Characterization of olfactory receptor neurons for pheromone candidate and plant volatile compounds in the clover root weevil,Sitona lepidus

Antennal olfactory receptor neurons (ORNs) for pheromone and plant volatile compounds were identified and characterized in male and female clover root weevil, Sitona lepidus (Gyllenhal), using the single sensillum recording technique with five pheromone-related compounds, and 40 host and non-host plant volatile compounds. Overall, seven different types of olfactory sensilla containing specialized ORNs were identified in each sex of S. lepidus. Among them, three different types of sensilla in the males and two types in the females housed ORNs specialized for pheromone-related compounds. The ORNs in males were specialized for 4-methyl-3,5-heptanedione or one or more of four stereoisomers of 5-hydroxy-4-methyl-3-heptanone. In contrast, female sensilla did not contain ORNs sensitive to 4-methyl-3,5-heptanedione while they contained ORNs sensitive to and specialized for the stereoisomers of (4S,5S)-5-hydroxy-4-methyl-3-heptanone. In addition to the pheromone-related ORNs, four types of olfactory sensilla contained ORNs responsive to plant volatile compounds in male S. lepidus, and five types in females. Most of the ORNs identified in S. lepidus showed a high degree of specificity to specific volatile compounds although some of the active compounds showed overlapping response spectra in the ORNs across different types of sensilla. The most active plant volatile compounds were the four green leaf volatile compounds, (E)-2-hexenol, (Z)-2-hexenol, (Z)-3-hexenol and (E)-2-hexenal, and isomers of two monoterpenols, (±)-linalool and (±)-α-terpineol, all eliciting strong responses from relatively large numbers of ORNs in male and female S. lepidus. Our study indicates that S. lepidus has a set of highly sensitive and selective ORNs for pheromone and plant volatile compounds. Further work is needed to elucidate the behavioral implications of these findings.     

Heterochromatic banding patterns on chromosomes of twelve weevil species (Insecta,Coleoptera, Curculionoidea: Apionidae,Curculionidae)

The C-banding patterns of twelve weevil species are presented. The obtained results confirm the existence of two groups of species: with a small or large amount of heterochromatin in the karyotype. The first group comprises seven species (Apionidae: Holotrichapion pisi; Curculionidae: Phyllobius urticae, Ph. pyri, Ph. maculicornis, Tanymecus palliatus, Larinodontes turbinatus, Cionus tuberculosus). In weevils with a small amount of heterochromatin, tiny grains on the nucleus in interphase are visible, afterwards in mitotic and meiotic prophase appearing as dark dots. The absence of C-bands does not indicate a lack of heterochromatin but heterochromatic regions are sometimes so small that the condensation is not visible during the cell cycle. The second group comprises five species (Otiorhynchus niger, O. morio, Polydrusus corruscus, Barypeithes chevrolati, Nedyus quadrimaculatus) which possess much larger heteropicnotic parts of chromosomes visible during all nuclear divisions. The species examined have paracentromeric C-bands on autosomes and the sex chromosome X, except for Otiorhynchus niger, which also has an intercalary bands on one pair of autososomes. All the species examined differ in the size of segments of constitutive heterochromatin. The y heterochromosome is dot-like and wholly euchromatic in all the studied species.     

Host plant recognition by the root feeding clover weevil, Sitona lepidus (Coleoptera: Curculionidae)

This study investigated the ability of neonatal larvae of the root-feeding weevil, Sitona lepidus Gyllenhal, to locate white clover Trifolium repens L. (Fabaceae) roots growing in soil and to distinguish them from the roots of other species of clover and a co-occurring grass species. Choice experiments used a combination of invasive techniques and the novel technique of high resolution X-ray microtomography to non-invasively track larval movement in the soil towards plant roots. Burrowing distances towards roots of different plant species were also examined. Newly hatched S. lepidus recognized T. repens roots and moved preferentially towards them when given a choice of roots of subterranean clover, Trifolium subterraneum L. (Fabaceae), strawberry clover Trifolium fragiferum L. (Fabaceae), or perennial ryegrass Lolium perenneL. (Poaceae). Larvae recognized T. repens roots, whether released in groups of five or singly, when released 25 mm (meso-scale recognition) or 60 mm (macro-scale recognition) away from plant roots. There was no statistically significant difference in movement rates of larvae.     

Using molecular genetics to identify immature specimens of the weevil Ceratapion basicorne (Coleoptera: Apionidae)

Molecular analyses can play a primary role in the process of host specificity evaluation at species and population levels. Here we present an example of their application with a promising candidate biological control agent for yellow starthistle, Centaurea solstitialis L. Although it is highly host specific, Ceratapion basicorne (Coleoptera: Apionidae) can develop on safflower in laboratory tests. A field experiment was conducted to further evaluate host plant specificity; however, it was not possible to rear all larvae to the adult stage, which was necessary for species determination. Therefore molecular genetic methods were used to identify immature specimens. A 731 bp fragment of mtDNA cytochrome C oxidase I gene (COI) was sequenced from 41 individuals of C. basicorne and four congeners: Ceratapion orientale, Ceratapion onopordi, Ceratapion penetrans and Ceratapion scalptum. Intraspecific variability ranged from 0.0% to 0.2%, and interspecific divergences ranged from 1.7% to 17.6%. All larvae that were sequenced from the field study, clearly matched one of the five species, enabling us to unambiguously identify them. Use of molecular genetics to identify larvae should also help the process of foreign exploration, enabling the identification of field-collected larvae, which often provide more reliable host plant associations than field collected adults.     

Mango seed weevil (Coleoptera: Curculionidae) and premature fruit drop in mangoes

The effect of infestations of mango seed weevil, Sternochetus mangiferae (F.), on premature fruit drop of mangoes was investigated. Mango fruits ('Haden') of equal size were collected both off the ground and from the tree at four times during the season (June-August). If weevil-infested fruit were more prone to dropping than uninfested fruit, the prediction was that a higher infestation rate would be found in fruit on the ground compared with fruit on the tree. Average fruit weight was used as an indicator of fruit maturity. The seed infestation rate was significantly higher in fruit collected off the ground compared with fruit collected from the tree in 38 g and 79 g (early-season) fruit but not significantly different in 207 g (midseason) and 281 g (late season) fruit. The age distribution of weevils and the number of insects in infested fruits were similar for ground and tree fruits on all dates. Results suggest that mango seed weevil infestation can increase fruit drop during early fruit development.     

Characterization of microsatellite loci in Brachyderes rugatus,the Canary Islands pine weevil (Coleoptera: Curculionidae)

The Canary Island pine weevil Brachyderes rugatus (Wollaston) consists of four allopatric subspecies that are thought to have arisen from several historic colonization events within the archipelago. We have isolated and optimized seven microsatellite loci from Brachyderes rugatus calvus from Gran Canaria. Six of these loci are polymorphic within B. rugatus (11–22 alleles per locus; heterozygosity between 0.43 and 0.84). There is no evidence for heterozygote deficit within populations or for linkage disequilibrium between pairs of loci. These molecular markers are likely to prove useful tools for quantifying the genetic variability of bottlenecked island populations.     

The Apionidae (Coleoptera: Curculionoidea) of New Caledonia, genus Caledonapion Wanat

The New Caledonian endemic weevil genus Caledonapion is revised, with one new species, C . assimile , described. Diagnoses, a key and distribution maps are provided for both species of the genus. An association of Caledonapion with the plant genus Hedycarya (Monimiaceae) is suggested from field observations.     

Effect of mango weevil (Coleoptera: Curculionidae) damage on mango seed viability in Hawaii

The mango weevil, Cryptorhynchus (= Sternochetus) mangiferae (F.), is a federally quarantined pest that prevents shipment of mangos from Hawaii into the continental United States. Although this monophagous weevil allegedly causes reduced seed germination, damage to the fruit pulp, and premature fruit drop in mangos, there are few studies examining these potential sources of crop loss. We conducted studies to assess the effect of mango weevil infestation on seed viability while making observations on the frequency of pulp feeding. Naturally infested seeds from mature fruit were planted in pots and scored for successful germination. Germination rates for infested seeds were equal to those of uninfested control seeds in a polyembryonic cultivar ('Common'), whereas germination was significantly reduced for infested seeds of a monoembryonic cultivar ('Haden') compared with uninfested control seeds but germination of infested seeds was still > 70%. To assess seed tolerance of damage, seeds were artificially damaged by cutting away 25, 50, or 75% of the cotyledon before planting and scored for germination. None of the damage treatments was significantly different from the undamaged controls, indicating that mango seeds can withstand substantial damage and still germinate successfully. Over the 2-yr period we conducted experiments, only four of 3,602 mango fruits (0.11%) showed evidence of direct feeding damage to the pulp. Results suggest that C. mangiferae is a less serious pest of mangos than previously thought.     

Effect of mango weevil (Coleoptera: Curculionidae) damage on mango seed viability in Tanzania

Studies were conducted at the horticulture unit of Sokoine University of Agriculture, Morogoro, Tanzania, to assess the incidence and effect of mango weevil, Cryptorhynchus mangiferae (F.) (Coleoptera: Curculionidae), infestation on mango, Mangifera indica L., seed viability. Three polyembryo mango cultivars ('Sindano nyeusi', 'Sindano nyeupe', and 'Dodo') as well as three monoembryo mango cultivars ('Ex-horticulture', 'Tango', and 'Bongwa') were collected and examined for the presence of C. mangiferae. The effect of seed damage on viability was assessed for both naturally and artificially damaged seeds. However, for artificially damaged seeds, the viability was assessed by cutting away 0, 25, 50, or 75% of the cotyledon before planting. In this experiment, only monoembryo mango cultivars were used. All the examined cultivars were infested by C. mangiferae, although at varying levels. Polyembryo mango cultivars were relatively more infested than monoembryo cultivars. Bongwa and Tango were least infested, whereas Sindano nyeusi recorded the highest C. mangiferae incidence. Germination rates of damaged seeds of polyembryonic cultivars differed significantly from the uninfested control, except for Sindano nyeusi. There were no significant differences in germination percentage among the three monoembryo cultivars, and all the cultivars differed significantly from the uninfested control. The germination rates of seeds with 25% of their cotyledons removed did not differ significantly from the undamaged seeds, indicating that monoembryo cultivar seeds can withstand up to 25% damage and germinate successfully.     

Hypersensitive response of beans to Apion godmani (Coleoptera: Curculionidae)

High levels of resistance to Apion godinani Wagner have been reported in bean, Phaseolus vulgaris L., landraces from Mexico. We report on the role of hypersensitivity to A. godmani in five resistant and three susceptible bean genotypes. In susceptible genotypes (cultivars 'Canario 107','Jamapa', and 'Zacatecas 45'), the eggs and first instars of A. godmani were embedded in the pod mesocarp and usually were surrounded by healthy tissue. In contrast, in resistant landraces ('Amarillo 154', 'Amarillo 155', 'J-117', 'Puebla 36', and 'Pinto 168'), necrotic tissues developed concentrically around the oviposition site, encapsulating eggs and dead larvae. An inverse relationship between percentage egg and larval encapsulation at the early immature pod stages and percentage of damaged seeds at harvest was found. Results indicate that hypersensitivity in developing pods plays an important role in antibiosis to A. godmani in beans. This information will facilitate future genetic and biochemical research and provide much needed information concerning the phenotypic basis of resistance to A. godmani in bean.     

Effects of carbon dioxide on the searching behaviour of the root-feeding clover weevil Sitona lepidus (Coleoptera: Curculionidae)

The respiratory emission of CO2 from roots is frequently proposed as an attractant that allows soil-dwelling insects to locate host plant roots, but this role has recently become less certain. CO2 is emitted from many sources other than roots, so does not necessarily indicate the presence of host plants, and because of the high density of roots in the upper soil layers, spatial gradients may not always be perceptible by soil-dwelling insects. The role of CO2 in host location was investigated using the clover root weevil Sitona lepidus Gyllenhall and its host plant white clover (Trifolium repens L.) as a model system. Rhizochamber experiments showed that CO2 concentrations were approximately 1000 ppm around the roots of white clover, but significantly decreased with increasing distance from roots. In behavioural experiments, no evidence was found for any attraction by S. lepidus larvae to point emissions of CO2, regardless of emission rates. Fewer than 15% of larvae were attracted to point emissions of CO2, compared with a control response of 17%. However, fractal analysis of movement paths in constant CO2 concentrations demonstrated that searching by S. lepidus larvae significantly intensified when they experienced CO2 concentrations similar to those found around the roots of white clover (i.e. 1000 ppm). It is suggested that respiratory emissions of CO2 may act as a 'search trigger' for S. lepidus, whereby it induces larvae to search a smaller area more intensively, in order to detect location cues that are more specific to their host plant.     

Boll weevil (Coleoptera: Curculionidae) survival through the seed cotton cleaning process in the cotton gin

There is concern that gins located in boll weevil, Anthonomus grandis grandis Boheman, eradication zones may become points of reintroduction when they process cotton grown in a neighboring infested area. We estimated boll weevil survival through two typical machine sequences used in commercial cotton gins to clean and dry the seed cotton in advance of the gin stand, as well as separately through two incline cylinder cleaners or one or two tower dryers operating at different temperatures. Large numbers of laboratory-reared adult boll weevils were marked with fluorescent powder, fed into the test system, and recovered with the assistance of blacklights. We found no evidence of survival through the seed cotton cleaning systems even when the dryers were not heated, or when passed separately through the two incline cleaners alone. Upper confidence limits (95%) were calculated for the observed zero recoveries based on sample size and the binomial distribution, and these represent the statistical worst-case (i.e., highest) survival potential. Survival through heated tower dryers declined rapidly to zero at higher temperatures, especially when two dryers were running. Although we conclude that the potential for survival of weevils in the seed cotton to the gin stand is zero or close to zero, a small percentage of live weevils was recovered in the green boll/rock trap, which may represent the greatest threat of reintroduction at the gin. Escape of live weevils with the gin trash is also possible, and studies addressing this issue will be presented elsewhere.     

Characterization of microsatellite loci in white pine weevil (Pissodes strobi)

Microsatellites are codominant and highly polymorphic genetic markers. They are widely used in studying parentage and reproductive success in many insect species. We developed six microsatellite markers in Pissodes storbi. The number of alleles per locus ranges from 5 to 16 alleles, with a mean number of 10. The high allelic diversity observed in this study showed that they could be useful in the study of parentage and reproductive success in P. strobi.     

A new species of genus Pseudaspidapion Wanat, 1990 (Coleoptera, Apionidae) from China

Pseudaspidapion botanicumsp. n. from China is described and figured. Its host plant is Grewia biloba G. Don var. parviflora (Bunge) Hand.-Mazz (Malvaceae: Grewioideae). The genus Harpapion Voss, 1966 is recorded as new for China and Vietnam and two comb. n. are proposed: Harpapion vietnamense (Korotyaev, 1985) (from Aspidapion) and Harpapion coelebs (Korotyaev, 1987) (from Pseudaspidapion). A key to the known species of the genus Pseudaspidapion from China is presented.     

Observations on the biology and ecology of Apion vorax (Coleoptera: Apionidae), a vector of broad bean stain and broad bean true mosaic viruses

In spring 1974 and 1975 many adult Apion vorax were found at Rothamsted in woods bordering fields that had been cropped with field beans (Vicia faba) the previous season. The weevils were most common on bramble (Rubus spp.), dog's mercury (Mercurialis perennis) and nettle (Urtica dioica). A decrease in the woodland population in May coincided with the appearance of the weevil on field bean crops in neighbouring areas. In laboratory feeding tests, adult weevils fed more on field bean leaves than on leaves of plants on which they were common in woodlands, and they fed more on field beans in May than at other times of the year. No difference was observed in the feeding of male and female weevils. Many A. vorax were caught on sticky traps in woodlands at Rothamsted in 1975, particularly in April and May, but very few were caught on sticky traps in bean crops. The numbers of A. vorax recorded on the upper foliage of bean crops at Rothamsted in June each year from 1970 to 1980 ranged from 0 to 10 per 10-m row. Eggs, larvae and pupae of A. vorax were found in field bean flowers collected from field crops. The larvae were feeding on the tips of the pistils and stamens. Adult weevils that emerged in mid-June from the flowers of autumn-sown beans mated and produced a second generation when caged on glasshouse-grown plants. Some Apion larvae, possibly A. vorax, were found inside flowers of common vetch (Vicia sativa). The name ‘bean flower weevil’ is considered a suitable common name for A. vorax.     

Primary culture of lobster (Homarus americanus) olfactory sensory neurons

Lobster olfactory sensory neurons have contributed to a number of advances in our understanding of olfactory physiology. To facilitate further study of their function, we have developed conditions allowing primary culture of the olfactory sensory neurons in a defined medium. The most common cells in the culture were round cell bodies with diameters of 10-15 micro m that often extended fine processes, features resembling olfactory sensory neurons. We discovered that acetylcholinesterase acted as a growth factor for these cells, improving their survival in culture. We also confirmed previous evidence from spiny lobsters that poly-D-lysine was a superior substrate for olfactory cells of this size and morphology. We then identified olfactory sensory neurons in the culture in two ways. Almost half the cells tested responded to application of a complex odorant with an inward current. An even more rigorous test was made possible by the development of an antiserum to OET-07, an ionotropic glutamate receptor homolog specifically expressed by Homarus americanus olfactory sensory neurons. It labeled a majority of the round cells in the culture, unequivocally identifying them as olfactory sensory neurons.     

Interactions between Apion species (Coleoptera: Curculionidae) and Polygonaceae. I. Apion curtirostre Germ, and Rumex acetosa L.

Abstract.

• 1 Apion curtirostre is the commonest weevil attacking Rumex acetosa in the Lancaster area.
• 2 The ovipositional behaviour, larval development, emergence and hibernation of A. curtirostre are described.
• 3 From dissection of samples of stems of R. acetosa, abundance curves for the stages of A. curtirostre and its chalcid parasitoids were constructed and the between-stage mortalities determined.
• 4 There is an exponential relationship between R. acetosa stem height and number of A. curtirostre eggs laid.
• 5 Male stems of R. acetosa appear earlier and are smaller than female stems. Taller stems, at least in female plants, live longer than shorter ones.
• 6 Apion survival is lower in male than in female stems.
• 7 Stem sex is the primary determinant of opposition choice with stem thickness a secondary determinant.
• 8 A. curtirostre eggs introduced experimentally into stems of four Rumex species survived significiantly better in R. acetosa than in any other species. Survival in R. acetosa female plants was twice that in males.
• 9 Survival of A. curtirostre eggs at densities of 4, 8 and 16 per R. acetosa stem was twice that at densities of 32 and 64 per stem.
• 10 The stem size of female R. acetosa was reduced as the number of attacking weevils increased. There was no significant effect in males. No adverse effect on seed production was detected.
• 11 The nature of the relationship between A. curtirostre and R. acetosa is discussed.

     

Inheritance of resistance to the bean-pod weevil (Apion godmani Wagner) in common beans from Mexico

The bean-pod weevil (BPW), Apion godmani Wagner, often causes heavy losses in crops of common bean (Phaseolus vulgaris L.). Farmers need resistant bean cultivars to minimize losses, cut production costs, stabilize seed yield, and reduce pesticide use and consequent health hazards. To design effective breeding methods, breeders need new and better sources of resistance and increased knowledge of their modes of inheritance. We therefore: (1) compared sources of resistance to BPW, (2) studied the inheritance of resistance, and (3) determined whether the sources possess similar or different genes for BPW resistance. The following sources of resistance, originating from the Mexican highlands, were evaluated for 3 years at INIFAP-Santa Lucía de Prias, Texcoco, Mexico: Amarillo 153, Amarillo 169, Hidalgo 58, J 117, Pinto Texcoco, Pinto 168, and Puebla 36. All except Puebla 36 were crossed with the susceptible cultivar Jamapa. Amarillo 153 and Puebla 36 were crossed with another susceptible cultivar, Bayo Mex. The parents, F₁ hybrids, and F₂ populations were evaluated for BPW damage in 1992. Backcrosses of the F₁ of Jamapa/Pinto 168 to the respective susceptible and resistant parents were also evaluated in 1992. All seven resistant accessions were crossed in all possible combinations, excluding reciprocals. The resulting 21 F₁ hybrids and 21 F₂ populations were evaluated for BPW damage in 1994. J 117 had the highest level of resistance to BPW. Pinto Texcoco and Puebla 36 had the highest mean damage score of all seven sources of resistance. The F₁ hybrids between susceptible parents and resistant sources were generally intermediate. Two genes segregating independently controlled the BPW resistance in each accession. One gene, Agm, has no effect when present alone, whereas the other gene, Agr, alone conferred intermediate resistance. When both genes were present, resistance to BPW was higher. Based on mean BPW damage scores, all 21 F₁ hybrids and their F₂ populations, derived from crosses among seven resistant accessions, were resistant. However, data from individual plant damage scores in F₂ populations of Amarillo 169/Pinto 168 and Pinto Texcoco/Pinto 168 suggested that at least one gene in each of the three accessions was non-allelic. Data also indicated that Amarillo 169 had a dominant gene that conferred high levels of BPW resistance, irrespective of the alleles at the other locus; and that Pinto Texcoco and Pinto 168 possessed two different genes for intermediate resistance.     

云南木蠹象的生物学研究

云南木蠹象Pissodes yunnanensis Langor et Zhang是我国西南地区近年来发现的森林蛀干害虫。该虫在云南西北部的丽江地区一年发生一代。成虫于6月下旬开始产卵, 产卵部位为当年生或头年生枝梢。7月上旬幼虫开始孵化,幼虫有4个龄期。11月中旬后,幼虫开始在受害枝内越冬,次年2月中下旬恢复活动。3月下旬至5月上旬为蛹期,4月中旬开始羽化。该虫主要危害云南松幼树,常造成受害树长势下降和树干畸形,连续危害2～3年可导致树木干枯死亡。