The two-spotted lady beetle, Adalia bipunctata L., displays warning colouration that is reinforced by the production of adaline and adalinine. These alkaloids are thought to provide defense against predation throughout all life stages of A. bipunctata and may play a role in the insect immune system. Vairimorpha (Nosema) adaliae, a microsporidium described from A. bipunctata, has minimal effects on its host (delayed larval development) when reared under optimum conditions but stress factors are shown to affect the development of microsporidiosis. The objectives of this study were to determine the effects of V. adaliae on relative alkaloid content (adaline) during A. bipunctata development, and to evaluate the combined effects of physical stress and infection on adult beetles (relative alkaloid content and infection load). First-instar larvae were isolated from uninfected and V. adaliae-infected colonies. Eggs and first-instar larvae were immediately prepared for alkaloid analysis, whereas late-instar larvae, pupae and adults were systematically processed when each reached their designated developmental stage. Upon eclosion, a subsample of beetles was exposed to varying amounts of physical agitation: control (no shaking), alternate shaking (every other day), and daily shaking. Immediately following these stress trials, alkaloid samples were collected for analysis and spore loads were assessed. Overall, relative adaline proportions increased from egg to adult. Uninfected individuals had significantly higher relative proportions of adaline than did infected individuals during early development; however, adaline content was higher in infected A. bipunctata from the third-instar onwards, when compared to their uninfected counterparts. Following exposure to physical agitation on alternate days, uninfected adults had a significantly higher relative proportion of adaline than did infected adults. Interestingly, exposure to different levels of agitation had no significant effect on alkaloid production for either uninfected or infected beetles. Mean spore counts were significantly higher for adults that were exposed to daily shaking when compared to individuals from the control and alternate shaking groups. From a biological perspective, one would expect to observe differences in alkaloid production through coccinellid development, as each successive life stage faces different external pressures and risks. When infected with the microsporidium V. adaliae, however, adaline production was reduced during early development but increased significantly in late life stages.

Honey is known for its antibacterial and antifungal properties. Manuka honey was examined for its potential to manage the microsporidium Vairimorpha (Nosema) adaliae infecting Adalia bipunctata larvae. Development time for uninfected larvae fed aphids and water was 13.0 +/- 0.2 days, which did not differ significantly from larvae fed aphids and manuka honey. Development of V. adaliae-infected larvae fed aphids and water was 16.3 +/- 0.5 days, compared to 15.0 +/- 0.2, 15.2 +/- 0.3, and 15.6 +/- 0.2 days for larvae fed aphids and 5 %, 10 %, or 15 % manuka honey, respectively. Development time was shorter for all honey treatments, but only those fed 5 % manuka differed significantly from the control. Control adults had 19.4 +/- 3.0 spores/120 mu m2, compared to 19.0 +/- 2.0, 19.1 +/- 2.1, and 14.3 +/- 2.2, for adults provided with 5 %, 10 %, and 15 % honey, respectively. Although spore loads did not differ significantly (p > 0.05), lighter infections were observed in the group fed 15 % manuka.

Nosema adaliae, a microsporidian pathogen described from the two-spotted lady beetle, Adalia bipunctata L., delays larval development when A. bipunctata is reared under laboratory conditions at 25 degrees C. In nature, lady beetles often experience a wide range of environmental temperatures, but little is known regarding the effects of microsporidian pathogens on lady beetles when they are reared at higher and lower temperatures. In this study, the effects of elevated rearing temperatures and microsporidiosis were observed on larval development time and mortality, sex ratios, alkaloid content (adaline and adalinine), and adult morphometrics. Uninfected larvae (24 h-old) were provided either an uninfected or N. adaliae-infected conspecific egg and subsequently reared at three temperatures (25 degrees C, 27.5 degrees C or 30 degrees C). After the egg was eaten, larvae were provided a diet of green peach aphids and their development was recorded daily. Following eclosion, a subsample of adults were photographed for microscopic measurements and reflex-fluid was collected for alkaloid analysis using gas chromatography-mass spectrometry. Afterwards, all individuals were examined to determine the number that had become infected, and the same subsample was used to assess the severity of infection. Development time was delayed significantly for larvae that consumed a N. adaliae-infected egg, and infected larvae took significantly longer to develop at 25 degrees C than at 27.5 degrees C or 30 degrees C. No differences were observed for larval mortality or sex ratios. The relative proportion of adaline increased when larvae were reared at 30 degrees C, but adaline concentration was not affected by the pathogen, N. adaliae. Adults reared at 25 degrees C had larger body measurements when compared to those reared at 27.5 degrees C and 30 degrees C. Overall, adults that had eaten a N. adaliae-infected A. bipunctata egg as larvae had similar body measurements to those that ate an uninfected egg. When comparing male and female measurements, a significant interaction between infection status, temperature and sex was observed for elytra length and head capsule width only. These measurements were similar for uninfected and N. adaliae-infected females across all temperatures. However, when reared at 25 degrees C, uninfected males had significantly smaller elytra and head capsules than did infected males; but when reared at 30 degrees C, no significant difference was observed for these measurements. Both percent infection and average spore count decreased at 27.5 degrees C and 30 degrees C. These results suggest that temperatures above 25 degrees C have a mitigating effect against N. adaliae in A. bipunctata.

The effects of food availability and infection with the microsporidium Nosema adaliae on alkaloid production in the two-spotted lady beetle, Adalia bipunctata L., was examined. Alkaloid production (relative percent adaline in reflex-fluid) and pathogen load (spore counts) were quantified for both uninfected and N. adaliae-infected A. bipunctata. Alkaloid content was significantly higher for beetles fed irregularly than for those fed daily. For beetles infected with N. adaliae, spore counts were significantly higher for those fed irregularly compared to those fed daily. These results suggest that adaline content in reflex-fluid is influenced by infection and that irregular food supply increases pathogen load.

Two species of microsporidia have been described from lady beetles that are commercially available for biological control: Nosema adaliae from the two-spotted lady beetle, Adana bipunctata L., and Tubulinosema hippodamiae from the convergent lady beetle, Hippodamia convergens Guerin-Meneville. These pathogens delay larval development under controlled conditions, but little is known regarding the effects that microsporidia cause when their hosts are subjected to stressful conditions that are often experienced in nature. In this study, the combined effects of microsporidiosis (N. adaliae on A. bipunctata and T. hippodamiae on H. convergens) and irregular food availability were observed on host fitness (larval development and mortality, sex ratios, and adult morphometrics). For each beetle species, 24 h-old larvae were provided either an uninfected or microsporidia-infected conspecific egg. After the egg was eaten, some larvae were provided an abundance of aphids daily, whereas others were provided aphids on an irregular basis. Development was delayed significantly for larvae that consumed a microsporidia-infected egg, and for those fed irregularly. For A. bipunctata, a significant interaction was observed between infection status and food availability. This suggests that N. adaliae-infected A. bipunctata larvae that have an irregular supply of aphids undergo further developmental delays than those with a generous food supply. This interaction was not observed for T. hippodamiae-infected H. convergens. For both species, larval mortality and sex ratios did not differ significantly, regardless of infection status or food availability. Adults that were fed daily as larvae were significantly larger than those fed irregularly. However, the elytra of N. adaliae-infected A. bipunctata were significantly larger than the elytra of their uninfected cohorts, and T. hippodamiae-infected H. convergens had wider pronota and head capsules than uninfected H. convergens. Because N. adaliae and T. hippodamiae prolong larval development of their respective hosts under controlled conditions, one would expect these pathogens to cause more profound effects when their hosts experience stressful conditions. The results from this study indicate that this was the case for A. bipunctata, but not for H. convergens.

Lacewing larvae are generalist predators that are commercially available for aphid control on a variety of crops in both Europe and North America. Although lacewings are known for their symbiotic association with yeasts and bacteria, there are few reports of microsporidia in these natural enemies. An undescribed microsporidium was found in Chrysoperla cornea (Stephens) during the routine examination of specimens that were obtained from a commercial insectary for biological pest control. The objective of this study was to describe the pathogen by means of ultrastructure, molecular characterization and tissue pathology. All stages of the microsporidium were diplokaryotic and developed in direct contact with the host cell cytoplasm. Merogony and sporogony were not observed. Mature spores measured 3.49 +/- 0.10 x 1.52 +/- 0.05 mu m and had an isofilar polar filament with 8-10 coils that were frequently arranged in a single row, although double rows were also observed. Spores contained a lamellar polaroplast and a relatively small and inconspicuous polar vacuole was observed in the posterior region of about half of the spores that were examined. Tubular structures, similar in appearance to those in Nosema granulosis were observed in both sporonts and in spores. A cluster of small tubules was also observed in the posterior region of some spores. Microsporidian spores were observed in cells of the proventriculus, diverticulum and in epithelial cells of the posterior midgut The Malpighian tubules, ileum, and rectum were heavily infected. Spores were also observed in the fat body, peripheral region of the ganglia, within and between the flight muscles, and beneath the cuticle. Although the tissues adjacent to the ovaries were heavily infected, microsporidian spores were not observed within the developing eggs. Pathogen transmission was not studied directly because it was difficult to maintain microsporidia-infected C. carnea in the laboratory. The presence of microsporidian spores in the alimentary canal suggests that the pathogen is transmitted per as and horizontal transmission may occur when infected larvae or adults are cannibalized by uninfected larvae. Molecular analysis of the microsporidian genome showed that the pathogen described in this study was 99% similar to Nosema bombycis, N. furnacalis, N. granulosis and N. spodopterae. Based on information gained during this study, we propose that the microsporidium in C. carnea be given the name Nosema chrysoperlae sp. nov. (C) 2013 Elsevier Inc. All rights reserved.

Two-spotted lady beetles, Adalia bipunctata L. are available for biological pest control in North America. Lady beetles are known to host microsporidia and although these pathogens are able to infect more than one host under laboratory conditions, little is known regarding the effects of more than one microsporidian pathogen on host fitness. In this study, egg cannibalism was used to examine the effects of the microsporidium Tubulinosema hippodamiae from Hippodamia convergens and an undescribed microsporidium from A. bipunctata (alone and in combination) on A. bipunctata host fitness (larval development and mortality, sex ratio, adult fecundity and longevity). Development was prolonged significantly for larvae that were infected with the undescribed microsporidium but T. hippodamiae had no effect and as a result, conclusions could not be made regarding the effects of both pathogens on larval development. The two microsporidia had no effect on sex ratios (1♀:1♂) or on adult fecundity and longevity. Spores were detected in the majority of smear preparations of individuals that were fed microsporidia-infected eggs and molecular analysis confirmed the identity of both pathogens in sampled individuals. T. hippodamiae spores were smaller than spores of the undescribed microsporidium (3.76±0.03×2.32±0.02 μm and 5.43±0.06×2.75±0.03 μm, respectively) and although the former stained less intensely than did those of latter, spores of the two pathogens are difficult to differentiate when examined by light microscopy alone. The ability of some microsporidia to infect more than one lady beetle host makes it difficult to conclude with certainty as to the number of species that are present in infected Adalia when specimens are examined solely by light microscopy.