Alkaloid content in microsporidia-infected Adalia bipunctata (Coleoptera: Coccinellidae) life stages, and pathogen spore load in adults after exposure to physical stress.

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.

Evaluation of manuka honey on the microsporidian pathogen Vairimorpha (Nosema) adaliae and its host, the two-spotted lady beetle, Adalia bipunctata L. (Coleoptera: Coccinellidae).

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 µm2, 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.

Effects of food availability on microsporidiosis and alkaloid production in the two-spotted lady beetle, Adalia bipunctata L.

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.

Effects of temperature on larval development, alkaloid production and microsporidiosis in the two-spotted lady beetle, Adalia bipunctata L. (Coleoptera: Coccinellidae).

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 °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 °C, 27.5 °C or 30 °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 °C than at 27.5 °C or 30 °C. No differences were observed for larval mortality or sex ratios. The relative proportion of adaline increased when larvae were reared at 30 °C, but adaline concentration was not affected by the pathogen, N. adaliae. Adults reared at 25 °C had larger body measurements when compared to those reared at 27.5 °C and 30 °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 °C, uninfected males had significantly smaller elytra and head capsules than did infected males; but when reared at 30 °C, no significant difference was observed for these measurements. Both percent infection and average spore count decreased at 27.5 °C and 30 °C. These results suggest that temperatures above 25 °C have a mitigating effect against N. adaliae in A. bipunctata.

Effects of the microsporidian pathogen, Nosema adaliae (Nosematidae) on the seven-spotted lady beetle, Coccinella septempunctata L. (Coleoptera: Coccinellidae).

Lady beetles are important predators in nature. Some species, including the two-spotted lady beetle, Adalia bipunctata L., are native to North America, whereas others, such as the seven-spotted lady beetle, Coccinella septempunctata L., have been introduced in North America for pest control on agriculture crops. Microsporidia are obligate pathogens that cause chronic disease, and these pathogens are known to infect several lady beetle species. Lady beetles are cannibalistic and, because many species share a given landscape, there is potential for microsporidia to infect susceptible coccinellids when infected eggs are eaten. The objective of this study was to examine the effects of the microsporidium Nosema adaliae isolated from A. bipunctata on C. septempunctata fitness (larval development and mortality, sex ratio, adult longevity and fecundity). Mortality was higher for C. septempunctata larvae that ate four A. bipunctata eggs (≥96% mortality) than for those that ate only one (<63.8%), suggesting that the mortality observed was influenced by the number of eggs eaten. A. bipunctata eggs contain adaline and adalinine, two species-specific alkaloids that have been shown to be detrimental to C. septempunctata larvae. Development of larvae that consumed one uninfected or one N. adaliae-infected A. bipunctata egg, did not differ significantly (20.5 ±â€¯0.2 d and 21.3 ±â€¯0.4 d, respectively) and, although mortality remained high for these larvae (53.5% and 65.6% mortality, respectively), these values also did not differ significantly (p = 0.05). Over a 60-d period, mean fecundity for C. septempunctata adults that ate one uninfected A. bipunctata egg as first-instar larvae was significantly greater (776.6 ±â€¯122.0 eggs) than those that ate one N. adaliae-infected egg (335.6 ±â€¯86.6 eggs, p = 0.005). Larvae from the former group also lived significantly longer (58.2 ±â€¯1.8 d) than did those from the latter group (38.4 ±â€¯6.4 d, p = 0.010). Sex ratios of adult beetles did not differ significantly. Because A. bipunctata and C. septempunctata share similar habitats, it is reasonable to expect these two coccinellids to encounter one another in nature. Results of this study show that the consumption of only one infected A. bipunctata egg by C. septempunctata larvae can result in high larval mortality and reduced fecundity.

Effects of microsporidiosis and food availability on the two-spotted lady beetle, Adalia bipunctata L., and convergent lady beetle, Hippodamia convergens Guérin-Méneville.

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, Adalia bipunctata L., and Tubulinosema hippodamiae from the convergent lady beetle, Hippodamia convergens Guérin-Méneville. 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.

The influence of microsporidian pathogens from commercially available lady beetles on larval development of the green lacewing, Chrysoperla carnea, in the absence of infection.

In North America, more than 70 species of natural enemies are available for pest control, including the aphid predators, Adalia bipunctata L. (two-spotted lady beetle) and Hippodamia convergens Guérin-Méneville (convergent lady beetle), and the generalist predator Chrysoperla carnea Stephens (green lacewing). The two lady beetle species are known to host microsporidian pathogens: Nosema adaliae was originally described from Adalia bipunctata and Tubulinosema hippodamiae from H. convergens. Microsporidia are spore-forming pathogens that typically produce chronic, debilitating disease. Because the spores of both pathogens are transovarially transmitted through beetle eggs, the predation behavior of lacewing larvae provides an opportunity for the transmission of these pathogens when infected lady beetles and lacewings share the same local environment. In this study, uninfected and microsporidia-infected eggs from A. bipunctata and H. convergens were offered to C. carnea larvae. The development of larvae that consumed N. adaliae-infected eggs was not affected, but larval development was prolonged by almost 3 days for those that consumed two or more T. hippodamiae-infected eggs. Prolonged larval development is considered to be costly because larvae remain vulnerable to cannibalization by sibling larvae or other predators. Longevity did not differ significantly between sexes of C. carnea, and the sex ratio of newly eclosed adults did not differ from the previously reported sex ratio of 1♂: 1♀. Upon examination by light microscopy at the end of the trial, two C. carnea larvae were infected with N. adaliae and none were infected with T. hippodamiae, suggesting that T. hippodamiae influenced lacewing larval development without establishing an infection.

Microsporidia - Emergent Pathogens in the Global Food Chain.

Intensification of food production has the potential to drive increased disease prevalence in food plants and animals. Microsporidia are diversely distributed, opportunistic, and density-dependent parasites infecting hosts from almost all known animal taxa. They are frequent in highly managed aquatic and terrestrial hosts, many of which are vulnerable to epizootics, and all of which are crucial for the stability of the animal-human food chain. Mass rearing and changes in global climate may exacerbate disease and more efficient transmission of parasites in stressed or immune-deficient hosts. Further, human microsporidiosis appears to be adventitious and primarily associated with an increasing community of immune-deficient individuals. Taken together, strong evidence exists for an increasing prevalence of microsporidiosis in animals and humans, and for sharing of pathogens across hosts and biomes.

Nosema adaliae sp. nov., a new microsporidian pathogen from the two-spotted lady beetle, Adalia bipunctata L. (Coleoptera: Coccinellidae) and its relationship to microsporidia that infect other coccinellids.

The two-spotted lady beetle, Adalia bipunctata L., is a tree-dwelling lady beetle endemic to parts of Europe, Central Asia and North America that is commercially available for aphid control in Europe and North America. Lady beetles host a wide variety of symbionts including parasitoids, viruses, eugregarines, fungi, bacteria, nematodes and microsporidia. Four species of microsporidia have been described from lady beetles, and an undescribed microsporidium was recently isolated from local populations of A. bipunctata in Nova Scotia, Canada. In a previous study, this pathogen prolonged the development of A. bipunctata larvae but had no effect on adult fecundity, longevity or sex ratios. The objective of this study was to formally describe the microsporidium by means of its ultrastructure, tissue pathology and molecular characterization. All stages of the microsporidium were diplokaryotic and developed in direct contact with the host cell cytoplasm. Mature spores measured 4.25±0.09×1.82±0.03µm (SE, n=49, from micrographs) and fresh spores measured 6.10±0.06×3.01±0.05µm (±SE, n=60; range: 5.0-6.9×2.18-3.86µm). The polar filament was isofilar with 10-18 coils that were frequently arranged in a single row. The lamellar polaroplast was not typically visible and spores contained a relatively small posterior vacuole. Both the flight muscles and fat body were heavily infected and a large number of spores were observed within and between the cells of these tissues. The ovaries, developing oocytes, spermatocytes and accessory glands within the testes, midgut epithelium, Malpighian tubules, ileum, colon, and ventral nerve cord were also infected but not as heavily. Connective tissue near the cuticle and surrounding the trachea were lightly infected. The presence of spores in both the alimentary canal and ovaries (particularly within developing oocytes) suggests that the microsporidium can be transmitted per os (horizontally) and transovarially (vertically). Molecular analysis of the genome of the microsporidium described in this study was 97% similar to Nosema bombi and 96% similar to Nosema thomsoni, Nosema vespula and Nosema oulemae. Based on information gained during this study, we propose that the microsporidium in A. bipunctata be given the name Nosema adaliae sp. nov.

Ultrastructure and molecular characterization of the microsporidium, Nosema chrysoperlae sp. nov., from the green lacewing, Chrysoperla carnea (Stephens) (Neuroptera: Chrysopidae) used for biological pest control.

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 carnea (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±0.10×1.52±0.05µm 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 os 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.

The convergent lady beetle, Hippodamia convergens Guérin-Méneville and its endoparasitoid Dinocampus coccinellae (Schrank): the effect of a microsporidium on parasitoid development and host preference.

Convergent lady beetles, Hippodamia convergens Guérin-Méneville are host to the braconid endoparasitoid, Dinocampus coccinellae (Schrank) and the microsporidian pathogen, Tubulinosema hippodamiae. The interrelationship between the endoparasitoid and the pathogen in H. convergens adults under laboratory conditions was examined by quantifying the effect of microsporidiosis on D. coccinellae development and host preference. Uninfected wasps were provided either uninfected or T. hippodamiae-infected beetles as hosts and the development of their progeny was observed over 30 days. The duration of endoparasitoid development from egg deposition in the host until adult eclosion for D. coccinellae did not differ significantly, regardless of the infection status of the host beetle. All wasp progeny that developed within, and emerged from, T. hippodamiae-infected beetles were infected with the microsporidian pathogen (n = 48; 100% transmission). Infected D. coccinellae adults were also provided either uninfected or T. hippodamiae infected host beetles so that the development of their progeny could be assessed over 30 days. Endoparasitoid development did not differ significantly; however, a significantly greater proportion of beetles stung by microsporidia-infected wasps did not contain an endoparasitoid larva when dissected at the end of the 30-day trial when compared to those stung by uninfected wasps. This suggests that the pathogen may reduce wasp fecundity or egg viability. Examination of paraffin-embedded D. coccinellae adult tissues revealed an extensive microsporidian infection throughout all major organs and tissues with exception of the ovary. During host choice trials, uninfected and microsporidia-infected D. coccinellae adults pursued, took an ovipositional stance, and attacked uninfected beetles more often than microsporidia-infected hosts but these observations did not differ significantly (P > 0.05).

The effects of two microsporidian pathogens on the two-spotted lady beetle, Adalia bipunctata L. (Coleoptera: Coccinellidae).

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.

Ultrastructure and molecular characterization of a microsporidium, Tubulinosema hippodamiae, from the convergent lady beetle, Hippodamia convergens Guérin-Méneville.

Hippodamia convergens, the convergent lady beetle, is available for aphid control in home gardens and in commercial food production systems throughout the United States and Canada. Beetles received from commercial insectaries for biological control are occasionally infected with a microsporidium. 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 in direct contact with the host cell cytoplasm. Early developmental stages were proximal to mature spores and both were observed throughout the tissue sections that were examined. Merogony resulted from binary fission. Early-stage sporoblasts were surrounded by a highly convoluted plasma membrane and contained an electron-dense cytoplasm and diplokaryon. Ovoid to elongated late-stage sporoblasts were surrounded by a relatively complete spore wall. The polar filament, polaroplast, and anchoring disk were readily observed within the cell cytoplasm. Mature spores were typical of terrestrial microsporidia, with a thickened endospore surrounded by a thin exospore. Spores contained well-defined internal structures, including a diplokaryon, lamellar polaroplast and a slightly anisofilar polar filament with 10-14 coils arranged in a single or double row. A prominent indentation was evident at the apical end of the spore wall proximal to the anchoring disk. Aberrant spores were also observed. These had a fully developed endospore and exospore but lacked any discernable internal spore structures, and were, instead, filled with lamellar or vesicular structures. Typical and aberrant spores measured 3.58 ± 0.2 × 2.06 ± 0.2 µm (n=10) and 3.38 ± 0.8 × 2.13 ± 0.2 µm (n=10), respectively. Spores were observed in longitudinal muscle surrounding the midgut and within the fat body, Malpighian tubules, pyloric valve epithelium, ventral nerve cord ganglia, muscles and ovaries. The hindgut epithelium was often infected but the connective tissues were rarely invaded. The life cycle and pathology of the microsporidium bears some resemblance to Nosema hippodamiae, the only microsporidium reported from H. convergens by Lipa and Steinhaus in 1959. Molecular characterization of the pathogen genomic DNA revealed that it is 99% similar to Tubulinosema acridophagus and T. ratisbonensis, two pathogens that infect Drosophila melanogaster and 98% similar to T. kingi from D. willistoni. Based on similarities in pathogen ultrastructure and the molecular information gained during this study, we propose that the microsporidium in H. convergens be given the name Tubulinosema hippodamiae.

Effects of a microsporidium from the convergent lady beetle, Hippodamia convergens Guérin-Méneville (Coleoptera: Coccinellidae), on three non-target coccinellids.

A microsporidium from Hippodamia convergens was transmitted horizontally to three non-target coccinellid hosts (Adalia bipunctata L., Coccinella septempunctata L. and Harmonia axyridis Pallas) under laboratory conditions. For all species examined, microsporidia-infected larvae took significantly longer to develop than did uninfected larvae but the microsporidium had no effect on larval mortality. Adult sex ratios of uninfected and microsporidia-infected adults were about 1:1 (female symbol:male symbol) and did not differ significantly. At the end of a 90-day trial, microsporidia-infected H. convergens produced significantly fewer eggs and did not live as long as uninfected individuals. Differences in fecundity and longevity were not observed for the three non-target coccinellids that were examined. Mean spore counts from smear preparations of microsporidia-infected A. bipunctata did not differ significantly from H. convergens, suggesting that A. bipunctata (a native coccinellid) is a suitable host for the microsporidium but infection was lighter in C. septempunctata and H. axyridis (introduced species). Vertical transmission of the pathogen was observed during the 90-day trial by examining eggs and larvae that were produced by microsporidia-infected adults. For all species examined, 100% vertical transmission of the pathogen was eventually observed. Three eugregarines were found in two adult A. bipunctata: Gregarine A trophozoites are similar in size to those of Gregarina katherina Watson (described earlier from Coccinella spp.), Gregarine B trophozoites are similar in size to those of Gregarine A but are morphology distinct, and Gregarine C trophozoites are similar in size to G. barbarara Watson (described earlier from A. bipunctata).

Effects of an unidentified microsporidium on the convergent lady beetle, Hippodamia convergens Guérin-Méneville (Coleoptera: Coccinellidae), used for biological control.

Convergent lady beetles, Hippodamia convergens Guérin-Méneville, are collected from overwintering sites in California and redistributed for aphid control in home gardens and agroecosystems. The effects of an unidentified microsporidium on the life history characteristics of commercially available H. convergens were examined. Mean development for microsporidia-infected and uninfected H. convergens was 15.40+/-0.14 and 14.76+/-0.16 days, respectively (P=0.01). Larval mortality did not differ significantly. Cumulative mean egg production for microsporidia-infected and uninfected females was 545.8+/-92.6 and 928.3+/-86.4 eggs, respectively (P=0.004) and mean survival was 64.5+/-5.6 and 77.1+/-4.5 days, respectively (P=0.04). Microsporidian spores (3.6x2.4 microm) are similar in size to those of Nosema hippodamiae.

Disease prevalence and transmission of Microsporidium phytoseiuli infecting the predatory mite, Phytoseiulus persimilis (Acari: Phytoseiidae).

Isolated colonies of the predatory mite, Phytoseiulus persimilis, were used to gain information regarding prevalence and transmission of Microsporidium phytoseiuli. Two colonies of P. persimilis were reared on spider mite (Tetranychus urticae)-infested bean plants in isolated cages. Disease prevalence of predators from Colony 1 remained relatively low (between 0 and 15%) over 57 weeks of observation whereas disease prevalence of predators from Colony 2 increased over 3 months (from 12 to 100%). Disease prevalence among predators from Colony 1 had increased to 100% 2 months after weekly sampling had ceased for this colony and periodic sampling confirmed that disease prevalence among individuals of both colonies remained at 100%. Microsporidian spores were not detected in randomly chosen samples of T. urticae prey mites that were removed and examined biweekly during this period. Although numerous microsporidian spores were observed in smear preparations of fecal pellets examined by light microscopy, spores were not observed on leaf surfaces or predator feces when examined by SEM. The latter appeared as intact aggregates composed of numerous dumbbell-shaped crystals and it is unlikely that spores are liberated from intact fecal pellets onto leaf surfaces. Vertical transmission of M. phytoseiuli was 100%; horizontal transmission was low (14.3%) and occurred only when immature P. persimilis were permitted to develop in contact with infected immature and adult predators. The mean number of eggs produced per mated pair was highest when uninfected females were mated with uninfected males (63.2 eggs per mated pair). Although mean egg production decreased when one or both parents were infected, not all differences were significant. Male predatory mites did not contribute to infection of their progeny. Results suggest that routine examination of P. persimilis for microsporidian spores is essential for the management of M. phytoseiuli within P. persimilis colonies. Low disease prevalence and lack of obvious disease signs or symptoms, as in the case of M. phytoseiuli, increase the probability that these pathogens will escape notice unless individuals are routinely examined for pathogens.

Once daily, high dose versus divided, low dose gentamicin for open fractures.

A prospective, randomized study was performed on 75 Gustilo Grades II and III open fractures to determine the efficacy of once daily, high dose aminoglycoside therapy, compared with more conventional dosing, in reducing the infection rate when used in conjunction with an aggressive operative treatment protocol. All patients enrolled in the study were treated with immediate irrigation, debridement, operative stabilization of the fracture, and 1 g of cefazolin every 8 hours. At the time of admission patients were randomized to two groups. Patients in Group I received gentamicin 5 mg/kg divided into twice daily doses, and patients in Group II received gentamicin 6 mg/kg given once daily. All patients were monitored for renal toxicity and observed for radiographic and clinical signs of infection until fracture union. The results of the study revealed no statistically significant difference between once daily, high dose versus divided, low dose gentamicin in infection rates. Thus, daily dosing of gentamicin was found to be safe, effective, and cost efficient in the treatment of open fractures when combined with a cephalosporin and aggressive operative debridement and stabilization.

Candida in pancreatic infection: a clinical experience.

Pancreatic infection remains a significant clinical problem, with substantial morbidity and mortality. Published case reports of Candida species identified in these infections prompted a review of 17 consecutive patients recently treated for peripancreatic infection by scheduled relaparotomy. Six patients were transferred from other hospitals, all having undergone prior operative intervention (median stay elsewhere: 58 days). The 11 other patients underwent initial operation an average of 14 days after admission. Candida species were identified in the initial operative cultures of 5 patients (29%), three of whom had undergone previous drainage at other hospitals. Two patients (11.7%) had Candida identified at subsequent operation. Six patients were treated with Amphotericin B for a median of 12 days (range 6-32) and a median dosage of 420 mg (range 225-830 mg). All patients were cleared of their Candida infection, but three subsequently died, for an overall mortality of 17.6%. Candida infected patients suffered a 42 per cent mortality. Our series supports the suspicion that Candida is much more frequent (41% of patients) than previously recognized in peripancreatic sepsis, and is commonly acquired after the initial operation. Amphotericin B therapy is effective in clearing Candida infection, but affected patients have a high associated mortality.