ABSTRACT
Social bees have been extensively studied for their gut microbial functions, but the significance of the gut microbiota in solitary bees remains less explored. Solitary bee, Megachile rotundata females provision their offspring with pollen from various plant species, harboring a diverse microbial community that colonizes larvae guts. The Apilactobacillus is the most abundant microbe, but evidence concerning the effects of Apilactobacillus and other provision microbes on growth and survival are lacking. We hypothesized that the presence of Apilactobacillus in abundance would enhance larval and prepupal development, weight, and survival, while the absence of intact microbial communities was expected to have a negative impact on bee fitness. We reared larvae on pollen provisions with naturally collected microbial communities (Natural pollen) or devoid of microbial communities (Sterile pollen). We also assessed the impact of introducing Apilactobacillus micheneri by adding it to both types of pollen provisions. Feeding larvae with sterile pollen + A. micheneri led to the highest mortality rate, followed by natural pollen + A. micheneri, and sterile pollen. Larval development was significantly delayed in groups fed with sterile pollen. Interestingly, larval and prepupal weights did not significantly differ across treatments compared to natural pollen-fed larvae. 16S rRNA gene sequencing found a dominance of Sodalis, when A. micheneri was introduced to natural pollen. The presence of Sodalis with abundant A. micheneri suggests potential crosstalk between both, shaping bee nutrition and health. Hence, this study highlights that the reliance on nonhost-specific environmental bacteria may not impact fitness of M. rotundata.IMPORTANCEThis study investigates the impact of environmentally acquired gut microbes of solitary bee fitness with insights into the microbial ecology of bee and their health. While the symbiotic microbiome is well-studied in social bees, the role of environmental acquired microbiota in solitary bees remains unclear. Assessing this relationship in a solitary pollinator, the leaf-cutting bee, Megachile rotundata, we discovered that this bee species does not depend on the diverse environmental bacteria found in pollen for either its larval growth or survival. Surprisingly, high concentrations of the most abundant pollen bacteria, Apilactobacillus micheneri did not consistently benefit bee fitness, but caused larval mortality. Our findings also suggest an interaction between Apilactobacillus and the Sodalis and perhaps their role in bee nutrition. Hence, this study provides significant insights that contribute to understanding the fitness, conservation, and pollination ecology of other solitary bee species in the future.
ABSTRACT
The solitary bee Osmia lignaria is a native pollinator in North America with growing economic importance. The life cycle of O. lignaria provides a unique opportunity to compare the physiological and molecular mechanisms underlying two ecologically contrasting dormancies within the same species. O. lignaria prepupae become dormant during the summer to avoid high temperatures. Shortly after adult eclosion, they enter a second dormancy and overwinter as diapausing adults. To compare these two dormancies, we measured metabolic rates and gene expression across development as bees initiate, maintain, and terminate both prepupal (summer) and adult (overwintering) dormancies. We observed a moderate temperature-independent decrease in gas exchange during both the prepupal dormancy after cocoon spinning (45 %) and during adult diapause after eclosion (60 %). We sequenced and assembled a high-quality reference genome from a single haploid male bee with a contiguous n50 of 5.5 Mbp to facilitate our transcriptomic analysis. The transcriptomes of dormant prepupae and diapausing adults clustered into distinct groups more closely associated with life stage than dormancy status. Membrane transport, membrane-bound cellular components, oxidoreductase activity, glutathione metabolism, and transcription factor activity increased during adult diapause, relative to prepupal dormancy. Further, the transcriptomes of adults in diapause clustered into two groups, supporting multiple phases of diapause during winter. Late adult diapause was associated with gene expression profiles supporting increased insulin/IGF, juvenile hormone, and ecdysone signaling.
Subject(s)
Diapause , Transcriptome , Bees/genetics , Male , Animals , Temperature , Gene Expression Profiling , Life Cycle StagesABSTRACT
Insects exposed to constant low temperatures (CLT) exhibit high rates of mortality as well as a variety of sublethal effects. In many species, interruptions of CLT with brief pulses of warm temperatures (fluctuating thermal regimes, FTR) lead to increases in survival and fewer sublethal effects. However, we still lack a complete understanding of the physiological mechanisms activated during FTR. In this review, we discuss recent advances in understanding FTR's underlying molecular mechanisms. We discuss knowledge gaps related to potential trade-offs between FTR's beneficial effects and the costs of these repairs to overwintering reserves and reproduction. We present the hypothesis that the warm pulse of FTR helps to maintain daily rhythmicity.
Subject(s)
Cold Temperature , Insecta , Animals , Temperature , Insecta/physiology , ReproductionABSTRACT
The Colorado potato beetle, Leptinotarsa decemlineata, (Coleoptera: Chrysomelidae) is an economically important pest insect of potatoes. Understanding how the mechanisms driving its invasiveness vary between sexes will be critical for developing modern control methods. However, the currently available methods for sexing adult Colorado potato beetles are either inefficient or unsuitable for projects that require RNA as an input, like those measuring gene expression. Therefore, the development of simple molecular tools that are tailored to these studies is important. In this study, we used publicly available RNA-seq data to select 5 candidate genes for sex-specific markers in adult Colorado potato beetles. We confirmed that our 5 marker candidates exhibit a sex-specific expression pattern and can be used as PCR markers for sex determination. This method of sex detection will allow researchers to distinguish the sex of the individual with a simple PCR reaction using cDNA as the template and assign sex to RNA-seq samples post hoc.
Subject(s)
Coleoptera , Solanum tuberosum , Animals , Female , Male , Coleoptera/genetics , Solanum tuberosum/genetics , Colorado , DNA, Complementary , Gene ExpressionABSTRACT
The success of agriculture relies on healthy bees to pollinate crops. Commercially managed pollinators are often kept under temperature-controlled conditions to better control development and optimize field performance. One such pollinator, the alfalfa leafcutting bee, Megachile rotundata, is the most widely used solitary bee in agriculture. Problematically, very little is known about the thermal physiology of M. rotundata or the consequences of artificial thermal regimes used in commercial management practices. Therefore, we took a broad look at the thermal performance of M. rotundata across development and the effects of commonly used commercial thermal regimes on adult bee physiology. After the termination of diapause, we hypothesized thermal sensitivity would vary across pupal metamorphosis. Our data show that bees in the post-diapause quiescent stage were more tolerant of low temperatures compared to bees in active development. We found that commercial practices applied during development decrease the likelihood of a bee recovering from another bout of thermal stress in adulthood, thereby decreasing their resilience. Lastly, commercial regimes applied during development affected the number of days to adult emergence, but the time of day that adults emerged was unaffected. Our data demonstrate the complex interactions between bee development and thermal regimes used in management. This knowledge can help improve the commercial management of these bees by optimizing the thermal regimes used and the timing of their application to alleviate negative downstream effects on adult performance.
Subject(s)
Cold Temperature , Medicago sativa , Bees , Animals , Temperature , Pupa , Metamorphosis, BiologicalABSTRACT
Megachile rotundata (F.) is an important pollinator of alfalfa in the United States. Enhancing landscapes with wildflowers is a primary strategy for conserving pollinators and may improve the sustainability of M. rotundata. Changing cold storage temperatures from a traditionally static thermal regime (STR) to a fluctuating thermal regime (FTR) improves overwintering success and extends M. rotundata's shelf life and pollination window. Whether floral resources enhance overwintering survival and/or interact with a thermal regime are unknown. We tested the combined effects of enhancing alfalfa fields with wildflowers and thermal regime on survival and macronutrient stores under extended cold storage (i.e., beyond one season). Megachile rotundata adults were released in alfalfa plots with and without wildflower strips. Completed nests were harvested in September and stored in STR. After a year, cells were randomly assigned to remain in STR for 6 months or in FTR for a year of extended cold storage; emergence rates were observed monthly. Macronutrient levels of emerged females were assessed. FTR improved M. rotundata survival but there was no measurable effect of wildflower strips on overwintering success or nutrient stores. Timing of nest establishment emerged as a key factor: offspring produced late in the season had lower winter survival and dry body mass. Sugars and glycogen stores increased under FTR but not STR. Trehalose levels were similar across treatments. Total lipid stores depleted faster under FTR. While wildflowers did not improve M. rotundata survival, our findings provide mechanistic insight into benefits and potential costs of FTR for this important pollinator.
Subject(s)
Hymenoptera , Female , Bees , Animals , Temperature , Medicago sativa , Trehalose , Nutrients , Glycogen , LipidsABSTRACT
Within the United States and Canada, the primary pollinator of alfalfa is the alfalfa leafcutting bee (ALCB), Megachile rotundata. Our previous findings showed that overwintering conditions impacted gene expression profile in ALCB prepupae that entered diapause early in the season. However, ALCB are a bivoltine species, which begs the question of whether bees entering diapause later in the season also show this trend. To better understand the effects of the timing of diapause initiation, we analyzed mRNA copy number of genes known to be involved in diapause regulation in early and late season diapausing ALCB that were overwintered in field conditions or using current agricultural management conditions. We hypothesized that overwintering conditions for late diapausing bees also affects gene expression profiles. Our results showed that expression profiles were altered by both overwintering condition and timing of diapause initiation, with bees that entered diapause earlier in the season showing different expression patterns than those that entered diapause later in the season. This trend was seen in expression of members of the cyclin family and several targets of the insulin signaling pathway, including forkhead box protein O (FOXO), which is known to be important for diapause regulation and stress responses. But, of the genes screened, the proto-oncogene, Myc, was the most impacted by the timing of diapause initiation. Under field conditions, there were significant differences in Myc expression between the early and late season samples in all months except for November and February. This same general trend in Myc expression was also seen in the laboratory-maintained bees with significant difference in expression in all months except for November, February, and May. These results support previous conclusions from our research showing that the molecular regulation of diapause development in ALCB is not a simple singular cascade of gene expression but a highly plastic response that varies between bees depending upon their environmental history.
ABSTRACT
Diapause is a complex physiological phenomenon that allows insects to weather stressful environmental conditions. The regulation of diapause is accordingly complex, including signaling pathways that involve both small RNA and mRNA and affect the cell cycle, stress resistance, and developmental timing. Transposable elements, mobile genetic elements that replicate within the genome, are also thought to be stress responsive and regulated by the small RNA pathway. Therefore, we asked what the relationship was between environmental stress, diapause status, and transposable element expression in two species of agriculturally important bees, Megachile rotundata and Osmia lignaria. We characterized the TE content of the genomes of both species, then evaluated the expression of TE families during temperature stress, general environmental stress, and diapause stage. We found that the genomic TE content of the two species was very different, and M. rotundata has a larger number of annotated TEs compared to O. lignaria. We also found that both diapause stage and temperature stress had large effects on TE expression. The fold change of TE famlies tended to be larger in those expressed during diapause, however there was only a small majority that were upregulated during diapause. This suggests that stress and diapause do not break down to a simple up-regulation or down-regulation of TEs, but rather that the TE family, the genomic position of its insertions, and the exact heterochromatin formation of the organism at any given environmental state or life stage may affect overall expression of TEs.
Subject(s)
DNA Transposable Elements , Diapause , Animals , Bees/genetics , Humans , Stress, Physiological , TemperatureABSTRACT
Megachile rotundata exhibits a facultative prepupal diapause but the cues regulating diapause initiation are not well understood. Possible cues include daylength and temperature. Megachile rotundata females experience changing daylengths over the nesting season that may influence diapause incidence in their offspring through a maternal effect. Juvenile M. rotundata spend their developmental period confined in a nesting cavity, potentially subjected to stressful temperatures that may affect diapause incidence and survival. To estimate the impact of daylength and nest cavity temperature on offspring diapause, we designed a 3D printed box with iButtons that measured nest cavity temperature. We observed nest building throughout the season, monitored nest cavity temperature, and followed offspring through development to measure diapause incidence and mortality. We found that daylength was a cue for diapause, and nest cavity temperature did not influence diapause incidence. Eggs laid during long days had a lower probability of diapause. Siblings tended to have the same diapause status, explaining a lot of the remaining variance in diapause incidence. Some females established nests that contained both diapausing and nondiapausing individuals, which were distributed throughout the nest. Nest cavities reached stressful temperatures, which decreased survival. Mortality was significantly higher in nondiapausing bees and the individuals that were laid first in the nest. In conclusion, we demonstrate a maternal effect for diapause that is mediated by daylength and is independent of nest box temperature.
Subject(s)
Bees/physiology , Diapause, Insect/physiology , Environment , Nesting Behavior/physiology , Animals , Larva/physiology , Medicago sativa/physiology , Seasons , TemperatureABSTRACT
Variation in body size has important implications for physical performance and fitness. For insects, adult size and morphology are determined by larval growth and metamorphosis. Female blue orchard bees, Osmia lignaria, (Say) provision a finite quantity of food to their offspring. In this study, we asked how provision-dependent variation in size changes adult morphology. We performed a diet manipulation in which some larvae were starved in the final instar and some were given unlimited food. We examined the consequences on adult morphology in two ways. First, allometric relationships between major body regions (head, thorax, abdomen) and total body mass were measured to determine relative growth of these structures. Second, morphometrics that are critical for flight (wing area, wing loading, and extra flight power index) were quantified. Head and thorax mass had hyperallometric relationships with body size, indicating these parts become disproportionately large in adults when larvae are given copious provisions. However, abdominal mass and wing area increased hypoallometrically with body size. Thus, large adults had disproportionately lighter abdomens and smaller wing areas than smaller adults. Though both males and females followed these general patterns, allometric patterns were affected by sex. For flight metrics, small adults had reduced wing loading and an increased extra flight power index. These results suggest that diet quantity alters development in ways that affect the morphometric trait relationships in adult O. lignaria and may lead to functional differences in performance.
Subject(s)
Bees , Body Size , Genetic Fitness/physiology , Wings, Animal , Animals , Bees/anatomy & histology , Bees/physiology , Body Size/physiology , Feeding Behavior , Female , Fertility , Hymenoptera/anatomy & histology , Hymenoptera/physiology , Larva/physiology , Male , Sex Factors , Wings, Animal/anatomy & histology , Wings, Animal/physiologyABSTRACT
Diapause is a non-feeding state that many insects undergo to survive the winter months. With fixed resources, overall metabolism and insulin signaling (IIS) are maintained at low levels, but whether those change in response to seasonal temperature fluctuations remains unknown. The focus of this study was to determine 1) how genes in the insulin signaling pathway vary throughout diapause and 2) if that variation changes in response to temperature. To test the hypothesis that expression of IIS pathway genes vary in response to temperature fluctuations during overwintering, alfalfa leafcutting bees, Megachile rotundata, were overwintered at either a constant 4 °C in the lab or in naturally fluctuating temperatures in the field. Expression levels of genes in the IIS pathway, cell cycle regulators, and transcription factors were measured. Overall our findings showed that a few key targets of the insulin signaling pathway, along with growth regulators, change during overwintering, suggesting that only cell cycle regulators, and not the IIS pathway as a whole, change across the phases of diapause. To answer our second question, we compared gene expression levels between temperature treatments at each month for a given gene. We observed significantly more differences in expression of IIS pathway targets, indicating that overwintering conditions impact insulin pathway gene expression and leads to altered expression profiles. With differences seen between temperature treatment groups, these findings indicate that constant temperatures like those used in agricultural storage protocols, lead to different expression profiles and possibly different diapause phenotypes for alfalfa leafcutting bees.
Subject(s)
Bees/physiology , Diapause , Gene Expression Regulation , Insulin/metabolism , Seasons , Animals , Bees/genetics , Signal TransductionABSTRACT
Interrupting the spring incubation of Megachile rotundata (F.) with a period of low-temperature storage for synchronizing the bees' emergence with crop bloom is an essential part of M. rotundata management. Previously, we demonstrated that bees exposed to thermoperiods (TPs) during low-temperature storage have higher survival rates than bees exposed to constant temperatures. But changing the temperature in the large mass of bees commonly found in most commercial settings would place considerable stress on the chambers' refrigeration system. Reducing the difference between a TP's cryophase and thermophase would decrease the stress on the refrigeration system. Therefore, we investigated a range of TPs with cryophases (12 h) of 6, 12, or 15°C and thermophases (12 h) of 15 or 18°C and compared the survival rates of these bees against bees exposed to constant temperatures of 12, 15, or 18°C. For eye-pigmented pupae, the TP 6-18°C and the control fluctuating thermal regime (FTR; 6°C with a daily 1-h pulse at 20°C) had the highest survival rates for the 2 yr tested. For the constant-temperature storage protocols, constant 15 and 18°C were either equivalent or lower survival than the control FTR. For emergence-ready adults, the 6-18°C TP had the highest survival rates. The constant 15°C and the control FTR had equivalent survival rates. Under the current constraints imposed by a commercial chamber's refrigeration system, interrupting M. rotundata spring incubation by exposing the developing bees to constant temperatures of 15-18°C is currently the best option for commercial operations.
Subject(s)
Hymenoptera , Animals , Bees , Cold Temperature , Medicago sativa , Pupa , TemperatureABSTRACT
In species that care for their young, provisioning has profound effects on offspring fitness. Provisioning is important in honeybees because nutritional cues determine whether a female becomes a reproductive queen or sterile worker. A qualitative difference between the larval diets of queens and workers is thought to drive this divergence; however, no single compound seems to be responsible. Diet quantity may have a role during honeybee caste determination yet has never been formally studied. Our goal was to determine the relative contributions of diet quantity and quality to queen development. Larvae were reared in vitro on nine diets varying in the amount of royal jelly and sugars, which were fed to larvae in eight different quantities. For the middle diet, an ad libitum quantity treatment was included. Once adults eclosed, the queenliness was determined using principal component analysis on seven morphological measurements. We found that larvae fed an ad libitum quantity of diet were indistinguishable from commercially reared queens, and that queenliness was independent of the proportion of protein and carbohydrate in the diet. Neither protein nor carbohydrate content had a significant influence on the first principle component 1 (PC1), which explained 64.4% of the difference between queens and workers. Instead, the total quantity of diet explained a significant amount of the variation in PC1. Large amounts of diet in the final instar were capable of inducing queen traits, contrary to the received wisdom that queen determination can only occur in the third instar. These results indicate that total diet quantity fed to larvae may regulate the difference between queen and worker castes in honeybees.
Subject(s)
Bees/physiology , Behavior, Animal , Diet , Social Behavior , AnimalsABSTRACT
The temperature of the nest influences fitness in cavity-nesting bees. Females may choose nest cavities that mitigate their offspring's exposure to stressful temperatures. This study aims to understand how cavity temperature impacts the nesting preference of the solitary bee Megachile rotundata (Fabricius) under field conditions. We designed and 3D printed nest boxes that measured the temperatures of 432 cavities. Nest boxes were four-sided with cavity entrances facing northeast, northwest, southeast, and southwest. Nest boxes were placed along an alfalfa field in Fargo, ND and were observed daily for completed nests. Our study found that cavity temperature varied by direction the cavity faced and by the position of the cavity within the nest box. The southwest sides recorded the highest maximum temperatures while the northeast sides recorded the lowest maximum temperatures. Nesting females filled cavities on the north-facing sides faster than cavities on the south-facing sides. The bees preferred to nest in cavities with lower average temperatures during foraging hours, and cavities that faced to the north. The direction the cavity faced was associated with the number of offspring per nest. The southwest-facing cavities had fewer offspring than nests on the northeast side. Our study indicates that the nesting box acts as a microclimate, with temperature varying by position and direction of the cavity. Variation in cavity temperature affected where females chose to nest, but not their reproductive investment.
Subject(s)
Hymenoptera , Animals , Bees , Female , Microclimate , Nesting Behavior , Reproduction , TemperatureABSTRACT
In insects, prolonged exposure to unseasonably low temperatures can lead to detrimental physiological effects known as chill injury. Changes to active and passive transport across epithelia during chilling likely drive the collapse of ion gradients, metabolic imbalance and potentially oxidative stress. In the alfalfa leafcutting bee, Megachile rotundata transcriptomic evidence provides support for these responses at the level of gene expression, but variable expression profiles between life stages in M. rotundata indicate that different mechanisms could be responsible for repairing and protecting against chill injuries across development. Herein, we test the hypotheses that 1) chill injury leads to oxidative stress and damage in insects and 2) exposure to a fluctuating thermal regime (FTR) promotes an increased oxidative stress response leading to a decrease in damage by reactive oxygen species. We measured the expression of transcripts with products known to have antioxidant properties in overwintering prepupae as well as total antioxidant capacity and lipid peroxidation during both extended overwintering in prepupae and low temperature stress during pupal development. We observed differential gene expression for the antioxidant glutathione peroxidase and several transcripts with putative antioxidant properties including vitellogenin, apolipoprotein D, glutathione S-transferase, and nuclear protein 1. However, the expression of transcripts coding for other enzymatic antioxidants did not change between treatments. Neither life stage varied in their capacity to cope with an induced oxidative stress after FTR exposure and we did not observe evidence of lipid peroxidation in chill injured (STR) prepupae. These results did not support our initial hypotheses and indicate that oxidative-stress-induced damage is neither a causal factor or symptom of chill injury.
Subject(s)
Bees/physiology , Cold Temperature/adverse effects , Oxidative Stress , Animals , Antioxidants , Bees/growth & development , Gene Expression , Life Cycle Stages , Lipid Peroxidation , Pupa/physiology , Reactive Oxygen Species , TranscriptomeABSTRACT
Insects exposed to low temperature stress can experience chill injury, but incorporating fluctuating thermoprofiles increases survival and blocks the development of sub-lethal effects. The specific parameters required for a protective thermoprofile are poorly understood, because most studies test a limited range of thermoprofiles. For example, thermoprofiles with a wave profile may perform better than a square profile, but these two profiles are rarely compared. In this study, two developmental stages of the alfalfa leafcutting bee, Megachile rotundata, eye-pigmented pupae, and emergence-ready adults, were exposed to one of eight thermoprofiles for up to 8 weeks. All the thermoprofiles had a base of 6°C and a peak temperature of either 12°C or 18°C. The duration at peak temperature varied depending on the shape of the thermoprofile, either square or wave form. Two other treatments acted as controls, a constant 6°C and a fluctuating thermal regime (FTR) with a base temperature of 6°C that was interrupted daily by a single, 1-h pulse at 20°C. Compared with constant 6°C, all the test thermoprofiles significantly improved survival. Compared with the FTR control, the thermoprofiles with a peak temperature of 18°C outperformed the 12°C profiles. Bees in the eye-pigmented stage exposed to the 18°C profiles separated into two groups based on the shape of the profile, with higher survival in the square profiles compared with the wave profiles. Bees in the emergence-ready stage exposed to 18°C profiles all had significantly higher survival than bees in the FTR controls. Counter to expectations, the least ecologically relevant thermoprofiles (square) had the highest survival rates and blocked the development of sub-lethal effects (delayed emergence).
Subject(s)
Bees/physiology , Longevity , Temperature , Animals , Bees/growth & development , Cold Temperature , Female , Male , Pupa/growth & development , Pupa/physiology , Time FactorsABSTRACT
The response of ectotherms to temperature stress is complex, non-linear, and is influenced by life stage and previous thermal exposure. Mortality is higher under constant low temperatures than under a fluctuating thermal regime (FTR) that maintains the same low temperature but adds a brief, daily pulse of increased temperature. Long term exposure to FTR has been shown to increase transcription of genes involved in oxidative stress, immune function, and metabolic pathways, which may aid in recovery from chill injury and oxidative damage. Previous research suggests the transcriptional response that protects against sub-lethal damage occurs rapidly under exposure to fluctuating temperatures. However, existing studies have only examined gene expression after a week or over many months. Here we characterize gene expression during a single temperature cycle under FTR. Development of pupating alfalfa leafcutting bees (Megachile rotundata) was interrupted at the red-eye stage and were transferred to 6°C with a 1-h pulse to 20°C and returned to 6°C. RNA was collected before, during, and after the temperature pulse and compared to pupae maintained at a static 6°C. The warm pulse is sufficient to cause expression of transcripts that repair cell membrane damage, modify membrane composition, produce antifreeze proteins, restore ion homeostasis, and respond to oxidative stress. This pattern of expression indicates that even brief exposure to warm temperatures has significant protective effects on insects exposed to stressful cold temperatures that persist beyond the warm pulse. Megachile rotundata's sensitivity to temperature fluctuations indicates that short exposures to temperature changes affect development and physiology. Genes associated with developmental patterning are expressed after the warm pulse, suggesting that 1 h at 20°C was enough to resume development in the pupae. The greatest difference in gene expression occurred between pupae collected after the warm pulse and at constant low temperatures. Although both were collected at the same time and temperature, the transcriptional response to one FTR cycle included multiple transcripts previously identified under long-term FTR exposure associated with recovery from chill injury, indicating that the effects of FTR occur rapidly and are persistent.
Subject(s)
Bees/growth & development , Hot Temperature , Transcription, Genetic , Animals , Bees/genetics , Pupa/genetics , Pupa/growth & developmentABSTRACT
Pollination services provided by the honey bee, Apis mellifera (Hymenoptera: Apidae, Linnaeus, 1758) have broad economic impacts and are necessary for production of a diversity of important crops. Hives may be transported multiple times per year to provide pollination. To test how temperature may contribute to transportation stress, temperature sensors were placed in hives in different locations and orientations on the trailer during shipping. Colony size prior to shipping significantly contributed to loss of population immediately after shipping which contributed to colony failure with smaller colonies more likely to fail and fail faster. Colony size also affects thermoregulation and temperature stress. Internal hive temperature varies significantly based on location and orientation. While colonies near the front and rear of the trailer and those oriented toward the center aisle had significantly different average internal temperatures, colony size best predicts loss of thermoregulation. Additionally, we profiled gene expression at departure, on arrival, and after a recovery period to identify transcriptional responses to transportation. Functional and enrichment analysis identified increased methylation and decreased ribosomal and protein-folding activity. Pheromone and odorant-binding transcripts were up-regulated after transportation. After recovery, transcripts associated with defense response, immune activity, and heat shock decreased, while production of antibiotic peptides increased. We conclude that hives experience considerable temperature stress possibly caused by turbulent airflow in exposed locations. Transportation stress should be considered an important component of annual colony losses which can be mitigated with improved management strategies.
Subject(s)
Hymenoptera , Animals , Bees , Crops, Agricultural , Pollination , TemperatureABSTRACT
Existing systems to measure insect emergence patterns have limitations; they are only partially automated and are limited in the maximum number of emerging insects they can detect. In order to obtain precise measurement of insect emergence, it is necessary for systems to be semi-automated and able to measure large numbers of emerging insects. We addressed these issues by designing and building a system that is automated and can measure emergence of up to 1200 insects. We modified the existing "falling-ball" system using Arduino microcontrollers to automate data collection and expand the sample size through multiple data channels. Multiple data channels enable the user to not only increase their sample size, but also allows for multiple treatments to be run simultaneously in a single experiment. Furthermore, we created an R script to automatically visualize the data as a bubble plot, while also calculating the median day and time of emergence. The current system was designed using 3D printing so the user can modify the system to be adjusted for different species of insects. The goal of this protocol is to investigate important questions in chronobiology and stress physiology, using this precise and automated system to measure insect emergence patterns.
Subject(s)
Insecta/physiology , AnimalsABSTRACT
Insects exposed to low temperature often have high mortality or exhibit sublethal effects. A growing number of recent studies have shown beneficial effects of exposing insects to recurrent brief warm pulses during low-temperature stress (fluctuating thermal regime, FTR). The physiological underpinnings of the beneficial effects of FTR on cold survival have been extensively studied over the past few years. Profiling with various '-omics' techniques has provided supporting evidence for different physiological responses between insects exposed to FTR and constant low temperature. Evidence from transcriptomic, metabolomic and lipidomic studies points to a system-wide loss of homeostasis at low temperature that can be counterbalanced by repair mechanisms under FTR. Although there has been considerable progress in understanding the physiological mechanisms underlying the beneficial effects of FTR, here we discuss how many areas still lack clarity, such as the precise role(s) of heat shock proteins, compatible solutes or the identification of regulators and key players involved in the observed homeostatic responses. FTR can be particularly beneficial in applied settings, such as for model insects used in research, integrated pest management and pollination services. We also explain how the application of FTR techniques in large-scale facilities may require overcoming some logistical and technical constraints. FTR definitively enhances survival at low temperature in insects, but before it can be widely used, we suggest that the possible fitness and energy costs of FTR must be explored more thoroughly. Although FTR is not ecologically relevant, similar processes may operate in settings where temperatures fluctuate naturally.