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Hermes, a functional non-Drosophilid insect gene vector from Musca domestica.

ABSTRACT: Hermes is a short inverted repeat-type transposable element from the house fly, Musca domestica. Using an extra-chromosomal transpositional recombination assay, we show that Hermes elements can accurately transpose in M. domestica embryos. To test the ability of Hermes to function in species distantly related to M. domestica we used a nonautonomous Hermes element containing the Drosophila melanogaster while (w+) gene and created D. melanogaster germline transformants. Transgenic G1 insects were recovered from 34.6% of the fertile G0 adults developing from microinjected w- embryos. This transformation rate is comparable with that observed using P or hobo vectors in D. melanogaster, however, many instances of multiple-element insertions and large clusters were observed. Genetic mapping, Southern blotting, polytene chromosome in situ hybridization and DNA sequence analyses confirmed that Hermes elements were chromosomally integrated in transgenic insects. Our data demonstrate that Hermes elements transpose at high rates in D. melanogaster and may be an effective gene vector and gene-tagging agent in this species and distantly related species of medical and agricultural importance.

SUBMITTER: O'Brochta DA

PROVIDER: S-EPMC1207027 | biostudies-other | 1996 Mar

REPOSITORIES: biostudies-other

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Hermes, a functional non-Drosophilid insect gene vector from Musca domestica.

O'Brochta D A DA Warren W D WD Saville K J KJ Atkinson P W PW

Genetics 19960301 3

Hermes is a short inverted repeat-type transposable element from the house fly, Musca domestica. Using an extra-chromosomal transpositional recombination assay, we show that Hermes elements can accurately transpose in M. domestica embryos. To test the ability of Hermes to function in species distantly related to M. domestica we used a nonautonomous Hermes element containing the Drosophila melanogaster while (w+) gene and created D. melanogaster germline transformants. Transgenic G1 insects were ...[more]

PMID: 8849896

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Project description:House flies are of major concern as vectors of food-borne pathogens to food crops. House flies are common pests on cattle feedlots and dairies, where they develop in and feed on animal waste. By contacting animal waste, house flies can acquire human pathogenic bacteria such as Escherichia coli and Salmonella spp., in addition to other bacteria, viruses, or parasites that may infect humans and animals. The subsequent dispersal of house flies from animal facilities to nearby agricultural fields containing food crops may lead to pre-harvest food contamination with these pathogens. We hypothesized that odors from honeydew, the sugary excreta produced by sucking insects feeding on crops, or molds and fungi growing on honeydew, may attract house flies, thereby increasing the risk of food crop contamination. House fly attraction to honeydew-contaminated plant material was evaluated using a laboratory bioassay. House flies were attracted to the following plant-pest-honeydew combinations: citrus mealybug on squash fruit, pea aphid on faba bean plants, whitefly on navel orange and grapefruit leaves, and combined citrus mealybug and cottony cushion scale on mandarin orange leaves. House flies were not attracted to field-collected samples of lerp psyllids on eucalyptus plants or aphids on crepe myrtle leaves. Fungi associated with field-collected honeydews were isolated and identified for further study as possible emitters of volatiles attractive to house flies. Two fungal species, Aureobasidium pullulans and Cladosporium cladosporioides, were repeatedly isolated from field-collected honeydew samples. Both fungal species were grown in potato dextrose enrichment broth and house fly attraction to volatiles from these fungal cultures was evaluated. House flies were attracted to odors from A. pullulans cultures but not to those of C. cladosporioides. Identification of specific honeydew odors that are attractive to house flies could be valuable for the development of improved house fly baits for management of this pest species.

Project description:BACKGROUND:It has been reported that there are more than ten antimicrobial peptides (AMPs) belonging to the cecropin family in Musca domestica; however, few of them have been identified, and the functions of the other molecules are poorly understood. METHODS:Sequences of the M. domestica cecropin family of genes were cloned from cDNA template, which was reverse-transcribed from total mRNA isolated from third-instar larvae of M. domestica that were challenged with pathogens. Sequence analysis was performed using DNAMAN comprehensive analysis software, and a molecular phylogenetic tree of the cecropin family was constructed using the Neighbour-Joining method in MEGA v.5.0 according to the mature peptide sequences. Antibacterial activity of the synthetic M. domestica cecropin protein was detected and the minimum inhibitory concentration (MIC) values were determined using broth microdilution techniques. Time-killing assays were performed on the Gram-negative bacteria, Acinetobacter baumannii, at the logarithmic or stabilizing stages of growth, and its morphological changes when treated with Cec4 were assessed by scanning electron microscopy (SEM) and detection of leakage of 260 nm absorbing material. RESULTS:Eleven cecropin family genes, namely Cec01, Cec02 and Cec1-9, show homology to the Cec form in a multigene family on the Scaffold18749 of M. domestica. In comparing the encoded cecropin protein sequences, most of them have the basic characteristics of the cecropin family, containing 19 conservative amino acid residues. To our knowledge, this is the first experimental demonstration that most genes in the Cec family are functional. Cec02, Cec1, Cec2, Cec5 and Cec7 have similar antibacterial spectra and antibacterial effects against Gram-negative bacteria, while Cec4 displays a more broad-spectrum of antimicrobial activity and has a very strong effect on A. baumannii. Cec4 eliminated A. baumannii in a rapid and concentration-dependent manner, with antibacterial effects within 24 h at 1× MIC and 2× MIC. Furthermore, SEM analysis and the leakage of 260 nm absorbing material detection indicated that Cec4 sterilized the bacteria through the disruption of cell membrane integrity. CONCLUSIONS:Although there are more than ten cecropin genes related to M. domestica, some of them have no preferred antibacterial activity other than Cec4 against A. baumannii.

Dataset Information

Hermes, a functional non-Drosophilid insect gene vector from Musca domestica.

Publications

Hermes, a functional non-Drosophilid insect gene vector from Musca domestica.

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