Honey Bee Brood Research Articles

Sucrose response thresholds of honey bee (Apis mellifera) foragers are not modulated by brood ester pheromone

Abstract Division of labor is a hallmark of eusocial insects and their ecological success can be attributed to it. Honey bee division of labor proceeds along a stereotypical ontogenetic path based on age, modulated by various internal and external stimuli. Brood pheromone is a major social pheromone of the honey bee that has been shown to affect honey bee division of labor. It elicits several physiological and behavioral responses; notably, regulating the timing of the switch from performing in-hive tasks to the initiation of foraging. Additionally, brood pheromone affects future foraging choice. In honey bees, sucrose response threshold is a physiological correlate of age of first foraging and foraging choice. Brood pheromone has been shown to modulate sucrose response threshold in young bees, but its effects on sucrose response thresholds of bees in advanced behavioral states (foragers) are not known. In this study we examined the sucrose response thresholds of two different task groups, foragers (pollen and non-pollen) and non-foraging bees, in response to honey bee brood pheromone. Sucrose response thresholds were not significantly different between brood pheromone treatment and controls among both non-pollen and pollen foragers. However, the sucrose response threshold of non-foraging bees was significantly higher in the brood pheromone treatment group than in the control group. The switch to foraging task is considered a terminal one, with honey bee lifespan being determined at least partially by risks and stress accompanying foraging. Our results indicate that foragers are physiologically resistant to brood pheromone priming of sucrose response thresholds.

Prevalence of American foulbrood in asymptomatic apiaries of Kurdistan, Iran.

Aim:Paenibacillus larvae subsp. larvae is the etiological agent of American foulbrood (AFB), the most virulent bacterial disease of honey bee brood worldwide. In many countries, AFB is a notifiable disease since it is highly contagious, in most cases incurable, and able to kill affected colonies. The aim of this study was to determine the prevalence of P. larvae subsp. larvae in Kurdistan province apiaries by polymerase chain reaction (PCR) technique.Materials and Methods:A total of 100 samples were randomly purchased from apiaries in Kurdistan, Iran. Apiaries were randomly sampled in accordance with the instructions of the veterinary organization from different provinces and were tested using PCR method and an exclusive primer of 16S rRNA for the presence of P. larvae subsp. larvae.Results:The results of this study indicated a low level of contamination with P. larvae subsp. larvae in the Kurdistan province. The number of positive samples obtained by PCR was 2%.Conclusion:Therefore, monitoring programs for this honeybee disease in Kurdistan should be developed and implemented to ensure that it is detected early and managed.

Population structure and antimicrobial susceptibility of Paenibacillus larvae isolates from American foulbrood cases in Apis mellifera in Japan.

Paenibacillus larvae is the causative agent of American foulbrood (AFB), the most destructive disease of the honey bee brood. In this study, we investigated the population structure and antimicrobial susceptibility of Japanese P. larvae using 100 isolates isolated between 1993 and 2017 in 17 prefectures. Using repetitive-element PCR and multilocus sequence typing, isolates from diverse origins were classified into six genotypes, including the novel genotype ERIC II-ST24. Among these genotypes, ERIC I-ST15 is the most common in Japan, while ERIC II-ST10 isolates were found to be increasing during the 2010s. Regardless of genotype or origin, all isolates were susceptible to the major antimicrobials used in the control of AFB, including mirosamicin and tylosin, which were approved for the prevention of AFB in Japan in 1999 and 2017 respectively. Despite nearly 20years of use, mirosamicin is still effective against Japanese P. larvae in vitro; however, the development of AFB in honey bee colonies may not always be suppressed by this drug. The case information collected in this study provides insight into the conditions under which prophylactic medicine may not exert sufficient preventive effects in vivo.

Bacterial Diseases in Honeybees

Purpose of Review American foulbrood (AFB) and European foulbrood (EFB) are widely distributed and highly infectious bacterial diseases of honeybee brood causing colony losses and considerable economic strain on apiculture globally. In this review, we synthesize the most recent discoveries and achievements made towards understanding the pathogenesis and epidemiology of these two bacterial diseases and present current efforts in finding ways to combat them. Recent Findings Advancements in molecular methods, such as next-generation sequencing, have provided high-resolution insight into the epidemiological parameters and factors of virulence for the foulbroods of honeybees. Summary The recently gained detailed knowledge of the diversity, biogeography, and relatedness of strains and sub-types of the causative bacteria of AFB and EFB provides a background to study their epidemiology at many scales. Such information will help provide a more global perspective on honeybee disease epidemiology for an increasingly international beekeeping industry.

Mineral composition of haemolymph of honey bee brood infested with varroatosis

Mineral composition of haemolymph of honey bee brood infested with varroatosis

Spermatozoa production in male Varroa destructor and its impact on reproduction in worker brood of Apis mellifera.

Reproduction in Varroa destructor exclusively takes place within the sealed honey bee brood cell and is, therefore, limited by the duration of the postcapping period. Oogenesis, ontogenetic development and mating must be optimized to ensure the production of as many mated daughter mites as possible. One adult male mite has to mate with up to five sister mites and transfer 30-40 spermatozoa to each female. We analyzed the production and transfer of male spermatozoa during a reproductive cycle by counting all spermatozoa in the genital tracts of the male and daughter mites in 80 worker brood cells at defined times after cell capping. We could show that spermatozoa production in male mites is an ongoing process throughout their adult lifetime starting after the adult molt. The spermatozoa are transferred to the females in an early non-capacitated stage and require further maturation within the female's genital tract. Our study points out that a Varroa male has at any time in the brood cell enough spermatozoa to inseminate all daughter mites but does not waste energy in producing a big surplus. In total one male produced, on average, 125 spermatozoa during a reproductive cycle in worker brood which is sufficient for successful matings with at least three daughter mites. Spermiogenesis in Varroa males represents therefore a further adaptation to the limited time available for reproduction.

Can Food Supplement Produced from Apilarnil be an Alternative to Testosterone Replacement Therapy

Apilarnil is male honey bee (Apis mellifera) brood homogenate, firstly used on elders have psychotic, neurodegenerative or sexual disorders in Romania by Nicola Iliesu in 1980. It has high nutritional value and a cheap source of biologically active substances. It consists of water (65–75%), proteins (9–12%), carbohydrates (6–12%), fatty acids and lipids (3.5–8%), minerals K, Na, Ca, Mg (1–1.5%), essential amino acids (threonine, leucine, isoleucine, methionine) and also it is rich in sex hormones such as testosterone, prolactin, progesterone and estradiol. Some defined chemicals in lipid portion and hormones are included in Apilarnil should be responsible by androgenic effect. So, it may enhance the androgenic activity for patients with androgen deficiency syndrome. Lower androgenic activity named as androgen deficiency syndrome. It causes reduced growth of penis end testes in early ages, failure to go through full normal puberty, gynaecomastia, poor development of facial, body or pubic hair and voice does not deepen in early teenagers, mood changes, reduced muscle strength, increased body fat, decreased libido, difficulty getting and keeping erections, low semen volume and gynaecomastia in adults. In this article, Apilarnil having an androgenic content and whether or not it is effective for the androgen deficiency syndrome has been prepared and interpreted by compiling studies on Apilarnil in the literature.

Transfer of the Active Ingredients of Some Plant Protection Products from Raspberry Plants to Beehives

Plant protection products (PPPs) have been found increasingly in the environment. They pose a huge threat to bees, contributing to honeybee colony losses and consequently to enormous economic losses. Therefore, this field investigation was designed to determine whether their active ingredients (AIs) were transferred from raspberry plants to beehives located in the immediate neighbourhood of the crop and to what extent they were transferred. Every week for 2 months, samples of soil, raspberry leaves, flowers and fruits, worker bees, honeybee brood, and honey were collected and analysed for the presence of propyzamide, chlorpyrifos, iprodione, pyraclostrobin, boscalid, cypermethrin, difenoconazole, azoxystrobin, and pyrimethanil residues. Five of these substances were found in the worker bee bodies. Chlorpyrifos, applied to only the soil through the irrigation system, also was detected in the brood. A small amount of boscalid was noted in the honey, but its residues did not exceed the maximum residue level. For chlorpyrifos, boscalid, and pyrimethanil, a positive correlation between the occurrence of PPPs in the crops and the beehives was found. Statistical methods confirmed that the application of PPPs on a raspberry plantation, as an example of nectar-secreting plants, was linked to the transfer of their AIs to beehives.

The honey bee brood test under semi-field conditions for the assessment of positive reference chemicals in Korea

Honey bee brood tests under semi-field conditions are higher-tier studies for investigating the potential impact of pesticides on brood development in honey bee colonies. Semi-field studies on the effects of pesticides on honey bees have been mainly carried out in the EU and USA, with only a relatively small number conducted in Korea and other Asian countries. Here, we report the first semi-field test in Korea, which was carried out from April to May 2016. The experiment included three treatment groups (control and two toxic reference chemicals), each with three replicate tunnels. The honey bee colonies were placed in tunnels covering 70 m2 and containing Brassica napus. Flight activity, mortality, the condition of the colonies, and brood development were assessed during the 28 days of testing period. The toxic reference treatments were 400 g dimethoate a.i./ha and 600 g diflubenzuron a.i./ha. Brood termination rates for marked eggs were 31.3% in the control group, 83.5 and 68.0% for dimethoate and diflubenzuron, respectively. These results confirm the sensitivity of the test method and indicate that these two chemicals could be used as appropriate toxic reference compounds in future semi-field tests in Korea.

Diagnosis and molecular detection of Paenibacillus larvae, the causative agent of American foulbrood in honey bees in Saudi Arabia

AbstractApis mellifera jemenitica, the only indigenous honey bee race of Saudi Arabia, is well adapted to the harsh local environmental conditions. A large-scale field survey was conducted to screen major Saudi Arabian beekeeping locations for infection byPaenibacillus larvae. Paenibacillus larvaeis one of the major bacterial pathogens of honey bee broods and is the causative agent of American foulbrood disease. Larvae from samples suspected of infection were collected from different apiaries and homogenized in phosphate-buffered saline. Bacteria were isolated on MYPGP agar medium. Two bacterial isolates, ksuPL3 and ksuPL5 (16S rRNA GenBank accession numbers, KR780760 and KR780761, respectively), were subjected to molecular identification usingP. larvae-specific primers. A BLAST sequence analysis revealed that the two isolates wereP. larvaewith more than 98% sequence identity. This detection ofP. larvaein the indigenous honey bee is the first recorded incidence of this pathogen in Saudi Arabia. This study emphasizes the need for the relevant authorities to take immediate steps towards treating and limiting the spread of this disease throughout the country.

Knowledge, attitudes and practices on edible insects in Lango sub-region, northern Uganda

The study was done to determine the knowledge, attitude and practices towards edible insects, their consumption and practices of harvesting and processing in Lango sub-region, with the aim of increasing their consumption and improving nutrition of the people. Data was collected using semi structured questionnaires and focus group discussions. Common insect orders used as food in the area included: Isoptera (termites), Orthoptera (grasshoppers, crickets), and Hymenoptera (honey bee broods). Winged termites (Macrotermes spp.), locally known asNgwen was consumed by more than 97% of the respondents. Soldier termites (Syntermes soldiers), locally calledokok, were consumed by 73% of the respondents and crickets (Brachytrupes spp.), locally known asodir, was consumed by 69% of respondents. These insects were seasonally abundant and consumed in households in seasons of availability. Winged and soldier termites were harvested by attraction to light at the termite mound and by ‘termite fishing’ technique, respectively. Grasshoppers were harvested by hand picking from cropland bushes. Women and children were more actively involved in collection of insects than men. Pan frying, roasting, boiling followed by sun drying were the most common methods of processing insects. Dried insects were either served as snacks or a side dish. A main dish locally known asalakena is prepared from freshly ground termites. Dried winged termites were preserved for consumption at a later date. Women and children consumed more edible insects than men. The results show that insects are important as a food resource to the rural poor. However, majority of the respondents did not know that insects are nutritious. Therefore there is need to popularise consumption of edible insects in and outside the subregion through research on nutritional values of insects and documenting the information and processing insects using modern techniques in order to increase its commercial value and availability of insect products in all seasons.

Toxicity of thiametoxam on in vitro reared honey bee brood

Pesticides are a possible cause of pollinator decline and honey bee colony losses experienced in several countries in recent years. In the past years, many north-west Italian beekeepers reported the presence of dead brood in field apiaries during neonicotinoid-coated maize sowing; therefore, a possible role of these insecticides was suspected. The objective of this study was to test this hypothesis. Laboratory repeated dose toxicity tests on in vitro reared larvae were carried out using thiamethoxam. The repeated treatment median lethal concentration (LC50) and the median lethal dose (LD50) 14 and 19 days after grafting were calculated and resulted of the same order of magnitude of realistic brood exposure under a worst-case scenario. Various sublethal effects, like brownish larvae, duplication of the pupal integument, delay in development, and deformed adults were also observed.

Agricultural Landscape and Pesticide Effects on Honey Bee (Hymenoptera: Apidae) Biological Traits.

Sixteen honey bee (Apis mellifera L.) colonies were placed in four different agricultural landscapes to study the effects of agricultural landscape and exposure to pesticides on honey bee health. Colonies were located in three different agricultural areas with varying levels of agricultural intensity (AG areas) and one nonagricultural area (NAG area). Colonies were monitored for their performance and productivity for one year by measuring colony weight changes, brood production, and colony thermoregulation. Palynological and chemical analyses were conducted on the trapped pollen collected from each colony and location. Our results indicate that the landscape’s composition significantly affected honey bee colony performance and development. Colony weight and brood production were significantly greater in AG areas compared to the NAG area. Better colony thermoregulation in AG areas’ colonies was also observed. The quantities of pesticides measured in the trapped pollen were relatively low compared to their acute toxicity. Unexplained queen and colony losses were recorded in the AG areas, while colony losses because of starvation were observed in the NAG area. Our results indicate that landscape with high urban activity enhances honey bee brood production, with no significant effects on colony weight gain. Our study indicates that agricultural crops provide a valuable resource for honey bee colonies, but there is a trade-off with an increased risk of exposure to pesticides.

Effects of steel foundation wire on elemental content and hygienic removal of honey bee (Apis mellifera) brood

Honey and honey bees are often contaminated with trace metals, some of which are associated with food resources. However, in some cases metal contamination may be coming from within the hive. Based on preliminary observations, we hypothesized that steel foundation wire negatively affects honey bee brood health. To test our hypothesis we quantified the elemental content and measured the removal rates of honey bee brood developing in cells overlapping foundation wires and in control cells adjacent to cells that overlapped foundation wires. We found evidence that brood positioned in cells overlapping the wires contained significantly more iron and was removed at a significantly higher rate compared to control brood. Our study identifies the harmfulness of the common modern beekeeping practice of placing steel wire in beeswax foundation, which if avoided may increase colony population size and improve the health of managed honey bee colonies.

Nutritional value of bee-collected pollens of hardy kiwi, Actinidia arguta (Actinidiaceae) and oak, Quercus sp. (Fagaceae)

Bee-collected pollens have gained attention as natural dietary supplement for human nutrition over the honeybee feed, and become important commercial hive products for beekeepers. Oak trees, Quercus sp. (Family: Fagaceae) and hardy kiwi, Actinidia arguta (Family: Actinidiaceae) are important pollen sources for honey bees. During collection and storage through the addition of nectar and salivary secretions the chemical composition of pollen is changed. We analyzed nutrient contents of bee collected oak and hardy kiwi pollens and compared with recommended requirement of essential amino acids for honey bee brood development. The results revealed that protein content was found 23.2% for oak and 26.5% for hardy kiwi pollen. All the determined essential amino acid contents of both oak and kiwi pollen satisfied the minimum individual amino acid requirement of honey bee. Threonine was present in marginal level in hardy kiwi pollen. Fat content of oak and hardy kiwi pollen were found 7 and 4.5% respectively. The higher value of polyunsaturated fatty acid/saturated fatty acid - stearic acid, lower AI and TI value, presence of essential fatty acid n-3 linolenic acid indicated these pollens as superior sources of fat. The most abundant mineral was found potassium in both the pollens followed by phosphorus. In contrast sodium content was found very less. In vitro digestibility of oak and kiwi pollen were found 69.5 and 37.8% suggesting the difficulty of nutrient uptake directly from pollen grains.

Synthesis of Enantiopure Alicyclic Ethers and Their Activity on the Chemosensory Organ of the Ectoparasite of Honey Bees, Varroa destructor.

The preparation of enantiopure conformationally restricted alicyclic ethers and their inhibitory activities on the chemosensory organ of the Varroa destructor, a parasite of honey bees, are reported in this article. We tested the effect of enantiopure ethers of cis-5-(2'-hydroxyethyl)cyclopent-2-en-1-ol on the Varroa chemosensory organ by electrophysiology, for their ability to inhibit the responses to two honey bee-produced odors that are important for the mite to locate its host: nurse bee head space odor and (E)-β-ocimene, a honey bee brood pheromone. Previous work with the racemic compounds showed that they suppress the mite's olfactory response to its bee host, which led to incorrect host choice. Based on a structure-activity relationship, we predicted that the two most active compounds-cis-1-butoxy-5-(2'-methoxyethyl)cyclopent-2-ene, cy{4,1}, and (cis-1-ethoxy-5-(2'ethoxyethyl)cyclopent-2-ene, cy{2,2}-could have opposite active enantiomers. Here we studied the enantiomers of both ethers, whose preparation involved enzymatic resolution of racemic diol cis-5-(2'-hydroxyethyl)cyclopent-2-en-1-ol using Lipase AK with vinyl acetate. The racemic diol was prepared from commercially available 2,5-norbornadiene. We observed that the responses of the chemosensory organ to honey bee head space volatiles were significantly decreased by both enantiomers of cy{4,1} and cy{2,2}, but that responses to (E)-β-ocimene were decreased significantly only by (+)-cy{4,1} (1R,5S) and (-)-cy{2,2} (1S,5R) and not by their respective enantiomers. The importance of this result is that the racemates could be used to inhibit olfactory detection of bee odors by mites, without a loss in activity relative to the more expensive enantiopure compounds.

Survey and molecular detection of Melissococcus plutonius, the causative agent of European Foulbrood in honeybees in Saudi Arabia.

A large-scale field survey was conducted to screen major Saudi Arabian beekeeping locations for infection by Melissococcus plutonius. M. plutonius is one of the major bacterial pathogens of honeybee broods and is the causative agent of European Foulbrood disease (EFB). Larvae from samples suspected of infection were collected from different apiaries and homogenized in phosphate buffered saline (PBS). Bacteria were isolated on MYPGP agar medium. Two bacterial isolates, ksuMP7 and ksuMP9 (16S rRNA GenBank accession numbers, KX417565 and KX417566, respectively), were subjected to molecular identification using M. plutonius -specific primers, a BLAST sequence analysis revealed that the two isolates were M. plutonius with more than 98% sequence identity. The molecular detection of M. plutonius from honeybee is the first recorded incidence of this pathogen in Saudi Arabia. This study emphasizes the need for official authorities to take immediate steps toward treating and limiting the spread of this disease throughout the country.

A descriptive sensory analysis of honeybee drone brood from Denmark and Norway

Honeybee (Apis mellifera) brood is enjoyed as food in many regions of the world. The Nordic region of Europe is not currently one of them, but it could be. The drone brood in particular constitute an untapped source of delicious, nutritious and potentially sustainable food. Currently, it is removed by many beekeepers as part of a strategy to lower the population of the destructive Varroa mite (Varroa destructor) in the hive, but most often it is disposed of. A descriptive sensory analysis of drone brood was conducted, using larvae and pupae from four hives, three in Denmark and one in Norway, from locations spanning a range of urban/rural and monocultural/polycultural surroundings. Data were collected with a panel of 10 subjects using fast sensory methods, and were analysed with analysis of variance-partial least squares regression. The first dimension accounted for 40% of the variance, and correlated closely with differences in developmental stage between larvae and pupae; the second dimension accounted for 16% of the variance, and most closely grouped samples from three of the locations apart from those of the fourth. Developmental stage was thus found to be the primary determinant of sensory profile, followed by variations in locality, possibly related to local forage. Further studies should investigate the different parameters of sensory profile in more detail to develop a more complex picture of drone brood’s taste and culinary potential – an important part of developing strategies to diversify our food sources and realise certain insects’ ecological, culinary, and economic potential for local food systems.

Biological Role of Paenilarvins, Iturin-Like Lipopeptide Secondary Metabolites Produced by the Honey Bee Pathogen Paenibacillus larvae.

The Gram-positive bacterium Paenibacillus larvae (P. larvae) is the causative agent of a deadly honey bee brood disease called American Foulbrood (AFB). AFB is a notifiable epizootic in most countries and, hence, P. larvae is of considerable relevance for veterinarians and apiculturists alike. Over the last decade, much progress has been made in the understanding of the (patho)biology of P. larvae. Recently, several non-ribosomally produced peptides (NRP) and peptide/polyketide (NRP/PK) hybrids produced by P. larvae were identified. Among these NRPs were iturin-like lipopeptides, the paenilarvins A-C. Iturins are known to exhibit strong anti-fungal activity; for some iturins, cytotoxic activity towards mammalian erythrocytes and human cancer cell lines are described. We here present our results on the analysis of the natural function of the paenilarvins during pathogenesis of P. larvae infections. We demonstrated production of paenilarvins in infected larvae. However, we could neither demonstrate cytotoxicity of paenilarvins towards cultured insect cells nor towards larvae in feeding assays. Accordingly, exposure bioassays performed with larvae infected by wild-type P. larvae and a knockout mutant of P. larvae lacking production of paenilarvins did not substantiate a role for the paenilarvins as virulence factor. Further experiments are necessary to analyze the relevance of the paenilarvins’ anti-fungal activity for P. larvae infections in the presence of fungal competitors in the larval midgut or cadaver.

Antibacterial Activity of Cordyceps militaris Waste Product Extract against Paenibacillus larvae

American foulbrood (AFB) is the most serious and dangerous bacterial disease of honeybee brood caused by the spore forming bacterium Paenibacillus larvae. The aim of the present work was to evaluate the antimicrobial effect of a waste product of Cordyceps militaris extract against P. larvae and its potential for the control of AFB. For the test, the waste product of C. militaris was extracted with methanol and partitioned. And then, the fractions were tested for antimicrobial activity against P. larvae by the radial diffusion assay. The result showed that the portions of the ethyl acetate, and water-saturated n-butanol presented inhibitory capacity against strains of P. larvae, whereas the portions of n-hexane and chloroform were presented no inhibitory capacity against P. larvae. One way ANOVA tests for radial diffusion assay data, indicated significant differences. The results demonstrated that waste product of C. militaris possess beneficial properties for honey bee health and this property could be attributed to the kind and percentage of the main components of waste product of C. militaris portions.