Staphylococcus,paramyxovirus-like,rickettsia-like and other structures in Peregrinus maidis (Homoptera,delphacidae)

In an ultrastructural study of the delphacid planthopper, Peregrinus maidis, vector of maize mosaic virus (MMV) and maize stripe virus (MStpV), the following structures were found in several organs of MMV-inoculative, MStpV-inoculative, and noninoculative insects: (a) paramyxovirus-like particles (PLP), (b) filamentous rhabdo-like structures (FRS), (c) spiked double-membrane structures (SDS) that were always associated, and sometimes contiguous, with FRS, and (d) rickettsia-like structures (RLS). In salivary gland acini, the PLP, FRS, and SDS were usually found in extracellular spaces between basal infoldings of plasma membranes, whereas the RLS were intracytoplasmic or intranuclear. The PLP and FRS appeared to bud through plasma membranes. The colony of P. maidis in which these structures were found suffered from retarded development and premature death. Limited trials to isolate a paramyxovirus from this colony were unsuccessful. However, further trials led to the isolation of Staphylococcus sciuri, which proved pathogenic when injected into P. maidis and two cicadellid leafhoppers; Dalbulus maidis and Graminella nigrifrons.     

Inoculation and acquisition of maize bushy stunt mycoplasma by its leafhopper vector Dalbulus maidis

Maize bushy stunt mycoplasma (MBSM), a mycoplasma-like organism, is transmitted in a persistent manner by the corn leafhopper, Dalbulus maidis, to maize (Zea mays). The influence of the duration of acquisition access and inoculation access periods on the transmission of MBSM by D. maidis was investigated. The proportion of plants infected by D. maidis increased significantly from 0 to 0.51 as the inoculation access time to a plant increased from 10 min to 72 h (X²= 101.5, P < 0.001). Likewise, the proportion of insects acquiring MBSM from infected plants increased from 0 to 0.19 as the acquisition access time to the source plant increased from 10 min to 72 h (X²= 53.2, P < 0.001). The data were fitted to a loglinear regression model. No significant association was found between the sex of the insects and vector ability.     

Transmission of rayado fino virus of maize (Zea mays) by Dalbulus tnaidis

Rayado fino virus (RFV) of maize (Zea mays) was transmitted by the leaf-hopper Dalbulus maidis in a manner characteristic of viruses that multiply in their insect vectors. Individual insects fed on infected plants transmitted the virus after incubation periods of 8–22 days; males had shorter incubation periods than females but died sooner. Insects retained infectivity for 1–20 days. Transmission by most insects was intermittent. Inoculativity by D. maidis decreased with time, but the virus was recovered from insects that had lost their ability to transmit. Extracts of plants infected with RFV and viruliferous insects were injected into healthy insects, which became viruli-ferous. Infectivity of the extracts was not affected by tetracycline hydrochloride (Achromycin). D. maidis was able to transmit simultaneously RFV and the corn stunt agent. Other than maize, Teosinte (Euchlaena mexicana) was the only plant susceptible to the virus, among a number of species of Gramineae tested.     

Inside the guts of wood-eating catfishes: can they digest wood?

To better understand the structure and function of the gastrointestinal (GI) tracts of wood-eating catfishes, the gross morphology, length, and microvilli surface area (MVSA) of the intestines of wild-caught Panaque nocturnus, P. cf. nigrolineatus “Marañon”, and Hypostomus pyrineusi were measured, and contrasted against these same metrics of a closely related detritivore, Pterygoplichthys disjunctivus. All four species had anatomically unspecialized intestines with no kinks, valves, or ceca of any kind. The wood-eating catfishes had body size-corrected intestinal lengths that were 35% shorter than the detritivore. The MVSA of all four species decreased distally in the intestine, indicating that nutrient absorption preferentially takes place in the proximal and mid-intestine, consistent with digestive enzyme activity and luminal carbohydrate profiles for these same species. Wild-caught Pt. disjunctivus, and P. nigrolineatus obtained via the aquarium trade, poorly digested wood cellulose (<33% digestibility) in laboratory feeding trials, lost weight when consuming wood, and passed stained wood through their digestive tracts in less than 4 h. Furthermore, no selective retention of small particles was observed in either species in any region of the gut. Collectively, these results corroborate digestive enzyme activity profiles and gastrointestinal fermentation levels in the fishes’ GI tracts, suggesting that the wood-eating catfishes are not true xylivores such as beavers and termites, but rather, are detritivores like so many other fishes from the family Loricariidae.     

“Endomicrobia” and Other Bacteria Associated with the Hindgut of Dermolepida albohirtum Larvae

Symbiotic bacteria residing in the hindgut chambers of scarab beetle larvae may be useful in paratransgenic approaches to reduce larval root-feeding activities on agricultural crops. We compared the bacterial community profiles associated with the hindgut walls of individual Dermolepida albohirtum third-instar larvae over 2 years and those associated with their plant root food source among different geographic regions. Denaturing gradient gel electrophoresis analysis was used with universal and Actinobacteria-specific 16S rRNA primers to reveal a number of taxa that were found consistently in all D. albohirtum larvae but not in samples from their food source, sugarcane roots. These taxa included representatives from the “Endomicrobia,” Firmicutes, Proteobacteria, and Actinobacteria and were related to previously described bacteria from the intestines of other scarab larvae and termites. These universally distributed taxa have the potential to form vertically transmitted symbiotic associations with these insects.     

Bacillus thuringiensis isolates from Korean forest environments

We investigated the distribution, toxicity, morphology, and protein profiles of Bacillus thuringiensis isolates from forests in Korea to isolate naturally occurring novel B. thuringiensis. A total of 170 B. thuringiensis isolates were obtained from 832 samples producing spore and parasporal inclusion bodies. In toxicity tests for lepidopteran, dipteran, and coleopteran insects, 57.6% isolates were toxic only to Lepidoptera, 5.3% were toxic only to Diptera, and 24.1% were toxic to both Diptera and Lepidoptera. The remaining collections (13.0%) were not toxic to the tested insects. The shapes of the parasporal crystals produced in B. thuringiensis isolates were bipyramidal, spherical, ovoid, or irregular. As their toxicities varied with parasporal crystal shape, B. thuringiensis isolates possessing bipyramidal or irregular parasporal crystals were largely toxic to lepidopteran species whereas those producing spherical parasporal crystals were mainly toxic to dipteran species. B. thuringiensis toxic to both dipteran and lepidopteran insects contained 130- and 70-kDa parasporal crystals, whereas B. thuringiensis toxic to lepidopteran insects expressed 130-kDa parasporal crystals. The results suggest that forest areas in Korea are a rich source of B. thuringiensis and need to be further explored to discover novel B. thuringiensis isolates.     

Praecocilenchus rhaphidophorus n. gen., n. sp. (Nematoda: Aphelenchoidea) Parasitizing Rhynchophorus bilineatus (Montrouzier) (Coleoptera: Curculionidae) in New Britain

Praecocilenchus rhaphidophorus n. gen., n. sp. is described as a new endoparasitic aphelenchoid nematode parasitizing adults of the palm weevil, Rhynchophorus bilineatus (Montrouzier). P. rhaphidophorus is unusual in that juveniles develop to maturity within the female uterus and thin, needle-shaped crystals form in the intestines of mature parasitic females. Hundreds of parasitic female nematodes were found in the body cavity of infected hosts. The role of this parasite as a biological control agent of Rhynchophorus weevils is discussed.     

Within‐field spatial distribution patterns of corn planthopper,Peregrinus maidis,and severity of hopperburn and Maize mosaic virus symptoms as influenced by sunn hemp intercropping

Habitat management (e.g., intercropping) may alter within‐field spatial distribution patterns of herbivores, from a typical pattern as observed in a monoculture, and may influence patterns of crop injury. Field trials were conducted to study the effect of intercropping maize, Zea mays L. (Poaceae), with sunn hemp, Crotalaria juncea L. (Fabaceae) strips on within‐field spatial distribution patterns of corn planthopper, Peregrinus maidis (Ashmead) (Hemiptera: Delphacidae), and combined severity of hopperburn and Maize mosaic virus (MMV) (Rhabdoviridae: Nucleorhabdovirus) symptoms. In each field trial, spatially explicit data on P. maidis counts and ratings of severity of symptoms were obtained by sampling maize plants at weekly intervals. These data were used to examine the spatial patterns of P. maidis and severity of symptoms in maize‐intercropped and monoculture plots with Spatial Analysis for Distance IndicEs (SADIE) methodology. Spatial aggregation patterns of P. maidis in each treatment plot were not consistent among the field trials and tended to be mediated by their population densities. Interpolation of local cluster indices showed that P. maidis were more often aggregated at the field edges, irrespective of treatment. At times of MMV incidence in field trials (fall 2010 and spring 2011), the patch clusters of P. maidis and symptomatic plants were located at the field edges, but were spatially unassociated in both treatment plots. The results provided an approximation of the unpredictability of P. maidis spatial patterns at different population densities and their association with severity of symptoms in two maize‐cropping systems. However, the gap clusters of symptomatic plants were primarily located at the field interiors and were larger in intercropped than in monoculture plots. Such spatial pattern of symptomatic plants resulted in the reduced incidence of MMV in the intercropped plot compared with the monoculture plot, suggesting intercropping sunn hemp can be a useful tool in the management of MMV in maize fields.     

Pathogenicity of fungal isolates (Ascomycota: Hypocreales) against Peregrinus maidis, Delphacodes kuscheli (Hemiptera: Delphacidae), and Dalbulus maidis (Hemiptera: Cicadellidae), vectors of corn diseases

Preliminary screening assays were carried out on 17 isolates from five fungal species Beauveria bassiana, Lecanicillium muscarium, Metarhizium anisopliae, Isaria farinosa, and I. fumosorosea. The three most effective isolates against Peregrinus maidis (Hemiptera: Delphacidae) were B. bassiana CEP 147, CEP 150, and CEP 189. There were no consistent differences found in males and females regarding fungal susceptibility. However, more females than males were proportionally infected. There was not a correlation between the percentage of conidial germination and the percentage of mortality caused by fungal infection in any of the treatments. Only B. bassiana CEP 147, which caused a cumulative mortality of 69.8 ± 6.4% after 7 days post-inoculation, was selected to be assayed against adults of P. maidis, Delphacodes kuscheli (Hemiptera: Delphacidae), and Dalbulus maidis (Hemiptera: Cicadellidae). In pathogenicity tests significant differences were observed among treatments. After 2 weeks post-inoculation, both D. kuscheli (cumulative mortality of 73.3 ± 9.0%) and P. maidis (cumulative mortality of 68.6 ± 6.7%) were significantly more susceptible than D. maidis (cumulative mortality of 49.9 ± 9.7%) to the selected isolate.     

Dispersal distances of the Caribbean fruit fly,corn planthopper and Cuban May beetle

Summary Adults of the Caribbean fruit fly,Anastrepha suspensa (Loew), and Cuban May beetle,Phyllophaga bruneri Chapin, were recovered at various distances from release sites. Corn planthopper,Peregrinus maidis (Ashmead), adults were numerous and more dead corn plants were found beside the grassy areas indicating that incidence of the disease was related to the insect abundance. Regression curves showed that rates of dispersion of insects or incidence of dead corn plants were related to distance.     

Solubilization, Activation, and Insecticidal Activity of Bacillus thuringiensis Serovar thompsoni HD542 Crystal Proteins

Cry15Aa protein, produced by Bacillus thuringiensis serovar thompsoni HD542 in a crystal together with a 40-kDa accompanying protein, is one of a small group of nontypical, less well-studied members of the Cry family of insecticidal proteins and may provide an alternative for the more commonly used Cry proteins in insect pest management. In this paper, we describe the characterization of the Cry15Aa and 40-kDa protein's biochemical and insecticidal properties and the mode of action. Both proteins were solubilized above pH 10 in vitro. Incubation of solubilized crystal proteins with trypsin or insect midgut extracts rapidly processed the 40-kDa protein to fragments too small to be detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, whereas the Cry15 protein yielded a stable product of approximately 30 kDa. Protein N-terminal sequencing showed that Cry15 processing occurs exclusively at the C-terminal end. Cry15 protein showed in vitro hemolytic activity, which was greatly enhanced by preincubation with trypsin or insect gut extract. Larvae of the lepidopteran insects Manduca sexta, Cydia pomonella, and Pieris rapae were susceptible to crystals, and presolubilization of the crystals enhanced activity to P. rapae. Activity for all three species was enhanced by preincubation with trypsin. Larvae of Helicoverpa armigera and Spodoptera exigua were relatively insensitive to crystals, and activity against these insects was not enhanced by prior solubilization or trypsin treatment. The 40-kDa crystal protein showed no activity in the insects tested, nor did its addition or coexpression in Escherichia coli increase the activity of Cry15 in insecticidal and hemolytic assays.     

Cloning of the spoT Gene of “Candidatus Phlomobacter fragariae” and Development of a PCR-Restriction Fragment Length Polymorphism Assay for Detection of the Bacterium in Insects

Marginal chlorosis is a new disease of strawberry in which the uncultured phloem-restricted proteobacterium “Candidatus Phlomobacter fragariae” is involved. In order to identify the insect(s) vector(s) of this bacterium, homopteran insects have been captured. Because a PCR test based on the 16S rRNA gene (rDNA) applied to these insects was unable to discriminate between “P. fragariae” and other insect-associated proteobacteria, isolation of “P. fragariae” genes other than 16S rDNA was undertaken. Using comparative randomly amplified polymorphic DNAs, an amplicon was specifically amplified from “P. fragariae”-infected strawberry plants. It encodes part of a “P. fragariae” open reading frame sharing appreciable homology with the spoT gene from other proteobacteria. A spoT-based PCR test combined with restriction fragment length polymorphisms was developed and was able to distinguish “P. fragariae” from other insect bacteria. None of the many leafhoppers and psyllids captured during several years in and around infected strawberry fields was found to carry “P. fragariae.” Interestingly however, the “P. fragariae” spoT sequence could be easily detected in whiteflies proliferating on “P. fragariae”-infected strawberry plants under confined greenhouse conditions but not on control whiteflies, indicating that these insects can become infected with the bacterium.     

Nuclear ribosomal internal transcribed spacer 1 (ITS1) variation in the Anastrepha fraterculus cryptic species complex (Diptera,Tephritidae) of the Andean region

The nuclear ribosomal internal transcribed spacer 1 (ITS1) was sequenced for Anastrepha fraterculus (Wiedemann, 1830) originating from 85 collections from the northern and central Andean countries of South America including Argentina (Tucumán), Bolivia, Perú, Ecuador, Colombia, and Venezuela. The ITS1 regions of additional specimens (17 collections) from Central America (México, Guatemala, Costa Rica, and Panamá), Brazil, Caribbean Colombia, and coastal Venezuela were sequenced and together with published sequences (Paraguay) provided context for interpretation. A total of six ITS1 sequence variants were recognized in the Andean region comprising four groups. Type I predominates in the southernmost range of Anastrepha fraterculus. Type II predominates in its northernmost range. In the central and northern Andes, the geographic distributions overlap and interdigitate with a strong elevational effect. A discussion of relationships between observed ITS1 types and morphometric types is included.     

Multiplication and persistence of Acholeplasma spp. in leafhoppers

Strains of Acholeplama spp., originally isolated from coconut palms, varied in their ability to multiply or persist in vivo following injection into the leafhoppers Dalbulus maidis and Euscelidius variegatus (Homoptera: Cicadellidae). Survival of the organisms in vivo was not related to their biochemical or serological properties and was a stable characteristic retained by clones that had been serially passaged in vitro. Tests with E. variegatus showed that multiplication of some of the isolates was dependent on the dose of organisms injected and suggested that growth occurred in the hemolymph where antibacterial proteins may play a part in survival of certain strains at low dose levels. This work confirms that Acholeplasma sp. have the ability to survive and even multiply in varied habitats, and under appropriate conditions, insects could play a role in their dissemination.     

Role of Proteolysis in Determining Potency of Bacillus thuringiensis Cry1Ac δ-Endotoxin

Bacillus thuringiensis protein δ-endotoxins are toxic to a variety of different insect species. Larvicidal potency depends on the completion of a number of steps in the mode of action of the toxin. Here, we investigated the role of proteolytic processing in determining the potency of the B. thuringiensis Cry1Ac δ-endotoxin towards Pieris brassicae (family: Pieridae) and Mamestra brassicae (family: Noctuidae). In bioassays, Cry1Ac was over 2,000 times more active against P. brassicae than against M. brassicae larvae. Using gut juice purified from both insects, we processed Cry1Ac to soluble forms that had the same N terminus and the same apparent molecular weight. However, extended proteolysis of Cry1Ac in vitro with proteases from both insects resulted in the formation of an insoluble aggregate. With proteases from P. brassicae, the Cry1Ac-susceptible insect, Cry1Ac was processed to an insoluble product with a molecular mass of ~56 kDa, whereas proteases from M. brassicae, the non-susceptible insect, generated products with molecular masses of ~58, ~40, and ~20 kDa. N-terminal sequencing of the insoluble products revealed that both insects cleaved Cry1Ac within domain I, but M. brassicae proteases also cleaved the toxin at Arg423 in domain II. A similar pattern of processing was observed in vivo. When Arg423 was replaced with Gln or Ser, the resulting mutant toxins resisted degradation by M. brassicae proteases. However, this mutation had little effect on toxicity to M. brassicae. Differential processing of membrane-bound Cry1Ac was also observed in qualitative binding experiments performed with brush border membrane vesicles from the two insects and in midguts isolated from toxin-treated insects.     

ELECTRON MICROSCOPIC STUDY OF THE CYTOPATHOLOGY OF ECHO VIRUS INFECTION IN CULTIVATED CELLS

The cultivated monkey kidney cell is subject to changes when infected with ECHO viruses 6, 9, and 19. The electron microscope reveals three stages of infection: (a) initial stage. The nucleus appears granular with chromatin condensation on the nuclear envelope. The cytoplasm contains electron transparent vesicles and vacuoles forming nests. (b) Intermediate stage. The nucleus seems to diminish, appearing more pycnotic and displaced toward the periphery. The cytoplasm is filled with electron transparent vacuoles and vesicles, and dense masses as well as some spiral bodies are seen. The mitochondria retain their shape. Dense particles are seen, which are possibly of viral nature. (c) Final stage. The nucleus is contracted to a narrow strip close to the cellular membrane or is completely destroyed. The cytoplasm shows no apparent changes. Crystals are frequently observed in cells infected with ECHO viruses 6 and 19, consisting of dense particles with an average diameter of 14.4 mµ ranging from approximately 13.2 to 15.6 mµ for ECHO virus 6, and 14.5 mµ ranging from approximately 12.5 to 16.5 mµ for ECHO virus 19. These particles are clustered in hexagonal packages forming angles of 75° and 105°. The particles in most crystals are arranged in rows separated by a constant distance, the latter varying from one crystal to another and being approximately 1.5 and 2.5 times the distance between particles. Other particles were observed which, however, are not considered to be of viral nature.     

Host plant interactions of the corn planthopper, Peregrinus maidis Ashm. (Homoptera: Delphacidae) in maize and sorghum agroecosystems

The corn planthopper, Peregrinus maidis (Ashmead) (Homoptera: Delphacidae) causes serious economic losses in corn and sorghum. The insect occurs mostly at humid low elevations in the tropics and coastal areas of subtropical and temperate regions of all continents, the Caribbean Islands, and islands in the Atlantic, Indian, and Pacific Oceans. This review provides a detailed compilation on the chronological progress made in basic and strategic aspects of research on the interactions between P. maidis and various host plants. The nature of damage by P. maidis and its economic impact, ecobiology in relation to host diversity, abiotic, and seasonal interactions; and life tables and alary polymorphism are discussed. Host plant resistance studies indicate that very few sources of resistance to P. maidis have been identified in maize, sorghum, or pearl millet, warranting a need to standardize rapid and reliable screening methods. The behavioral responses vis-à-vis mechanisms of resistance show the predominance of antixenosis for colonization and/or oviposition with variable degrees of antibiosis affecting life cycle parameters of P. maidis on maize and sorghum. The role of morphological traits, physiological mechanisms, and biochemical factors governing resistance are described. Population dynamics based on density-dependent and density-independent interactions are also discussed. In addition, aspects of P. maidis on chemical control, biological control, and trophobiosis interactions are listed. Future thrusts on research approaches are also discussed. Genetic engineering techniques involving lectin genes in the development of transgenic plants, and the molecular mapping of genes conferring resistance to both P. maidis and its transmitted virus diseases may stimulate further research and lead to better understanding of P. maidis—host plant interactions. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Cry1Ab protein levels in phytophagous insects feeding on transgenic corn: implications for secondary exposure risk assessment

Enzyme-linked immunosorbent assays (ELISA) and bioassays were used to estimate levels of Cry1Ab protein in four species of phytophagous insects after feeding on transgenic Bt-corn plants expressing Cry1Ab protein or artificial diets containing Cry1Ab protein. The level of Cry1Ab in insects feeding on sources containing the Cry1Ab protein was uniformly low but varied with insect species as well as food source. For the corn leaf aphid, Rhopalosiphum maidis (Fitch), feeding on diet solutions containing Cry1Ab protein, the level of the protein in the aphid was 250–500 times less than the original levels in the diet, whereas no Cry1Ab was detected by ELISA in aphids feeding on transgenic Bt-Corn plants. For the lepidopteran insects, Ostrinia nubilalis (Hübner), Helicoverpa zea (Boddie), and Agrotis ipsilon (Hufnagel), levels of Cry1Ab in larvae varied significantly with feeding treatment. When feeding for 24 h on artificial diets containing 20 and 100 ppm of Cry1Ab, the level of Cry1Ab in the larvae was about 57 and 142 times lower, respectively, than the original protein level in the diet for O. nubilalis, 20 and 34 times lower for H. zea, and 10 to 14 times lower for A. ipsilon. Diet incorporation bioassays with a susceptible insect (first instar O. nubilalis) showed significant Cry1Ab bioactivity present within whole body tissues of R. maidis and O. nubilalis that had fed on diet containing a minimum of 20 ppm or higher concentrations (100 or 200 ppm) of Cry1Ab, but no significant bioactivity within the tissues of these insects after feeding on transgenic Bt-corn plants. The relevance of these findings to secondary exposure risk assessment for transgenic Bt crops is discussed.     

The saurian malarias of Venezuela: Haemosporidian parasites of gekkonid lizards

Telford S. P., Jr. 1978. The saurian malarias of Venezuela: haemosporidian parasites of gekkonid lizards. International Journal for Parasitology8: 341–353. Five haemosporidian species were found among 185 gekkonid lizards from Estados Portuguesa, Cojedes and Aragua, Venezuela, four of which were new to science. A pigmented Plasmodium species is described from Gonatodes taniae of Estado Aragua. It produces 8–20 merozoites in variably shaped schizonts, and elongate, irregularly margined prematuration gametocytes which contract to form round to broadly elongate mature gametocytes. Phyllodactylus ventralis of Estado Portuguesa is parasitized by two new unpigmented malarial species. One produces 11–35 merozoites in schizonts which are often rounded or elongated, occasionally fan-shaped. Gametocytes are always elongated and usually lie diagonally across one end of the host cell or laterally to the nucleus. The second species forms rounded mature schizonts nearly filled with 14–32 merozoites. The sexual stages are usually round or oval, rarely elongate. Plasmodium aurulentum Telford, 1971 was found in Thecadactylus raplcaudus of Estados Portuguesa and Cojedes. A single Thecadactylus from Cojedes was infected by a haemosporidian species of uncertain generic identity which resembles a parasite found earlier in a Panamanian gecko.     

Interspecies bacterial competition regulates community assembly in the C. elegans intestine

From insects to mammals, a large variety of animals hold in their intestines complex bacterial communities that play an important role in health and disease. To further our understanding of how intestinal bacterial communities assemble and function, we study the C. elegans microbiota with a bottom-up approach by feeding this nematode with bacterial monocultures as well as mixtures of two to eight bacterial species. We find that bacteria colonizing well in monoculture do not always do well in co-cultures due to interspecies bacterial interactions. Moreover, as community diversity increases, the ability to colonize the worm gut in monoculture becomes less important than interspecies interactions for determining community assembly. To explore the role of host–microbe adaptation, we compare bacteria isolated from C. elegans intestines and non-native isolates, and we find that the success of colonization is determined more by a species’ taxonomy than by the isolation source. Lastly, by comparing the assembled microbiotas in two C. elegans mutants, we find that innate immunity via the p38 MAPK pathway decreases bacterial abundances yet has little influence on microbiota composition. These results highlight that bacterial interspecies interactions, more so than host–microbe adaptation or gut environmental filtering, play a dominant role in the assembly of the C. elegans microbiota.Subject terms: Microbiome, Microbial ecology