Matrix-assisted laser desorption/ionization mass spectrometry biotyping, an approach for deciphering and assessing the identity of the honeybee pathogen Nosema.

RATIONALE

The microsporidia are obligate intracellular pathogenic fungi that parasitize a wide range of invertebrate and vertebrate hosts and have important impacts on health, food security and the economy. In this paper, we focus on Nosema ceranae and N. apis, which chronically infect the digestive tract of honeybees, altering their physiology and lifespan.

METHODS

We applied matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) for rapid molecular profiling of extracts of Nosema spores in order to identify the species and the geographical origin, and assess the viability status of Nosema microsporidia in conjunction with a flow cytometric approach. Pure solutions of spores were prepared for flow cytometric analysis and MALDI-MS profiling. A mechanical extraction of viable or heat-killed Nosema spores was conducted to obtain mass fingerprints of peptides/proteins for samples of microsporidia from different geographical origins (MBO.NC01, MBO.NC02 and MBO.NA01).

RESULTS

A distinction in the peptide/protein profiles between two isolates with different geographical origins was observed. Mass fingerprints of viable and experimentally killed spores were also clearly distinguishable, regardless of Nosema species. Finally, using our computational models on the different Nosema species, we were able to classify five independent isolates of Nosema microsporidia.

CONCLUSIONS

We have shown that MALDI-MS is a rapid, cost-effective and simple method for identifying Nosema species. We demonstrated that MALDI Biotyping could represent a valuable surveillance tool of nosemosis in apiaries for sanitary services and beekeepers.