No impact of hygienic behavior and viral coinfection on the development of European foulbrood in honey bee (Apis mellifera) colonies during blueberry pollination in Michigan.

European foulbrood (EFB) is a severe disease of honey bee (Apis mellifera) larvae caused by the bacterium Linnaeus [Hymenoptera: Apidae]) Melissococcus plutonius (ex White) Bailey and Collins (Lactobacillales: Enterococcaceae). Many beekeepers in North America report severe EFB following blueberry pollination, but it is not clear what factors during pollination are related to clinical disease. Additionally, the impact that other factors such as viral load and hygienic behavior have on EFB has not been studied. In Spring of 2020 we enrolled 60 commercial honey bee colonies in a prospective cohort study. Colonies were inspected 3 times over the season with hive metrics and samples taken for viral testing. Each colony was tested for hygienic behavior twice and the score was averaged. Viral loads were determined by qPCR for deformed wing virus (DWV) A and B. We found no statistical difference in the EFB prevalence or severity between the 2 yards at any timepoint; 50% (n = 16) of the colonies in the holding yard and 63% (n = 17) in blueberry developed moderate to severe EFB over the study period. When colonies from both yards were pooled, we found no relationship between viral load or hygienic behavior and development of EFB. These results suggest that other factors may be responsible for driving EFB virulence and hygienic behavior is not likely helpful in managing this disease.

Mite non-reproduction, recapping behavior, and hygienic behavior (freeze-kill method) linked to Varroa destructor infestation levels in selected Apis mellifera colonies.

The genetic selection of honey bees (Apis mellifera) possessing specific social hygienic behaviors offers the beekeeping industry the possibility of controlling the Varroa destructor parasite and thus reducing its dependence on acaricides. However, the links between these behavioral traits are not yet well defined, which limits genetic progress in breeding programs. We measured the following behavioral varroa resistance traits: freeze-kill brood (FKB) and pin-kill brood (PKB) assays, varroa-sensitive hygiene (VSH), pupae removal, mite non-reproduction (MNR), and recapping activity. We found 2 negative and significant relationships: 1) between the recapping of cells infested with varroa and the total number of recapped cells, and 2) between the recapping of cells infested with varroa and VSH. We also selected the best predictive model of varroa infestation levels using the "step-wise" approach based on the Akaike information criterion. Our model revealed that MNR and FKB were significantly related to the varroa population levels, with a negative relationship; recapping was significantly related to mite infestation levels, with a positive relationship. Thus, a higher MNR or FKB score was linked to lower levels of mite infestation in colonies on August 14 (prior to fall infestation treatments); a higher recapping activity was linked to a higher level of mite infestation. Recapping behavior could be a useful trait to aid the selection of varroa-resistant bee lineages.

Observation of Genetic Gain with Instrumental Insemination of Honeybee Queens.

Controlling mating in the honeybee (Apis mellifera) is part of one of the greatest challenges for the beekeeping industry's genetic selection programs due to specific characteristics of their reproduction. Several techniques for supervising honeybee mating with relative effective control have been developed over the years to allow honeybee selection. As part of this project, we compared the genetic gains for several colony performance traits, obtained using the BLUP-animal method, according to the selection pressure applied in controlled reproduction (directed fertilization versus instrumental insemination). Our results show similar genetic gains for hygienic behavior and honey production between colonies whether queens were fertilized naturally or via instrumental insemination, as well as similar or lower genetic gains for colonies with queens inseminated for spring development. In addition, we noticed greater fragility in queens following insemination. These findings show that instrumental insemination is an effective tool for reproductive control in genetic selection and for estimating breeding values more precisely. However, this technique does not result in queens of superior genetic quality for commercial purposes.

A Reciprocal Transplant Experiment Confirmed Mite-Resistance in a Honey Bee Population from Uruguay.

In the past few years there has been an increasing interest for the study of honey bee populations that are naturally resistant to the ectoparasitic mite Varroa destructor, aiming to identify the mechanisms that allow the bees to limit the reproduction of the mite. In eastern Uruguay there are still bees resistant to mites that survive without acaricides. In order to determine if the differential resistance to V. destructor was maintained in other environments, a reciprocal transplant experiment was performed between the mite-resistant bee colonies and the mite-susceptible bee colonies from the east and the west of the country, respectively, infesting bees with local mites. In both regions, the mite-resistant colonies expressed a higher hygienic behavior and presented a higher phoretic mites/reproductive mites and mites in drone cells/mites in worker cells ratio than the mite-susceptible colonies. All the mite-susceptible colonies died during fall-winter, while a considerable number of mite-resistant colonies survived until spring, especially in the east of the country. This study shows that the bees in the east of the country maintain in good measure the resistance to V. destructor in other regions and leaves open the possibility that the mites of the two populations have biases in the reproductive behavior.

Tailoring Can Improve Consumers' Hygienic Food-Handling Behavior to Reduce the Transmission Risk of Antimicrobial-Resistant Bacteria through Food.

Antimicrobial-resistant (AMR) bacteria spread via food to humans and can seriously impair infection treatment. Hygienic food handling is an effective measure to avoid the transmission of bacteria. Therefore, we tested three types of interventions (tailored, generic, and no intervention) for their effectiveness in improving consumers' hygienic food handling against the spread of antimicrobial-resistant bacteria through foods in a longitudinal randomized control trial. We based the determinants of hygienic food-handling behavior on the Health Action Process Approach (HAPA). The tailored intervention raised self-reported hygienic food handling, self-efficacy, and perceived likelihood of risk compared to no intervention. Moreover, interventions yielded different effects for participants with high vs. low intentions to improve their food-handling behavior. However, there were no differences between the tailored and generic interventions. More research is needed to find out whether including other behavior change techniques in the tailored intervention may increase the effect of tailoring.

Medical Decision Style and COVID-19 Behavior.

Given the flood of health-related information stirred up by the coronavirus disease 2019 (COVID-19) pandemic, it is important to understand the factors that influence people to engage in protective public health measures so that medical communication can be tailored to be effective. Following the idea that people have a general inclination toward health care utilization, which is either more passive (i.e., medical minimizer) or more aggressive (i.e., medical maximizer), we assess if this inclination extends to being more or less willing to engage in protective public health behavior. We investigate the effect of individual differences in medical minimizing and medical maximizing orientation on COVID-19-related protective behaviors and attitudes. We used the validated Medical Maximizer-Minimizer Scale (MMS) and surveyed a diverse opt-in sample of the Swedish population (n = 806). Our results show that the MMS significantly predicts a wide range of self-reported behaviors and attitudes in relation to COVID-19. Participants with a stronger minimization orientation were significantly less likely to practice social distancing, follow hygiene recommendations, and be supportive of strict COVID-19 policies. Participants with a stronger maximization orientation had a larger discrepancy between perceived own risk and others getting infected. Thus, they perceived themselves as being less at risk for getting infected compared to the average person. Our findings imply that the MMS can be effectively used to predict who is more or less reluctant to follow public health recommendations.JEL codes: D70 E71 I12 I18.

Sublethal pesticide exposure induces larval removal behavior in honeybees through chemical cues.

We were intrigued by reported observations of reduced brood production and a high number of empty brood cells in bee colonies exposed to sublethal pesticide doses, which could suggest an active removal of larvae. Higher numbers of oenocytes, insect cells responsible for lipid processing and detoxification, were also found in pesticide-exposed larvae. Oenocytes are involved in hydrocarbon metabolism and chemical communication, and we hypothesized that these larvae could display altered cuticular hydrocarbon (CHC) profiles when exposed to pesticides as compared to control larvae. In addition, we proposed that these chemical cues could trigger specific behavioral responses in colony nurses. To test these hypotheses, we analyzed the CHC profiles of artificially reared larvae that had been fed sublethal doses of either dimethoate or clothianidin or fed on lipopolysaccharide (LPS) using gas chromatography-mass spectrometry. We found significant differences in the CHC profiles of these differently treated larvae. In a subsequent behavioral experiment, we transferred clothianidin-treated or LPS-treated larvae into the brood combs of surrogate colonies. Larvae that had been fed either the pesticide or LPS were removed at a significantly higher rate than control larvae. Our results demonstrate that larvae exposed to clothianidin possess altered CHC profiles, are detected in the colony by nurse bees via chemical cues and are actively removed.

Hygienic Behavior of Apis mellifera and Its Relationship with Varroa destructor Infestation and Honey Production in the Central Highlands of Ecuador.

The aim of this research was to analyze the relationship among hygienic behavior (HB), Varroa destructor infestation, and honey production in the central highlands of Ecuador. Overall, 75 honey bee colonies were evaluated before, during, and after production at three altitude levels (2600-2800, 2801-3000, and >3000 m.a.s.l.). The hygienic behavior percentage of the colonies was determined by the pin-killing method, and the colonies were classified into three groups: high HB (>85%), mid HB (60.1-85%), and low HB (≤60%). Varroa infestation was diagnosed as well, and honey production was evaluated only during production. HB was high and heterogeneous, averaging 80% ± 9.7%. Its highest expression was observed at lower altitudes. The infestation degree was low (3.47% ± 1.56%), although the mite was detected in all colonies upon sampling. A negative correlation was observed between HB and Varroa infestation in the first sampling (-0.49 **), suggesting that the high- and mid-altitude HB colonies underwent the lowest infestation rates, regardless of sampling. The correlations between HB and production were significant (0.26 *), indicating a positive effect of HB on production, meaning that colonies with high HB obtained the highest honey production (25.08 ± 4.82 kg/hive). The HB of bees showed an inverse relationship with altitude and it tended to reduce the effect of Varroa infestation, favoring honey production and, thus, suggesting the feasibility of selecting colonies with high HB.

Hygiene-Eliciting Brood Semiochemicals as a Tool for Assaying Honey Bee (Hymenoptera: Apidae) Colony Resistance to Varroa (Mesostigmata: Varroidae).

Despite numerous interventions, the ectoparasitic mite Varroa (Varroa destructor Anderson and Trueman [Mesostigmata: Varroidae]) and the pathogens it vectors remain a primary threat to honey bee (Apis mellifera Linnaeus [Hymenoptera: Apidae]) health. Hygienic behavior, the ability to detect, uncap, and remove unhealthy brood from the colony, has been bred for selectively for over two decades and continues to be a promising avenue for improved Varroa management. Although hygienic behavior is expressed more in Varroa-resistant colonies, hygiene does not always confer resistance to Varroa. Additionally, existing Varroa resistance selection methods trade efficacy for efficiency, because those achieving the highest levels of Varroa resistance can be time-consuming, and thus expensive and impractical for apicultural use. Here, we tested the hypothesis that hygienic response to a mixture of semiochemicals associated with Varroa-infested honey bee brood can serve as an improved tool for predicting colony-level Varroa resistance. In support of our hypothesis, we demonstrated that a mixture of the compounds (Z)-10-tritriacontene, (Z)-8-hentriacontene, (Z)-8-heptadecene, and (Z)-6-pentadecene triggers hygienic behavior in a two-hour assay, and that high-performing colonies (hygienic response to ≥60% of treated cells) have significantly lower Varroa infestations, remove significantly more introduced Varroa, and are significantly more likely to survive the winter compared to low-performing colonies (hygienic response to <60% of treated cells). We discuss the relative efficacy and efficiency of this assay for facilitating apiary management decisions and selection of Varroa-resistant honey bees, as well as the relevance of these findings to honey bee health, pollination services, and social insect communication.

The Impact of Trust and Risk Perception on the Acceptance of Measures to Reduce COVID-19 Cases.

The SARS-CoV-2 virus that causes COVID-19 has had a large impact on the lives of many people worldwide. At the peak of confirmed COVID-19 cases during the first wave in Switzerland (March-April 2020), we conducted a survey in the German-speaking part of the country (N = 1,585). The results suggest that the implemented measures are accepted. The survey participants are more concerned that other family members could catch the virus compared with themselves, and they are worried about its economic impact. The results suggest that how trust is measured is crucial because general trust and social trust have opposite effects on the participants' risk perceptions. People with high general trust perceive less risks associated with COVID-19 compared with people who have low general trust, and people with high social trust perceive more risks compared with people who have low social trust. The results further indicate that perceived risks are important drivers for the acceptance of the government's implemented measures to control COVID-19 and for more precautionary behavior (i.e., contact with fewer people and more hygienic behavior). Theoretical and practical implications of these findings are discussed.

Non-Destructive Genotyping of Honeybee Queens to Support Selection and Breeding.

In traditional bee breeding, the honeybee queen is chosen for breeding based on the performance of the colony produced by its mother. However, we cannot be entirely certain that a specific queen will produce offspring with desirable traits until we observe the young queen's new colony. Collecting the queen's genetic material enables quick and reliable determination of the relevant information. We sampled exuviae, feces, and wingtips for DNA extraction to avoid fatally injuring the queen when using tissue samples. Quantity and purity of extracted DNA were measured. Two mitochondrial markers were used to determine the lineage affiliation and exclude possible contamination of DNA extracts with non-honeybee DNA. dCAPS (derived Cleaved Amplified Polymorphic Sequences) markers allowed detection of single nucleotide polymorphisms (SNPs) in nuclear DNA regions presumably associated with Varroa sensitive hygiene and set the example of successful development of genotyping protocol from non-destructive DNA sources. One of the logical future steps in honeybee breeding is introducing genomic selection and non-destructive sampling methods of genetic material may be the prerequisite for successful genotyping. Our results demonstrate that the extraction of DNA from feces and exuviae can be introduced into practice. The advantage of these two sources over wingtips is reducing the time window for processing the samples, thus enabling genotyping directly after the queen's emergence.

Descriptive Analysis of the Varroa Non-Reproduction Trait in Honey Bee Colonies and Association with Other Traits Related to Varroa Resistance.

In the current context of worldwide honey bee colony losses, among which the varroa mite plays a major role, the hope to improve honey bee health lies in part in the breeding of varroa resistant colonies. To do so, methods used to evaluate varroa resistance need better understanding. Repeatability and correlations between traits such as mite non-reproduction (MNR), varroa sensitive hygiene (VSH), and hygienic behavior are poorly known, due to practical limitations and to their underlying complexity. We investigate (i) the variability, (ii) the repeatability of the MNR score, and (iii) its correlation with other resistance traits. To reduce the inherent variability of MNR scores, we propose to apply an empirical Bayes correction. In the short-term (ten days), MNR had a modest repeatability of 0.4, whereas in the long-term (a month), it had a low repeatability of 0.2, similar to other resistance traits. Within our dataset, there was no correlation between MNR and VSH. Although MNR is amongst the most popular varroa resistance estimates in field studies, its underlying complex mechanism is not fully understood. Its lack of correlation with better described resistance traits and low repeatability suggest that MNR needs to be interpreted cautiously, especially when used for selection.

Effect of Immune Inducers on Nosema ceranae Multiplication and Their Impact on Honey Bee (Apis mellifera L.) Survivorship and Behaviors.

Nosema disease is a major disease of honey bees caused by two species of microsporidia, Nosema apis and N. ceranae. Current control involves using antibiotics, which is undesirable because of possible antibiotic resistance and contamination. In this study, flagellin, zymosan, chitosan, and peptidoglycan were investigated as alternatives for controlling N. ceranae infections and for their effect on bee survivorship and behaviors. Chitosan and peptidoglycan significantly reduced the infection, and significantly increased survivorship of infected bees, with chitosan being more effective. However, neither compound altered the bees' hygienic behavior, which was also not affected by the infection. Chitosan significantly increased pollen foraging and both compounds significantly increased non-pollen foraging compared to healthy and infected bees. Memory retention, evaluated with the proboscis extension reflex assay, was temporarily impaired by chitosan but was not affected by peptidoglycan, nor was it affected by N. ceranae infection compared to the non-infected bees. This study indicates that chitosan and peptidoglycan provide benefits by partially reducing N. ceranae spore numbers while increasing survivorship compared to N. ceranae infected bees. Also, chitosan and peptidoglycan improved aspects of foraging behavior even more than in healthy bees, showing that they may act as stimulators of important honey bee behaviors.

Facing death: How does the subterranean termite Coptotermes gestroi (Isoptera: Rhinotermitidae) deal with corpses?

Social insects display a range of sophisticated behaviors to deal with cadavers, which together act to guarantee the health and social homeostasis of their colonies. In termites, cadaver management involves an entire repertoire of activities, such as antennation, grooming, retreat, alarm and agonistic behaviors. We performed different bioassays to evaluate whether postmortem-age, origin and caste of a cadaver influence the behavior of Coptotermes gestroi. Quantitative analyses of corpse management behaviors indicated that C. gestroi is able to discriminate the origin and caste of cadavers. Cadavers of worker nestmates were preferentially cannibalized while corpses of alien workers were buried. In addition, soldiers that had been dead for 24hours were buried while freshly dead soldiers were either buried, consumed or ignored, indicating a plastic behavioral response that depends on postmortem age. Corpse consumption is an important component of corpse management repertoire because this process eliminates contaminant and enables nutrient recycling. The physical isolation of the termite corpse through burial after grooming also helps to prevent other colony members from contacting the cadaver, further mitigating against the potential spread of disease.

Laurus nobilis L. Extracts against Paenibacillus larvae: Antimicrobial activity, antioxidant capacity, hygienic behavior and colony strength.

The aim of this work was to compare the antimicrobial activity against Paenibacillus larvae and the antioxidant capacity of two Laurus nobilis L. extracts obtained by different extraction methods. The hydroalcoholic extract was moreover added as supplementary diet to bees in field conditions to test behavioural effects and colony strength. Both laurel extracts were subjected to different phytochemical analysis to identify their bioactive compounds. Antimicrobial activity was analyzed by the minimal inhibitory concentration (MIC) determination by means the agar dilution method. The hydroalcoholic extract (HE) was able to inhibit the bacterial growth of all P. larvae strains, with 580 µg/mL mean value. This better antibacterial activity in relation to the essential oil (EO) could be explained by the presence of some phenolic compounds, such as flavonoids, evidenced by characteristic bands resulting from the Fourier Transform Infrared Spectroscopy (FTIR) analysis. Antioxidant activities of the extracts were evaluated by 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical-scavenging ability and ferric reducing antioxidant power (FRAP) assays. The HE showed the highest antioxidant activity as measured by DPPH, with IC50 values of 257 ±â€¯12 µg/mL. The FRAP assay method showed that the HE was 3-fold more effective reducing agent than the EO. When the bee colonies were supplied with laurel HE in sugar paste an improvement in their general condition was noticed, although neither the hygienic behavior nor the proportions of the breeding cells varied statistically due to the treatment. In conclusion, the inhibition power against P. larvae attributable to the phenolic compounds, the antioxidant capacity of the HE, and the non-lethal effects on adult honey bees on field trials suggest the HE of laurel as a promising substance for control American foulbrood disease.

Sanitary behavior in queenright and queenless ant colonies.

Waste disposal is important for maintaining the health of animal societies. Adults and off ;spring produce large amounts of waste and feces that could contain pathogens or toxins and may need to be stored away from the young or adult individuals. In social insects, the worker caste is responsible for nest maintenance, including sanitary behavior, and waste disposal strategies vary between species. However, individual task allocation is generally affected by queen presence and worker efficiency often decreases in the absence of a queen. Here we show that most (74%) colonies of the cavity-dwelling ant Temnothorax crassispinus construct up to two localized indoor 'latrines', which are used for defecation and only very rarely also as waste dumps. Restriction of defecation to designated areas affects the growth of mold inside the nest. Defecation strategies of colonies are furthermore affected by queen presence, with workers from queenless nests more frequently defecating outside the nest and forming latrines. As colonies do not actively avoid moldy nests, mold seems to not necessarily be a threat to the colony. While solid waste management has been more extensively studied in social insects, this study contributes a rare insight into the organization of non-easily transportable fecal waste.

Nurse honeybee workers tend capped brood, which does not require feeding, around the clock.

'Nurse' honeybees tend brood around the clock with attenuated or no circadian rhythms, but the brood signals inducing this behavior remain elusive. We first tested the hypothesis that worker circadian rhythms are regulated by brood pheromones. We monitored locomotor activity of individually isolated nurse bees that were exposed to either various doses of larval extract or synthetic brood ester pheromone (BEP). Bees orally treated with larval extract showed attenuated circadian rhythms in one of four tested colonies; a similar but statistically non-significant trend was seen in two additional colonies. Nurse bees treated with synthetic BEP showed rhythm attenuation in one of three tested colonies. Next, we tested the hypothesis that capped brood, which does not require feeding, nevertheless induces around-the-clock activity in nurses. By combining a new protocol that enables brood care by individually isolated nurse bees, detailed behavioral observations and automatic high-resolution monitoring of locomotor activity, we found that isolated nurses tended capped brood around the clock with attenuated circadian rhythms. Bees individually isolated in similar cages but without brood showed strong circadian rhythms in locomotor activity and rest. This study shows for the first time that the need to feed hungry larvae is not the only factor accounting for around-the-clock activity in nurse bees. Our results further suggest that the transition between activity with and without circadian rhythms is not a simple switch triggered by brood pheromones. Around-the-clock tending may enhance brood development and health in multiple ways that include improved larval feeding, thermoregulation or hygienic behavior.

Behavioral and molecular studies of quantitative differences in hygienic behavior in honeybees.

BACKGROUND: Hygienic behavior (HB) enables honeybees to tolerate parasites, including infection with the parasitic mite Varroa destructor, and it is a well-known example of a quantitative genetic trait. The understanding of the molecular processes underpinning the quantitative differences in this behavior remains limited. RESULTS: We performed gene expression studies in worker bees that displayed quantitative genetic differences in HB. We established a high and low genetic source of HB performance and studied the engagements into HB of single worker bees under the same environmental conditions. We found that the percentage of worker bees that engaged in a hygienic behavioral task tripled in the high versus low HB sources, thus suggesting that genetic differences may mediate differences in stimulated states to perform HB. We found 501 differently expressed genes (DEGs) in the brains of hygienic and non-hygienic performing workers in the high HB source bees, and 342 DEGs in the brains of hygienic performing worker bees, relative to the gene expression in non-hygienic worker bees from the low HB source group. "Cell surface receptor ligand signal transduction" in the high and "negative regulation of cell communication" in the low HB source were overrepresented molecular processes, suggesting that these molecular processes in the brain may play a role in the regulation of quantitative differences in HB. Moreover, only 21 HB-associated DEGs were common between the high and low HB sources. CONCLUSIONS: The better HB colony performance is primarily achieved by a high number of bees engaging in the hygienic tasks that associate with distinct molecular processes in the brain. We propose that different gene products and pathways may mediate the quantitative genetic differences of HB.

Genome-Wide Association Study of a Varroa-Specific Defense Behavior in Honeybees (Apis mellifera).

Honey bees are exposed to many damaging pathogens and parasites. The most devastating is Varroa destructor, which mainly affects the brood. A promising approach for preventing its spread is to breed Varroa-resistant honey bees. One trait that has been shown to provide significant resistance against the Varroa mite is hygienic behavior, which is a behavioral response of honeybee workers to brood diseases in general. Here, we report the use of an Affymetrix 44K SNP array to analyze SNPs associated with detection and uncapping of Varroa-parasitized brood by individual worker bees (Apis mellifera). For this study, 22 000 individually labeled bees were video-monitored and a sample of 122 cases and 122 controls was collected and analyzed to determine the dependence/independence of SNP genotypes from hygienic and nonhygienic behavior on a genome-wide scale. After false-discovery rate correction of the P values, 6 SNP markers had highly significant associations with the trait investigated (α < 0.01). Inspection of the genomic regions around these SNPs led to the discovery of putative candidate genes.

Cryptic extended brood care in the facultatively eusocial sweat bee Megalopta genalis.

As a result of different brood cell provisioning strategies, nest-making insects may differ in the extent to which adults regularly provide extended parental care to their brood beyond nest defense. Mass-provisioning species cache the entire food supply needed for larval development prior to the oviposition and typically seal the brood cell. It is usually assumed that there is no regular contact between the adult(s) and brood. Here, we show that the bee, Megalopta genalis, expresses a form of cryptic brood care, which would not be observed during normal development. Following experimental injections of different provisioning materials into brood cells, foundresses reopened manipulated cells and the brood were aborted in some cases, implying that the foundresses assessed conditions within the cells. In aborted cells, foundresses sometimes laid a second egg after first removing dead larvae, previously stored pollen and contaminants. Our results show that hygienic brood care can be cryptic and hence may be more widespread than previously believed, lending support to the hypothesis that extended parental care is a preadaptation toward eusociality.