Molecular Screening of Varroa-Resistant Trait of Honey Bee Colonies based on NorpA2 Candidate Gene Polymorphism: A Genetic Case-Control Study

Subject Areas : Camel

B. Sepehri ¹ , S. Alijani ² , A. Javanmard ³ , H. Johnmohammadi ⁴ , K. Hasanpur ⁵

1 - Department of Animal Science, Faculty of Agriculture, University of Tabriz, Tabriz, Iran
2 - Department of Animal Science, Faculty of Agriculture, University of Tabriz, Tabriz, Iran
3 - Department of Animal Science, Faculty of Agriculture, University of Tabriz, Tabriz, Iran
4 - Department of Animal Science, Faculty of Agriculture, University of Tabriz, Tabriz, Iran
5 - Department of Animal Science, Faculty of Agriculture, University of Tabriz, Tabriz, Iran

Received: 2022-02-23 Accepted : 2022-07-16 Published : 2023-03-01

Keywords: Resistance, single nucleotide polymorphism, candidate gene, <i>Apis mellifera</i>, <i>Varroa destructor</i>,

Abstract :

In recent years the Varroa destructor mite has been a dominant challenge for honey bee colonies, worldwide. Varroa- sensitive hygiene, a behavioral characteristic of honey bee, involves the detection and removal of Varroa larvae from the colony. It has been exploited as a recordable trait for Varroa resistance. The sense of smell is a key element in Varroa sensitive hygiene behavior. In this scenario, NorpA2 candidate gene, a putative olfactory receptor, is associated with vision and smell in honey bee and is known as a candidate gene for Varroa sensitive hygiene. With this motivation, the main goal of the current work was to determine some single nucleotide polymorphism (SNP) markers of NorpA2 candidate gene employing PCR sequencing in a case-control approach. To this end, after the identification of resistant (RES) and susceptible (SUS) colonies according to the percentage of infections of the drone pupae to Varroa mite, a total of 10 drone pupae (5 from SUS and 5 from RES) were selected and were subjected for DNA isolation. The polymerase chain reaction (PCR) was performed based on two pairs of specific primers for the amplification of 5¢ untranslated region (5¢UTR) and promoter regions of NorpA2 gene. After expected size bands were observed, the purification process and Sanger sequencing of the PCR products were carried out. The sequencing results were quality checked and the alignment and clustering were done using the BLAST and MAFTT software, respectively. Sequences of both UTR and promoter regions displayed multiple variations (SNP, deletion). Interestingly, the results showed the existence of three specific differences in sequence in the form of SNP (C/T) at position 308 and the form of SNP/deletion at positions 504 and 563 of the nucleotide sequence region in the promoter between the SUS and RES groups. To our knowledge, this is the first report on the identification of biomarkers to control host-mite interaction in the honey bee Apis mellifera. Further research is required for the reported SNPs to be validated as biomarkers of resistance to Varroa.

References:

Behrens D., Huang Q., Geßner C., Rosenkranz P., Frey E., Locke B. and Kraus F.B. (2011). Three QTL in the honey bee Apis mellifera L. suppress reproduction of the parasitic mite Varroa destructor. Ecol. Evol. 1, 451-458.

Broeckx B.J., De Smet L., Blacquière T., Maebe K., Khalenkow M., Van Poucke M., Dahle P., Neumann P., Bach Nguyen K., Smagghe G., Deforce D., Van Nieuwerburgh F., Peelman L. and de Graaf D.C. (2019). Honey bee predisposition of resistance to ubiquitous mite infestations. Sci. Rep. 9, 1-11.

Cornelissen M. (2015). Insight into the genetic basis of Varroa destructor resistance in Apis mellifera. Laboratory of Genetics, Wageningen University, Wageningen, the Netherlands.

Danka R.G., Rinderer T.E., Spivak M. and Kefuss J. (2013). Comments on: Varroa destructor, research avenues towards sustainable control. J. Apic. Res. 52, 69-71.

Dick D.M., Agrawal A., Keller M.C., Adkins A., Aliev F., Monroe S., Hewitt J.K., Kendler K.S. and Sher K.J. (2015). Candidate gene-environment interaction research: reflections and recommendations. Perspect. Psychol. Sci. 10, 37-59.

Dietemann V., Pflugfelder J., Anderson D., Charrière J.D., Chejanovsky N., Dainat B., de Miranda J., Delaplane K., Dillier F.X., Fuch S., Gallmann P., Gauthier L., Imdorf A., Koeniger N., Kralj J., Meikle W., Pettis J., Rosenkranz P., Sammataro D., Smith D., Yanez O. and Neumann P. (2012). Varroa destructor: Research avenues towards sustainable control. J. Apic. Res. 51, 125-132.

Elmi M. (2019). QTL detection for Varroa mite resistance trait in Azerbaijani bee colonies. PhD. Thesis, University of Tabriz, Tabriz, Iran.

Fries I., Huazhen W., Wei S. and Jin C.S. (1996). Grooming behavior and damaged mites (Varroa jacobsoni) in Apis cerana cerana and Apis mellifera ligustica. Apidologie. 27, 3-11.

Haddad N., Mahmud Batainh A., Suleiman Migdadi O., Saini D., Krishnamurthy V., Parameswaran S. and Alhamuri Z. (2016). Next generation sequencing of Apis mellifera syriaca identifies genes for Varroa resistance and beneficial bee keeping traits. Insect Sci. 23, 579-590.

Jack C.J. and Ellis J.D. (2021). Integrated pest management control of Varroa destructor (Acari: Varroidae), the most damaging pest of (Apis mellifera L. (Hymenoptera: Apidae)) colonies. J. Insect Sci. 21, 6-12.

Kefuss J., Vanpoucke J., De Lahitte J.D. and Ritter W. (2004). Varroa tolerance in France of intermissa bees from Tunisia and their naturally mated descendants: 1993-2004. Am. Bee J. 144, 563-568.

Kirrane M.J., de Guzman L.I., Holloway B., Frake A.M., Rinderer T.E. and Whelan P.M. (2015). Phenotypic and genetic analyses of the Varroa sensitive hygienic trait in Russian honey bee (Hymenoptera: Apidae) colonies. PLoS One. 10, e0116672.

Kurze C., Routtu J. and Moritz R.F. (2016). Parasite resistance and tolerance in honeybees at the individual and social level. Zoology. 119, 290-297.

Le Conte Y., Alaux C., Martin J.F., Harbo J.R., Harris J.W., Dantec C., Séverac D., Cros-Arteil S. and Navajas M. (2011). Social immunity in honeybees (Apis mellifera): Transcriptome analysis of varroa-hygienic behaviour. Insect Mol. Biol. 20, 399-408.

Le Conte Y., De Vaublanc G., Crauser D., Jeanne F., Rousselle J.C. and Bécard J.M. (2007). Honey bee colonies that have survived Varroa destructor. Apidologie. 38, 566-572.

Locke B., Conte Y.L., Crauser D. and Fries I. (2012). Host adaptations reduce the reproductive success of Varroa destructor in two distinct European honey bee populations. Ecol. Evol. 2, 1144-1150.

Meyertholen E.P., Stein P.J., Williams M.A. and Ostroy S.E. (1987). Studies of the Drosophila norpA phototransduction mutant. J. Comp. Physiol. A. 161, 793-798.

Mondet F., Alaux C., Severac D., Rohmer M., Mercer A.R. and Le Conte Y. (2015). Antennae hold a key to Varroa-sensitive hygiene behaviour in honey bees. Sci. Rep. 5, 1-12.

Mondet F., Beaurepaire A., McAfee A., Locke B., Alaux S., Blanchard S., Danka B. and Le Conte Y. (2020). Honey bee survival mechanisms against the parasite Varroa destructor: A systematic review of phenotypic and genomic research efforts. Int. J. Parasitol. 50, 433-447.

Mondet F., Blanchard S., Barthes N., Beslay D., Bordier C., Costagliola G. and Le Conte Y. (2021). Chemical detection triggers honey bee defense against a destructive parasitic threat. Nat. Chem. Biol. 17, 524-530.

Navajas M., Migeon A., Alaux C., Martin-Magniette M.L., Robinson G.E., Evans J.D., Cros-Arteil S., Crauser D. and Le Conte Y. (2008). Differential gene expression of the honey bee Apis mellifera associated with Varroa destructor infection. BMC Genomics. 9, 1-11.

Noël A., Le Conte Y. and Mondet F. (2020). Varroa destructor: How does it harm Apis mellifera honey bees and what can be done about it? Emerging Top. Life Sci. 4, 45-57.

Oddie M.A., Burke A., Dahle B., Le Conte Y., Mondet F. and Locke B. (2021). Reproductive success of the parasitic mite (Varroa destructor) is lower in honeybee colonies that target infested cells with recapping. Sci. Rep. 11, 1-7.

Oxley P.R., Spivak M. and Oldroyd B.P. (2010). Six quantitative trait loci influence task thresholds for hygienic behaviour in honeybees (Apis mellifera). Mol. Ecol. 19, 1452-1461.

Pak W.L. (1979). Study of photoreceptor function using Drosophila mutants. Pp. 67-99 in Neurogenetics: Genetic Approaches to the Nervous System. X. Breakfield, Ed., Elsevier North-Holland, Amsterdam, New York.

 Pollock V.P., Radford J.C., Pyne S., Hasan G., Dow J.A. and Davies S.A. (2003). NorpA and itpr mutants reveal roles for phospholipase C and inositol (1, 4, 5)-trisphosphate receptor in Drosophila melanogaster renal function. J. Exp. Biol. 206, 901-911.

Riesgo-Escovar J., Raha D. and Carlson J.R. (1995). Requirement for a phospholipase C in odor response: overlap between olfaction and vision in Drosophila. Proc. Natl. Acad. Sci. USA. 92, 2864-2868.

Rinderer T.E., Harris J.W., Hunt G.J. and De Guzman L.I. (2010). Breeding for resistance to Varroa destructor in North America. Apidologie. 41, 409-424.

Rosenkranz P., Aumeier P. and Ziegelmann B. (2010). Biology and control of Varroa destructor. J. Invertebr. Pathol. 103, 96-119.

Rosenkranz P., Tewarson N.C., Singh A. and Engels W. (1993). Differential hygienic behaviour towards Varroa jacobsoni in capped worker brood of Apis cerana depends on alien scent adhering to the mites. J. Apic. Res. 32, 89-93.

Schmid-Hempel P. (2021). Evolutionary Parasitology: The Integrated Study of Infections, Immunology, Ecology, and Genetics. Oxford University Press, United Kingdom.

Seeley T.D. (2007). Honey bees of the Arnot Forest: A population of feral colonies persisting with Varroa destructor in the northeastern United States. Apidologie. 38, 19-29.

Tantillo G., Bottaro M., Di Pinto A., Martella V., Di Pinto P. and Terio V. (2015). Virus infections of honeybees Apis mellifera. Italian J. Food Saf. 4, 5364-5375.

Thaduri S., Stephan J.G., de Miranda J.R. and Locke B. (2019). Disentangling host-parasite-pathogen interactions in a varroa-resistant honeybee population reveals virus tolerance as an independent, naturally adapted survival mechanism. Sci. Rep. 9, 1-10.

Traynor K.S., Mondet F., de Miranda J.R., Techer M., Kowallik V., Oddie M.A.Y., Chantawannakul P. and McAfee A. (2020). Varroa destructor: A complex parasite, crippling honey bees worldwide. Trends Parasitol. 36, 592-606.

Tsuruda J.M., Harris J.W., Bourgeois L., Danka R.G. and Hunt G.J. (2012). High-resolution linkage analyses to identify genes that influence Varroa sensitive hygiene behavior in honey bees. PLoS One. 7, e48276.

Underwood R. and López-Uribe M. (2019). Methods to Control Varroa mites: An Integrated Pest Management Approach. Pennsylvania State University Extension, University Park, USA.