Taxonomic classification of the bacterial endosymbiont Wolbachia based on next-generation sequencing: is there molecular evidence for its presence in tardigrades?

Abstract

We used high-throughput sequencing of 16S rRNA to test whether tardigrade species are infected with Wolbachia parasites. We applied SILVA and Greengenes databases that allowed taxonomic classification of bacterial sequences to OTUs. The results obtained from both databases differed considerably in the number of OTUs, and only the Greengenes database allowed identification of Wolbachia (infection was also supported by comparison of sequences to NCBI database). The putative bacterial endosymbiont Wolbachia was discovered only in adult eutardigrades, while bacteria identified down to the order Rickettsiales were detected in both eutardigrade eggs and adult specimens. Nevertheless, the frequency of Wolbachia in the bacterial communities of the studied eutardigrades was low. Similarly, in our positive control, i.e. a fairy shrimp Streptocephalus cafer, which was found to be infected with Wolbachia in our previous study using Sanger sequencing, only the Rickettsiales were detected. We also carried out phylogenetic reconstruction using Wolbachia sequences from the SILVA and Greengenes databases, Alphaproteobacteria putative endosymbionts and Rickettsiales OTUs obtained in the previous studies on the microbial community of tardigrades as well as Rickettsiales and Wolbachia OTUs obtained in the current study. Our discovery of Wolbachia in tardigrades can fuel new research to uncover the specifics of this interaction.

Tardigrade community microbiomes in North American orchards include putative endosymbionts and plant pathogens

“…The life cycle of tardigrades poses a unique selective pressure on any permanent residents of the microbiota, as these organisms would also have to survive within the tardigrade during cryptobiosis. This would be especially true for the obligate endosymbiotic taxa Rickettsiales and Polynucleobacter previously observed in tardigrades (Vecchi et al 2018;Roberto Guidetti et al 2020;Kaczmarek et al 2020;Mioduchowska et al 2021), as well as for the Rickettsia identified in the current study (see below).…”

“…In this study, the five most abundant phyla were Proteobacteria, Firmicutes, Bacteroidota, Actinobacteria, and Acidobacteria (Table S10). All of these except Acidobacteria were previously reported as highly abundant in at least two of the three previous tardigrade microbiome surveys that presented results at a phylum level, with Proteobacteria identified as the most abundant phylum in all cases (Vecchi et al 2018;Kaczmarek et al 2020;Mioduchowska et al 2021). Combined, the tardigrades in these studies represent a diverse set of species, including wild and laboratory-reared specimens isolated from multiple continents, suggesting that the predominance of these phyla is broadly characteristic of the microbiome of Tardigrada, regardless of species or location.…”

“…This study examined the microbiota of the full tardigrade community from a particular substrate sample, in contrast to previous surveys that studied isolated species, often from laboratory cultures (Vecchi et al 2018;Kaczmarek et al 2020;Mioduchowska et al 2021;Zawierucha et al 2022). It is of course desirable to identify species-specific microbiota, as Vecchi et al (2018) found that tardigrade-associated bacteria varied among tardigrade species.…”

“…All tardigrade microbiome surveys, including the current study, employed laboratory technique to reduce contamination by washing the tardigrades in sterile water before DNA extraction (Vecchi et al 2018;Mioduchowska et al 2021;Kaczmarek et al 2020;Zawierucha et al 2022). Working in a sterile environment further decreases contamination; therefore,…”

“…A third paper conducted 16S rRNA amplicon sequencing on four tardigrade species: (Mioduchowska et al 2021).…”