title: Expression and extracellular release of a functional anti-trypanosome Nanobody® in Sodalis glossinidius, a bacterial symbiont of the tsetse fly.
abstract: BACKGROUND
  Sodalis glossinidius, a gram-negative bacterial endosymbiont of the tsetse fly, has been proposed as a potential in vivo drug delivery vehicle to control trypanosome parasite development in the fly, an approach known as paratransgenesis. Despite this interest of S. glossinidius as a paratransgenic platform organism in tsetse flies, few potential effector molecules have been identified so far and to date none of these molecules have been successfully expressed in this bacterium.
  
  
  RESULTS
  In this study, S. glossinidius was transformed to express a single domain antibody, (Nanobody®) Nb_An33, that efficiently targets conserved cryptic epitopes of the variant surface glycoprotein (VSG) of the parasite Trypanosoma brucei. Next, we analyzed the capability of two predicted secretion signals to direct the extracellular delivery of significant levels of active Nb_An33. We show that the pelB leader peptide was successful in directing the export of fully functional Nb_An33 to the periplasm of S. glossinidius resulting in significant levels of extracellular release. Finally, S. glossinidius expressing pelBNb_An33 exhibited no significant reduction in terms of fitness, determined by in vitro growth kinetics, compared to the wild-type strain.
  
  
  CONCLUSIONS
  These data are the first demonstration of the expression and extracellular release of functional trypanosome-interfering Nanobodies® in S. glossinidius. Furthermore, Sodalis strains that efficiently released the effector protein were not affected in their growth, suggesting that they may be competitive with endogenous microbiota in the midgut environment of the tsetse fly. Collectively, these data reinforce the notion for the potential of S. glossinidius to be developed into a paratransgenic platform organism.
authors.affiliation: Department of Biomedical Sciences, Unit of Veterinary Protozoology, Institute of Tropical Medicine Antwerp, Antwerp, Belgium.
authors.forename: Linda
authors.initials: L
authors.lastname: De Vooght
authors.forename: Guy
authors.initials: G
authors.lastname: Caljon
authors.forename: Benoît
authors.initials: B
authors.lastname: Stijlemans
authors.forename: Patrick
authors.initials: P
authors.lastname: De Baetselier
authors.forename: Marc
authors.initials: M
authors.lastname: Coosemans
authors.forename: Jan
authors.initials: J
authors.lastname: Van den Abbeele
mesh_terms: D000818:Animals; D000913:Antibodies, Protozoan / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D004755:Enterobacteriaceae / Q000378:metabolism*; D015870:Gene Expression; D006023:Glycoproteins / Q000276:immunology; D019897:Periplasm / Q000378:metabolism; D021382:Protein Sorting Signals / Q000235:genetics; D015800:Protozoan Proteins / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D013559:Symbiosis; D014346:Trypanosoma brucei brucei / Q000378:metabolism; D014370:Tsetse Flies / Q000382:microbiology*
chemical_list: D000913:Antibodies, Protozoan; D006023:Glycoproteins; D021382:Protein Sorting Signals; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins