2.2.6. In vitro rearing of Nosema spp.: cell culture systems
Cell and tissue cultures are indispensable for the propagation and study of obligate intracellular pathogens like viruses and microsporidia. Bee pathogens comprise viruses and microsporidia as obligate intracellular parasites. However, studies on cellular and molecular aspects of pathogen-host interactions of these pathogens and their target cells have been hampered in the past by the complete lack of permanent bee cell lines. Recently, protocols for the prolonged although limited maintenance of primary honey bee cells have been described (Bergem et al., 2006; Hunter, 2010, but these cells have not been used for infection experiments. Several hundreds of non-honey bee insect cell lines are commercially available (Lynn, 2007; van Oers and Lynn, 2010) and have been proven to be valuable tools for elucidating attachment, entry, and replication of several intracellular insect pathogens and for analysing cellular reactions towards infection (Smagghe et al., 2009; van Oers and Lynn, 2010). However, these cell lines were considered unsuitable for the study of bee pathogens due to the assumed host specificity of bee pathogenic viruses and microsporidia. We here describe both, the infection of primary ventricular cells established from honey bee pupae as well as the infection of commercially available insect cell lines established from several lepidoptera with N. ceranae and N. apis. General techniques for cell cultures are described by Genersch et al. (2013) in the BEEBOOK.