Infected Bees Research Articles

Detection of honey bee viruses in larvae of Vespa orientalis.

The Oriental hornet (Vespa orientalis) is one of the major predators of honey bees. It has been demonstrated that adults of V. orientalis can harbor honey bee viruses, however the transmission route of infection is still not clear. The aim of this study was to study the possible presence of honey bee viruses in V. orientalis larvae and honey bees collected from the same apiary. Therefore, 29 samples of V. orientalis larvae and 2 pools of honey bee (Apis mellifera). samples were analyzed by multiplex PCR to detect the presence of six honeybee viruses: Acute Bee Paralysis Virus (ABPV), Black Queen Cell Virus (BQCV), Chronic Bee Paralysis Virus (CBPV), Deformed Wing Virus (DWV), Kashmir Bee Virus (KBV) and Sac Brood Virus (SBV). Biomolecular analysis of V. orientalis larvae revealed that DWV was present in 24/29 samples, SBV in 10/29, BQCV in 7/29 samples and ABPV in 5/29 samples, while no sample was found positive for CBPV or KBV. From biomolecular analysis of honey bee samples DWV was the most detected virus, followed by SBV, BQCV, ABPV. No honey bee sample was found positive for CBPV or KBV. Considering the overlapping of positivities between V.orientalis larvae and honey bee samples, and that V.orientalis larvae are fed insect proteins, preferably honey bees, we can suggest the acquisition of viral particles through the ingestion of infected bees. However, future studies are needed to confirm this hypothesis and rule out any other source of infection.

Microsporidiosis Causing Necrotic Changes in the Honeybee Intestine

Background: Microsporidia from the Nosema (Vairimorpha) genus are pathogenic fungi that complete their life cycle in the honeybee intestine. Therefore, the aim of this study was to determine the impact of the course of infection on the viability of honeybee intestine cells. Methods and Results: Intestines isolated from healthy and N. ceranae-infected honeybees were stained using two dyes, SYTO 9 and propidium iodide, and analyzed under an Axiovert 200M fluorescence microscope immediately after the isolation of the intestines. The ImageJ program was used for the quantitative analysis of the cell structure parameters. Our study demonstrated for the first time that healthy bees have a higher number of live cells in their intestines than infected bees, and that the intestines of N. ceranae-infected honeybees contain dead cells concentrated in spots. The results obtained for these two cases differed significantly, and were confirmed by statistical tests. Conclusions: The intestines of infected honeybees contain dead cells concentrated in red/dead spots, which can lead to necrotic changes, the interruption of the host’s intestinal continuity, intestinal leaking and the increased mortality of the host.

Differential bumble bee gene expression associated with pathogen infection and pollen diet

BackgroundDiet and parasitism can have powerful effects on host gene expression. However, how specific dietary components affect host gene expression that could feed back to affect parasitism is relatively unexplored in many wild species. Recently, it was discovered that consumption of sunflower (Helianthus annuus) pollen reduced severity of gut protozoan pathogen Crithidia bombi infection in Bombus impatiens bumble bees. Despite the dramatic and consistent medicinal effect of sunflower pollen, very little is known about the mechanism(s) underlying this effect. However, sunflower pollen extract increases rather than suppresses C. bombi growth in vitro, suggesting that sunflower pollen reduces C. bombi infection indirectly via changes in the host. Here, we analyzed whole transcriptomes of B. impatiens workers to characterize the physiological response to sunflower pollen consumption and C. bombi infection to isolate the mechanisms underlying the medicinal effect. B. impatiens workers were inoculated with either C. bombi cells (infected) or a sham control (un-infected) and fed either sunflower or wildflower pollen ad libitum. Whole abdominal gene expression profiles were then sequenced with Illumina NextSeq 500 technology.ResultsAmong infected bees, sunflower pollen upregulated immune transcripts, including the anti-microbial peptide hymenoptaecin, Toll receptors and serine proteases. In both infected and un-infected bees, sunflower pollen upregulated putative detoxification transcripts and transcripts associated with the repair and maintenance of gut epithelial cells. Among wildflower-fed bees, infected bees downregulated immune transcripts associated with phagocytosis and the phenoloxidase cascade.ConclusionsTaken together, these results indicate dissimilar immune responses between sunflower- and wildflower-fed bumble bees infected with C. bombi, a response to physical damage to gut epithelial cells caused by sunflower pollen, and a strong detoxification response to sunflower pollen consumption. Identifying host responses that drive the medicinal effect of sunflower pollen in infected bumble bees may broaden our understanding of plant-pollinator interactions and provide opportunities for effective management of bee pathogens.

Pollen as Bee Medicine: Is Prevention Better than Cure?

To face environmental stressors such as infection, animals may display behavioural plasticity to improve their physiological status through ingestion of specific food. In bees, the significance of medicating pollen may be limited by their ability to exploit it. Until now, studies have focused on the medicinal effects of pollen and nectar after forced-feeding experiments, overlooking spontaneous intake. Here, we explored the medicinal effects of different pollen on Bombus terrestris workers infected by the gut parasite Crithidia bombi. First, we used a forced-feeding experimental design allowing for the distinction between prophylactic and therapeutic effects of pollen, considering host tolerance and resistance. Then, we assessed whether bumble bees favoured medicating resources when infected to demonstrate potential self-medicative behaviour. We found that infected bumble bees had a lower fitness but higher resistance when forced to consume sunflower or heather pollen, and that infection dynamics was more gradual in therapeutic treatments. When given the choice between resources, infected workers did not target medicating pollen, nor did they consume more medicating pollen than uninfected ones. These results emphasize that the access to medicating resources could impede parasite dynamics, but that the cost-benefit trade-off could be detrimental when fitness is highly reduced.

Small hive beetle (Aethina tumida) threat on the horizon

Beekeeping is an agricultural industry that deals with breeding bees, obtaining honey, wax, and other products. Bees are used for pollination of many types of crops to increase their harvest. Beekeeping products are also used in medicine, pharmacology, chemical, and food industries. One of the essential aspects of successful beekeeping development is ensuring sustainable epizootic well-being from infectious and invasive bee diseases in Ukraine. About 3 million bee colonies are owned by private farms and amateur beekeepers, which give approximately 50-60 thousand tons of commercial honey and 1.2–1.4 thousand tons of wax per year at the current stage of beekeeping development in Ukraine. Honey and bee products from Ukraine received 15 prestigious awards in various categories at the 77th National Honey Show, held in London among many countries in 2008. In recent years, there has been a constant increase in the threat of new little-known invasive and infectious diseases that pose a significant danger to beekeeping worldwide. The invasion of the small hive beetle Aethina tumida belongs to quarantine and hazardous invasive diseases, which have destroyed many bee colonies in several countries worldwide (USA, Canada, African countries, Australia, some European countries – Italy, and Portugal). There are no effective methods of preventing and controlling the small hive beetle. The only way to prevent the invasion of the small hive beetle in Ukraine is constant monitoring and control at the state border of the movement of bee packages and bee products, which may pose a threat to introducing this invasive species into Ukraine.

The effects of thymol, oxalic acid (Api-Bioxal) and hops extract (Nose-Go) on viability, the Nosema sp. spore load and the expression of vg and sod-1 genes in infected honey bees

This study was done to investigate the effects of thymol, fumagillin, oxalic acid (Api-Bioxal) and hops extract (Nose-Go) on Nosema sp. spore load, the expression of vitellogenin (vg) and superoxide-dismutase-1 (sod-1) genes and mortality of bees infected with N. ceranae. Five healthy colonies were assigned as the negative control, and 25 Nosema sp. infected colonies were assigned to five treatment groups including: the positive control: no additive to sirup; fumagillin 26.4 mg/L, thymol 0.1 g/L, Api-Bioxal 0.64 g/L and Nose-Go 5.0 g/L sirup. The reduction in the number of Nosema sp. spores in fumagillin, thymol, Api-Bioxal and Nose-Go compared to the positive control was 54, 25, 30 and 58%, respectively. Nosema sp. infection in all infected groups increased (p < .05) Escherichia coli population compared to the negative control. Nose-Go had a negative effect on lactobacillus population compared to other substances. Nosema sp. infection decreased vg and sod-1 genes expression in all infected groups compared to the negative control. Fumagillin and Nose-Go increased the expression of vg gene, and Nose-Go and thymol increased the expression of sod-1 gene than the positive control. Nose-Go has the potential to treat nosemosis if the necessary lactobacillus population is provided in the gut.

Virus transmission via honey bee prey and potential impact on cocoon-building in labyrinth spiders (Agelena labyrinthica).

Interspecies transmission of RNA viruses is a major concern for human and animal health. However, host-range, transmission routes and especially the possible impact of these viruses on alternative hosts are often poorly understood. Here, we investigated the role of the labyrinth spider, Agelena labyrinthica, as a potential alternative host of viruses commonly known from western honey bees, Apis mellifera. Field-collected spiders were screened for Acute bee paralysis virus (ABPV), Black queen cell virus, Chronic bee paralysis virus, Deformed wing virus type A and B (DWV-B), Israeli acute paralysis virus, Lake Sinai virus and Sacbrood virus. In a laboratory experiment, labyrinth spiders were fed with ABPV and DWV-B infected honey bees or virus free control food. Our results show that natural infections of A. labyrinthica with these viruses are common in the field, as 62.5% of the samples were positive for at least one virus, supporting their wide host range. For DWV-B, the laboratory data indicate that foodborne transmission occurs and that high virus titres may reduce cocoon building, which would be the first report of clinical symptoms of DWV in Araneae. Since cocoons are tokens of fitness, virus transmission from honey bees might affect spider populations, which would constitute a concern for nature conservation.

Unraveling the Bombus terrestris Hemolymph, an Indicator of the Immune Response to Microbial Infections, through Complementary Mass Spectrometry Approaches

Pollinators, including Bombus terrestris, are crucial for maintaining biodiversity in ecosystems and for agriculture. Deciphering their immune response under stress conditions is a key issue for protecting these populations. To assess this metric, we analyzed the B. terrestris hemolymph as an indicator of their immune status. Hemolymph analysis was carried out using mass spectrometry, MALDI molecular mass fingerprinting was used for its effectiveness in assessing the immune status, and high-resolution mass spectrometry was used to measure the impact of experimental bacterial infections on the "hemoproteome". By infecting with three different types of bacteria, we observed that B. terrestris reacts in a specific way to bacterial attacks. Indeed, bacteria impact survival and stimulate an immune response in infected individuals, visible through changes in the molecular composition of their hemolymph. The characterization and label-free quantification of proteins involved in specific signaling pathways in bumble bees by bottom-up proteomics revealed differences in protein expression between the non-experimentally infected and the infected bees. Our results highlight the alteration of pathways involved in immune and defense reactions, stress, and energetic metabolism. Lastly, we developed molecular signatures reflecting the health status of B. terrestris to pave the way for diagnosis/prognosis tools in response to environmental stress.

Effect of bee bread on Africanized honey bees infected with spores of Nosema spp.

Floral diversity allows honey bees, Apis mellifera L. (including its hybrids between subspecies) (Hymenoptera: Apidae), to meet their nutritional requirements. However, infection by Nosema spp. (Microsporidia: Nosematidae) destroys a bee's digestive epithelium, hindering the ability to absorb nutrients. In some subtropical regions, such as the Yucatan Peninsula, Mexico, nosemosis is one of the most widespread diseases in commercial apiaries. To date, there have been no reports of colony deaths attributable to the disease. It is possible that the naturally occurring diet in the Yucatan Peninsula sustains honey bee health. We tested this hypothesis by evaluating the effect of four diets – bee bread collected in the Yucatan's rainy and dry seasons, sucrose syrup, and a pollen substitute – on the response of caged Africanized honey bees (A. m. scutellata × A. m. adansonii × A. mellifera) to nosemosis infection (treatment with spores of Nosema apis Zander and Nosema ceranae Fries). We found that infected and uninfected bees consumed food in similar amounts regardless of the diet, although, overall, they fed at significantly lower rates when provided the pollen-substitute diet. Bees fed either of the bee bread diets had relatively low fat-body weight and expression of Vg, proPO, and GOx genes, three genes related to immune function. Bees fed sucrose syrup had low fat-body weight but relatively high gene expression. Infection from Nosema spores only reduced the individual survival probability for bees fed the bee bread collected in the dry season, yet bees fed the dry-season bee bread had the lowest infection rates. Our results indicate that natural stored bee bread in honey bee colonies from the Yucatan Peninsula could be one of the main factors influencing their survival and reducing Nosema spp. spore load.

Laboratory Cultivation of Vairimorpha (Nosema) ceranae (Microsporidia: Nosematidae) in Artificially Infected Worker Bees.

Nosemosis type C is a dangerous and widespread disease of the adult European honey bee Apis mellifera and is caused by the spore-forming intracellular parasite Vairimorpha (Nosema) ceranae. The search for new ways of therapy for this disease is complicated due to the seasonal availability of V. ceranae-infected insects as well as the lack of a developed system for the pathogen's cultivation. By carrying out trials which used different infectious dosages of the parasite, spore storage protocols, host age, and incubation temperatures, we present a simple, safe, and efficient method of V. ceranae propagation in artificially infected worker bees in the laboratory. The method is based on feeding the groups of adult worker bees with microsporidian spores and insect maintenance in plastic bottles at 33 °C. The source of the spores originated from the cadavers of infected insects from the previous round of cultivation, in which the infective spores persist for up to six months. An analysis of five independent cultivation rounds involving more than 2500 bees showed that the proposed protocol exploiting the dosage of one million spores per bee yielded over 60 million V. ceranae spores per bee, and most of the spore samples can be isolated from living insects.

First Detection of Honeybee Pathogenic Viruses in Butterflies.

Simple SummaryA decline in the number and diversity of pollinators has been observed worldwide. Various butterfly species are also threatened by extinction. One of the less studied reasons for the pollinator decline are pathogens, although it is known that they can be transmitted between cultivated and some wild pollinator species. The objective of our study was to determine whether pathogenic honeybee viruses can be transmitted to butterflies. Butterfly and bee samples from four locations in Slovenia were analyzed using the molecular method RT-PCR. We considered six bee viruses and found very low levels of four of them, i.e., acute bee paralysis virus (ABPV), black queen cell virus (BQCV), Lake Sinai virus 3 (LSV3), and Sacbrood bee virus (SBV), in the butterfly samples compared to the bee samples, although these viruses were present in large amounts in honeybees from the same locations. The results of our study indicate that butterflies are probably contaminated with bee viruses when they visit the same flowers as honeybees. Therefore, based on the very low viral load, we believe that butterflies are unlikely to be threatened by bee viruses and that they are not important as vectors of viral infection to other pollinators.Several pathogens are important causes of the observed pollinator decline, some of which could be transmitted between different pollinator species. To determine whether honeybee viruses can be transmitted to butterflies, a total of 120 butterflies were sampled at four locations in Slovenia. At each location, butterflies from three families (Pieridae, Nymphalidae, Hesperiidae/Lycenidae) and Carniolan honeybees (Apis mellifera carnica) were collected. The RNA of six honeybee viruses, i.e., acute bee paralysis virus (ABPV), black queen cell virus (BQCV), chronic bee paralysis virus (CBPV), deformed wing virus A (DWV-A), Sacbrood bee virus (SBV), and Lake Sinai virus 3 (LSV3), was detected by a specific quantitative method (RT-PCR). The presence of ABPV, BQCV, LSV3, and SBV was detected in both butterflies and honeybees. All butterfly and bee samples were negative for CBPV, while DWV-A was detected only in honeybees. The viral load in the positive butterfly samples was much lower than in the positive bee samples, which could indicate that butterflies are passive carriers of bee viruses. The percentage of positive butterfly samples was higher when the butterflies were collected at sampling sites with a higher density of apiaries. Therefore, we believe that infected bees are a necessary condition for the presence of viruses in cohabiting butterflies. This is the first study on the presence of pathogenic bee viruses in butterflies.

A mathematical model for the spread of Varroa mites in honeybee populations: two simulation scenarios with seasonality

We formulate and study a mathematical model for a honeybee colony infected with Varroa mites which describes the parallel phenomena of the spread of both the mites and the virus transmitted by them. We extend our previous model by including infected forager bees and considering model parameters as time-periodic functions. Firstly, we study the autonomous model and show the stability of equilibria. We present two simulation scenarios to study the impact of seasonality on the spread of Varroa mites and the disease they carry. Numerical studies are given to show how the parameter changes might lead to the colony's failure.

Effect of amide protoporphyrin derivatives on immune response in Apis mellifera

The intracellular microsporidian parasite Nosema ceranae is known to compromise bee health by induction of energetic stress and downregulation of the immune system. Porphyrins are candidate therapeutic agents for controlling Nosema infection without adverse effects on honeybees. In the present work, the impact of two protoporphyrin IX derivatives, i.e. PP[Asp]2 and PP[Lys]2, on Apis mellifera humoral immune response has been investigated in laboratory conditions in non-infected and N. ceranae-infected honeybees. Fluorescence spectroscopy analysis of hemolymph showed for the first time that porphyrin molecules penetrate into the hemocoel of honeybees. Phenoloxidase (PO) activity and the expression of genes encoding antimicrobial peptides (AMPs: abaecin, defensin, and hymenoptaecin) were assessed. Porphyrins significantly increased the phenoloxidase activity in healthy honeybees but did not increase the expression of AMP genes. Compared with the control bees, the hemolymph of non-infected bees treated with porphyrins had an 11.3- and 6.1-fold higher level of PO activity after the 24- and 48-h porphyrin administration, respectively. Notably, there was a significant inverse correlation between the PO activity and the AMP gene expression level (r = − 0.61696, p = 0.0143). The PO activity profile in the infected bees was completely opposite to that in the healthy bees (r = − 0.5118, p = 0.000), which was related to the changing load of N. ceranae spores in the porphyrin treated-bees. On day 12 post-infection, the spore loads in the infected porphyrin-fed individuals significantly decreased by 74%, compared with the control bees. Our findings show involvement of the honeybee immune system in the porphyrin-based control of Nosema infection. This allows the infected bees to improve their lifespan considerably by choosing an optimal PO activity/AMP expression variant to cope with the varying level of N. ceranae infection.

Use of Thymol in Nosema ceranae Control and Health Improvement of Infected Honey Bees.

Simple SummaryIn the European Union, there is no registered product for the control of the honey bee endoparasite Nosema ceranae. Thus, researchers are looking for options for Nosema treatment. The aim of this study was to investigate the effect of a natural essential-oil ingredient (thymol) derived from Thymus vulgaris on honey bees infected with N. ceranae. Thymol exerted certain positive effects (increasing bee survival, immunity, and antioxidative protection), as well as positively affecting the spore loads in Nosema-infected bees. However, when applied to Nosema-free bees, thymol caused certain health disorders; therefore, beekeepers should be careful with its use.Nosema ceranae is the most widespread microsporidian species which infects the honey bees of Apis mellifera by causing the weakening of their colonies and a decline in their productive and reproductive capacities. The only registered product for its control is the antibiotic fumagillin; however, in the European Union, there is no formulation registered for use in beekeeping. Thymol (3-hydroxy-p-cymene) is a natural essential-oil ingredient derived from Thymus vulgaris, which has been used in Varroa control for decades. The aim of this study was to investigate the effect of thymol supplementation on the expression of immune-related genes and the parameters of oxidative stress and bee survival, as well as spore loads in bees infected with the microsporidian parasite N. ceranae. The results reveal mostly positive effects of thymol on health (increasing levels of immune-related genes and values of oxidative stress parameters, and decreasing Nosema spore loads) when applied to Nosema-infected bees. Moreover, supplementation with thymol did not induce negative effects in Nosema-infected bees. However, our results indicate that in Nosema-free bees, thymol itself could cause certain disorders (affecting bee survival, decreasing oxidative capacity, and downregulation of some immune-related gene expressions), showing that one should be careful with preventive, uncontrolled, and excessive use of thymol. Thus, further research is needed to reveal the effect of this phytogenic supplement on the immunity of uninfected bees.

Mathematical modelling of between hive transmission of Nosemosis by drifting

Nosema ceranae is a microsporidian parasite of the Western honeybee (Apis mellifera). Building on a previous model of within-hive transmission we formulate a mathematical model for the spread of this disease between hives in an apiary. The underlying honeybee population model considers hive and forager bees separately; to account for seasonal variations in honeybee and disease biology, the model parameters vary in time with a period of one year. In the hive, two transmission routes are considered, one is indirect (infected hive bees deposit Nosema spores that are ingested by other bees), the other one is direct and follows the usual mass action kinetics of SIR models. Inter colony transmission is assumed to be caused by drifting. The resulting ecoepidemiological metapopulation model is studied numerically. Our simulations suggest that in the case of indirect within-hive transmission the longterm behaviour is not very sensitive to drifting intensity, and that across the apiary the individual hives synchronise and stratify quickly. In the case of direct within-hive transmission very irregular and complex behaviour can be observed, for example some colonies might survive in working order whereas neighbouring hives might fail. This is due to the underlying single hive model. If both within-hive transmission routes are combined the solutions of the metapopulation model, depending on parameters, can show facets of both transmission routes.

Effect of Nosema ceranae infection and season on the gut bacteriome composition of the European honeybee (Apis mellifera)

Nosema ceranae is an intracellular parasite that infects honeybees’ gut altering the digestive functions; therefore, it has the potential of affecting the composition of the gut microbiome. In this work, individual bees of known age were sampled both in spring and autumn, and their digestive tracts were assessed for N. ceranae infection. Intestinal microbiome was assessed by sequencing the bacterial 16S rRNA gene in two different gut sections, the anterior section (AS; midgut and a half of ileum) and the posterior section (PS; second half of ileum and rectum). A preliminary analysis with a first batch of samples (n = 42) showed that AS samples had a higher potential to discriminate between infected and non-infected bees than PS samples. As a consequence, AS samples were selected for subsequent analyses. When analyzing the whole set of AS samples (n = 158) no changes in α- or β-diversity were observed between infected and non-infected bees. However, significant changes in the relative abundance of Proteobacteria and Firmicutes appeared when a subgroup of highly infected bees was compared to the group of non-infected bees. Seasonality and bees’ age had a significant impact in shaping the bacteriome structure and composition of the bees’ gut. Further research is needed to elucidate possible associations between the microbiome and N. ceranae infection in order to find efficient strategies for prevention of infections through modulation of bees’ microbiome.

Consuming sunflower pollen reduced pathogen infection but did not alter measures of immunity in bumblebees.

Certain diets can benefit bee health by reducing pathogens, but the mechanism(s) driving these medicinal effects are largely unexplored. Recent research found that sunflower (Helianthus annuus) pollen reduces the gut pathogen Crithidia bombi in the common eastern bumblebee (Bombus impatiens). Here, we tested the effects of sunflower pollen and infection on two bee immune metrics to determine whether sunflower pollen diet drives changes in host immunity that can explain this medicinal effect. Bees were infected with C. bombi or not and given either sunflower or wildflower pollen. Subsequently, bees received a benign immune challenge or were left naive to test the induced and constitutive immune responses, respectively. We measured haemolymph phenoloxidase activity, involved in the melanization cascade, and antibacterial activity. Sunflower pollen reduced C. bombi infection, but we found no significant pollen diet effect on either immune measure. Phenoloxidase activity was also not affected by C. bombi infection status; however, uninfected bees were more likely to have measurable constitutive antibacterial activity, while infected bees had higher induced antibacterial activity. Overall, we found that sunflower pollen does not significantly affect the immune responses we measured, suggesting that the mechanisms underlying its medicinal effect do not involve these bee immune parameters. This article is part of the theme issue 'Natural processes influencing pollinator health: from chemistry to landscapes'.

Effect of Chronic Exposure to Sublethal Doses of Imidacloprid and Nosema ceranae on Immunity, Gut Microbiota, and Survival of Africanized Honey Bees.

Large-scale honey bee colony losses reported around the world have been associated with intoxication with pesticides, as with the presence of pests and pathogens. Among pesticides, neonicotinoid insecticides are the biggest threat. Due to their extensive use, they can be found in all agricultural environments, including soil, water, and air, are persistent in the environment, and are highly toxic for honey bees. In addition, infection by different pests and pathogens can act synergistically, weakening bees. In this study, we investigated the effects of chronic exposure to sublethal doses of imidacloprid alone or combined with the microsporidia Nosema ceranae on the immune response, deformed wing virus infection (DWV), gut microbiota, and survival of Africanized honey bees. We found that imidacloprid affected the expression of some genes associated with immunity generating an altered physiological state, although it did not favor DWV or N. ceranae infection. The pesticide alone did not affect honey bee gut microbiota, as previously suggested, but when administered to N. ceranae infected bees, it generated significant changes. Finally, both stress factors caused high mortality rates. Those results illustrate the negative impact of imidacloprid alone or combined with N. ceranae on Africanized honey bees and are useful to understand colony losses in Latin America.

Genomic characterisation of an entomopathogenic strain of Serratia ureilytica in the critically endangered phasmid Dryococelus australis.

Between 2014 and 2019, unexpected mortalities were observed in a colony of Dryococelus australis, an endangered stick-insect kept at the Melbourne Zoo for a breeding and conservation program. Pure cultures of Serratia spp. were obtained from the haemolymph of moribund and recently deceased individuals. The combined bacteriological and histopathological observations suggested an infectious cause of these mortalities. Genotyping of Serratia sp. isolated from the insects and their environment revealed a predominant strain profile. A representative isolate, AM923, was entirely sequenced and compared to 616 publicly available Serratia spp. genomes, including 37 associated with insects. The genomes were distributed into 3 distinct groups, with 63% of the insect-associated isolates within a single clade (clade A) containing AM923, separated from most environmental/plant-associated strains (clade B) and human isolates (clade C). Average nucleotide identity and phylogenetic analyses identified AM923 as S. ureilytica and revealed similarities with putatively entomopathogenic strains. An experimental infection model in honey bees (Apis mellifera) confirmed the pathogenic potential of AM923. A urease operon was found in most insect isolates and a PCR assay, based on the ureB gene sequence, was used to confirm the presence of AM923 in experimentally infected bees. This species-specific PCR could be applied to detect entomopathogenic Serratia spp. in infected insects or their environment.

Poison or Potion: Effects of Sunflower Phenolamides on Bumble Bees and Their Gut Parasite.

Simple SummaryBee declines have been reported worldwide, partly due to parasite spread induced by human activities. However, bees may forage on specific floral resources to face parasite infection. Such natural resources are comparable to ‘natural pharmacies’ and may be favoured in bee conservation strategies. Consumption of sunflower pollen, despite being detrimental for larval development, has been recently shown to reduce the load of a widespread bumble bee gut parasite in the common eastern bumble bee. Although the underlying mechanisms remain unknown, it has been suggested that sunflower phenolamides—a family of molecules found in most flowering plants—may be responsible for such a reduction in parasite load. Here, we tested the impacts of sunflower phenolamides on healthy and infected buff-tailed bumble bees. Expectedly, sunflower pollen had harmful consequences on bumble bee development but surprisingly, it did not alter parasite load. By contrast, sunflower phenolamides had milder effects on bumble bee development but unexpectedly increased parasite load. Phenolamide effects may stem from the physiological stress they induced or the gut microbial community alteration they may have triggered. Since biological models and experimental framework differ greatly in related studies tackling plant–bee–parasite interplays, we challenged the definition of medicinal effects and questioned the way to assess them in controlled conditions.Specific floral resources may help bees to face environmental challenges such as parasite infection, as recently shown for sunflower pollen. Whereas this pollen diet is known to be unsuitable for the larval development of bumble bees, it has been shown to reduce the load of a trypanosomatid parasite (Crithidia bombi) in the bumble bee gut. Recent studies suggested it could be due to phenolamides, a group of compounds commonly found in flowering plants. We, therefore, decided to assess separately the impacts of sunflower pollen and its phenolamides on a bumble bee and its gut parasite. We fed Crithidia-infected and -uninfected microcolonies of Bombus terrestris either with a diet of willow pollen (control), a diet of sunflower pollen (natural diet) or a diet of willow pollen supplemented with sunflower phenolamides (supplemented diet). We measured several parameters at both microcolony (i.e., food collection, parasite load, brood development and stress responses) and individual (i.e., fat body content and phenotypic variation) levels. As expected, the natural diet had detrimental effects on bumble bees but surprisingly, we did not observe any reduction in parasite load, probably because of bee species-specific outcomes. The supplemented diet also induced detrimental effects but by contrast to our a priori hypothesis, it led to an increase in parasite load in infected microcolonies. We hypothesised that it could be due to physiological distress or gut microbiota alteration induced by phenolamide bioactivities. We further challenged the definition of medicinal effects and questioned the way to assess them in controlled conditions, underlining the necessity to clearly define the experimental framework in this research field.