2.3.1 Morphological description

The typical symptoms observed in a colony affected by stonebrood are not very different from chalkbrood symptoms and includes irregular capping of the brood. Infected brood, also called "mummies”, can be seen in the combs. Stonebrood mummies turn hard and they resemble small stones, not sponge-like as chalkbrood mummies. Stonebrood mummies are difficult to remove from the cells with forceps and removal by the worker bees is also difficult. Infected brood becomes covered with powdery yellow, brown, green or black fungal spores depending on the species. In some cases infected or deceased larvae looked dry, but they do not produce visible conidia within a 48 hrs after pathogen inoculation (Vojvodic, unpublished)

Stonebrood can be diagnosed by its gross symptoms, but positive identification requires its cultivation in the laboratory and subsequent microscopic examination. Structures of the conidiophores (spore forming structures) are very important for identification of Aspergillus spp. The conidiophores originate from a basal cell located on the supporting hyphae and terminate in a vesicle (Fig. 8). The morphology, colour and roughness of the conidiophores vary from species to species. Additionally, the position of the flask-shaped phialides (spore producing cells) on the vesicle is an important character. The phialides can cover the vesicle surface entirely ("radiate" head) or partially ("columnar" head) and the phialids can be attached to the vesicle directly (uniseriate) or attached via a supporting cell, called metula (biseriate). The phialides produce round conidia (2-5 µm in diameter) that form radial chains (Fig. 8). The conidia of the different species can have different colours (Table 4) (See Fig. 9 for Asp. flavus in vitro infected larvae). We however recommend contacting a mycologist for correct species identification of the Aspergillus specimens.

Table 4. Microscopic characters of three Aspergillus species most often reported to cause stonebrood.

Species

Conidia colour

Conidiophores

Phialides

Vesicle

Asp. flavus

Yellow-green

Colourless. Rough

Uni/biseriate

Round, radiate head

Asp. fumigatus

Blue-green to grey

Short (<300µm), smooth, colourless-green

Uniseriate

Round, columnar head

Asp. niger

Black

Long, smooth, colourless or brown

Biseriate

Round, radiate head

 

Fig. 8. Structures of importance for identification of Aspergillus species.

Figure 8

Fig. 9. In vitro reared honey bee larvae infected with Aspergillus flavus: the bottom left cell contains larvae in an early stage of disease visible by the change in larval colour; bottom right cell contains later stage of the diseases with the visible fungal body and conidia protruding out of the larval cuticle. The two upper cells contain healthy larvae. Photo: S Vojvodic.

Figure 9