High and low temperatures differently affect infection density and vertical transmission of male-killing Spiroplasma symbionts in Drosophila hosts

Abstract

We investigated the vertical transmission, reproductive phenotype, and infection density of a male-killing Spiroplasma symbiont in two Drosophila species under physiological high and low temperatures through successive host generations. In both the native host Drosophila nebulosa and the nonnative host Drosophila melanogaster, the symbiont infection and the male-killing phenotype were stably maintained at 25 degrees C, rapidly lost at 18 degrees C, and gradually lost at 28 degrees C. In the nonnative host, both the high and low temperatures significantly suppressed the infection density of the spiroplasma. In the native host, by contrast, the low temperature suppressed the infection density of the spiroplasma whereas the high temperature had little effect on the infection density. These results suggested that the low temperature suppresses both the infection density and the vertical transmission of the spiroplasma whereas the high temperature suppresses the vertical transmission preferentially. The spiroplasma density was consistently higher in the native host than in the nonnative host, suggesting that the host genotype may affect the infection density of the symbiont. The temperature- and genotype-dependent instability of the symbiont infection highlights a complex genotype-by-genotype-by-environment interaction and may be relevant to the low infection frequencies of the male-killing spiroplasmas in natural Drosophila populations.

FIG. 1.

Phylogenetic relationships of the spiroplasmas associated with Drosophila species, inferred from the sequences of the 16S rRNA gene (1,429 aligned nucleotide sites) (A), the p58 gene (798 sites) (B), the spoT gene (513 sites) (C), and the dnaA gene (474 sites) (D). Neighbor-joining trees are shown, while maximum-parsimony trees and maximum-likelihood trees exhibit almost the same topologies. Bootstrap values higher than 50% are indicated on the nodes. Sequence accession numbers are in brackets, and host organism names are in parentheses.

FIG. 2.

Infection frequencies of the spiroplasma in the nonnative host D. melanogaster strain ORNSRO and the native host D. nebulosa strain G87 maintained under different temperature conditions. The infection frequency was calculated as 100 × (number of infected females)/(number of all females examined) (%).

FIG. 3.

Intensity of male-killing phenotype caused by the spiroplasma in the nonnative host D. melanogaster strain ORNSRO and the native host D. nebulosa strain G87 maintained under different temperature conditions. When the number of males was less than or the same as that of females, the intensity of male killing was formulated as 100 × [(number of females) − (number of males)]/(number of females) (%), which ranges from 0% for a 1:1 sex ratio to 100% for all females. When the number of males was more than that of females, the intensity of male killing was regarded as 0%.

FIG. 4.

Infection density of the male-killing spiroplasma in the D. melanogaster strain ORNSRO (A) and the D. nebulosa strain G87 (B) maintained under different temperature conditions. Relative spiroplasma densities are indicated in terms of symbiont dnaA gene copies per μg (dry weight) of host. The medians of 8 to 20 measurements are shown for generations 1, 2, 3, and 8. Sample sizes are labeled on the columns. Error bars show the interquartile ranges. Asterisks indicate statistically significant differences among temperature treatments in the same generation (asterisks above the columns) or between the strains ORNSRO and G87 at the same temperature in the same generation (asterisks below the columns of upper graphs) after adjustment by the standard Bonferroni method (generalized linear models; *, P < 0.05; **, P < 0.01; ***, P < 0.001). “N.S.” indicates no significant difference.

FIG. 5.

Relationship between infection density and infection frequency of the spiroplasma in the D. melanogaster strain ORNSRO and the D. nebulosa strain G87 maintained under different temperature conditions. Sample sizes are shown on the plots in parentheses.