FluorescenceIn SituHybridization Provides Evidence for the Presence of the Endosymbiotic Bacterial GenusRickettsiain Tardigrades

Many ecdysozoans harbor endosymbiotic bacteria within their microbiota, and these endosymbionts can have a range of positive and negative effects on their hosts. Recent 16S rRNA gene amplicon sequencing studies have provided evidence for endosymbionts within the tardigrade microbiota. In a previous study amplicon study, we determined that sequences corresponding to the endosymbiotic genusRickettsiawere significantly more associated with tardigrades than with the substrate from which they were isolated. Here, we performed fluorescencein situhybridization (FISH) using aRickettsia-specific probe, RickB1, to determine ifRickettsiacould be found in tardigrades. RickB1 and a probe targeting most bacteria, EUB338, colocalized within the tissues of tardigrades, indicating the presence ofRickettsia. We also performed FISH using RickB1 and a nonsense probe which allowed us to distinguish between false positives and true positives. This method revealed RickB1 signals in tardigrades that were not due to erroneous probe binding, providing further evidence thatRickettsiais present in tardigrades. Future research will be necessary to determine the effects, if any, of this endosymbiont on its tardigrade host.