Lane 1, 33277; lane 2, KDP164 (hbp35 insertion mutant); lane 3, K

Lane 1, 33277; lane 2, KDP164 (hbp35 insertion mutant); lane 3, KDP166 (hbp35 deletion mutant). (PPT 390 KB) Additional file 2: Preparation of the anti-HBP35-immunoreactive 27-kDa protein for PMF analysis. Immunoprecipitates of lysates of KDP164 (hbp35 insertion mutant) with anti-HBP35 antibody was analyzed by SDS-PAGE followed by staining with CBB (left)

or immunoblot analysis with anti-HBP35 antibody (right). A 27-kDa protein band on the gel indicated was subjected to PMF analysis. (PPT 222 KB) Additional file 3: Structures of the HBP35 protein GSK2118436 molecular weight and the hbp35 gene. A. Domain organization of HBP35 protein. HBP35 contains a signal peptide region, a thioredoxin domain and a C-terminal domain. B. The hbp35 gene loci in various buy BI-D1870 mutant strains. Mutated hbp35 genes of KDP164 (hbp35

insertion mutant), KDP168 (hbp35 [M115A] insertion mutant), KDP169 (hbp35 [M135A] insertion mutant) and KDP170 (hbp35 [M115A M135A] insertion mutant) were depicted. (PPT 170 KB) Additional file 4: N-terminal amino acid sequencing of the recombinant 27-kDa protein produced in an E. coli expressing the hbp35 gene. rHBP35 products, which were partially purified using a C-terminal histidine-tag, were analyzed by SDS-PAGE followed by staining with CBB (left) or immunoblot analysis with anti-HBP35 PF 2341066 antibody (right). The N-terminal amino acid sequence of the recombinant 27-kDa protein was determined Resveratrol by Edman sequencing, resulting in M135 as an N-terminal residue. (PPT 320 KB) Additional file 5: Bacterial strains and plasmids used in this study. (XLS 32 KB) Additional file 6: Oligonucleotides used in this study. (DOC 35 KB) References 1. Roper JM, Raux E, Brindley AA, Schubert HL, Gharbia SE, Shah HN, Warren MJ: The enigma of cobalamin (Vitamin B12) biosynthesis in Porphyromonas gingivalis . Identification and characterization of a functional corrin pathway. J Biol Chem 2000,275(51):40316–40323.PubMedCrossRef 2. Kusaba A, Ansai T, Akifusa S, Nakahigashi K, Taketani S, Inokuchi H, Takehara T: Cloning and expression of a Porphyromonas gingivalis gene for protoporphyrinogen oxidase by complementation of a hemG mutant of Escherichia

coli . Oral Microbiol Immunol 2002,17(5):290–295.PubMedCrossRef 3. Nelson KE, Fleischmann RD, DeBoy RT, Paulsen IT, Fouts DE, Eisen JA, Daugherty SC, Dodson RJ, Durkin AS, Gwinn M, et al.: Complete genome sequence of the oral pathogenic bacterium Porphyromonas gingivalis strain W83. J Bacteriol 2003,185(18):5591–5601.PubMedCrossRef 4. Olczak T, Simpson W, Liu X, Genco CA: Iron and heme utilization in Porphyromonas gingivalis . FEMS Microbiol Rev 2005,29(1):119–144.PubMedCrossRef 5. Potempa J, Sroka A, Imamura T, Travis J: Gingipains, the major cysteine proteinases and virulence factors of Porphyromonas gingivalis : structure, function and assembly of multidomain protein complexes. Curr Protein Pept Sci 2003,4(6):397–407.PubMedCrossRef 6.

Phys Rev B 2009, 79:205211 CrossRef 3 Kumar M, Singh

Phys Rev B 2009, 79:205211.CrossRef 3. Kumar M, Singh Selleck Roscovitine VN, Mehta BR, Singh JP: Tunable synthesis of indium oxide octahedra, nanowires and tubular nanoarrow structures under oxidizing and reducing ambients. Nanotechnology 2009, 20:235608.CrossRef 4. Han SY, Herman GS, Chang CH: Low-temperature, high-performance, solution-processed indium oxide thin-film transistors. J Am Chem Soc 2011, 133:5166–5169.CrossRef 5. Elouali S, Bloor LG, Binions

R, Parkin IP, Carmalt CJ, Darr JA: Gas sensing with nano-indium oxides (In 2 O 3 ) prepared via continuous hydrothermal flow synthesis. Langmuir 2012, 28:1879–1885.CrossRef 6. Lee D, Ondrake J, Cui T: A conductometric indium oxide semiconducting nanoparticle enzymatic biosensor array. Sensors 2011, 11:9300–9312.CrossRef 7. Reyes-Gil KR, Reyes-Garcia EA, Raftery D: Nitrogen-doped In 2 O 3 thin film electrodes for photocatalytic water splitting. J Phys Chem C 2007, 111:14579–14588.CrossRef 8. Gan J, Lu X, Wu J, Xie S, Zhai T, Yu M, Zhang Z, Mao Y, Wang SCL, Shen Y, Tong Y: Oxygen vacancies promoting photoelectrochemical performance of In 2 O 3 nanocubes. GS-9973 clinical trial Sci Rep 2013, 3:1021. 9. Shao D, Qin L, Sawyer S: High responsivity, bandpass near-UV photodetector fabricated from PVA-In 2 O 3 nanoparticles on a GaN substrate. IEEE Photon J 2012, 4:715–720.CrossRef

10. Zhang D, Li C, Han S, Liu X, Tang T, Jin W, Zhou C: Ultraviolet photodetection properties of indium oxide nanowires. Appl Phys A 2003, 77:163–166.CrossRef 11. Al-Dahoudi N, Aegerter MA: Comparative study of transparent conductive selleck In 2 O 3 :Sn (ITO) coatings

made using a sol and a nanoparticle suspension. Thin Solid Films 2006, 502:193–197.CrossRef 12. Cheong DS, Yun DH, Kim DH, Han KR: Indium tin oxide (ITO) coatings fabricated using nanoparticle slurry and sol. J Korean Ceram Soc 2011, 48:516–519.CrossRef 13. Flores-Mendoza MA, Castanedo-Perez R, Torres-Delgado G, Marquez Marin J, Zelaya-Angel O: Influence of Selleckchem HSP inhibitor annealing temperature on the properties of undoped indium oxide thin films obtained by the sol–gel method. Thin Solid Films 2008, 517:681–685.CrossRef 14. Kim S, Kim S, Srisungsitthisunti P, Lee C, Xu M, Ye PD, Qi M, Xu X, Zhou C, Ju S, Janes DB: Selective contact anneal effects on indium oxide nanowires transistors using femtosecond laser. J Phys Chem C 2011, 115:17147–17153.CrossRef 15. Wu CC, Wu CI, Sturm JC, Kahn A: Surface modification of indium tin oxide by plasma treatment: an effective method to improve the efficiency, brightness, and reliability of organic light emitting devices. Appl Phys Lett 1997, 70:1348–1350.CrossRef 16. Remashan K, Hwang DK, Park SD, Bae JW, Yeom GY, Park SJ, Jang JH: Effect of N 2 O plasma treatment on the performance of ZnO TFTs. Electrochem Solid-State Lett 2008, 11:H55-H59.CrossRef 17. Murali A, Barve A, Leppert VJ, Risbud SH: Synthesis and characterization of indium oxide nanoparticles. Nano Lett 2001, 1:287–289.CrossRef 18.

brasiliensis (Figure 1B), at least a 3-fold increase in compariso

brasiliensis (Figure 1B), at least a 3-fold increase in comparison with the control. Also, the proportion of internalized yeast cells (23%) was higher than the proportion of yeast cells adhered to macrophage surfaces (6%). In contrast, we found that 0.25 μM alexidine dihydrochloride caused an 8-fold inhibition in the levels of phagocytosis by MH-S cells compared with the control (Figure 1B). No effects of alexidine dihydrochloride or pulmonary surfactant on adhesion and internalization of heat-killed P. brasiliensis were observed (data not shown).

Table 1 Phospholipase B activities secreted under the experimental conditions used for Phagocytic test Treatment Specific activity of PLB (μmol buy BTSA1 min-1mg-1protein) Untreated control 1.21 ± 0.02 Pulmonary surfactant (100 μg mL-1) buy Cilengitide 1.55 ± 0.06* (28% activation) Alexidine dihydrochloride (0.25 μM) 0.41 ± 0.08* (66% inhibition) Phospholipase B activities were assayed after 6 h of co-cultivation

of alveolar macrophage (MH-S) cells with P. brasiliensis yeast cells with pulmonary surfactant (100 μg mL-1) and alexidine dihydrochloride (0.25 μM), as well as without treatment (untreated control), as described in Materials and Methods. *Significantly different from the untreated control, P < 0.05 by the paired 2-tailed Student's t-test. Results are means ± SEM of triplicate assays. A role for learn more PLB activity in adhesion of C. neoformans to lung epithelial cells has already been proposed [9]; DPPC is predicted to be the favored lipid substrate for PLB, leading to the production of glycerophosphocholine and free palmitic acid. In this context, it is hypothesized that the addition of pulmonary surfactant (rich in DPPC) would increase the adhesion of P. brasiliensis yeast cells to MH-S cells. These results strongly suggest that PLB activity is important in P. brasiliensis adhesion to and/or internalization by MH-S cells. In the present study, enzyme activities were tested under conditions

used for adhesion Acetophenone (Table 1). P. brasiliensis produced high levels of PLB at 6 h post-infection. 0.25 μM Alexidine dihydrochloride selectively inhibited PLB activity by 66%. In contrast, PLB activity in the presence of 100 μg mL-1 pulmonary surfactant was significantly increased (28%) compared to the control experiment. Modulation of P. brasiliensis and MH-S genes in the host-pathogen interaction Real-time quantitative reverse-rranscriptase-polymerase chain reaction (qRT-PCR) analysis confirmed that the plb1 (PLB), sod3 (Cu, Zn superoxide dismutase – SOD), and icl1 (isocitrate lyase) genes were up-regulated in P. brasiliensis yeast cells during 6 h of interaction with MH-S cells in the presence of pulmonary surfactant. The sod3 gene presented a 4.1-fold increase in expression (Figure 2) and under these conditions a higher percentage of yeast cell internalization was observed (Figure 1B).

In general, the proteins of any one (sub)family are distributed f

In general, the proteins of any one (sub)family are distributed fairly equally between these three segments with few exceptions. Arm1 includes 17% of the total chromosome and encodes 16% of the transport proteins. The core

includes 57% of the chromosome and encodes 54% of the transport proteins. Arm2 includes 26% of the chromosome and encodes 30% of the transport proteins. Thus, transporter genes exhibit nearly uniform density within the three chromosomal segments. Three (sub)families (2.A.1.67, 2.A.39 and 3.A.1.3) have five members in S. coelicolor. The distributions of the encoding genes within arm1, arm2, PI3K Inhibitor Library concentration and core are 0/1/4, 1/2/2 and 0/0/5. Subfamily 3.A.1.3 is concerned exclusively with the uptake of polar amino acids and therefore probably serves housekeeping functions. Five subfamilies

have six proteins, and all but one are represented in all three chromosomal segments. Two subfamilies have seven proteins and two have eight. All four are also represented in all three segments. Two subfamilies Selleckchem Mocetinostat (3.A.1.2 and 3.A.1.105) have ten members, and while the former has representation in all three segments, the latter has all ten genes in the core. These proteins catalyze drug export. Subfamily 2.A.1.2 has eleven members distributed throughout the chromosome. Two (sub)families have seventeen members. Family 2.A.3 amino acid uptake porters and subfamily 3.A.1.5 peptide and oligosaccharide uptake systems are distributed about equally on arm2 and the core with little or no representation on arm1. Finally, the 45 members of the MFS polar amino acid porters (subfamily 2.A.1.3) show equal representation in arm 2 and the core, but poor representation in arm1. Conversely, ABC sugar transporters of subfamily of 3.A.1.1 with 75 members have nearly equal distribution in the three chromosomal segments. In this case the gene density is somewhat highest on arm1. These results show that while the transporters in general are distributed in accordance with expectation based on the sizes of these segments, some (sub)families are asymmetrically distributed. However, seldom are the members of a single (sub)family localized to a single segment.

Identification of distant transport proteins in Sco In the analyses reported above, the cutoff point for proteins retrieved using the GBLAST program was an e-value of 0.001. In order to determine if more distant transport protein homologues could be Adenosine identified, all sequences brought up with e-values between 0.001 and 0.1 were examined. In Sco, over 300 sequences were retrieved, almost all of which proved to be false positives. However, careful examination revealed that a few true transport protein homologues were included in this list. The following 14 proteins, all of which have been included in TCDB, were obtained (see Table 3). Table 3 Distant Sco transport proteins Assigned TC number UniProt acc number Size (number of aas) Number of TMSs Family assignment 2.A.1.21.18 Q9KXM8 463 12 MFS Superfamily 2.A.1.21.

Aquat Microb Ecol 37:295–304 Tianpanich K, Prachya S, Wiyakrutta

Aquat Microb Ecol 37:295–304 Tianpanich K, Prachya S, Wiyakrutta S, Mahidol C, Ruchirawat S, Kittakoop P (2011) Radical scavenging and antioxidant activities of isocoumarins and a phthalide from the endophytic fungus Colletotrichum sp. J Nat Prod 74:79–81PubMed Vadassery J, Oelmüller R (2009) Calcium signaling in pathogenic and beneficial plant microbe interactions. Plant Signal Behav 4:1024–1027PubMed Vadassery J, Ritter C, Venus Y, Camehl I, Varma A, Shahollari B, Novák O, Strnad M, Ludwig-Müller J, Oelmüller R (2008) The role of auxins and cytokinins in the mutualistic interaction between Arabidopsis and Piriformospora indica. Mol Plant Microbe Interact 21:1371–Selleckchem Ganetespib 1383PubMed van Oppen

MJH, Leong JA, Gates RD (2009) Coral-virus interactions: a double-edged sword? Symbiosis 47:1–8 Varughese SHP099 in vivo T, Rios N, Higginbotham S, Arnold AE, Coley PD, Kursar TA, Gerwick WH, Rios LC (2012) Antifungal depsidone metabolites from Cordyceps dipterigena, an endophytic fungus antagonistic to the phytopathogen Gibberella fujikuroi. Tetrahedron Lett 53:1624–1626PubMed Verma SA, Varma A, Rexer KH, Hassel A, Kost G, Sarbhoy A, Bisen P, Bütehorn B, Franken P (1998)

Piriformospora indica, gen. et sp. nov., a new root-colonizing fungus. learn more Mycologia 90:898–905 Waller F, Achatz B, Baltruschat H, Fodor J, Becker K, Fischer M, Heier T, Hückelhoven R, Neumann C, von Wettstein D, Franken P, Kogel KH (2005) The endophytic fungus Piriformospora indica reprograms barley to salt-stress tolerance, disease resistance and higher yield. Proc Natl Acad Sci USA 102:13386–13391PubMed Wang LW, Xu BG, Wang JY, Su ZZ, Lin FC, Zhang CL, Kubicek CP (2012a) Bioactive

metabolites from Phoma species, an endophytic fungus from the Chinese medicinal plant Arisaema erubescens. Appl Microbiol Biotechnol 93:1231–1239PubMed Wang Y, Xu L, Ren W, Zhao D, Zhu Y, Wu X (2012b) Bioactive metabolites from Chaetomium globosum L18, an endophytic fungus in the medicinal plant Curcuma wenyujin. Phytomedicine 19:364–368PubMed Webster NS, Taylor MW (2012) Marine sponges Phospholipase D1 and their microbial symbionts: love and other relationships. Environ Microbiol 14:335–346PubMed Weinl S, Held K, Schlücking K, Steinhorst L, Kuhlgert S, Hippler M, Kudla J (2008) A plastid protein crucial for Ca2+-regulated stomatal responses. New Phytol 179:675–686PubMed Williams RB, Henrikson JC, Hoover AR, Lee AE, Cichewicz RH (2008) Epigenetic remodeling of the fungal secondary metabolome. Org Biomol Chem 6:1895–1897PubMed Xia X, Zhang J, Zhang Y, Wei F, Liu X, Jia A, Liu C, Li W, She Z, Lin Y (2012) Pimarane diterpenes from the fungus Epicoccum sp. HS-1 associated with Apostichopus japonicas. Bioorg Med Chem Lett 22:3017–3019PubMed Yang G, Sandjo L, Yun K, Leutou AS, Kim G-D, Choi HD, Kang JS, Hong J, son BW (2011) Flavusides A and B, antibacterial cerebrosides from the marine-derived fungus Aspergillus flavus.

aureus 43300 without interference from the nasal flora was needed

aureus 43300 without interference from the nasal flora was needed. Hence, nutrient agar plates with different concentrations of ampicillin (4, 8, 16, 20 and 32 μg/ml)

were prepared. All the nasal isolates (NS-1, NS-2, NS-3, S. aureus 29213 as well as S. aureus 43300) were spread #Idasanutlin mw randurls[1|1|,|CHEM1|]# plated respectively. Nutrient agar plates with no antibiotic were used as control. All the plates were incubated for 24 h at 37°C. Next day, growth was observed on plates and the ampicillin concentration showing complete inhibition of growth (no colonies on selective plates) was noted. Ampicillin at a concentration of ≥16 μg/ml completely inhibited the growth of NS-1, NS-2 and NS-3 however MRSA 43300 growth was inhibited at 32 μg/ml. Hence, a GSK2118436 dose of 20 μg/ml ampicillin was selected to be added to nutrient agar for preparing selective plates which allowed the growth of MRSA 43300 colonies only with no interference from nasal flora strains. Nasal carriage model of S. aureus 43300 S. aureus 43300 was cultivated for 24 h at 37°C in brain heart infusion broth. Next day,

cells were pelleted and washed twice with phosphate-buffered saline (PBS). Bacterial suspension prepared in PBS was adjusted at 600 nm so as to achieve a cell density corresponding to a range of bacteria inoculums (105,106 and 107 CFU/ml). The number of CFU/ml was confirmed by quantitative plate count. Mice were grouped randomly into three groups (N = 3) with twenty mice (n = 20) per group. For intranasal instillation, a 50 μl inoculum of respective bacterial dose was instilled into the nasal opening while holding the mice upright. The mouse was held upright for at least 2 minutes to allow the mice to take the inoculum with minimum loss. After an interval of 48 hours, second dose of inoculum was again instilled into the nares of

mice in the same way as described above. Four mice from each group were sacrificed on day 2, 5, 7, 10 and 12 post inoculum administrations. After disinfecting the nasal area with 70% alcohol, the nasal tissue was dissected from each mouse and washed twice in PBS (pH 7.2). The tissue was homogenized, and dilutions of the homogenates were plated on nutrient agar plates to evaluate total bacterial flora. The homogenate dilutions were also plated on nutrient agar plates RVX-208 containing ampicillin (20 μg/ml) so as to check the load of S. aureus 43300 colonised in the nasal tissue. Phage and mupirocin protection studies Therapeutic potential of bacteriophage, MR-10 alone as well as in combination with mupirocin was evaluated for its ability to reduce the nasal carriage in BALB/c mice. Male BALB/c mice were used and randomly divided into four groups (N = 4) with each group containing 20 mice each (n = 20). The infection and treatment schedule is depicted in Figure 1. Figure 1 Schematic representation of the infection and treatment schedule followed for establishing nasal colonization model in BALB/c mice.

Step 8: Select suitable survey methods For most measurement endpo

Step 8: Select suitable survey methods For most measurement endpoints, several survey methods exist (Table 3) but not all methods are equally effective for all species or species groups. We recommend survey methods that monitor multiple species simultaneously to provide more information for similar effort. We also recommend using more than one survey method for each species, because combining methods can decrease bias and provide better estimates Lazertinib chemical structure of

the variable of interest. Consistent use of the same methods and personnel over time and across control/mitigation sites is important to provide comparable results. Table 3 Potential survey method(s) for each measurement endpoint Assessment endpoint Measurement endpoint Potential survey methods Human casualties Number of NCT-501 humans killed or injured due to wildlife-vehicle collisions or due to collision avoidance Questionnaire Insurance money spent on material/immaterial damage due to wildlife-vehicle collisions Questionnaire Number of hospitalizations

due to vehicle-animal GM6001 mouse collisions Questionnaire Number of wildlife-vehicle collisions, concerning species that potentially impact human safety, regardless of whether they resulted in human injury or death Road surveys Wildlife health and mortality Number of animals killed or injured while crossing roads Road surveys Number of animals killed or with ill-health due to isolation from needed resources through the barrier effect of roads Field surveys Population viability Trend in population size/density Capture-mark-recapture, Point/Transect counts or calling surveys, Pellet counts, Nest/den counts, Tracking arrays, e.g. photo/video cameras, track pads Number of animals killed Road surveys Reproductive success Counts of eggs/young Age before structure Capture, Direct observation Sex ratio Capture, Direct observation Between-population movements Capture-Mark-Recapture, Radio-tracking, Direct observation, Tracking arrays Genetic differentiation Invasive DNA sampling after capture, Non-invasive DNA sampling, e.g. through hair traps, scat collection, antler/skin collection Genetic variability Invasive DNA sampling after capture, Non-invasive DNA sampling

The list provides primarily some examples of frequently used survey methods and is not aimed at being complete Step 9: Determine costs and feasibility A comprehensive evaluation of road mitigation measures will require a substantial budget. However, other resources that may not have direct costs are equally important, e.g., sufficient time, or stakeholder support. The need for both economic and non-economic resources demands detailed organization and planning, including clear deadlines for decisions, and strong consensus among the research team, the funding organization and other stakeholders. For example, if a land owner refuses access to a sampling site during a long-term study, resources spent on sampling that location will have been wasted.

“Background At the forefront of many lines of research in

“Background At the forefront of many lines of research in drug delivery are the endless possibilities of gold nanoparticles (AuNPs) [1–4]. These molecules are selleck kinase inhibitor readily taken up by cells, and they therefore provide a valuable means for drug delivery, with reports of efficient transport across the blood–brain barrier in mice [5] and nuclear penetration in the human HeLa cell line [6]. At nanoscale, the properties conferred upon such an otherwise inert metal in its bulk form are surprising. It is precisely these unique properties that offer potential KPT-8602 mouse in fields as diverse as diagnostics, anti-cancer therapies, catalysts and fuel cells. One avenue that has been

studied exhaustively in recent years is the use of coatings and capping agents in the rational design of NPs, both to stabilise and functionalise these nanoparticles. Specific capping agents can lead to the self-assembly of NPs into ordered ‘superstructures’ creating different shapes [7], and by altering the capping structure, different arrangements can be achieved. In terms of biocompatibility, when using a polyvinyl alcohol capping agent, AuNPs do not show toxicity in zebrafish, despite being taken up into embryos and evidence of bioaccumulation [8]. These observations highlight INK1197 mouse the

use of capping agents as an approach to achieve safer NPs. We recently proposed the use of peptide-biphenyl hybrid (PBH) ligands as capping agents [9]. PBHs have a biphenyl system and two amino acid/peptide fragments, and they present key characteristics, such as dynamic

properties in solution [10], ordered structures in the solid phase [11] and biological activity as calpain inhibitors [12]. Some of these properties arise from the presence of amino acid residues, as well as aromatic rings, that are able to participate in a variety of non-covalent bonds, including hydrogen bonds [13, 14] and arene interactions [15, 16]. In addition, the conformational flexibility around the aryl-aryl single bond allows the PBH to adopt its structure in order to obtain the most favourable interactions with other chemical Tryptophan synthase species, thus achieving high biological activity [17]. In peptidomimetics, this approach is considered a novel way to tailor NPs to have desired physico-chemical properties, which could contribute, for example, to advances in biomedical applications for AuNPs as drug delivery systems. A molecule can be designed in such a way as to benefit from structure-activity relationships and to attain higher levels of stability and/or biocompatibility. In a study on the design of peptide capping ligands for AuNPs, Lévy et al. [18] reported that peptide chain length, hydrophobicity and charge strongly influence NP stability. Here, we capped AuNPs with various PBH ligands and studied how the ligand structures influence the stability and the physico-chemical properties of the AuNPs under cell culture conditions and how they affect the biological response.

Thus it seems that this novel serotype has already appeared in na

Thus it seems that this novel serotype has already appeared in natural infections. Although serotype 1 d represented less than 1% of the isolates, it would be important to monitor this new serotype epidemiologically, considering that novel S. flexneri serotypes such as 1c and Xv achieved its dominance among the S. flexneri serotypes in a very short time frame [5, 16, 17] SfI and SfX integrated in tandem into the same site of host chromosome PF01367338 It has been observed that the serotype-converting phages, except for Sf6, usually integrate into the tRNA-thrW

gene of the host chromosome, which is adjacent to proA upstream [15]. However, the gene downstream the integrated phage have not been consistently identified [6, 7]. Genomic analysis of S. flexneri serotype 2a strain 301 (NC_004337), 2457 T (NC_004741) and serotype Xv strain 2002017 (CP001383) showed that the serotype-converting

phages were all integrated upstream of host gene yaiC. Thus cross-bridging PCR analyses of S. flexneri 036, 036_X, learn more and 036_1d across the proA-yaiC region were conducted using a series of primers and found that both phages SfX and SfI were integrated into the tRNA-thrW site, which is Selleckchem QNZ immediately downstream of gene proA, and upstream of gene yaiC (Figure 2). The phage SfI was found to be integrated immediately upstream of SfX genome, with an att site at both ends (Figure 2). By comparing the joining sequences between the serotype-converting phage genomes,

we found that the phage SfI was integrated at the attL site of phage SfX enough (see Additional file 1). The integration site for the 24 serotype X isolates converted by SfI was also found to be the same site and thus it appears that the integration is very site specific. Figure 2 Genetic organization of prophage genomes of SfX and/or SfI in S. flexneri 036_X and 036_1d. The prophage genomes of SfX and/or SfI are highlighted in yellow and pink respectively. The conserved genes of the host strain were shown in different colors: proA, gray; yaiC, yellow; IS600 ORF1 and ORF2, brown; IS629 ORF1 and ORF2, orange; the putative integrase gene (int), white. The integration sites attB, attL and attR are indicated in thick line. After strain name in brackets is the serotype of the strain. Conclusions A novel serotype 1 d was constructed by sequentially infecting a serotype Y strain of S. flexneri with phage SfX and SfI, or by infecting clinical serotype X isolates with SfI. These results indicate that serotype conversion with phages SfI and SfX could occur in nature. However, the observation that the order of infection by the 2 phages affects convertibility of a strain indicates that serotype conversion is not only determined by the modification specific genes but also constrained by the properties of the serotype-converting phages. Our findings provide possible mechanisms how new serotypes of S. flexneri could emerge in nature.

The most frequently PHA produced is poly(3-hydroxybutyrate) or PH

The most frequently PHA produced is poly(3-hydroxybutyrate) or PHB [2]. The ability to produce PHB has been correlated with improved survival under stress conditions or in competitive environments [5, 6]. PHB is generally produced in conditions of carbon oversupply and low levels of other nutrients such as nitrogen, phosphate or oxygen [7]. The biosynthesis of PHB is dependent on the activity of the following enzymes: (i) a 3-ketothiolase which condenses two acetyl-CoA yielding acetoacetyl-CoA (encoded by phbA), (ii) a NADPH-dependent acetoacetyl-CoA

reductase which reduces acetoacetyl-CoA to (R)-3-hydroxybutyryl-CoA AZD8931 (encoded by phbB) and (iii) the PHB synthase (encoded by phbC) that catalyses the polymerization of (R)-3-hydroxybutyryl-CoA to form the polymer [8, 9]. This polymer is stored intracellularly as insoluble inclusion bodies called PHB granules [1] which also contain about 2% protein as well as phospholipids [10]. The main protein associated with the PHB granules is phasin (encoded

by phaP) which prevents coalescence of buy AZD2171 PHB granules by coating the granule surfaces [11–14]. However, other proteins have also been found associated with the granules, including transcriptional regulators such as PhaF from Pseudomonas oleovorans GPo1, PhaR from Paracoccus denitrificans, and PhaR from Ralstonia eutropha H16 [15–17]. Expression of enzymes involved in PHA/PHB biosynthesis and the granule-associated phasin are reported to be regulated at the transcriptional level [15, 16, 18–26]. This regulation may include repressors as well as find more activators [21]. The proteins PhbR from Azotobacter vinelandii UW136 [22] and PhaD from Pseudomonas putida KT2442 [24] are transcription activators. In contrast, PhaR of P. denitrificans represses phaR expression by

binding to a TGC rich region which overlaps the -35/-10 promoter [16]. In R. eutropha H16 the PhaR protein binds to the -35/-10 phaP promoter at two sites: the transcriptional start site and upstream from the -35 at the promoter region, thereby blocking RNA polymerase [17]. The PhaR binding site determined in R. eutropha comprises two 12 bp O-methylated flavonoid repeated sequences not related to those observed in P. denitrificans, suggesting that DNA-binding sites for PhaR recognition and the mechanisms of regulation may vary. The β-Proteobacterium Herbaspirillum seropedicae SmR1 is a plant-endophytic diazotroph found in association with economically important graminaceous species such as sugar cane, sorghum, rice and maize [27]. H. seropedicae SmR1 has been already described as a PHB producer using glucose as carbon source [28], however the molecular aspects of its PHB metabolism have not been addressed. The H.