Place: Milano, italy, Europe
Sustainable management of natural resources Sustainable management of natural resources
Total Budget: € 0,00 | Period: From January 2010 To December 2012


Honeybees play the crucial ecosystem service of pollination guaranteeing not only the production of many crops and wild plants, but also the maintenance of the biodiversity. Recently they are facing a severe decline due to the occurrence of several biotic and abiotic factors, among which the outbreak of diseases and the use of pesticides. The project proposed a microbial formulation, referred as MICRO4BEE (M4B), as a solution to improve the honeybee health status. M4B is a mix of bacteria with probiotic activity for honeybees: laboratory tests conducted in vitro, in vivo and by the use of field trials have shown that the administration of M4B is effective in preventing the bacterial disease American Foulbrood. Currently, M4B is a patent of the University of Milan (PCT/IB2012/053459) licensed to the spin-off Micro4yoU and tested on the experimental apiaries of the beekeeping company Mario Bianco (Caluso). The patent is following the PCT procedure in different countries around the world.


Department of Food, Environmental and Nutritional Sciences, University of Milan - Italy

Lead applicant

Micro4yoU srl - Italy

Initiative partner

Beekeeping Company Mario Bianco - Italy

Initiative partner

The use of probiotic symbionts is proposed for the development of a sustainable strategy aimed to improve the honeybee and hive's health status. The concept arises from the most famous one related to the human resource management. In this view, as in a company the staff is managed to optimize the efficiency of the company, in a natural organism the symbiotic microbial resource could be managed to obtain a positive effect on the functioning of the organism itself.

The proposed technology is within the ambit of the development of a strategy alternative to the traditional ones. It foresees the use of microorganisms with probiotic activities to improve the health status of honeybees and hives against diseases and stressors. Micro4Bee (M4B) represents an innovative product composed of a mix of symbiotic bacteria, naturally present in the gut of bees, harmless to humans and with probiotic activities towards honeybees. M4B functions with multiple mechanisms: 1) direct antagonism against the pathogen; 2) stimulation of the immune system; 3) restoration of the optimal conditions of the intestinal flora. By the use of in vitro and in vivo tests and field trials it has been evaluated the ability of M4B to inhibit the pathogen of the bee larval disease American Foulbrood (AFB) and to reduce the larval mortality when bee larvae are challenged with the pathogen.

Honeybees are one of the most important pollinators for crops worldwide. It has been estimated that one third of food production can be connected, directly or indirectly, to pollination and that many crops for food production are strictly dependent to the activity of animal pollinators.  Another important issue exerted by honeybees is the maintenance of the biodiversity, which has an inestimable value and importance. Thus, the recent decline of honeybees is a worldwide problem, with significant impact on the biodiversity, threatening also the economy of the beekeeping and agricultural sectors. Biotic factors, such as viruses, bacteria and parasites, together with abiotic factors, such as the use of pesticide, the lack of forage and the climate change (and the combination of both kinds of stressors), are impacting the honeybee life and productivity. It is essential that the public and private bodies collaborate in coordinated efforts to develop strategies to prevent further losses.

In vitro and in vivo experiments, performed in laboratory conditions, showed that i) the bacteria, isolated and selected among the bacterial community members of the honeybee, inhibited the growth of Paenibacillus larvae, the etiological agent of the American Foulbrood (AFB) disease; and that ii) there was a reduction of the larval mortality when bee larvae were challenged with the pathogen, after a previous administration with the bacterial symbionts. These experiments constituted the basis for a field application, which confirmed the observed reduction of the larval mortality when hives were treated with the bacterial symbionts and the collected larvae were then challenged with the pathogen, in laboratory conditions. Experiments performed to elucidate the mechanisms exerted in the protection mediated by symbionts involved i) the direct inhibition of the pathogen by antibiosis, ii) the elicitation of the host immune system and iii) the competitive exclusion.

Laboratory in vivo rearing tests Laboratory in vivo rearing tests: the picture shows a multiwell plate in which bee larvae are reared with the artificial diet

The application of this technology could bring significant benefits to the beekeeping sector, improving the health status of honeybees and hives. In order to give an informative picture of the characteristics and needs of future beneficiaries (beekeepers), M4U carried out a survey sampling 41 beekeepers. From the survey it emerged that about three quarters of beekeepers lost recently more than half of their hives. The major causes of death are Varroa (parasitic mites) and the bacterial diseases. For the bacterial diseases they denounced that there are no products on the market. In reply to the question related to the possibility to use in field treatments a microbiological formulation with probiotic properties on bees, about 70% gave a positive answer.
The developed technology could positively influenced also the crop production in the frame of the sustainable agriculture

UNIMI: Dr. L. Sansonno, PhD (microbiologist, technical responsible) was a former PhD student working on the microbial diversity associated to honeybees to develop a biocontrol strategy based on the use of symbionts; Dr. E. Crotti, PhD (microbiologist, research responsible) is a researcher at DeFENS, specialized in the study and the biotechnological applications of associations between bacteria and insects.
Micro4yoU Srl: Dr. A. Balloi, PhD (Managing Director and technology transfer responsible) is co-founder and CEO of the spin-off Micro4yoU. Her research focuses on the associations between bacteria and insects and technology transfer in biotechnology.
Azienda Apistica Mario Bianco: Dr. A. Bianco (Beekeeper and clinical trials responsible) is the heir of an old family of beekeepers, faithful to the principles of quality craftsmanship. His company is among the most prestigious Italian honey exporters in Japan.
In the project other people from academic entities have been involved.

Field clinical trials The picture shows Annalisa Balloi and Andrea Bianco applying MICRO4BEE on bee hives

The hardest part of the project was to obtain the permission by the Ministry of Health for field clinical trials.

The main strength of managing the symbiotic resource associated to honeybees is that it foresees to preserve honeybee health by exploiting natural mechanisms of protection. The proposed technology is an example of a sustainable approach towards the environment by enhancing the beehive and honeybee productivity, using a strategy alternative to the traditional ones, based on chemicals. The technology includes positive effects on the quality and quantity of crop production.

Ecological Sustainability: The product makes possible to control the spread of bee diseases by improving the honeybee health using probiotic symbionts that show a direct antagonistic activity against pathogens and the capacity to stimulate the host immune system.

Economic sustainability: To ensure the profitability of the product and, at the same time, providing our customers with a product compatible to their budget, the microbial formulate will have a commercial cost similar to other food additives commonly used in beekeeping.

Transferability and Reproducibility: The project had an "evolution": it has gone from tests carried out in vitro, in vivo and finally in field at the Mario Bianco’s beekeeping company. Repeated experiments have been planned in three different climatic zones: Alpine climate (Trentino), continental climate (Piedmont) and a Mediterranean climate (Sardinia).

1) Unconventional dissemination

  • “Che Tempo Che Fa” (23rd March 2014): http://www.rai.tv/dl/RaiTV/programmi/media/ContentItem-39cc35f3-f070-48c1-b179-877edafa9c9c.html
  • “SuperQuark” (21st September 2014): http://www.rai.tv/dl/RaiTV/programmi/media/ContentItem-e2ab4dad-5ec1-4b75-87a1-698f005c3381.html#p=0

In these occasions, Dr Balloi led the public debate toward scientific topics (the concept of probiosis not only related to human health and the application of bacterial based products in several human activities) till then discussed only in Academic contexts.
2) Dissemination in the Beekeepins sector
The results of the research has been presented in 2 numbers of Apitalia (a specialized magazine for beekeepers) and in the conference “I Giorni del Miele” – 5/10/2014 - Lazise (Verona)
3) Accademic dissemination
The results were presented in several congresses and international meetings.
The obtained results are going to be published in 'peer-reviewed' international scientific journals.