Playing Peekaboo with a Master Manipulator: Metagenetic Detection and Phylogenetic Analysis of Wolbachia Supergroups in Freshwater Invertebrates

Abstract

The infamous “master manipulators”—intracellular bacteria of the genus Wolbachia—infect a broad range of phylogenetically diverse invertebrate hosts in terrestrial ecosystems. Wolbachia has an important impact on the ecology and evolution of their host with documented effects including induced parthenogenesis, male killing, feminization, and cytoplasmic incompatibility. Nonetheless, data on Wolbachia infections in non-terrestrial invertebrates are scarce. Sampling bias and methodological limitations are some of the reasons limiting the detection of these bacteria in aquatic organisms. In this study, we present a new metagenetic method for detecting the co-occurrence of different Wolbachia strains in freshwater invertebrates host species, i.e., freshwater Arthropoda (Crustacea), Mollusca (Bivalvia), and water bears (Tardigrada) by applying NGS primers designed by us and a Python script that allows the identification of Wolbachia target sequences from the microbiome communities. We also compare the results obtained using the commonly applied NGS primers and the Sanger sequencing approach. Finally, we describe three supergroups of Wolbachia: i) a new supergroup V identified in Crustacea and Bivalvia hosts; ii) supergroup A identified in Crustacea, Bivalvia, and Eutardigrada hosts, and iii) supergroup E infection in the Crustacea host microbiome community.

Morphological and genetic variability in cosmopolitan tardigrade species—Paramacrobiotus fairbanksi Schill, Förster, Dandekar & Wolf, 2010

“…Furthermore, Mioduchowska et al 87 provided molecular evidence of the presence of the bacterial endosymbiont Wolbachia based on next generation sequencing in tardigrades. Wolbachia have an effect on the evolution as well as the ecology of their hosts and have been found to cause effects including cytoplasmic incompatibility, feminization, male killing, and induced parthenogenesis 110 It has been noted that at the intraspecific level, even individuals from the same population can undergo morphological changes in their characters to diversify within niches available to the species 111 . Similarly, Kihm et al 104 proposed epigenetic factors as a main cause for variability in tardigrade Dactylobiotus ovimutans 112 Kihm, Kim, McInnes, Zawierucha, Rho, Kang & Park, 2020 egg morphology, although the population was cultured under controlled laboratory conditions.…”

A Review on the Genus Paramacrobiotus (Tardigrada) with a New Diagnostic Key

“…Indeed, many metazoan species appear to harbour their own specific microbiome community [48]. However, our understanding of the microbiome composition of Tardigrada, based on next-generation sequencing (NGS) methods targeting the standard 16S rRNA bacterial barcoding gene fragment, is limited to a very small number of published articles [84][85][86][87][88][89][90].…”

“…The genus Wolbachia is an intracellular bacterium belonging to the order Rickettsiales, and it infects various invertebrates, particularly terrestrial insects [92]. However, recent studies have identified infections of this bacterial endosymbiont in various freshwater invertebrate species [90,93,94]. Generally, this bacterium is transmitted vertically from mother to offspring and/or through horizontal transfer directly from the environment or between different hosts [95].…”

“…In 2023, Mioduchowska et al [90] described new molecular and bioinformatic tools for detecting Wolbachia in freshwater invertebrates. In this study, Wolbachia was detected in Pam.…”

Protecting Complex Biodiversity Systems: Ciliophora Endosymbionts of the Threatened Freshwater Mussel Species Unio crassus

“…They can harbor endosymbionts which role is not recognized yet, but potentially with large impacts on hosts, like bacteria (e.g. Wolbachia - [20]) or for microorganisms which may have negative effects on their progeny (like glochidia eating ciliate - [21]). This is a rich community of organisms adapted to within-shell ecosystem of freshwater mussels, which diversity and relation to the host and other organisms is not recognized yet.…”