Acute Imidacloprid Exposure Alters Mitochondrial Function in Bumblebee Flight Muscle and Brain

Mitochondria are intracellular organelles responsible for cellular respiration with one of their major roles in the production of energy in the form of ATP. Activities with increased energetic demand are especially dependent on efficient ATP production, hence sufficient mitochondrial function is fundamental. In bees, flight muscle and the brain have particularly high densities of mitochondria to facilitate the substantial ATP production required for flight activity and neuronal signalling. Neonicotinoids are systemic synthetic insecticides that are widely utilised against crop herbivores but have been reported to cause, by unknown mechanisms, mitochondrial dysfunction, decreasing cognitive function and flight activity among pollinating bees. Here we explore, using high-resolution respirometry, how the neonicotinoid imidacloprid may affect oxidative phosphorylation in the brain and flight muscle of the buff-tailed bumblebee, Bombus terrestris. We find that acute exposure increases routine oxygen consumption in the flight muscle of worker bees. This provides a candidate explanation for prior reports of early declines in flight activity following acute exposure. We further find that imidacloprid increases the maximum electron transport capacity in the brain, with a trend towards increased overall oxygen consumption. However, intra-individual variability is high, limiting the extent to which apparent effects of imidacloprid on brain mitochondria are shown conclusively. Overall, our results highlight the necessity to examine tissue-specific effects of imidacloprid on respiration and energy production.

A Candidate RNAi Screen Reveals Diverse RNA-Binding Protein Phenotypes in Drosophila Flight Muscle

The proper regulation of RNA processing is critical for muscle development and the fine-tuning of contractile ability among muscle fiber-types. RNA binding proteins (RBPs) regulate the diverse steps in RNA processing, including alternative splicing, which generates fiber-type specific isoforms of structural proteins that confer contractile sarcomeres with distinct biomechanical properties. Alternative splicing is disrupted in muscle diseases such as myotonic dystrophy and dilated cardiomyopathy and is altered after intense exercise as well as with aging. It is therefore important to understand splicing and RBP function, but currently, only a small fraction of the hundreds of annotated RBPs expressed in muscle have been characterized. Here, we demonstrate the utility of Drosophila as a genetic model system to investigate basic developmental mechanisms of RBP function in myogenesis. We find that RBPs exhibit dynamic temporal and fiber-type specific expression patterns in mRNA-Seq data and display muscle-specific phenotypes. We performed knockdown with 105 RNAi hairpins targeting 35 RBPs and report associated lethality, flight, myofiber and sarcomere defects, including flight muscle phenotypes for Doa, Rm62, mub, mbl, sbr, and clu. Knockdown phenotypes of spliceosome components, as highlighted by phenotypes for A-complex components SF1 and Hrb87F (hnRNPA1), revealed level- and temporal-dependent myofibril defects. We further show that splicing mediated by SF1 and Hrb87F is necessary for Z-disc stability and proper myofibril development, and strong knockdown of either gene results in impaired localization of kettin to the Z-disc. Our results expand the number of RBPs with a described phenotype in muscle and underscore the diversity in myofibril and transcriptomic phenotypes associated with splicing defects. Drosophila is thus a powerful model to gain disease-relevant insight into cellular and molecular phenotypes observed when expression levels of splicing factors, spliceosome components and splicing dynamics are altered.

Rapid recovery by fat- and muscle-depleted Blackpoll Warblers following trans-oceanic migration is driven by time-minimization

Nonstop endurance flights are a defining characteristic of many long-distance migratory birds, but subsequent recovery phases are not typically distinguished from fueling phases (collectively “stopovers”), despite endurance flights inducing marked physiological changes including flight muscle atrophy and gastrointestinal tract reductions. Here, we hypothesize that recovery requires unique behavioral adaptations, leading to departures from the predictions of optimal migration theory for time-minimizing migrants. We predict that recovering birds will (1) select (moist) food-rich habitats on arrival; (2) have slow initial fueling rates due to decreased gastrointestinal capacity; (3) show a negative correlation between stopover duration and arrival condition instead of a negative correlation with fuel deposition rate (FDR); (4) stopover longer than required to store energy reserves for subsequent flights; and (5) show evidence of rebuilding flight muscles. To test these predictions, we studied Blackpoll Warblers (Setophaga striata) in northern Colombia following trans-oceanic flights >2,250 km. Birds selected dry seasonal habitats, despite the proximity of moist forests, and among 1,227 captured individuals, 14–21% were emaciated and 88% had atrophied flight muscles. We recaptured 74 individuals, revealing net positive mass gains and, contrary to prediction (2), no evidence for slow initial recovery rates. Contrary to prediction (3), stopover duration was only weakly correlated with arrival condition and birds with high FDR (4.9% lean body mass day–1) had shorter durations (3 days) relative to birds with slower rates (7 days): both groups accumulated sufficient fuel to reach nonbreeding (over-wintering) grounds 500–1,000 km away. Mass increases were largely attributable to fat deposition but some birds improved flight muscle condition (31.9%), consistent with prediction (5). Together these results reveal a strong selection for time-minimization in the decisions made by Blackpoll Warblers following trans-oceanic flights, likely mediated through advantages to early arrival on nonbreeding grounds, contrary to our hypothesis of recovery imposing unique selection pressures.

A Candidate RNAi Screen Reveals Diverse RNA-Binding Protein Phenotypes in Drosophila Flight Muscle

The proper regulation of RNA processing is critical for muscle development and the fine-tuning of contractile ability between muscle fiber-types. RNA binding proteins (RBPs) regulate the diverse steps in RNA processing including alternative splicing, which generates fiber-type specific isoforms of structural proteins that confer contractile sarcomeres with distinct biomechanical properties. Alternative splicing is disrupted in muscle diseases such as myotonic dystrophy and dilated cardiomyopathy, and is altered after intense exercise as well as with aging. It is therefore important to understand splicing and RBP function, but currently only a small fraction of the hundreds of annotated RBPs expressed in muscle have been characterized. Here we demonstrate the utility of Drosophila as a genetic model system to investigate basic developmental mechanisms of RBP function in myogenesis. We find that RBPs exhibit dynamic temporal and fiber-type specific expression patterns in mRNA-Seq data and display muscle-specific phenotypes. We performed knockdown with 105 RNAi hairpins targeting 35 RBPs and report associated lethality, flight, myofiber and sarcomere defects, including flight muscle phenotypes for Doa, Rm62, mub, mbl, sbr, and clu. Interestingly, knockdown phenotypes of spliceosome components, as highlighted by phenotypes for A-complex components SF1 and Hrb87F (hnRNPA1), revealed level- and temporal-dependent myofibril defects. We further show that splicing mediated by SF1 and Hrb87F is necessary for Z-disc stability and proper myofibril development, and strong knockdown of either gene results in impaired localization of Kettin to the Z-disc. Our results expand the number of RBPs with a described phenotype in muscle and underscore the diversity in myofibril and transcriptomic phenotypes associated with splicing defects. Drosophila is thus a useful model to gain disease-relevant insight into cellular and molecular phenotypes observed when expression levels of splicing factors, spliceosome components and splicing dynamics are altered.

Proline as a Sparker Metabolite of Oxidative Metabolism during the Flight of the Bumblebee, Bombus impatiens

Several insect species use the amino acid proline as a major energy substrate. Although initially thought to be limited to blood-feeding dipterans, studies have revealed this capability is more widespread. Recent work with isolated flight muscle showed that the bumblebee Bombus impatiens can oxidize proline at a high rate. However, its role as a metabolic fuel to power flight is unclear. To elucidate the extent to which proline is oxidized to power flight and how its contribution changes during flight, we profiled 14 metabolites central to energy and proline metabolism at key time points in flight muscle and abdominal tissues. Ultra-high performance liquid chromatography-electrospray ionization-quadrupole time of flight mass spectrometry (UPLC-ESI-QTOF MS) analysis revealed that proline is likely used as a sparker metabolite of the tricarboxylic acid cycle at the onset of flight, whereby it supplements the intermediates of the cycle. Carbohydrates are the major energy substrates, which is evidenced by marked decreases in abdominal glycogen stores and a lack of alanine accumulation to replenish flight muscle proline. The time course of fuel stores and metabolites changes during flight highlights homeostatic regulation of energy substrates and patterns of changes in metabolic intermediates within pathways. This study clarifies the role of proline and carbohydrate metabolism during flight in hymenopterans, such as B. impatiens.

Flight muscle and flight activity of melon fly, Bactrocera cucurbitae (Diptera: Tephritidae)

The flight activity and flight muscle of the melon fly, Bactrocera cucurbitae (Coquillett) (Diptera: Tephritidae) were observed. The Tethered technique was used to observe the flight activity in this study. The flight activity, and wing and flight muscles were compared between male and female melon flies. The results indicate that the female was relatively better and strong flier than the male. The mean duration of the flight activity of the females was 13.90 min/hour and of the males was 7.12 min./hour. The mean length, width, volume of wings of the males were 6.07 mm, 2.67 mm and 10.99 mm³, respectively. On the other hand, the mean length, width and volume of the wings of females were 7.07 mm, 2.87 mm and 15.60 mm³, respectively. In case of wing muscles, the mean volume of dorsal longitudinal muscle (DLM) in male and female was found 5.20 mm³ and 5.67 mm³, respectively. The mean length of flight wing muscle of male and female was 2.22 and 2.23 mm, respectively and the mean breadth of male and female was 1.65 and 1.77 mm, respectively. Dhaka Univ. J. Biol. Sci. 30(2): 179-185, 2021 (July)

Tissue-specific transcriptional patterns underlie seasonal phenotypes in honey bees (Apis mellifera)

Faced with adverse conditions, such as winter in temperate regions or hot and dry conditions in tropical regions, many insect species enter a state of diapause, a period of dormancy associated with a reduction or arrest of physical activity, development, and reproduction. Changes in common physiological pathways underlie diapause phenotypes in different insect species. However, most transcriptomic studies of diapause have not simultaneously evaluated and compared expression patterns in different tissues. Honey bees (Apis mellifera) represent a unique model system to study the mechanisms underpinning diapause. In winter, honey bees exhibit a classic diapause phenotype, with reduced metabolic activity, increased physiological nutritional resources, and altered hormonal profiles. However, winter bees actively heat their colony by vibrating their wing muscles; thus, this tissue is not quiescent. Here, we evaluated the transcriptional profiles of flight muscle tissue and fat body tissue (involved in nutrient storage, metabolism and immune function) of winter bees. We also evaluated two behavioral phenotypes of summer bees: nurses, which exhibit high nutritional stores and low flight activity, and foragers, which exhibit low nutritional stores and high flight activity. We found winter bees and nurses have similar fat body transcriptional profiles compared to foragers, whereas winter bees and foragers have similar flight muscle transcriptional profiles compared to nurses. Additionally, differentially expressed genes were enriched in diapause-related GO terms. Thus, honey bees exhibit tissue-specific transcriptional profiles associated with diapause, laying the groundwork for future studies evaluating the mechanisms, evolution, and consequences of this tissue-specific regulation.