DAVIDSON, S.K.; KOROPATNICK, T.; KOSSMEHL, R.; MCFALL-NGAI, M.J.: Nitric oxide as a specificity determinant in the squid-Vibrio association

During colonization of the Euprymna scolopes light organ, the symbiont Vibrio fischeri aggregates in mucus that is secreted by cells of a complex ciliated field on the surface of the nascent light organ. Once aggregated, the symbionts migrate through ducts into epithelia-lined crypts where the symbionts reside throughout the life history of the host. In this study, we provide evidence that nitric oxide (NO) acts as a specificity determinant during the colonization process. Nitric oxide synthase was detected by NADPH-diaphorase staining and immunocytochemistry in high concentration in both the cells of the ciliated field and in the cells that line the ducts. The location and levels of NO production were monitored by confocal microscopy with the diaminofluorescein fluorochrome DAF-FM, which fluoresces in the presence of NO. These analyses revealed that NO was present not only in the ciliated field of cells and the ducts, but also in vesicles within the symbiont-containing mucus aggregates. In the presence of NO scavengers, nonsymbiotic vibrio species formed unusually large aggregates outside of the light organ. In addition, the bacterial symbionts irreversibly attenuated NO production in the cells of the ciliated field and ducts within 12-14 h following crypt colonization. Taken together, these data suggest that nitric oxide plays a role in the specific establishment of the association between E. scolopes and V. fischeri, and that the symbionts influence host production of NO. Supported by NSF IBN 9904601 and NIH RR12294 to MM-N and NIH NRSA GM20177 to SKD.