Distinct effects of the nephridial symbionts Verminephrobacter and Candidatus Nephrothrix on reproduction and maturation of its earthworm host Eisenia andrei.

Verminephrobacter, the most common specific symbionts in the nephridia (excretory organs) of lumbricid earthworms, have been shown to improve reproduction of the garden earthworm Aporrectodea tuberculata under nutrient limitation. It is unknown how general this beneficial trait is in the Verminephrobacter-earthworm symbiosis, whether other nephridial symbionts also affect host fitness and what the mechanism of the fitness increase is. Here we report beneficial effects of Verminephrobacter and Candidatus Nephrothrix on life history traits of the compost worm Eisenia andrei, which in addition to these two symbionts also hosts Agromyces-like bacteria in its mixed nephridial community: while growth was identical between control, Verminephrobacter-free and aposymbiotic worms, control worms produced significantly more cocoons and offspring than both Verminephrobacter-free and aposymbiotic worms, confirming the reproductive benefit of Verminephrobacter in a second host with different ecology and feeding behavior. Furthermore, worms with Verminephrobacter and Ca. Nephrothrix, or with only Ca. Nephrothrix present, reached sexual maturity significantly earlier than aposymbiotic worms; this is the first evidence for a beneficial role of Ca. Nephrothrix in earthworms. Riboflavin content in cocoons and whole earthworms was unaffected by the presence or absence of nephridial symbionts, suggesting that nutritional supplementation with this vitamin does not play a major role in this symbiosis.

Biparental transmission of Verminephrobacter symbionts in the earthworm Aporrectodea tuberculata (Lumbricidae).

Most lumbricid earthworms harbor species-specific Verminephrobacter symbionts in their excretory organs (nephridia). These symbionts are vertically transmitted via the cocoon, where they colonize the embryos. Despite cospeciation for >100 million years with their hosts, Verminephrobacter lack genome reduction and AT bias typical of evolutionary old, vertically transmitted symbionts, caused by recurring bottlenecks. We hypothesized that biparental symbiont transmission into the cocoon enabled genetic mixing and relieved the bottleneck, and tested biparental transmission experimentally for V. aporrectodeae subsp. tuberculata, the specific symbiont of the earthworm Aporrectodea tuberculata, for which aposymbiotic worm lines are available. Virgin symbiotic and aposymbiotic adult worms were tagged, mated in pairs, separated before the start of cocoon production and their offspring assessed for Verminephrobacter. Specific PCR detected the symbionts in 41.5% of 188 juveniles produced by 20 aposymbiotic worms; fluorescence in situ hybridization showed a patchy but successful colonization of their nephridia. Symbionts were present in the mucus but absent in feed, soil, and spermatophora/nephridia of the aposymbiotic partner, suggesting symbiont transfer via mucus during mating. These results are consistent with the hypothesis that genome evolution in Verminephrobacter is distinct from other vertical-ly transmitted symbionts due to genetic mixing during transmission, partially facilitated by biparental transmission.