0 was suspended in 0.8 ml of 50 mM Tris-HCl (pH 6.8). A sample of 15 μl of the protein extracts was analysed

on NuPAGE® 4-12% Bis-Tris gels (Invitrogen) Depsipeptide manufacturer using the X Cell SureLock® Mini-Cell system (Invitrogen) as recommended by the supplier. The gels were Coomassie stained using GelCode® Blue Stain Reagent (Pierce). DNA-binding analysis Gel retardation analysis were performed as described by Nan et al by mixing 100 ng of plasmid DNA (pBluescript II SK+(Stratagene)) with increasing amounts of peptide in 20 μl binding buffer (5% glycerol, 10 mM Tris, 1 mM EDTA, 1 mM dithiothreitol, 20 mM KCL and 50 μg ml-1 bovine serum albumin) [28]. Reaction mixtures were incubated 1 h at room temperature and subjected LEE011 price to 1% agarose gel electrophoresis and visualised using ethidium bromide. Transposon library in L. monocytogenes and S. aureus Transposon mutagenesis of L. monocytogenes 4446 was performed with the temperature-sensitive plasmid pLTV1 as described, but with modifications [29]. L. monocytogenes 4446 harbouring pLTV1 was grown overnight

at 30°C in BHI containing 5 μg/ml erythromycin. The bacterial culture was then diluted 1:200 in BHI containing 5 μg/ml erythromycin and grown for 6 h at 42°C. Aliquots were plated onto BHI containing 5 μg/ml erythromycin plates and incubated at 42°C. Colonies were harvested from the plates in BHI and stored in 30% glycerol at -80°C. To determine the transposition frequency, the transposon library was plated onto BHI containing 5 μg/ml erythromycin. One hundred colonies were picked and streaked

onto BHI plates containing 5 μg/ml erythromycin, 10 μg/ml chloramphenicol, and 12.5 μg/ml tetracycline, respectively, and CHIR-99021 clinical trial incubated at 30°C for 48 h. The transposition frequency was calculated as the percentage of colonies growing only on BHI + 5 μg/ml erythromycin and BHI+10 μg/ml chloramphenicol (harbouring only the transposon) but not on BHI+12.5 μg/ml tetracycline (still harbouring the plasmid). Transposon mutagenesis of S. aureus 8325-4 with bursa aurealis was performed as described [30]. Screening of transposon library for plectasin resistant mutants The transposon mutant libraries were screened on agar plates for increased resistance to plectasin as compared to wild-type sensitivity. Wild-type sensitivity was determined by plating approx. 1.0 × 107 CFU/ml on TSB agar containing plectasin (S. aureus) and approx. 1.0 × 105 CFU/ml on Muller Hinton Broth agar plates (MHB, 212322 Becton Dickinson) with plectasin (L. monocytogenes). Plates were incubated at 37°C for 3 days and inspected for growth. The transposon libraries were screened on TSB agar with 300 μg/ml, 500 or 750 μg/ml plectasin (S. aureus) or MHB plates with 250 μg/ml or 500 μg/ml plectasin (L. monocytogenes) at 37°C for up to 7 days. Identification of transposon mutant Chromosomal DNA was purified from resistant mutants using FAST DNA kit, Bio101, Qiagen, Germany).

Germany) fitted with a Zeiss LSM 510 META Confocal scan head. Imaging was carried out using the

458/477/488 nm Argon and 543 nm HeNe laser lines and a 63× C-Apochromat® water immersion lens. Live and dead cells in the stained biofilms were quantified using COMSTAT software [18] with the viability of the biofilm obtained by averaging the number of live cells over the entire z-stack [15]. Biofilm thickness was also measured using light microscopy [15]. Total RNA extraction P. gingivalis W50 biofilm and planktonic samples (40 mL) were immediately added to 0.125 volume of ice-cold Phenol solution (phenol saturated with 0.1 M citrate buffer, pH 4.3, Sigma-Aldrich, Inc. Saint Louis, MO). The mixture was centrifuged and the pellet suspended in 800 μL of ASE lysis buffer (20 mM Na acetate, 0.5% SDS, 1 mM EDTA pH 4.2) and transferred check details into a 2 mL microcentrifuge tube. An equal buy H 89 volume of ice cold Phenol solution was added and the mixture

was vortexed for 30 s before incubation at 65°C for 5 min. The mixture was then chilled on ice for 3 min after which of 200 μL of chloroform was added and mixed by brief vortexing. The mixture was centrifuged at 16,100 × g and the aqueous phase collected and extracted using a Phenol solution/chloroform (1:1 vol:vol) mix. The RNA in the aqueous phase was precipitated by addition of 700 μL of 4 M LiCl and incubated overnight at -20°C. Samples were then thawed and the total RNAs were pelleted by centrifugation. The pellet was washed with cold 70% ethanol, air dried and suspended in 50 μL of 0.1% diethylpyrocarbonate treated water. The samples were then treated with DNase I (Promega, Madison, WI) and purified using RNeasy Mini columns (Qiagen, Valencia, CA) according to protocols supplied by the manufacturer. The quality of the total RNA was verified by analytical agarose gel electrophoresis and the concentration was determined spectrophotometrically. Microarray analyses Reverse transcription reactions contained

Ribonucleotide reductase 10 μg of total RNA, 5 μg of random hexamers, the first strand buffer [75 mM KCl, 50 mM Tris-HCl (pH 8.3), 3 mM MgCl2], 0.63 mM each of dATP, dCTP, and dGTP, 0.31 mM dTTP (Invitrogen Life Technologies, Carlsbad, CA) and 0.31 mM aminoallyl dUTP (Ambion, Austin TX), 5 mM DTT, and 800 u of SuperScript III reverse transcriptase (Invitrogen). The reaction mixture was incubated at 42°C for 2 h. The RNA was hydrolysed by incubation with 0.5 M EDTA and 1 M NaOH at 65°C for 15 min and the sample neutralized with 1 M HCl before purification of the cDNA with QIAquick columns (Qiagen). The cDNAs were coupled with monoreactive Cy3 or Cy5 (40 nmol) (Amersham Biosciences, Piscataway, NJ) in the presence of 0.1 M NaHCO3 for 60 min at room temperature. The labeled cDNAs were purified using QIAquick columns (Qiagen), combined and vacuum dried. Samples were then suspended in hybridization buffer containing 50% formamide, 10× SSC (150 mM sodium citrate, pH 7.0 and 1.5 M NaCl), 0.

A similar crystallographic disorder, with an approximately 2-nm thickness, between the film and underlayer was shown in the perovskite LSMO and SrTiO3 epilayers grown on lattice mismatched

Sorafenib ic50 materials [15]. This crystallographic disorder region is associated with a lattice strain relief between the film and the underlayer. The fast Fourier transformation (FFT) patterns in Figure 2d shows two misoriented nanograins. Depending on the relative rotation among the different grains during thin-film growth, the subgrain boundaries are formed among the nanograins. The TEM image shows that the subgrain boundaries on the nanometric scale combine the discrete-oriented crystallites to form a continuous LSMO nanolayer. Quantization of the spectrum in Figure 2e shows that the contents of La, Sr, Mn, and O are approximately 12.45, 7.85, 22.11, and 57.59 at %, respectively, for the LSMO thin layer. Therefore, approximately 38.7 at % of Sr dopant was achieved within the LSMO. Figure 2f exhibits that the element contents of the In2O3 layer

are slightly oxygen deficient (the contents of In and O are approximately 46.19 and 53.81 at %, respectively). This is because the In2O3 epitaxy was selleck screening library grown under an oxygen-deficient atmosphere. Figure 2 TEM and HRTEM images and EDS spectra of LSMO nanolayer and In 2 O 3 epitaxy. (a) Low-magnification TEM image of the LSMO nanolayer with In2O3 epitaxial buffering on the sapphire substrate. The HRTEM image was taken from the interface of the In2O3 epitxay-sapphire substrate (white Cyclic nucleotide phosphodiesterase square region), and the inset shows the corresponding electron diffraction pattern at the heterointerface. (b) HRTEM image taken from the local single LSMO nanograin on the In2O3 epitaxy. (c, d) HRTEM images taken from the different local regions containing two neighboring LSMO nanograins on the In2O3 epitaxy. The corresponding FFT patterns taken from

the different oriented LSMO nanograins are also shown in the insets of (d). (e) EDS spectrum taken from the LSMO nanolayer. (f) EDS spectrum taken from the In2O3 epitaxy. Figure 3a shows the cross-sectional TEM morphology of the LSMO nanolayer grown on the bare sapphire substrate. A similarly damaged thin-layer was observed herein. Notably, granular LSMO layer contrast changes suggest that the film is composed of different LSMO crystallite orientations. Comparatively, the LSMO on the sapphire substrate experienced a relatively small degree of contrast changes, which cause the film structure to be more homogeneous than that on the In2O3 epitaxy. The insets show HR lattice fringes taken from different local regions at the interfaces between the LSMO nanograins and the sapphire substrate. Two types of heterointerface between the LSMO and substrate were presented. In the left inset, a thin (approximately 2 nm thick) transition layer formed at the heterointerface.

J Microbial Biotech 2007, 17:364–368. 10. Corti G, Panunzi I, Losco M, Buzzi R: Post-surgical osteomyelitis caused by Enterobacter sakazakii in a healthy young man. J Chemotherapy 2007, 19:94–94. 11. Forsythe SJ:Enterobacter sakazakii

and other bacteria in powdered infant milk formula. J Matern Child Nutr 2005, 1:44–50.CrossRef 12. Gallagher PG:Enterobacter bacteremia in pediatric patients. Rev Infect Dis 1990, 12:808–812.PubMed 13. Kothary MH, McCardell BA, SB203580 supplier Frazar CD, Deer D, Tall BD: Characterization of the zinc-containing metalloprotease encoded by zpx and development of a species-specific detection method for Enterobacter sakazakii. Appl Environ Microbiol 2007, 73:4142–4151.CrossRefPubMed 14. Lehner A, Stephan R: Microbiological, epidemiological, and food safety aspects of Enterobacter sakazakii. J Food Prot 2004, 67:2850–2857.PubMed 15. Mullane NR, Iverson C, Healy B, Walsh C, Whyte P, Wall PG, Quinn T, Fanning S:Enterobacter

sakazakii an emerging bacterial pathogen with implications for infant health. Minerva Pediatrica 2007, 59:137–148.PubMed 16. Mullane NR, Whyte P, Wall PG, Quinn T, Fanning S: Application of pulse field gel electrophoresis to characterize and trace the prevalence of Enterobacter sakazakii in an infant formula processing facility. Int J Food Microbiol 2007, 116:73–81.CrossRefPubMed 17. Muytjens HL, Zanen Torin 1 price HC, Sonderkamp HJ, Kollee LA, Wachsmuth IK, Farmer JJ: Analysis of eight cases of neonatal meningitis and sepsis due to Enterobacter sakazakii. J Clin Microbiol 1983, 18:115–120.PubMed Mannose-binding protein-associated serine protease 18. Gurtler JB, Kornacki JL, Beuchat L:Enterobacter sakazakii : A coliform of increased concern to infant health. Int J Food Microbiol 2005, 104:1–34.CrossRefPubMed 19. Farmer JJ, Asbury MA, Hickman FW, Brenner DJ: The Enterobacteriaceae study group. Enterobacter sakazakii : a newspecies of Enterobacteriaceae’ ‘ isolated from clinical specimens. Int J Syst Bacteriol 1980, 30:569–584.CrossRef 20. Muytjens HL, Roelofs-Willemse H, Jaspar GHJ: Quality of powdered substitutes for breast milk with regard to members of the family Enterobacteriaceae. J Clin Microbiol 1988, 26:743–746.PubMed 21. Restaino L, Frampton EW, Lionberg

WC, Becker RJ: A chromogenic plating medium for the isolation and identification of Enterobacter sakazakii from foods, food ingredients, and environmental sources. J Food Prot 2006, 69:315–322.PubMed 22. Shaker R, Osaili T, Al-Omary W, Jaradat Z, Al-Zuby M: Isolation of Enterobacter sakazakii and other Enterobacter sp. from food and food production environments. Food Control 2007, 18:1241–1245.CrossRef 23. Bar-Oz B, Preminger A, Peleg O, Block C, Arad I:Enterobacter sakazakii infection in the newborn. Acta Paediatr 2001, 90:356–358.CrossRefPubMed 24. Block C, Peleg O, Minster N, Bar-Oz B, Simhon A, Arad I, Shapiro M: Cluster of neonatal infections in Jerusalem due to unusual biochemical variant of Enterobacter sakazakii. Eur J Clin Microbiol Infect Dis 2002, 21:613–616.

This 696 nm band is now assigned to originate in chlorophyll–protein complex (CP-47) in Photosystem II. George Papageorgiou recently wrote to Govindjee about another interesting topic (photodynamic action of hypericin on cyanobacteria) on which he and Steve had worked together at Demokritos, Greece in the 1990s (see Papageorgiou et al. 1996; Brody et al. 1997).

George remarked Epigenetics Compound Library manufacturer that during a short visit to his lab, Steve had impressed all his collaborators, and added “Steve was a great scientist, a great guy, a great human being of our times.” Govindjee ends this short snippet of Steve by mentioning that Steve was a very friendly person; he was the only one to call me “Go”, first 2 letters of my name. When I spoke in Hindi on the telephone with my family and friends, he picked up one word “Accha”; it implies “OK” or “good”. In good humor, he would often use it in conversation with me. After receiving his PhD, and after only one semester of lessons from the School of Aviation, at the University of Illinois at Urbana, he obtained his private pilot license. He would rent one of the University airplanes and fly members of the

Emerson-Rabinowitch Lab (as he would put it “those who would dare”) to conferences. Jean Lavorel recently wrote, “I vividly remember that in February, selleck chemicals 1957, we had all gone in an airplane piloted by Steve to Columbus (Ohio) to participate in a Biophysical Society meeting there. It was a fascinating experience”. However, neither Rabinowitch, nor Emerson ever flew with him. I was too scared to fly with him although I did take a short ride once. Joint Research with Marcia Brody GO Marcia Brody was a former PhD student of Robert Emerson, and was also senior to me; she is currently Professor Emeritus of Hunter College, New York. Marcia is an accomplished scientist and had made major discoveries

in the area of two-light effect and two photosystems in the red alga Porphyridium cruentum (see e.g., M. Brody and Emerson 1959a, b). Historically, it is important to point out that oxyclozanide Marcia was a coauthor of an early abstract of a presentation by Robert Emerson (Emerson et al. 1956) that had some of the first hints on what led to the concepts of the two-light effect and two pigment systems of photosynthesis, based on the Emerson Enhancement Effect (Emerson et al. 1957; Rabinowitch and Govindjee 1960; R. Govindjee et al. 1960. (Both Govindjee and Rajni Govindjee were students of Emerson, but became students of Rabinowitch after Emerson died in a plane crash on Feb. 4, 1959.) Steve Brody collaborated with Marcia (see Biographical Portrait below) extensively since 1959 for a little more than 10 years. We mention only a few of their collaborative studies here. This collaboration included studies on dynamic changes in the efficiency of excitation energy transfer (Brody and Brody 1959; M.

References Aasen PA (2009) Abrodd—Artemisia abrotanum L.http://​www.​plantearven.​no/​abbrodd.​htm Baade PN (1768/1901) Tronhiemske Have-Planter. In: Nøvik P (ed) Samlinger til Havebrugets

historie i Norge. Udgivet af Selskabet “Havedyrkningens venner”. Gröndahl & Sön, Christiania, pp 75–87 Balvoll G, Weisæth G (1994) Horticultura. Norsk hagebok fra 1694 av Christian Gartner. Landbruksforlaget, Otta Berentsen VD, Eek A, Grefsrød E-E (2007) Sansehager for personer med demens. Utforming og bruk. Aldring og helse, Tønsberg Hammer C (1772) Norsk Huusholdings-Kalender. Første Deel. S. C. Schwach, Christiania Jacquin NJ (1764) Observationum botanicarum. Vindobonae, Vienna Kålås JA, Viken Å, Bakken T

(eds) (2006) 2006 Norwegian red list. The Norwegian Biodiversity Information GS 1101 Centre, Trondheim Kaplan R, Kaplan S (1989) The experience of nature. Cambridge Press, Cambridge The Linnaean correspondence, an electronic edition prepared by the Swedish Linnaeus Society, Uppsala, and published by the Centre international d’étude du XVIIIe siècle, Ferney-Voltaire. Giovanni Antonio Scopoli to Carl Linnaeus, 1 September 1760, The Linnaean Correspondence, linnaeus.c18.net, Ferrostatin-1 cost letter L2798 (consulted 19 August 2009). Carl Linnaeus to Nicolaus Joseph von Jacquin, 1 April 1764, The Linnaean Correspondence, linnaeus.c18.net, letter L3397 (consulted 19 August 2009). Carl Linnaeus to Nicolaus Joseph von Jacquin, 24 August 1767, The Linnaean Correspondence, linnaeus.c18.net, letter L3945 (consulted 19 August 2009) Marstein M (2009)

Galnebær—Scopolia carniolicaJacq.http://​www.​plantearven.​no/​galnebær.​htm Rathke J (1823) Enumeratio plantarum horti botanici Universitatis Regiae Fredericianae Christianiensis. Gröndahl, Christiania Reichborn-Kjennerud, I (1922) Våre folkemedisinske lægeurter. Kristiania Schübeler FC (1886–1889) Viridarium norwegicum. Norges Væxtrige. Et bidrag til Nord-Europas natur og Culturhistorie, I–III. Christiania Scopoli GA (1760) Flora Carniolica, ed.1. Vienna Stafleu FA, Cowan however RS (1985) Taxonomic literature, 5th edn. Sal-Ste. W. Junk b.v., The Hague, Boston”
“Introduction Insects associated with plant galls have been a key model system for understanding host-parasite interactions, trophic cascades, host specificity, and other aspects of community ecology, as these multitrophic systems represent natural microcosms that are tractable for ecologists (Stone et al. 2002). Gall inducers manipulate their host plant to produce structures of varying complexity in which the gall inducer develops (Rohfritsch 1992). The most complex and species rich group of gall-inducing organisms are the cynipid gall wasps of the tribe Cynipini, which produce complex galls on various tissues of oaks.

loti R7A and MAFF303099 has shown that T4SS is involved in the symbiosis stabilization, increasing or decreasing the nodulation phenotype, according to the host involved [53]. The homologous proteins of virB, AvhB8, AvhB9, and AvhB10 genes identified in R. tumefaciens and VirB8, VirB9, and VirB10 of E. meliloti are located on plasmids. Although there is a considerable Ceritinib synteny between R. tumefaciens

and E. meliloti chromosomes [5, 26], conservation in the gene order among the plasmids of these microorganisms is not expected, due to the high frequency of horizontal gene transfer between plasmids of species of the Rhizobiales order. However, the grouping observed between the symbiont E. meliloti and the pathogen R. tumefaciens in the reconstruction trees generated with VirB8, VirB9, and VirB10 is in agreement with the topologies of VirB/Trb presented by Frank et al. (2005) [54], which examined

the functional divergence and horizontal transfer of the T4SS. According to these authors, the coexistence of the AvhB conjugation protein with VirB translocation effectors in the same clade, as well as the location of these proteins in plasmids and the presence of multiple copies in some species, is indicative of the occurrence of multiple events of horizontal gene transfer, the process believed to be responsible for spreading the virB operon Fenbendazole between the alpha-Proteobacteria, representing the dominant mechanism in the evolution of the conjugation selleck kinase inhibitor systems for secretion. Regarding the proximity of the X. autotrophicus with R. radiobacter, and of Bradyrhizobium BTAi1 with

B. quintana or R. vitis, there is no data in the literature that could allow inferences about such relationships. In these organisms, the virB operon is located between hypothetical and Tra conjugation proteins (data not shown). However, proteins involved in integration, transposition, and/or DNA recombination were not identified close to VirB8, VirB9, and VirB10 (database), which might allow inferences that these genes could have arisen from horizontal gene transfer. Conclusions In this study, the genomic comparison has shown that symbiotic and pathogenic bacteria belonging to the order Rhizobiales may share several similar strategies of host interaction, inference taken from the high similarity on several proteins identified – e.g., FixNOPQ, NodN and VirB8910. However, it should be noted that some common clusters obtained are formed by protein families which may possess different functions in each process. The presence of symbiotic and virulence genes in both pathogens and symbionts does not seem to be the only determinant factor for lifestyle evolution in these microorganisms, although they may act in common stages of host infection.

SR contributed to sample collection and microbiological analysis. MA provided direction on available means of data analyses. RS conceived the study, analysed the data and wrote the manuscript. All authors contributed to the general content and structure of the final manuscript.”
“Background The enormous impact of horizontal gene transfer (HGT) on the evolution of bacterial Y-27632 datasheet species has only been recognized during the past years [1]. Among the mobile genetic elements involved in HGT genomic islands are of particular relevance since they can comprise large genomic regions encoding accessory factors required by the bacteria to thrive in specific environments. For example, many virulence related factors of pathogenic

bacteria are encoded on so-called pathogenicity islands, while metabolic islands frequently encode factors required for detoxification of poisonous compounds or for the utilization of specific carbon sources such as aromatic compounds [2, 3]. The genus Bordetella harbours several important pathogens infecting humans and various animals. While B. pertussis and B. parapertussis are etiological agents of whooping cough in man, B. bronchiseptica and B. avium can cause respiratory infections in various mammalian species and birds, respectively [4]. B. petrii was the first Bordetella species isolated from the environment, while all other

Bordetella species so far could only be found in obligate association with host organisms [5]. Phylogenetically, B. petrii appears to be closely Montelukast Sodium FK506 related to a common ancestor of the pathogenic Bordetellae and links the genus with other environmental bacteria of the genera Achromobacter and Alcaligenes [5, 6]. B. petrii was repeatedly isolated from contaminated soil [7, 8]. However, recently, several isolates from clinical specimens associated with bone degenerative disease or cystic fibrosis were found to be closely related to B. petrii, although the underlying etiology is not

clear in any of the cases [9–11]. The pathogenic Bordetellae encode a multitude of virulence factors including several toxins and adhesins [4]. The evolutionary origin of these factors is unclear, since in contrast to many virulence genes of other pathogens they are not located on mobile genetic elements such as pathogenicity islands or prophages. In fact, so far only few presumptive horizontal gene transfer events are known among the pathogenic members of the genus, e.g. a 66 kb island encoding iron transport genes that presumably has been exchanged between B. pertussis and B. holmesii, a pathogenic species mainly found in immunocompromised individuals [12]. A prevalent feature in the evolution of virulence in this genus is reductive genome evolution, since strains specialized on particular host organisms such as the exclusive human pathogen B. pertussis have presumably evolved from a B. bronchiseptica-like ancestor.

(XLS 43 KB) Additional file 4: Figure S2: Predicted T7G translational

frameshift sites in Smp131 and closely related prophages from Xanthomoas and Stenotrophomonas. (A) T7G (enclosed by a rectangle) and the surrounding regions including genes p27, p27.1 and p28 of Smp131. Stop codons are denoted by three dots after the amino acids. Predicted start codon ATG of p27.1 is underlined, whereas ribosomal binding site AGAGG for gene p28 is in gray background. (B) DNA sequence alignment of the regions surrounding T7G translational frameshift sites (enclosed in rectangles) from Smp131 and the related prophages from X. campestris pv. campestris 33913, X. oryzae pv. oryzae strains KACC10331, MAFF311018 and PXO99A. An asterisk indicates identical nucleotides in all phages. (PPT 1 MB) Additional file 5: Figure S3: Comparison of tyrosine integrase of Smp131 and its homologues. Identical residues found in 3-deazaneplanocin A concentration more than 3 residues are highlighted. Active sites determined for XerD are indicated by downward arrowhead and the RKHRH pentad conserved

residues are indicated above. The α-helix (empty rectangle) and β-sheet (empty arrow) structural motifs under the alignments are based on the crystal structure of E. coli XerD. Abbreviations: Smp131, integrase deduced from Smp131 orf43; P2, integrase of Enterobacteria phage P2 (GenBank:P36932); 186, integrase of Enterobacteria phage 186 (GenBank:P06723); XerD, site-specific recombinase selleck kinase inhibitor of E. coli (GenBank:1A0P_A). (PPT 2 MB) Additional file 6: Table S3: Identities of amino acid sequence shared between the proteins deduced from Smp131 and those from bacteriophages. (XLS 44 KB) Additional file 7: Table S4: Positions and sequences of att sites and tRNA of Smp131 and prophages in Xanthomonas and Stenotrophomonas. (XLS 26 KB) Additional file 8: Figure S4: Strategy for cloning the host-prophage junctions from Smp131-lysogenized S. maltophilia T13. (A) Sketch depicting the circular Smp131 Bay 11-7085 genome and genes near the predicted attP site. Arrows represent the genes and predicted attP site. (B) Sketch showing the host S. maltophilia

T13 chromosome and its attB site. (C) Map showing relative positions of genes after Smp131 integration into host S. maltophilia T13. Primers used in PCR were: L1; 5′-TGAAAGGTGCCATGACCACACG-3′; L2, 5′-GCGTTGCCAAGGTCAGATCGG-3′; L3; 5′-CGCATCGCACTCTAGGAAGTGAAG-3′; L4, 5′-AACTGCCAGAACCTCTGCAGTG-3′; R1, 5′-CTCTTGTCCTCGCTGTCGGT-3′; R2, 5′-TGATAGCCCTATTTTCAAGGGC-3′; R3, 5′-AGGCCCAGCAGCGCA-3′; R4, 5′-TGCCTGCCGCCAGCT-3′. S. maltophilia T13 chromosome containing prophage Smp131 was digested with HincII and NaeI. The fragments were self-ligated and the circularized DNA was then used as the templates for inverse PCR. Amplicons obtained were sequenced for comparison. (PPT 183 KB) References 1. Palleroni NJ, Bradbury JF: Stenotrophomonas, a new bacterial genus for Xanthomonas maltophilia (Hugh 1980) Swings et al. 1983.

Briefly, 100 mg of extracted and purified rhamnolipids were suspended in 5 ml of 50 mM sodium acetate buffer, pH 4.1. To this solution was added 100 mg of naringinase from Penicillum decumbens (Sigma). The mixture was then kept at 50°C for 2 h with gyratory shaking (240 rpm), at which point 20 ml of buffer were added. After 24 h, another 150 mg of naringinase were added as well as 25 ml of buffer. The reaction was kept under these conditions for 8 days. A final 50 mg of naringinase

in 20 ml of buffer were added to the mixture and was left for another 24 h. Thereafter, the solution was acidified to pH 3-4 using concentrated HCl and extracted three times with ethyl acetate. The fatty acid moieties generated by naringinase cleavage were then analyzed by LC/MS after the extract had been dried and evaporated. CMC – Surface tension assay Critical micelle concentration and surface tension were measured by the du Noüy ring Selleck Lumacaftor method [50] using a surface tensiometer (Fisher). The instrument was calibrated against water and assays were performed in triplicate at room temperature. Swarming motility For swarming MG-132 order assays, cultures were grown overnight, diluted

in fresh medium and subcultured until OD600~6.0 was reached. Swarm plates were prepared as follows: freshly autoclaved medium consisting of NB supplemented with 0.5% dextrose (Fisher) and 0.5% Bacto-agar (Difco) was poured into standard Petri dishes and dried under laminar flow for 30 min, as before [42]. Immediately following the drying period, plates were inoculated at their center with 5 μl of bacterial culture and placed at 30°C. For swarming phenotype restoration, 1, 5, 10 and 25 mg/L of purified B. thailandensis E264 rhamnolipids were deposited (10 μl) at the

center of respective plates and left to dry for 15 minutes before spot inoculation with swarming-deficient ΔrhlA mutant strains. For cross-feeding experiments, either equal parts of the cultures were mixed before being plated at the center on the swarm plate, or cultures were simply spotted side-by-side. Acknowledgements Special thanks to Marie-Christine Groleau and Ludovic Vial for insightful O-methylated flavonoid comments and technical assistance as well as all members of ED laboratory for helpful discussions. This work was funded by NSERC discovery grants to FL and ED. DD was recipient of a Master’s Degree scholarship from The Fondation Armand-Frappier. References 1. Jarvis FG, Johnson MJ: A glyco-lipid produced by Pseudomonas aeruginosa. J Am Oil Chem Soc 1949,71(12):4124–4126. 2. Edwards JR, Hayashi JA: Structure of a rhamnolipid from Pseudomonas aeruginosa. Arch Biochem Biophys 1965,111(2):415–421.CrossRefPubMed 3. Kitamoto D, Isoda H, Nakahara T: Functions and potential applications of glycolipid biosurfactants–from energy-saving materials to gene delivery carriers. J Biosci Bioeng 2002,94(3):187–201.PubMed 4. Rahman PKSM, Gakpe E: Production, characterisation and applications of biosurfactants – Review.