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Larva of the greater wax moth, Galleria mellonella, is a suitable alternative host for studying virulence of fish pathogenic Vibrio anguillarum.

McMillan S, Verner-Jeffreys D, Weeks J, Austin B, Desbois AP - BMC Microbiol. (2015)

Bottom Line: However, disease studies performed in fish hosts often require specialist infrastructure (e.g., aquaria), adherence to strict legislation and do not permit high-throughput approaches; these reasons justify the development of alternative hosts.The G. mellonella model is simple, more ethically acceptable than experiments on vertebrates and, crucially, does not necessitate liquid systems, which reduces infrastructure requirements and biohazard risks associated with contaminated water.The G. mellonella model may aid our understanding of microbial pathogens in aquaculture and lead to the timely introduction of new effective remedies for infectious diseases, while adhering to the principles of replacement, reduction and refinement (3Rs) and considerably reducing the number of vertebrates used in such studies.

View Article: PubMed Central - PubMed

Affiliation: Marine Biotechnology Research Group, Institute of Aquaculture, School of Natural Sciences, University of Stirling, Stirling, UK. stuart.mcmillan@stir.ac.uk.

ABSTRACT

Background: Microbial diseases cause considerable economic losses in aquaculture and new infection control measures often rely on a better understanding of pathogenicity. However, disease studies performed in fish hosts often require specialist infrastructure (e.g., aquaria), adherence to strict legislation and do not permit high-throughput approaches; these reasons justify the development of alternative hosts. This study aimed to validate the use of larvae of the greater wax moth (Galleria mellonella) to investigate virulence of the important fish pathogen, Vibrio anguillarum.

Results: Using 11 wild-type isolates of V. anguillarum, these bacteria killed larvae in a dose-dependent manner and replicated inside the haemolymph, but infected larvae were rescued by antibiotic therapy. Crucially, virulence correlated significantly and positively in larva and Atlantic salmon (Salmo salar) infection models. Challenge studies with mutants knocked out for single virulence determinants confirmed conserved roles in larva and fish infections in some cases (pJM1 plasmid, rtxA), but not all (empA, flaA, flaE).

Conclusions: The G. mellonella model is simple, more ethically acceptable than experiments on vertebrates and, crucially, does not necessitate liquid systems, which reduces infrastructure requirements and biohazard risks associated with contaminated water. The G. mellonella model may aid our understanding of microbial pathogens in aquaculture and lead to the timely introduction of new effective remedies for infectious diseases, while adhering to the principles of replacement, reduction and refinement (3Rs) and considerably reducing the number of vertebrates used in such studies.

No MeSH data available.


Related in: MedlinePlus

Pearson correlation of virulence of 11 wild-type Vibrio anguillarum isolates in the Galleria mellonella larvae (calculated as cumulative larval survival; present study) and in a Salmo salar infection model (median lethal doses determined after intraperitoneal injection [25]). Median lethal doses for Vib64, Vib87, Vib88 and Vib93 were >108 CFU (the highest dose administered) but these have been plotted as 1 × 108 CFU. The numbers beside the points designate each ‘Vib’ isolate. n = 11
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Fig4: Pearson correlation of virulence of 11 wild-type Vibrio anguillarum isolates in the Galleria mellonella larvae (calculated as cumulative larval survival; present study) and in a Salmo salar infection model (median lethal doses determined after intraperitoneal injection [25]). Median lethal doses for Vib64, Vib87, Vib88 and Vib93 were >108 CFU (the highest dose administered) but these have been plotted as 1 × 108 CFU. The numbers beside the points designate each ‘Vib’ isolate. n = 11

Mentions: The relative virulence of each isolate in the larva was compared with virulence determined previously for each isolate in a S. salar infection model [25]. Interestingly, there was highly significant positive correlation (p < 0.01) between relative virulence of the 11 wild-type isolates in larva and fish models of infection, indicating that more virulent V. anguillarum strains in S. salar were also more virulent in the insect (Fig. 4). Closer examination of the growth of each isolate in the larva revealed that more virulent strains replicated faster and reached greater burden in the haemolymph than less virulent isolates (Fig. 3). Indeed, within 48 h the more virulent isolates had increased to approximately 109–1010 CFU/mL in the larval haemolymph, whereas less virulent strains reached approximately 107–108 CFU/mL by this time (Fig. 3). After 48 h it was not possible to obtain data for the more virulent strains as most larvae were dead, while the haemolymph burden of less virulent strains remained at approximately 107–108 CFU/mL for the duration of the experiment (Fig. 3). Additionally, mean bacterial burden in the haemolymph of dead larvae was determined and there was always greater than 5.72 × 109 CFU/mL in these insects for the seven strains and times examined, indicating the likely breaching of a burden threshold before larval death ensued (data not shown).Fig. 4


Larva of the greater wax moth, Galleria mellonella, is a suitable alternative host for studying virulence of fish pathogenic Vibrio anguillarum.

McMillan S, Verner-Jeffreys D, Weeks J, Austin B, Desbois AP - BMC Microbiol. (2015)

Pearson correlation of virulence of 11 wild-type Vibrio anguillarum isolates in the Galleria mellonella larvae (calculated as cumulative larval survival; present study) and in a Salmo salar infection model (median lethal doses determined after intraperitoneal injection [25]). Median lethal doses for Vib64, Vib87, Vib88 and Vib93 were >108 CFU (the highest dose administered) but these have been plotted as 1 × 108 CFU. The numbers beside the points designate each ‘Vib’ isolate. n = 11
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4477312&req=5

Fig4: Pearson correlation of virulence of 11 wild-type Vibrio anguillarum isolates in the Galleria mellonella larvae (calculated as cumulative larval survival; present study) and in a Salmo salar infection model (median lethal doses determined after intraperitoneal injection [25]). Median lethal doses for Vib64, Vib87, Vib88 and Vib93 were >108 CFU (the highest dose administered) but these have been plotted as 1 × 108 CFU. The numbers beside the points designate each ‘Vib’ isolate. n = 11
Mentions: The relative virulence of each isolate in the larva was compared with virulence determined previously for each isolate in a S. salar infection model [25]. Interestingly, there was highly significant positive correlation (p < 0.01) between relative virulence of the 11 wild-type isolates in larva and fish models of infection, indicating that more virulent V. anguillarum strains in S. salar were also more virulent in the insect (Fig. 4). Closer examination of the growth of each isolate in the larva revealed that more virulent strains replicated faster and reached greater burden in the haemolymph than less virulent isolates (Fig. 3). Indeed, within 48 h the more virulent isolates had increased to approximately 109–1010 CFU/mL in the larval haemolymph, whereas less virulent strains reached approximately 107–108 CFU/mL by this time (Fig. 3). After 48 h it was not possible to obtain data for the more virulent strains as most larvae were dead, while the haemolymph burden of less virulent strains remained at approximately 107–108 CFU/mL for the duration of the experiment (Fig. 3). Additionally, mean bacterial burden in the haemolymph of dead larvae was determined and there was always greater than 5.72 × 109 CFU/mL in these insects for the seven strains and times examined, indicating the likely breaching of a burden threshold before larval death ensued (data not shown).Fig. 4

Bottom Line: However, disease studies performed in fish hosts often require specialist infrastructure (e.g., aquaria), adherence to strict legislation and do not permit high-throughput approaches; these reasons justify the development of alternative hosts.The G. mellonella model is simple, more ethically acceptable than experiments on vertebrates and, crucially, does not necessitate liquid systems, which reduces infrastructure requirements and biohazard risks associated with contaminated water.The G. mellonella model may aid our understanding of microbial pathogens in aquaculture and lead to the timely introduction of new effective remedies for infectious diseases, while adhering to the principles of replacement, reduction and refinement (3Rs) and considerably reducing the number of vertebrates used in such studies.

View Article: PubMed Central - PubMed

Affiliation: Marine Biotechnology Research Group, Institute of Aquaculture, School of Natural Sciences, University of Stirling, Stirling, UK. stuart.mcmillan@stir.ac.uk.

ABSTRACT

Background: Microbial diseases cause considerable economic losses in aquaculture and new infection control measures often rely on a better understanding of pathogenicity. However, disease studies performed in fish hosts often require specialist infrastructure (e.g., aquaria), adherence to strict legislation and do not permit high-throughput approaches; these reasons justify the development of alternative hosts. This study aimed to validate the use of larvae of the greater wax moth (Galleria mellonella) to investigate virulence of the important fish pathogen, Vibrio anguillarum.

Results: Using 11 wild-type isolates of V. anguillarum, these bacteria killed larvae in a dose-dependent manner and replicated inside the haemolymph, but infected larvae were rescued by antibiotic therapy. Crucially, virulence correlated significantly and positively in larva and Atlantic salmon (Salmo salar) infection models. Challenge studies with mutants knocked out for single virulence determinants confirmed conserved roles in larva and fish infections in some cases (pJM1 plasmid, rtxA), but not all (empA, flaA, flaE).

Conclusions: The G. mellonella model is simple, more ethically acceptable than experiments on vertebrates and, crucially, does not necessitate liquid systems, which reduces infrastructure requirements and biohazard risks associated with contaminated water. The G. mellonella model may aid our understanding of microbial pathogens in aquaculture and lead to the timely introduction of new effective remedies for infectious diseases, while adhering to the principles of replacement, reduction and refinement (3Rs) and considerably reducing the number of vertebrates used in such studies.

No MeSH data available.


Related in: MedlinePlus