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Amitraz Resistance in French Varroa Mites: Not Related to a Single Mutation

Table of Contents

In this study, varroa mites from French apiaries were collected in 2020 and 2021 to assess their laboratory sensitivity to amitraz (CL90). Then, 240 of these varroa mites were genotyped to identify if the mutation at position 260 of the ORβ-2R-L gene (previously linked to amitraz resistance) was present or not. Finally, amitraz-based strip treatments were applied in the hives where the mites were collected to measure the treatment efficacy.

The findings revealed regional occurrences of amitraz resistance, with a less severe decrease in treatment efficacy compared to other pesticides. Notably, the genetic analysis showed that the previously identified mutation at position 260 may not directly cause amitraz resistance, questioning the role of this genetic marker and suggesting a more complex resistance mechanism.

 

The study can be viewed in its entirety at this link.

Note: The publication mentions the term “Single nucleotide polymorphism (SNP)” which is a small genetic variation that occurs when a single nucleotide (the building blocks of DNA) in the genome sequence is altered. However, as this term might not be familiar to everyone, we have sometimes replaced it with the word “mutation” in the following article, a term frequently used in recent publications on resistance. It is important to note, though, that scientifically speaking, a SNP is more accurately described as a “variation” rather than a true mutation.

Introduction

This study investigated the emergence of amitraz resistance in Varroa destructor mite populations across different regions of France. Amitraz, a widely used formamidine acaricide, has been a cornerstone in varroa mite control, but recent reports of diminishing treatment efficacy, have sparked concerns about developing resistance. Unlike the rapid spread of pyrethroid (tau-fluvalinate and flumethrin) resistance, amitraz resistance appears to manifest as “islands of resistance” with initially less severe declines in efficacy, creating a unique and intriguing pattern of resistance emergence.

Materials and Methods

In 2020 and 2021, laboratory bioassays were conducted to assess varroa mite susceptibility to amitraz using mites collected from commercial apiaries across France. Two hundred and forty mites classified as “sensitive” or “resistant” were genotyped, focusing on a single nucleotide polymorphism (SNP) at position 260 of the Orβ-2R-L gene (octopamine receptor), previously linked to amitraz resistance. Additionally, a field trial evaluating Apivar® (Amitraz) efficacy was performed.

To sum up, the study was done in 3 steps, performed on the same mite populations:

  • Laboratory Susceptibility Testing:

Mites were exposed to a lethal amitraz concentration (LC90, Lethal Concentration for 90% of the study population) integrated into paraffin capsules following a standardized procedure. Mite mortality rate was measured to indicate susceptibility. The mite populations were classified as “sensitive”, “moderate” and “resistant”.

  • Genotyping of Orβ-2R-L Gene and Potential Amitraz Resistance Links:

The classified mite populations were genotyped, focusing on the SNP at position 260 of the Orβ-2R-L gene, which encodes the β-adrenergic-like octopamine receptor, the primary target of amitraz. This SNP causes an amino acid substitution (N87S) that may alter amitraz binding.

  • Field Efficacy of Apivar®:

Apivar® efficacy was evaluated on 50 colonies (previously sampled for the laboratory assay and genotyping analysis) in two regions of France (Occitanie and PACA) with strips inserted in late August/mid-September 2021.

Results

The results confirm the presence of amitraz resistance “islands” with a less severe decline in efficacy compared to pyrethroids.

However, the genetic analysis raises doubts about the direct role of the mutation at position 260 in resistance. Indeed, the mutation at position 260 was present in both “resistant” and “sensitive” mite populations. In other words, this mutation was found in mites that died from the amitraz exposure (= sensitive), and mites that survived amitraz exposure (= resistant). This means that the presence of this mutation is not fully responsible for the development of amitraz resistance.

Regarding the field efficacy, the average efficacy was 93%, with 95.5% efficacy in Occitanie region and 90.5% in PACA region.

Discussion

While amitraz resistance is confirmed in some varroa mite populations in France, its spread and impact differ significantly from the rapid proliferation of pyrethroid resistance.

The results of this study challenge the hypothesis that the mutation at position 260 of the Orβ-2R-L gene is the sole direct cause of amitraz resistance. This contrasts with earlier findings by Hernández-Rodríguez et al. (2021); Rinkevich et al. (2023), which identified this mutation as a key genetic marker for resistance. Rinkevich et al. observed a strong correlation between the presence of the 260 A>G mutation and mite survival after amitraz exposure in U.S. populations. Similarly, González-Cabrera et al. linked this mutation to resistance in Spanish Varroa populations.

However, this study did not establish a direct link between the mutation 260 and amitraz survival, suggesting that other factors and more complex mechanisms are involved in resistance development.

Additional variations were identified at positions 344-345 of the Orβ-2R-L gene, more common in “resistant” mites, indicating a potential role in resistance that needs further exploration.

The findings suggest that the varroa mite genome is more variable than initially assumed, with ongoing adaptations to the host and environment, including pesticide exposure. Amitraz resistance, therefore, appears to involve complex evolutionary mechanisms rather than being driven by a single polymorphism.

The authors propose that the previously identified mutation at position 260 is not solely responsible for amitraz resistance, which likely arises from additional genetic variations and evolutionary processes. This complexity makes detecting resistance through genetic markers more challenging than simply identifying the presence of a single mutation.

Key Takeaways

  • Regional occurrences of amitraz resistance were identified in French varroa mite populations.
  • The reduction in amitraz treatment efficacy was less severe compared to resistance against other pesticides like pyrethroids.
  • Genetic analysis challenged the idea that the previously identified mutation (or SNP) at position 260 is the direct cause of amitraz resistance, indicating a more complex resistance mechanism.

References

  • Marsky U, Rognon B, Douablin A, Viry A, Rodríguez Ramos M.A, Hammaidi A. Amitraz Resistance in French Varroa Mite Populations—More Complex Than a Single-Nucleotide Polymorphism. Insects 2024, 15, 390. https://doi.org/10.3390/insects15060390
  • Hernández-Rodríguez CS, Moreno-Martí S, Almecija G, Christmon K, Johnson JD, Ventelon M et al., Resistance to amitraz in the parasitic honey bee mite varroa destructor is associated with mutations in the β-adrenergic-like octopamine receptor. J Pest Sci 95:1179-1195 (2021).
  • Rinkevich FD, Moreno-Martí S, Hernández-Rodríguez CS, González-Cabrera J. Confirmation of the Y215H mutation in the β2 -octopamine receptor in Varroa destructor is associated with contemporary cases of amitraz resistance in the United States. Pest Manag Sci. 2023 Aug;79(8):2840-2845. doi: 10.1002/ps.7461. Epub 2023 Apr 11. PMID: 36947601.
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