Quantitative estimation of Dehalococcoides mccartyi at laboratory and field scale: comparative study between CARD-FISH and Real Time PCR.

Dehalococcoides mccartyi detectability in the field is a fundamental tool to assess the efficiency of natural attenuation or engineered bioremediation in chlorinated solvent-contaminated sites. This study reports on the direct comparison of quantitative data obtained by Real Time PCR (qPCR) and CAtalyzed Reporter Deposition-Fluorescence In situ Hybridization (CARD-FISH) over a wide range of Dehalococcoides concentrations (10-10(8) cells mL(-1)) both in three independent 10-fold serial dilutions of a laboratory dechlorinating enrichment and in 49 groundwater samples from 6 different contaminated sites. Dehalococcoides enumeration by CARD-FISH yielded a linear curve in the analyzed concentration range which was consistent with the expected concentrations and showed good reproducibility in triplicate assays. Alternatively, qPCR did not allow for the discrimination of 16S rRNA gene concentrations lower than 10(3) gene copies mL(-1) either in the dechlorinating mixed culture or in field samples. Overall this study highlights the limits of qPCR quantification, especially in samples where low concentrations of this microorganism may be expected, and suggests the use of a confirmatory methodology under these particular conditions.

Comparative proteomics of Dehalococcoides spp. reveals strain-specific peptides associated with activity.

Anaerobic reductive dehalogenation by Dehalococcoides spp. is an ideal system for studying functional diversity of closely related strains of bacteria. In Dehalococcoides spp., reductive dehalogenases (RDases) are key respiratory enzymes involved in the anaerobic detoxification of halogenated compounds at contaminated sites globally. Although housekeeping genes sequenced from Dehalococcoides spp. are >85% identical at the amino acid level, different strains are capable of dehalogenating diverse ranges of compounds, depending largely on the suite of RDase genes that each strain harbors and expresses. We identified RDase proteins that corresponded to known functions in four characterized cultures and predicted functions in an uncharacterized Dehalococcoides-containing mixed culture. Homologues within RDase subclusters containing PceA, TceA, and VcrA were among the most frequently identified proteins. Several additional proteins, including a formate dehydrogenase-like protein (Fdh), had high coverage in all strains and under all growth conditions.

Lignocaine aerosol spray in outpatient hysteroscopy: a randomized double-blind placebo-controlled trial.

OBJECTIVE: To assess the efficacy of lignocaine spray during outpatient hysteroscopy in reducing the need for additional anesthesia and reducing the discomfort of the procedure. DESIGN: A randomized double-blind, placebo-controlled trial. SETTING: An undergraduate university teaching hospital in London. PATIENT(S): One hundred twenty patients undergoing outpatient hysteroscopy. INTERVENTION(S): Application of lignocaine spray to the cervix, cervical canal, and uterine cavity during outpatient hysteroscopy. MAIN OUTCOME MEASURE(S): The need to use additional anesthesia and the pain experienced at various steps of the procedure. RESULT(S): Women treated with active spray experienced significantly less pain when the cervix was grasped with a tenaculum at the start of hysteroscopy. There were no other significant differences in the outcome of hysteroscopy between the placebo and lignocaine groups, although there was a significant reduction in the use of additional anesthesia in both groups compared with historical controls. CONCLUSION(S): Lignocaine spray has beneficial effects on cervical but not uterine sensation. Pretreatment with either lignocaine or placebo seems to reduce the need for additional intracervical anesthesia during hysteroscopy.

Vaginal hysterectomy for the large uterus.

OBJECTIVE: To assess the feasibility and safety of performing vaginal hysterectomy on enlarged uteri the equivalent of 14 to 20 weeks of gestation in size. DESIGN: A prospective observational study. SETTING: The Royal Free Hospital, London. PARTICIPANTS: Fourteen consecutive women undergoing vaginal hysterectomy for uterine fibroids up to 20 weeks in size. INTERVENTIONS: Vaginal hysterectomy with or without bilateral salpingo-oophorectomy or oophorectomy. MAIN OUTCOME MEASURES: Uterine size and weight, techniques used to reduce uterine size, surgical outcome, operative time, estimated operative blood loss, intra- and post-operative complications, duration of hospitalisation. RESULTS: The mean uterine size was 16.3 weeks (range 14 to 20 weeks). All hysterectomies were completed successfully by the vaginal route. The uteri weighed 380 to 1100 g, with a mean of 638.7 g. Bisection combined with myomectomy and morcellation were used in most cases to obtain reduction in uterine size, whereas coring was only utilised in two cases. The mean operating time was 84.3 min with a range of 30 to 150 min. The only complications were transient haematuria (n = 6) and superficial vaginal grazes (n = 5). One of the women required a blood transfusion. The mean post-operative hospital stay was 3.7 days (range 2 to 9 days). CONCLUSION: Enlargement of the uterus to a size equivalent to 20 weeks of gestation should no longer be considered a contraindication to vaginal hysterectomy. Many more hysterectomies should be carried out vaginally without resorting to abdominal or laparoscopic surgery.

Is laparoscopic hysterectomy a waste of time?

Laparoscopic hysterectomy (LH) is a way to avoid laparotomy. However, there is evidence that most women treated by abdominal hysterectomy are suitable for vaginal surgery. To test this hypothesis, and to determine the relative merits of laparoscopic and vaginal hysterectomy (VH) and the best technique for LH, we prospectively studied 98 women who had relative contraindications for vaginal surgery by traditional criteria. 75 underwent LH and 23 VH. The LH group included 22 women who had been assigned to this route of surgery as part of a prospective randomised controlled comparison with VH (23 women). Surgery was completed with the intended technique in 93.9% of cases. 5 women in the LH group (6.7%) and 2 in the VH group required laparotomy or additional procedures. In the prospective randomised study LH took longer than VH (mean duration 131 vs 77 min). VH was the faster procedure, irrespective of uterine size and need for oophorectomy. With LH, the operative time increased as more of the hysterectomy was carried out with laparoscopic rather than vaginal dissection. Complication rates, blood loss, analgesia requirements, and recovery were similar for the two techniques. Our study confirms that most hysterectomies could be performed vaginally, and that LH is a much slower procedure. If LH is done, it should be converted to a vaginal procedure as early as possible to reduce the overall operating time. LH does seem to be a waste of time for most patients.

Transvaginal endoscopic oophorectomy.

Four women underwent transvaginal endoscopic oophorectomy during vaginal hysterectomy. The adnexa were visualized with a laparoscope inserted into the upper vagina. Bilateral salpingo-oophorectomy or oophorectomy was carried out with standard laparoscopic instruments introduced through the vagina without a pneumoperitoneum; Endoloop sutures and bipolar electrocoagulation were used for hemostasis.

Analysis of reciprocal creatinine plots by two-phase linear regression.

The progression of renal diseases is often monitored by the serial measurement of plasma creatinine. The slope of the linear relation that is frequently found between the reciprocal of creatinine concentration and time delineates the rate of change in renal function. Minor changes in slope, perhaps indicating response to therapeutic intervention, can be difficult to identify and yet be of clinical importance. We describe the application of two-phase linear regression to identify and characterise changes in slope using a microcomputer. The method fits two intersecting lines to the data by computing a least-squares estimate of the position of the slope change and its 95% confidence limits. This avoids the potential bias of fixing the change at a preconceived time corresponding with an alteration in treatment. The program then evaluates the statistical and clinical significance of the slope change and produces a graphical output to aid interpretation.

Comparison of two strengths of iohexol and iothalamate in urography.

Iohexol and iothalamate were studied in 2 strengths and compared with results from a previous trial of Iopamidol and 4 ionic agents. Scoring was consistent from trial to trial. No worthwhile increase in density was achieved at high doses using nonionics and pyelographic distention was less than for ionic media. The timing of the nephrogram was the same for ionics and nonionics. There is no need to adopt a different film sequence for nonionic media. Urticarial reactions were identical for all 4 media studied in this trial. Nonionic media perform as well as other media; at lower doses they are much better than meglumine salts of ionic media.

The old and the new: a study of five contrast media for urography.

Five contrast media, Conray 280 and 420, Urografin 370, Uromiro Sodium 300 and Niopam 370, were compared in a randomised trial involving a total of 482 patients. The best urographic agent was Conray 420 and the worst Conray 280, these control agents defining the ends of the scoring system. Uromiro Sodium 300 was very nearly as good as Conray 420. A non-ionic agent, Niopam 370, scored nearly equal with Urografin 370; both were rather better than Conray 280. There was little difference in minor reactions between the media. No reason was found to prefer non-ionic to ionic agents for general use in urography; indeed for a diagnostic examination the sodium salt of an ionic agent is preferable.

A rigorous comparison of real time and B scan cephalometry.

Real time cephalometry has been compared with B scan cephalometry by a rigorous method with complete partition of variables. Analysis was by standard analysis of variance techniques. Systematic and random errors between and within observers, machines and days have been evaluated. The largest systematic error has been shown to be between machines. This is due to failure to align the beam precisely at right angles to the mid-line of the fetal head using B scanners. Real time scanners with bar calipers are, therefore, superior to B scanners for cephalometry. There is a lesser difference between observers. An additional element of variability arises when measurements are made on different days but this is smaller than other sources of variation and smaller than the random variation. Random errors are described by a standard deviation of 1 mm and systematic errors are all numerically smaller than this.

Dynamic fetal cephalometry.

An iterative method of fetal cephalometry for use with real time scanners is described using a flow chart. The method is shown to be capable of yielding measurements having a standard deivation of less than 1 mm and we have found it to be superior to the combined A and B scan technique from which it is derived.

Folates of rat tissue. Bioassay of tissue folylpolyglutamates and a relationship of liver folypolyglutamates to nutritional folate sufficiency.

The folate content of young rat tissues extracted into boiling ascorbate was assayed by Lactobactillus casei both without and after treatment by a folate-free preparation of conjugase. The total folate content of various tissues was: liver, 8.9 microgram/g; kidney, 2.6; adrenal, 2.6; bone marrow, 2.4; spleen, 0.9; erythrocytes, 0.8; small intestinal mucosa, 0.7; small intestinal smooth muscle, 0.8; heart, 0.6; brain, 0.4, and skeletal muscle, 0.1 microgram/g tissue. For most tissues, with the exception of muscle and kidney, approximately 80% of the total folates assayed as longer chain length folylpolyglutamates. When liver folates were analyzed from rats fed folate-supplemented, control and folate-deficient diets, a relationship was found between folate nutrition and distribution of folylpolyglutamates. The proportion of total folates in the form of longer chain length folylpolyglutamates was greatest in the livers of folate-deficient rats and least in the livers of folate-supplemented rats.

Fetal cephalometry. An improved method and an evaluation.

A method of fetal cephalometry is described which is an advance on the combined A and B scan technique previously used. A flow chart has been prepared which clearly specifies the procedure to be followed. The criteria used when making an estimation are reported. The method is applicable to various types of machine and it can be taught to a good degree of accuracy in a short time to people with limited obstetric knowledge. There may be a significant difference between operators but this difference is roughly the same from patient to patient and decreases with training. It is considered that the chief source of variation is the failure to measure the same diameter on each occasion.