Standardization of G. mellonella Larvae to Provide Reliable and Reproducible Results in the Study of Fungal Pathogens
Abstract
:1. Introduction
2. Reported Variability of Fungal Infection Models
3. Standardization of G. mellonella Larvae
4. Standardization of Challenge and Dosing
4.1. Subcutaneous Microinjection
4.2. Feeding Larvae
4.3. Oral Gavage
5. Standardised Scoring
6. High Throughput Screens
6.1. High Throughput Screening of Mutant Libraries
6.2. Screening for Antifungal Agents
7. Discussion
Author Contributions
Funding
Conflicts of Interest
References
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Model | Whole Animal Model | Use at 37 °C | Precise Dosing | Immune System | Cost of Maintenance by User | Regulated Use in the UK |
---|---|---|---|---|---|---|
Monolayer cell cultures | no | yes | yes | no | medium | no |
3D cell cultures | no | yes | yes | no | high | no |
Caenorhabditis elegans (nematode) | yes | no | no | yes | low | no |
Panagrellus redivivus (nematode) | yes | yes | no | yes | low | no |
Zebra fish (and embryos) | yes | no | yes | yes | high | yes (fish and older embryos) |
Drosophila melanogaster (fruit fly) | yes | yes | no | yes | low | no |
G. mellonella | yes | yes | yes | yes | low | no |
Manduca sexta (tobacco hornworm) | yes | yes | yes | yes | low | regulated as a crop pest. |
Category | Description | Score |
---|---|---|
activity | no movement | 0 |
minimal movement on stimulation | 1 | |
move when stimulated | 2 | |
move without stimulation | 3 | |
cocoon formation | no cocoon | 0 |
partial cocoon | 0.5 | |
full cocoon | 1 | |
melanisation | black larvae | 0 |
black spots on brown larvae | 1 | |
≥3 spots on beige larvae | 2 | |
<3 spots on beige larvae | 3 | |
no melanisation | 4 | |
survival | dead | 0 |
alive | 2 |
Model Limitations | Consequences | Solution |
---|---|---|
Differences in age, weight and health status of larvae | Lack of reproducibility between experiments | Use age and weight defined larvae |
Antibiotic and hormone residues | Lack of reproducibility between experiments. May distort the results of tests of antimicrobial efficacy | Use larvae bred without the use of antibiotics or hormones |
Genetic diversity of G. mellonella | Lack of reproducibility between experiments | Use inbred breeding colony |
Larvae have a surface flora of pathogenic microorganisms | Deaths in controls when injected with PBS | Surface decontaminate larvae |
Larvae normally available do not feed | Difficult to dose orally with pathogens or chemicals | Use early instar stage larvae or Oral gavage |
Scoring or morbidity or mortality can be subjective | End points are not well defined | Use Heath Index Scoring System |
Dosing involves injection of small volumes | Dose of pathogen given is not precisely determined | Use Hamilton syringes |
High throughput screening is limited by operator ability to inject large numbers of larvae | Screens are size limited | Reduce cohort size using standardised larvae or Develop automated screens |
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Champion, O.L.; Titball, R.W.; Bates, S. Standardization of G. mellonella Larvae to Provide Reliable and Reproducible Results in the Study of Fungal Pathogens. J. Fungi 2018, 4, 108. https://doi.org/10.3390/jof4030108
Champion OL, Titball RW, Bates S. Standardization of G. mellonella Larvae to Provide Reliable and Reproducible Results in the Study of Fungal Pathogens. Journal of Fungi. 2018; 4(3):108. https://doi.org/10.3390/jof4030108
Chicago/Turabian StyleChampion, Olivia L., Richard W. Titball, and Steven Bates. 2018. "Standardization of G. mellonella Larvae to Provide Reliable and Reproducible Results in the Study of Fungal Pathogens" Journal of Fungi 4, no. 3: 108. https://doi.org/10.3390/jof4030108