Protocol for measuring the effects of an inhibitory signal associated with danger on honey bee dopamine levels.

The stop signal is produced in response to negative experiences at the food source and inhibits honey bee (Apis mellifera) waggle dancing. Here, we present a protocol for measuring the effects of an inhibitory signal associated with danger on honey bee dopamine levels. We describe steps for observing honey bee colonies, training them with artificial nectar, and simulating hornet attacks. We then detail procedures for recording waggle dancing and stop signals and measuring brain dopamine levels during different treatments. For complete details on the use and execution of this protocol, please refer to Dong et al.1.

Identification of characteristic flavor compounds of boletus edulis from different regions based on by E-nose, HS-GC-IMS and HS-SPME-GC-MS.

In this study, E-nose, HS-GC-IMS, and HS-SPME-GC-MS technologies were used to evaluate the flavor characteristics of the pileus and stipe of Boletus edulis from eight origins. 23 key Volatile organic compounds (VOCs) with odor activity values (OAVs) > 1 were identified, and 19 aroma types have been identified in Boletus edulis at the same time. Vegetable and earthy were defined as the dominant aroma types for all pileus and stipe samples. Balsamic and musty were the main and characteristic aroma types for the pileus. The highest concentrations of VOCs in the pileus and stipe were originated from Chuxiong Prefecture and Aba Prefecture, respectively. 19 and 16 key VOCs were detected Chuxiong pileus and Aba stipe, respectively, and Methional was the decisive compound that influenced the vegetable aroma type. The results of this study could be helpful for flavor identification and application of pileus and stipe from Boletus edulis.

A Method for Studying Social Signal Learning of the Waggle Dance in Honey Bees.

Honey bees use a complex form of spatial referential communication. Their waggle dance communicates to nestmates the direction, distance, and quality of a resource by encoding celestial cues, retinal optic flow, and relative food value into motion and sound within the nest. This protocol was developed to investigate the potential for social learning of this waggle dance. Using this protocol, we showed that correct waggle dancing requires social learning. Bees (Apis mellifera) that did not follow any dances before they first danced produced significantly more disordered dances, with larger waggle angle divergence errors, and encoded distance incorrectly. The former deficits improved with experience, but distance encoding was set for life. The first dances of bees that could follow other dancers had none of these impairments. Social learning, therefore, shapes honey bee signaling, as it does communication in human infants, birds, and multiple other vertebrate species. However, much remains to be learned about insects' social learning, and this protocol will help to address knowledge gaps in the understanding of sophisticated social signal learning, particularly in understanding the molecular bases for such learning. Key features It was unclear if honey bees (Apis mellifera) could improve their waggle dance by following experienced dancers before they first waggle dance. Honey bees perform their first waggle dances with more errors if they cannot follow experienced waggle dancers first. Directional and disorder errors improved over time, but distance error was maintained. Bees in experimental colonies continued to communicate longer distances than control bees. Dancing correctly, with less directional error and disorder, requires social learning. Distance encoding in the honey bee dance is largely genetic but may also include a component of cultural transmission.

An inhibitory signal associated with danger reduces honeybee dopamine levels.

Positive and negative experiences can alter animal brain dopamine levels.1 When first arriving at a rewarding food source or beginning to waggle dance and recruit nestmates to food, honeybees have increased brain dopamine levels, indicating a desire for food.2 We provide the first evidence that an inhibitory signal, the stop signal, which counters waggle dancing and is triggered by negative events at the food source, can decrease head dopamine levels and dancing, independent of the dancer having any negative experiences. The hedonic value of food can therefore be depressed simply by the receipt of an inhibitory signal. Increasing the brain dopamine levels reduced the aversive effects of an attack, increasing the time that bees spent subsequently feeding and waggle dancing and decreasing their stop signaling and time spent in the hive. Because honeybees regulate food recruitment and its inhibition at the colony level, these results highlight the complex integration of colony information with a basic and highly conserved neural mechanism in mammals and insects.2 VIDEO ABSTRACT.

Social signal learning of the waggle dance in honey bees.

Honey bees use a complex form of spatial referential communication. Their "waggle dance" communicates the direction, distance, and quality of a resource to nestmates by encoding celestial cues, retinal optic flow, and relative food value into motion and sound within the nest. We show that correct waggle dancing requires social learning. Bees without the opportunity to follow any dances before they first danced produced significantly more disordered dances with larger waggle angle divergence errors and encoded distance incorrectly. The former deficit improved with experience, but distance encoding was set for life. The first dances of bees that could follow other dancers showed neither impairment. Social learning, therefore, shapes honey bee signaling, as it does communication in human infants, birds, and multiple other vertebrate species.

Surface and back-side defects identification combined with magnetic flux leakage and boundary magnetic perturbation.

In magnetic flux leakage (MFL) detection, the identification of surface and back-side defects is required to obtain more accurate defect quantification and risk assessment results. However, current MFL techniques can detect both surface and back-side defects but are generally unable to distinguish between them. Therefore, this paper proposes a new boundary magnetic perturbation (BMP) testing method, combining the results of MFL to distinguish between surface and back-side defects. First, the detection mechanism of the BMP testing method and the impact of the tested magnetic flux density components are presented and analyzed by simulations to further develop an identification method. Then, the influences of the BMP sensor's lift-off and installation position are investigated by experiments to improve distinguishing performance. Finally, the repeated measurements show that the surface and back-side defects within the wide range of sizes can be identified accurately, even when the defect depths are in the range of 12.5%-87.5% of the sample thickness. Furthermore, the BMP testing method neither increases the length of the detection device nor requires additional magnetizers or signal generators. Therefore, the proposed method is highly suitable for the existing MFL detection devices to distinguish between surface and back-side defects.

Identification of giant hornet Vespa mandarinia queen sex pheromone components.

The Vespidae is a diverse family of wasps and hornets that are formidable predators of insects, including social bees1, and includes a number of invasive species2. Recently, the world's largest hornet, Vespa mandarinia Smith (Hymenoptera: Vespidae), which occurs naturally in the Indomalayan region, has been found in Canada and the United States2. Some simulations indicate that it could rapidly spread throughout Washington and Oregon in the western US, as well as some eastern parts of the country2,3, threaten native bees and honeybees, and harm bee-pollinated crop production worth over $100 million annually3. There is consequently an urgent need to learn more about V. mandarinia's reproductive biology and to develop trapping methods to locate its nests and to control its reproduction. We identified V. mandarinia queen-produced sex pheromone from the 5th and 6th intersegmental sternal glands of virgin queens. The major active compounds were hexanoic acid, octanoic acid, and decanoic acid. When placed in field traps, the synthetic compounds and a queen-equivalent mixture rapidly attracted hundreds of males but no females or other species.

Lethality of Honey Bee Stings to Heavily Armored Hornets.

The heat ball defense of honey bees against their sympatric hornet predators is a classic and spectacular outcome of a co-evolutionary race. Hundreds of bees can encapsulate a hornet within a large ball that kills it with elevated heat. However, the role of stinging in this defense has been discounted, even though sting venom is an important weapon in bees. Surprisingly, no studies have tested the role of bee sting venom alone or in conjunction with elevated temperature on hornet survival. We surveyed dead Vespa velutina hornets found near and inside Apis cerana colonies and found stings retained in hornet bodies, most often in an intersegmental neck-like region, the veracervix. Experimentally stinging hornets in this region with A. cerana and Apis mellifera guards significantly increased hornet mortality. The combination of sting venom and elevated heat ball temperature (44 °C) was the most lethal, although there was no synergistic interaction between sting venom and temperature. As expected, hornet mortality increased when they were stung more often. The average amount of venom per insect species and the length of stinger lancets correlated with insect mass. Sting venom thus remains important in the arms race between bees and their hornet predators.

The Nasonov gland pheromone as a potential source of death cue in Apis cerana.

The ability to detect and remove dead adult bees is an essential part of honeybee colony fitness that prevents the spread of pathogens. Fatty acid olfactory cues stimulate undertaking behavior among different social species within Hymenoptera, but the chemicals responsible for the death cue in Apis cerana have not yet been identified. We explored the Nasonov gland as a potential source of these chemicals in A. cerana. Gas chromatography indicated that unlike A. mellifera, the A. cerana Nasonov gland does not contain any volatile terpenes, only fatty acids. As a bioassay, dead honeybees were rinsed free of their individual cuticular hydrocarbons via dichloromethane and two concentrations of oleic acid and a synthetic blend of the Nasonov pheromone in A. cerana were applied to the dummies. Results showed that oleic acid did not stimulate corpse removal in A. cerana. However, the synthetic pheromone blend of A. cerana Nasonov did stimulate removal.

Early rehabilitation after lung transplantation with extracorporeal membrane oxygenation (ECMO) of COVID-19 patient: a case report.

Even when severe acute respiratory syndrome coronavirus 2-related coronavirus disease 2019 (COVID-19) is treated with first-line drugs, it progresses and leads to irreversible loss of lung function in some critically ill patients, and lung transplantation is an effective treatment for end-stage chronic pulmonary disease. This case report mainly describes the rehabilitation of a 66-year-old female patient with severe COVID-19 after bilateral lung transplantation. The old patient had a body mass index of 31.2 kg/m2. She underwent bilateral lung transplantation due to severe and irreversible injury of both lungs. Long-term mechanical ventilation and extracorporeal membrane oxygenation (ECMO) treatment and preoperative and postoperative high-dose corticosteroid therapy and due to the size of the donor lung does not match the size of the recipient's diseased lung, and the right middle lobe of the graft is removed before transplantation. Weaning from the ventilator failed due to weak neuromuscular drive, and muscle strength. A full, personalized pulmonary rehabilitation program was initiated with the help of the physical therapists, the respiratory therapy, the doctors, the nurses and psychotherapist team based on the functional levels. The rehabilitation intervention was conducted on postoperative day 4, This included posture management, airway clearance techniques, respiratory training, muscle strength training, transfer training, daily therapeutic bronchoscopy and psychological support. The ECMO was removed successfully on the fifth day. the patient's physical function, muscle strength and the quality of life has been improved. The good prognosis after rehabilitation indicates that early rehabilitation intervention is effective and feasible and safety for patients after lung transplantation.

Visual contagion in prey defence signals can enhance honest defence.

The co-evolutionary arms race between predators and their prey has led to complex signalling, especially in groups that benefit from the social transmission of alarm signals. In particular, pursuit deterrence signals can allow individuals and groups to indicate, at relatively low cost, that a predator's further approach is futile. Pursuit deterrence signals are usually more effective if amplified, for example, by becoming contagious and rapidly spreading among prey without requiring individual prey to confirm predator presence. However, this can also lead to runaway false signalling. We provide the first evidence of a contagious pursuit deterrence signal in social insects. The Asian honey bee Apis cerana, performs an I See You (ISY) signal that deters attacking hornets. We show that these signals enhance defensive signalling by also attracting guard bees and that the visual movements of appropriate stimuli alone (hornets and ISY signalling bees, but not harmless butterflies) provide sufficient stimuli. Olfaction and other potential cues are not necessary. The ISY signal is visually contagious and is buffered from runaway false signals because it is specifically triggered and by likely selection for honesty within the highly cooperative bee colony. These results expand our understanding of contagious signals and how they can be honestly maintained in highly cooperative collectives.

Floral tea polyphenols can improve honey bee memory retention and olfactory sensitivity.

Animal-pollinated plants face a common problem, how their defensive anti-herbivore compounds may impair or alter pollinator behavior. Evolution has tailored multiple solutions, which largely involve pollinator tolerance or manipulation, to the benefit of the plant, not the removal of these compounds from pollen or nectar. The tea plant, Camilla sinensis, is famous for the caffeine and tea polyphenols (TP) that it produces in its leaves. However, these compounds are also found in its nectar, which honey bees readily collect. We examined the effects of these compounds on bee foraging choices, learning, memory, and olfactory sensitivity. Foragers preferred a sucrose feeder with 100 µg or 10 µg TP/ml over a control feeder. Caffeine, but not TP, weakly increased honey bee learning. Both caffeine and TP significantly increased memory retention, even when tested 7 d after the last learning trial. In addition, TP generally elevated EAG responsiveness to alarm pheromone odors. These results demonstrate that other secondary plant compounds, not only caffeine, can attract pollinators and influence their learning and memory.

[Source Apportionment and Heath Risk Quantification of Heavy Metals in PM2.5 in Yangzhou, China].

Recently, a new method combining positive matrix factorization (PMF) and heavy metal health risk (HMHR) assessment was proposed to apportion sources of heavy metals in ambient particulate matter and the associated heavy metal cancer health risk (HMCR), which has been applied to data collected in Yangzhou, China. The annual average concentrations of six measured heavy metals were Pb (64.4 ng·m-3), followed by Cr (25.24 ng·m-3), As (6.36 ng·m-3), Ni (5.36 ng·m-3), Cd (3.34 ng·m-3), and Co (1.21 ng·m-3). The results showed that the major sources of PM2.5 were secondary sources (37.7%), followed by coal combustion (19.4%), resuspended dust (17.5%), vehicle emissions (16.9%), construction dust (5.2%), and industrial emissions (3.4%). As was primarily emitted from coal combustion, vehicle emissions, and resuspended dust. Co originated from industry emissions. Pb was mainly emitted from coal combustion. Ni and Cd were from industrial emissions. The major sources that contributed to HMCR were resuspended dust, coal combustion, vehicle emissions, industry emissions, and construction dust. The high contributions of resuspended dust and coal to HMCR were likely due to the high heavy metals concentrations in coal and the resuspended dust profile as well as high emissions of these sources.

PIK3CD induces cell growth and invasion by activating AKT/GSK-3ß/ß-catenin signaling in colorectal cancer.

The catalytic subunit p110δ of phosphoinositide 3-kinase (PI3K) encoded by PIK3CD has been implicated in some human solid tumors. However, its roles in colorectal cancer (CRC) remain largely unknown. Here we found that PIK3CD was overexpressed in colon cancer tissues and CRC cell lines and was an independent predictor for overall survival (OS) of patients with colon cancer. The ectopic overexpression of PIK3CD significantly promoted CRC cell growth, migration and invasion in vitro and tumor growth in vivo. In contrast, inhibition of PIK3CD by specific small-interfering RNA or idelalisib dramatically suppressed CRC cell growth, migration and invasion in vitro and tumor growth in vivo. Moreover, PIK3CD overexpression increased AKT activity, nuclear translocation of ß-catenin and T-cell factor/lymphoid enhancer factor (TCF/LEF) transcriptional activity and decreased glycogen synthase kinase 3ß (GSK-3ß) activity, whereas PIK3CD inhibition exhibited the opposite effects. Furthermore, PIK3CD-mediated cell growth, migration and invasion were reversed by blockade of AKT signaling or depletion of ß-catenin. In addition, PIK3CD expression in colon cancer tissues positively correlated with ß-catenin abnormal expression, which was an independent predictor for OS of colon cancer patients. Taken together, our findings demonstrate that PIK3CD is an independent prognostic factor in CRC and that PIK3CD induces CRC cell growth, migration and invasion by activating AKT/GSK-3ß/ß-catenin signaling, suggesting that PIK3CD might be a novel prognostic biomarker and a potential therapeutic target for CRC.

Interval colorectal cancer: a challenging field in colorectal cancer.

Accumulated evidence has shown that colonoscopy may not be a perfect tool in screening and reducing the incidence of the colorectal cancer (CRC), because interval CRC (I-CRC), a specific subgroup of CRCs, has been challenging the traditional detection technology in recent years. I-CRC is accounting for an increasing proportion in CRCs. However, the effective procedures to prevent and supervise I-CRC need to be explored. In this review, we summarized the incidence, causes, risk factors, characteristics and management of I-CRC. It would promote the awareness of the special value in the education and training for the gastroenterologists, which plays an important role in conquering CRC.

The pesticide flupyradifurone impairs olfactory learning in Asian honey bees (Apis cerana) exposed as larvae or as adults.

Relatively little attention has focused on how pesticides may affect Asian honey bees, which provide vital crop pollination services and are key native pollinators. We therefore studied the effects of a relatively new pesticide, flupyradifurone (FLU), which has been developed, in part, because it appears safer for honey bees than neonicotinoids. We tested the effects of FLU on Apis cerana olfactory learning in larvae (lower dose of 0.033 µg/larvae/day over 6 days) and, in a separate experiment, adults (lower dose of 0.066 µg/adult bee/day) at sublethal, field-realistic doses given over 3 days. A worst-case field-realistic dose is 0.44 µg/bee/day. Learning was tested in adult bees. The lower larval dose did not increase mortality, but the lower adult dose resulted in 20% mortality. The lower FLU doses decreased average olfactory learning by 74% (larval treatment) and 48% (adult treatment) and reduced average memory by 48% (larval treatment) and 22% (adult treatment) as compared to controls. FLU at higher doses resulted in similar learning impairments. The effects of FLU, a pesticide that is reported to be safer than neonicotinoids for honey bees, thus deserve greater attention.

The sex pheromone of a globally invasive honey bee predator, the Asian eusocial hornet, Vespa velutina.

The Asian hornet, Vespa velutina, is an invasive, globally-distributed predator of European honey bees and other insects. To better under its reproductive biology and to find a specific, effective, and low-impact control method for this species, we identified and tested the key compounds in V. velutina sex pheromone. Virgin gynes (reproductive females) produced this sex pheromone in the sixth intersegmental sternal glands of their abdomens. The active compounds were 4-oxo-octanoic acid (4-OOA, 10.4 µg bee-1) and 4-oxo-decanoic acid (4-ODA, 13.3 µg bee-1) at a 0.78 ratio of 4-OOA/4-ODA. We synthesized these compounds and showed that male antennae were highly sensitive to them. Moreover, males were only strongly attracted to a 4-OOA/4-ODA blend at the natural ratio produced by gynes. These results provide the first demonstration of an effective way to lure V. velutina males, and the first chemical identification of a sex pheromone in the eusocial hornets.

Corrigendum: Resisting majesty: Apis cerana, has lower antennal sensitivity and decreased attraction to queen mandibular pheromone than Apis mellifera.

This corrects the article DOI: 10.1038/srep44640.

Resisting majesty: Apis cerana, has lower antennal sensitivity and decreased attraction to queen mandibular pheromone than Apis mellifera.

In highly social bees, queen mandibular pheromone (QMP) is vital for colony life. Both Apis cerana (Ac) and Apis mellifera (Am) share an evolutionarily conserved set of QMP compounds: (E)-9-oxodec-2-enoic acid (9-ODA), (E)-9-hydroxydec-2-enoic acid (9-HDA), (E)-10-hydroxy-dec-2-enoic acid (10-HDA), 10-hydroxy-decanoic acid (10-HDAA), and methyl p-hydroxybenzoate (HOB) found at similar levels. However, evidence suggests there may be species-specific sensitivity differences to QMP compounds because Ac workers have higher levels of ovarian activation than Am workers. Using electroantennograms, we found species-specific sensitivity differences for a blend of the major QMP compounds and three individual compounds (9-HDA, 10-HDAA, and 10-HDA). As predicted, Am was more sensitive than Ac in all cases (1.3- to 2.7- fold higher responses). There were also species differences in worker retinue attraction to three compounds (9-HDA, HOB, and 10-HDA). In all significantly different cases, Am workers were 4.5- to 6.2-fold more strongly attracted than Ac workers were. Thus, Ac workers responded less strongly to QMP than Am workers, and 9-HDA and 10-HDA consistently elicited stronger antennal and retinue formation responses [corrected].

The dynamic association between ovariole loss and sterility in adult honeybee workers.

In the social insects, ovary state (the presence or absence of mature oocytes) and ovary size (the number of ovarioles) are often used as proxies for the reproductive capacity of an individual worker. Ovary size is assumed to be fixed post-eclosion whereas ovary state is demonstrably plastic post-eclosion. Here, we show that in fact ovary size declines as honeybee workers age. This finding is robust across two honeybee species: Apis mellifera and A. cerana The ovariole loss is likely to be due to the regression of particular ovarioles via programmed cell death. We also provide further support for the observation that honeybee workers with activated ovaries (mature oocytes present) most commonly have five ovarioles rather than a greater or smaller number. This result suggests that workers with more than five ovarioles are unable to physiologically support more than five activated ovarioles and that workers with fewer than five ovarioles are below a threshold necessary for ovary activation. As a worker's ovariole number declines with age, studies on worker ovariole number need to take this plasticity into account.