Wolbachia infections in world populations of bean beetles (Coleoptera: Chrysomelidae: Bruchinae) infesting cultivated and wild legumes

Wolbachia endosymbionts are widespread among insects and other arthropods, often causing cytoplasmic incompatibility and other reproductive phenotypes in their hosts. Recently, possibilities of Wolbachia-mediated pest control and management have been proposed, and the bean beetles of the subfamily Bruchinae are known as serious pests of harvested and stored beans worldwide. Here we investigated Wolbachia infections in bean beetles from the world, representing seven genera, 20 species and 87 populations. Of 20 species examined, Wolbachia infections were detected in four species, Megabruchidius sophorae, Callosobruchus analis, C. latealbus and C. chinensis. Infection frequencies were partial in M. sophorae but perfect in the other species. In addition to C. chinensis described in the previous studies, C. latealbus was infected with two distinct Wolbachia strains. These Wolbachia strains from the bean beetles were phylogenetically not closely related to each other. Among world populations of C. chinensis, some Taiwanese populations on a wild leguminous plant, Rhynchosia minima, exhibited a peculiar Wolbachia infection pattern, suggesting the possibility that these populations comprise a distinct host race or a cryptic species.     

Feeding behavior of leaf beetles (Coleoptera,Chrysomelidae)

The feeding behavior of adult leaf beetles (41 species from 18 genera and 8 subfamilies) was studied for the first time. Beetles of the genera Chrysolina, Chrysomela, Cryptocephalus, Galeruca, Gastrophysa, Labidostomis, Leptinotarsa, Timarcha, and Cassida stigmatica gnaw a leaf from the edge, whereas the representatives of Donacia, Galerucella, Lema, Lilioceris, Oulema, Phyllobrotica, Plagiodera, Zeugophora, Hypocassida, and most species of Cassida gnaw the leaf plane. In addition, adults of Lilioceris merdigera and Donacia clavipes feed on young leaves rolled into a tube. New host plants are reported for the first time: Hyoscyamus niger for the larvae of the Colorado potato beetle and Naumburgia thyrsiflora for Galerucella grisescens.     

Molecular phylogeny of rootworms and related galerucine beetles (Coleoptera: Chrysomelidae)

The Galerucinae (Coleoptera: Chrysomelidae) sensu stricto (true galerucines) comprise a large assemblage of diverse phytophagous beetles containing over 5000 described species. Together with their sister taxon, the flea beetles, which differ from true galerucines by having the hind femora usually modified for jumping, the Galerucinae sensu lato comprises over 13 000 described species and is the largest natural group within the Chrysomelidae. Unlike the flea beetles, for which robust hierarchical classification schemes have not been erected, an existing taxonomic structure exists for the true galerucines, based mostly on the works of the late John Wilcox. In the most recent taxonomic list of the Galerucinae sensu stricto, five tribes were established comprising 29 sections housing 488 genera. The majority of the diversity within these tribes is found within the tribe Luperini, in which two genera, Monolepta and Diabrotica, are known to contain over 500 described species. Here, we extend the work from previous phylogenetic studies of the Galerucinae by analysing four amplicons from three gene regions (18S and 28S rRNA; COI) representing 249 taxa, providing the largest phylogenetic analysis of this taxon to date. Using two seven‐state RNA models, we combine five maximum likelihood models (RNA + DNA for the rRNAs; three separate DNA models for the COI codon positions) for these partitions and analyse the data under likelihood using Bayesian inference. The results of these two analyses are compared with those from equally weighted parsimony. Instead of choosing the results from one optimality criterion over another, either based on statistical support, tree topology or philosophical predisposition, we elect to draw attention to the similar results produced by all three analyses, illustrating the robustness of the data to these different analytical methods. In general, the results from all three analyses are consistent with each other and previous molecular phylogenetic reconstructions for Galerucinae, except that increased taxon sampling for several groups, namely the tribes Hylaspini and Oidini, has improved the phylogenetic position of these taxa. As with previous analyses, under‐sampled taxa, such as the Old World Metacyclini and all sections of the subtribe Luperina, continue to be unstable, with the few taxa representing these groups fluctuating in their positions based on the implemented optimality criterion. Nonetheless, we report here the most comprehensive phylogenetic estimation for the Galerucinae to date.     

Anthophagy in the leaf beetles (Coleoptera,Chrysomelidae)

Laboratory experiments were performed on leaf beetles collected on flowers. Flower consumption (anthophagy) was demonstrated for adults of Donacia bicolora, D. brevitarsis, D. obscura, D. thalassina, Plateumaris discolor, Cryptocephalus laetus, C. sericeus, C. solivagus, Labidostomis longimana, Hydrothassa marginella, Phaedon concinnus, Galerucella nymphaeae, Neocrepidodera femorata, Altica oleracea, Aphthona lutescens, Longitarsus pellucidus, and larvae of Entomoscelis adonidis, H. marginella, and G. nymphaeae for the first time for all the species except for P. discolor, L. longimana, and C. sericeus. The feeding behavior was described, and the mouthparts of adults were studied. The mandibles and maxillae of the Donacia, Plateumaris, and Orsodacne species mentioned are adapted to consuming pollen. Some questions of ecology and evolution of anthophagy are discussed. An overview of literature on anthophagy in the leaf beetles is presented.     

Comparative analysis of microbial diversity in Longitarsus flea beetles (Coleoptera: Chrysomelidae)

Herbivorous beetles comprise a significant fraction of eukaryotic biodiversity and their plant-feeding adaptations make them notorious agricultural pests. Despite more than a century of research on their ecology and evolution, we know little about the diversity and function of their symbiotic microbial communities. Recent culture-independent molecular studies have shown that insects possess diverse gut microbial communities that appear critical for their survival. In this study, we combined culture-independent methods and high-throughput sequencing strategies to perform a comparative analysis of Longitarsus flea-beetles microbial community diversity (MCD). This genus of beetle herbivores contains host plant specialists and generalists that feed on a diverse array of toxic plants. Using a deep-sequencing approach, we characterized the MCD of eleven Longitarsus species across the genus, several of which represented independent shifts to the same host plant families. Database comparisons found that Longitarsus-associated microbes came from two habitat types: insect guts and the soil rhizosphere. Statistical clustering of the Longitarsus microbial communities found little correlation with the beetle phylogeny, and uncovered discrepancies between bacterial communities extracted directly from beetles and those from frass. A Principal Coordinates Analysis also found some correspondence between beetle MCD and host plant family. Collectively, our data suggest that environmental factors play a dominant role in shaping Longitarsus MCD and that the root-feeding beetle larvae of these insects are inoculated by soil rhizosphere microbes. Future studies will investigate MCD of select Longitarsus species across their geographic ranges and explore the connection between the soil rhizosphere and the beetle MCD.     

Toxicity toChrysomela tremulae (Coleoptera: Chrysomelidae) of transgenic poplars expressing a cysteine proteinase inhibitor

The aim of this study was to test the potential of proteinase inhibitors to controlChrysomela tremulae, a beetle that causes severe damage in young plantations and in short-rotation intensive culture (SRIC) of poplar. As a first step, cysteine proteinases were determined to be the major digestive proteinases ofC. tremulae and oryzacystatin OCI, a cysteine proteinase inhibitor, was shown to inhibit this activityin vitro. The gene encoding OCI was introduced into poplar (Populus tremula ×P. tremuloides) and transgenic plants expressing OCI at a high level were selected. Feeding tests on these transgenic plants demonstrate the toxicity of OCI-producing poplar leaves againstC. tremulae larvae.J.C. Leplé and M. Bonadé-Bottino contributed equally to the research presented in this paper.     

Genetics of resistance to transgenic Bacillus thuringiensis poplars in Chrysomela tremulae (Coleoptera: Chrysomelidae)

The area under genetically engineered plants producing Bacillus thuringiensis (Bt) toxins is steadily increasing. This increase has magnified the risk of alleles conferring resistance to these toxins being selected in natural populations of target insect pests. The speed at which this selection is likely to occur depends on the genetic characteristics of Bt resistance. We selected a strain of the beetle Chrysomela tremulae Fabricius on a transgenic Bt poplar clone Populus tremula L. x Populus tremuloides Michx producing high levels of B. thuringiensis Cry3Aa toxin. This strain was derived from an isofemale line that generated some F2 offspring that actively fed on this Bt poplar clone. The resistance ratio of the strain was >6400. Susceptibility had decreased to such an extent that the mortality of beetles of the strain fed Bt poplar leaves was similar to that of beetles fed nontransgenic poplar leaves. Genetic crosses between susceptible, resistant, and F1 hybrids showed that resistance to the Cry3Aa toxin was almost completely recessive (D(LC) = 0.07) and conferred by a single autosomal gene. The concentration of Cry3Aa produced in the transgenic Bt poplar used in this study was 6.34 times higher than the LC99 of the F1 hybrids, accounting for the complete recessivity (D(ML) = 0) of survival on Bt poplar leaves. Overall, the genetic characteristics of the resistance of C. tremulae to the Cry3Aa toxin are consistent with the assumptions underlying the high-dose refuge strategy, which aims to decrease the selection of Bt resistance alleles in natural target pest populations.     

Effect of a phloxine B-cucurbitacin bait on diabroticite beetles (Coleoptera: Chrysomelidae)

Cucurbitacin E glycoside, extracted from a bitter mutant of Hawkesbury watermelon [Citrulls lanatus (Thunb.) Matsum. & Nakai (Syn. Citrullus vulgaris Schrad)] is the active ingredient of a feeding stimulant for the corn rootworm complex. It is the primary component of a water-soluble bait that can be combined with toxins for adult diabroticite control. Studies were conducted using phloxine B (D&C Red 28), a xanthene dye, as the toxin. This dye was efficacious against Diabrotica undecimpunctata howardi Barber, spotted cucumber beetle, and Acalymma vittatum (F.), striped cucumber beetle, in cucumber plots and could be recovered from cucumber leaves for 8 d after treatment. The average amount of dye recovered per dead spotted cucumber beetle at 8 d after treatment was 0.173 microg. Concentrated and sugar-free fermented forms of the watermelon extract were developed and compared with the fresh juice in field applications on cucumber plants. There was no significant difference in mortality of beetles from phloxine B-bait prepared with fresh, fermented, or concentrated extract, although in laboratory studies, fermented juice had higher feeding stimulant activity.     

Optomotor reaction and fixation of walking colorado beetles (Coleoptera: Chrysomelidae): Course control aspects

The paths of Colorado beetles (Leptinotarsa decemlineata Say)in a featureless environment are circular. This behavior is explained by an internal asymmetry. To stabilize the path, the fixation reaction or the optomotor response must work against this asymmetry. The turning behavior was examined in stationary patterns of vertical stripes different at spatial wavelengths (). The internal asymmetry was tested in a horizontally striped pattern. A stable fixation reaction was found only for 120 °. The results suggest that larger intrinsic turning tendencies shifts the stable point of the fixation reaction. The same vertically striped patterns were rotated to examine the following reaction of the beetle. It is concluded that the fixation component of the response of these insects, in particular, does not differ in the two situations.     

New species of the leaf beetles (Coleoptera,Chrysomelidae) from China: VII

The following new taxa of leaf beetles are described from Chinese provinces Gansu, Sichuan, and Xinjiang based on the material of the Zoological Institute, Russian Academy of Sciences, St. Petersburg, where the types are deposited: Chrysolina confucii sp. n., Ch. daosana sp. n., Ch. infernalis sp. n., Ch. gruevi sp. n., Ch. songpana sp. n., Phratora alternata sp. n., Taipinus rotundatus gen. et sp. n., Oreomela milae sp. n. (Chrysomelinae) and Zangastra sichuanica sp. n. (Galerucinae).     

Efficiency of flight interception traps for adult Colorado potato beetles (Coleoptera: Chrysomelidae)

Interception traps have been used to monitor and sample Colorado potato beetle, Leptinotarsa decemlineata (Say), populations in flight, but the efficiency of these traps was unknown. The efficiency of interception window traps is limited because flying adults avoid the traps and may bounce off the trap without being collected. All trap types tested were avoided in flight chamber tests, including those constructed of transparent Plexiglas or yellow wood boards. A larger screen trap and a harp trap designed to reduce detection were also avoided by the beetles in flight. None of the traps provide a direct estimate of the number of adults randomly flying in the trap area. The highest level of efficiency for window traps was obtained with the yellow trap, which caught 16% of the expected flying population. Harp and screen traps without frames intercepted 60-62% of the expected flying population. The presence of a frame did not increase significantly the avoidance of the screen and harp traps by beetles in flight. Recommendations are made to maintain or increase the efficiency of some traps.     

On the laboratory rearing of green dock leaf beetles Gastrophysa viridula (Coleoptera: Chrysomelidae)

Abstract Leaf beetles Gastrophysa viridula have attracted recently increased research interest from various points of view, since they are: (i) pest insects in rhubarb crops; (ii) potential biocontrol agents of dock plants Rumex spp. in grasslands; and (iii) a model species in ecological studies on insect population dynamics, biochemistry, behavior, biomechanics and biomimetics. The continuous rearing of beetles at standardized conditions may help to unify the fitness state of different individuals, allowing a better comparison of experimental results. The present communication suggests a modular space‐ and time‐saving rearing method of G. viridula in stackable faunariums under laboratory conditions, which has been successfully established and continuously used over the last 5 years. Several developmental stages were kept in separate boxes, and multiple generations were kept simultaneously, depending on the required number of beetles.     

Inbreeding depression in two seed-feeding beetles, Callosobruchus maculatus and Stator limbatus (Coleoptera: Chrysomelidae)

Inbreeding depression is well documented in insects but the degree to which inbreeding depression varies among populations within species, and among traits within populations, is poorly studied in insects other than Drosophila. Inbreeding depression was examined in two long-term laboratory colonies of the seed beetle, Callosobruchus maculatus (Fabricius), which are used frequently as models for experiments in ecology, evolution and behaviour. Inbreeding depression in these laboratory colonies are compared with one recently field-collected population of a different seed beetle, Stator limbatus Horn. Inbreeding reduced embryogenesis, egg hatch and larval survival in both species, such that eggs produced by sib matings were >17% less likely to produce an adult offspring. Inbred larvae also took 4-6% longer to develop to emergence in both species. Inbreeding depression varied among the measured traits but did not differ between the two populations of C. maculatus for any trait, despite the large geographic distance between source populations (western Africa vs. southern India). Inbreeding depression was similar in magnitude between C. maculatus and S. limbatus. This study demonstrates that these laboratory populations of C. maculatus harbour substantial genetic loads, similar to the genetic load of populations of S. limbatus recently collected from the field.     

Contributions to the knowledge of the leaf beetles (Coleoptera, Chrysomelidae) of Russia

A new species, Macroplea skomorokhovi sp. n., is described from the Russian Far East, and a key to species of the genus Macroplea Curtis is given. Cryptocephalus oxysternus Jacobson (Primorskii Territory) and Longitarsus truncatellus Weise (northern Caucasus) are recorded for the Russian fauna for the first time.     

Leaf beetles are ant-nest beetles: the curious life of the juvenile stages of case-bearers (Coleoptera,Chrysomelidae, Cryptocephalinae)

Although some species of Cryptocephalinae (Coleoptera: Chrysomelidae) have been documented with ants (Hymenoptera: Formicidae) for almost 200 years, information on this association is fragmentary. This contribution synthesizes extant literature and analysizes the data for biological patterns. Myrmecophily is more common in the tribe Clytrini than in Cryptocephalini, but not documented for Fulcidacini or the closely-related Lamprosomatinae. Myrmecophilous cryptocephalines (34 species in 14 genera) primarily live among formicine and myrmecines ants as hosts. These two ant lineages are putative sister-groups, with their root-node dated to between 77–90 mya. In the New World tropics, the relatively recent radiation of ants from moist forests to more xeric ecosystems might have propelled the association of cryptocephalines and ant nests. Literature records suggest that the defensive behavioral profile or chemical profile (or both) of these ants has been exploited by cryptocephalines. Another pattern appears to be that specialized natural enemies, especially parasitoid Hymenoptera, exploit cryptocephaline beetles inside the ant nests. With the extant data at hand, based on the minimum age of a fossil larva dated to 45 mya, we can infer that the origin of cryptocephaline myrmecophily could have arisen within the Upper Cretaceous or later. It remains unknown how many times myrmecophily has appeared, or how old is the behavior. This uncertainty is compounded by incongruent hypotheses about the origins of Chrysomelidae and angiosperm-associated lineages of cryptocephalines. Living with ants offers multiple advantages that might have aided the colonization of xeric environments by some cryptocephaline species.     

An alternative sampling technique for cucumber beetles (Coleoptera: Chrysomelidae) and diurnal beetle activity on muskmelon

Field studies were conducted to investigate the effectiveness of yellow sticky traps as an alternative sampling technique for striped, Acalymma vittatum (F.), and spotted, Diabrotica undecimpunctata howardi Barber, cucumber beetles (Coleoptera: Chrysomelidae) and the diurnal beetle activity on muskmelon, Cucumis melo L., near Vincennes, IN, in 2003 and 2004. The experimental design included six replications of seven 20-m-long rows each of muskmelon with 1.5 m between rows. On each sampling date, two yellow sticky traps were placed randomly between rows in each replication. One sticky trap was placed vertically with the lower edge even with the top of the canopy, whereas the other trap was placed horizontally, even with the top of the canopy. After traps were placed in the field, number of beetles on plants was counted in situ from 0800 to 1600 hours at 2-h intervals the next day. After 48 h in the field, the number of cucumber beetles adhering on traps was counted. Analyses of variance and Tukey's multiple comparison procedure were used to compare the densities of beetles among sampling times, and regression analyses were applied to correlate the numbers of beetles on traps and the numbers of in situ counts. Results show that both species of cucumber beetles were most active from 1200 to 1400 hours, and 20 beetles on the vertically positioned sticky trap were equivalent to one beetle per plant in the field. The application of the sampling technique and scouting time for cucumber beetle management are discussed.     

A new genus of seed beetles (Coleoptera,Chrysomelidae, Bruchinae) in Baltic amber

The new genus and new species of seed beetles (Copeoptera, Chrysomelidae, Bruchinae) Electrocaryedon poinari gen. et sp. nov. are described from the Late Eocene Baltic amber. This new genus differs from the genus Caryedon Schoenherr, 1823 in the completely separate procoxal cavities, not shortened lateral carina on pronotum, and wider body with sparse setae.     

Cytological studies on three species of neotropical cassidines (Coleoptera,Chrysomelidae)

Chromosome numbers, meiosis and sex-determining mechanisms of three species of Chrysomelidae (Coleoptera) were studied. They belong to the tribe Stolaini of Neotropical Cassidinae. Stolas lacordairei and Botanochara sp. have high chromosome numbers (2n=30♂), the first with a primitive coleopteran sex chromosome system Xy _p and the other with a very complex one X _p ¹ X _p ² neo Xneo Y. Botanochara bonariensis shows an even higher chromosome number (2n=44♂) and multiple sex chromosomes as does B. sp. The possible tendencies in the karyological evolution of the Stolaine Cassidines are discussed.