J Bacteriol 1998, 180:2373–2378.PubMedCentralPubMed 6. Arthofer W, Riegler M, Schneider D, Krammer M, Miller WJ, Stauffer C: Hidden

Wolbachia diversity in field populations of the European cherry fruit fly, Rhagoletis cerasi (Diptera, Tephritidae). Mol Ecol 2009, 18:3816–3830.PubMedCrossRef 7. Masui S, Kamoda S, Sasaki T, Ishikawa H: The first detection of the insertion sequence ISW1 in the intracellular reproductive parasite Wolbachia . Plasmid 1999, 42:13–19.PubMedCrossRef 8. Wu M, Sun LV, Vamathevan J, Riegler M, Deboy R, Brownlie JC, McGraw EA, Martin W, Esser C, Ahmadinejad N, Wiegand C, Madupu R, Beanan MJ, Brinkac LM, Daugherty SC, Durkin AS, Kolonay JF, Nelson WC, Mohamoud Y, Lee P, Berry K, Young MB, Utterback mTOR inhibitor AR-13324 T, Weidman J, Nierman WC,

Paulsen IT, Nelson KE, Tettelin H, O’Neill SL, Eisen JA: Phylogenomics of the reproductive parasite Wolbachia pipientis w Mel: a streamlined genome overrun by mobile genetic elements. PLoS Biol 2004, 2:E69.PubMedCentralPubMedCrossRef 9. Riegler M, Sidhu M, Miller WJ, O’Neill SL: Evidence for a global Wolbachia replacement in Drosophila melanogaster . Curr Biol 2005, 15:1428–1433.PubMedCrossRef 10. Cordaux R: ISWpi1 from Wolbachia pipientis defines a novel group of insertion sequences within the IS5 family. Gene 2008, 409:20–27.PubMedCrossRef 11. Miller WJ, Ehrman L, Schneider D: Infectious speciation revisited: impact of symbiont-depletion on female fitness and mating behavior of Drosophila paulistorum . PLoS Pathog 2010, 6:e1001214.PubMedCentralPubMedCrossRef 12. Schneider DI, Inflammation inhibitor Garschall KI, Parker AG, Abd-Alla AM, Miller WJ: Global Wolbachia prevalence, titer fluctuations and their potential of causing cytoplasmic incompatibilities in tsetse flies and hybrids of Glossina morsitans subgroup species. PIK3C2G J Invertebr Pathol 2013,112(Suppl):S104-S115.PubMedCentralPubMedCrossRef 13. Riegler M, Iturbe-Ormaetxe I, Woolfit M, Miller WJ, O’Neill SL: Tandem repeat markers as

novel diagnostic tools for high resolution fingerprinting of Wolbachia . BMC Microbiol 2012,12(Suppl 1):S12.PubMedCentralPubMedCrossRef 14. Brelsfoard C, Tsiamis G, Falchetto M, Gomulski L, Telleria E, Alam U, Ntountoumis E, Swain M, Malacrida A, Bourtzis K, Aksoy S: Wolbachia symbiont genome sequence and extensive chromosomal insertions described from the tsetse fly Glossina morsitans . 2014. 15. Stahlhut JK, Desjardins CA, Clark ME, Baldo L, Russell JA, Werren JH, Jaenike J: The mushroom habitat as an ecological arena for global exchange of Wolbachia . Mol Ecol 2010, 19:1940–1952.PubMedCrossRef 16. Belda E, Moya A, Bentley S, Silva FJ: Mobile genetic element proliferation and gene inactivation impact over the genome structure and metabolic capabilities of Sodalis glossinidius , the secondary endosymbiont of tsetse flies. BMC Genom 2010, 11:449.CrossRef 17.

The wells in the second plate were carefully washed three times with PBS and then

used to determine the total number of adherent bacteria. All assays were performed in duplicate and repeated independently four times. Murine models of infection Six- to eight-week-old female CFW1 mice (Harlan) https://www.selleckchem.com/products/napabucasin.html were used for intestinal colonization experiments as described previously [64]. Briefly, mice were provided with drinking water containing 5 g/l streptomycin sulphate for 24 h and fed a 100 μl suspension containing ~109 CFU of each strain in 20% sucrose. On indicated days, faecal pellets were collected, weighed and homogenised in 0.9% NaCl and dilutions plated onto MacConkey agar supplemented MG-132 in vitro with appropriate antibiotics for faecal CFU counts.

A previously described intranasal infection model was used in a co-infection format [23]. Six- to eight-week-old female NMRi mice (Harlan) were anaesthetized and hooked on a string by their front teeth. 50 μl of bacterial suspension containing ~5 × 107 CFU of each strain was dropped onto the nares to allow for aspiration. Mice were left hooked on the string for 10 min before being returned to their cages. At sacrifice lungs, spleen and liver were collected in 0.9% NaCl and homogenised. Serial dilutions were plated on selective media for CFU counts. The ascending urinary tract infection model in which C3H mice (Harlan) were inoculated transurethrally

many with 50 μl of bacterial suspension containing ~5 × 108 CFU bacteria has been described in detail previously [22, 65]. All animal experiments were conducted under the auspices of the Animal Experiments Inspectorate, the Danish Ministry of Justice. Data analysis, statistics and nucleotide accession number Nucleotide sequences were annotated and analysed using the Integrative Services for Genomic Analysis software and manually curated [66]. The competitive index (CI) was calculated by dividing the ratio of fim2-positive to fim2-negative bacteria recovered from selleck screening library infected organs by the ratio of the corresponding bacteria in the initial inoculum. The non-parametric Mann–Whitney U test was used to analyse infection data. Biofilm and cell-adhesion data were analysed using the non-parametric Kruskal-Wallis test and Dunn’s posthoc analysis. The nucleotide sequence of KpGI-5 has been deposited online [GenBank: JN181158]. Acknowledgements We thank Jean-Marc Ghigo, Unité de Génétique des Biofilms, Institut Pasteur, France, for providing pKOBEG-Apra and Stefan Hyman, Centre for Core Biotechnology Services, University of Leicester, for electron microscopy analysis. This study was supported by a Medisearch research grant. JJvA was supported by a University of Leicester, 50th Anniversary PhD Scholarship. SGS was partially supported by the Danish Research Agency grant 2101-06-0009.