The Diadenylate Cyclase CdaA Is Critical for Borrelia turicatae Virulence and Physiology.

Relapsing fever (RF), caused by spirochetes of the genus Borrelia, is a globally distributed, vector-borne disease with high prevalence in developing countries. To date, signaling pathways required for infection and virulence of RF Borrelia spirochetes are unknown. Cyclic di-AMP (c-di-AMP), synthesized by diadenylate cyclases (DACs), is a second messenger predominantly found in Gram-positive organisms that is linked to virulence and essential physiological processes. Although Borrelia is Gram-negative, it encodes one DAC (CdaA), and its importance remains undefined. To investigate the contribution of c-di-AMP signaling in the RF bacterium Borrelia turicatae, a cdaA mutant was generated. The mutant was significantly attenuated during murine infection, and genetic complementation reversed this phenotype. Because c-di-AMP is essential for viability in many bacteria, whole-genome sequencing was performed on cdaA mutants, and single-nucleotide polymorphisms identified potential suppressor mutations. Additionally, conditional mutation of cdaA confirmed that CdaA is important for normal growth and physiology. Interestingly, mutation of cdaA did not affect expression of homologs of virulence regulators whose levels are impacted by c-di-AMP signaling in the Lyme disease bacterium Borrelia burgdorferi Finally, the cdaA mutant had a significant growth defect when grown with salts, at decreased osmolarity, and without pyruvate. While the salt treatment phenotype was not reversed by genetic complementation, possibly due to suppressor mutations, growth defects at decreased osmolarity and in media lacking pyruvate could be attributed directly to cdaA inactivation. Overall, these results indicate CdaA is critical for B. turicatae pathogenesis and link c-di-AMP to osmoregulation and central metabolism in RF spirochetes.

Assessing the Contribution of an HtrA Family Serine Protease During Borrelia turicatae Mammalian Infection.

Tick-borne relapsing fever (TBRF), characterized by recurring febrile episodes, is globally distributed and among the most common bacterial infections in some African countries. Despite the public health concern that this disease represents, little is known regarding the virulence determinants required by TBRF Borrelia during infection. Because the chromosomes of TBRF Borrelia show extensive colinearity with those of Lyme disease (LD) Borrelia, the exceptions represent unique genes encoding proteins that are potentially essential to the disparate enzootic cycles of these two groups of spirochetes. One such exception is a gene encoding an HtrA family protease, BtpA, that is present in TBRF Borrelia, but not in LD spirochetes. Previous work suggested that btpA orthologs may be important for resistance to stresses faced during mammalian infection. Herein, proteomic analyses of the TBRF spirochete, Borrelia turicatae, demonstrated that BtpA, as well as proteins encoded by adjacent genes in the B. turicatae genome, were produced in response to culture at mammalian body temperature, suggesting a role in mammalian infection. Further, transcriptional analyses revealed that btpA was expressed with the genes immediately upstream and downstream as part of an operon. To directly assess if btpA is involved in resistance to environmental stresses, btpA deletion mutants were generated. btpA mutants demonstrated no growth defect in response to heat shock, but were more sensitive to oxidative stress produced by t-butyl peroxide compared to wild-type B. turicatae. Finally, btpA mutants were fully infectious in a murine relapsing fever (RF) infection model. These results indicate that BtpA is either not required for mammalian infection, or that compensatory mechanisms exist in TBRF spirochetes to combat environmental stresses encountered during mammalian infection in the absence of BtpA.

Intrinsically disordered protein.

Proteins can exist in a trinity of structures: the ordered state, the molten globule, and the random coil. The five following examples suggest that native protein structure can correspond to any of the three states (not just the ordered state) and that protein function can arise from any of the three states and their transitions. (1) In a process that likely mimics infection, fd phage converts from the ordered into the disordered molten globular state. (2) Nucleosome hyperacetylation is crucial to DNA replication and transcription; this chemical modification greatly increases the net negative charge of the nucleosome core particle. We propose that the increased charge imbalance promotes its conversion to a much less rigid form. (3) Clusterin contains an ordered domain and also a native molten globular region. The molten globular domain likely functions as a proteinaceous detergent for cell remodeling and removal of apoptotic debris. (4) In a critical signaling event, a helix in calcineurin becomes bound and surrounded by calmodulin, thereby turning on calcineurin's serine/threonine phosphatase activity. Locating the calcineurin helix within a region of disorder is essential for enabling calmodulin to surround its target upon binding. (5) Calsequestrin regulates calcium levels in the sarcoplasmic reticulum by binding approximately 50 ions/molecule. Disordered polyanion tails at the carboxy terminus bind many of these calcium ions, perhaps without adopting a unique structure. In addition to these examples, we will discuss 16 more proteins with native disorder. These disordered regions include molecular recognition domains, protein folding inhibitors, flexible linkers, entropic springs, entropic clocks, and entropic bristles. Motivated by such examples of intrinsic disorder, we are studying the relationships between amino acid sequence and order/disorder, and from this information we are predicting intrinsic order/disorder from amino acid sequence. The sequence-structure relationships indicate that disorder is an encoded property, and the predictions strongly suggest that proteins in nature are much richer in intrinsic disorder than are those in the Protein Data Bank. Recent predictions on 29 genomes indicate that proteins from eucaryotes apparently have more intrinsic disorder than those from either bacteria or archaea, with typically > 30% of eucaryotic proteins having disordered regions of length > or = 50 consecutive residues.

Quantitative swab culture versus tissue biopsy: a comparison in chronic wounds.

Soft tissue infection present a significant obstacle to the healing of chronic wounds. Historically, the gold standard for determining wound bacterial bioburden has been the quantitative tissue biopsy. Nevertheless, tissue biopsies are not universally used in today's healthcare setting. Likely reasons include damage to healing tissue, the lack of facilities to process tissue biopsies, significant pain in sensate soft tissue, and increased expense with this modality. More recently, quantitative tissue swab culture has been suggested as a means to determine the wound bioburden. The authors prospectively studied 38 patients with chronic wounds of various etiologies to evaluate the correlation between quantitative wound biopsy and swab culture. Of the 38 biopsies performed, 74% indicated infection. Simultaneous swab culture of these 28 biopsies indicated infection in 22 of the 28 cases for a correlation of 79%. The authors concluded that quantitative swab culture provides a valuable adjunct in the management of chronic wounds.

Frequency of peristomal complications.

Practicing wound ostomy continence (WOC) nurses see peristomal complications in their ostomy patient population. The exact frequency and cause of these complications are unknown. Research into problems arising from prolonged use of ostomy appliances is lacking. To ascertain the frequency of peristomal complications, two WOC nurses at a major medical center prospectively assessed the peristomal skin of all new ostomy patients returning for their 2-month postoperative check-up using a peristomal complications tool. The study was conducted from August 1999 to August 2000. Descriptive statistics were used to summarize the data. In the course of the study, 161 new ostomy patients were seen, 10 with peristomal complications, for a frequency of 6%. The frequency of peristomal complications was the highest in people with ileal conduits (five of 34, 15%) and ileostomies (four of 46, 9%). Only one of the 81 people with a colostomy developed peristomal complications. All 10 patients had a retracted stoma. Eight had chemical damage (six with irritant dermatitis, two with pseudoverrucous lesions) and two had Candida infections. Although the study sample size is small, the results suggest that patients with ileal conduits and patients with retracted stomas may require more frequent follow-up visits to monitor skin conditions and pouching modifications.

A brief history of advanced practice nursing and its implications for WOC advanced nursing practice.

Although the evolution of clinical nurse specialists and nurse practitioners during the previous several decades has led to a sense of novelty regarding advanced practice, it is important to remember that nurses have engaged in such roles for nearly 100 years. This article will review the history of advanced nursing practice, summarize current definitions of specialty and advanced practice, discuss the evolution of advanced practice and its impact on contemporary nursing, and provide a background for advanced practice considerations for the WOC nurse, which are discussed in detail in subsequent articles by Janice Beitz (pp 55-64) and Dorothy Doughty (pp 65-8) in this issue.

TCOMs as a screening tool for hyperbaric oxygen therapy.

Transcutaneous oximetry (TcPO2), often referred to as TCOM, is a noninvasive measurement of oxygen tension in the tissue via a heated electrode. TCOMs can be used prior to beginning hyperbaric oxygen (HBO) treatment to determine if the patient has adequate blood flow to the wounded area to benefit from HBO. A case study of how it can be used in a problematic wound is presented.

Pressure ulcer assessment and management.

A pressure ulcer is an area of localized tissue destruction directly related to prolonged pressure. The loss of skin integrity produces significant consequences not only for the individual, but also for the community, with reported costs of $8.5 billion for pressure-ulcer care. Because of these costs, health care providers should be seeking prevention programs that identify high-risk individuals and implement preventive measures before the ulcer begins. Once the individual develops an ulcer, assessment of healing is critical to determine the appropriate treatment. Successful treatment depends on the principles of debridement, cleansing, bacterial control, wound dressing, and occasionally, surgical intervention.

Silicon granulomas and dermatomyositis-like changes associated with chronic eyelid edema after silicone breast implant.

Silicone medical products may be associated with various complications. Recent attention has focused particularly on health risks associated with silicone breast implants. The authors present the clinical and histopathological findings in a 71-year-old woman who had previously undergone breast reconstruction with a silicone gel-filled elastomer envelope-type breast implant and who developed severe chronic eyelid edema and inflammation. Extensive systemic evaluation and serologic studies were unrevealing, prompting biopsy of the eyelids. Histopathologic evaluation of the upper eyelids revealed scattered noncaseating granulomas throughout the dermis, an atrophying interface dermatitis with a sparse perivascular lymphocytic infiltrate, and focal perivascular and interstitial lymphocytic infiltrates of the muscles. Scanning electron microscopy demonstrated particles of material within the granulomas, which were revealed by electron probe x-ray microanalysis to contain silicon. The presence of silicon-containing material within areas of eyelid inflammation supports a role for silicon in the pathogenesis of this patient's condition. In the absence of other predisposing conditions, migration of silicone breast implant material must be considered.

Pilot study of a unique film dressing for the treatment of donor site wounds.

The clinical benefits of using the VENTEX Wound Dressing System (VWDS) (The Kendall Co.) in the management of donor site wounds were evaluated in this pilot study involving 10 informed and consenting patients. The study was a prospective, randomized, controlled study in which one donor site wound was managed with VWDS, and the other donor site wound on the same patient was managed by our standard of care, which involved the use of Xeroform gauze (The Kendall Co.). Each donor site wound was independently assessed daily for pain, rate of reepithelialization, adverse reactions, and ease of dressing use while patients were hospitalized. Quality of scar was assessed during regularly scheduled follow-up visits. The use of VWDS eliminated donor site pain and accelerated reepithelialization compared with Xeroform gauze. There were no adverse reactions associated with the use of VWDS. In contrast, there were two cases of suspected infection at the donor sites treated with Xeroform gauze. The VWDS was more complicated to apply and use compared with Xeroform gauze. There were no differences observed in scar quality of donor sites treated with these dressings.

The development and implementation of an integrated multidisciplinary clinical pathway.

Clinical pathways, linear time-related representations of patient care processes, are widely encouraged as a mechanism to outline efficient, cost-effective, multidisciplinary care. The translation of pathways from concept to reality is, however, predictably difficult. All caregivers are dedicated to a common goal, but organizational, personal, and professional perspectives are barriers to development of a common tool. Moreover, the building process requires the discovery, articulation, and communication of previously tacit patient care processes. Although no prescription can work for all pathway development, some strategies can help ensure the best possible chance of pathway success. Participants must recognize that between-patient variability can be expected to decrease with pathway implementation, and educational processes must support that aim. "Stakeholder groups" must be identified and their investment must be assessed, with careful attention paid to acquiring the unconditional support of institutional leadership. Planning of precise building, implementation, and piloting processes, with provision for facilitation of building and implementation processes, is critical. Those charged with pathway development must commit to the establishment and explication of clear goals (economic and quality outcomes) and to careful integration of the pathway with planned and existing continuous quality improvement processes. These strategies are illustrated with actual experiences in implementing cystectomy and pressure ulcer pathways in one academic medical center.

Development and implementation of a clinical pathway for radical cystectomy and urinary system reconstruction.

A multidisciplinary pathway and patient guide for radical cystectomies is described. Various forms of urinary diversion may be employed after cystectomy for bladder cancer. A clinical pathway for the management of patients undergoing radical cystectomy and urinary diversion or neobladder construction has proved beneficial to patient care, to the nursing and medical staffs, and to the institution.

Sexual adjustment of patients undergoing gracilis myocutaneous flap vaginal reconstruction in conjunction with pelvic exenteration.

BACKGROUND: Although the technique for gracilis myocutaneous vaginal reconstruction was first described in the mid-1970s and has been used in conjunction with pelvic exenteration since that time, there is little available information regarding sexual adjustment after such a procedure. The purpose of this study was to assess the sexual adjustment of women who underwent pelvic exenteration and gracilis myocutaneous vaginal reconstruction at the study institution. METHODS: In a prospective study design, 95 patients were identified who underwent pelvic exenteration and gracilis myocutaneous vaginal reconstruction at the study institution from 1977 through 1989 and a convenience sample was selected of 44 patients who completed a modified version of the Sexual Adjustment Questionnaire (SAQ) when they returned to the gynecologic oncology outpatient clinic for routine follow-up care. A vaginal assessment was also performed by the attending physician. RESULTS: Twenty-one of 40 patients (52.5%) completing the questionnaire reported not resuming sexual activity after surgery; 19 patients reported sexual activity between 1.5 months to 12 years postoperatively. Of the patients who resumed sexual activity, 84% did so within 1 year of surgery. The most common problems noted by patients in adjusting to sexual activity after surgery were self-consciousness about the urostomy or colostomy and being seen in the nude by their partner, vaginal dryness, and vaginal discharge. The mean rank of preexenteration SAQ scores was 66.4, and the mean rank of postexenteration scores was 48.7 (P < 0.0001), demonstrating that sexual adjustment after exenteration was significantly poorer than before the surgery. On the basis of data gathered from a vaginal assessment form, 31 of 44 patients (70.4%) were judged to have a potentially functional neovagina. CONCLUSIONS: Based on the findings of this questionnaire study, sexual adjustment is often significantly impaired in women after pelvic exenteration and gracilis myocutaneous vaginal reconstruction. Future modifications in surgical technique, more realistic patient counseling and aggressive postoperative support will hopefully minimize such problems.

Evaluating data acquisition and smoothing functions of currently available videokeratoscopes.

PURPOSE: To compare the accuracy of computerized videokeratography systems using identical, calibrated test objects. SETTING: Lions Eye Institute, Albany, New York. METHODS: We evaluated the accuracy and smoothing of raw data acquisition (axial solution) of seven commercially available videokeratoscopes: Alcon EyeMap, Computed Anatomy TMS, EyeSys CAS, Humphrey MasterVue, Topcon CM-1000, Optikon Keratron, and TechnoMed C-Scan. We used six calibrated test objects to simulate clinical settings: spherical, spherocylindrical, simulated myopic ablation, hyperopic ablation, and a simulated central island. RESULTS: None of the systems accurately imaged all objects. Although all systems imaged spherical objects with reasonable accuracy, errors greater than 4.0 diopters (D) frequently occurred in the central 6.0 mm optical zone (maximum error 10.0 D) Sources of error included excessive raw data smoothing, inability to read large transitions, loss of accuracy in the periphery, and poor central coverage. CONCLUSION: The clinician should be aware of the potential limitations of corneal topography when making clinical decisions.

PAR Corneal Topography System (PAR CTS): the clinical application of close-range photogrammetry.

The PAR Corneal Topography System (CTS) is a computer-driven corneal imaging system which uses close-range photogrammetry (rasterphotogrammetry) to measure and produce a topographic map of the corneal surface. The PAR CTS makes direct point-by-point measurements of surface elevation using a stereo-triangulation technique. The CTS uses a grid pattern composed of horizontal and vertical lines spaced about 0.2 mm (200 microns) apart. Each grid intersection comprises a surface feature which can be located in multiple images and used to generate an (x,y,z) coordinate. Unlike placido disc-based videokeratoscopes, the PAR CTS requires neither a smooth reflective surface nor precise spatial alignment for accurate imaging. In addition to surface elevation, the PAR CTS computes axial and tangential curvatures and refractive power. Difference maps are available in all curvatures, refractive power, and in absolute elevation.

Alignment success following medical rectus recessions in normal and delayed children.

Others have suggested that developmentally delayed children, who are frequently esotropic, have a poor prognosis following esotropia surgery. To date, no comparison of success rates in normal and delayed children following similar surgery has been made. We compared our long-term results following graded bilateral medial rectus recessions in consecutive normal (n = 62) and delayed (n = 29) children. We defined success as alignment maintained within 10 delta of orthophoria. Survival curves were compared using the Mantel-Haenszel statistic. Delayed children had a significantly poorer outcome (p = .05) throughout follow up, which ranged from 12 to 120 months (mean = 24 months). Early in the period of study, we observed a large proportion of overcorrections among the delayed children. Subsequent modification in the amount of surgery performed appeared to improve the initial and long-term success in these children. We conclude that delayed children do have a poorer prognosis than normal children following medial rectus recession. This prognosis may be improved by more conservative amounts of surgery.

The chronic wound care clinic: "one-stop shopping".

The principal goals of wound management are the provision of effective care and cost containment. An outpatient clinic dedicated to the management of chronic wounds offers several advantages over traditional, more fragmented approaches. The Chronic Wound Care Clinic at the University of Virginia Health Sciences Center, a multidisciplinary outpatient clinic, provides "one-stop shopping," allowing patients to benefit from coordinated care provided in a single setting. Overviews of the clinic's history, philosophy, and structure are presented. A case study illustrates the rapid wound healing and cost savings of one patient cared for at the clinic.

Silicone intubation without intranasal fixation for treatment of congenital nasolacrimal duct obstruction.

PURPOSE: We assessed the efficacy and morbidity of silicone intubation without intranasal fixation to treat congenital nasolacrimal duct obstruction. METHODS: Forty eyes of 30 consecutive patients underwent silicone intubation to treat congenital nasolacrimal duct obstruction. The ends of the silicone tubing were tied together with a single square knot and were allowed to retract into the nose without intranasal fixation. Tubing removal was performed in the office via the medial canthus in all patients. No attempt was made to reposition prematurely dislodged tubing. RESULTS: The success rate for this series was 38 (95%) of 40 eyes in 29 (97%) of 30 patients. Of seven eyes that required unplanned early tubing removal for dislocated tubing, five eyes nevertheless showed complete resolution of nasolacrimal duct obstruction. Only one patient underwent a second silicone intubation under general anesthesia. CONCLUSIONS: Silicone intubation without the use of intranasal fixation is an effective strategy for treating congenital nasolacrimal duct obstruction. It spares most patients from a second procedure under general anesthesia and achieves success rates comparable to previous studies in which more complex techniques were used.

Associations of HLA-DR and HLA-DQ types with anti-GBM nephritis by sequence-specific oligonucleotide probe hybridization.

Twenty-three patients from the United States with anti-glomerular basement membrane nephritis (Goodpasture's syndrome) were HLA class I and class II typed by serology and HLA-DR and HLA-DQB typed using sequence-specific oligonucleotide (SSO) probes. Protocols used were those of the 11th International HLA Workshop. We were able to confirm the strong association of DR2 with the disease and, using the molecular techniques, showed that all DR2 patients were restricted to DRB1-1501 and DQB1-0602 alleles. We also found highly significant associations between disease and the alleles of DRB1-0301 and DQB1-0201. DR4 alleles as a group were not significantly more frequent among patients, but the allele DRB1-0404 did appear to be associated with the disease. The use of SSO typing provides an expanded opportunity for refined analysis of HLA and disease associated alleles.