Rrh: Wolbachia Superinfection in Aedes Albopictus Title: Origin of Wolbachia Superinfection in Aedes Albopictus by Sequential Population Replacement

A reproductive advantage afforded to female hosts by obligate intracellular Wolbachia infections can result in the spread of the vertically-inherited bacteria into the host population, replacing the uninfected cytotype (population replacement). Here we examine Wolbachia infection dynamics relevant to the origin of Wolbachia superinfection (i.e., host individuals that are co-infected with two or more Wolbachia types). Possible routes of origin for Wolbachia superinfection are: 1) the simultaneous invasion of an uninfected host population by the superinfected cytotype, or 2) the sequential invasion of an uninfected host population by a single-infection followed by the subsequent superinfection invasion. Relative to uninfected females, superinfected Aedes albopictus females are at a reproductive advantage due to both cytoplasmic incompatibility (CI) and a female fitness increase associated with Wolbachia infection. A host fitness increase combined with CI is predicted to reduce the threshold Wolbachia infection frequency required for population replacement. However, this prediction applies only to the simultaneous invasion hypothesis. To better understand events associated with sequential invasion, we have examined cytoplasmic incompatibility levels, host longevity, egg hatch rates and fecundity in introgressed A. albopictus strains that are uninfected, single-infected, and superinfected with Wolbachia. Relative to uninfected females, infected females are at a reproductive advantage due to both cytoplasmic incompatibility and a fitness increase associated with Wolbachia infection. In contrast, no fitness advantage was observed in comparisons of singleand superinfected females. We discuss the observed results in regard to the proposed origin of Wolbachia superinfection in A. albopictus via sequential population replacement. The hypothesized evolution of Wolbachia superinfection in A. albopictus by mutation and horizontal transmission is also discussed. Comparisons of similarly-infected A. albopictus strains that differ in host genotype demonstrate host fitness varies with host genotpe, but that cytoplasmic incompatibility and Wolbachia-induced host fitness effects are consistent regardless of the host genotype.