Transmission of Spiroplasma citri by the aster leafhopper Macrosteles fascifrons (Homoptera: Cicadellidae)

The aster leafhopper (Macrosteles fascifrons), injected with an isolate of Spiroplasma citri obtained from brittle root-diseased horseradish (Armoracia rusticana), transmitted the spiroplasma to horseradish and China aster (Callistephus chinensis.) After feeding on plants infected with S. citri, M. fascifrons transmitted the spiroplasma from aster to aster and horseradish, from yellow rocket (Barbarea vulgaris) to aster, and from turnip (Brassica rapa) to turnip. Symptoms in infected horseradish were chlorosis and stunting of newly formed leaves, discoloration of root phloem, and reduced plant growth typical of brittle root disease. Chlorosis, stunting, and asymmetry of young leaves occurred in affected aster and turnip. Flowers of infected aster were small and pale in colour and occasionally showed other symptoms including asymmetry, petal distortion, or light green petals. Spiroplasmas were isolated from all plants showing symptoms. Transmission rates by M. fascifrons which acquired S. citri by feeding on infected plants were very low, but injected leafhoppers transmitted more frequently. This is the first report of the transmission of S. citri from diseased to healthy plants by M. fascifrons.     

Multiplication and morphology of Spiroplasma citri in the leafhopper Euscelis plebejus

Spiroplasma citri multiplied in all Euscelis plebejus leaf hoppers injected and sometimes reached titres of over 1 × 10⁷ colony forming units per insect. Spiroplasmas could be isolated from the haemolymph at all times although helices were only apparent for a few days after injection. The salivary glands of injected insects contained membrane bound pockets densely packed with mycoplasma-like bodies. These bodies were frequently infected with virus-like particles similar to those found in cultures of S. citri. Spiroplasmas had little effect on the longevity of the leafhoppers.     

Spiralin is not essential for helicity, motility, or pathogenicity but is required for efficient transmission of Spiroplasma citri by its leafhopper vector Circulifer haematoceps

Spiralin is the most abundant protein at the surface of the plant pathogenic mollicute Spiroplasma citri and hence might play a role in the interactions of the spiroplasma with its host plant and/or its insect vector. To study spiralin function, mutants were produced by inactivating the spiralin gene through homologous recombination. A spiralin-green fluorescent protein (GFP) translational fusion was engineered and introduced into S. citri by using an oriC-based targeting vector. According to the strategy used, integration of the plasmid by a single-crossover recombination at the spiralin gene resulted in the expression of the spiralin-GFP fusion protein. Two distinct mutants were isolated. Western and colony immunoblot analyses showed that one mutant (GII3-9a5) did produce the spiralin-GFP fusion protein, which was found not to fluoresce, whereas the other (GII3-9a2) produced neither the fusion protein nor the wild-type spiralin. Both mutants displayed helical morphology and motility, similarly to the wild-type strain GII-3. Genomic DNA analyses revealed that GII3-9a5 was unstable and that GII3-9a2 was probably derived from GII3-9a5 by a double-crossover recombination between plasmid sequences integrated into the GII3-9a5 chromosome and free plasmid. When injected into the leafhopper vector Circulifer haematoceps, the spiralinless mutant GII3-9a2 multiplied to high titers in the insects (1.1 x 10(6) to 2.8 x 10(6) CFU/insect) but was transmitted to the host plant 100 times less efficiently than the wild-type strain. As a result, not all plants were infected, and symptom production in these plants was delayed for 2 to 4 weeks compared to that in the wild-type strain. In the infected plants however, the mutant multiplied to high titers (1.2 x 10(6) to 1.4 x 10(7) CFU/g of midribs) and produced the typical symptoms of the disease. These results indicate that spiralin is not essential for pathogenicity but is required for efficient transmission of S. citri by its insect vector.     

Disruption of a gene predicted to encode a solute binding protein of an ABC transporter reduces transmission of Spiroplasma citri by the leafhopper Circulifer haematoceps

Spiroplasma citri is transmitted from plant to plant by phloem-feeding leafhoppers. In an attempt to identify mechanisms involved in transmission, mutants of S. citri affected in their transmission must be available. For this purpose, transposon (Tn4001) mutagenesis was used to produce mutants which have been screened for their ability to be transmitted by the leafhopper vector Circulifer haematoceps to periwinkle plants. With one mutant (G76) which multiplied in leafhoppers as efficiently as S. citri wild-type (wt) strain GII-3, the plants showed symptoms 4 to 5 weeks later than those infected with wt GII-3. Thirty to fifty percent of plants exposed to leafhoppers injected with G76 remained symptomless, whereas for wt GII-3, all plants exposed to the transmission showed severe symptoms. This suggests that the mutant G76 was injected into plants by the leafhoppers less efficiently than wt GII-3. To check this possibility, the number of spiroplasma cells injected by a leafhopper through a Parafilm membrane into SP4 medium was determined. Thirty times less mutant G76 than wt GII-3 was transmitted through the membrane. These results suggest that mutant G76 was affected either in its capacity to penetrate the salivary glands and/or to multiply within them. In mutant G76, transposon Tn4001 was shown to be inserted into a gene encoding a putative lipoprotein (Sc76) In the ABCdb database Sc76 protein was noted as a solute binding protein of an ABC transporter of the family S1_b. Functional complementation of the G76 mutant with the Sc76 gene restored the wild phenotype, showing that Sc76 protein is involved in S. citri transmission by the leafhopper vector C. haematoceps.     

Disruption of a Gene Predicted To Encode a Solute Binding Protein of an ABC Transporter Reduces Transmission of Spiroplasma citri by the Leafhopper Circulifer haematoceps

Spiroplasma citri is transmitted from plant to plant by phloem-feeding leafhoppers. In an attempt to identify mechanisms involved in transmission, mutants of S. citri affected in their transmission must be available. For this purpose, transposon (Tn4001) mutagenesis was used to produce mutants which have been screened for their ability to be transmitted by the leafhopper vector Circulifer haematoceps to periwinkle plants. With one mutant (G76) which multiplied in leafhoppers as efficiently as S. citri wild-type (wt) strain GII-3, the plants showed symptoms 4 to 5 weeks later than those infected with wt GII-3. Thirty to fifty percent of plants exposed to leafhoppers injected with G76 remained symptomless, whereas for wt GII-3, all plants exposed to the transmission showed severe symptoms. This suggests that the mutant G76 was injected into plants by the leafhoppers less efficiently than wt GII-3. To check this possibility, the number of spiroplasma cells injected by a leafhopper through a Parafilm membrane into SP4 medium was determined. Thirty times less mutant G76 than wt GII-3 was transmitted through the membrane. These results suggest that mutant G76 was affected either in its capacity to penetrate the salivary glands and/or to multiply within them. In mutant G76, transposon Tn4001 was shown to be inserted into a gene encoding a putative lipoprotein (Sc76) In the ABCdb database Sc76 protein was noted as a solute binding protein of an ABC transporter of the family S1_b. Functional complementation of the G76 mutant with the Sc76 gene restored the wild phenotype, showing that Sc76 protein is involved in S. citri transmission by the leafhopper vector C. haematoceps.     

Production and characteristics of antisera to Spiroplasma citri and clover phyllody-associated antigens derived from plants

Antisera were produced to clover phyllody- and Spiroplasma citri-associated antigens partially purified from infected Vinca rosea plants. Separate antisera were made to ‘membrane fraction’ (MF) preparations comprising the resuspended pellet obtained by high speed centrifugation, and to ‘soluble fraction’ (SA) preparations, comprising the supernatant from high speed centrifugation concentrated by freeze-drying. All antisera showed considerable activity against normal plant antigens but after cross-absorption with extracts of healthy plants the MF antisera were used in F(ab')₂based ELISA tests to detect S. citri- or clover phyllody-associated antigens in infected plants. The ‘clover phyllody’ antiserum also reacted specifically with extracts of clover plants with phyllody, and with naturally-infected strawberry plants showing symptoms of green petal disease. Both the ‘clover phyllody’ and S. citri antisera were specific for their respective homologous antigens. No cross-reactions were observed in heterologous tests or between either antiserum and extracts of V. rosea infected with various MLOs obtained from different host plants.     

Infection and replication sites of Spiroplasma kunkelii (Class: Mollicutes) in midgut and Malpighian tubules of the leafhopper Dalbulus maidis

Spiroplasma kunkelii distribution and infection mechanisms in the intestines and Malpighian tubules of Dalbulus maidis were investigated by transmission electron microscopy. Spiroplasmas were found between microvilli and in endocytic vesicles of the midgut epithelium. At the basal part, cytoplasmic vesicles contained multiple spiroplasmas with tube-like extensions and spiroplasmas accumulated between the laminae rara and densa of the basal lamina. Tip structures of flask-shaped spiroplasmas pierced the lamina densa that was discontinuous in close proximity to spiroplasmas. Spiroplasmas were found in hemolymph, crossed the basal lamina of Malpighian tubule epithelium and accumulated at high numbers in muscle cells that had cytopathogenic changes. S. kunkelii had perithrochous approximately 8nm diameter structures determined to be fimbriae protruding from the cell surface, and similar structures were adhering to the basal lamina of midgut epithelium and to external lamina of muscle cells. Further, spiroplasmas had pili-like appendages at one or both cell poles and appeared to conjugate. This is the first time that fimbriae and pili have been observed in a mollicutes.     

A remarkable new opsiine leafhopper genus (Hemiptera: Cicadellidae: Deltocephalinae) from Southern China,with notes on its phylogenetic position

A remarkable new genus of tribe Opsiini (Deltocephalinae), Yinformibus gen. nov. including Yinformibus menglaensis sp. nov. is described from Yunnan of China. The new genus is placed in the tribe Opsiini based on male adults having a pair of aedeagal shafts, each with its own gonopore. Partial 28S rDNA and Histone H3 sequences are provided for the new species, and a phylogenetic analysis based on these markers for Deltocephalinae retrieved from GenBank suggests Opsiini is monophyletic and supports the placement of Yinformibus gen. nov. in Opsiini. The molecular results placed the new genus in a clade comprising Hishimonus Ishihara, Opsius Fieber, and an undescribed genus from Zambia. Additionally, we also analysed the relationships of Yinformibus gen. nov. with other genera based on morphology.

http://www.zoobank.org/urn:lsid:zoobank.org:pub:85C23AC3-8B19-4F29-9F55-BFC392242B51     

A new species and new records of the leafhopper genus Taperus Li & Wang, 1994 (Hemiptera,Cicadellidae, Evacanthinae) from China

The paper deals with the species of the Oriental leafhopper genus Taperus Li & Wang. A new species, Taperus daozhenensissp. n., from Guizhou Province, China is described and new records for other Chinese species are given together with a key for their separation. The type specimens of the new species are deposited in the Institute of Entomology, Guizhou University (GUGC).     

A new species of the leafhopper genus Diomma Motschulsky (Hemiptera, Cidadellidae, Typhlocybinae) from China

In the present paper, a new species is added to the genus Diomma Motschulsky (Hemiptera: Cicadellidae: Typhlocybinae) from Southwest China, Diomma pincersasp. n. At the same time, a key can distinguish all Chinese species of the genus is provided.     

Description of two new species of the leafhopper subgenus Pediopsoides (Pediopsoides) (Hemiptera,Cicadellidae, Macropsinae) from Guangxi Province,Southern China

Two new species of the Macropsinae leafhopper subgenus Pediopsoides (Pediopsoides) Matsumura, 1912, Pediopsoides (Pediopsoides) damingshanensis Li, Dai & Li, sp. n. and Pediopsoides (Pediopsoides) tishetshkini Li, Dai & Li sp. n., are described and illustrated from Guangxi Province of southern China. A key to males is provided to distinguish the species of the subgenus along with a map showing the distribution of the new species.     

假眼小绿叶蝉卵的寄生蜂种类及种群动态

通过田间采梢、室内镜下查卵和饲养观察,初步获得假眼小绿叶蝉Empoasca vitis Gthe卵寄生蜂2种,分别隶属于三棒缨小蜂属Stethynium sp.和裂骨缨小蜂属Schizophragma sp.,其中三棒缨小蜂属Stethyniumsp.是优势种。2种缨小蜂的寄生动态调查结果显示,除8月和12月寄生率较低外,其它月份的自然寄生率都在30%以上,特别是10~11月寄生率最高达65%,对小绿叶蝉的虫口数量具有重要控制作用,值得进一步研究和保护利用。     

柑桔木虱及其防治

柑砧术虱是柑桔新梢期重要害虫,刺吸为害。福建福州在柑桔上一年发生8代,在月桔、九里香上一年约发生10代左右,世代重叠,以成虫在柑桔叶背群集越冬,无完全滞育。4—5月气温22．3℃世代历期42．5天．6-7月27．2℃为24．4天．8月28．1℃为23．2天。10-12月19.6℃为52．9天。成虫寿命长,约历1个月半,越冬代长达半年。田间种群消长与柑桔芽梢抽发期相对应．一年中虫口数量出现3个高峰期,第一峰期3-4月为柑桔春梢主要抽发期,第二峰期5-6月为夏梢主要抽发期,第三峰期8～9月为秋梢主要抽发期,其中以秋梢期虫口数量最大．秋芽受害最重,次为春梢期。本还对柑桔术虱天敌进行调查,并对柑桔术虱卵、若虫和成虫进行药剂防治试验。     

Detection of a Concanavalin A binding protein in the mollicute Spiroplasma citri and purification from the plasma membrane

A protein that binds Concanavalin A (Con A) was detected on Western blots of Spiroplasma citri proteins. Its apparent molecular weight was 84000. It was localized in the plasma membrane. Affinity chromatography on Con A-agarose was used to isolate this protein. The glycosylation inhibitor, tunicamycin, inhibits S. citri growth and seems to block the glycosylation of the Con A-binding protein.     

棉铃虫种群能量动态及其为害特征分析

根据棉铃虫种群数量密度、年龄结构、存活率及虫体含能量,系统地分析了８种不同类型棉田生态系统中棉铃虫种群生产力、摄入量及其为害特征．结果表明,棉铃虫种群的能量生产主要集中于第３代;其所引起棉花繁殖器官的被害量在第２、３、４代分别为１１．８３、１６．６５和９．５２个·ｍ－２．随着播种期的推后,２代棉铃虫种群生产力和摄入量减少,３、４代值增加．实行套作种植使２、４代值减少,而３代值增加．棉铃虫种群摄食利用效率随世代增加而减小,其净生态效率以第３代最高,第４代居中,第２代最小．由此进一步探讨了不同时空类型棉田棉铃虫各代管理的对策．     

Expression in Spiroplasma citri of an epitope carried on the G fragment of the cytadhesin P1 gene from Mycoplasma pneumoniae.

We have previously described the use of the replicative form (RF) of Spiroplasma citri virus SpV1 as a vector for cloning and expressing foreign genes in S. citri, an organism which reads UGA as a tryptophan codon (C. Stamburski, J. Renaudin, and J.M. Bové, J. Bacteriol. 173:2225-2230, 1991). We now report cloning and expression in S. citri of the G fragment of cytadhesin P1 gene from Mycoplasma pneumoniae. The G fragment was inserted in the SpV1 RF downstream of a synthetic ribosome binding site and introduced into S. citri by electroporation. Northern (RNA) blot analyses showed that in S. citri, the G fragment was transcribed from an SpV1 RF promoter as a 1.2-kb mRNA. The translation product was detected by Western blotting (immunoblotting) with a rabbit antiserum raised against total proteins from M. pneumoniae (strain FH) and was proved to be P1 specific by using monoclonal antibodies specific for the G region of the P1 protein. The apparent molecular mass of the polypeptide (24.5 kDa) indicates that in S. citri, the G fragment was fully translated in spite of the seven UGA codons present in the reading frame.     

The effect of food quantity and quality on the growth,birth-rate and longevity of Daphnia hyalina Leydig

To study experimentally the relation between zooplankton and phytoplankton, laboratory cultures of Daphnia hyalina Leydig were set up. The combined influence of food quality and quantity on growth, birth-rate and longevity was measured. The effect of seven different food regimes was tested. Natural unfiltered lake water from the eutrophic lake Tjeukemeer was used in one regime. The food value of the natural unfiltered lake water appeared to be relatively low, which was most likely caused by the abundance of large sized algae in the lake water.