The insect endosymbiont Sodalis glossinidius utilizes a type III secretion system for cell invasion.
Journal: 2001/April - Proceedings of the National Academy of Sciences of the United States of America
ISSN: 0027-8424
Abstract:
Sodalis glossinidius is a maternally transmitted secondary endosymbiont residing intracellularly in tissues of the tsetse flies, Glossina spp. In this study, we have used Tn5 mutagenesis and a negative selection procedure to derive a S. glossinidius mutant that is incapable of invading insect cells in vitro and is aposymbiotic when microinjected into tsetse. This mutant strain harbors Tn5 integrated into a chromosomal gene sharing high sequence identity with a type III secretion system invasion gene (invC) previously identified in Salmonella enterica. With the use of degenerate PCR, we have amplified a further six Sodalis inv/spa genes sharing high sequence identity with type III secretion system genes encoded by Salmonella pathogenicity island 1. Phylogenetic reconstructions based on the inv/spa genes of Sodalis and other members of the family Enterobacteriaceae have consistently identified a well-supported clade containing Sodalis and the enteric pathogens Shigella and Salmonella. These results suggest that Sodalis may have evolved from an ancestor with a parasitic intracellular lifestyle, possibly a latter-day entomopathogen. These observations lend credence to a hypothesis suggesting that vertically transmitted mutualistic endosymbionts evolve from horizontally transmitted parasites through a parasitism-mutualism continuum.
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Proc Natl Acad Sci U S A 98(4): 1883-1888

The insect endosymbiont <em>Sodalis glossinidius</em> utilizes a type III secretion system for cell invasion

Sir Alexander Robertson Center for Tropical Veterinary Medicine, Royal Dick School of Veterinary Studies, University of Edinburgh, Easter Bush, Roslin, Midlothian EH25 9RG, Scotland, United Kingdom
To whom reprint requests should be addressed. E-mail: ku.ca.de@eladc.
Edited by Margaret G. Kidwell, University of Arizona, Tucson, AZ, and approved November 16, 2000
Edited by Margaret G. Kidwell, University of Arizona, Tucson, AZ, and approved November 16, 2000
Received 2000 Sep 20

Abstract

Sodalis glossinidius is a maternally transmitted secondary endosymbiont residing intracellularly in tissues of the tsetse flies, Glossina spp. In this study, we have used Tn5 mutagenesis and a negative selection procedure to derive a S. glossinidius mutant that is incapable of invading insect cells in vitro and is aposymbiotic when microinjected into tsetse. This mutant strain harbors Tn5 integrated into a chromosomal gene sharing high sequence identity with a type III secretion system invasion gene (invC) previously identified in Salmonella enterica. With the use of degenerate PCR, we have amplified a further six Sodalis inv/spa genes sharing high sequence identity with type III secretion system genes encoded by Salmonella pathogenicity island 1. Phylogenetic reconstructions based on the inv/spa genes of Sodalis and other members of the family Enterobacteriaceae have consistently identified a well-supported clade containing Sodalis and the enteric pathogens Shigella and Salmonella. These results suggest that Sodalis may have evolved from an ancestor with a parasitic intracellular lifestyle, possibly a latter-day entomopathogen. These observations lend credence to a hypothesis suggesting that vertically transmitted mutualistic endosymbionts evolve from horizontally transmitted parasites through a parasitism–mutualism continuum.

Abstract

Although parasitism and mutualism may have radically different implications for host fitness, endosymbiotic bacteria participating in these relationships are known to share many similarities, including an intracellular habitat (1). The exploitation of an intracellular habitat is thought to have been one of the most important events in bacterial evolution, permitting significant environmental niche expansion and defining the arrival of intracellular pathogens and mutualistic endosymbionts (2, 3). Although there is a good understanding of the mechanisms contributing to bacterial pathogenesis, very little is known about interactions between bacterial endosymbionts and their host cells. Theoretical studies assume that there may be a tradeoff between the effectiveness of horizontal and vertical modes of transmission (4, 5). It has been predicted that mutualists evolve from parasites through an evolutionary continuum in which parasite virulence is attenuated and transmission strategy switches from horizontal to vertical (6). According to this theory, we might expect to find that pathogens and mutualistic endosymbionts harbor similar virulence determinants and utilize the same machinery to facilitate invasion and survival in host cells. In the present study, we explore these issues by investigating genes that coordinate insect cell invasion in Sodalis glossinidius, an intracellular secondary endosymbiont of the tsetse fly (Glossina spp.).

Three distinct endosymbiotic bacteria have been identified previously in the tissues of tsetse (7). Whereas one of these bacteria is known to be a parasitic Wolbachia, the remaining two are thought to be mutualists and have been classified as the primary and secondary endosymbionts of tsetse (named Wigglesworthia glossinidia and S. glossinidius, respectively) (8, 9). Sodalis is a bacterium found exclusively in tsetse flies residing both inter- and intracellularly in a number of different host tissues, including midgut, fat body, and hemolymph (9, 10). The symbiotic role of Sodalis remains unclear, because it has proved difficult to selectively eliminate either Sodalis or Wigglesworthia from tsetse without inducing sterility in the host. Phylogenetic reconstructions based on the 16S rDNA locus reveal that Sodalis is a member of the family Enterobacteriaceae, which is closely related to other intracellular secondary bacterial endosymbionts found in other insects such as the flour weevil Sitophilus zeamais and the aphid Acrythosiphon pisum (1113). We are particularly interested in Sodalis as a study model because it is known that the association between this bacterium and tsetse has only recently been established. This association is evident from symbiont–host coevolution studies demonstrating the absence of phylogenetic congruence in the evolution of Sodalis and tsetse (11). Sodalis provides an excellent model for the study of host–symbiont interactions because of the availability of an in vitro Sodalis–insect cell coculture system (14). In addition, Sodalis is the only maternally transmitted insect endosymbiont to have been isolated and maintained in pure culture (9). In this study, we demonstrate the use of Tn5 transposon mutagenesis as a tool for generating random Sodalis mutants. With the use of an in vitro negative selection procedure, we have identified Sodalis mutants deficient in their ability to attach to and invade insect cells both in vitro and in vivo. Characterization of a noninvasive Tn5 Sodalis mutant has revealed that Sodalis relies on components of a type III secretion system to facilitate entry into insect cells.

Acknowledgments

We gratefully acknowledge Roeland van Ham (Instituto Nacional de Tecnica Aeroespacial, Madrid, Spain), who provided a critique of this manuscript. This document is an output from a project funded by the Department for International Development for the benefit of developing countries (C.D.). The views expressed are not necessarily those of the Department for International Development. D.T.H. and S.C.W. are supported by the Wellcome Trust.

Acknowledgments

Abbreviation

MMMitsuhashi–Maramorosch medium
Abbreviation

Footnotes

This paper was submitted directly (Track II) to the PNAS office.

Data deposition: The sequences reported in this paper have been deposited in the GenBank database (accession nos. {"type":"entrez-nucleotide","attrs":{"text":"AF306649","term_id":"12082751","term_text":"AF306649"}}AF306649 and {"type":"entrez-nucleotide","attrs":{"text":"AF306650","term_id":"12082756","term_text":"AF306650"}}AF306650).

See commentary on page 1338.

Article published online before print: Proc. Natl. Acad. Sci. USA, 10.1073/pnas.021450998.

Article and publication date are at www.pnas.org/cgi/doi/10.1073/pnas.021450998

Footnotes

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