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      Coxiella-Like Endosymbiont of Rhipicephalus sanguineus Is Required for Physiological Processes During Ontogeny

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          Abstract

          Obligatory hematophagous arthropods such as lice, bugs, flies, and ticks harbor bacterial endosymbionts that are expected to complement missing essential nutrients in their diet. Genomic and some experimental evidence support this expectation. Hard ticks (Acari: Ixodidae) are associated with several lineages of bacterial symbionts, and very few were experimentally shown to be essential to some aspects of tick’s fitness. In order to pinpoint the nature of interactions between hard ticks and their symbionts, we tested the effect of massive elimination of Coxiella-like endosymbionts (CLE) by antibiotics on the development and fitness of the brown dog tick ( Rhipicephalus sanguineus). Administration of ofloxacin to engorged (blood fed) nymphs resulted in significant and acute reduction of their CLE loads – an effect that also persisted in subsequent life stages (aposymbiotic ticks). As a result, the post-feeding development of aposymbiotic female (but not male) nymphs was delayed. Additionally, aposymbiotic adult females needed a significantly prolonged feeding period in order to replete (detach from host), and had reduced engorgement weight and a lower capacity to produce eggs. Consequently, their fecundity and fertility were significantly reduced. Eggs produced by aposymbiotic females were free of CLE, and the resulting aposymbiotic larvae were unable to feed successfully. Our findings demonstrate that the observed fitness effects are due to CLE reduction and not due to antibiotic administration. Additionally, we suggest that the contribution of CLE is not mandatory for oocyte development and embryogenesis, but is required during feeding in females, when blood meal processing and tissue buildup are taking place. Presumably, under these extreme physiological demands, CLE contribute to R. sanguineus through supplementing essential micro- and macronutrients. Further nutrient complementary studies are required to support this hypothesis.

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          Wolbachia as a bacteriocyte-associated nutritional mutualist.

          Many insects are dependent on bacterial symbionts that provide essential nutrients (ex. aphid-Buchnera and tsetse-Wiglesworthia associations), wherein the symbionts are harbored in specific cells called bacteriocytes that constitute a symbiotic organ bacteriome. Facultative and parasitic bacterial symbionts like Wolbachia have been regarded as evolutionarily distinct from such obligate nutritional mutualists. However, we discovered that, in the bedbug Cimex lectularius, Wolbachia resides in a bacteriome and appears to be an obligate nutritional mutualist. Two bacterial symbionts, a Wolbachia strain and an unnamed gamma-proteobacterium, were identified from different strains of the bedbug. The Wolbachia symbiont was detected from all of the insects examined whereas the gamma-proteobacterium was found in a part of them. The Wolbachia symbiont was specifically localized in the bacteriomes and vertically transmitted via the somatic stem cell niche of germalia to oocytes, infecting the incipient symbiotic organ at an early stage of the embryogenesis. Elimination of the Wolbachia symbiont resulted in retarded growth and sterility of the host insect. These deficiencies were rescued by oral supplementation of B vitamins, confirming the essential nutritional role of the symbiont for the host. The estimated genome size of the Wolbachia symbiont was around 1.3 Mb, which was almost equivalent to the genome sizes of parasitic Wolbachia strains of other insects. These results indicate that bacteriocyte-associated nutritional mutualism can evolve from facultative and prevalent microbial associates like Wolbachia, highlighting a previously unknown aspect of the parasitism-mutualism evolutionary continuum.
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            Reproductive Effort and Terminal Investment in Iteroparous Animals

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              The Recent Evolution of a Maternally-Inherited Endosymbiont of Ticks Led to the Emergence of the Q Fever Pathogen, Coxiella burnetii

              Q fever is a highly infectious disease with a worldwide distribution. Its causative agent, the intracellular bacterium Coxiella burnetii, infects a variety of vertebrate species, including humans. Its evolutionary origin remains almost entirely unknown and uncertainty persists regarding the identity and lifestyle of its ancestors. A few tick species were recently found to harbor maternally-inherited Coxiella-like organisms engaged in symbiotic interactions, but their relationships to the Q fever pathogen remain unclear. Here, we extensively sampled ticks, identifying new and atypical Coxiella strains from 40 of 58 examined species, and used this data to infer the evolutionary processes leading to the emergence of C. burnetii. Phylogenetic analyses of multi-locus typing and whole-genome sequencing data revealed that Coxiella-like organisms represent an ancient and monophyletic group allied to ticks. Remarkably, all known C. burnetii strains originate within this group and are the descendants of a Coxiella-like progenitor hosted by ticks. Using both colony-reared and field-collected gravid females, we further establish the presence of highly efficient maternal transmission of these Coxiella-like organisms in four examined tick species, a pattern coherent with an endosymbiotic lifestyle. Our laboratory culture assays also showed that these Coxiella-like organisms were not amenable to culture in the vertebrate cell environment, suggesting different metabolic requirements compared to C. burnetii. Altogether, this corpus of data demonstrates that C. burnetii recently evolved from an inherited symbiont of ticks which succeeded in infecting vertebrate cells, likely by the acquisition of novel virulence factors.
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                Author and article information

                Contributors
                Journal
                Front Microbiol
                Front Microbiol
                Front. Microbiol.
                Frontiers in Microbiology
                Frontiers Media S.A.
                1664-302X
                22 April 2020
                2020
                : 11
                : 493
                Affiliations
                [1] 1Koret School of Veterinary Medicine, The Robert H. Smith Faculty of Agriculture, Food and Environment, The Hebrew University of Jerusalem , Rehovot, Israel
                [2] 2Kimron Veterinary Institute , Bet Dagan, Israel
                Author notes

                Edited by: Christoph Vorburger, Swiss Federal Institute of Aquatic Science and Technology, Switzerland

                Reviewed by: Joseph James Gillespie, University of Maryland, Baltimore, United States; Alejandro Manzano Marín, University of Vienna, Austria

                *Correspondence: Yuval Gottlieb, gottlieb.yuval@ 123456mail.huji.ac.il

                This article was submitted to Microbial Symbioses, a section of the journal Frontiers in Microbiology

                Article
                10.3389/fmicb.2020.00493
                7188774
                32390951
                0c796dad-d124-4d0c-acad-5e46362604cb
                Copyright © 2020 Ben-Yosef, Rot, Mahagna, Kapri, Behar and Gottlieb.

                This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

                History
                : 01 December 2019
                : 06 March 2020
                Page count
                Figures: 6, Tables: 0, Equations: 0, References: 45, Pages: 16, Words: 0
                Funding
                Funded by: Israel Science Foundation 10.13039/501100003977
                Funded by: United States - Israel Binational Science Foundation 10.13039/100006221
                Categories
                Microbiology
                Original Research

                Microbiology & Virology
                ticks,arthropod symbiosis,hematophagy,antibiotic treatment,reproductive fitness

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