Prevalence of a Non-Male-Killing Spiroplasma in Natural Populations of Drosophila hydei

Abstract

Male-killing phenotypes are found in a variety of insects and are often associated with maternally inherited endosymbiotic bacteria. In several species of Drosophila, male-killing endosymbionts of the genus Spiroplasma have been found at low frequencies (0.1 to 3%). In this study, spiroplasma infection without causing malekilling was shown to be prevalent (23 to 66%) in Japanese populations of Drosophila hydei. Molecular phylogenetic analyses showed that D. hydei was infected with a single strain of spiroplasma, which was closely related to male-killing spiroplasmas from other Drosophila species. Artificial-transfer experiments suggested that the spiroplasma genotype rather than the host genotype was responsible for the absence of the male-killing phenotype. Infection densities of the spiroplasma in the natural host, D. hydei, and in the artificial host, Drosophila melanogaster, were significantly lower than those of the male-killing spiroplasma NSRO, which was in accordance with the hypothesis that a threshold infection density is needed for the spiroplasma-induced male-killing expression.Incidences of male-specific death at early developmental stages, called male killing, have been found in a variety of insects, such as ladybird beetles, fruit flies, butterflies, and moths (16). In many of these cases, the causal agents have been identified as maternally inherited endosymbiotic bacteria, such as Spiroplasma, Rickettsia, Wolbachia, and Arsenophonus (18).Members of the genus Spiroplasma, belonging to the class Mollicutes, are spiral, wall-less, actively motile, very small (0.1 to 0.2 m thick and 4 to 5 m long), and associated with plants and arthropods. For instance, Spiroplasma citri, Spiroplasma phoeniceum, and Spiroplasma kunkelii are known as plant pathogens transmitted by insect vectors; Spiroplasma apis and Spiroplasma melliferum as pathogens of the honeybee; and Spiroplasma poulsonii as a maternally inherited male killer in Drosophila fruit flies (44).In Central and South American countries, S. poulsonii and its relatives with a male-killing phenotype have been found in the Drosophila willistoni species group, including D. willistoni, Drosophila nebulosa, Drosophila equinoxialis, and Drosophila paulistorum, and have been called by the acronyms WSRO (from willistoni sex ratio organism), NSRO, ESRO, and PSRO, respectively (45). Recently, a male-killing spiroplasma that was indistinguishable from NSRO on the basis of several gene sequences was found in Drosophila melanogaster in Brazil and designated MSRO (27,28). Infection frequencies of the malekilling spiroplasmas are generally low: 0.1 to 3% in the D. willistoni species group (45) and 2.3% in D. melanogaster (28).In a Japanese population of Drosophila hydei, Ota et al. (31) microscopically observed a high frequency (46%) of spiroplasma infection without the male-killing phenotype. To date, this case has been the only report of a non-male-killing spiroplasma occurring in natural Drosophila populations. It was of great interest to discover whether the high...

Effects of host genotype against the expression of spiroplasma-induced male killing in Drosophila melanogaster

“…Although the nuclear factors of Sevelen and Hikone suppressed the expression of male killing to considerable levels, they could not accomplish complete suppression of male killing (that is, when mothers become sufficiently old, suppression of male killing is attenuated and eventually lost). Similar results were obtained in an earlier study using the spiroplasma strain WSRO (male-killing spiroplasma derived from D. willistoni), which is closely related to NSRO (Montenegro et al, 2005;Kageyama et al, 2006). Specifically, microinjection of WSRO-laden hemolymph into D. melanogaster revealed that the male-killing intensity was high in Oregon-R, but weak in Sevelen (Sakaguchi and Poulson, 1963).…”

“…For artificial infection of NSRO, microinjection of NSROladen hemolymph was performed as described earlier (Kageyama et al, 2006). Briefly, collection and injection of hemolymph was conducted under a dissecting microscope with thin glass capillary tubes made with a microelectrode puller (PN-3; Narishige, Tokyo, Japan).…”

Evolution of early male-killing in horizontally transmitted parasites

“…The observed spiroplasma is a close relative to MK spiroplasmas from other Drosophila species and, at the same time, the most basal taxon of the clade. Two parsimonious scenarios for the evolution of MK in this spiroplasma clade were suggested [17]. First, a common non-MK ancestor of the clade could evolve MK ability, which was subsequently lost in the spiroplasma lineage of D. hydei, a scenario indicative of the evolutionary branching we observed in our study.…”

“…However, highly efficient male-killers might have evolved gradually from less-efficient male-killers, and from bacteria that did not possess any MK ability [17]. Our analysis reveals that less efficient male-killers are not viable where highly efficient male-killers can persist (electronic supplementary material, appendix S3).…”

“…Second, insects are known to be infected by close relatives of MK bacteria, which do not cause MK per se. For example, Kageyama et al [17] report Spiroplasma infection not causing MK to be prevalent (23-66%) in Japanese populations of Drosophila hydei. The observed spiroplasma is a close relative to MK spiroplasmas from other Drosophila species and, at the same time, the most basal taxon of the clade.…”