Pedro Negri

Effects of the organic acids produced by a lactic acid bacterium in Apis mellifera colony development, Nosema ceranae control and fumagillin efficiency.

The European honey bee Apis mellifera is known to be affected by many parasites and pathogens that have great impact over the insect development. Among parasites affecting bee health, Nosema ceranae is one of the main biotic factors affecting colony populations. As honey bee populations decline, interest in pathogenic and mutualistic relationships between bees and microorganisms has increased. The main goal of the current study was to assess the effect of the oral administration of the metabolites produced by Lactobacillus johnsonii CRL1647 (mainly organic acids) supplemented in syrup, on: (I) N. ceranae sporulation dynamics before and after fumagillin application, and (II) performance of A. mellifera colonies. Different experiments were conducted to evaluate the effects of these bacterial metabolites on bees: in vitro administration revealed no toxic effects against bees. Colonies fed with the lactic acids incremented their beehive population and also the amount of fat bodies per bee. Finally, the organic acids reduced the intensity of the pathogen after the second application of treatment as well as enhanced the fumagillin efficiency. This study provides important information for the development of new control substances against nosemosis.

Abscisic acid enhances the immune response in Apis mellifera and contributes to the colony fitness

The primary food of adult honey bees (Apis mellifera) is honey prepared by bees from nectar, provided by plants in order to stimulate the bee’s pollination service. Nectar consists of carbohydrates, amino acids and water, as well as other minor compounds whose proportion varies among plant species and whose biological implications in the honey bee physiology require intense research. Several environmental stressors are causing the decline of bee colonies, and thereby, we tried to connect the nutritional quality of bee’s diet with the strength of the bee’s immune system. The phytohormone abscisic acid (ABA) is present in nectar, honey and adult honey bees. It has been demonstrated that ABA stimulates innate immune defences in animal cells. However, the influence of ABA on A. mellifera’s health and fitness is unknown. Here, we show that honey bees fed with an ABA supplement in field experiments resulted in (i) the appearance of ABA in larvae and adult bees, (ii) enhanced haemocyte response to non-self recognition, (iii) improved wound healing and granulocyte and plasmatocyte activation and (iv) maximum adult bee population after the winter and increased pesticide tolerance. The results indicate that the naturally occurring compound ABA has a positive influence in honey bee immunity. ABA emerges as a potent booster of immune defence in A. mellifera and may be useful in addressing the colony losses threatening apiculture and pollination service worldwide.

Honeybee health in South America

Honeybees are essential components to modern agriculture and economy. However, a continuous increase in cases of colony losses and colony depopulation are being reported worldwide. This critical situation has put the fragile equilibrium between bees and plants on the edge. As a consequence, several scientists have begun to focus their lines of research on this issue. Most researchers agree that there is no single explanation for the observed colony losses. Instead, these losses result from a synergistic interaction between different stressors. South America is not the exception; several cases of colony losses and colony depopulation were reported by beekeepers throughout the continent, yet no accurate data has been published to date. Therefore, this article attempts to analyze the past and present situation of honeybee health in South America, specifically in Argentina, Chile, Uruguay, Brazil, and Venezuela. Furthermore, it is intended to serve as a comparison to future colony losses, as well as to provide guidance for future hypothesis-driven research on the causes of colony mortality. We evaluate the impact of the main parasites and pathogens affecting honeybee colonies and discuss the role of each with respect to reported honeybee losses. We also contemplate the main challenges that each nation must confront with regards to honeybee health.

Apis mellifera haemocytes in-vitro, What type of cells are they? Functional analysis before and after pupal metamorphosis

Summary The honey bee Apis mellifera is affected by many parasites posing a serious threat to our ecosystem, agriculture and apiculture. The study of A. meliifera immune response represents a great complement for developing integrated disease management strategies. This work presents updated data concerning the in vitro cellular immune response of A. meliifera fifth instar larvae (L5) and newly emerged workers (W). Haemocytes from A. mellifera L5 and W clearly displayed different appearance in-vitro. The blood cells from these two developmental stages yielded behavioural differences regarding attachment and spreading. However, haemocytes extracted from L5 and W exhibited strong similarities with respect to their general (multicellular) behavioural pattern. This multi-haemolytic process, triggered after glass contact, could result from an encapsulation-like response. At the same time, an experimental system for studying A. mellifera cellular immunity in-vitro upon recognition of glass surfaces and haemolytic encapsulation response is herein developed.

Apis mellifera hemocytes generate increased amounts of nitric oxide in response to wounding/encapsulation

Apis mellifera populations are being threatened by several pathogens and parasitosis. Several authors have proposed that honey bee colonies may suffer from a compromised immune system leading to colony loss. This is why the study of A. mellifera immune system has become a topic of pressing concern. Nitric oxide (NO) is a signaling and immune effector molecule that has been proposed as a key molecule in invertebrate immunity, and that plays a part in A. mellifera cellular defenses. This paper deals with NO participation in the response to wounding/encapsulation challenge in A. mellifera fifth instar (L5) larvae. Challenging A. mellifera L5 larvae with nylon implants enhanced NO production and spreading in granulocyte-like hemocytes and increased the number of this NO-producing hemocyte type. However, AmNOS expression levels were not influenced by the insult. These results reveal that NO participates in the wound healing/encapsulation response as a signal molecule, possibly by the activation of a constitutively expressed AmNOS in honey bees.

Nitric oxide participates at the first steps of Apis mellifera cellular immune activation in response to non-self recognition

The honey bee Apis mellifera (Hymenoptera) is being affected by many diseases. The elimination of organisms in the insect hemocoel requires hemocytes recognition and response to the invader. After recognizing a surface as “foreign,” hemocytes “spread.” Spreading on glass surfaces by insect hemocytes is used as a measure of immune activation. Nitric oxide (NO) is a signaling and immune effector molecule in response to microbial infection that has been proposed as a key molecule in invertebrate immunity. The participation of NO in the hemocytic response of A. mellifera upon recognition of non-self is herein analyzed. Glass-adherent hemocytes produce large amounts of NO. Treatment with NO donor sodium nitroprusside enhanced hemocyte spreading, while NO scavenger carboxyPTIO reduced hemocyte immune activation. These results are indicative of NO participation at the beginning of A. mellifera immune response to non-self.

Cellular immunity in Apis mellifera: studying hemocytes brings light about bees skills to confront threats

Honey bee colonies are threatened by different stress factors around the world. Considerable efforts are being devoted to understanding honey bee defences to confront different kinds of stress factors. Despite the importance of honey bee hemocytes in resisting disease, detailed information about their role in response to challenge is still scarce. This manuscript combines the results obtained in studies which focused on the cellular defences of Apis mellifera, aiming to show how the understanding of the cellular components of the immune system is central to developing new strategies to enhance bees’ fitness.

Abscisic acid enhances cold tolerance in honeybee larvae

The natural composition of nutrients present in food is a key factor determining the immune function and stress responses in the honeybee (Apis mellifera). We previously demonstrated that a supplement of abscisic acid (ABA), a natural component of nectar, pollen, and honey, increases honeybee colony survival overwinter. Here we further explored the role of ABA in in vitro-reared larvae exposed to low temperatures. Four-day-old larvae (L4) exposed to 25°C for 3 days showed lower survival rates and delayed development compared to individuals growing at a standard temperature (34°C). Cold-stressed larvae maintained higher levels of ABA for longer than do larvae reared at 34°C, suggesting a biological significance for ABA. Larvae fed with an ABA-supplemented diet completely prevent the low survival rate due to cold stress and accelerate adult emergence. ABA modulates the expression of genes involved in metabolic adjustments and stress responses: Hexamerin 70b, Insulin Receptor Substrate, Vitellogenin, and Heat Shock Proteins 70. AmLANCL2, the honeybee ABA receptor, is also regulated by cold stress and ABA. These results support a role for ABA increasing the tolerance of honeybee larvae to low temperatures through priming effects.

Assessing in Vitro Acaricidal Effect and Joint Action of a Binary Mixture Between Essential Oil Compounds (Thymol, Phellandrene, Eucalyptol, Cinnamaldehyde, Myrcene, Carvacrol) Over Ectoparasitic Mite Varroa Destructor (Acari: Varroidae)

Abstract Varroa destructor (Anderson & Trueman, 2000) causes the most important parasitosis of beekeeping in the world. For this reason, prevention is needed to avoid colony death. The most typical treatments involve synthetic acaricides. However, the use of these acaricides results in the emergence of resistant populations of mites to these products and in the appearances of drug residues in products of the hives. Compounds of essential oils have emerged as an alternative to traditional acaricides; however the toxicity produced by these mixtures is currently poorly explored. The aim of this work was to assess, by means of in vitro tests with adult bees, how acaricidal action and toxic interactions in a binary mixture of essential oil compounds (Thymol, Phellandrene, Eucalyptol, Cinnamaldehyde, Myrcene, and Carvacrol) affect V. destructor. Calculations of LC50 ’s of the individual compounds on A. mellifera and V. destructor made clear that the toxic effect of each compound is different for both species. Thymol and Phellandrene turned out to be lethal for mites at lower concentrations than for bees. The binary mixture of these two substances presented a different toxicity than one produced by each pure compound, as it was highly selective for mites in bioassays at 24 hours through complete exposure to both A. mellifera and V. destructor. These results make such formulations optimal substances to be considered as alternative controls for the parasitosis.

Dietary Supplementation of Honey Bee Larvae with Arginine and Abscisic Acid Enhances Nitric Oxide and Granulocyte Immune Responses after Trauma

Many biotic and abiotic stressors impact bees’ health, acting as immunosupressors and contribute to colony losses. Thus, the importance of studying the immune response of honey bees is central to develop new strategies aiming to enhance bees’ fitness to confront the threats affecting them. If a pathogen breaches the physical and chemical barriers, honey bees can protect themselves from infection with cellular and humoral immune responses which represent a second line of defense. Through a series of correlative studies we have previously reported that abscisic acid (ABA) and nitric oxide (NO) share roles in the same immune defenses of Apis mellifera (A. mellifera). Here we show results supporting that the supplementation of bee larvae’s diet reared in vitro with l-Arginine (precursor of NO) or ABA enhanced the immune activation of the granulocytes in response to wounding and lipopolysaccharide (LPS) injection.