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<h3>Project Basis</h3>
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<div class="a"><p align="justify"> The foundation of the project was evolving, improving and solidifying thanks to the interactions we experienced in our human practices. There were fundamental changes such as the production chassis of the peptides and the application method of the product, as well as recommendations for the experimental plan and insights about the risk factors we should consider.
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<div class="a"><p align="justify"> Excretion of antimicrobial peptides in <i>Pichia pastoris for</i> <i>Paenibacillus larvae</i> and <i>Melissococcus plutonius</i> inhibition. The initial plan was to apply genetically modified yeasts engineered to excrete AMP’s and surfactin in a solution with hexane extract of the plant A. satureioides; surfactin has a synergistic effect with the hexane extract. This solution would be applied to the bee bread we knew larvae ingested in their diet.
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<div class="a"><p align="justify">Learned obstacles regarding the method:</p></div>
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<ol>
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<li>Presence of genetically modified yeats would complicate the exportation process of honey to the European Union increasing the risks of having the product rejected.</li>
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<li>The hexane extract of <i>A.satureioides</i> is oxidized by exposure to air; therefore, it may lose its efficiency.</li>
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<li>An overdose of yeasts would cause fermentation of the honey and an alteration in the microbiota of the bees which could be counterproductive since this would cause diseases such as Nosema and American foulbrood to arrive.</li>
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<div class="a"><p align="justify">Production of antimicrobial peptides in Escherichia coli for Paenibacillus larvae and Melissococcus plutonius inhibition. The second plan consisted in applying the purified antimicrobial peptides, after their production in E. coli BL21 (DE3), in the bee bread we knew larvae ingested in their diet.</div></p>
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<div class="a"><p align="justify">Learned obstacles regarding the method:</p></div>
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<ol>
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<li>Bee bread is mainly stored in the hive so there is little chance that the peptides reach the larva.</li>
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</ol>
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<div class="a"><p align="justify">The third plan consisted in applying the purified peptides sprinkled as a liquid solution or as a powder on the hives.</p></div>
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<div class="a"><p align="justify">Learned obstacles regarding the method:</p></div>
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<ol>
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<li>When the product is found scattered in the hive, it can be easily affected by external factors such as air or other insects.</li>
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</ol>
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<div class="a"><p align="justify">We were able to arrive at the <b>final application method</b> with the recommendation of <b>MVZ Ernesto Tanús</b> who considered that the most suitable way to apply our product would be through the liquid food beekeepers provide to the hives and that nurse bees ingest to feed the larvae subsequently.</p></div><br>
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<h4>Experimentation plan</h4>
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<div class="a"><p align="justify">We integrated three aspects discussed during our interviews within the <a href="https://2018.igem.org/Team:Tec-Chihuahua/Results#future">future  experimentation plan</a> of the project:</p></div>
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<li>Add an innocuous color dye to the nanospheres to make the track in the bee system and ensure that the product reaches the larvae. This topic was discussed during the meeting with <b>MVZ Ernesto Tanús Sánchez, MVZ Adolfo Arroyo,</b> and <b>Eng. José Luis González Miranda.</b></li>
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<li>Test the research on the pure Italian Apis mellifera ligustica, as it is the most widespread in the world. This information was considered thanks to the video call with FAI presidential staff member <b>Lorenzo Della Morte.</b> </li>
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<li>Extract the hypopharyngeal glands of the nurse bees after they consume the liquid food where nanospheres were added in order to determine if there is presence of AMP's. Consideration taken from the conversation with <b>Ph.D. David de Jong.</b></li>
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<h4>Important considerations for the project</h4>
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<div class="a"><p align="justify"><b>MVZ Ernesto Tanús</b> considered it was essential to take into consideration the factor of the microbiota of the bee, regarding the fact that there would be AMP's supplied that can alter the natural microbiota. Regarding this same subject, <b>Ph.D. David De Jong</b> considered that our product works like the antibiotics humans use; there is the negative part that the intestinal flora will be affected, but this is in search for a greater good, attacking pathogens. The experimentation is necessary in order to be able to know what would happen with the microbiota of the bee and what consequences this situation could bring.</p></div><br>
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<div class="a"><p align="justify">Another of the considerations of <b>Ph.D. David De Jong</b>, was the analysis on how when you give certain substance to an animal, there is a proclivity for it to produce less of this substance; risk addressed in the <a href="#1">bioethical essay</a> and mentioned in the <a href="#2">legal plan</a>. </p></div><br>
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<h3>Enviromental</h3>
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<div class="a"><p align="justify"> The importance of the environment for the subsistence of life on earth is unquestionable. The magnitude of its value is represented by the human right to a healthy environment, a prerogative that protects the environment as a fundamental legal right and expresses the relevance of this to have a dignified life. This human right constitutes a guarantee for the realization and validity of the rest, since human being are in an indissoluble relationship with its environment and nature<sup>1</sup>. For this reason, achieving sustainable development would ensure the quality of present and future life. In this sense, the dignity, autonomy, and sanctity of the person depend on the health of the planet, its resources, and species. </p></div><br>
 
<div class="a"><p align="justify"> The importance of the environment for the subsistence of life on earth is unquestionable. The magnitude of its value is represented by the human right to a healthy environment, a prerogative that protects the environment as a fundamental legal right and expresses the relevance of this to have a dignified life. This human right constitutes a guarantee for the realization and validity of the rest, since human being are in an indissoluble relationship with its environment and nature<sup>1</sup>. For this reason, achieving sustainable development would ensure the quality of present and future life. In this sense, the dignity, autonomy, and sanctity of the person depend on the health of the planet, its resources, and species. </p></div><br>
 
<div class="a"><p align="justify"> Food Security is a central topic in the global agenda. The Food and Agriculture Organization (FAO) estimates that by the year 2050, world population will exceed 9,000 million, of which 150 million will dwell in Mexico. This should increase agricultural production by 60% in the country by that year <sup>2</sup>. </p></div><br>
 
<div class="a"><p align="justify"> Food Security is a central topic in the global agenda. The Food and Agriculture Organization (FAO) estimates that by the year 2050, world population will exceed 9,000 million, of which 150 million will dwell in Mexico. This should increase agricultural production by 60% in the country by that year <sup>2</sup>. </p></div><br>
 
<div class="a"><p align="justify"> Beekeeping is a fundamental activity to achieve Food Security in Mexico and the world, which is why authorities, researchers, farmers, beekeepers, and members are aware of the importance of bees. The Food and Agriculture Organization (FAO) says 100 crop species provide 90% of the food worldwide, and bees pollinate 71% of these<sup>3</sup>. It is for this reason, given by the engineer <b>Manuel Ramirez Salcido</b>, that bees have been presented as a parameter of environmental health, talking about ecology.</p></div><br>
 
<div class="a"><p align="justify"> Beekeeping is a fundamental activity to achieve Food Security in Mexico and the world, which is why authorities, researchers, farmers, beekeepers, and members are aware of the importance of bees. The Food and Agriculture Organization (FAO) says 100 crop species provide 90% of the food worldwide, and bees pollinate 71% of these<sup>3</sup>. It is for this reason, given by the engineer <b>Manuel Ramirez Salcido</b>, that bees have been presented as a parameter of environmental health, talking about ecology.</p></div><br>
 
<div class="a"><p align="justify">The Ministry of Agriculture, Livestock, Rural Development, Fisheries and Food (SAGARPA) says that if bees disappear there would be a food crisis; 84% of cash crops depend on their pollination <sup>4</sup>. In the same way, SAGARPA states that multiple varieties of fruit trees depend on cross-pollination that uses animals as vectors, such as bees; that is to say that there is an enormous amount of zoophilic plants.</p></div><br>
 
<div class="a"><p align="justify">The Ministry of Agriculture, Livestock, Rural Development, Fisheries and Food (SAGARPA) says that if bees disappear there would be a food crisis; 84% of cash crops depend on their pollination <sup>4</sup>. In the same way, SAGARPA states that multiple varieties of fruit trees depend on cross-pollination that uses animals as vectors, such as bees; that is to say that there is an enormous amount of zoophilic plants.</p></div><br>
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<div class="a"><p align="justify"> According to the Ministry of Environment and Natural Resources (SEMARNAT), the links between the population, environment, and natural resources are mediated by multiple factors. The immediate environmental impact of urban settlements derives from a change in land purpose<sup>5</sup>, reducing the size of crops that can be pollinated by bees, generating an effect similar to that of habitat loss and with this, nutritional complications in bees, conferring susceptibility to diseases. This is one of the points suggested by the engineer Alejandro Bianchi during our meeting on June 15, 2018. </p></div><br>
 
<div class="a"><p align="justify"> According to the Ministry of Environment and Natural Resources (SEMARNAT), the links between the population, environment, and natural resources are mediated by multiple factors. The immediate environmental impact of urban settlements derives from a change in land purpose<sup>5</sup>, reducing the size of crops that can be pollinated by bees, generating an effect similar to that of habitat loss and with this, nutritional complications in bees, conferring susceptibility to diseases. This is one of the points suggested by the engineer Alejandro Bianchi during our meeting on June 15, 2018. </p></div><br>
 
<div class="a"><p align="justify"> However, there are actions that seek to provide a healthy life for bees and their existence. There is a concern for the negative impact on anthophiles by irresponsible human activities; it is vital to prevent and attack the causes of bee reduction or loss at their roots. As we learned thanks to <b>MEXMUN 2018</b>, the objectives of the Convention on Biological Diversity should be used. These objectives are: </p></div>
 
<div class="a"><p align="justify"> However, there are actions that seek to provide a healthy life for bees and their existence. There is a concern for the negative impact on anthophiles by irresponsible human activities; it is vital to prevent and attack the causes of bee reduction or loss at their roots. As we learned thanks to <b>MEXMUN 2018</b>, the objectives of the Convention on Biological Diversity should be used. These objectives are: </p></div>
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<div class="a"><p align="justify">Currently, the importance of carrying out a risk analysis is integrated in our legal plan, as per the <b>Enviromental Risk Assesment</b> Guide. In addition to the aspects mentioned in the assessment of the quality control within the legal plan, the following steps are involved, such as the guidelines for animal health risk analysis according to the Regulation of the Federal Animal Health Law, which are the following: </p></div>
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<div class="a"><p align="justify">Currently, the importance of carrying out a risk analysis is integrated in our legal plan, as per the <b>Enviromental Risk Assesment Guide</b>. In addition to the aspects mentioned in the assessment of the quality control within the legal plan, the following steps are involved, such as the guidelines for animal health risk analysis according to the Regulation of the Federal Animal Health Law, which are the following: </p></div>
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<ol>
 
<ol>
<li> Identification of the possible risks.
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<li>Identification of the possible risks.
 
</li>
 
</li>
<li> Assessment of the chances that these risks arise.
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<li>Assessment of the chances that these risks arise.
 
</li>
 
</li>
<li> Evaluation of the consequences.
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<li>Evaluation of the consequences.
 
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<li>Estimation of the possible global risk.
 
<li>Estimation of the possible global risk.
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   <ol>
 
   <ol>
       <li>Elzen, P. J., Westervelt, D., Causey, D., Ellis, J., Hepburn, H. R., & Neumann, P. (2002). <i>Method of Application of Tylosin, an Antibiotic for American Foulbrood Control, with Effects on Small Hive Beetle</i> (Coleoptera: Nitidulidae) Populations. Journal of Economic Entomology, 95(6), 1119–1122. doi:10.1603/0022-0493-95.6.1119</li>
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       <li>National Human Rights Commission (Mexico). (2014). The Human Right to a Healthy Environment for Development and Wellbeing. Mexico: National Human Rights Commission (Mexico).</li>
      <li>Grout, R. (1949). <i>The Hive and the Honey Bee</i>. Hamilton, IL: Dandant & Sons. p.504-509.</li>
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       <li>Crop Protection, Science and Technology. (2018). Beekeeping; as a fundamental activity to reach food security in Mexico. October 10th, de Crop Protection, Science and Technology Web site: http://proccyt.org.mx/noticias/303-la-apicultura-actividad-fundamental-para-alcanzar-la-seguridad-alimentaria-en-mexico-2</li>
       <li>Makadia, H. K., & Siegel, S. J. (2011). <i>Poly Lactic-co-Glycolic Acid (PLGA) as Biodegradable Controlled Drug Delivery Carrier</i>. Polymers, 3(3), 1377–1397.https://doi.org/10.3390/polym3031377</li>
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       <li>Food and Agriculture Organization of the United Nations. (2016). The power of pollinators: why more bees means better food. October 10th, de Food and Agriculture Organization of the United Nations Sitio web: http://www.fao.org/zhc/detail-events/en/c/428504/ </li>
       <li>Budhian, A., Siegel, S. J., & Winey, K. I. (2008). <i>Controlling the in vitro release profiles for a system of haloperidol-loaded PLGA nanoparticles</i>. International Journal of Pharmaceutics, 346(1–2), 151–159. https://doi.org/10.1016/j.ijpharm.2007.06.011</li>
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       <li>Delegación SAGARPA Yucatán. (2017). Apicultura, actividad de gran importancia económica y social en Yucatán [Beekeeping, activity of great economic and social importance in Yucatan]. October 10th, de SAGARPA Web site: https://www.gob.mx/sagarpa%7Cyucatan/articulos/apicultura-actividad-de-gran-importancia-económica-y-social-en-yucatan</li>
      <li>Siepmann, J., Elkharraz, K., Siepmann, F., & Klose, D. (2005). <i>How Autocatalysis Accelerates Drug Release from PLGA-Based Microparticles:  A Quantitative Treatment</i>. Biomacromolecules, 6(4), 2312–2319. doi:10.1021/bm050228k</li>
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       <li>SEMARNAT. (2010). Compendio de Estadísticas Ambientales 2010. October 10th, de SEMARNAT Sitio web: http://aplicaciones.semarnat.gob.mx/estadisticas/compendio2010/10.100.13.5_8080/ibi_apps/WFServlet1bdcb.html </li>
       <li>Ford Versypt, A. N., Pack, D. W., & Braatz, R. D. (2013). <i>Mathematical modeling of drug delivery from autocatalytically degradable PLGA microspheres — A review</i>. Journal of Controlled Release, 165(1), 29–37. https://doi.org/10.1016/j.jconrel.2012.10.015</li>
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<br>
      <li>Rojas-López, C. A., Padilla-Salas, V. R., López-Cortés, G. I., Ordaz-Briseño, R., & Islas-Rodríguez, A. E. (2017). <i>Sistema inmune innato de las abejas</i>. Revista De Ciencias Ambientales Y Recursos Naturales, 3(7), 25-32.</li>
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  </ol>
      <li>Nation, J. L., Sr. (2015). <i>15.4.5 Hemolymph pH and hemplymph buffers</i>. In Insect Physiology and Biochemistry (3rd ed., pp. 424-425). CRC Press</li>
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</div>
      <li>Chen, C. (1995). <i>Changes in protein components and storage stability of Royal Jelly under various conditions</i>. Food Chemistry, 54(2), 195–200. https://doi.org/10.1016/0308-8146(95)00031-d</li>
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       <li>Sanhueza-Rojas, O.H. (2016). <i>Caracterización química multifactorial de miel en relación a la infección por nosema ceranae en abejas, actividad antimicrobiana y origen geográfico del producto</i>. Thesis. Universidad de Chile.</li>
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      <li>Tautz J (2008) <i>The Buzz about Bees</i>. Springer-Verlag, Berlin Heidelberg.</li>
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      <li>Heinrich, B. (1979). <i>Bumblebee economics</i>. Cambridge: Harvard University Press</li>
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      <li>Genersch, E. (2010).<i> American Foulbrood in honeybees and its causative agent, Paenibacillus larvae</i>. Journal of Invertebrate Pathology, 103, S10–S19. doi:10.1016/j.jip.2009.06.015</li>
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      <li>Herbert, E. W., Jr., & Shimanuki, H. (1983). <i>Effect of diet pH on the consumption, brood rearing, and pH of worker jelly produced by caged honey bees</i>. Apidologie, 14(3), 191-196. Retrieved from https://hal.archives-ouvertes.fr/hal-00890597.</li>
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<li>Khilnani, J. C. (2015). The Effects of Honeybee (Apis mellifera) Antimicrobial Peptides on Paenibacillus larvae. UNLV Theses, Dissertations, Professional Papers and Capstones, 2486. Retrieved from https://digitalscholarship.unlv.edu/thesesdissertations/2486/.</li><br>
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<h3>Economic</h3>
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<div class="a"><p align="justify">All modern human endeavors are, one way or another, economic activities. Beekeeping is no different. This income-generating activity plays a crucial role in the food industry, thanks to both pollination services and honey production.
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<div class="a"><p align="justify">Classic economics tells us that it is time put into a product or service which gives it its value. Beekeepers know about this; they work all the time, with a lot of hives. They must take care of each colony, all while creating smoke to keep the bees passive. At the <b>IV State Beekeeping Forum</b>, emphasis was made on what a laborious activity this job is.</p></div><br>
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<center><img src="https://static.igem.org/mediawiki/2018/2/27/T--Tec-Chihuahua--EcoWagon.png"></center>
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<div class="a"><p align="justify">UN risk assessment researcher, <b>Maria Mercedes Roca, Ph.D. </b>, mentioned how economic activities are central to the functioning of our interconnected world. Beekeeping must be propelled to make for economic gains, especially in countries such as Mexico where a large part of its economy is based on the primary sector. <b>Mr. Manuel Ramirez</b> also referred to this topic; Our country plays a significant role as a honey trader, with most of our exports making their way to the EU. However, there are several obstacles. EU imports can have neither genetically modified content nor most antibiotics. One can’t add anything to a product that would alter its sensorial characteristics without expecting a negative impact. <b>Dr. David de Jong</b> considered we should check what amount of AMPs make their way to honey products, aspect revised in the <b>IV State Beekeeping Forum</b> with <b>Dr. Sóstenes Rafael Rodríguez</b> where we ensured AMP’s are not a problem when it comes to EU honey exportation analysis. Since the national Secretariat of Agriculture, Livestock, Rural Development, Fisheries and Food (<b>SAGARPA</b>) is also strict on contaminants, <b>Guillermo Rodriguéz</b> told us, a natural product against foulbrood is needed both nationally and internationally.</p></div><br>
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<center><img src="https://static.igem.org/mediawiki/2018/6/6b/T--Tec-Chihuahua--EcoHoney.png"></center>
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<div class="a"><p align="justify">As explained in the chronology of the aforementioned facts, while the project was being developed and basing us on the comments and concerns expressed by professionals, we were able to ensure that our product would not impede exportations and importations of honey. The world honey market reported historical records during 2015, with volumes of operation exceeding 2.3 billion dollars<sup>1</sup> and it represents substantial revenues not only for Mexico but countries like China, Argentina, Spain, etc. More than ensuring that highest AMP concentrations in honey would not result in an exportation problem, our project fosters to make exportations go up since there will be no problem with antibiotic residues in honey when beekeepers have to deal with diseases.</p></div><br>
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<div class="a"><p align="justify">The economic part of our project is also made up of the development of the <a href="https://2018.igem.org/Team:Tec-Chihuahua/Entrepreneurship">entrepreneurial area</a> where the implications for the sale of the final product are analyzed.</p></div><br>
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<div class="a"><p align="justify">Our project will make sure Foulbrood no longer generates huge economic losses (by killing bees and necessitating antibiotics) but instead become a small nuisance in an otherwise thriving industry.</p></div><br>
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<button class="collapsible3"><h4 align="center">References</h4></button>
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      <li>PROCCyT. (2018). La apicultura; actividad fundamental para alcanzar la seguridad alimentaria en México. Retrieved from http://proccyt.org.mx/noticias/303-la-apicultura-actividad-fundamental-para-alcanzar-la-seguridad-alimentaria-en-mexico-2</li>
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<h3>Social</h3>
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<div class="a"><p align="justify">While we interacted with beekeepers and researchers, it was not difficult to realize the importance of considering social aspects within the activities and plans for our project.</p></div><br>
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<div class="a"><p align="justify">Rider University explains “sociological imagination” as the ability to see the connection between the larger events affecting the whole society and the individual consequences for each of us in our own lives. Personal troubles may be our fault; but in others, they may be a result of social forces beyond our control as individuals; society shape us as we shape our society.<sup>1</sup></p></div><br>
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<div class="a"><p align="center"><i>Analyzing and acting upon social implications lies in living together in an organized and beneficial way.</i></p></div><br>
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<center><img src="https://static.igem.org/mediawiki/2018/8/8f/T--Tec-Chihuahua--SocWorld.png"></center>
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<div class="a"><p align="justify">In the first approach we had with a beekeeper, <b>Mr. Guillermo Rodríguez</b>, it was explained how beekeeping varies around the world and how many times the needs are different for each region. In our effort to consider all these needs in our project, we managed to have interviews with professionals related to the topic in Brazil, Chile, Mexico, Italy, Valencia, and Canadá, taking into account from beekeepers who manage few hives to complete associations, investigators, governmental institutions and international agencies. We considered specific needs to obtain a universal solution.</p></div><br>
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<div class="a"><p align="justify">In the <b>IV State’s Beekeeping Forum</b> and the <b>CIAD</b> visit, we were able to experience what beekeepers’ work involves. Working with few beehives lessen the job, however in the case of beekeepers like the Chilean Bryaan Saaverdra, that works with more than 1000 hives the work notably intensifies, as you must attend beehive by beehive whatever the weather condition is. Through the application method of our product, which was transformed over the months basing on the concerns and needs of the beekeepers, we foster the welfare of this community taking in consideration for their working days not to become intensified when experiencing the diseases and applying the product. </p></div><br>
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<div class="a"><p align="justify">In the meeting with <b>Mr. Manuel Ramírez Salcido</b>, he commented on how there are almost no young people who are interested in beekeeping and that there is a significant concern for the future of this sector. While developing our project, we saw the enthusiasm of beekeepers for seeing young people interested in the area. Throughout all these months we have shared our project in many places, we have tried to bring the youth to be interested in bee’s importance and beekeeping. For the <b>IV State’s Beekeeping Forum</b> we made an open invitation to different year students of our biotechnology career, encouraging them to attend, to listen to the conferences and learn about apiaries. By proudly conducting this project we are directly involving with beekeeping and fostering an important approach to this sector in the young people that surround us in our daily lives as college students.</p></div><br>
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<center><img src="https://static.igem.org/mediawiki/2018/e/ed/T--Tec-Chihuahua--SocStats.png"></center>
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<div class="a"><p align="justify">After all the research made, the ambiguities that were found when doing the evaluation of the presence of Foulbrood in the world and the different conclusions that were given, (some resulting from the video call with <b>FAI presidential staff member Lorenzo Della Morte</b> and others from the video call with <b>MVZ Juan Diego Pérez de la Rosa</b>) showed that there is a problem in the health and statistical assessment studies regarding bee diseases. Many times, beekeepers who have affected hives, do not report it because of the negative implications that this entails; burning of colonies, the need of using antibiotics that disturb the innocuity of honey and finally the risk of losing investment made over many years. Our product promotes this social problem to be resolved; by having a profitable and useful product, which displace conventional antibiotics usage and the burning of hives. It encourages that the beekeeper openly reports if their beehives suffer from the disease having the peace of mind that he will not have to lose large quantities of money or their hives nor he will need to alter their bee products with antibiotics. </p></div><br>
 +
<div class="a"><p align="justify">By addressing individual concerns as well as general insights such as the problem of statistical assessment studies, throughout the development of our project and the implementation of the product, we seek to address aspects that are often out of the hands of beekeepers, and that may also have an impact on communities outside of the beekeeping sector. Attention to the previously mentioned issues is based on the fact of building an organized society that interacts effectively within itself and with external communities.</p></div><br>
 +
<div class="a"><p align="justify">The social area covers the ethical and legal aspects:</p></div><br>
 +
 +
<p id="1"></p>
 +
<center><img src="https://static.igem.org/mediawiki/2018/b/ba/T--Tec-Chihuahua--Ethics.png"></center>
 +
<h4 align="center">Ethical</h4>
 +
 +
<br>
 +
<div class="a"><p align="justify">Three of our human practices activities stood out in determining points that led us to reflect and to investigate more in the situation. Is saving bees our duty? Is our project good? Is it worth taking risks involved? Bioethical dilemmas that will be addressed below.</p></div><br>
 +
 +
<div class="a"><p align="justify">From the meeting we had with <b>Dr. Roca</b>, we discussed the importance of not only considering human rights but also environmental rights. The next question arose: Should humanity intervene and conserve biodiversity by any means?</p></div><br>
 +
 +
<div class="a"><p align="justify">The participation in <b>MEXMUN</b> helped us to realize the importance of being responsible for the risks involved in the creation of any biotechnological tool; from there these questions arose: Should we develop the treatment even though it could result in the development of pathogen resistance? Or, in the worst case, damage the innate immune system leaving bees unable to defend themselves?</p></div><br>
 +
 +
<div class="a"><p align="justify">Based on comments from <b>Ph.D. David de Jong, Dr. Ernesto Guzmán Novoa, M.S. Alejandro Romo Chacón and M.S. Adrián Ponce de León</b>. We realized that there are no regulations to experiment with bees in Canada, Brazil, and Mexico. This led us to question whether the use of bee larvae is justified in order to obtain relevant data for the project to advance. Can even be considered morally relevant?
 +
</p></div><br>
 +
<center><img src="https://static.igem.org/mediawiki/2018/2/24/T--Tec-Chihuahua--EthicsPoints.png"></center>
 +
<br>
 +
<object data="https://static.igem.org/mediawiki/2018/0/08/T--Tec-Chihuahua--Bioethics.pdf"  type="application/pdf" width="100%" height="700px" align="middle"> </object>
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<br>
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<p id="2"></p>
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<center><img src="https://static.igem.org/mediawiki/2018/1/19/T--Tec-Chihuahua--Legal.png"></center>
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<h4 align="center">Legal</h4>
 +
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<br>
 +
<div class="a"><p align="justify">From the interviews we had with <b>Mr. Guillermo Rodríguez</b> and <b>Mr. Manuel Ramírez Salcido</b> we realized the importance of considering both national and international governmental agencies involved in the assurance of honey innocuousness. Nevertheless, the legal validation of the development of our project took a big step after the meeting we had with <b>MVZ Ernesto Tanús Sánchez, MVZ Adolfo Arroyo, and Mr. José Luis González Miranda</b>. In this meeting, different legislation that would be ideal to consider was mentioned; concerning innocuity, exportation, Federal Animal Health Law, Zoosanitary regulations as well as Regulations of the European Union regarding food safety.</p></div><br>
 +
 +
<div class="a"><p align="justify">In addition, our participation in activities of international nature, being, in this case, <b>MEXMUN</b> and the <b>Environmental Risk Assessment Guide for Genetically Modified Organisms</b>, led us to evaluate aspects of biosecurity and risk analysis. We considered it appropriate and necessary to build a legal plan in accordance with the regulations and norms pertinent to our project, covering the local and international scope for ensuring that our project is attached to safety and legality.</p></div><br>
 +
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<object data="https://static.igem.org/mediawiki/2018/0/08/T--Tec-Chihuahua--Bioethics.pdf"  type="application/pdf" width="100%" height="700px" align="middle"> </object>
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      <li>Raider University. (n.d.). <i>Why Study Sociology?</i> Retrieved from https://www.rider.edu/academics/colleges-schools/college-liberal-arts-education-sciences/liberal-arts-programs/sociology/why-study-sociology</li>
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Latest revision as of 16:50, 17 October 2018

Erwinions


Project Basis

The foundation of the project was evolving, improving and solidifying thanks to the interactions we experienced in our human practices. There were fundamental changes such as the production chassis of the peptides and the application method of the product, as well as recommendations for the experimental plan and insights about the risk factors we should consider.


Excretion of antimicrobial peptides in Pichia pastoris for Paenibacillus larvae and Melissococcus plutonius inhibition. The initial plan was to apply genetically modified yeasts engineered to excrete AMP’s and surfactin in a solution with hexane extract of the plant A. satureioides; surfactin has a synergistic effect with the hexane extract. This solution would be applied to the bee bread we knew larvae ingested in their diet.


Learned obstacles regarding the method:

  1. Presence of genetically modified yeats would complicate the exportation process of honey to the European Union increasing the risks of having the product rejected.
  2. The hexane extract of A.satureioides is oxidized by exposure to air; therefore, it may lose its efficiency.
  3. An overdose of yeasts would cause fermentation of the honey and an alteration in the microbiota of the bees which could be counterproductive since this would cause diseases such as Nosema and American foulbrood to arrive.

Production of antimicrobial peptides in Escherichia coli for Paenibacillus larvae and Melissococcus plutonius inhibition. The second plan consisted in applying the purified antimicrobial peptides, after their production in E. coli BL21 (DE3), in the bee bread we knew larvae ingested in their diet.

Learned obstacles regarding the method:

  1. Bee bread is mainly stored in the hive so there is little chance that the peptides reach the larva.

The third plan consisted in applying the purified peptides sprinkled as a liquid solution or as a powder on the hives.

Learned obstacles regarding the method:

  1. When the product is found scattered in the hive, it can be easily affected by external factors such as air or other insects.

We were able to arrive at the final application method with the recommendation of MVZ Ernesto Tanús who considered that the most suitable way to apply our product would be through the liquid food beekeepers provide to the hives and that nurse bees ingest to feed the larvae subsequently.


Experimentation plan

We integrated three aspects discussed during our interviews within the future experimentation plan of the project:

  1. Add an innocuous color dye to the nanospheres to make the track in the bee system and ensure that the product reaches the larvae. This topic was discussed during the meeting with MVZ Ernesto Tanús Sánchez, MVZ Adolfo Arroyo, and Eng. José Luis González Miranda.
  2. Test the research on the pure Italian Apis mellifera ligustica, as it is the most widespread in the world. This information was considered thanks to the video call with FAI presidential staff member Lorenzo Della Morte.
  3. Extract the hypopharyngeal glands of the nurse bees after they consume the liquid food where nanospheres were added in order to determine if there is presence of AMP's. Consideration taken from the conversation with Ph.D. David de Jong.

Important considerations for the project

MVZ Ernesto Tanús considered it was essential to take into consideration the factor of the microbiota of the bee, regarding the fact that there would be AMP's supplied that can alter the natural microbiota. Regarding this same subject, Ph.D. David De Jong considered that our product works like the antibiotics humans use; there is the negative part that the intestinal flora will be affected, but this is in search for a greater good, attacking pathogens. The experimentation is necessary in order to be able to know what would happen with the microbiota of the bee and what consequences this situation could bring.


Another of the considerations of Ph.D. David De Jong, was the analysis on how when you give certain substance to an animal, there is a proclivity for it to produce less of this substance; risk addressed in the bioethical essay and mentioned in the legal plan.


Enviromental


The importance of the environment for the subsistence of life on earth is unquestionable. The magnitude of its value is represented by the human right to a healthy environment, a prerogative that protects the environment as a fundamental legal right and expresses the relevance of this to have a dignified life. This human right constitutes a guarantee for the realization and validity of the rest, since human being are in an indissoluble relationship with its environment and nature1. For this reason, achieving sustainable development would ensure the quality of present and future life. In this sense, the dignity, autonomy, and sanctity of the person depend on the health of the planet, its resources, and species.


Food Security is a central topic in the global agenda. The Food and Agriculture Organization (FAO) estimates that by the year 2050, world population will exceed 9,000 million, of which 150 million will dwell in Mexico. This should increase agricultural production by 60% in the country by that year 2.


Beekeeping is a fundamental activity to achieve Food Security in Mexico and the world, which is why authorities, researchers, farmers, beekeepers, and members are aware of the importance of bees. The Food and Agriculture Organization (FAO) says 100 crop species provide 90% of the food worldwide, and bees pollinate 71% of these3. It is for this reason, given by the engineer Manuel Ramirez Salcido, that bees have been presented as a parameter of environmental health, talking about ecology.


The Ministry of Agriculture, Livestock, Rural Development, Fisheries and Food (SAGARPA) says that if bees disappear there would be a food crisis; 84% of cash crops depend on their pollination 4. In the same way, SAGARPA states that multiple varieties of fruit trees depend on cross-pollination that uses animals as vectors, such as bees; that is to say that there is an enormous amount of zoophilic plants.


According to the Ministry of Environment and Natural Resources (SEMARNAT), the links between the population, environment, and natural resources are mediated by multiple factors. The immediate environmental impact of urban settlements derives from a change in land purpose5, reducing the size of crops that can be pollinated by bees, generating an effect similar to that of habitat loss and with this, nutritional complications in bees, conferring susceptibility to diseases. This is one of the points suggested by the engineer Alejandro Bianchi during our meeting on June 15, 2018.


However, there are actions that seek to provide a healthy life for bees and their existence. There is a concern for the negative impact on anthophiles by irresponsible human activities; it is vital to prevent and attack the causes of bee reduction or loss at their roots. As we learned thanks to MEXMUN 2018, the objectives of the Convention on Biological Diversity should be used. These objectives are:

  1. Conservation of biodiversity.
  2. Sustainable use of the components of biological diversity.
  3. Appropriate transference of technologies.

Currently, the importance of carrying out a risk analysis is integrated in our legal plan, as per the Enviromental Risk Assesment Guide. In addition to the aspects mentioned in the assessment of the quality control within the legal plan, the following steps are involved, such as the guidelines for animal health risk analysis according to the Regulation of the Federal Animal Health Law, which are the following:

  1. Identification of the possible risks.
  2. Assessment of the chances that these risks arise.
  3. Evaluation of the consequences.
  4. Estimation of the possible global risk.
  5. The resolution of whether the risks are acceptable or not, as a whole, as well as the strategies for the management of possible risks.

Dr. Ernesto Tanus Sánchez mentioned the importance of avoiding the burning of hives, due to the limited accessibility to the wood with which they are made. Cedar, specifically red cedar (Cedrela odorata), is considered the best type of wood to make a hive. However, this species is subject to special protection according to NOM-059-SEMARNAT-2010, which means factors that negatively affect its viability may threaten it. Obtaining permission to perform the practice of incinerating hives becomes an arduous and even deficient process. Due to the virulence of Paenibacillus larvae, American Foulbrood sweeps entire hives before there is an answer by the authorities.


In other Latin American countries, the action plan presents variations. Ph.D. David De Jong explained us how in Brazil, it is not permissible for someone to incinerate a beehive if it is not a beekeeper, sometimes the firefighters do it in an emergency. Bryan Saavedra, mentioned how in Chile, they have the Agricultural and Livestock Service (SAG) who is in charge of incinerating hives in case the disease is present.



  1. National Human Rights Commission (Mexico). (2014). The Human Right to a Healthy Environment for Development and Wellbeing. Mexico: National Human Rights Commission (Mexico).
  2. Crop Protection, Science and Technology. (2018). Beekeeping; as a fundamental activity to reach food security in Mexico. October 10th, de Crop Protection, Science and Technology Web site: http://proccyt.org.mx/noticias/303-la-apicultura-actividad-fundamental-para-alcanzar-la-seguridad-alimentaria-en-mexico-2
  3. Food and Agriculture Organization of the United Nations. (2016). The power of pollinators: why more bees means better food. October 10th, de Food and Agriculture Organization of the United Nations Sitio web: http://www.fao.org/zhc/detail-events/en/c/428504/
  4. Delegación SAGARPA Yucatán. (2017). Apicultura, actividad de gran importancia económica y social en Yucatán [Beekeeping, activity of great economic and social importance in Yucatan]. October 10th, de SAGARPA Web site: https://www.gob.mx/sagarpa%7Cyucatan/articulos/apicultura-actividad-de-gran-importancia-económica-y-social-en-yucatan
  5. SEMARNAT. (2010). Compendio de Estadísticas Ambientales 2010. October 10th, de SEMARNAT Sitio web: http://aplicaciones.semarnat.gob.mx/estadisticas/compendio2010/10.100.13.5_8080/ibi_apps/WFServlet1bdcb.html



Economic


All modern human endeavors are, one way or another, economic activities. Beekeeping is no different. This income-generating activity plays a crucial role in the food industry, thanks to both pollination services and honey production.


Classic economics tells us that it is time put into a product or service which gives it its value. Beekeepers know about this; they work all the time, with a lot of hives. They must take care of each colony, all while creating smoke to keep the bees passive. At the IV State Beekeeping Forum, emphasis was made on what a laborious activity this job is.


UN risk assessment researcher, Maria Mercedes Roca, Ph.D. , mentioned how economic activities are central to the functioning of our interconnected world. Beekeeping must be propelled to make for economic gains, especially in countries such as Mexico where a large part of its economy is based on the primary sector. Mr. Manuel Ramirez also referred to this topic; Our country plays a significant role as a honey trader, with most of our exports making their way to the EU. However, there are several obstacles. EU imports can have neither genetically modified content nor most antibiotics. One can’t add anything to a product that would alter its sensorial characteristics without expecting a negative impact. Dr. David de Jong considered we should check what amount of AMPs make their way to honey products, aspect revised in the IV State Beekeeping Forum with Dr. Sóstenes Rafael Rodríguez where we ensured AMP’s are not a problem when it comes to EU honey exportation analysis. Since the national Secretariat of Agriculture, Livestock, Rural Development, Fisheries and Food (SAGARPA) is also strict on contaminants, Guillermo Rodriguéz told us, a natural product against foulbrood is needed both nationally and internationally.


As explained in the chronology of the aforementioned facts, while the project was being developed and basing us on the comments and concerns expressed by professionals, we were able to ensure that our product would not impede exportations and importations of honey. The world honey market reported historical records during 2015, with volumes of operation exceeding 2.3 billion dollars1 and it represents substantial revenues not only for Mexico but countries like China, Argentina, Spain, etc. More than ensuring that highest AMP concentrations in honey would not result in an exportation problem, our project fosters to make exportations go up since there will be no problem with antibiotic residues in honey when beekeepers have to deal with diseases.


The economic part of our project is also made up of the development of the entrepreneurial area where the implications for the sale of the final product are analyzed.


Our project will make sure Foulbrood no longer generates huge economic losses (by killing bees and necessitating antibiotics) but instead become a small nuisance in an otherwise thriving industry.



  1. PROCCyT. (2018). La apicultura; actividad fundamental para alcanzar la seguridad alimentaria en México. Retrieved from http://proccyt.org.mx/noticias/303-la-apicultura-actividad-fundamental-para-alcanzar-la-seguridad-alimentaria-en-mexico-2



Social


While we interacted with beekeepers and researchers, it was not difficult to realize the importance of considering social aspects within the activities and plans for our project.


Rider University explains “sociological imagination” as the ability to see the connection between the larger events affecting the whole society and the individual consequences for each of us in our own lives. Personal troubles may be our fault; but in others, they may be a result of social forces beyond our control as individuals; society shape us as we shape our society.1


Analyzing and acting upon social implications lies in living together in an organized and beneficial way.


In the first approach we had with a beekeeper, Mr. Guillermo Rodríguez, it was explained how beekeeping varies around the world and how many times the needs are different for each region. In our effort to consider all these needs in our project, we managed to have interviews with professionals related to the topic in Brazil, Chile, Mexico, Italy, Valencia, and Canadá, taking into account from beekeepers who manage few hives to complete associations, investigators, governmental institutions and international agencies. We considered specific needs to obtain a universal solution.


In the IV State’s Beekeeping Forum and the CIAD visit, we were able to experience what beekeepers’ work involves. Working with few beehives lessen the job, however in the case of beekeepers like the Chilean Bryaan Saaverdra, that works with more than 1000 hives the work notably intensifies, as you must attend beehive by beehive whatever the weather condition is. Through the application method of our product, which was transformed over the months basing on the concerns and needs of the beekeepers, we foster the welfare of this community taking in consideration for their working days not to become intensified when experiencing the diseases and applying the product.


In the meeting with Mr. Manuel Ramírez Salcido, he commented on how there are almost no young people who are interested in beekeeping and that there is a significant concern for the future of this sector. While developing our project, we saw the enthusiasm of beekeepers for seeing young people interested in the area. Throughout all these months we have shared our project in many places, we have tried to bring the youth to be interested in bee’s importance and beekeeping. For the IV State’s Beekeeping Forum we made an open invitation to different year students of our biotechnology career, encouraging them to attend, to listen to the conferences and learn about apiaries. By proudly conducting this project we are directly involving with beekeeping and fostering an important approach to this sector in the young people that surround us in our daily lives as college students.


After all the research made, the ambiguities that were found when doing the evaluation of the presence of Foulbrood in the world and the different conclusions that were given, (some resulting from the video call with FAI presidential staff member Lorenzo Della Morte and others from the video call with MVZ Juan Diego Pérez de la Rosa) showed that there is a problem in the health and statistical assessment studies regarding bee diseases. Many times, beekeepers who have affected hives, do not report it because of the negative implications that this entails; burning of colonies, the need of using antibiotics that disturb the innocuity of honey and finally the risk of losing investment made over many years. Our product promotes this social problem to be resolved; by having a profitable and useful product, which displace conventional antibiotics usage and the burning of hives. It encourages that the beekeeper openly reports if their beehives suffer from the disease having the peace of mind that he will not have to lose large quantities of money or their hives nor he will need to alter their bee products with antibiotics.


By addressing individual concerns as well as general insights such as the problem of statistical assessment studies, throughout the development of our project and the implementation of the product, we seek to address aspects that are often out of the hands of beekeepers, and that may also have an impact on communities outside of the beekeeping sector. Attention to the previously mentioned issues is based on the fact of building an organized society that interacts effectively within itself and with external communities.


The social area covers the ethical and legal aspects:


Ethical


Three of our human practices activities stood out in determining points that led us to reflect and to investigate more in the situation. Is saving bees our duty? Is our project good? Is it worth taking risks involved? Bioethical dilemmas that will be addressed below.


From the meeting we had with Dr. Roca, we discussed the importance of not only considering human rights but also environmental rights. The next question arose: Should humanity intervene and conserve biodiversity by any means?


The participation in MEXMUN helped us to realize the importance of being responsible for the risks involved in the creation of any biotechnological tool; from there these questions arose: Should we develop the treatment even though it could result in the development of pathogen resistance? Or, in the worst case, damage the innate immune system leaving bees unable to defend themselves?


Based on comments from Ph.D. David de Jong, Dr. Ernesto Guzmán Novoa, M.S. Alejandro Romo Chacón and M.S. Adrián Ponce de León. We realized that there are no regulations to experiment with bees in Canada, Brazil, and Mexico. This led us to question whether the use of bee larvae is justified in order to obtain relevant data for the project to advance. Can even be considered morally relevant?




Legal


From the interviews we had with Mr. Guillermo Rodríguez and Mr. Manuel Ramírez Salcido we realized the importance of considering both national and international governmental agencies involved in the assurance of honey innocuousness. Nevertheless, the legal validation of the development of our project took a big step after the meeting we had with MVZ Ernesto Tanús Sánchez, MVZ Adolfo Arroyo, and Mr. José Luis González Miranda. In this meeting, different legislation that would be ideal to consider was mentioned; concerning innocuity, exportation, Federal Animal Health Law, Zoosanitary regulations as well as Regulations of the European Union regarding food safety.


In addition, our participation in activities of international nature, being, in this case, MEXMUN and the Environmental Risk Assessment Guide for Genetically Modified Organisms, led us to evaluate aspects of biosecurity and risk analysis. We considered it appropriate and necessary to build a legal plan in accordance with the regulations and norms pertinent to our project, covering the local and international scope for ensuring that our project is attached to safety and legality.





  1. Raider University. (n.d.). Why Study Sociology? Retrieved from https://www.rider.edu/academics/colleges-schools/college-liberal-arts-education-sciences/liberal-arts-programs/sociology/why-study-sociology