Examination of the success rate of secondary symbiont manipulation by microinjection methods in the pea aphid system

Insects harbor a wide range of microbial symbionts, but their influence on host phenotypes is described in a limited number of biological models. One experimental approach to gain knowledge on the effects of symbionts to their hosts is to create insect lines with and without symbionts and examine their phenotypes. However, the success rate of symbiont elimination and introduction methods is dependent on several parameters that are scarcely tested or described. The pea aphid, Acyrthosiphon pisum Harris (Hemiptera: Aphididae), is a model insect of symbiosis studies. It harbors a primary symbiont that supplies the host with essential amino acids, and an array of secondary symbionts whose effects have been assessed by manipulating their presence/absence in the insect. Here, we describe the influence of key parameters on the success rate of symbiont manipulation using the pea aphid-secondary symbiont system. We compared two elimination methods differing in antibiotic treatment using several aphid-symbiont combinations. We also created new aphid host-symbiont combinations by secondary symbiont introduction and examined the effects of larval stage of recipient aphids on introduction success. Our study revealed that the aphid-symbiont combination has strong influence on both symbiont introduction and elimination success rates, and that the type of antibiotics and the larval stage of recipient aphids influence the elimination and introduction success rate, respectively.