Apoptosis in the pathogenesis of Nosema ceranae (Microsporidia: Nosematidae) in honey bees (Apis mellifera)

Abstract

Nosema ceranae is a parasite of the epithelial ventricular cells of the honey bee that belongs to the microsporidian phylum, a biological group of single-cell, spore-forming obligate intracellular parasites found in all major animal lineages. The ability of host cells to accommodate a large parasitic burden for several days suggests that these parasites subvert the normal host cells to ensure optimal environmental conditions for growth and development. Once infected, cells can counteract the invasive pathogen by initiating their own death by apoptosis as a defence strategy. To determine whether N. ceranae blocks apoptosis in infected ventricular cells, cell death was assessed in sections of the ventriculum from experimentally infected honey bees using the TUNEL assay and by immunohistochemistry for caspase-3. Ventricular epithelial cells from infected bees were larger than those in the uninfected control bees, and they contained N. ceranae at both mature and immature stages in the cytoplasm. Apoptotic nuclei were only observed in some restricted areas of the ventriculum, whereas apoptosis was typically observed throughout the epithelium in uninfected bees. Indeed, the apoptotic index was higher in uninfected versus infected ventriculi. Our results suggested that N. ceranae prevents apoptosis in epithelial cells of infected ventriculi, a mechanism possible designed to enhance parasite development.

Short communication: Survival of honey bees (Apis mellifera) infected with Crithidia mellificae (Langridge and McGhee: ATCC® 30254™) in the presence of Nosema ceranae

“…mellificae was significantly higher than that of those that received C. mellificae alone and that of the control worker bees (Kaplan-Meier Long-Rank test, p<0.001). Conversely, the survival of the control and the C. mellificae infected bees was significantly higher than that of the references tion with N. ceranae is consistent with that seen previously (reviewed by Higes et al, 2013;Williams et al, 2014;Huang et al, 2015;Milbrath et al, 2015), confirming that bees infected with N. ceranae have a shorter life expectancy than uninfected bees. The incubation temperature used may explain the distinct evolution in the survival of infected honey bees relative to other laboratory experiments performed in our laboratory (Martín-Hernandez et al, 2009).…”

“…The DNA from those colonies was extracted as described previously in Cepero et al (2014) and the trypanosomatid species was confirmed by PCR (Meeus et al, 2010) and subsequent sequencing (Cepero et al, 2014). Freshly purified N. ceranae spores for experimental infection were obtained as described elsewhere (Martín-Hernández et al, 2011Higes et al, 2013), counting both the N. ceranae spores and C. mellificae spheroid forms in a haemocytometer. DNA was extracted from an aliquot of Nosema spores to confirm the species by PCR (Martín-Hernández et al, 2012).…”

“…DNA was extracted from an aliquot of Nosema spores to confirm the species by PCR (Martín-Hernández et al, 2012). Nosema-free and trypanosomatid-free honey bees were obtained as described previously (Higes et al, 2013). Emerging worker bees from three different colonies were removed carefully, confined to cages in groups of 25 and kept in the incubator for 5 days at 27ºC, the optimum temperature for C. mellificae growth (Langridge & McGhee, 1967).…”

“…Honey bees (Apis mellifera iberiensis) were infected as described elsewhere (Higes et al, 2013), administering 1 µL of a sucrose solution containing 50,000 N. ceranae spores or 50,000 C. mellificae spheroid forms. The co-infected bees received 1 µL of each pathogen solution, while uninfected control bees were fed with 2 µL of the 50% sucrose solution alone.…”

“…However, most of the studies published in recent years have confirmed that N. ceranae has a pathogenic effect on this host, expressed as a shortening of the honey bee (workers) lifespan (Higes et al, 2013). Significantly, the longevity of infected bees varies strongly among these studies, probably due to the many factors that influence the development and the course of the infection.…”