; Vous pouvez consulter sur ce serveur de podcast plus de 800 ressources pédagogiques en ligne, principalement constituées de conférences, colloques, mais aussi de documentaires, films et didacticiels sous formats audio ou vidéo. After dilution to an optical density at 550 nm (OD550) of 0.07, optical densities were measured, and CFU/ml were enumerated from each culture every hour for 3 h. Growth curves were not extended past 3 h since Gc begins to undergo autolysis as it approaches the stationary phase. Gels were silver stained using the Pierce silver stain kit (Fisher) according to the manufacturer's protocols or transferred to polyvinylidene difluoride (PVDF) and immunoblotted with the 3F11 monoclonal antibody, obtained from Michael Apicella (University of Iowa), followed by horseradish peroxidase-coupled goat anti-mouse IgM antibody (Southern Biotech). Opaless Gc behaves similarly to phenotypically Opa. Extensive database of ORAC food values. We do not retain these email addresses. The University of Technology Sydney Library provides support for the teaching, learning and research needs of the students and staff at UTS. *, P < 0.025 (Student's two-tailed t test) for Opaless Gc ± cytochalasin D. Complementation of Opaless Gc with non-phase-variable OpaD restores Opa-dependent phenotypes in Gc.Opaless Gc was complemented with a constitutively expressed allele of the FA1090 opaD gene. Hope you guys can identify them as in the past. 2B to G) and yielded equivalent numbers of CFU per colony after 24 h of growth on solid medium (see Fig. Scale bar = 10 μm. Thus, FA1090 Opaless Gc and nonvarying Opa derivatives can be used in place of predominantly Opa+ and Opa− bacteria to avoid the confounding effects of Opa variation during infection. Investigating the contribution of Opa proteins to gonococcal pathogenesis is complicated by high-frequency phase variation of the opa genes. Statistics.Differences between Gc strains were compared using two-tailed Student's t test. 181 p. 22 cm Date: 2010 Availability: Copies available: College Library Filipiniana Section [ PE1130 .B928 2010 ] (3), In this work, we addressed colonization of the nasopharyngeal niche by focusing on the interplay between meningococci and the airway mucus that lines the mucosa of the host. 4.4 out of 5 stars 2,707. All Rights Reserved. The Gram-negative bacterium Neisseria gonorrhoeae (the gonococcus [Gc]) causes the sexually transmitted infection gonorrhea, which is notable for its prevalence—approximately 106 million cases of gonorrhea occur worldwide each year—and its increasing resistance to antibiotics (1–3). OpaB expression was verified by Western blotting (see below). $6.99 $ 6. We first determined how genetically opa-deficient Gc modulated the PMN oxidative burst. Manage and improve your online marketing. We are an international, independent publisher headquartered in Berlin. La réponse est peut-être ici ! Password Potential opa-deleted clones were verified by PCR, using genomic DNA as the template and the primer combinations listed in Table S3 and/or by Southern blotting. Gc growth measurements. However, OpaD expression reduced Gc survival after exposure to primary human PMNs. Experiments were performed 3 to 4 times. The deletion construct (middle) was transformed into the chromosome of FA1090 Gc to produce opa-deleted progeny. (A) Primary human PMNs were exposed to Opaless Gc, OpaD+var Gc, or OpaD+nv Gc, and ROS production was measured as described for Fig. Bottle Height: 2.75". 1A). Scale bar = 1 mm. Username. 3A. OpaD is an OpaCEA protein that confers a strongly opaque phenotype on Gc colonies. Opaless Gc survived exposure to primary human PMNs and suppressed the PMN oxidative burst akin to parental, Opa− bacteria. Parental and ΔopaBEGK Gc were predominantly Opa− by Western blotting and colony opacity (data not shown). 6B). We gratefully acknowledge members of the Criss laboratory for their input, especially Jonathan Handing for advice on polymyxin B. SLUth searches for print books, ebooks, journals, journal articles, audio and video media, microfilm, and more including the full text for many ebooks and journal articles. Fiore R, Schratt G. EMBO J. We thank Michael Davis for advice on overlap extension PCR and LOS preparation. (A) ROS production was measured in human PMNs exposed to S. aureus, ΔopaBEGK Gc (Opa− Gc), or ΔopaBEGK Gc phase-ON for OpaA, OpaD, OpaF, or OpaI, as described for Fig. Suivez l'évolution de l'épidémie de CoronaVirus / Covid19 en France département. Opaless Gc colonies appeared uniformly translucent (Fig. To assess how expression of OpaD influenced FA1090 Gc survival after challenge with PMNs, adherent, primed human PMNs were exposed to Opaless, OpaD+var, and OpaD+nv Gc. One mutant clone served as the parent for the next round of transformation, until all 11 opa genes were sequentially deleted. Opa expression was validated by Western blotting with the 4B12 pan-Opa antibody (see below). The Neisseria gonorrhoeae (the gonococcus [Gc]) opacity-associated (Opa) proteins mediate bacterial binding and internalization by human epithelial cells and neutrophils (polymorphonuclear leukocytes [PMNs]). Powered by Bow and Baan 4). 2A), and opaque Gc colonies were detected in cultures of parental Gc (Fig. Continued. Identification of surface proteins of, Up-regulation of human neutrophil receptors for, The specific innate immune receptor CEACAM3 triggers neutrophil bactericidal activities via a Syk kinase-dependent pathway, Opa (protein II) influences gonococcal organization in colonies, surface appearance, size and attachment to human fallopian tube tissues, Multiple gonococcal pilin antigenic variants are produced during experimental human infections, An alternative DNA structure is necessary for pilin antigenic variation in, Gene splicing and mutagenesis by PCR-driven overlap extension, Gonococci exit apically and basally from polarized epithelial cells and exhibit dynamic changes in type IV pili, Purification and visualization of lipopolysaccharide from Gram-negative bacteria by hot aqueous-phenol extraction, Loss of both Holliday junction processing pathways is synthetically lethal in the presence of gonococcal pilin antigenic variation, Experimental gonococcal genital tract infection and opacity protein expression in estradiol-treated mice, Anaerobic growth of gonococci does not alter their Opa-mediated interactions with human neutrophils, Investigation of oxidative stress defenses of, A strain-specific catalase mutation and mutation of the metal-binding transporter gene, Experimental gonococcal infection in male volunteers: cumulative experience with, Estradiol-treated female mice as surrogate hosts for, Construction of Opa-positive and Opa-negative strains of, Macropinocytosis: an endocytic pathway for internalising large gulps, Antigonococcal activity of human neutrophil cathepsin G, Studies on gonococcus infection. Rebond de l'épidémie de Covid-19 : le maire de Reims "propose de reconfiner" après Noël et jusqu'à... 22/12/2020 08:26:33. Loss of opa is indicated by the faster-migrating PCR product for each primer set. Most Opa proteins interact with one or more human carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) (OpaCEA) (reviewed in reference 13). The parent of Opaless and OpaD+ nonvariable (OpaD+nv) Gc has a point mutation in the G-quadruplex region upstream of pilE that abrogates pilin antigenic variation (40) (FA1090 1-81-S2 P+nv), to ensure all bacteria express the same pilin. It's a collection of multiple types of lists used during security assessments, collected in one place. Serum survival.Gc grown in supplemented GCBL was pelleted at 10,000 × g for 3 min. Opa+ Gc may show more sensitivity to killing by PMN serine proteases, which have antigonococcal activity in vitro and in PMN phagosomes (48, 61), as Opa expression makes Gc more sensitive to the serine proteases trypsin and chymotrypsin (50). Opa proteins can also recognize lacto-N-neotetraose moieties on LOS (18, 19). Sensitivity of OpaD+ Gc to killing by human PMNs but not polymyxin B. Top Requirement: 20/410. 2011 Oct 19;30(20):4116-8. doi: 10.1038/emboj.2011.356. The latest wellness and longevity research. When a non-phase-variable, in-frame allele of FA1090 opaD was reintroduced into Opaless Gc, the bacteria induced the PMN oxidative burst, and OpaD+ Gc survived less well after exposure to PMNs compared to Opa− bacteria. Substance Search. One of the main reasons for creating Opaless Gc was to avoid changes in expression of Opa proteins during bacterial culture that adversely affect Gc-host interaction studies. While translucent colonies were occasionally detected in the OpaD+var Gc population, OpaD+nv Gc colonies were uniformly opaque (Fig. Thus, Gc does not need to express Opa proteins to be internalized by PMNs in this assay system. (C) Opaless Gc, OpaD+var Gc, and OpaD+nv Gc were exposed to polymyxin B at the indicated concentrations. These in vivo observations indicate that factors other than PMNs select for bacterial expression of Opa proteins during acute infection. Your session has expired. *, P < 0.05 (Student's two-tailed t test) for Opaless Gc compared to either OpaD+var Gc or OpaD+nv Gc at matched time points. While Opa variation may enable bacterial evasion of host immunity and adaptation to new hosts or sites within a host, it complicates the assessment of how Opa expression influences specific aspects of Gc pathogenesis. XIV. ROS production was measured as luminol-dependent chemiluminescence produced over 1 h. (B) PMNs were left untreated or exposed to opsonized S. aureus, alone or with an equivalent multiplicity of infection (MOI) of phenotypically translucent parental Gc (parent), ΔopaBEGK Gc, or Opaless Gc. Twenty nine (29) units of computers are dedicated for internet, online database searching and document processing. Given the small-colony phenotype of a previously constructed FA1090 opa deletion strain (50), we measured the in vitro growth of FA1090 parental, ΔopaBEGK, and Opaless Gc. Gc does not require Opa expression for nonopsonic internalization by primary human PMNs.Several groups have shown that PMN phagocytosis of unopsonized Gc requires bacterial expression of OpaCEA proteins (28–32). Bacterial survival over time was calculated as the CFU enumerated from PMN lysates at each time point, divided by the CFU present at 0 min. Finally, the contribution of putative (non-Opa) Gc virulence factors can be investigated in constitutively Opa+ or Opa− genetic backgrounds, to determine if and how the presence of Opa affects their function. The mutated sequence's codons appear at the same frequency in the Gc genome as the parental, phase-variable codons ( Consultez les chiffres clés, l'identité, les dirigeants de toutes les entreprises immatriculées au RNCS (Registre National du Commerce des Sociétés). A few customers have noted that the seal on the bag doesn't close properly. The remaining seven were then deleted to yield the complete opa deletion, here called Opaless. (ii) Constitutively expressed OpaD+ Gc (OpaD+nv).The CTCTT repeat-rich region of FA1090 opaD was replaced with a sequence encoding the same amino acids but unable to undergo phase variation (see Fig. We took advantage of the redundancy of the genetic code to engineer an OpaD allele in which the CTCTT repeat tract was replaced with a DNA sequence that encoded the same amino acids but cannot phase vary (Fig. New Look, Same Trusted Formula, KILZ Original 13 New Look, Same Trusted Formula, KILZ Original 13 oz. (ii) Liquid medium.Gc was grown in GCBL modified with Kellogg's supplements I and II and 0.042% sodium bicarbonate as previously described (33). 1B). This effect was likely due to a subset of bacteria that phase varied ON Opa expression in the parental and ΔopaBEGK Gc cultures. Study Habits and Academic Performance in English Plus of Freshmen Maritime Students of LB By: Dr. Precy Guerra (2007) Show Abstract Status and Prospect for the Sustainable Development of the Maritime Education Program of LB By: Dr. Rolando Lontoc Sr. (2007) Show Abstract. Therefore, phenotypically translucent Gc may in fact express one or more Opa proteins. These results indicate that opa-deficient Gc, like Opa phase-OFF parental Gc, retains the ability to suppress PMN ROS production. Dr. Ph. Human PMNs in suspension can bind and phagocytose unopsonized Gc only if the bacteria express OpaCEA proteins (28–32). 6C). [Epub ahead of print] Releasing a tiny molecular brake may improve memory. Arrows point to translucent colonies in the OpaD+var population. FA1090 1-81-S2 Gc predominantly expressing OpaB was previously described (35). OpaD-expressing Gc survived significantly less well after PMN challenge compared with Opaless bacteria (Fig. 4D). These derivatives provide insight into Opa-mediated interactions of Gc with human PMNs and make a system available for uniformly investigating the contribution of Opa proteins to Gc biology and pathogenesis. We thank Joanna Goldberg for critical reading of the manuscript. Human feces is the solid or semisolid remains of food that could not be digested or absorbed in the small intestine of humans, but has been further broken down by bacteria in the large intestine. For subsequent experiments, parental Gc was subcloned to yield an isolate that was predominantly Opa negative (Opa−) by Western blotting and colony morphology (data not shown). Découvrez tout notre assortiment ci-dessus, personnalisez votre tote bag en ligne et recevez rapidement votre commande au bureau ! Opa proteins also affect Gc phagocytosis and killing by PMNs. TOP10 E. coli (Invitrogen) was grown in Luria-Bertani (LB) (BD Difco) broth or agar supplemented with 50 μg/ml kanamycin (Sigma-Aldrich) at 37°C for plasmid propagation. No differences were observed in protein or LOS composition among parental, ΔopaBEGK, and Opaless Gc (see Fig. Lun-Ven 9h30-19h [email protected] nous contacter. Please login again. Bacterial survival at each concentration was calculated as the ratio of the number of CFU recovered after 45 min divided by the number of CFU present at the start of the experiment and is presented relative to survival in the absence of polymyxin B (100%). Experiments were performed 3 times. These and other approaches will help reveal the contribution of Opa proteins, individually or in combination, to Gc-host interactions and enhance understanding of how these interactions influence the course of human disease. Opaless Gc and an intermediate isolate in which the genes encoding translucent Opa proteins were deleted (ΔopaBEGK) behaved similarly to Opa phase-OFF bacteria in in vitro growth and survival after exposure to primary human PMNs. CFU arising after nonselective transformation were screened by PCR for replacement of the full-length opa with the deletion construct. Pendant la crise sanitaire, 70 % des parents ont ressenti plus de fatigue que d’habitude 22/12/2020 08:45:34. Ebenso Texte aus dem E-Repository der Hochschule und aus den Nationallizenzen.Die Verfügbarkeit der einzelnen Titel wird in der Trefferliste angezeigt. Saba R, Störchel PH, Aksoy-Aksel A, Kepura F, Lippi G, Plant TD, Schratt GM. This observation suggested that Opa expression, even residual expression in predominantly Opa− bacteria, was required for Gc growth. 5A). To begin to understand the mechanism underlying the increased killing of OpaD+ Gc by PMNs, we measured bacterial survival after exposure to the model cationic peptide polymyxin B. For freshness, you possibly need to use a sturdy kitchen clip or something similar. Bacteria were resuspended in 100% freshly isolated normal human serum for 30 min. Search. In contrast, we reported that predominantly Opa− Gc can be internalized by adherent, IL-8-treated human PMNs (35). PMN oxidative burst.Reactive oxygen species generation from PMNs exposed to Gc was measured by luminol-dependent chemiluminescence, using a Victor 3 multilabel plate reader (Perkin-Elmer), as described in reference 33. It also contains bacteria and a relatively small amount of metabolic waste products such as bacterially altered bilirubin, and the dead epithelial cells from the lining of the gut. Putative transformants were selected by colony photoopacity, and successful reintegration of opaD+nv into the opaD locus of Opaless Gc was confirmed by PCR. NLB manages the National Library, 26 Public Libraries and the National Archives. Interestingly, there was a small increase in ROS production from PMNs exposed to cultures of predominantly translucent parental and ΔopaBEGK Gc, whereas Opaless Gc stimulated no detectable ROS from PMNs. To examine the survival of genetically opa-deficient Gc after PMN challenge, adherent, IL-8-primed human PMNs were infected with ΔopaBEGK and Opaless Gc. 6A). There was no statistical difference in survival between OpaD+nv and OpaD+var Gc (Fig. Opaque colonies arising in the predominantly translucent culture were streak purified and stored at −80°C. The Opa protein each expressed was determined by Western blotting with a panel of FA1090 Opa-specific antibodies (see below). Study Habits and Academic Performance in English Plus of Freshmen Maritime Students of LB By: Dr. Precy Guerra (2007) Show Abstract . (B) The 5′ end of FA1090 opaD was mutated to replace the phase-variable CTCTT repeats (line above the DNA sequences) with nucleotides encoding the same amino acids (written below the DNA sequences) yet unable to phase vary (black boxes). © 2020 Visvesvaraya Technological University. All Content in this webport is for the using stakeholder of VTU Only. For Southern blotting, genomic DNA was restriction digested (AvaI for opaA, C, D, G, and J; ClaI for opaB; SalI for opaE; XbaI for opaI; XbaI and XhoI for opaK; all restriction enzymes were from New England BioLabs) and probed with an opa-specific gene probe labeled with digoxigenin (Roche). Opa+ colonies (arrows) are optically reflective when illuminated with the mirrored substage and dark in color when illuminated with the frosted substage. Tous les décès depuis 1970, évolution de l'espérance de vie en France, par département, commune, prénom et nom de famille ! DIRECT. Global incidence and prevalence of four curable sexually transmitted infections (STIs): new estimates from WHO, The 2nd Global HIV/AIDS Surveillance Meeting, A bacterial siren song: intimate interactions between, Studies on gonococcus infection. Création de site internet, Référencement Google, Publicité sur les réseaux sociaux. Absence of Opa protein expression in Opaless Gc. Welcome to Common Chemistry ™ from Chemical Abstracts Service (CAS), a web resource that contains CAS Registry Numbers for approximately 7,900 chemicals of widespread general public interest. - danielmiessler/SecLists Phenotypically translucent ΔopaBEGK Gc did not express any Opa proteins by Western blotting (Fig. De Gruyter publishes first-class scholarship and has done so for more than 270 years. Opa expression also facilitates Gc transcytosis of polarized epithelial monolayers, allowing the bacteria to gain access to subepithelial regions (64). We previously showed that OpaB+ Gc survives less well than Opa− bacteria after PMN exposure (35). Second, genetically opa-deficient Gc survives exposure to primary human PMNs and suppresses the PMN oxidative burst, analogous to predominantly Opa− bacteria. Other value added services offered by the Electronic Resources Section (ERS) include cd-burning, printing and document scanning. Bacterial survival.Gc survival after exposure to adherent, IL-8-primed PMNs was measured as previously described (47). Mit dem Onlinekatalog recherchieren Sie (nahezu) rund um die Uhr in den Beständen der Stadtbibliothek, stöbern in unseren neu erworbenen Titeln oder entdecken Medientipps zu aktuellen Themen. Opa+ Gc engagement of CEACAM1 also leads to B cell death (67). CFU/ml were enumerated from the culture before and after serum exposure, and bacterial survival was expressed as (CFU/ml)t = 30 min/(CFU/ml)t = 0 min × 100%. Inhalte wie Buchkapitel, Beiträge in Sammelwerken und Artikel in Zeitschriften finden Sie in unserem Opac plus oder in Fachdatenbanken. Mapúa University, Muralla St., Intramuros, Manila 1002 Philippines © 2013-2019 All Rights Reserved We therefore created a derivative of strain FA1090 Gc carrying in-frame deletions in all opa genes, which is termed Opaless. Opaless Gc remained uniformly Opa negative (Opa−), whereas cultures of predominantly Opa− parental Gc and an intermediate lacking the “translucent” subset of opa genes (ΔopaBEGK) stochastically gave rise to Opa-positive (Opa+) bacterial colonies. Feedback on Strategic Plan, FET JU Congratulation message of Hon'ble MIC for the good academic attainments of the students of Jadavpur University in the 2018 All India GATE examination and also for performing exceedingly well in the Association of Indian University's East … Immunofluorescence.Binding and phagocytosis of Gc after exposure to interleukin-8 (IL-8)-primed PMNs, attached to serum-coated glass coverslips, were measured as previously described (48). Its powerful stain-blocking formula also blocks most pet and smoke odors and creates an … 3C. Thus, reintroduction of a nonvarying opa gene into Opaless Gc allows for constitutive Opa protein expression, without affecting Gc growth. This UPC 18" 5mm heavy wall straight tube water pipe is a practical piece. Pädagogische Hochschulbibliothek Ludwigsburg. We thank Christof Hauck, Ann Jerse, and Michael Apicella for providing antibodies and the University of Virginia Lymphocyte Culture Center for production of the 4B12 monoclonal antibody. 2D, G) and remained translucent for 25 nonselective passages (data not shown). When nonselectively passaged, ΔopaBEGK Gc sporadically yielded opaque colonies (Fig. While the magnitude of the oxidative burst was slightly larger in response to OpaD+nv Gc, this difference was not statistically significant. (A) PMNs were left untreated or exposed to opsonized S. aureus, phenotypically translucent parental Gc (parent), ΔopaBEGK Gc, or Opaless Gc. List types include usernames, passwords, URLs, sensitive data patterns, fuzzing payloads, web shells, and many more. The construct was transformed into Opaless Gc to yield OpaD+nv (nonvariable) Gc. Screening and validation of opa gene deletions in FA1090 Gc.Deletion constructs were sequentially introduced into FA1090 Gc by natural transformation on a solid medium (43). 3A). The plasmids were digested with HpaI along with a plasmid-specific restriction enzyme and ligated together to create the deletion product. Il appartient au groupe 13 du tableau périodique ainsi qu'à la famille des métaux pauvres.. 3A. The translucent Opa proteins confer a poorly opaque phenotype on Gc colonies grown on commonly used media, and Opa expression can be reliably detected only by immunoblot (38). Care was taken to minimize time between PMN isolation and use in experiments. A subset of Opa proteins bind heparan sulfate proteoglycans on epithelial cells or indirectly engage integrins by binding vitronectin (14–17). Fourth, Opa expression is dispensable for Gc phagocytosis by adherent, chemokine-primed human PMNs. Experiments were performed 3 times. Gc-PMN coculture.All human subject research was conducted in accordance with a protocol approved by the University of Virginia Institutional Review Board for Health Sciences Research. Jacquard Products Textile Color Fabric Paint 2.25-Ounce, Super Opaque White Julie Boyer. Opaless Gc and Opa-expressing derivatives provide a robust system for studies of Opa biology and Gc pathogenesis. 10 - 70173 Stuttgart - Telefon +49 711-212-4454 P S. This adult dry food is great for your senior cat. Gc constitutively expressing the OpaCEA OpaD recapitulated phenotypes associated with OpaD phase-ON Gc in induction of the PMN oxidative burst and bacterial sensitivity to PMNs. Each of the 11 transformation events that yielded Opaless occurred at a frequency of ≥0.02 per CFU, which is similar to the frequency we measure for transformation of non-opa constructs (our unpublished observations). (i) CFU/colony.The CFU arising per colony of FA1090 Gc with and without opa deletions was enumerated as previously described (46). Activated PMNs assemble the NADPH oxidase enzyme to generate reactive oxygen species (ROS), e.g., the oxidative burst (27). TPL is one of the world's busiest urban public library systems, with 100 branches across the city and over 10 million books, movies, and other items to borrow or download online. Neisseria meningitidis is an inhabitant of the nasopharynx, from which it is transmitted from person to person or disseminates in blood and becomes a harmful pathogen. Polymyxin B survival.Mid-logarithmic phase Gc was exposed to increasing concentrations of polymyxin B (Alexis Biochemicals) as described in reference 49, except that incubations occurred in 0.5× GCBL. All Content in this webport is for the using stakeholder of VTU Only. Opaless, ΔopaBEGK, and parental Opa phase-OFF Gc were statistically indistinguishable from PMNs in their survival profile, and all survived significantly better than OpaB+ bacteria (Fig. Opaless Gc behaves similarly to phenotypically Opa− bacteria in interactions with primary human PMNs.To compare the functionality of opa-deleted Gc to Opa phase-OFF bacteria, we assessed parental, ΔopaBEGK, and Opaless Gc interactions with primary human PMNs. . However, OpaD+ Gc showed no increased sensitivity to polymyxin B, a surrogate for the cationic antimicrobial peptides produced by PMNs. Investigating the contribution of Opa proteins to gonococcal pathogenesis is complicated by high-frequency phase variation of the opa genes. Introducing a non-phase-varying allele of OpaD from strain FA1090 into Opaless Gc allowed us to examine the effect of constitutive Opa protein expression on Gc interactions with human PMNs. Martin's 400032-XXX Bleedproof White, 1.0 oz. TISS is a unique institution that brings together high quality scholars and practitioners from Social, Economic, Political, Physical, Habitat, Engineering, Health, and Environmental Sciences to create interdisciplinary teaching and research programmes combined with field action to address the most critical current and emerging issues of the nation.