By Clarence Collison
Reproducing Varroa females lay the first egg in the brood cell approximately 70 hours after host cell capping.
The life cycle of the female Varroa mite is subdivided into a phoretic phase in which she lives on adult bees and a reproductive phase occurring within worker or drone brood cells. The reproductive phase is initiated when the female mite leaves the adult host and enters a brood cell with a 5th instar larva shortly before the cell is capped. This foundress female passes between the larva and the cell wall to the bottom of the cell and becomes stuck within the larval food (larval jelly). Approximately five hours after cell capping, the bee larva has consumed the rest of the larval food which frees the mite (Ifantidis 1988). At that time the female mite has already started oogenesis (creation of eggs) in the terminal oocyte (Steiner et al. 1994; Garrido et al. 2000).
After leaving the larval food, the mite begins feeding on hemolymph of the bee prepupa. The Varroa mother prepares a feeding site by making a wound in the prepupa cuticle, which is used by all mite offspring including the male. This feeding site is critical for the survival of all developmental stages, because their mouthparts are not strong enough to pierce the soft cuticle of the bee pupae. The infesting mother mite also forms a rendezvous site with her feces on the cell wall on which all mobile individuals aggregate and on which matings preferentially occur (Donzé and Guerin 1994). In cells infested by more than one Varroa foundress mite, no aggressiveness between them has been observed and the members of the different families construct and cohabit the feeding punctures and fecal accumulations. The increased number of progeny in such cells does, however, leads to competition at the feeding site.
Female mites may invade worker or drone brood cells when worker bees bring them in close contact with brood cells. The attractive period of drone brood cells is two to three times longer than that of worker brood cells. The attractiveness of brood cells is related to the distance between the larva and the cell rim and the age of the larva. The moment of invasion of the mite into a brood cell is not related to the duration of its stay on adult bees. The fraction of the phoretic mites that invade brood cells is determined by the ratio of the number of suitable brood cells and the size of the colony. The distribution of mites over worker and drone brood in a colony is determined by the specific rates of invasion and the numbers of both brood cells (Beetsma et al. 1999).
Garrido et al. (2000) determined the moment of activation of oocyte growth in Varroa females. Ovaries of the mites were dissected and stained with toluidine blue. The coloration of the terminal oocyte indicates the uptake of euplasmatic and/or yolk material and therefore, the initiation of the reproductive phase. In phoretic mites removed from adult bees, no staining of the ovary was detected. Females artificially introduced into freshly capped brood cells and removed for dissection six hours later already showed clear blue staining of the terminal oocyte. The ovaries of female mites introduced 14 hours after capping of the brood cell, however, remained uncolored after incubation in toluidine blue. In phoretic mites, oogenesis is apparently arrested in a previtellogenic phase. Immediately after invasion of the brood cell, reproduction is activated by some factor. This factor is present in freshly capped brood cells but not in brood cells 14 hours after capping. Oocyte growth in reproductive mites depends on the consumption of hemolymph from freshly sealed larvae (Donzé and Guerin 1994; Tewarson and Engels 1982).
Reproducing Varroa females lay the first egg in the brood cell approximately 70 hours after host cell capping (Ifantidis 1983; Steiner et al. 1994). This egg is unfertilized and develops into a male, while the three to four subsequent eggs that are laid at approximately 30 hour intervals are fertilized and develop into female offspring (Rehm and Ritter 1989; Martin 1994). However, the last eggs laid will usually not reach maturity, because the developmental time of the immature bee in the capped cell is too short to allow completion of mite development. Since the capped stage of drone cells is about two days longer than that of worker cells (Jay 1963), drone cells are in principle more rewarding in terms of mite reproduction than worker cells because more young mites can reach maturity. In the European honey bee, mites produce on average two to three viable female offspring in drone cells and one or two viable female offspring in worker cells (Schulz 1984; Fuchs and Langenbach 1989).
The mite larva develops within the egg during the first hours after ovipositon. During the period of time from egg hatch until adult molt, the mite offspring pass through protonymphal and deutonymphal stages. The total development time is about 5.8 and 6.6 days for female and male mites, respectively (Donzé and Guerin 1994; Martin 1994; Rehm and Ritter 1989).
Using transfer experiments, Garrido and Rosenkranz (2003) examined whether the sequence of sexes (first egg unfertilized, followed by fertilized eggs) in the brood cell is triggered by a host stimulus. When reproducing Varroa females were transferred from white-eyed worker pupae into freshly capped worker brood cells, 77% of the fertile mites after the transfer began a new reproductive cycle by laying an unfertilized egg. The proportion of brood cells with male offspring was similar to naturally infested brood cells. Varroa females transferred into brood cells with young pupae reproduced, but only 6% of the fertile mites after the transfer produced male offspring. This was significantly different from male production in naturally reproducing Varroa females and those transferred into freshly capped brood cells. They concluded that a host stimulus present in freshly capped brood cells triggers both the start of reproduction and the sequence of sexes.
The reproductive cycle of the Varroa mite is closely linked to the development of the honey bee host larvae. Using a within colony approach, phoretic Varroa females were introduced into brood cells of different ages in order to analyze the capacity of certain stages of the honey bee larva to either activate or interrupt the reproduction of Varroa females (Frey et al. 2013). Only larvae within 18 hours (worker) and 36 hours (drones), respectively, after cell capping were able to stimulate the mite’s oogenesis. Stage specific volatiles of the larval cuticle are at least part of these activation signals. This is confirmed by the successful stimulation of presumably non-reproducing mites to oviposition by the application of a larval extract into the sealed brood cells. Preliminary quantitative gas chromatography-mass spectrometry analyses suggest certain fatty acid ethyl esters which make up brood pheromone, as candidate compounds. If Varroa females that have just started egg formation are transferred to brood cells containing host larvae of an elder stage, two-thirds of these mites stopped their oogenesis. This confirms the presence of an additional signal in the host larvae allowing the reproducing mites to adjust their own reproductive cycle to the ontogenetic development of the host. From an adaptive point of view, that sort of a stop signal enables the female mite to save resources for a next reproductive cycle if their own egg development is not sufficiently synchronized with the development of the host.
The reproduction of Varroa mites during successive honey bee brood cycles was investigated (de Ruijter 1987). Newly capped worker brood cells were identified and into each cell an adult female mite was introduced. After 10 days the cells were opened and the contents examined. Those females still present and alive were once again introduced into newly capped brood cells and so on. Varroa mites were capable of reproducing up to seven times under these experimental conditions. The maximum number of eggs laid was 30 per female. Females that produced only male offspring because they were unmated, kept doing so in subsequent brood cycles. Though in contact with adult males several times, no successful mother mite matings occurred. Probably only young females mate successfully.
The male mates with the female offspring of the mother mite in the brood cell and only the mother and daughter females emerge from the cell. Protandry (appearance of males prior to females) in Varroa enables the fertilization of a maximum number of daughters within the limited time span of the capped brood. To be successful, however, the newly emerged adult daughters must encounter a male. However, adult males are scarce, occurring in only 60% of single infested cells due to developmental mortality (Fuchs and Langenbach 1989).
The mating of Varroa daughters after ecdysis (molting) and as soon as they arrive on the fecal accumulation prepared by the mother mite (Donzé et al. 1996). Such females are remated for as long as no other freshly molted daughter arrives on the fecal accumulation. The number of spermatozoa stored in the mite’s spermatheca increases with remating, a strong indication for sperm mixing when brood cells contain more than one Varroa foundress. The number of daughters per infesting mother decreases at higher rates of infestation per cell, but the proportion of such daughters with a mate rises sharply due to the higher probability of finding a male within multi-infested cells. The number of mated daughters per mother is maximal in cells with two foundress Varroa females.
Martin (1995) investigated the developmental times and mortality of Varroa in drone cells. The position and time of capping of 2671 naturally infested drone cells were recorded. Six hours after the cell was capped, 90% of the mites were free from the brood food to start feeding on the developing drone. The developmental time of the mite’s first three female offspring (133±3 hours) and the male offspring (150 hours) and the intervals between egg laying (20-32 hours) were similar to those found in worker cells. However, the mortality of the offspring was much lower in drone cells than worker cells. The mode number of eggs laid were six and five in drone and worker cells, respectively. All offspring had ample time to develop fully in drone cells with the sixth offspring reaching maturity approximately one day before the drone bee emerged. Normal mites (those which lay five or six viable eggs) produced on average four female adult offspring but since only around approximately 55% of the mite population produced viable offspring the mean number of viable adult female offspring per total number of mother mites was two to 2.2 in drone cells.
Within any mite population, large numbers of mites fail to produce fertile female offspring despite entering a suitable host cell. These can be classed into those that do not lay eggs, those that lay non-viable eggs and those that only produce viable male offspring. Another cause which leads to the production of non-fertile females is the premature death of the male offspring before it is able to mate with its sisters. This situation arises because female mites only produce a single male during each reproductive cycle and this male needs to fertilize all of his sisters (Martin et al. 1997).
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Clarence Collison is an Emeritus Professor of Entomology and Department Head Emeritus of Entomology and Plant Pathology at Mississippi State University, Mississippi State, MS.