Standard methods for European foulbrood research

Authors: Eva Forsgren, Giles E Budge, Jean-Daniel Charrière and Michael A Z Hornitzky.

Table of contents

 

 

Authors
Summary
1. Introduction

   1.1. Background
   1.2. Disease symptoms
   1.3. Secondary bacteria
   1.4. Diagnosis
2. Bio-safety recommendations
3. Reference strains of Melissococcus plutonius
4. Sampling

   4.1. Brood
   4.2. Adults
   4.3. Honey and pollen
5. Cultivation of M. plutonius
   5.1. Basal medium (modified from Bailey, 1957)
   5.2. M110 agar (from BCCM/LMG bacteria catalogue)

      5.2.1. Agar base
      5.2.2. Agar plates
   5.3. Anaerobic incubation
   5.4. Medium for long-term storage

6. Microscopy
   6.1. Carbol fuchsin staining
   6.2. Gram staining
7. Immunology-based methods
   7.1. ELISA (Enzyme Linked Immuno Sorbent Assay)
      7.1.2. Sample processing
      7.1.3. Indirect ELISA
   7.2. Lateral flow immunoassay (LFI)
8. PCR-based methods
   8.1. Processing
   8.2. DNA extraction

      8.2.1. Adults
      8.2.2. Larvae / pupae
      8.2.3. Honey
   8.3. PCR
      8.3.1. Qualitative PCR
      8.3.2. Quantitative PCR, qPCR
9. Exposure bioassays using in vitro rearing of larvae
   9.1. Estimating the concentration of bacteria
      9.1.1. Plate count
      9.1.2. Total or microscopic count
   9.2. Protocol for inducing EFB infection in honey bee larvae reared in vitro
10. Measuring susceptibility / resistance to antibiotics of Melissococcus plutonius
11. Conclusions
12. References