Summary

والحشرات<em> جاليريا mellonella</em> نموذجا عدوى قوية المعنية بالتحقيق في إمراض البكتيرية

Published: December 11, 2012
doi:

Summary

haemocolic العدوى عن طريق الفم وداخل يرقات العث من الشمع أكبر<em> جاليريا mellonella</emيوصف>. ويمكن استخدام هذه الحشرة لدراسة العوامل الممرضة للحشرات من الفوعة وكذلك البكتيريا الانتهازية الثدييات. تربية الحشرات، وطرق العدوى والأمثلة على<em> في الجسم الحي</emموصوفة> التحليل.

Abstract

The study of bacterial virulence often requires a suitable animal model. Mammalian models of infection are costly and may raise ethical issues. The use of insects as infection models provides a valuable alternative. Compared to other non-vertebrate model hosts such as nematodes, insects have a relatively advanced system of antimicrobial defenses and are thus more likely to produce information relevant to the mammalian infection process. Like mammals, insects possess a complex innate immune system1. Cells in the hemolymph are capable of phagocytosing or encapsulating microbial invaders, and humoral responses include the inducible production of lysozyme and small antibacterial peptides2,3. In addition, analogies are found between the epithelial cells of insect larval midguts and intestinal cells of mammalian digestive systems. Finally, several basic components essential for the bacterial infection process such as cell adhesion, resistance to antimicrobial peptides, tissue degradation and adaptation to oxidative stress are likely to be important in both insects and mammals1. Thus, insects are polyvalent tools for the identification and characterization of microbial virulence factors involved in mammalian infections.

Larvae of the greater wax moth Galleria mellonella have been shown to provide a useful insight into the pathogenesis of a wide range of microbial infections including mammalian fungal (Fusarium oxysporum, Aspergillus fumigatus, Candida albicans) and bacterial pathogens, such as Staphylococcus aureus, Proteus vulgaris, Serratia marcescens Pseudomonas aeruginosa, Listeria monocytogenes or Enterococcus faecalis4-7. Regardless of the bacterial species, results obtained with Galleria larvae infected by direct injection through the cuticle consistently correlate with those of similar mammalian studies: bacterial strains that are attenuated in mammalian models demonstrate lower virulence in Galleria, and strains causing severe human infections are also highly virulent in the Galleria model8-11. Oral infection of Galleria is much less used and additional compounds, like specific toxins, are needed to reach mortality.

G. mellonella larvae present several technical advantages: they are relatively large (last instar larvae before pupation are about 2 cm long and weight 250 mg), thus enabling the injection of defined doses of bacteria; they can be reared at various temperatures (20 °C to 30 °C) and infection studies can be conducted between 15 °C to above 37 °C12,13, allowing experiments that mimic a mammalian environment. In addition, insect rearing is easy and relatively cheap. Infection of the larvae allows monitoring bacterial virulence by several means, including calculation of LD5014, measurement of bacterial survival15,16 and examination of the infection process17. Here, we describe the rearing of the insects, covering all life stages of G. mellonella. We provide a detailed protocol of infection by two routes of inoculation: oral and intra haemocoelic. The bacterial model used in this protocol is Bacillus cereus, a Gram positive pathogen implicated in gastrointestinal as well as in other severe local or systemic opportunistic infections18,19.

Protocol

1. تربية الحشرات دورة كاملة من البيض إلى يرقات الطور مشاركة يستمر حوالي 5 أسابيع في C. ° 25 وهناك حاجة إلى واحد أو 2 أسابيع إضافية للحصول على الفراشات الكبار. وضع لا يقل عن 100 الشرانق أو…

Representative Results

haemocoelic الحقن داخل البكتيريا إلى G. وقد ثبت mellonella مفيدة جدا لتحديد عوامل الفوعة التعامل مع العديد من تلف الأنسجة والمقاومة لعوامل المناعة الفطرية من مسببات الأمراض البشرية عدة. على سبيل المثال، يمثل الرقم 2A فيات الحشرات بعد حقن جرعات مختلفة من B.</em…

Discussion

استخدام الحشرات وخاصة مرحلة اليرقات، ونماذج عدة أنواع العدوى، أصبحت متكررة. نموذج المفضل لبعض الجوانب غير ذبابة الفاكهة (ذبابة النموذج) تستخدم كل من البالغين واليرقات 1،2 المرحلة. الحشرة lepidopteran G. كما تم mellonella تستخدم أساسا لمعايرة الفوعة البكتيري…

Disclosures

The authors have nothing to disclose.

Acknowledgements

نود أن نشكر اليزابيث Guillemet، وكريستوف بويسون Bridoux لودوفيك للمساعدة التقنية الممتازة. نحن مدينون كثيرا لSalamitou سيلفي وFedhila SINDA لإعداد النظام الأولي.

Materials

Name of the reagent Company Catalogue number Comments (optional)
Wax and pollen La Ruche Roanaise 303000 Any honey producer
Automated syringe pump KD Scientific KDS 100  
Syringe 1 ml Terumo BS 01T  
Needle 0.45 x 12 mm Terumo NN 2613R  
Petri dish 5 cm VWR 89000-300  
Needle 30G, 25 mm hypodermic Burkard Mfg. Co. Ltd. PDE0005  

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Cite This Article
Ramarao, N., Nielsen-Leroux, C., Lereclus, D. The Insect Galleria mellonella as a Powerful Infection Model to Investigate Bacterial Pathogenesis. J. Vis. Exp. (70), e4392, doi:10.3791/4392 (2012).

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