The estimation of population density of the green rice leafhopper,Nephotettix cinticeps Uhler,in spring field by the capture recapture method

The population parameters of green rice leafhopper, Nephotettix cincticeps in hibernated generation was estimated by the capture-recapture method on the gramineous weeds of resting paddy field from late in April to mid-May, 1962–1965. The difficulty of applying the capture-recapture method to this insect is caused from the low density and the low activity of the leafhopper, so that it is necessary to make the suitable plan of capture-recapture series and to construct the special method to detect the population parameters. The number of adults differed greatly among the years and among the plots in the same year. But, in general, the density was relatively high late in April, and decreased rapidly in May. The number of males was higher than that of females late in April, but decreased rapidly to become lower in May. The sampling efficiency with sweep-net depends upon the many factors, such as weather and floristic conditions; especially the atmospheric temperature and the force of wind are considered to affect greatly to the efficiency. In general, low temperature and strong wind are the cause of low sampling efficiency.     

Estimating the survival rate and mean longevity for adults in a field population of the green rice leafhopper,Nephotettix cincticeps Uhler,by the application ofHokyo andKiritani’s method

Application of the female dissection method proposed byHokyo andKiritani (1967) was attempted in both 1968 and 1969 to estimate the daily survival rate and the mean longevity for the adult population of the green rice leafhopper, Nephotettix cincticeps, in a paddy field. The estimated mean longevity for females was far shorter than the physiological longevity of this species, ranging from 4 to 7 days with some variation between different generations. This could explain the remarkable discontinuity among successive generations which proved to form an important feature of the pattern of seasonal population changes of this insect. Between the two years, the estimated survival rate (and hence the mean longevity also) was negatively correlated to the estimated population size of adults that emerged in either of the two successive generations. This apparent density dependence suggests the possibility that the adult survival, including the effect of dispersal, plays some critical role in bringing about the remarkable population stability from year to year shown by the field population of N. cincticeps.     

Life table studies on the paddy field population of the green rice leafhopper,Nephotettix cincticeps Uhler (Hemiptera: Cicadellidae), with special reference to the mechanism of population regulation

The population growth of the green rice leafhopper, Nephotettix cincticeps, in the paddy field was analyzed based on the life table data accumulated for six years. The paddy field population, which stems from the invading adults of the first generation (G-I), repeats two complete generations, and the hatchlings of the fourth generation (G-IV) enter diapause and overwinter as the fourth instar nymphs in fallow paddy fields. It was clarified that the density dependent reduction in the mean longevity and oviposition rate of adult females in G-II and III played a primary role in stabilizing the annual population densities. The annual average of the mean longevity of G-II females (3.9 days) was much shorter than that of G-III ones (7.7 days) and thus the density dependent reduction in the mean longevity induced a more prompt regulatory effect on the oviposition of G-II females compared with G-III ones. As the result, two equilibrium densities of eggs were obtained, e.g., ca 100 and 700 eggs per hill in G-III and IV, respectively. Density dependent decrease in the proportion of mature females in the adult population was especially conspicuous in G-II, and this was closely associated with the density dependent reduction in the mean longevity and fecundity. Thus, the density dependent dispersal (emigration) of the adult females by flight in G-II and III was the most convincing factor in the process of population regulation. The density dependent dispersal of the adult females is effective in avoiding the deleterious effects of nymphal crowding in a breeding habitat unit (a paddy field), and may result in a more even distribution of the population over a continuous habitat units in a locality than otherwise.     

The outbreak mechanisms of the green rice leafhopper,Nephotettix cincticeps Uhler,in northern Japan

Population Ecology - Intraspecific regulatory processes keep the population ofNephotettix cincticeps stable at a low density in southern Japan. In northern Japan, however, the yearly population...     

Descriptive models for the system of the natural spread of infection of rice dwarf virus (RDV) by the green rice leafhopper,Nephotettix cincticeps Uhler (Hemiptera: Deltocephalidae)

Seasonal changes in the densities of eggs and of 1st instar nymphs of N. cincticeps, a vector of the rice dwarf virus (RDV) and those in the proportion of virus-infected rice hills were examined by regular field censuses during the period of 1964–69 at two localities in Kochi, Nangoku and Ino. The proportion of virus-infected insects in the vector population was measured by the serological test. Mean fecundity of adult females of N. cincticeps in different generations was assessed by the method proposed byHokyo andKiritani (1968) in 1966 and 1967. Using observed variables as mentioned above, two mathematical models were built up for the system of the natural spread of infection of RDV. Submodels describing generation-to-generation changes in densities of eggs, 1st instar nymphs and leafhopper adults were involved in both Models 1 and 2. In Model 1, the proportion (P) of infected insects in the vector population was assumed to be determined directly by the amount (I_v) of infected rice plants in the field. In Model 2, instead of relating P directly to I_v,P was related to the acquisition rate (W) of RDV by the insects and to the effect of RDV (αr) on fecundity and survival of infected insects. Then, W was related to I_v. The estimates of P by these models fitted well to the observed values for 1st to 3rd generations but a great deviation was noted in 4th and 5th generations. For the percentage of infected rice hills, both models gave relatively good estimates in the early planted rice, though they gave overestimated values in the late planted rice.     

Molecular mapping of a novel gene, Grh5, conferring resistance to green rice leafhopper (Nephotettix cincticeps Uhler) in rice, Oryza sativa L.

The green rice leafhopper (GRH), Nephotettix cincticeps Uhler, is one of the most serious insect pests affecting cultivated rice (Oryza sativa L.) in temperate regions of East Asia. An accession of the wild rice species, Oryza rufipogon Griff. (W1962), was found to be highly resistant to GRH by an antibiosis test. To understand the genetic basis of the GRH resistance, a BC₁F₁ population derived from a cross between a susceptible Japonica variety, Taichung 65 (T65), and a highly resistant accession W1962 was analyzed by quantitative trait loci (QTL) mapping. A single major QTL for GRH resistance was detected on rice chromosome 8. A nearly isogenic population containing segments of the targeted QTL region derived from W1962 was then developed through advanced backcrossing with marker-assisted selection. Further molecular mapping using a BC₄F₂ population revealed that a new resistance gene, designated as Green rice leafhopper resistance 5 (Grh5), was located on the distal region of the long arm of chromosome 8 and tightly linked to the simple sequence repeat markers RM3754 and RM3761. A nearly isogenic line (NIL) carrying Grh5 was subsequently developed in the progeny of the mapping population. The resistance level of Grh5-NIL was compared with those of developed NILs for GRH resistance and was found to have the highest resistance. The DNA markers found to be closely linked to Grh5 would be useful for marker-assisted selection for the improvement of resistance to GRH in rice.     

Laccase-type phenoloxidase in salivary glands and watery saliva of the green rice leafhopper, Nephotettix cincticeps

The activity and composition of leafhopper saliva are important in interactions with the host rice plant, and it may play a physiological role in detoxifying toxic plant substances or ingesting sap. We have characterized diphenoloxidase in the salivary glands of Nephotettix cincticeps, its activity as a laccase, and its presence in the watery saliva with the objective of understanding its function in feeding on rice plants. Nonreducing SDS-PAGE of salivary gland homogenates with staining by the typical laccase substrate 2,2'-azino-bis (3-ethylbenzthiazoline-6-sulfonic acid) (ABTS), hydroquinone or syringaldazine revealed a band at a molecular mass of approximately 85 kDa at pH 5. A band also appeared at a molecular mass of approximately 200 kDa when the gels were treated with dopamine, L-3,4-dihydroxyphenylalanine (DOPA) or catechol at pH 7. The ABTS-oxidizing activity of the homogenates was drastically inhibited by N-hydroxyglycine, a specific inhibitor of laccase. However, the dopamine-oxidizing activity was not inhibited by N-hydroxyglycine, while it was inhibited by phenylthiourea (PTU). Thus, the salivary glands of N. cincticeps contain two types of phenoloxidases: a laccase (85 kDa) and a phenoloxidase (200 kDa). Laccase activity was detected in a holidic sucrose diet that was fed on for 16 h by two females, but only a trace of catechol oxidase activity was observed, suggesting that the laccase-type phenoloxidase was the predominant phenoloxidase secreted in watery saliva. The laccase exhibited an optimum pH of 4.75-5 in McIlvaine buffer and had a PI of 4.8. Enzyme activity was histochemically localized in V cells of the posterior lobe of the salivary glands. It remained at the same level throughout the adult stage from 2 days after eclosion. A possible function of N. cincticeps salivary laccase may be rapid oxidization of potentially toxic monolignols to nontoxic polymers during feeding on the rice plant. This is the first report proving that laccase occurs in the salivary glands of Hemiptera species and is secreted in the watery saliva.     

Absence of protein polymorphism attributable to insecticide-insensitivity of acetylcholinesterase in the green rice leafhopper, Nephotettix cincticeps

The cDNA sequence of acetylcholinesterase (AChE) from the green rice leafhopper, Nephotettix cincticeps, was amplified, based on conserved peptide sequences of AChEs. A 2.3 kb contiguous sequence, containing an ORF encoding an AChE precursor with 677 amino acid residues was obtained. The deduced protein sequence showed the most similarity to that of AChE in the Colorado potato beetle, having common features in the primary AChE structure. cDNA sequences of individual leafhoppers from an insecticide susceptible strain and the resistant strain Nakagawara, whose methylcarbamate-insensitive AChEs show 10(2) or more I(50) ratio for propoxur, were compared. No fixed inter-strain difference was identified in the protein sequence, though amino acid substitution polymorphism was found at one position in the susceptible strain. Insecticide-insensitivity of leafhopper AChE does not result from changes in the protein primary structure that is encoded by the AChE gene sequence isolated in this study.     

Analytical studies of light trap records in the Hokuriku district,II the green rice leafhopper,Nephotettix cincticeps

Light-trap records on the green rice leafhopper, Nephotettix cincticeps, were dealt with to study its population fluctuations in the Hokuriku district. Crude data were modified for distinguishing years of the low intensity of infestation by the insect from the rest of years. It is then clearly demonstrated that the low intensity of infestation were ordinarily preceded by heavy snowfall, although heavy snowfall could not be regarded as the only factor checking the vigorous multiplication of the populations.     

Molecular cloning of a novel calcium-binding protein in the secreted saliva of the green rice leafhopper Nephotettix cincticeps

Green rice leafhoppers (Nephotettix cincticeps) secrete watery and coagulable saliva in the feeding process. In our study, the watery salivary secretion was concentrated by ultrafiltration from “fed diet” and subjected to SDS-PAGE. The N-terminal amino acid sequence of the most predominant band at 84 kDa (designated NcSP84) was analyzed by Edman degradation. This sequence was completely consistent with the most abundant protein in the salivary gland extracts, which was separated by two-dimensional gel electrophoresis. Based on the N-terminal amino acid sequence, the complete cDNA of this protein was cloned by 5′- and 3′-RACE using degenerate primers. The deduced NcSP84 contained an open reading frame of 2061 bp encoding a putative 687 amino acids with a putative signal sequence composed of 19 amino acids. The nucleotide and amino acid sequences of NcSP84 did not share statistically significant homology with any sequences in public databases. Motif search predicted that this protein had EF-hands, the most common motif found in Ca²⁺ -binding proteins. As predicted, NcSP84 exhibited Ca²⁺-binding activity. The SDS-PAGE mobility of purified NcSP84 bound to Ca²⁺ tended to decline discretely, depending on the concentration of CaCl₂ with which it was mixed for 1 h before adding SDS buffer. In situ hybridization and immunohistochemistry showed that the NcSP84 gene and gene product were expressed and stored in type III cells, which are the largest lobes in the primary salivary glands. The NcSP84 protein was detected in the phloem sap of rice exposed to leafhoppers, verifying that the NcSP84 protein was injected into the sieve tubes. These results suggest that NcSP84 could be secreted into the sieve tubes during feeding, which might bind Ca²⁺ ions that flow into sieve tubes in response to stylet puncturing. This might suppress sieve-element clogging and facilitate continuous ingestion from sieve tubes.     

A systems model and computer simulation of the green rice leafhopper populations in control programmes

A simulation model of the green rice leafhopper-spiders system was presented. The validity of this model for simulation purpose was tested by comparing the calculated values with observed ones (Fig. 4). The effectiveness of various control measures against leafhoppers was evaluated by computer simulations. The computer simulation demonstrated that the wrong use of selective insecticide, contrary to expectation, brought an increase in the pest density, i. e., that the egg densities of leafhoppers in the 2nd and 3rd generations are increased by the insecticidal application in February, while they are decreased by the July application (Table 2). To obtain satisfactory control by sterile-male release, 320,000 sterile-male per square kilometer should be released even in the combined use with insecticides (Fig. 5). The escape of leafhoppers from predation by spiders was demonstrated by the simulation. It is suggested that spiders are able to suppress the leafhopper populations at a low density when there is a very favorable balance between spiders and leafhoppers, and this condition may be realized by sophisticated use of selective insecticides (Fig. 6). Factors and/or processes which have to be involved in a more improved systems model are discussed.     

水稻矮缩病毒对黑尾叶蝉取食及产卵偏好性的影响

水稻矮缩病毒（Rice dwarf virus, RDV）是水稻普通矮缩病（Rice dwarf disease）的病原,主要依赖黑尾叶蝉Nephotettix cincticeps经卵以持久增殖方式传播。本研究采用室内试验观测了带毒与无毒的黑尾叶蝉在健康及感病水稻间的取食、产卵选择趋性。取食偏好性试验发现,无毒叶蝉对感病水稻趋性更强,从观测的第8小时开始达显著差异水平;带毒叶蝉则对健康水稻趋性更强,在观测的第24小时开始达到显著差异水平,表明RDV侵染水稻会引起黑尾叶蝉取食偏好性的改变。产卵偏好性试验发现无毒及带毒黑尾叶蝉在健康及感病水稻间3个观测指标（每植株卵块数、每卵块的卵量及每植株上的总卵量）均无显著差异,表明RDV侵染水稻对黑尾叶蝉产卵选择偏好性无显著影响。本文初步明确RDV对黑尾叶蝉取食和产卵偏好性的影响,为揭示介体昆虫-病毒-寄主植物三者之间的互作机制提供佐证。     

High mortality caused by high dose of dsRNA in the green rice leafhopper Nephotettix cincticeps (Hemiptera: Cicadellidae)

RNA interference (RNAi) is a useful tool for gene functional studies in non-model insects and pest insects. Establishment of experimental conditions for RNAi, which differ from insect to insect, is important for evaluating the effect of dsRNA injection of relevant genes. When injecting dsRNA into the green rice leafhopper Nephotettix cincticeps (Uhler), high mortality was observed. Therefore, the adverse effects of injection of dsRNA on leafhopper development were examined to assess the suitable conditions for RNAi in this species. Injection manipulation by using a glass capillary did not affect leafhopper survival but delayed the molt of the corresponding instar. High mortality was observed when large amounts of dsRNA were administered. This adverse effect of dsRNA was examined in 2 genes, exogenous EGFP gene and endogenous peptidoglycan recognition protein gene (NcPGRP12). Injection of a high dose (60 ng/insect) caused high mortality in all stages tested: 4th instar, 5th instar, and female adult. A relatively low dose (6 ng/insect) did not cause high mortality, retaining a high potential for gene silencing. Since RNAi is highly effective in this species, the deleterious effect of large amounts of dsRNA could be avoided by administering a low dsRNA dose.     

Numerous peptidoglycan recognition protein genes expressed in the bacteriome of the green rice leafhopper Nephotettix cincticeps (Hemiptera,Cicadellidae)

Applied Entomology and Zoology - Peptidoglycan recognition proteins (PGRPs) contribute to the immune response of vertebrates and higher invertebrates. Animals have a varying number of PGRP genes: 4...     

Life tables and population parameters of the green leafhopper,Nephotettix cincticeps (Uhler), in the southwestern district of Japan with special reference to the first generation on foxtail grass

The three year (1988–1990) life tables of Nephotettix cincticeps were constructed, and the daily survival rate and longevity of female adults were estimated by Hokyo and Kiritani's (1967) method for the overwintering and the first-generations on the foxtail grass in Okayama, southwestern Japan. The life tables and the population parameter values estimated were compared with those in the other generations on rice. The FARMCOP suction sampler was employed to survey the population density. The durations of pre-ovarial maturation of female adults of the 1st generation on foxtail grass and rice seedling were similar. Longevity of adults of the overwintering and the first generations which emerged on the wild host was longer than that of the other generations (2nd and 3rd generations) on the rice plants. Fecundity of females decreased successively as the generation proceeded and it became lowest in the final 3rd generation. Only about 3.5 percent of first-instar nymphs of the 1st generation emerged as adults in the fallow field. The survival rate of nymphs on foxtail grass was always lower in comparison with that on rice plants. However, the survival rates of nymphs on foxtail grass and rice seedling were not significantly different from each other under laboratory conditions. In the fields, senescence of foxtail grass occurred in the midst of nymphal period of the 1st generation. The survival rate of nymphs on foxtail grass decreased with the increasing in the nymphal density. Abundance of spiders during the 1st generation was higher than that in the early stage of rice plants.     

Population dynamics of the green leafhopper,Nephotettix virescens Distant (Hemiptera: Cicadellidae) in synchronized and staggered transplanting areas of paddy fileds in Indonesia

The population dynamics of Nephotettix virescens, a vector of rice tungro virus disease was investigated in a synchronized transplanting area at Jatisari (1984–1986), West Java and in a staggered transplanting area at Sidan (1986–1988), Bali, Indonesia. The FARMCOP suction sampler was employed for population censuses of N. virescens and its natural enemies. The population growth pattern was affected by transplanting pattern: In the staggered transplanting area, the population density increased from the immigrant generation to the first generation, and sharply decrease thereafter, while in the synchronized transplanting area the population density often reached the highest peak in the second generation. The degree of contageousness in the spatial distribution of N. virescens was negatively correlated with population density of the immigrant generation.     

黑尾叶蝉卵黄原蛋白受体基因cDNA的克隆、序列分析及表达模式

【目的】卵黄原蛋白受体(vitellogenin receptor,VgR)属于低密度脂蛋白受体,通过介导内吞作用为发育中的卵母细胞摄取卵黄原蛋白,为胚胎发育提供营养物质,在昆虫生殖过程中发挥关键作用。为研究黑尾叶蝉Nephotettix cincticeps VgR(NcVgR)基因的生理功能及其在生殖中的作用,本研究克隆并解析了NcVgR基因的序列,并对其时空表达进行了研究。【方法】根据黑尾叶蝉转录组数据信息,利用RT-PCR克隆了NcVgR基因,并进行了生物信息学分析;利用实时荧光定量PCR研究了不同发育时期、成虫不同组织NcVgR的表达水平。【结果】NcVgR c DNA序列全长6 676 bp,开放阅读框长度5 568 bp,编码1 855个氨基酸,预测编码蛋白的分子量为206 k D,N端前17个氨基酸为信号肽。序列分析显示,NcVgR具有低密度脂蛋白家族的5个经典保守域,即:配体结合域(ligand-binding domain,LBD)、表皮生长因子前体同源域(EGF-precursor homology domain,EGFP)、O-糖链结构域(O-linked sugar domain,OLSD)、跨膜域(transmembrane domain,TMD)和胞质尾域(cytoplasmic domain)。系统发育分析表明,NcVgR与褐飞虱N.lugens VgR亲缘关系最近。实时荧光定量PCR结果显示,NcVgR转录起始时间为5龄若虫,羽化后转录水平逐渐上升,至羽化后8 d达到峰值,随后下降。有意思的是,随着黑尾叶蝉产卵,NcVgR转录水平再次上升,至羽化后16 d达到最高水平。组织定位结果显示,NcVgR在黑尾叶蝉雌成虫卵巢中特异性高表达,而在雌成虫脂肪体和肠道中微量表达,在雌成虫脑及雄成虫中均未检测到表达。【结论】NcVgR在黑尾叶蝉雌成虫卵巢中特异性表达,并且不同发育时期具有不同的表达量,这为研究黑尾叶蝉的生殖调控机理提供了分子信息。