Living in the endosymbiotic world of Wolbachia: A centennial review

Abstract

The most widespread intracellular bacteria in the animal kingdom are maternally inherited endosymbionts of the genus Wolbachia. Their prevalence in arthropods and nematodes worldwide and stunning arsenal of parasitic and mutualistic adaptations make these bacteria a biological archetype for basic studies of symbiosis and applied outcomes for curbing human and agricultural diseases. Here, we conduct a summative, centennial analysis of living in the Wolbachia world. We synthesize literature on Wolbachia's host range, phylogenetic diversity, genomics, cell biology, and applications to filarial, arboviral, and agricultural diseases. We also review the mobilome of Wolbachia including phage WO and its essentiality to hallmark reproductive phenotypes in arthropods. Finally, the Wolbachia system is an exemplar for discovery-based science education using biodiversity, biotechnology, and bioinformatics lessons. As we approach a century of Wolbachia research, the interdisciplinary science of this symbiosis stands as a model for consolidating and teaching the integrative rules of endosymbiotic life.

Keywords: Wolbachia; cytoplasmic incompatibility; evolution; feminization; male killing; parthenogenesis; phage WO; vector control.

Figure 1.. Phylogeny and evolution of Wolbachia.

Schematic representation of a) phylogenetic relationships of Wolbachia and members of the family Anaplasmataceae, with Rickettsia shown as an outgroup, and b) an unrooted 16S rRNA-based consensus phylogenetic tree of the major and well-established Wolbachia supergroups. The phylogenetic positions of the supergroups are currently tentative based on previously-published single gene and multigene analyses. Colors correspond to different patterns of host-Wolbachia associations across the supergroups. Supergroup E is multicolor labeled since the evidence for Wolbachia-associated phenotypes in the host are suggestive but inconclusive. A phage symbol represents Wolbachia supergroups with genomic evidence of intact or relic prophage WO presence. Specific examples of phage-association and reproductive phenotypes are listed in supplemental table 2.

Figure 2.. The tripartite symbiosis of bacteriophage WO, Wolbachia, and arthropods.

a) Schematic representation of two phage WO life cycles – lytic and lysogenic - in Wolbachia cells (blue and orange) living in endosymbiosis within an arthropod host cell. During the lytic cycle, phage WO particles are produced and can exit and enter Wolbachia cells to establish new infections, while in the lysogenic cycle, prophage WO is stably integrated into the Wolbachia chromosome. b) Relic prophage WO regions, such as in the phage regions of WORecB and WOAlbB, likely arose by invasion of a fully intact phage WO, chromosome integration, and erosion over time that led to loss of structural genes and the ability to form phage particles. In a process termed domestication, Wolbachia can retain selected phage-associated genes for adaptive functions such as cytoplasmic incompatibility encoded by the cifA and cifB genes and male killing encoded by the candidate gene wmk. c) Genomic maps of the newly-discovered pWCP plasmid in the wPip Wolbachia strain of mosquitoes and the wMel chromosome from flies with its two integrated prophages, WOMelA and WOMelB. Prophage regions are highlighted in light gray on the Wolbachia chromosome, phage structural genes are in dark gray, and key reproductive parasitism genes, cifA;cifB (pink) and wmk (blue), are highlighted in their respective colors within the eukaryotic association module (EAM). pWCP and phage WO are absent in the nematode Wolbachia.

Figure 3.. Wolbachia tissue tropism in arthropods and nematodes.

a) Somatic and reproductive tissues with Wolbachia are labeled in the arthropod host Drosophila and nematode host Brugia malayi. b) During Drosophila oogenesis, Wolbachia (red) are present in germline stem cells where cell differentiation gives rise to a Wolbachia-infected egg chamber composed of an oocyte and nurse cells that interconnect by ring canals. During early oogenesis, Wolbachia utilize microtubules to move into the oocyte. They localize to the posterior pole during mid oogenesis and remain throughout development in the mature egg. c) During Drosophila spermatogenesis, Wolbachia are present in the germline stem cells, which divide mitotically to give rise to 16 interconnected spermatocytes with unevenly distributed Wolbachia. The spermatocytes then undergo meiosis to create a 64-cell cyst of interconnected spermatids that undergo differentiation and individualization. During individualization, excess cytoplasmic components including Wolbachia are removed from the mature sperm into a cytoplasmic waste bag.

Figure 4.. Reproductive phenotypes in arthropods.

Wolbachia induce four, well-established reproductive parasitism phenotypes that assist its spread in a range of arthropod hosts. a) Cytoplasmic incompatibility (CI) causes embryonic death in crosses between infected males and uninfected females. In D. melanogaster males, expression of Wolbachia (red dots) prophage WO genes, cytoplasmic incompatibility factors (cif) cifA and cifB causes CI and results in embryonic death. Female expression of cifA in Wolbachia-infected eggs rescues the CI phenotype, leading to normal development of embryos. b) Male killing results in a female-biased sex ratio in various arthropods by selectively killing males. In D. melanogaster, transgenic expression of candidate gene wmk in the embryos causes partial, male-specific embryonic lethality. c) Parthenogenesis causes virgin mothers to produce all female offspring from their unfertilized eggs. In Trichogramma wasps, Wolbachia in the unfertilized eggs double the haploid set of maternal chromosomes, causing them to develop into diploid females. d) Feminization results in genetic males that phenotypically develop as females. The molecular mechanism of feminization is unknown; however, on an evolutionary scale, Wolbachia inserted a fragment of their DNA called ‘f-element’ into the pillbug host genome that effectively results in the evolution of a new female-sex determining chromosome.

Figure 5.. Wolbachia-mediated applications to control insect vectors.

a) Population replacement strategy commences with release of both male and female mosquitoes where CI-inducing Wolbachia spread throughout uninfected target populations, thus replacing the native species with pathogen-resistant, Wolbachia-infected mosquitoes, no longer capable of transmitting disease. b) Incompatible insect technique entails release of CI-causing Wolbachia-infected male mosquitoes that do not produce viable embryos after mating with wild-type uninfected females, thus reducing the total number of disease-transmitting mosquitoes in natural populations. Figure is adapted from Yen and Failloux, 2020.