Drosophila
 as a Model Host for
 Pseudomonas aeruginosa
 Infection

D’Argenio, David A.; Gallagher, Larry A.; Berg, Celeste A.; Manoil, Colin

doi:10.1128/jb.183.4.1466-1471.2001

Cited by 206 publications

(164 citation statements)

References 48 publications

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“…The Ecc15, Escherichia coli, and Micrococcus luteus strains were described in ref. 17. The P. aeruginosa PA01, P. putida KT2440, and P. syringae pv.…”

Section: Methodsmentioning

confidence: 99%

Drosophila host defense after oral infection by an entomopathogenic Pseudomonas species

Vodovar

Vinals

Liehl

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

405

485

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Drosophila has been shown to be a valuable model for the investigation of host-pathogen interactions. Study of the Drosophila immune response has been hampered, however, by the lack of true Drosophila pathogens. In nearly all studies reported, the bacteria used were directly injected within the body cavity of the insect, bypassing the initial steps of a natural interaction. Here, we report the identification of a previously uncharacterized bacterial species, Pseudomonas entomophila (Pe), which has the capacity to induce the systemic expression of antimicrobial peptide genes in Drosophila after ingestion. In contrast to previously identified bacteria, Pe is highly pathogenic to both Drosophila larvae and adults, and its persistence in larvae leads to a massive destruction of gut cells. Using this strain, we have analyzed the modulation of the larval transcriptome upon bacterial infection. We found that natural infection by Pe induces a dramatic change in larval gene expression. In addition to immunity genes, our study identifies many genes associated with Pe pathogenesis that have been previously unreported.innate immunity ͉ microarray ͉ host-microbe interaction

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“…The Ecc15, Escherichia coli, and Micrococcus luteus strains were described in ref. 17. The P. aeruginosa PA01, P. putida KT2440, and P. syringae pv.…”

Section: Methodsmentioning

confidence: 99%

Drosophila host defense after oral infection by an entomopathogenic Pseudomonas species

Vodovar

Vinals

Liehl

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

405

485

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“…Transposon mutagenesis of M. extorquens AM1 was performed using the ISphoA|hah-Tc delivery plasmid pCM639 (D'Argenio et al, 2001;Marx et al, 2003). pCM639 was introduced into wild-type M. extorquens AM1 by biparental mating using Escherichia coli SM10 l pir (Miller & Mekalanos, 1988).…”

Section: Methodsmentioning

confidence: 99%

Reconstruction of C3 and C4 metabolism in Methylobacterium extorquens AM1 using transposon mutagenesis

et al. 2003

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The growth of Methylobacterium extorquens AM1 on C 1 compounds has been well-studied, but little is known about how this methylotroph grows on multicarbon compounds. A Tn5 transposon mutagenesis procedure was performed to identify genes involved in the growth of M. extorquens AM1 on succinate and pyruvate. Of the 15 000 insertion colonies screened, 71 mutants were found that grew on methanol but either grew slowly or were unable to grow on one or both of the multicarbon substrates. For each of these mutants, the chromosomal region adjacent to the insertion site was sequenced, and 55 different genes were identified and assigned putative functions. These genes fell into a number of predicted categories, including central carbon metabolism, carbohydrate metabolism, regulation, transport and non-essential housekeeping functions. This study focused on genes predicted to encode enzymes of central heterotrophic metabolism: 2-oxoglutarate dehydrogenase, pyruvate dehydrogenase and NADH : ubiquinone oxidoreductase. In each case, the mutants showed normal growth on methanol and impaired growth on pyruvate and succinate, consistent with a role specific to heterotrophic metabolism. For the first two cases, no detectable activity of the corresponding enzyme was found in the mutant, verifying the predictions. The results of this study were used to reconstruct multicarbon metabolism of M. extorquens AM1 during growth on methanol, succinate and pyruvate.

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“…The injection method described here bypasses the need for a nematode vector, thus uncoupling the effects of bacteria and nematode on the insect. This method allows for accurate enumeration of infectious material (cells or protein) within the inoculum, which is not possible using other existing methods for analyzing entomopathogenesis, including nicking 11 and oral toxicity assays 12 . Also, oral toxicity assays address the virulence of secreted toxins introduced into the digestive system of larvae, whereas the direct injection method addresses the virulence of wholecell inocula.…”

mentioning

confidence: 99%

Rearing and Injection of Manduca sexta Larvae to Assess Bacterial Virulence

Hussa

Goodrich-Blair

2012

JoVE

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Manduca sexta, commonly known as the tobacco hornworm, is considered a significant agricultural pest, feeding on solanaceous plants including tobacco and tomato. The susceptibility of M. sexta larvae to a variety of entomopathogenic bacterial species [1][2][3][4][5] , as well as the wealth of information available regarding the insect's immune system [6][7][8] , and the pending genome sequence 9 make it a good model organism for use in studying host-microbe interactions during pathogenesis. In addition, M. sexta larvae are relatively large and easy to manipulate and maintain in the laboratory relative to other susceptible insect species. Their large size also facilitates efficient tissue/hemolymph extraction for analysis of the host response to infection.The method presented here describes the direct injection of bacteria into the hemocoel (blood cavity) of M. sexta larvae. This approach can be used to analyze and compare the virulence characteristics of various bacterial species, strains, or mutants by simply monitoring the time to insect death after injection. This method was developed to study the pathogenicity of Xenorhabdus and Photorhabdus species, which typically associate with nematode vectors as a means to gain entry into the insect. Entomopathogenic nematodes typically infect larvae via natural digestive or respiratory openings, and release their symbiotic bacterial contents into the insect hemolymph (blood) shortly thereafter 10 . The injection method described here bypasses the need for a nematode vector, thus uncoupling the effects of bacteria and nematode on the insect. This method allows for accurate enumeration of infectious material (cells or protein) within the inoculum, which is not possible using other existing methods for analyzing entomopathogenesis, including nicking 11 and oral toxicity assays 12 . Also, oral toxicity assays address the virulence of secreted toxins introduced into the digestive system of larvae, whereas the direct injection method addresses the virulence of wholecell inocula.The utility of the direct injection method as described here is to analyze bacterial pathogenesis by monitoring insect mortality. However, this method can easily be expanded for use in studying the effects of infection on the M. sexta immune system. The insect responds to infection via both humoral and cellular responses. The humoral response includes recognition of bacterial-associated patterns and subsequent production of various antimicrobial peptides 7 ; the expression of genes encoding these peptides can be monitored subsequent to direct infection via RNA extraction and quantitative PCR 13 . The cellular response to infection involves nodulation, encapsulation, and phagocytosis of infectious agents by hemocytes 6 . To analyze these responses, injected insects can be dissected and visualized by microscopy 13,14 . Video LinkThe video component of this article can be found at

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Drosophila as a Model Host for Pseudomonas aeruginosa Infection

Cited by 206 publications

References 48 publications

Drosophila host defense after oral infection by an entomopathogenic Pseudomonas species

Drosophila host defense after oral infection by an entomopathogenic Pseudomonas species

Reconstruction of C3 and C4 metabolism in Methylobacterium extorquens AM1 using transposon mutagenesis

Rearing and Injection of <em>Manduca sexta</em> Larvae to Assess Bacterial Virulence

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