Confirmation of QTL effects and evidence of genetic dominance of honeybee defensive behavior: results of colony and individual behavioral assays

The stinging and guarding components of the defensive behavior of European, Africanized, hybrid, and backcross honeybees (Apis mellifera L.) were compared and analyzed at both colony and individual levels. Hybrid and Africanized backcross colonies stung as many times as Africanized ones. European backcross colonies stung more than European bees but not as many times as Africanized or Africanized backcross colonies. The degree of dominance for the number of times that worker bees stung a leather patch was estimated to be 84.3%, 200.8%, and 145.8% for hybrid, backcross European, and backcross Africanized colonies, respectively. Additionally, both guards at the colony entrance and fast-stinging workers of one European backcross colony had a significantly higher frequency of an Africanized DNA marker allele, located near sting1, a QTL previously implicated in stinging behavior at the colony level. However, guards and fast-stinging bees from a backcross to the Africanized parental colony did not differ from control bees in their frequency for the Africanized and European markers, as would be expected if large genetic dominance effects for sting1 exist. These results support the hypothesis that genetic dominance influences the defensive behavior of honeybees and confirm the effect of sting1 on the defensiveness of individual worker bees.     

Quantitative trait loci that influence the expression of guarding and stinging behaviors of individual honey bees

This study was conducted to test for the effect of three stinging behaviors QTLs (sting-1, sting-2 and sting-3) on the expression of guarding and stinging behavior of individual honey bees, and to determine if results of defensive behavior QTLs found in studies with Africanized honey bees could be extended to other populations of bees. Samples of guards, stingers, foragers and nurse bees were taken from two backcross colonies derived from a defensive colony and a gentle colony. The genotype of each bee for both types of colonies was determined for two sequence tagged site (STS) markers linked to sting-1 and for another two STSs, one linked to sting-2 and one linked to sting-3. Results showed that sting-1 had an effect on the expression of both stinging and guarding behaviors, sting-2 and sting-3 influenced the expression of guarding behavior. These results indicate that division of labor is influenced by specific QTLs. Results also show that QTLs mapped in a population of Africanized honey bees using colony level phenotypes also influenced the expression of guarding and stinging behavior of individual bees of other populations.     

A genetic analysis of the stinging and guarding behaviors of the honey bee

In order to identify genes that are influencing defensive behaviors, we have taken a new approach by dissecting colony-level defensive behavior into individual behavioral measurements using two families containing backcross workers from matings involving European and Africanized bees. We removed the social context from stinging behavior by using a laboratory assay to measure the stinging response of individual bees. A mild shock was given to bees using a constant-current stimulator. The time it took bees to sting in response to this stimulus was recorded. In addition, bees that were seen performing guard behaviors at the hive entrance were collected. We performed QTL mapping in two backcross families with SNP probes within genes and identified two new QTL regions for stinging behavior and another QTL region for guarding behavior. We also identified several candidate genes involved in neural signaling, neural development and muscle development that may be influencing stinging and guarding behaviors. The lack of overlap between these regions and previous defensive behavior QTL underscores the complexity of this behavior and increases our understanding of its genetic architecture.     

Biochemical variability of venoms from individual European and Africanized honeybees (Apis mellifera).

To study biochemical differences between venom from individual honeybees, venom sacs from 103 European (EU) bees and 92 Africanized bees representing 12 different colonies were dissected, and the dry weight (DW) of venom from each bee was determined. Venom from each of these bees was studied with isoelectric focusing and functional assays for phospholipase A2 and melittin. Phospholipase concentrations in individual EU bee venoms varied between 1.8% and 27.4% (wt/wt). The melittin concentration in EU bee venom varied less and, on the average, was found to be much lower than previously reported. There was an eightfold to ninefold difference between lowest and highest venom sac DW contents, suggesting the possibility of highly variable venom delivery from bee stings. One EU bee contained greater than 300 micrograms of venom, three times the recommended maintenance dose for venom immunotherapy. Isoelectric focusing also demonstrated large differences between individual bees, with respect to major and minor components of their venoms. Africanized bees contained significantly less venom but more phospholipase than did EU bees. Bee venoms from different colonies differed in their DW content and in their concentrations of phospholipase and melittin. The results are relevant to the uncertainty of responses from sting challenges and field stings in allergic patients and massive stinging attacks on normal subjects.     

The behavioral genetics of colony defense in honeybees: Genetic variability for guarding behavior

Guard honeybees stand at the entrance of colonies and facilitate the exclusion of nonnestmates from the colony. In this study, we examined the hypothesis that genetic variability among individuals in colonies might explain variability in guarding activity. To do this, we cross-fostered honey bees between colonies with high-defensive responses and colonies with low-defensive responses in alarm pheromone tests. Individuals from high-defensive colonies were more likely to guard in their own colonies (controls) than cross-fostered bees from low-defensive colonies. Cross-fostered high-defensive bees also were more likely to guard in low-defense colonies. These results support the hypothesis that interindividual differences in guarding behavior are at least partially under genetic control. A positive correlation between number of guards and response to alarm pheromone demonstrates a link between behaviorally separated components of the overall defensive response.This work was supported by NSF Grant BNS 8605604.     

Olfactory stimulation of Africanized honey bee (Hymenoptera: Apidae) attacks by insect repellents

Three common insect repellents (N,N-diethyl-m-toluamide [DEET], Pyranha, and Repel X) were tested to determine whether they affected Africanized honey bee attack behavior. Eight Africanized honey bee (Apis mellifera L.) colonies were exposed in an alternating series to the test repellents or blank controls delivered in a stream of air directed toward the colony entrances. The response generated by the repellents and the controls was measured as the number of attacking honey bees recorded with an electronic temper tester. Neither a citronella-based repellent (Pyranha) nor DEET had any effect on colony behavior; however, Repel X consistently caused a greater attack response after exposure.     

Avoidance of bee and wasp stings: an entomological perspective

PURPOSE OF REVIEW: Clinicians and researchers in allergy and immunology are often unaware of aspects of stinging insect biology that would be of practical interest to their patients. This review discusses entomological literature pertaining to avoidance of bee and wasp stings, with emphasis on risk factors associated with provoking individual foragers versus disturbing colonies and preventive measures for both circumstances. RECENT FINDINGS: Recent work pertaining to sting avoidance has mostly been concerned with the development and testing of attractants, insecticides and delivery systems for toxic baiting programs to control vespine wasps. SUMMARY: Sting risks and avoidance measures associated with bee and wasp foragers are different from those posed by disturbing colonies. Despite widespread advice to the contrary, no evidence currently exists that wearing perfume or bright, floral-colored clothing elevates sting risk. Foragers usually have to be firmly touched before they will sting; therefore, personal protection largely involves guarding against accidental direct contact. Although still under development, the most effective means for reducing local populations of foraging vespine wasps are toxic baiting programs. Preventing stings from colonies is more problematic and depends mostly on personal awareness when disturbing vegetation. The most effective measure in mitigating the severity of a mass attack is probably the wearing of white or light-colored clothing.     

Genetic study of the aggressiveness of two subspecies ofApis mellifera in Bazil. IV. Number of stings in the gloves of the observer

Data are analyzed on an aspect of aggressiveness in workers from colonies of Africanized bees (Apis mellifera adansonii), Italian bees (Apis mellifera ligustica), their F ₁ hybrids, and backcrosses of the F ₁ to the parental stocks (Rothenbuhler method). The segregation values (3:1) in the backcrosses to the Africanized stock and nonsegregation in the backcrosses to the Italian stock suggest the existence of two pairs of genes (F₁/F₁;F₂/F₂ in the Italian bees and f₁/f₁;f₂/f₂ in the Africanized bees) which control a character defined as the number of stings in the gloves of the observer.This work was carried out in the Department of Genetics, Faculdade de Medicina de Ribeirão Preto, and in the Laboratory of Biology, Faculdade de Filosofia, Ciências, e Letras de Araraquara, with the financial assistance of Fundação de Amparo à Pesquisa do Estado de São Paulo. This paper is part of the author's doctoral dissertation.     

Genetic differences in learning behavior in honeybees (Apis mellifera capensis)

Workers from colonies of Cape honeybees show marked phenotypic differences in performance in proboscis extension reflex (PER) conditioning. Analysis of these differences using parthenogenetic offspring groups permitted the estimation of genotypic values and revealed a high degree of genetic variability that is evident among related as well as unrelated bees. The results obtained from related groups are of particular importance, since they demonstrated the existence of strong genetic variability among individuals of the same colony. Quantitative analysis yielded high estimates of additive genetic effects and low estimates of dominance effects. Selection of individual workers resulted in an explicit increase in genetic variance of the next generation (G1). However, selection of bees from the parthenogenetic G1 generation, which was done to obtain parthenogenetic G2 offspring, did not lead to further improvement in selection. This observation suggests that recombination of linked genes underlying proboscis extension reflex was neglible during selection in parthenogenetic groups. Taken together with further behavioral analysis (Brandes and Menzel, 1990; Brandes et al., 1988), results from these quantitative genetic experiments suggest that additive genetic factors contribute significantly to variability among individuals for associative learning.This work was supported by a fellowship from the Deutsche Forschnungsgemmeinschaft (Br 827/1, Br 827/2).     

Intergenerational transfers may have decoupled physiological and chronological age in a eusocial insect

Life-history theory generally predicts that there should be no selection for longevity beyond the limit of reproductive capacity. However, the capacity to increase fitness may not end when individuals reach a state of functional sterility. Recent studies show that intergenerational transfers of resources from post-reproductive parents can increase the offspring's fitness, and analytical theory shows that age-trajectories of transfers may shape the course of senescence in social organisms. In eusocial insects, female roles are partitioned so that one phenotype or "caste" reproduces while another is responsible for resource transfers: the reproductive "queens" are arrested in a continuous reproductive mode, while transfer-activities such as hygienic behaviors, guarding, foraging and further food processing ("nursing") that increases the nutritional value of provisions are conducted by sterile "workers". Worker honey bees normally perform these tasks in a sequence so that nursing inside the protected nest is conducted prior to more risky exterior hive activities such as guarding and foraging. However, foragers may revert to nurse-activity in response to demographic changes, and worker bees can also develop into a stress resistant survival form with a 10-fold increase in lifespan. This elastic division of parental functions is believed to increase colony fitness. Further, it generates a stage-dependent trajectory of senescence that is difficult to address with established theories of aging. In the following, we show how a recent theory that includes resource transfers can be used to elucidate patterns of senescence in eusocial, non-reproducing individuals like the honey bee worker.     

Analgesia on electrical stimulation of mesencephalic nuclei in rats with a pain syndrome of spinal origin

Effects of electrical stimulation of the periventricular gray matter and dorsal nucleus raphe in the midbrain on physiological pain induced by nociceptive stimulation (crushing the tail or skin of the limbs with forceps) and on pathological pain (a pain syndrome of spinal origin) were studied in experiments on albino rats. This last pain syndrome was produced by creating a generator of pathologically enhanced excitation in the posterior horns of the spinal cord with the aid of tetanus toxin, which disturbs various types of inhibition. Electrical stimulation of the above structure was shown to depress both physiological and pathological pain. It is concluded that analgesia during electrical stimulation of brain structures is connected not only with the strengthening of descending inhibition in the spinal cord, as in the case of physiological pain caused by peripheral nociceptive stimulation (as several workers have shown), but also with blocking the spread of excitation at the supraspinal level. This mechanism must play the decisive role in the production of analgesia in pain syndromes of central origin, including those arising under natural conditions.Laboratory of General Pathology of the Nervous System, Institute of General Pathology and Pathological Physiology, Academy of Medical Sciences of the USSR, Moscow. Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 84, No. 7, pp. 16–19, July, 1977.     

Learning and memory in workers reared by nutritionally stressed honey bee (Apis mellifera L.) colonies

Chronic nutritional stress can have a negative impact on an individual's learning ability and memory. However, in social animals that share food among group members, such as the honey bee (Apis mellifera L.), it is unknown whether group-level nutritional stress is manifested in the learning performance of individuals. Accordingly, we examined learning and memory in honey bee workers reared by colonies exposed to varying degrees of long-term pollen stress. Pollen provides honey bee workers with almost all of the proteins, lipids, vitamins, and minerals that they require as larvae and adults. Colonies were created that were either chronically pollen poor or pollen rich, or were intermediate in pollen supply; treatments altered colonies' pollen stores and brood-rearing capacity. Workers from these colonies were put through a series of olfactory-conditioning assays using proboscis-extension response (PER). PER thresholds were determined, then workers learned in olfactory-conditioning trials to associate two floral odors (one novel and the other presented previously without reward) with stimulation with sucrose and a sucrose reward. The strength of the memory that was formed for the odor/sucrose association was tested after olfactory-conditioning assays ended. Colony-level nutritional status had no effect on worker learning or memory (response threshold of workers to sucrose, acquisition of the odor/sucrose association, occurrence of latent inhibition, or memory retention over 72 h). We conclude that potential effects of chronic, colony-wide nutrient deprivation on learning and memory are not found in workers, probably because colonies use brood-rearing capacity to buffer nutrient stress at the level of the individual.     

Blood pressure reflexes to stimulation of tibial nerve a fibers in mesencephalic and bulbar cats

The role of the various subgroups of A fibers of the tibial nerve (pulse frequency of electrical stimulation 10/sec) in the formation of reflex changes in blood pressure (BP) was investigated in unanesthetized cats with total transection of the brain stem at the level of the pontomedullary junction (bulbar animals) or at the rostral border of the mesencephalon (mesencaphalic animals), and also in anesthetized cats with an intact brain. The lowest thresholds for the reflexes were found in anesthetized animals with an intact brain, the highest in bulbar cats. Excitation of A fibers in anesthetized cats with an intact brain evoked only depressor reflexes. In some bulbar and mesencephalic animals only pressor reflexes appeared. In the experiments of this group excitation of fibers with a conduction velocity of over 15 m/sec in mesencephalic cats evoked reflexes of near maximal strengths, whereas in bulbar cats excitation of thinner A fibers also was necessary. In unanesthetized animals disconnection of the suprabulbar structures thus lowers the sensitivity of the central mechanisms of vasomotor regulation to impulses in lowthreshold A fibers. No such effect was found in another group of experiments in which depressor reflexes appeard in response to stimulation of fast-conducting A fibers only. In these experiments, if slower A fibers also were stimulated, the reflexes became pressor but the difference between their magnitude in the bulbar and mesencephalic cats was not signicant.Laboratory of Biophysics and Pathophysiology of the Circulation, Institute of General Pathology and Pathological Physiology, Academy of Medical Sciences of the USSR, Moscow. (Presented by Academician of the Academy of Medical Sciences of the USSR A. M. Chernukh.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 84, No. 10, pp. 393–396, October, 1977.     

Antagonism between L-dopa and apomorphine in their effects on rat behavior

The effect of apomorphine and L-dopa on the apomorphine behavioral stereotype, on aggressiveness, and on the thresholds of emotional reactivity and aggressiveness induced by painful electrical stimulation, and on orienting motor activity was studied in male albino rats. Whereas the action of these substances given separately was similar in direction, definite antagonism was observed between apomorphine and L-dopa when given together with respect to all behavioral tests. Apomorphine (5 mg/kg), given after L-dopa, increased the dopamine concentration even higher in the forebrain and diencephalon without affecting the noradrenalin level. It is suggested that an increased concentration of functionally active mediator inhibits the activity of postsynaptic receptors sensitive to it.Department of Pharmacology, Tartu University. (Presented by Academician of the Academy of Medical Sciences of the USSR S. V. Anichkov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 84, No. 7, pp. 51–53, July, 1977.     

Role of interoceptive impulses in the modulation of emotional and behavioral responses

Chronic experiments on rabbits with a gastric fistula and electrodes implanted into deep brain structures showed that stimulation of the gastric receptors leads to modulation of emotional and behavioral responses evoked by electrical stimulation of the hypothalamus, amygdala, and hippocampus. The effect depends on the intensity of interoceptive stimulation and on nature of the emotional response, which has its own cerebral control systems.Department of Physiology, Ivan-Frankovsk Medical Institute. (Presented by Academician V. N. Chernigovskii.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 81, No. 3, pp. 264–266, March, 1976.     

Nociceptive muscle reflex responses in human forearms

In healthy volunteers, 5-min noxious stimulation with rectangular electrical pulses applied transcutaneously to the phalanges enhanced tonic activity in the palmar and finger flexor muscles resulting in a specific electromyographic pattern consisting of two successive bursts of activity which appeared after a period of inhibition. In patients with chronic pain in the arm, significantly lower thresholds for the first and the second waves of reflex activity have been found. This electromyographic pattern of the forearm muscle reflex responses is supposed to be similar to the nociceptive flexor reflex in the leg. It can be useful for objective assessment of the effectiveness of analgesia and pain syndrome therapy in patients with cervical spinal cord injury. Translated fromByulleten' Eksperimental'noi Biologii I Meditsiny, Vol. 126, No. 9, pp. 278–282, September, 1998     

Effects of functional electrical stimulation on the urethral closing muscles

The importance of the selection of patients with urinary incontinence suitable for treatment with functional electrical stimulation (f.e.s.) was tested. 40 patients with urinary incontinence of different kinds (mostly stress incontinence) were selected on the basis of urological, neurophysiological and urodynamic examinations. The positive response to optimal f.e.s. was a criterion for urodynamic selection. Mechanical properties of the unstimulated and electrically stimulated urethral wall tissue in the functional part of the urethra were studied. The differences between the acute effects of different types of stimulation, namely monophasic and biphasic f.e.s. with vaginal or different types of anal plugs and mechanical stimulation produced by these plugs, were stated. An attempt was made to quantify the success of treatment with f.e.s. Besides the routine urological, neurophysiological and urodynamic methods, the investigative methods of measuring the urethral pressure profile (u.p.p.) and measuring the pressure at one point of the urethra, by means of Mikro-Tip pressure transducers, were used in our study. Errors due to the dimensions of the measuring device causing a nonphysiological dilatation of the urethra were considered in the method of measuring the u.p.p.     

Lactic acid as a factor in changes of vascular reactivity

Results showing that contractile responses of isolated vascular segments to electrical stimulation are replaced by relaxation if the preparations are contained in a solution with lactic acid are described. Ability to contract is not completely lost under these circumstances but it is manifested only during very strong stimulation. The changes in the character of response to electrical stimulation are connected both with changes in the pH of the solution and with the specific action of the lactic acid.Laboratory of Physiology and Pathophysiology of the Circulation, A. L. Myasnikov Institute of Cardiology, Academy of Medical Sciences of the USSR, Moscow. (Presented by Academician of the Academy of Medical Sciences of the USSR E. I. Chazov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 82, No. 8, pp. 923–926, August, 1976.     

Electrical cochlear stimulation in the deaf cat: comparisons between psychophysical and central auditory neuronal thresholds

Cochlear prostheses for electrical stimulation of the auditory nerve ("electrical hearing") can provide auditory capacity for profoundly deaf adults and children, including in many cases a restored ability to perceive speech without visual cues. A fundamental challenge in auditory neuroscience is to understand the neural and perceptual mechanisms that make rehabilitation of hearing possible in these deaf humans. We have developed a feline behavioral model that allows us to study behavioral and physiological variables in the same deaf animals. Cats deafened by injection of ototoxic antibiotics were implanted with either a monopolar round window electrode or a multichannel scala tympani electrode array. To evaluate the effects of perceptually significant electrical stimulation of the auditory nerve on the central auditory system, an animal was trained to avoid a mild electrocutaneous shock when biphasic current pulses (0.2 ms/phase) were delivered to its implanted cochlea. Psychophysical detection thresholds and electrical auditory brain stem response (EABR) thresholds were estimated in each cat. At the conclusion of behavioral testing, acute physiological experiments were conducted, and threshold responses were recorded for single neurons and multineuronal clusters in the central nucleus of the inferior colliculus (ICC) and the primary auditory cortex (A1). Behavioral and neurophysiological thresholds were evaluated with reference to cochlear histopathology in the same deaf cats. The results of the present study include: 1) in the cats implanted with a scala tympani electrode array, the lowest ICC and A1 neural thresholds were virtually identical to the behavioral thresholds for intracochlear bipolar stimulation; 2) behavioral thresholds were lower than ICC and A1 neural thresholds in each of the cats implanted with a monopolar round window electrode; 3) EABR thresholds were higher than behavioral thresholds in all of the cats (mean difference = 6.5 dB); and 4) the cumulative number of action potentials for a sample of ICC neurons increased monotonically as a function of the amplitude and the number of stimulating biphasic pulses. This physiological result suggests that the output from the ICC may be integrated spatially across neurons and temporally integrated across pulses when the auditory nerve array is stimulated with a train of biphasic current pulses. Because behavioral thresholds were lower and reaction times were faster at a pulse rate of 30 pps compared with a pulse rate of 2 pps, spatial-temporal integration in the central auditory system was presumably reflected in psychophysical performance.