to their progeny.     

Feeding and dispersion of Spodoptera exigua larvae and its relevance for control with a nuclear polyhedrosis virus

On chrysanthemum crops the larvae of the beet armyworm Spodoptera exigua (Hübner) (Lep.: Noctuidae) moved upwards to the top of plants after hatching and predominantly fed on the upper foliage layers. On tomato, however, the larvae did not move upwards and mainly fed on the lower leaves. On chrysanthemums up to the fourth instar most feeding occurred at the underside of foliage while the upper-epidermis remained intact. Larvae dispersing from a single egg batch of 35 eggs damaged about 90 small and 50 tall chrysanthemum plants during their development. The successive larval stages contributed respectively 0.1, 0.4, 4, 20 and 75 percent to the total foliage consumption.The results indicated that the virus preferably should be applied to the lower leaves of tomatoes and chrysanthemums when young instars are present, but to the upper-middle and top leaves of chrysanthemums when the larvae are older than second instars.

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Résumé La consommation et la dispersion des chenilles de Spodoptera exigua sont examinées sur chrysanthèmes et tomates de serre. Les adults déposent les oeufs généralement au feuilles près du sol. Apres l'éclosion les chenilles graduellement se mouvent en haute dans les chrysanthèmes et consomment principalement les feuilles les plus hautes. Dans tomate, cependant, les chenilles s'alimentent principalement au feuilles près du sol.Pendant leur dévelopment les chenilles originaires d'une seule pond de 35 oeufs peuvent endommager environ go petits ou so hautes crysanthè mes. Jusqu'à la quatrième stade larvale les chenilles s'alimentent principalement avec le surface inférieure des feuilles, sans consommer l'épiderme supérieure.Les cinq stades larvales contribuent respectivement 0,1, 0,4, 4, 20 et 75% a la consommation totale des feuilles de chrysanthème.Les resultats des expériments sur la conduite larvale suggestent que les viruses de la polyédrose nucléaire doivent être appliqués préférablement sur la face inférieure des feuilles bas de chrysanthème et de tomate avant que les chenilles se développent a la troisième stade larvale. Quand les larves sur chrysanthèmes sont déja dans le troisième stade larvale, application du virus sur les feuilles plus haute probablement donnait des resultats optimal.

     

Resistance to a nuclear polyhedrosis virus in the light-brown apple moth Epiphyas postvittana (Lepidoptera: Tortricidae)

Measurements were made of the relative susceptibility to a nuclear polyhedrosis virus of three populations of light-brown apple moth Epiphyas postvittana: a resistant laboratory strain (CAN), a susceptible laboratory strain (BAR), and a field population. CAN was found to be 50 times more resistant than BAR and 160 times more resistant than the field line. Experiments on hybrid crosses of resistant and susceptible strains showed that resistance is genetically determined. This serves as a warning of the possible selection of virus-resistant strains of insect pests, where viral insecticides are being used in the field.     

Electron microscope observations of Autographa californica (Noctuidae) nuclear polyhedrosis virus replication in the midgut of the saltmarsh caterpillar, Estigmene acrea (Arctiidae)

The nuclear polyhedrosis virus from Autographa californica was studied with the electron microscope in the midgut of the salt marsh caterpillar, Estigmene acrea. The results of the present study were compared with a previous study in which the same inoculum was fed to Spodoptera exigua. In Estigmene acrea polyhedra were produced, but virions were not occluded. Nonoccluded virions were found throughout the midgut cytoplasm and budding into the hemocoel. Within the cytoplasm, the rough endoplasmic reticulum was observed to contain paracrystalline proteinaceous bodies. Fibrous bodies and annulate lamellae were also found in the cytoplasm of infected cells.     

New cell lines from larval fat bodies of Spodoptera exigua: characterization and susceptibility to baculoviruses (Lepidoptera: Noctuidae)

Two new cell lines, designated IOZCAS-Spex-II and IOZCAS-Spex-III, were initiated from the fat bodies of larvae of the beet armyworm, Spodoptera exigua (Lepidoptera: Noctuidae) in TNM-FH medium containing 10% fetal bovine serum. The spherical cells were predominant among the various cell types and measures approximately 15 microm in diameter. The cell lines were mainly composed of tetraploid cells with chromosome numbers ranging from 116 to 131 (n=31). The cell lines were confirmed to have originated from the S. exigua by DAF-PCR technique. They were susceptible to the multiple nucleocapsid nuclear polyhedrosis viruses from S. exigua.     

甜菜夜蛾SeGluCl剪接变异体的敲除及其对杀虫剂敏感性的影响

【目的】本研究旨在利用CRISPR/Cas9基因编辑技术,探索甜菜夜蛾Spodoptera exigua谷氨酸门控氯离子通道(glutamate-gated chloride channels, GluCls)基因不同转录变异体编码的SeGluCls对杀虫剂敏感性是否存在差异。【方法】采用RT-PCR和RACE技术获得甜菜夜蛾SeGluCl的全长cDNA序列,并对其进行生物信息学分析;利用CRISPR/Cas9基因编辑技术敲除SeGluCl两个剪接变异体SeGluCl 3a和SeGluCl 3b,并构建甜菜夜蛾敲除品系(3a-KO和3b-KO),生物测定甜菜夜蛾敏感品系WH-S(对照品系)及3a-KO和3b-KO品系的3龄幼虫对阿维菌素、甲维盐和氟虫腈3种杀虫剂敏感性的差异。【结果】获得了甜菜夜蛾SeGluCl全长cDNA序列,并发现其2种剪接变异体(SeGluCl 3a和SeGluCl 3b),SeGluCl 3a(GenBank登录号：OM304353)的开放阅读框(open reading frame, ORF)长1 362 bp,编码454个氨基酸,SeGluCl 3b(Gen...     

幼虫密度对甜菜夜蛾生长发育与繁殖的影响

为了研究甜菜夜蛾Spodoptera exigua(Hübner)幼虫的密度对其发育及繁殖的影响,本实验观察了5种幼虫密度下（1,5,10,20,30头/瓶）,幼虫发育和成虫繁殖情况。结果表明：幼虫和蛹历期、存活率和蛹重均差异显著。幼虫和蛹历期均以20头/瓶的最短,1头/瓶的最长,其余随幼虫密度增加而延长;幼虫至蛹存活率以10头/瓶的最高,其余随幼虫密度增加而降低;1头/瓶的蛹最重,显著高于其他密度的,其余随幼虫密度增加而下降。尽管密度间成虫羽化率和产卵前期均无显著差异,但成虫产卵量、寿命和畸形率差异显著。1头/瓶的产卵量最多,其次为10头/瓶的,其余随幼虫密度增加而减少,30头/瓶的产卵量显著少于其他密度的; 密度在1～20头/瓶范围内,雌蛾寿命均较短,显著短于30头/瓶的,而雄蛾寿命以5头/瓶的最短,显著短于其他密度的（10头/瓶除外）,10头/瓶的次之,其余密度间差异不显著;不同幼虫密度下羽化的成虫畸形率差异显著,10头/瓶的最低,其余随幼虫密度增加而升高; 生命表结果表明甜菜夜蛾在10头/瓶下世代存活率和种群增长指数均最高,幼虫密度过低或过高均不利于种群增长;世代存活率（S）和种群增长指数（I）与幼虫密度之间的关系均呈抛物线关系：S =－0.2087x²+2.5694x+211.52 (R²=0.88),I=－0.0552x²+0.9166x+54.168 (R²=0.95)。结果提示幼虫密度影响甜菜夜蛾种群动态的重要生态因子之一。     

An RNA virus in Autographa californica nuclear polyhedrosis virus preparations: Incidence and influence on baculovirus activity

A small RNA virus infectious to Trichoplusia ni larvae (TRV) was observed as a contaminant of several Autographa californica nuclear polyhedrosis virus preparations (AcMNPV). The extent of contamination in various AcMNPV preparations was studied by means of serial enrichment passages through T. ni larvae and enzyme-linked immunosorbent assay (ELISA). TRV could not be detected by ELISA in the original preparation of AcMNPV polyhedra prepared in 1968 even after five enrichment passages. Antibody inactivation offers a possible prophylactic method against TRV but temperature inactivation (55°C) does not. Although TRV reduced larval weight, it had little or no effect on bioassays of AcMNPV to T. ni and Heliothis virescens.