# Lock and Key: Why Rickettsia Endosymbionts Do Not Harm Vertebrate Hosts?

Alejandro Cabezas-Cruz, Andréa Cristina Fogaça

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Alejandro Cabezas-Cruz, Andréa Cristina Fogaça. Lock and Key: Why Rickettsia Endosymbionts Do Not Harm Vertebrate Hosts?. Pathogens, 2022, 11 (12), pp.1494. 10.3390/pathogens11121494. hal-04009589

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# Lock and Key: Why *Rickettsia* Endosymbionts Do Not Harm Vertebrate Hosts?

Alejandro Cabezas-Cruz <fnref n="1" />,*,† ID and Andréa Cristina Fogaça <fnref n="2" />,*,†

Are tick endosymbionts transmitted to and able to injure vertebrate hosts? This is a long-standing question concerning both infectiology and microbiology, as microbes (parasites, mutualists, or commensals) shift back and forth from parasitic to mutualistic lifestyles in an evolutionary process dubbed “transitions along the parasite–mutualist continuum” by Drew et al. [1]. Tick-associated microbes are not an exception to this phenomenon [2]. Notable examples include the evolutionary transitions that led a pathogenic strain of *Francisella* to evolve into a *Francisella*-like endosymbiont in the Gulf Coast tick *Amblyomma maculatum* [3], or that caused a maternally inherited *Coxiella*-like endosymbiont to become the Q fever pathogen, *Coxiella burnetii* [4].

The changes in microbial lifestyle involve genomic rearrangements, for instance, size alterations, as indicated for *Coxiella* [4], *Francisella* [3], and *Rickettsia* [5–7], among other tick-borne microbes [8]. In general, during their evolutionary transitions from a parasitic to a mutualistic lifestyle, bacterial genomes undergo size reduction [5] and loss in coding capacity [3] but retain their extensive metabolic capabilities [3,4,9]. The latter is an essential trait defining entrance into the “evolutionary rabbit hole” of heritable symbiosis [10]. Although carrying commensal/mutualist microorganisms may impose an energetic burden on the host [10], they can also benefit the host, for instance, by producing essential vitamins and conferring resistance to tick-borne pathogens [2].

It was previously reported that the tick intra-mitochondrial symbiont *Midichloria* is present in both tick salivary glands and saliva [11]. Remarkably, bacterial DNA and anti-*Midichloria* antibodies were detected in vertebrate hosts after they were infested by ticks [12], showing that tick endosymbionts can be transmitted to vertebrate hosts. Less clear, however, are some other questions, namely can tick endosymbionts cause disease in vertebrate hosts? What are the lock and key mechanisms to bacteria–host cell interactions that enable a tick-borne microbe to cause disease? A research paper that tackles these questions was published by Kristof et al. [13].

In their work, Kristof et al. [13] remarkably showed that certain bacteria in the *Rickettsia* genus recognized as human pathogens (i.e., *R. rickettsii* *R. parkeri*, *R. africana*, and *R. akari*) can grow within both endothelial and differentiated THP-1 cells (a human leukemia monocytic cell line used as a human macrophage model). In contrast to pathogenic rickettsiae, *Rickettsia bellii*, a tick endosymbiont [6], and the avirulent Iowa strain of *R. rickettsii* can invade both cell types but proliferate only within endothelial cells. They further found that pathogenic rickettsiae grow within THP-1 cytosol but are not localized in the lysosomal compartments, while *R. bellii* was found to be associated with lysosomal compartments. Macrophage xenophagy, the process of the selective capture and degradation of intracellular bacteria by lysosomes, is an essential mechanism of innate immunity to kill and clear infectious bacterial agents [14]. In addition, cell surface recognition and

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**Citation:** Cabezas-Cruz, A.; Fogaça, A.C. Lock and Key: Why *Rickettsia* Endosymbionts Do Not Harm Vertebrate Hosts? *Pathogens* **2022**, *11*, 1494. https://doi.org/10.3390/pathogens11121494

Received: 22 November 2022

Accepted: 5 December 2022

Published: 8 December 2022

**Publisher's Note:** MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

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**Copyright:** © 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).

<footnote n="1">1 ANSES, INRAE, Ecole Nationale Vétérinaire d'Alfort, UMR BIPAR, Laboratoire de Santé Animale, F-94700 Maisons-Alfort, France</footnote>

<footnote n="2">2 Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo, São Paulo 05508-000, SP, Brazil</footnote>

<footnote>* Correspondence: alejandro.cabezas@vet-alfort.fr (A.C.-C.); deafog@usp.br (A.C.F.)</footnote>

<footnote>† These authors contributed equally to this work.</footnote>
the cytosolic sensing of pathogen-associated molecular patterns (PAMPs) and pathogen-
induced damage-associated molecular patterns (DAMPs) result in signaling cascades that
promote autophagosome formation and subsequent bacterial targeting for lysosome degra-
dation [14]. In this context, these findings by Kristof et al. [13] are relevant to understand
the evolutionary transitions within *Rickettsia*.

The reconstruction of the evolutionary history of *Rickettsia* using genomic comparisons suggested that its ancestor initiated intracellular parasitism in unicellular eukaryotic-like amoebae [5], a trait still conserved in *R. bellii* [6]. Later, it adapted to multicellular eukaryotes [7], first as arthropod symbionts and subsequently as a vector-borne pathogen that affects vertebrates. *Rickettsia bellii* is known to be (i) the earliest diverging species of bacteria in the genus *Rickettsia*, (ii) the most common rickettsia found in ticks in the American continent, (iii) a transovarially transmitted bacterium, and (iv) the sole rickettsia found in both soft and hard ticks [6]. The results by Kristof et al. [13] suggest that the tick endosymbiont *R. bellii* may lack or have lost or not acquired one of the essential features of its pathogenic relatives: the mastery to hijack and manipulate the machinery of defensive cells in their vertebrate hosts [15]. Intriguingly, *R. bellii* is able to cause disease in vertebrates, as experimentally proven in guinea pigs [6,16] and rabbits [6], suggesting that it is capable of somehow crossing this apparent “barrier” to infection. Indeed, the intradermal injection of low infectious doses of *R. bellii* is sufficient to cause inflammatory reactions, while high infectious doses induce a black necrotic eschar [6], which is a typical clinical sign of rickettsial pathogenicity [17]. However, notably, the severity of the disease triggered by presumed endosymbionts, including *R. bellii*, but also *Rickettsia amblyommatis* and *Rickettsia montanensis*, is significantly milder than those of recognized rickettsial pathogens such as *R. rickettsii* [16]. Other findings suggested the natural infection of dogs by *R. bellii* [18] and humans by *R. montanensis* and *R. amblyommatis* [19].

It was previously shown that the highly lethal *R. rickettsii* is capable of inhibiting apoptosis in host endothelial cells [20–22] and tick cells [23]. Indeed, apoptosis inhibition promotes rickettsial growth in tick cells [23]. On the other hand, *R. parkeri*, the causative agent of a mild form of spotted fever, activates apoptosis in tick cells, which also seems to favor infection [24]. Hence, additional comparative studies on the molecular interactions among bacteria in the genus *Rickettsia* and their host/vector cells, such as that reported by Kristof et al. [13], are warranted and may reveal further mechanisms that define the pathogenicity level of rickettsiae. Regardless of the answers to all of the questions presented above, *Rickettsia* spp., independent of their current status in the transitions along “the parasite–mutualist continuum”, should be considered as potential emerging pathogens posing a potential risk to human and animal health.

**Funding:** UMR BIPAR is supported by the French Government’s Investissement d’Avenir program, Laboratoire d’Excellence “Integrative Biology of Emerging Infectious Diseases” (grant no. ANR-10-LABX-62-IBEID).

**Conflicts of Interest:** The authors declare no conflict of interest.

**References**

1. Drew, G.C.; Stevens, E.J.; King, K.C. Microbial evolution and transitions along the parasite-mutualist continuum. *Nat. Rev. Microbiol.* **2021**, *19*, 623–638. [CrossRef] [PubMed]

2. Bonnet, S.I.; Binetruy, F.; Hernández-Jarguín, A.M.; Duron, O. The Tick Microbiome: Why Non-pathogenic Microorganisms Matter in Tick Biology and Pathogen Transmission. *Front. Cell. Infect. Microbiol.* **2017**, *7*, 236. [CrossRef] [PubMed]

3. Gerhart, J.G.; Moses, A.S.; Raghavan, R. A *Francisella*-like endosymbiont in the Gulf Coast tick evolved from a mammalian pathogen. *Sci. Rep.* **2016**, *6*, 33670. [CrossRef] [PubMed]

4. Duron, O.; Noël, V.; McCoy, K.D.; Bonazzi, M.; Sidi-Boumedine, K.; Morel, O.; Vavre, F.; Zenner, L.; Jourdain, E.; Durand, P.; et al. The Recent Evolution of a Maternally-Inherited Endosymbiont of Ticks Led to the Emergence of the Q Fever Pathogen, *Coxiella burnetii*. *PLoS Pathog.* **2015**, *11*, e1004892. [CrossRef]

5. Blanc, G.; Ogata, H.; Robert, C.; Audic, S.; Suhre, K.; Vestris, G.; Claverie, J.M.; Raoult, D. Reductive genome evolution from the mother of *Rickettsia*. *PLoS Genet.* **2007**, *3*, e14. [CrossRef]
6. Ogata, H.; La Scola, B.; Audic, S.; Renesto, P.; Blanc, G.; Robert, C.; Fournier, P.E.; Claverie, J.M.; Raoul, D. Genome sequence of *Rickettsia bellii* illuminates the role of amoebae in gene exchanges between intracellular pathogens. *PLoS Genet.* **2006**, *2*, e76. [CrossRef] [PubMed]

7. El Karkouri, K.; Ghigo, E.; Raoult, D.; Fournier, P.E. Genomic evolution and adaptation of arthropod-associated *Rickettsia*. *Sci. Rep.* **2022**, *12*, 3807. [CrossRef]

8. Hotopp, J.C.D.; Lin, M.; Madupu, R.; Crabtree, J.; Angiuoli, S.V.; Eisen, J.; Seshadri, R.; Ren, Q.; Wu, M.; Utterback, T.R.; et al. Comparative genomics of emerging human ehrlichiosis agents. *PLoS Genet.* **2006**, *2*, e21.

9. Duron, O.; Morel, O.; Noël, V.; Buysse, M.; Binetruy, F.; Lancelot, R.; Loire, E.; Ménard, C.; Bouchez, O.; Vavre, F.; et al. Tick-Bacteria Mutualism Depends on B Vitamin Synthesis Pathways. *Curr. Biol.* **2018**, *28*, 1896–1902. [CrossRef]

10. Bennett, G.M.; Moran, N.A. Heritable symbiosis: The advantages and perils of an evolutionary rabbit hole. *Proc. Natl. Acad. Sci. USA* **2015**, *112*, 10169–10176. [CrossRef]

11. Di Venere, M.; Fumagalli, M.; Cafiso, A.; De Marco, L.; Epis, S.; Plantard, O.; Bardoni, A.; Salvini, R.; Viglio, S.; Bazzocchi, C.; et al. *Ixodes ricinus* and Its Endosymbiont *Midichloria mitochondrii*: A Comparative Proteomic Analysis of Salivary Glands and Ovaries. *PLoS ONE* **2015**, *10*, e0138842. [CrossRef]

12. Bazzocchi, C.; Mariconti, M.; Sassera, D.; Rinaldi, L.; Martin, E.; Cringoli, G.; Urbanelli, S.; Genchi, C.; Bandi, C.; Epis, S. Molecular and serological evidence for the circulation of the tick symbiont *Midichloria* (Rickettsiales: Midichloriaceae) in different mammalian species. *Parasit. Vectors* **2013**, *6*, 350. [CrossRef] [PubMed]

13. Kristof, M.N.; Allen, P.E.; Yutzy, L.D.; Thibodaux, B.; Paddock, C.D.; Martinez, J.J. Significant Growth by *Rickettsia* Species within Human Macrophage-Like Cells Is a Phenotype Correlated with the Ability to Cause Disease in Mammals. *Pathogens* **2021**, *10*, 228. [CrossRef] [PubMed]

14. Bah, A.; Vergne, I. Macrophage Autophagy and Bacterial Infections. *Front. Immunol.* **2017**, *8*, 1483. [CrossRef]

15. Voss, O.H.; Rahman, M.S. *Rickettsia*-host interaction: Strategies of intracytosolic host colonization. *Pathog. Dis.* **2021**, *79*, ftab015. [CrossRef] [PubMed]

16. Snellgrove, A.N.; Krapiunaya, I.; Scott, P.; Levin, M.L. Assessment of the Pathogenicity of *Rickettsia amblyommatis*, *Rickettsia bellii*, and *Rickettsia montanensis* in a Guinea Pig Model. *Vector Borne Zoonotic Dis.* **2021**, *21*, 232–241. [CrossRef] [PubMed]

17. Jakimovski, D.; Mateska, S.; Simin, V.; Bogdan, I.; Mijatović, D.; Estrada-Peña, A.; Mateos-Hernández, L.; Foucault-Simonin, A.; Moutailler, S.; Cabezas-Cruz, A.; et al. Mediterranean spotted fever-like illness caused by *Rickettsia sibirica mongolitimonae*, North Macedonia, June 2022. *Eurosurveillance* **2022**, *27*, 2200735. [CrossRef]

18. Fortes, F.S.; Silveira, I.; Moraes-Filho, J.; Leite, R.V.; Bonacim, J.E.; Biondo, A.W.; Labruna, M.B.; Molento, M.B. Seroprevalence of *Rickettsia bellii* and *Rickettsia felis* in dogs, São José dos Pinhais, State of Paraná, Brazil. *Rev. Bras. Parasitol. Vet.* **2010**, *19*, 222–227. [CrossRef]

19. Delisle, J.; Mendell, N.L.; Stull-Lane, A.; Bloch, K.C.; Bouyer, D.H.; Moncayo, A.C. Human Infections by Multiple Spotted Fever Group Rickettsiae in Tennessee. *Am. J. Trop. Med. Hyg.* **2016**, *94*, 1212–1217. [CrossRef]

20. Clifton, D.R.; Goss, R.A.; Sahni, S.K.; Van Antwerp, D.; Baggs, R.B.; Marder, V.J.; Silverman, D.J.; Sporn, L.A. NF-kappa B-dependent inhibition of apoptosis is essential for host cell survival during *Rickettsia rickettsii* infection. *Proc. Natl. Acad. Sci. USA* **1998**, *95*, 4646–4651. [CrossRef]

21. Joshi, S.G.; Francis, C.W.; Silverman, D.J.; Sahni, S.K. Nuclear factor kappa B protects against host cell apoptosis during *Rickettsia rickettsii* infection by inhibiting activation of apical and effector caspases and maintaining mitochondrial integrity. *Infect. Immun.* **2003**, *71*, 4127–4136. [CrossRef]

22. Joshi, S.G.; Francis, C.W.; Silverman, D.J.; Sahni, S.K. NF-kappaB activation suppresses host cell apoptosis during *Rickettsia rickettsii* infection via regulatory effects on intracellular localization or levels of apoptogenic and anti-apoptotic proteins. *FEMS Microbiol. Lett.* **2004**, *234*, 333–341.

23. Martins, L.A.; Palmisano, G.; Cortez, M.; Kawahara, R.; de Freitas Balanco, J.M.; Fujita, A.; Alonso, B.I.; Barros-Battesti, D.M.; Braz, G.R.C.; Tirloni, L.; et al. The intracellular bacterium *Rickettsia rickettsii* exerts an inhibitory effect on the apoptosis of tick cells. *Parasit. Vectors* **2020**, *13*, 603. [CrossRef]

24. Wang, X.R.; Burkhardt, N.Y.; Kurtti, T.J.; Oliver, J.D.; Price, L.D.; Cull, B.; Thorpe, C.J.; Thiel, M.S.; Munderloh, U.G. Mitochondrion-Dependent Apoptosis Is Essential for *Rickettsia parkeri* Infection and Replication in Vector Cells. *mSystems* **2021**, *6*, e01209-20. [CrossRef]