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Reduced polysialic acid capsule expression in Escherichia coli K1 mutants with chromosomal defects in kpsF.

Neuroinvasive Escherichia coli K1 synthesizes and assembles a polysialic acid capsule virulence factor on the external leaflet of the outer membrane. This capsule functions in pathogenesis by blocking non-immune host defence mechanisms and acting as a relatively non-immunogenic molecular mimic of the polysialic acid chains found in high concentrations on neural cell adhesion molecules of the human embryo and neonate. The synthetic, regulatory and export components for capsule expression are encoded in three functionally distinct gene blocks or regions of the 20 kb kps 'pathogenicity island'. These regions are organized as two convergently transcribed operons inserted into the monocistronic tRNA gene, pheV. The six genes of the so-called region 1 operon are transcribed in the same direction as pheV, and at least four of these genes are required for polysialic acid export. Expression of this operon is thermoregulated by transcriptional control of its first gene, kpsF. To investigate the function of region 1 further, two independent chromosomal disruptions were engineered by inserting promoterless, terminatorless kanamycin or chloramphenicol resistance cassettes into the HindIII site of the kpsF coding sequence. The chromosomal insertions were regulated by temperature in the same way as the wild-type operon, demonstrating that this control mechanism remained intact in these mutants. Chemical, immunological and ultrastructural microscopical methods demonstrated that full-length polysialic acid chains were synthesized but not exported by the kpsF mutants. This phenotype was correlated with decreased plaque diameter when the mutants were infected with the capsule-specific bacteriophage K1F. The export defect could not be complemented in trans with kpsF+ containing its cis-regulatory region because of titration of an apparent positive regulator of region 1 expression, whereas complementation was observed with a plasmid expressing kpsF from a physiologically irrelevant promoter. An N-terminal polyhistidine peptide was attached to KpsF and used to purify the overproduced polypeptide. Antibodies raised against KpsF identified at least one of its paralogues in E. coli, GutQ, suggesting that KpsF and its homologues are membrane associated. The results indicate the requirement for a precise balance between region 1 components of the capsule export machinery, and that KpsF plays a positive role in the assembly, operation or regulation of this apparatus.

Bacterial Proteins↗

Transcriptional organization and regulation of the Escherichia coli K30 group 1 capsule biosynthesis (cps) gene cluster.

Escherichia coli group 1 capsules are important virulence determinants, yet little is known about the transcriptional organization or regulation of their biosynthetic (cps) operons. Transcription of the prototype serotype K30 cluster is modulated by the JUMPStart-RfaH antitermination mechanism, with the cps promoter being localized to a region immediately upstream of the JUMPStart sequence. A putative stem-loop structure located within the K30 cps cluster separates conserved genes with products that are required for surface expression of capsule from serotype-specific genes encoding enzymes for polymer repeat-unit synthesis and polymerization. This putative stem-loop structure significantly reduces transcription in a termination-probe vector and may contribute to differential expression of the cps genes. Previous work indicated that increased amounts of group 1 capsular polysaccharide synthesis resulted from the overexpression of the Rcs (regulator of capsule synthesis) proteins. However, neither overexpression of the transcriptional activator RcsB nor an rcsB::aadA chromosomal insertion altered the level of transcription measured by cps::lacZ fusions. In the group 1 strains examined, an RcsAB box was found immediately upstream of galF, a gene involved in the production of sugar nucleotide precursors. Overexpression of RcsB was found to result in a threefold increase in transcription of a galF::lacZ chromosomal fusion. Moreover, overexpression of GalF gave rise to a two- to threefold increase in cell-free as well as cell-associated capsule, without affecting cps::lacZ activity. These results indicate that transcription of the E. coli group 1 capsule cluster itself is not regulated by the Rcs system and may, in fact, be constitutive. However, the Rcs system can potentially influence levels of capsular polysaccharide production by increasing galF transcription and influencing the available pool of biosynthetic precursors.

Bacterial Proteins↗

[Histological observations of the capsules induced by Contortylenchus diplogaster v. Lins (Nematoda: Allantonematidae) in the adipose tissue of Ips sexdentatus Boern. (Coleoptera: Scolytidae)].

The evolution of the capsules induced by C. diplogaster in the fat body of I. sexdentatus is followed by histological methods from the pupal stage to the oviposition of the host. Early in the pupal stage of the insect, the parasitic female of C. diplogaster has not oviposited yet and the nuclei of the capsule that surrounds it are gathered in tetrads; some of these nuclei are in a well forwarded degenerescence. These observations suggest that the syncytial capsule should result both from an amitotic division of the nuclei followed by a degenerescence of some of them and from a fusion of cells. Late in the pupal stage, the nuclei are set apart from each other and distributed at the periphery of the capsule. The oviposition of the nematode takes place when the host is an adult. After the development of the larvae has been made into the capsule, this one ends in breaking, generally when the bark beetle has attacked a new host-tree. Then, the larvae of the nematode leave the insect and invade the galleries. The parasitic development of C. diplogaster always takes place quite in the fat body of the beetle.

Adipose Tissue↗

Extraction of entrapped capsules from the small bowel by means of push-and-pull enteroscopy with the double-balloon technique.

The new technique of push-and-pull enteroscopy using the double-balloon technique (double-balloon enteroscopy) makes it possible to remove swallowed foreign bodies causing intestinal obstruction deep in the small bowel without the need for surgical laparotomy. This report describes two cases of enteroscopic removal of entrapped capsules. In one patient with acute recurrent intestinal bleeding and recurrent abdominal pain, Crohn's disease had been suspected on capsule endoscopy. The second patient, with known Crohn's disease, was suffering from abdominal pain and underwent capsule endoscopy for investigation of the small bowel. Prior enteroclysis had not revealed stenoses in either patient. An oral approach was chosen and the capsules were identified approximately 140 cm and 310 cm from the pylorus, respectively, in front of stenoses. The capsule endoscopes were removed successfully in both patients by means of push-and-pull enteroscopy using a new enteroscope device.

Device Removal↗

The yield of capsule endoscopy in patients with abdominal pain or diarrhea.

BACKGROUND AND STUDY AIMS: Capsule endoscopy, proven effective for evaluation of obscure gastrointestinal bleeding and suspected Crohn's disease, is increasingly used to investigate other small-intestine disorders, but its yield for other indications is not well known. We sought to evaluate its yield and findings for abdominal pain or diarrhea. PATIENTS AND METHODS: Medical records of patients with abdominal pain or diarrhea (> 6 weeks' duration) who underwent capsule endoscopy between August 2001 and June 2004 were retrospectively reviewed for demographic data, indications, findings, diagnoses, complications, and radiologic studies. All patients had previous endoscopic or radiologic examinations (colonoscopy, enteroscopy, upper endoscopy, small-bowel series, computed tomography enterography, or computed tomography) demonstrating no abnormalities sufficient for diagnosis. RESULTS: 64 patients (26 men; 38 women; mean age, 43 years; age range, 19 - 83 years) who met study criteria had 68 capsule endoscopy studies. Indications were abdominal pain (35 patients), diarrhea (14), or both (15). Complete small-bowel visualization with identification of the cecum was achieved in 81 %; yield of positive findings was 9 % (6 patients). By indications, the yield was 6 % for abdominal pain, 14 % for diarrhea, and 13 % for both. Diagnoses included Crohn's disease (3), enteropathy induced by nonsteroidal anti-inflammatory drugs (2), and submucosal tumor (1). Capsule retention occurred in two patients, requiring surgical removal. CONCLUSIONS: Capsule endoscopy had a low yield for evaluation of abdominal pain or diarrhea and cannot be recommended as a first-line test without further study. Nonetheless, it facilitated diagnosis in 9 % of patients with negative endoscopic and radiologic examinations.

Abdominal Pain↗

[Results of mobilization under anesthesia in adhesive capsulitis in relation to stage of the disease].

PROBLEM: Mobilisation under anaesthesia is a well approved method for the treatment of idiopathic adhesive capsulitis, the best time to perform the mobilisation, however, is difficult to define. MATERIAL AND METHODS: Thirty-nine patients who had been treated with mobilisation under anaesthesia for idiopathic adhesive capsulitis stage II or III were examined at an average follow-up time of 3.4 years. RESULTS: This therapy failed in 10.3% of the patients. There were no differences in the follow-up results between both groups of patients, limitations in the range of motion remained in 20% of all patients. At follow up the average CONSTANT Score of the stage II patients was 86.8, that of the stage III patients was 89.9 points. However, manipulation under anaesthesia showed a significantly better influence on the course of the disease in patients with stage III than in patients with stage II adhesive capsulitis. CONCLUSION: Mobilisation under anaesthesia is a successful therapy in adhesive capsulitis stage III, but should be performed in adhesive capsulitis stage II only in cases of decreasing pain in which the limitation in the range of motion is the main symptom.

Adult↗

Experimental and computational probes of the space in a self-assembled capsule.

Self-assembled capsules are hosts that recognize and surround smaller molecule guests of appropriate size, shape, and chemical surfaces. The space available inside is a cage of fixed solvent molecules, many of which are aromatic. These aromatics provide anisotropic shielding to guests, and a map of induced magnetic shielding for the inner space can be obtained through nucleus-independent chemical shift calculations. Experimental values of the magnetic environment can be determined by NMR spectra of the guests inside. We describe here the environment in a cylindrical capsule with tapered ends. A series of terminal acetylenes -- the narrowest of organic structures -- was synthesized and used to probe the magnetic shielding of the capsule's ends. Their NMR spectra showed that the acetylenic hydrogen experiences deshielding as it is forced deeper into the tapered end of the capsule where four benzene rings converge. Modeling and density functional theory calculations provided excellent agreement with the experimental values and established a molecular ruler to explore steric and magnetic environments inside the capsule.

Acetylene↗

Critical role of the group A streptococcal capsule in pharyngeal colonization and infection in mice.

To study the role of the group A streptococcal capsule in pharyngeal colonization, we used two acapsular mutants derived from a type 24 strain of group A Streptococcus by transposon mutagenesis. One mutant had a stable acapsular phenotype due to a transposon-associated chromosomal deletion of essential capsule synthetic genes, while the second mutant could revert to the encapsulated phenotype at a low frequency (< 10(-4)) upon spontaneous excision of the transposon from the capsule-synthesis region of the chromosome. Both acapsular mutants were sensitive to phagocytic killing in vitro and had reduced virulence in mice after intraperitoneal challenge. Mice inoculated intranasally with the stable acapsular mutant rapidly cleared the organisms from the pharynx, and no mice died. In contrast, throat cultures of animals challenged with the revertible mutant yielded many encapsulated revertants, and mortality was similar to that of animals challenged with the parent strain. The rapid emergence of a population of encapsulated revertants in the pharynx implies that the capsule conferred a powerful selective advantage in this environmental niche. Together with the complete avirulence of the stable acapsular mutant, these observations indicate that the hyaluronic acid capsule plays a critical role in colonization and infection of the pharynx by group A streptococci.

Animals↗

Effect of growth factors on the activation of human Tenon's capsule fibroblasts.

PURPOSE: To investigate stimulatory effects of PDGF-AA, PDGF-AB, PDGF-BB, bFGF, IL-1beta, TGF-beta1 and TGF-beta2 on the proliferation and myofibroblast transformation of cultured human Tenon's capsule fibroblasts and to characterize expression of PDGF- and TGF-beta-receptors in these cells. METHODS: To determine cell proliferation, cell number of 2nd passage cultured human Tenon's capsule fibroblasts was measured before and after addition of growth factors using a computer-based cell counter system. Immunoblotting was used to detect and quantitate alpha-smooth-muscle actin (alpha-SMA) expression. Expression of PDGF- and TGF-beta-receptor mRNA was detected by RT-PCR, expression of the corresponding protein was demonstrated using Western blot. RESULTS: A significant increase in proliferation (p < or = 0.05) was detected after exogenous stimulation with PDGF-AA (10 ng/ml and 100 ng/ml), PDGF-AB (10 ng/ml and 100 ng/ml), PDGF-BB (10 ng/ml and 100 ng/ml), bFGF (100 ng/ml), IL-1beta (1 ng/ml and 10 ng/ml), TGF-beta1 (0.5 ng/ml) and TGF-beta2 (0.5 ng/ml). Both TGF-beta1 and TGF-beta2 stimulated expression of alpha-SMA in a dose dependent manner with peak activity at a concentration of 50 ng/ml (TGF-beta1) and 500 ng/ml (TGF-beta2). Protein and mRNA of PDGF-receptor type alpha and type beta and TGF-beta-receptors type I, II and III are expressed in cultured human Tenon's capsule fibroblasts. CONCLUSIONS: The present investigation strongly supports the hypothesis that PDGF-isoforms are major stimulators of proliferation of Tenon's capsule fibroblasts after glaucoma filtering surgery while TGF-beta-isoforms are essential for the transformation of Tenon's capsule fibroblasts into myofibroblasts.

Actins↗

Capsule endoscopy: role of bowel preparation in successful visualization.

BACKGROUND: Lesions of the small bowel that cannot be detected by other techniques can be indentified by capsule endoscopy. Good preparation before any endoscopic procedure is the most important factor for patient safety, quality of care and cost effectiveness. This should also apply to capsule endoscopy. The aim of this study was to evaluate the quality and efficacy of preparation with oral sodium phosphate for capsule endoscopy. METHODS: A retrospective cohort study design was used. The study sample consisted of 32 consecutive patients scheduled for capsule endoscopy in two periods; the first 10 patients were prepared with overnight fasting only, and the next 22 with oral sodium phosphate. An experienced endoscopist and gastroenterology nurse, who were blinded to the method used, graded the quality of preparation. RESULTS: Small-bowel cleansing was significantly better in patients given sodium phosphate. Five patients (50%) received a poor preparation rating in the first period compared with only 1 (4.5%) of the 22 patients given oral sodium phosphate (P = 0.01). Intraluminal fluid was noted in 9 patients (90%) in the first group after a mean of 53 +/- 60 min, and in 17 patients (77%) in the second group after a mean of 158 +/- 71 min (P < 0.001). The proportion of non-ideal preparation was significantly higher in group 1 than in group 2, significant visualization disturbances being more prevalent in the first period. CONCLUSIONS: Bowel preparation with sodium phosphate before capsule endoscopy offers better visualization than overnight fasting alone and induces fewer disturbances by intraluminal turbid fluid.

Administration, Oral↗

Demonstration of the sustained anthelmintic activity of a controlled-release capsule formulation of ivermectin in ewes under field conditions in New Zealand.

Ten field trials were conducted in the North and South Islands of New Zealand to evaluate the anthelmintic efficacy of an intraruminal controlled-release capsule formulation of ivermectin. A total of 810 Coopworth, Perendale, Romney or Coopworth ' Romney ewes, weighing on average 42-70 kg, were used. Ewes were either untreated or treated shortly before lambing in late winter-early spring (eight trials) or in late spring (two trials) with an ivermectin controlled-release capsule which delivers ivermectin at 1.6 mg per day for 100 days (minimum dose rate 20microg/kg/day). Bodyweights, faecal nematode egg counts and dag scores were determined before treatment and at about 2 and/or 4, 6 or 8, 10 or 12, 14 and 16 weeks after treatment. Ewes treated with the ivermectin controlled-release capsule gained on average 1.1kg more than untreated sheep over the 16 weeks of the trials, but this difference was not significant (p > 0.10). Before treatment, faecal strongylid egg counts were equivalent (p > 0.10), but at each time point thereafter, egg counts in ivermectin controlled-release capsule treated sheep were significantly lower (p < 0.01; p < 0.05 at Week 2). Dag scores were not different at the start of the trial (p > 0.10), but at the end of the trial ivermectin controlled-release capsule treated ewes had significantly lower scores (p < 0.01) than untreated ewes. These findings indicated that treated animals shed significantly fewer nematode eggs and therefore pasture contamination with nematode eggs should be significantly reduced for at least 112 days. The control of dags should result in reduced direct losses due to the decreased value of dag wool, and indirect losses due to the cost of dagging sheep and the cost associated with the treatment and control of flystrike initiated by dags in the breech area.

Journal Article↗

An electron microscopic India ink technique for demonstrating capsules on microorganisms: studies with Streptococcus pneumoniae, Staphylococcus aureus, and Neisseria gonorrhoeae.

A technique using India ink in electron microscopic preparations was used to study bacterial capsules. Capsules were demonstrated on Streptococcus pneumoniae and Staphylococcus aureus (strain M and the Smith diffuse variant) from in vitro cultures. Two types of false capsules were observed on Neisseria gonorrhoeae grown in vitro or obtained from human urethral exudates. No true capsules were definitively identified on gonococci from these sources. The technique described does not require the use of specific anticapsular antibody or specific polysaccharide stains and permits the detection of artifacts that may, by the use of light microscopic methods, be misinterpreted as capsules.

Antigens, Bacterial↗

The effects of hematoma on implant capsules.

Hematoma surrounding an implant is one of the many factors that have been suggested as possible causes for scar capsule contracture. In this study, experiments were designed to determine the influence of hematoma on the incidence and severity of capsule contracture in rabbits. Two implants were placed in each animal, 1 with a surrounding hematoma and 1 control. Capsules were evaluated subjectively and compared objectively with measurements of deformability, surface area, and capsule thickness. No differences were found with any of the objective criteria, which suggests that hematoma may not be a noteworthy cause of implant capsule contracture.

Animals↗

Types of collagen in breast capsules.

Representative sections of the capsules that envelop surgically implanted silicone were analyzed for their collagen content. The specimens were biopsied (as incidental procedures) as the opportunities arose. All four classes of capsules as codified clinically and those surrounding tissue expanders are represented. The ages of the capsules ranged from 6 weeks to 13 years. Eighteen specimens were examined by slab gel electrophoresis. Type I collagen predominated in all of the electrophoretic patterns. Type III collagen was consistently present in lesser concentrations. Type V collagen was consistently present as a small fraction. There was no evident correlation between the concentrations of the component collagens and the clinically assessed class of the capsules or of those from around tissue expanders, but there was a correlation to the age of the capsules. The electrophoretic patterns of the collagens of this study are indistinguishable from the (previously reported) analyses of (cutaneous) scar.

Adult↗

Analysis of silicon levels in capsules of gel and saline breast implants and of penile prostheses.

Although a potential link between silicone-gel breast implants and autoimmune connective tissue disease has been suggested, none has been proven. The potential role of silicone as an immune adjuvant remains very controversial. Currently available techniques do not allow precise measurements of silicone in tissues. However, all compounds containing silicon (including silicone) can be measured accurately. The present study was designed to measure silicon levels in the fibrous capsules of patients with silicone-gel breast implants, saline breast implants, and silicone inflatable penile prostheses. Baseline control silicon levels were obtained from the breast tissue of patients undergoing breast reduction, who had no exposure to breast implants. All silicon measurements were carried out using atomic absorption spectrometry with a graphite furnace. Silicon was measured in a normal heptane extract of silicone from dried tissue. The mean silicon levels in 16 breast tissue control samples from 8 patients undergoing breast reduction varied from 0.025 to 0.742 micrograms/gm with the median mean being 0.0927. The median silicon level in capsules from six patients with saline implants was 7.7 micrograms/gm (range, 1.9-36.6 micrograms/gm). The median silicon level in capsules from five patients with silicone inflatable penile prostheses was 19.5 micrograms/gm (range, 1.9-34.8 micrograms/gm). Although the levels of silicon in capsules of patients with saline breast prostheses and penile implants were higher than in control samples, they were much lower than those from the capsules of the 58 gel implants (median, 9,979 micrograms/gm; range, 371-152,000 micrograms/gm).(ABSTRACT TRUNCATED AT 250 WORDS)

Breast↗

The fate of the fibrous capsule after saline implant removal.

After removal of a silicone breast implant, if a capsulectomy is not performed, the residual capsule may persist, become calcified, and appear on routine mammograms. The fate of the capsule around saline implants is less clear. The purpose of this study was to determine the fate of the capsule around saline-filled implants in an animal model. Rats were implanted with 6-ml tissue expanders, which were left in place for 4 months. The implants were then removed and the capsules around the injection port (smooth surface) and tissue expander (textured surface) were examined sequentially. The capsules contracted and dissipated gradually over a year in association with a pericapsular vascular proliferation. It may not be necessary to perform a capsulectomy at the time of saline implant removal.

Animals↗

Unilateral trigeminal nerve branch lesion (V1 and V2) at the skull base and nasal capsule growth in rabbits.

The effects of long-term afferent denervation on midfacial and nasal capsule growth after craniofacial or skull base surgery are unknown. Experimental studies suggest that afferent nerves may play a role in regulating craniofacial growth through neurotrophic influences. The present study was designed to assess the effects of nasal capsule afferent (V1 and V2) denervation on nasal capsule growth in the rabbit. Eighteen 1 1/2-week-old rabbits were evenly divided into three groups: unoperated controls, sham controls, and animals with unilateral anterior ethmoid and sphenopalatine nerve lesions. The nerves were lesioned at the skull base with electrocautery through an intraorbital approach at 1 1/2 weeks of age. Lateral head radiographs and dental study models were taken at 1 1/2, 3 1/2, 6, and 12 weeks of age. By 3 1/2 weeks of age, sham controls and animals with nerve lesions exhibited significant (p < 0.05) reductions in nasal capsule area and height compared with unoperated control animals; the lesioned animals were more severely affected. However, by 6 weeks no differences were noted between the sham and lesioned groups, which were both smaller than unoperated controls through 12 weeks of age. Results suggest that acute nasal capsule growth deficits are a consequence of surgery as well as afferent denervation, with a greater deficit in the latter group, possibly through neurotrophic interruption. Such deficits may not be a permanent effect in the rabbit model.

Afferent Pathways↗

Effects of local instillation of triamcinolone on the capsules around silicone bag-gel prostheses in animals.

"Soft" silicone bag-gel miniprostheses were implanted in 103 rats. The experimental animals had periprosthetic instillations of zero, 0.2, or 2 mg of triamcinolone acetonide at implantation. The compressibility, the architecture, and the chemical composition of the surrounding tissues were observed at 10 and at 120 days. The compressibility was also measured by mechanical means at 10 day intervals. At 120 days all capsules were similar. All the capsules gradually became firmer with time after the implantation. After 60 days, the steroid-treated capsules were very slightly more compressible (softer) than the controls. This slight difference in compressibility could not be correlated with the other changes in the capsule properties we examined. It is suggested that the structure of these capsules may be similar to that of immature scar, and that it is kept in this condition by the continued stimulus of the foreign implant.

Animals↗