As is well known, the female Varroa mite feeds during its reproductive phase on the internal fluids of bees undergoing nymphosis. To do so, it must perforate the integument of its host, a process that takes nearly one hour; the resulting wound, usually located on the second abdominal segment, must remain open for the entire period required to feed both the foundress and her entire progeny. Young Varroa mites are in fact unable to pierce the bee’s cuticle, and it is therefore up to the foundress to create the feeding site that will sustain the whole family. This represents a considerable challenge for the parasite, as it must penetrate a layer that is extremely resistant at its scale, avoid triggering an immune response in the parasitised bee, and at the same time protect the host from other pathogens that could kill it and thus deprive the mite of the resource necessary to complete its reproductive cycle.

In parasitic arthropods, saliva generally plays an important role in this respect. In Varroa specifically, earlier studies have shown that saliva is involved in the dissolution of fat body tissue, on which the phoretic form feeds on adult bees [1], and that it is toxic to female larvae of Apis cerana—for this reason, Varroa destructor naturally avoids parasitising female brood of this bee species (Zhang and Han 2018). These observations prompted a scientific team to investigate the possible effects of an enzyme present in the saliva of this mite.

The researchers first analysed the genome [2] of Varroa destructor (decoded in 2017) to identify all genes potentially encoding proteins found in the venom or saliva of parasitic arthropods. They then searched among these genes for those that were overexpressed in the mite’s saliva, by comparing expression levels in the salivary glands with those in the rest of the body.[1]

A single protein clearly emerged from this analysis: a chitinase, that is, an enzyme capable of degrading chitin, which is expressed about 340 times more strongly in the salivary glands than in the rest of the Varroa organism (Figure 1) [2]. This enzyme likely facilitates the parasite’s penetration of the bee cuticle [3] and contributes to keeping the wound open for the entire period required to feed the foundress and her offspring.

 

Figure 1: Light microscopy image of the anterior part of a stained Varroa mite. Sg: salivary glands; C: chelicerae; P: pedipalps; B: brain. The blue dye “recognises” the RNA responsible for expression of the gene encoding chitinase. As shown, staining is confined to the salivary glands. This is one of the methods by which researchers identified chitinase as a saliva-specific component, enabling subsequent study of its effects in Varroa and in the bee. The scale bar represents 1/10 of a millimetre (100 µm). Source: Becchimanzi et al. 2020, image under a Creative Commons Attribution licence.

 

The researchers then silenced the chitinase gene in groups of ten Varroa mites. Specifically, the mites were immersed in a bath containing RNA that blocks the DNA encoding the enzyme; treated mites were therefore unable to produce chitinase. Bee pupae were then infested with groups of untreated Varroa mites and with groups of treated, chitinase-deficient mites.

They observed that mortality in Varroa mites lacking chitinase exceeded that of control mites by more than 60%, demonstrating that this enzyme plays a crucial role in parasite survival. Furthermore, examination of the parasitised bee nymphs revealed an effect of chitinase on host immunity. Several genes associated with immune responses were expressed at much higher levels when the nymph was parasitised by a chitinase-deficient Varroa than when it was parasitised by an untreated mite. These genes include those encoding abaecin, apidaecin and hymenoptaecin, antimicrobial substances essential in defence against fungal and bacterial diseases. Production of these antimicrobial compounds is therefore suppressed by the chitinase normally present in Varroa saliva. This confirms and helps to explain (at least in part) the well-established observation that infestation by Varroa destructor weakens the bee’s immune defences. Finally, chitinase may also play a role in regulating microbial proliferation around the parasite’s feeding site.

 

Figure 2: Scanning electron microscopy image of a Varroa feeding site located on the intersegmental membrane of a bee. The mite was removed for the photograph. The feeding orifice is clearly visible (black arrow), as is the imprint left by the mite on the membrane and the adhesive pads at the ends of the forelegs (white arrows), which remained attached when the Varroa was removed. This remarkable image was taken at the USDA-ARS in Beltsville, USA. Source: Ramsey SD et al. 2019, reproduced with the kind permission of S. Ramsey.

 

This represents a further step towards the development of biocontrol strategies against Varroa. In theory, bees could be treated with interfering RNA capable of blocking a parasite gene—here, the gene encoding chitinase. This approach is all the more promising given that RNA has been shown to undergo bidirectional transfer, from treated bees to Varroa and from Varroa to other bees (Garbian et al. 2012), which would ensure effective distribution of the treatment throughout the hive.

 

[1] See “Lu pour vous” in LSA No. 291, pp. 223–226.

[2] The genome is the complete set of genes of a species; concretely, it is a series of molecular sequences (each represented by a letter) that contain, in encoded form, the instructions for all the proteins that an organism of that species must be able to synthesise in order to survive, develop and reproduce.

[3] Chitin, a substance similar in composition to cellulose, forms fibres that contribute greatly to the mechanical strength of the cuticle.

 

You may also be interested in:

•  Varroa does not feed on blood

•  Varroa control: summer brood interruption

•  Resistance to Varroa

•  Varroa explained by Dr Joseph Létondal

•  “Varroa-resistant” bees?

•  Varroa, master of cuticular mimicry

 

Bibliography

Source article: Becchimanzi A et al. (2020): A salivary chitinase of Varroa destructor influences host immunity and mite’s survival, PLoS Pathog 16(12): e1009075. https://doi.org/10.1371/journal.Ppat.1009075

Garbian Y et al. (2012): Bidirectional Transfer of RNAi between Honey Bee and Varroa destructor: Varroa Gene Silencing Reduces Varroa Population, PLoS Pathog 8: e1003035. https://doi.org/10.1371/journal.ppat.1003035 PMID: 23308063

Zhang Y and Han R (2018): A Saliva Protein of Varroa Mites Contributes to the Toxicity toward Apis cerana and the DWV Elevation in A. mellifera, Scientific Reports 8. https://doi.org/10.1038/s41598-018-21736-9 PMID: 29467400