Detection of Bacteroides fragilis Enterotoxin Gene by PCR

Bacteroides fragilis constitutes about 1% of the bacterial flora in intestines of normal humans. Enterotoxigenic strains of B. fragilis have been associated with diarrheal diseases in humans and animals. The enterotoxin produced by these isolates induces fluid changes in ligated intestinal loops and an in vitro cytotoxic response in HT-29 cells. We developed a nested PCR to detect the enterotoxin gene of B. fragilis in stool specimens. After DNA extraction, a 367-bp fragment was amplified with two outer primers. The amplicon from this reaction was subjected to a second round of amplification with a set of internal primers. With these inner primers, a 290-bp DNA fragment was obtained which was confirmed as part of the B. fragilis enterotoxin gene by Southern blotting with a nonradioactive internal probe and a chemiluminescence system. By this approach, B. fragilis enterotoxin gene sequences were detected in eight known enterotoxigenic human isolates and nine enterotoxigenic horse isolates. No amplification products were obtained from DNA extracted from 28 nonenterotoxigenic B. fragilis isolates or B. distasonis, B. thetaiotaomicron, B. uniformis, B. ovatus, Escherichia coli, or Clostridium difficile. The sensitivity of this assay allowed us to detect as little as 1 pg of enterotoxin DNA sequences or 100 to 1,000 cells of enterotoxigenic B. fragilis/g of stool. Enterotoxin production of all isolates was confirmed in vitro in HT-29 cells. A 100% correlation was obtained between enterotoxin detection by cytotoxin assay and the nested PCR assay. This rapid and sensitive assay can be used to identify enterotoxigenic B. fragilis and may be used clinically to determine the role of B. fragilis in diarrheal diseases.     

Neonatal mortality in Jordan: secondary analysis of Jordan Population and Family Health Survey (JPFHS) data

Objective: This study aimed to analyze the 2009 Jordan Population and Family Health Survey (JPFHS) data to determine the level, trend, and distribution of neonatal mortality (NNM) in Jordan and determine its associated factors.

Methods: Nationally representative data on NNM were extracted from the JPFHS data. Using multivariate analyses, the strength of associations between 12 clinical/sociodemographic variables and neonatal mortality were quantified after controlling for potential confounders.

Results: The weighted NNM rate for 2005–2009 period was 16 deaths per 1000 live births, with the early NNM rate and late NNM rates were 10 deaths per 1000 live births and six deaths per 1000 live births, respectively. Fluctuations of NNM according to year of birth and geographic variations were noted. Risk of NNM increased among male newborns, as mother’s education level decreased, in mothers 40–49 years old, in multiple gestations-low birth weight neonates, and as birth interval was <3 years.

Conclusions: The NNR rate for 2005–2009 period of 16 deaths per 1000 live births indicates that there are opportunities to decrease it. Risk factors of neonatal mortality with respect to predictors of death during first days of life and variables related to geographic variations require particular focus to improve the quality of obstetric and neonatal health services and to decrease neonatal mortality.     

Ximelagatran,the New Oral Anticoagulant: Would Warfarin Survive the Challenge?

The last decade witnessed major advances in the prevention and treatment of venous as well as of arterial thrombosis. Limitations of existing anticoagulants led to the development of novel therapeutic approaches. Ximelagatran is a new direct thrombin inhibitor (DTI) that is given orally, without the need for close monitoring. This compound was tried in the treatment of active venous thromboembolism, and the results were encouraging. Randomized trials suggest that ximelagatran is not inferior to warfarin in the prevention of stroke in patients with nonvalvular atrial fibrillation. Multiple controlled, prospective trials compared ximelagatran with low molecular weight heparin or warfarin in prevention of venous thromboembolism in patients undergoing major orthopedic procedures. The results of these clinical trials are reviewed in this article. Because of certain safety concerns (increased liver enzymes, potential hepatonecrosis, and increased coronary events) ximelagatran has not yet been approved by the FDA. Additional studies may be required to address these concerns. Ximelagatran has been approved, however, by the European regulatory authorities for short‐term thromboprophylaxis. The success of ximelagatran or other oral antithrombin agents would provide significant proof of the concept for the long‐term use of oral antithrombins in the prevention and treatment of both arterial and venous thrombosis.     

Prevalence of enterotoxigenic Bacteroides fragilis in hospital-acquired diarrhea

Stool specimens from 152 hospitalized patients with diarrhea were analyzed for the presence of enterotoxigenic Bacteroides fragilis (ETBF) by a nested polymerase chain reaction (PCR) assay. ETBF gene sequences were directly detected in 14/152 (9.21%) stools of patients. The prevalence of ETBF in hospital-acquired diarrhea was statistically significant when compared to a prevalence of 2.3% in control subjects (P = 0.04). B. fragilis was cultured from 19.7% (30/152) patients with diarrhea; 4 of these isolates were enterotoxigenic. To determine whether colonization with B. fragilis is heterogeneous in nature, multiple colonies from 17 individual patients were analyzed for enterotoxin gene sequences and genotyped by arbitrarily primed PCR. Of these 17 patients, 13 harbored multiple strain types suggesting heterogeneity of colonization with both enterotoxigenic and non-enterotoxigenic strains. Identification of ETBF in the stools of 10 patients in the absence of a positive culture is likely due to the noted heterogeneity and suggests that detection of enterotoxin by PCR should be performed directly in the stool. These preliminary data indicate that ETBF may play a role in hospital-acquired diarrhea of unknown origin and suggest the need for further studies.     

Invasive meningococcal disease in adolescents and young adults

Context  Incidence of invasive meningococcal disease has increased recently in persons aged 15 through 24 years. Objective  To characterize meningococcal infection in adolescents and young adults in Maryland during the 1990s. Design and Setting  Population-based surveillance study for meningococcal disease from January 1, 1990, through December 31, 1999, in Maryland. Patients  Maryland residents diagnosed as having invasive meningococcal disease. Main Outcome Measure  Invasive meningococcal infection. Results  Of 295 total cases, 71 (24.1%) occurred among persons aged 15 through 24 years. Sixteen (22.5%) of these cases were fatal. The annual incidence rate increased from 0.9 to 2.1 cases per 100 000 among 15 through 24 year olds (P = .01). The proportion of all disease increased from 16.0% to 28.9% (P = .03). The incidence and proportion of cases subsequently decreased to 1.0 and 16.4% in 1998 through 1999, respectively. Infection in 15 through 24 year olds was more likely to be fatal than infection in those younger than age 15 years (22.5% vs 4.6%; P = .001). Infection in 15 through 24 year olds, compared with those aged 25 years or older, was more likely to be associated with male sex (66.2% vs 34.8%; P<.001) and serogroup C infection (46.9% vs 20.2%; P<.001), respectively. Infections were potentially preventable with the licensed meningococcal vaccine in 82.8% of 15 through 24 year olds, 68.1% of those younger than 15 years, and 76.8% of adults aged 25 years or older. Conclusions  Incidence of meningococcal infection in 15 through 24 year olds in Maryland increased and then declined during the 1990s. Infection in this age group was associated with an unusually high case-fatality ratio, and the vast majority of cases were potentially vaccine preventable.      

Benefits of Zataria multiflora Boiss in Experimental Model of Mouse Inflammatory Bowel Disease

Inflammatory bowel disease (IBD) is a chronic condition of theintestine with unknown etiology involving multiple immune, geneticand environmental factors. We were interested to examine theeffect of total extract from Zataria multiflora Boiss, a folkmedicinal plant on prevention and treatment of experimentalIBD. Z. multiflora was administered (400, 600, 900 p.p.m.) throughdrinking water to IBD mice induced by intrarectal administrationof acetic acid. Prednisolone was used as the standard drug forcomparison. Biochemical, macroscopic and microscopic examinationsof colon were performed. Biochemical evaluation of inflamedcolon was done using assay of myeloperoxidase (MPO) activityand thiobarbituric acid reactive substances (TBARS) concentrationas indicators of free radical activity and cell lipid peroxidation.The activity of MPO and lipid peroxidation products (TBARS)increased in acetic acid-treated groups while recovered by pretreatmentof animals with Z. multiflora (400–900 p.p.m.) and prednisolone.Z. multiflora (600 and 900 p.p.m.) and prednisolone-treatedgroups showed significantly lower score values of macroscopicand microscopic characters when compared with the acetic acid-treatedgroup. The beneficial effect of Z. multiflora (900 p.p.m.) wascomparable with that of prednisolone. The antioxidant, antimicrobialand anti-inflammatory potentials of Z. multiflora might be themechanisms by which this herbal extract protects animals againstexperimentally induced IBD. Proper clinical investigation shouldbe carried out to confirm the activity in human.