The EXIT Strategy: an Approach for Identifying Bacterial Proteins Exported during Host Infection.

Exported proteins of bacterial pathogens function both in essential physiological processes and in virulence. Past efforts to identify exported proteins were limited by the use of bacteria growing under laboratory (in vitro) conditions. Thus, exported proteins that are exported only or preferentially in the context of infection may be overlooked. To solve this problem, we developed a genome-wide method, named EXIT (exported in vivotechnology), to identify proteins that are exported by bacteria during infection and applied it to Mycobacterium tuberculosis during murine infection. Our studies validate the power of EXIT to identify proteins exported during infection on an unprecedented scale (593 proteins) and to reveal in vivo induced exported proteins (i.e., proteins exported significantly more during in vivo infection than in vitro). Our EXIT data also provide an unmatched resource for mapping the topology of M. tuberculosis membrane proteins. As a new approach for identifying exported proteins, EXIT has potential applicability to other pathogens and experimental conditions.IMPORTANCE There is long-standing interest in identifying exported proteins of bacteria as they play critical roles in physiology and virulence and are commonly immunogenic antigens and targets of antibiotics. While significant effort has been made to identify the bacterial proteins that are exported beyond the cytoplasm to the membrane, cell wall, or host environment, current methods to identify exported proteins are limited by their use of bacteria growing under laboratory (in vitro) conditions. Because in vitro conditions do not mimic the complexity of the host environment, critical exported proteins that are preferentially exported in the context of infection may be overlooked. We developed a novel method to identify proteins that are exported by bacteria during host infection and applied it to identify Mycobacterium tuberculosis proteins exported in a mouse model of tuberculosis.

Inhaled Pyrazinoic Acid Esters for the Treatment of Tuberculosis.

PURPOSE: Analog development of existing drugs and direct drug delivery to the lungs by inhalation as treatments for multiple and extensively drug resistant (MDR and XDR) tuberculosis (TB) represent new therapeutic strategies. Pyrazinamide (PZA) is critical to drug sensitive TB therapy and is included in regimens for MDR TB. However, PZA-resistant Mycobacterium tuberculosis (Mtb) strains threaten its use. Pyrazinoic acid esters (PAEs) are PZA analogs effective against Mtb in vitro, including against the most common PZA resistant strains. However, PAEs require testing for TB efficacy in animal models. METHODS: PAEs were delivered daily as aqueous dispersions from a vibrating mesh nebulizer to Mtb infected guinea pigs for 4 weeks in a regimen including orally administered first-line TB drugs. RESULTS: PAEs tested as a supplement to oral therapy significantly reduced the organ bacterial burden in comparison to infected, untreated control animals. Thus, PAE aerosol therapy is a potentially significant addition to the regimen for PZA resistant MDR-TB and XDR-TB treatment. Interestingly, low dose oral PZA treatment combined with standard therapy also reduced bacterial burden. This observation may be important for PZA susceptible disease treatment. CONCLUSION: The present study justifies further evaluation of PZA analogs and their lung delivery to treat TB.

Comparison of the prognostic value of the 6th and 7th editions of the Union for International Cancer Control TNM staging system in patients with lower esophageal cancer undergoing neoadjuvant chemotherapy followed by surgery.

Carcinoma of the esophagus is classified according to the Union for International Cancer Control (UICC) TNM staging system. The 7th edition of the UICC TNM staging system was published in 2009. This is the first study to compare the prognostic value of the TNM 6th and 7th editions in patients with esophageal carcinoma treated with chemotherapy followed by surgery. Two hundred forty-three patients with esophageal carcinoma were retrospectively selected from two referral centers. All patients received chemotherapy before surgery. Histopathologic data from the resection specimens were retrieved and restaged according to the TNM 7th edition. Disease-specific survival curves were plotted for depth of tumor invasion (ypT), lymph node status (ypN), and ypTNM stage and then compared. Median follow-up after surgery was 2.5 years (range 0.2-9 years). Survival analysis using the log-rank method revealed that there was a significant difference in survival between ypT4 disease and ypT3 disease (P= 0.003), but no difference between ypT0, ypT1, ypT2, and ypT3 categories irrespective of TNM edition used. Survival probability was significantly different between ypN0 and ypN1 (P= 0.001 for TNM 6th and 7th edition), as well as ypN2 and ypN3 (TNM 7th edition, P= 0.004), but not between ypN1 and ypN2 (TNM 7th edition, P= 0.89). Neither the TNM 6th nor 7th edition T staging provides accurate survival probability stratification. However, the advantage of the 7th edition is the introduction of a third tier in survival stratification for patients with nodal involvement.

Readability in the British Journal of Surgery.

BACKGROUND: Readability scores predict the ease with which a document can be read and understood. Identification of the factors that affect readability might improve the quality of surgical literature. METHODS: Electronic copies of submitted, peer reviewed and edited manuscripts of 189 articles published in the British Journal of Surgery from March 2006 to April 2007 were studied. Flesch reading ease scores were calculated for each version. Type of article (meta-analysis, review, randomized trial or other original paper), first language of principal author, number of peer revisions and editor were recorded. RESULTS: Flesch score varied according to type of article (P = 0.004). The mean readability score was lower when English was the first language of the principal author (P = 0.016). There was no significant difference in mean Flesch score between the submitted and accepted (peer reviewed) manuscripts, but a significant increase occurred after editing (P < 0.001), which did not vary between editors. CONCLUSION: Several factors influence readability. Knowledge of such factors might help authors to improve their scientific writing.

Minimally invasive management of chylous fistula after esophagectomy.

Chyle leak is an unwelcome complication of esophagectomy that is associated with a high mortality. The diagnosis of this condition may be difficult or delayed and requires a high index of suspicion. Management varies from conservative treatment with drainage, intravenous nutrition, treatment and prevention of septic complications, to re-operation, either by thoracotomy or laparotomy to control the fistula. To reduce the mortality, early surgical intervention is advised and a minimally invasive approach has recently been reported in several cases. From June 2002 through August 2005 we have used video-assisted thoracoscopic surgery to diagnose and treat chyle fistulas from 6/129 (5%) patients who underwent esophagectomy for resectable carcinoma of the esophagus or high-grade dysplasia. The fistula was successfully controlled in 5/6 cases by direct thoracoscopic application of a suture, clips or fibrin glue. One patient required a laparotomy and ligation of the cysterna chyli after thoracoscopy failed to identify an intrathoracic source of the leak. An early minimally invasive approach can be safely and effectively applied to the diagnosis and management of post-esophagectomy chylous fistula in the majority of cases. Open surgery may be appropriate where minimally invasive approaches fail or where the availability of such skills is limited.

Right-sided Bochdalek hernia obstructing in an adult: case report and review of the literature.

Bochdalek hernias on the right side of the diaphragm are very rarely diagnosed in adults. We review a case of a 35-year-old female who presented acutely with intestinal obstruction. Plain and cross-sectional imaging identified a large right-sided Bochdalek hernia, containing colon, causing a mechanical obstruction and, surprisingly, concurrent appendicitis. The patient underwent an emergency laparotomy. At surgery the colon was reduced and was viable. The diaphragmatic defect was repaired using non-absorbable sutures and an appendicectomy was then performed for purulent appendicitis. She made an uneventful recovery and remains well at 9-month follow-up. We discuss what we believe to be the first reported case of an obstructed right-sided Bochdalek's hernia associated with appendicitis in an adult and review the published literature on this rare condition.

Illness behavior and laparoscopic antireflux surgery: tailoring the wrap to suit the patient.

Psychological factors are believed to play a role in gastroesophageal reflux disease. It has previously been shown that preoperative illness behavior influences the outcome after laparoscopic Nissen fundoplication. Between August 2001 and June 2004 we considered a partly subjective assessment of illness behavior when selecting patients with gastroesophageal reflux disease for laparoscopic anterior partial (n = 77) or total fundoplication (n = 90). A prospective questionnaire study of illness behavior was also undertaken and the results were correlated with clinical follow up after 12 months. There was a statistically significant difference in age (P < 0.001), primary esophageal peristalsis on manometry (P = 0.037) and two illness behavior category scores related to hypochondriasis (P = 0.041 and P = 0.025) between laparoscopic anterior partial fundoplication and Nissen total fundoplication groups. Despite these differences, there was no significant correlation between preoperative illness behavior score and patient satisfaction in either group. There was a statistically significant negative correlation between the ability to express personal feelings and postoperative heartburn score in those who had a laparoscopic anterior partial fundoplication (P = 0.048). The clinical outcome in both groups was good to excellent in terms of postoperative heartburn and satisfaction scores. A tailored approach in the choice of wrap, taking into account psychological factors preoperatively, is an appropriate strategy for laparoscopic fundoplication.

Effect on iron deficiency anemia of laparoscopic repair of large paraesophageal hernias.

Patients with iron deficiency anemia sometimes have a large paraesophageal hernia and no other explanation for their chronic blood loss. The management of these patients can be a dilemma, especially when the hernia is otherwise asymptomatic. We aimed to determine whether a laparoscopic repair of the hernia could cure the anemia. We reviewed a consecutive series of 11 cases of iron deficiency anemia associated with a large paraesophageal hernia, many without associated linear gastric erosions, managed by laparoscopic repair and fundoplication. There was one conversion in a patient with dense adhesions from previous upper abdominal surgery. Another patient required a laparoscopic reoperation for an early recurrence. Major morbidity occurred in three patients and there was no mortality. There was no recurrence of anemia after a median follow-up of more than 2 years. Iron deficiency anemia in association with a large paraesophageal hernia can be treated by laparoscopic repair with acceptable morbidity and minimal mortality. The complications of a large paraesophageal hernia are also prevented.

A prospective study of ambulatory laparoscopic cholecystectomy: training economic, and patient benefits.

BACKGROUND: Even though ambulatory laparoscopic cholecystectomy (ALC) is safe and cost effective, this approach has yet to gain acceptance in the United Kingdom. We report our 5-year experience of ALC with emphasis on its appropriateness for higher surgical training. METHODS: Between July 1997 and July 2002, patients with symptomatic cholelithiasis who met with appropriate criteria underwent ALC. Surgery was performed either by a consultant surgeon or a higher surgical trainee (HST) under direct supervision in our dedicated day surgery unit. Data were recorded prospectively and patients were interviewed postoperatively by an independent researcher. RESULTS: There were 269 patients (231 female and 38 male) with a median age of 46 years (range 17-76). Conversion to open cholecystectomy was necessary in three cases (1%). Of the patients, 79% (213) were discharged within 8 hours of surgery; 95% (256) were discharged on the same day. Thirteen patients (5%) required overnight admission as inpatients. An HST performed 166 (62%) of the procedures. There was a statistically significant difference in operating time between consultants (41 min) and trainees (47 min, P = 0.001) but no significant difference in clinical outcome or patient satisfaction. The mean procedural cost to the hospital was 768 pound sterling for ALC compared with 1430 pound sterling for an inpatient operation. Of patients, 87% expressed satisfaction with the day case operation. CONCLUSION: Our results for ALC compare favorably with published series. In addition, we have demonstrated that the operation can be performed safely by HST under direct supervision without compromising operating lists or safety.

Benign febrile myoclonus in childhood.

BACKGROUND: Myoclonus is often associated with progressive myoclonic epilepsy or neurodegenerative conditions. Febrile myoclonus is a benign phenomenon, which has only been reported previously in one child. METHODS: The clinical features of three children with fever-induced myoclonus are described. RESULTS: Fever-induced myoclonus is characterized by frequent myoclonus, which resolves with resolution of the fever in otherwise healthy children. CONCLUSIONS: Recognition of fever-induced myoclonus as a benign phenomenon may prevent unnecessary investigations and interventions.

Early pin failure following external femoral fracture fixation in a head-injured child.

External fixation facilitates nursing care and early rehabilitation of head-injured children with femoral shaft fractures. We report an unusual complication of early fatigue failure of the external fixator pins without the patient ever bearing weight.

A comparison of microsatellite instability in early onset gastric carcinomas from relatively low and high incidence European populations.

We have investigated the genetic basis of gastric carcinomas occurring in patients aged under 40 years from a Portuguese population with a relatively high incidence of gastric cancer. We analysed a panel of 12 microsatellite loci in DNA extracted from gastric carcinomas arising in 16 patients aged 24-39 years from Braga, Portugal. Overall, microsatellite instability (MI) in at least 1 locus was detected in 44% (7 of 16) of carcinomas. A single patient demonstrated a mutator phenotype suggestive of the hereditary nonpolyposis colorectal cancer syndrome with instability in 82% of loci. This carcinoma showed loss of expression of the hMLH1 mismatch repair protein. In a previous study, we found no evidence of MI among 10 cases of early onset gastric carcinomas from an English population, which has a relatively low incidence of gastric cancer. Comparing the 2 series, we found that there was a significant difference (p = 0.04) in the prevalence of MI (at least 1 marker affected). This geographical difference in low-level MI may be related to a significantly higher prevalence of background chronic atrophic gastritis (8 of 16 vs. 0 of 8) and Helicobacter pylori infection (15 of 16 vs. 2 of 8) in Portuguese carcinomas compared with English cases. Genetic mechanisms underlying the hereditary non-polyposis colorectal cancer syndrome may play a role in a small number of early onset gastric carcinomas. The difference in prevalence of low-level MI between these relatively high and low incidence European populations requires further investigation.

Diaphragmatic rupture resulting from gastrointestinal barotrauma in a scuba diver.

A fit young man sustained a ruptured diaphragm during a recreational scuba dive three months after undergoing an uncomplicated laparoscopic Nissen fundoplication for gastro-oesophageal reflux disease. It is proposed that this rare occurrence was attributable to gastrointestinal barotrauma. The injury was treated by laparotomy, mobilisation of herniated abdominal viscera back into the abdomen, repair of the crura and gastropexy. He made a full postoperative recovery. It is concluded that scuba diving should be avoided in patients who have undergone fundoplication.

Assessment of microsatellite alterations in young patients with gastric adenocarcinoma.

BACKGROUND: Genetic factors are probably important in the development of gastric carcinoma in young patients (younger than 40 years). The authors investigated early onset primary gastric adenocarcinomas for the presence of microsatellite instability, which is a phenotypic marker for the hereditary nonpolyposis colon carcinoma syndrome. METHODS: DNA was extracted from archival microdissected carcinoma and corresponding normal tissue from 10 British gastric carcinoma patients age 19 to 39 years at the time of diagnosis. A panel of 12 microsatellite loci were amplified by fluorescent polymerase chain reaction and analyzed using an automated DNA sequencer. RESULTS: There was no evidence of microsatellite instability. In contrast, allelic imbalance was recorded at D3S966, D3S1076, D10S197, D11S904, P53, NM23, and DCC microsatellite loci. CONCLUSIONS: The authors reported ten cases of early onset gastric carcinoma that demonstrated allelic imbalance but no evidence of instability at microsatellite loci. It is unlikely that defective DNA mismatch repair is important in this group of young patients.

Prognostic significance of microsatellite instability in patients with gastric carcinoma.

A proportion of gastric adenocarcinomas exhibit replication errors manifested as microsatellite instability. The clinicopathological and prognostic significance of this abnormality remains uncertain. This study aimed to determine the importance of microsatellite instability by analysing a large series of gastric carcinomas from an English population. Using a novel fluorescent polymerase chain reaction technique, we amplified 11 microsatellite sequences from paired normal and carcinoma DNA from 101 patients who underwent a potentially curative resection for gastric carcinoma. Overall, 21% of cases demonstrated microsatellite instability in at least one locus. At least four loci were examined in each case. A replication error positive phenotype (minimum of 29% of loci affected) was detected in 9% of cases. There was no statistically significant association between the presence of microsatellite instability or replication error positive phenotype and the patient's age, sex, tumour site, stage, node status, histological subtype or grade. Carcinomas confined to the mucosa or submucosa (T1) showed a significantly higher frequency of instability and replication error positive phenotypes than T3 lesions (P = 0.03 and P = 0.05, respectively). A larger proportion of patients who were microsatellite instability or replication error positive were alive at 5 years compared with those who were negative but this did not reach statistical significance (P = 0.15 and P = 0.16, respectively). We identified a subset of gastric carcinomas from a relatively low-risk population which showed evidence of microsatellite instability. There were no statistically significant 5-year survival advantages in cases demonstrating microsatellite instability or replication error positive phenotypes. The detection of microsatellite instability is of limited prognostic value in gastric carcinoma.