Molecular identification of a male‐killing agent in the ladybird Harmonia axyridis (Pallas) (Coleoptera: Coccinellidae)

Inherited bacteria that manipulate the reproduction of their hosts are common in invertebrates. Of four known manipulative strategies caused by bacteria in insects, three, namely cytoplasmic incompatibility, feminization and parthenogenesis induction, appear to be caused solely by members of the genus Wolbachia . In contrast, the fourth, male-killing, is associated with a variety of bacteria. Within the coccinellids, which are particularly prone to invasion by male-killing endosymbionts (Majerus & Hurst, 1997), bacteria of four groups, Rickettsia (Werren et al ., 1994), Spiroplasma (Hurst et al ., 1999a), Flavobacteria (Hurst et al ., 1997) and Wolbachia (Hurst et al ., 1999b) have been reported to be associated with the male-killing trait. That one species of coccinellid, Adalia bipunctata , harbours four different bacteria, a Rickettsia , a Spiroplasma and two Wolbachia , demonstrates the susceptibility of these beetles to male-killers. Two questions arise from these observations. First, are there constraints on the evolution of a male-killing strategy within inherited bacteria, or can all such bacteria evolve this phenotype? Second, is invasion by more than one malekiller a general feature of those insect host species which are susceptible to male-killing? The Asian ladybird Harmonia axyridis (Pallas) harbours a maternally inherited, female-biased sex ratio trait, taken to result from male-killing (Matsuka et al ., 1975; Gotoh & Niijima, 1986; Majerus et al ., 1998). This species shows all the features suggested by Majerus & Hurst (1997) to be necessary for the invasion and establishment of malekillers in a coccinellid: aphidophagy, laying eggs in tight batches, sibling egg cannibalism and significant levels of neonate larval mortality due to starvation. Previous studies have shown variation in the vertical transmission efficiencies between male-killing matrilines (Matsuka et al ., 1975; Majerus et al ., 1998), and both progressive sex ratio families (progenic sex ratio becomes progressively female biased) and revertant families (initial female bias in progenic sex ratio is lost) (Matsuka et al ., 1975). Differences in the phenotypic expression of female-biased traits might be indicative of different causative agents of male-killing, making H. axyridis a suitable candidate in which to seek diversity in male-killers over geographical distance. Furthermore, establishment of the identity of the male-killer(s) in this species may help indicate which bacteria are most prone to evolving male-killing.