Site-specific recombinase and integrase activities of a conjugative relaxase in recipient cells

Conjugative relaxases are the proteins that initiate bacterial conjugation by a site-specific cleavage of the transferred DNA strand. In vitro, they show strand-transferase activity on single-stranded DNA, which suggests they may also be responsible for recircularization of the transferred DNA. In this work, we show that TrwC, the relaxase of plasmid R388, is fully functional in the recipient cell, as shown by complementation of an R388 trwC mutant in the recipient. TrwC transport to the recipient is also observed in the absence of DNA transfer, although it still requires the conjugative coupling protein. In addition to its role in conjugation, TrwC is able to catalyze site-specific recombination between two origin of transfer (oriT) copies. Mutations that abolish TrwC DNA strand-transferase activity also abolish oriT-specific recombination. A plasmid containing two oriT copies resident in the recipient cell undergoes recombination when a TrwC-piloted DNA is conjugatively transferred into it. Finally, we show TrwC-dependent integration of the transferred DNA into a resident oriT copy in the recipient cell. Our results indicate that a conjugative relaxase is active once in the recipient cell, where it performs the nicking and strand-transfer reactions that would be required to recircularize the transferred DNA. This TrwC site-specific integration activity in recipient cells may lead to future biotechnological applications.     

Carriage of Neisseria meningitidis and Neisseria lactamica in infants and children.

Asymptomatic carriage of Neisseria meningitidis and Neisseria lactamica was studied in a total of 2,969 healthy infants and children in Danbury, Conn., between October 1971 and June 1975. The prevalence of N. meningitidis averaged 0.71% during the first four years of life and increased to 5.4% by 14--17 years. Rates of carriage of N. lactamica increased from 3.8% in three-month-old infants to a peak of 21.0% at 18 months and then declined to 1.8% by 14--17 years of age. Of the children who acquired N. lactamica, 66% developed fourfold or greater rises in titers of IgG antibody to groups A, B, and/or C meningococci as determined by immunofluorescence compared with only 5% of control children. Of new carriers of N. lactamica, 40% developed increased titers of bactericidal antibody to groups A, B, and/or C meningococci as compared with 7% of noncarriers. Carriage of N. lactamica may assist in the development of natural immunity to N. meningitidis by induction of cross-reactive antibodies.     

High yield criteria for emergency cranial computed tomography in adult patients with no history of head injury

Objectives—A recent American study identified clinical factors which effectively predicted those patients who would have significant findings on cranial computed tomography. It was proposed to apply these criteria in a UK setting and to determine whether modifications could be made to improve their efficiency.

Methods—A prospective observational study was conducted over a four month period including all non-trauma adult patients referred from the accident and emergency (A&E) department for urgent cranial computed tomography. Presenting symptoms and signs were analysed for ability to predict clinically significant computed tomography findings, namely: acute infarct, malignancy, acute hydrocephalus, intracranial haemorrhage, or intracranial infection.

Results—Sixty two patients were included; 22 (35%) had significant findings on computed tomography. Applying the original criteria (any of: age 60 years or older, focal neurology, headache with nausea or vomiting, altered mental status) to the study population showed that no clinically significant tomograms would have been omitted but only 11% fewer performed. Modifying the criteria by removing "age 60 years or older" and replacing "altered mental status" with a Glasgow coma score <14, still ensured 100% sensitivity and would have resulted in 19% fewer scans being performed.

Conclusion—Simple clinical criteria can be usefully applied to patients presenting to an A&E department in this country to target patients most likely to have clinically significant findings on urgent cranial computed tomography.

     

The East Anglia Deanery Communication Skills Teaching Project-six years on

This paper describes the Cascade Communication Skills Teaching Project, which is a programme of facilitator training enabling communication skills teaching in the consultation to general practitioners to be cascaded throughout the former East Anglia Deanery. The paper also explores the project's educational and organizational effectiveness. The programme is based on an ongoing training programme for a cohort of around 30 district-based communication skills facilitators, and was set up in the autumn of 1995. These facilitators are now able to act as a resource to cascade high-quality communication skills teaching into vocational training schemes, trainer education and the continuing professional development of established general practitioners throughout each district in the region. The project has now been extended into medical student teaching, specialist teaching at junior and senior level, and multi-disciplinary teaching.     

The use and implications of do not resuscitate orders in intensive care units

To describe current "do not resuscitate" (DNR) order writing practices, we studied 7,265 intensive care unit (ICU) admissions at 13 hospitals. All of the ICUs used DNR orders and 39% of all in-unit deaths were preceded by them. Patients with DNR orders were often elderly and in severely failing health. They were more severely ill than other patients in ICUs, and often had multiple organ failure. Most patients with DNR orders (94%) died in the hospital, and 86% died or were discharged from the ICU three days after a DNR order. The frequency of DNR orders ranged from 0.4% to 13.5%, and the mean interval from ICU admission to DNR order was from 5.4 to 24 days. These variations could not be explained by differences in patient characteristics, and may reflect varying physician attitudes. Do not resuscitate orders are now an accepted practice in ICUs and their use follows basic ethical and scientific guidelines. The brief interval between writing a DNR order and death or ICU discharge suggests that they often represent a decision point for placing broader limits on therapy.     

Identification of N-{varepsilon}-(2-propenal)lysine as the main form of malondialdehyde in food digesta

The form(s) in which malondialdehyde (MDA), a mutagenic productof the oxidative decomposition of polyunsaturated fatty acids(PUFA), occurs in foods was investigated. Several foods of animalorigin (ground beef, smoked fish, chicken, sausages) were digestedin vitro using pepsin and porcine intestinal fluid and the occurrenceof MDA derivatives in the digesta was investigated by thin-layer,column and high-performance liquid chromatography. The predominantform of MDA was shown to be identical to synthetic N--propenallysine. This compound is apparently formed by a reaction betweenfree MDA generated as a product of the oxidative rancidity ofPUFA in foods and the free -amino groups of proteins, from whichit is released in the course of digestion. It has been shownto be excreted in rat and human urine partially in the unchangedform and partially as the N--acetyl derivative. The resultsof this study serve to mitigate concern over the possible carcinogenicityof MDA in the diet, since <10% of the MDA in several foodscontaining highly unsaturated fatty acids was found in the freeform.