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A subset of Opa proteins bind heparan sulfate proteoglycans on epithelial cells or indirectly engage integrins by binding vitronectin (14–17). These derivatives provide a robust system for assessing the role of Opa proteins in Gc biology. 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). These results indicate that opa-deficient Gc, like Opa phase-OFF parental Gc, retains the ability to suppress PMN ROS production. Inhalte wie Buchkapitel, Beiträge in Sammelwerken und Artikel in Zeitschriften finden Sie in unserem Opac plus oder in Fachdatenbanken. Together, these results conclusively show that adherent, primed human PMNs can phagocytose Gc, in the absence of Opa protein expression or known opsonins, by an actin-dependent process. Luminescence was measured in 1-s intervals every 5 min for 30 to 60 min. Experiments were performed 3 times. When cultured in rich liquid medium, parental, ΔopaBEGK, and Opaless Gc grew with kinetics identical to those of midexponential phase, as measured by optical density (see Fig. OpaD+nv and OpaD+var Gc expressed similar amounts of Opa protein (Fig. 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. 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. Liquid - 50 oz (3.12 lb) - Free & Clear Scent - 1 / Each - Clear. Therefore, we assessed the ability of adherent, primed PMNs to internalize genetically Opa-deficient Gc by immunofluorescence. A third possibility is that Opa+ bacteria are more sensitive to PMN antimicrobial proteins. Its powerful stain-blocking formula also blocks most pet and smoke odors and creates an … Since phagocytosis of Opaless Gc requires the PMN actin cytoskeleton, PMNs may be internalizing Opa− bacteria by actin-dependent macropinocytosis. WeBoost - Agence Web. That should be in your record, but it’s still a good idea to mention it. While the magnitude of the oxidative burst was slightly larger in response to OpaD+nv Gc, this difference was not statistically significant. Opac Recherchieren, Kontoeinsicht, Vormerken, Verlängern In unserem Online-Katalog suchen Sie nach gedruckten Büchern, E-Books, Zeitschriften und audiovisuellen Medien. DPD relais s’appuie sur le réseau Pickup constitué de plus de 6000 points relais en France sélectionnés sur des critères stricts comme l’amplitude horaire (98% des points sont ouverts le samedi) et l’espace de stockage dédié. Loss of Opa expression did not affect Gc growth. Hinweise: Mit einem * können Sie beliebige Wortendungen mit suchen. Notre service DPD Relais vous permet d’offrir plus de liberté à vos clients destinataires. There was no statistical difference in survival between OpaD+nv and OpaD+var Gc (Fig. . 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. In contrast to parental and ΔopaBEGK Gc, Opaless Gc was negative for Opa protein expression by Western blotting (Fig. 3B). While it is possible that we selected for a mutation that allows for Gc growth in the absence of all Opa expression, opa-deficient derivatives of strain MS11 Gc and strain H44/76 Neisseria meningitidis also show no in vitro growth defect (19, 59). One mutant clone served as the parent for the next round of transformation, until all 11 opa genes were sequentially deleted. We circumvented this issue by creating Opaless Gc in which all opa genes were deleted from the chromosome of strain FA1090 bacteria and complementing it with a nonvariable opaD allele. Lire plus. In contrast, we reported that predominantly Opa− Gc can be internalized by adherent, IL-8-treated human PMNs (35). Bücher & mehr enthält alle Bücher, Zeitschriften, E-Books, E-Journals und Medien der PH Ludwigsburg. Given the potent ROS response of PMNs to OpaD+ Gc, one possibility is that Opa+ bacteria are killed by PMNs' extensive oxidative arsenal. 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). Opaless Gc also remained resistant to freshly isolated normal human serum, which is characteristic of strain FA1090 (51) (146% ± 27% survival of parental Gc versus 119% ± 31% survival of Opaless Gc after 30 min of incubation in 100% serum). Opaless Gc and Opa-expressing derivatives provide a robust system for studies of Opa biology and Gc pathogenesis. Approximately 60% of PMN-associated Opaless Gc was intracellular, a percentage statistically indistinguishable from the Opa− parental or ΔopaBEGK Gc (Fig. 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. Opa proteins also affect Gc phagocytosis and killing by PMNs. Fourth, Opa expression is dispensable for Gc phagocytosis by adherent, chemokine-primed human PMNs. Opa expression was validated by Western blotting with the 4B12 pan-Opa antibody (see below). https://e-multimedia.inalco.fr; 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. *, 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. The dopamine-regulated microRNA, miR-181a, controls GluA2 surface expression in hippocampal neurons. However, the Opa− Gc population in our study could have contained a subpopulation of Opa expressors that represented the internalized population. FA1090 has been used extensively in human urethral challenge studies (57) and the murine genital tract model of gonorrhea (58). *, P < 0.05 (Student's two-tailed t test) for OpaB+ Gc compared to the parent, ΔopaBEGK, or Opaless Gc at matched time points. The Neisseria gonorrhoeae (the gonococcus [Gc]) opacity-associated (Opa) proteins mediate bacterial binding and internalization by human epithelial cells and neutrophils (polymorphonuclear leukocytes [PMNs]). FA1090 1-81-S2 Gc predominantly expressing OpaB was previously described (35). 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). One clone with the opa deletion was transformed with the next opa deletion construct, until all 11 opa genes were replaced in the genome. (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). 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. 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. Découvrez tout notre assortiment ci-dessus, personnalisez votre tote bag en ligne et recevez rapidement votre commande au bureau ! XIV. 4). This UPC 18" 5mm heavy wall straight tube water pipe is a practical piece. (C) Opaless Gc, OpaD+var Gc, and OpaD+nv Gc were exposed to polymyxin B at the indicated concentrations. 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. Opa proteins are 25- to 30-kDa outer membrane proteins that were named for their ability to confer an opaque morphology on Gc colonies (6). Powered by Bow and Baan Württembergische Landesbibliothek - Konrad-Adenauer-Str. 3C. Bacterial survival.Gc survival after exposure to adherent, IL-8-primed PMNs was measured as previously described (47). S2B) and CFU/ml of culture (see Fig. Interactions of Opaless Gc with primary human PMNs. Best Seller in Fabric & Textile Paints. (i) CFU/colony.The CFU arising per colony of FA1090 Gc with and without opa deletions was enumerated as previously described (46). Bacterial survival was calculated as the CFU after 45 min of exposure, divided by the input CFU. In contrast, the Opaless FA1090 derivative described in this work grew with similar kinetics to the opa-intact parent. Your source for achieving better health through nutrition. Assuming the reduced ROS in response to OpaD+var Gc is due to a minor fraction of Opa− bacteria, these findings highlight the advantage of using Gc expressing a nonvarying opaD allele to examine Opa-dependent bacterial interactions with host cells. The University of Technology Sydney Library provides support for the teaching, learning and research needs of the students and staff at UTS. Additionally, complementation with nonvarying opaD did not change the growth characteristics of Opaless Gc. Images were acquired from a Nikon Eclipse E800 upright fluorescence microscope with a Hamamatsu digital camera using Openlab software (PerkinElmer). Percent internalization (E) was calculated as the number of intracellular Gc divided by the total number of cell-associated Gc. Interactions between Gc and human cells are modulated by a variety of bacterial surface structures, including type IV pili, lipooligosaccharide (LOS), porin, and opacity-associated (Opa) proteins (5). 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). The three strains produced similarly sized colonies (Fig. My Cloud Web access. Julie Boyer. Gc does not require Opa expression for nonopsonic internalization by primary human PMNs. Continued. Moreover, it is not always possible to identify Opa expression by colony photoopacity. 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. Assuming the minimal ROS production in PMNs exposed to parental and ΔopaBEGK Gc was due to a subpopulation of Opa+ bacteria, this finding illustrates that Opaless Gc is a preferable genetic background in situations where Opa variation influences the experimental readout. 181 p. 22 cm Date: 2010 Availability: Copies available: College Library Filipiniana Section [ PE1130 .B928 2010 ] (3), 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. Saba R, Störchel PH, Aksoy-Aksel A, Kepura F, Lippi G, Plant TD, Schratt GM. The Opa protein each expressed was determined by Western blotting with a panel of FA1090 Opa-specific antibodies (see below). Près de 270 acheteurs diffusent leurs consultations sur cette plateforme. CFU arising after nonselective transformation were screened by PCR for replacement of the full-length opa with the deletion construct. Opa proteins can also recognize lacto-N-neotetraose moieties on LOS (18, 19). (D to G) Colonies of OpaD+var Gc (D, F) and OpaD+nv Gc (E, G) were imaged as described for Fig. (B) Genomic DNA from parental, ΔopaBEGK, and Opaless Gc was used as the template for PCR, using primer sets homologous to the sequences flanking each of the 11 opa genes. The use of these derivatives provides a robust and flexible system for quantifying the effect of Opa proteins on bacteria-bacteria and bacteria-host interactions. Saba R, Störchel PH, Aksoy-Aksel A, Kepura F, Lippi G, Plant TD, Schratt GM. 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%. 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. Votre identifiant correspond à l'email que vous avez renseigné lors de l'abonnement. FA1090 opa gene deletion construction.The locations and characteristics of all opa genes in FA1090 Gc are listed in Table S1 in the supplemental material. All Content in this webport is for the using stakeholder of VTU Only. The remaining seven were then deleted to yield the complete opa deletion, here called Opaless. Differences were considered statistically significant at a P value of <0.05. 2B and E). The mutated sequence's codons appear at the same frequency in the Gc genome as the parental, phase-variable codons (http://exon.gatech.edu/metagenome/CodonUsageDatabase/display.cgi?accession=NC_002946). 6B). Opa-expressing Gc. Therefore, we conclude that Opa expression is not required for Gc growth in vitro. NLB promotes reading, learning and information literacy by providing a trusted, accessible and globally-connected library and information service through the National Library and a comprehensive network of Public Libraries. We selected Gc strain FA1090 for this study for several reasons. Um effizient in Opac plus zu recherchieren, muss Ihr Rechner am Hochschulnetz angemeldet sein. 2E to G). Blown by UPC, this water pipe comes to you from Toluca Lake CA. 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. Supplemental material for this article may be found at http://dx.doi.org/10.1128/JB.00171-13. Enter multiple addresses on separate lines or separate them with commas. Phenotypically translucent ΔopaBEGK Gc did not express any Opa proteins by Western blotting (Fig. S1B in the supplemental material), and OpaD+nv Gc retained the LOS profile of Opaless Gc (see Fig. K-LINE, fabricant français de fenêtres, baies coulissantes, portes d’entrée sur mesure en neuf et rénovation. Entrons en contact. Occurrence of color/opacity colonial variants in clinical cultures, Opa binding to cellular CD66 receptors mediates the transcellular traversal of, Neisserial binding to CEACAM1 arrests the activation and proliferation of CD4+ T lymphocytes, Submission, Review, & Publication Processes, http://exon.gatech.edu/metagenome/CodonUsageDatabase/display.cgi?accession=NC_002946, Constitutively Opa-Expressing and Opa-Deficient Neisseria gonorrhoeae Strains Differentially Stimulate and Survive Exposure to Human Neutrophils. 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. Julie Boyer Docteure en droit (PhD in law), Enseignante Paris et périphérie. (A) PMNs were left untreated or exposed to opsonized S. aureus, phenotypically translucent parental Gc (parent), ΔopaBEGK Gc, or Opaless Gc. This question is for testing whether or not you are a human visitor and to prevent automated spam submissions. 3C). Generating Opaless Gc in strain FA1090.To delete the opa genes from piliated Gc of strain FA1090, the majority of each open reading frame was replaced in frame with a restriction enzyme recognition sequence (Fig. Username. SLUth Search Plus. Zugriff auf lizenzierte E-Medien erhalten Sie im Campusnetz, Angehörige der Hochschule auch außerhalb des Hochschulnetzes via VPN oder Shibboleth. Over 2,000 bacteria were examined for their localization. Hope you guys can identify them as in the past. Les plus grandes marques de chaussures de randonnée sont Salomon, Meindl, La Sportiva, Merrell, Salewa et Scarpa. 5D to G). $6.99 $ 6. 1A). We conclude that expression of the OpaCEA OpaD renders FA1090 Gc more sensitive to PMNs. LOS preparation and Western blotting.LOS was prepared from Gc as described by Davis and Goldberg (45), except Gc was collected after overnight growth on GCB, and the protocol was stopped after the proteinase K treatment. Opa expression is phase variable due to slipped-strand mispairing at pentameric repeats in the 5′ end of opa genes (20, 21). Gefällt 1.295 Mal. --Fir0002 July 1, 2005 01:03 (UTC) That fruit reminds me of longan, but I'm not sure it's related to it.The leaves don't look right, and the rind looks sort of fuzzy. The plasmids were digested with HpaI along with a plasmid-specific restriction enzyme and ligated together to create the deletion product. Made in the U.S.A. 4.8 out of 5 stars 1,212. Investigating the contribution of Opa proteins to gonococcal pathogenesis is complicated by high-frequency phase variation of the opa genes. In support of this notion, we and others have shown that expression of CEACAM-binding Opa proteins (for instance, FA1090 OpaB) enhances the association of Gc with PMNs (30–32, 35). 3A). (A) The internal portion of each opa gene (top) was replaced in frame with a HpaI restriction enzyme recognition site (H) by overlap extension PCR, using primers with homology to the 5′ (solid line) and 3′ (dotted line) ends of opa. Approximately 0.5 to 1 kb of sequence upstream (fragment A) and downstream (fragment B) of each opa locus was amplified by PCR from FA1090 1-81-S2 P+nv genomic DNA (see Table S2 in the supplemental material for primer sequences and Table S3 in the supplemental material for PCR conditions). Other value added services offered by the Electronic Resources Section (ERS) include cd-burning, printing and document scanning. Search. Results are the averages ± standard errors from 11 to 14 replicate samples per strain. As overlap extension PCR products from opaF, opaG, and opaJ could not be generated, fragments A and B were separately cloned into pSmart or pCR-Blunt TOPO. For freshness, you possibly need to use a sturdy kitchen clip or something similar. La réponse est peut-être ici ! We thank Michael Davis for advice on overlap extension PCR and LOS preparation. However, OpaD+ Gc showed no increased sensitivity to polymyxin B, a surrogate for the cationic antimicrobial peptides produced by PMNs. 99 $12.00 $12.00. The latest wellness and longevity research. 5B). Opaless Gc behaves similarly to phenotypically Opa. Securely access your photos, music and files from anywhere. (A) Lysates from equal CFU of parental Gc, phenotypically translucent ΔopaBEGK (ΔopaBEGK translucent) Gc, phenotypically opaque ΔopaBEGK (ΔopaBEGK opaque) Gc, and Opaless Gc were incubated at 37°C or 100°C and then separated by SDS-PAGE and immunoblotted with a pan-Opa antibody. Password Substance Search. We therefore engineered a derivative of Gc strain FA1090 in which all opa genes were deleted in frame, termed Opaless. PH Bibliothek Ludwigsburg, Ludwigsburg (Ludwigsburg, Germany). Lumière, isolation, design, finesse, couleur. Here, the ΔopaBEGK Gc background was used instead of the opa-intact parent to ensure the OpaD phase-ON bacteria did not coexpress any translucent Opa proteins. No significant difference in sensitivity to polymyxin B was measured between Opaless and OpaD+ Gc (Fig. (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. S3 in the supplemental material). Opacity-associated phenotypes and growth characteristics of Opa-deficient Gc.Parental Gc and opa-deleted progeny were examined for Opa expression by Western blotting and colony photoopacity. Opaless Gc can serve as a recipient for Opa variants expressed by other strains of Neisseria or Opa proteins synthesized in vitro, in order to evaluate these proteins' receptor binding profiles and other biochemical properties. Manage and improve your online marketing. Complementation of Opaless Gc with non-phase-variable OpaD restores Opa-dependent phenotypes in Gc. For opsonization, S. aureus was incubated with 20% freshly isolated normal human serum for 20 min at 37°C. 4.4 out of 5 stars 2,707. Plant-derived cleaning agents and triple enzymes to make short work of even tough stains, but is gentle to your world; Powers through tough wash-day challenges without dyes, optical brighteners or synthetic fragrances SOCIETE.COM, service gratuit d'information sur les entreprises. 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). 3A). 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). P S. This adult dry food is great for your senior cat. --HappyCamper 1 July 2005 02:55 (UTC) To me it looks kinda like a sycamore. DIRECT. Targeting of the Arpc3 actin nucleation factor by miR-29a/b regulates dendritic spine morphology. Human PMNs in suspension can bind and phagocytose unopsonized Gc only if the bacteria express OpaCEA proteins (28–32). 5B). What am I searching? 3A. 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. Although most colonies of parental Gc were phenotypically translucent, the population contained some Opa-expressing bacteria. XIII. Parental and ΔopaBEGK Gc were predominantly Opa− by Western blotting and colony opacity (data not shown). 5C). Mapúa University, Muralla St., Intramuros, Manila 1002 Philippines © 2013-2019 All Rights Reserved 5A). S1C and D). The parental bacterial lysate contained one or more heat-modifiable proteins that reacted with a pan-Opa antibody by Western blotting (Fig. Although expression of Opa proteins like OpaD is detrimental to Gc survival after exposure to PMNs, Opa+ Gc is recovered from the male urethra and the female cervix during the follicular phase of the menstrual cycle (25, 26, 52, 62). Vous avez besoin d'aide ? Opaless Gc colonies appeared uniformly translucent (Fig. 3F11 recognizes the terminal N-acetyllactosamine of the Gc oligosaccharide α chain of LOS (18). The opaD+nv amplicon was generated by overlap extension PCR using the primers in Table S2 and conditions in Table S3 in the supplemental material and then transformed into Opaless Gc. Securely access your photos, music and files from anywhere. Experiments were performed 3 times. 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). Powered by Bow and Baan De plus, vous pouvez compter sur nous pour toujours utiliser des matériaux et une impression de la plus haute qualité. © 2020 Visvesvaraya Technological University. Most Opa-positive (Opa+) Gc stimulate primary human PMNs and promyelocytic HL-60 cells to produce ROS (28–32). In contrast, PMNs exposed to Opaless Gc were identical to uninfected PMNs and generated no ROS (Fig. Therefore, phenotypically translucent Gc may in fact express one or more Opa proteins. (C) Parental Gc that was predominantly Opa− by Western blotting (parent) or expressed the translucent OpaB (OpaB+), ΔopaBEGK Gc, and Opaless Gc were exposed to adherent, IL-8-primed PMNs. Jacquard Products Textile Color Fabric Paint 2.25-Ounce, Super Opaque White. Additionally, studies in mice have shown that Opa+ Gc is more resistant to serum bactericidal activity and to as-yet unidentified factors in genital tract secretions (50). Username. Investigating the contribution of Opa proteins to gonococcal pathogenesis is complicated by high-frequency phase variation of the opa genes. 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. 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. Opa proteins have high isoelectric points (pI of mature OpaD = 9.83 [available via http://web.expasy.org]), which may help the negatively charged bacterial surface resist binding of positively charged antimicrobial peptides. Another pressure may stem from the human adaptive immune system. 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. PCR analysis showed the four intended opa genes and no others were deleted in ΔopaBEGK Gc, while all 11 opa genes were deleted in Opaless Gc (Fig. The FA1090 genome has been sequenced and annotated (www.genome.ou.edu/gono.html; http://stdgen.northwestern.edu), the locations of all 11 opa genes in the FA1090 chromosome are known (8) (see Table S1 in the supplemental material), and the colony opacity phenotype for each FA1090 Opa protein has been elucidated (25, 53).

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