Benefits of Propolis to Honey Bee Health and Immunity
The goal of this research is to test the hypothesis that honey bees’ construction of a propolis envelope within the nest cavity is a vital component of individual and social immunity, enhancing bee health and colony survivorship. The benefit of a natural propolis envelope to colony health and immune system functioning is being quantified by measuring immune-related gene transcripts, bacterial loads, and colony-level measures of diseases, mites, population strength and winter survival. If a heavy propolis envelope is a vital component to a healthy bee colony, we can modify beekeeping equipment currently used by beekeepers nationwide. Such modifications will encourage the bees’ natural construction of a antimicrobial propolis envelope in the nest cavity. A long-term outcome of this research is to promote honey bee health, which will directly support local, regional and national beekeepers by having stronger colonies to produce more honey.
Quantify the relative benefit of these propolis configurations to the immune system of individual bees and to viral levels within the colony. Thirty-six colonies were established in new 10-frame equipment in April 2012. Sister queens were introduced into each colony to reduce genetic variation. Twelve colonies were provided with commercial propolis traps stapled to the four inner walls of each bee box to encourage the bees to build a propolis envelope (Figure 1a). Another 12 colonies were provided with the propolis traps only on top of the frames, as is done to collect propolis commercially (Figure 1b). The last 12 colonies served as controls; no propolis trap was provided and the bees deposited propolis in the cracks and crevices within the box where they could (Figure 1c). Seven day old, marked bees were collected in July 2012, September 2012 and May 2013 to compare the relative immune-gene expression of bees from among the treatments. Candidate genes for immune-related functions were selected based on their relevance for the honey bee immune system (Evans et al. 2009), and included: the antimicrobial peptides (AMPs) hymenoptaecin, abaecin, denfensin-2, defensin-1; NF-κB-like Relish and a candidate for cellular immunity (prophenoloxidase activated enzyme). The transcription factor levels of blood storage protein (vitellogenin) and the three most commonly identified viruses in bee colonies were also measured; Deformed Wing Virus (DWV), Black Queen Cell Virus (BQCV) and Israeli Acute Paralysis Virus (IAPV). To assess bacterial loads, we used a generic primer for 16S ribosomal DNA amplification. Total RNA was isolated from individual bees using TRIzol (Life Technologies). RNA was reverse transcribed using Superscript II (Invitrogen) and cDNA was quantified through real-time PCR. Colony strength (disease levels, number of adult bees and brood) was also assayed to analyze additional benefits of propolis to colony health and survivorship. The experiment was repeated on a new set of colonies in 2013.
Benefits of propolis to colony health and strength
- There was no significant difference in levels of Varroa mite or Nosema in the colonies among the treatments in either replication of the experiment; all levels were low and no colonies received treatments for Nosema.
- There was no significant difference in the population size of adult worker bees in colonies that survived in May 2013 or 2014.
- Colonies with a propolis envelope had significantly more brood in May 2013, and slightly more brood in 2014 compared to colonies in the other two treatments (control and propolis trap colonies)
Relative benefit of propolis to the immune system of 7-d old bees:
- Summer and Fall: As expected, the transcription levels of hymenoptaecin and abaecin in bees from colonies with a propolis envelope were significantly lower compared to bees from control colonies in July 2012. In September 2012, all candidates for the immune system response were been expressed at significantly lower levels in bees from the propolis envelope treatment compared to controls. In September 2013, significant differences were observed for hymenoptaecin and abaecin. Bees in colonies with propolis traps on top of the colony showed immune responses that were either intermediate, or more variable relative to bees in the propolis envelope and control colonies.
- Spring of the following year: All colonies were wintered in the boxes according to the treatment they had received. In May 2013, bees in the propolis envelope treatment showed higher transcription of three immune-related genes (defensin-1, relish, prophenoloxidase activated enzyme) and vitellogenin compared to control colonies. Expression levels in bees from colonies with propolis traps varied among the genes. In contrast, in May 2014, only vittelogenin showed significant differences in expression level among the treatments. As in the previous year, it was found higher in bees from the colonies with a propolis envelope. Vittelogenin (Vg) level is a good marker of nutrition status because it is the main storage protein for bees and a precursor for many other proteins. The significantly higher levels of Vg in bees from propolis envelope colonies in May 2013 and 2014 suggests that these bees had more protein storage in the spring compared to bees in control colonies.
These preliminary results suggest that the propolis envelope was more beneficial to the bees’ immune system during the summer and fall compared bees in colonies with no propolis envelope. The propolis traps on top of the colonies yielded more variable results. The first set of colonies that survived the winter in May 2013 had very different immune gene transcript results compared to the second set in May 2014. It may be the propolis did not retain its activity over the winter, which we are testing with further laboratory tests.
Relative benefit of propolis to viral levels within the colony:
- There was no significant difference in levels of all three viruses (DWV, IAPV and BQCV) in bees from colonies with the propolis envelope compared to bees from control colonies.
- In May 2014, bees from colonies with a propolis envelope showed significantly lower levels of BQCV compared to propolis trap treatment.
- Bees from colonies with a propolis trap on top of the frames had significantly higher levels of DWV compared to bees in control and propolis envelope colonies in September 2012, May 2013 and May 2014.
The presence of high levels of virus has been correlated with individual immunosuppression and colony death. In this study, we analyzed the expression of the three most common viruses in honey bee colonies. We hypothesize that the presence of a propolis trap on top of the colony could have altered the microenvironment of the colony (e.g. increasing humidity levels or affecting air circulation within the nest), leading to favorable conditions to the growth of pathogens and maybe viruses. Thus, it appears that leaving a propolis trap on top of a colony over the winter is not beneficial to bee health. The propolis envelope was neutral in its effect, and did not appear to reduce virus levels.
Impacts and Contributions/Outcomes
Currently, the honey bee population is experiencing a rapid decline in the United States due to the combined effects of diseases, parasites, pesticides and nutritional deficiencies (Cox-Foster et al., 2007; Johnson et al., 2009, 2010; Runckel et al. 2011; Spivak et al., 2011). Since the term Colony Collapse Disorder was coined in 2006 (Skokstad, 2007), national surveys reveal that 30-35% of all honey bee colonies die every winter across the United States (vanEngelsdorp et al., 2007, 2008, 2010, 2011). Beekeepers make up losses by splitting surviving colonies or by purchasing new colonies from national bee breeders and distributors. However, this loss is not sustainable and it is critical that research is undertaken in order to develop methods to improve the natural defenses of honey bees. To our knowledge, this is the first study on the seasonal benefits of propolis for honey bee health and immunity. This research reveals the benefits of propolis to honey bee immune system throughout the year and after the winter, which has a high potential to positively impact bee health, colony strength and consequently, the beekeeping industry. The preliminary results of this research have been presented by R. Borba in nationwide (2013 Entomological Society of America in Austin-TX and 2012 annual North American Pollinator Partnership Conference in Washington-DC, 2014 North Central Branch-ESA in Des Moine-IA) and local meetings (Minnesota Hobby Beekeepers Association, Minnesota Honey Producers Association and Wisconsin Honey Producers Association). M. Spivak has presented on this topic to over 5 different professional and public meetings of beekeepers, scientists and the general public across the US. Beekeeping short-courses offered by the Spivak lab also provide beekeepers easy access to the findings and provide a way to educate beekeepers on sustainable ways to encourage propolis collection. A national survey on bee health has been initiated through the Bee Informed Partnership (www.BeeInformed.org) and beekeepers can evaluate and provide anonymous feedback on the effectiveness of our findings through this website.
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