Flow cytometry analysis of Nosema species to assess spore viability and longevity.

Nosema apis and Nosema ceranae are microsporidia which present resistant spores for the transmission stage (environmental spores) that play an important role for epidemiology and for laboratory studies of honey bee microsporidiosis. In this study, the long-term longevity of N. apis and N. ceranae spores exposed to 4 °C, room temperature (mean 25 °C) and 35 °C for 6-month long and to -20 °C for 10-month long has been assessed by flow cytometry. Storage temperature and the length of storage duration had adverse effects on spore viability of both Nosema spores, with significant differences between the two species. The greatest increase in spore mortality was observed in N. apis spores stored at 33 °C (64, 89%) and in N. ceranae spores at -20 °C (53.55%) and at 33 °C (51.97%). For N. ceranae spores at -20 °C, the loss in viability was very quick, getting an increase over 20% just after 6 days of exposure. Results on viability were confirmed by the infectivity tests where the lowest infectivity for N. ceranae was observed with spores stored for 10 months at -20 °C (79%; P < 0.05) and for N. apis with spores stored at 33 °C (71%; P < 0.05). For both Nosema species, the best storage temperatures were 25 and 4 °C, especially for N. apis that was almost unaffected at those temperatures.