Overproduction of isoprenoids by Saccharomyces cerevisiae in a synthetic grape juice medium in the absence of plant genes.

The objective of this work is to demonstrate if the hexaprenyl pyrophosphate synthetase Coq1p might be involved in monoterpenes synthesis in Saccharomyces cerevisiae, although its currently known function in yeast is to catalyze the first step in ubiquinone biosynthesis. However, in a BY4743 laboratory strain, the presence of an empty plasmid in a chemically defined grape juice medium results in a statistically significant increase of linalool, (E)-nerolidol and (E,E)-farnesol. When COQ1 is overexpressed from a plasmid, the levels of the volatile isoprenoids are further increased. Furthermore, overexpression of COQ1 in the same genetic context but with a mutated farnesyl pyrophosphate synthetase (erg20 mutation K197E), results in statistically significant higher levels of linalool (above 750 µg/L), geraniol, α-terpineol, and the sesquiterpenes, farnesol and nerolidol (total concentration of volatile isoprenoids surpasses 1300 µg/L). We show that the levels of monoterpenes and sesquiterpenes that S. cerevisiae can produce, in the absence of plant genes, depend on the composition of the medium and the genetic context. To the best of our knowledge, this is the highest level of linalool produced by S. cerevisiae up to now. Further research will be needed for understanding how COQ1 and the medium composition might interact to increase flavor complexity of fermented beverages.

Endorectal mucosal advancement flap: the preferred method for complex cryptoglandular fistula-in-ano.

Cryptoglandular fistula-in-ano is a common affliction that usually responds well to conventional surgical procedures such as fistulectomy, fistulotomy, and seton placement. These procedures, however, can be associated with varying degrees of fecal incontinence. Endorectal mucosal advancement flap has been advocated as an alternative procedure that avoids this problem. This study was undertaken to determine the risks and benefits associated with endorectal mucosal advancement flap in the treatment of complex fistula-in-ano. One hundred sixty-four patients underwent 167 endorectal mucosal advancement flap procedures for complex cryptoglandular fistula-in-ano between January 1982 and December 1990. There were 126 men and 38 women whose mean age was 42.1 years (range 20 to 79 years). The majority of the patients (70%) had complex fistulas (transsphincteric, suprasphincteric, or extrasphincteric). Fifteen patients (9%) had an intersphincteric fistula. All patients were available for short-term follow-up (6 weeks). Postoperative morbidity was minimal and included urinary retention in 13 patients (7.8%) and bleeding in one patient. Healing time averaged 6 weeks. Long-term follow-up, ranging from 19 to 135 months, was carried out in 61 patients. There were two recurrences (3.28%). Nine patients (15%) complained of varying degrees of fecal incontinence. Six patients complained of incontinence to flatus and three patients complained of incontinence to liquid stool. No patient was incontinent of solid stool. Sixty patients (98%) rated their functional result as excellent or good. Endorectal mucosal advancement flap is a safe and effective technique for the treatment of complex cryptoglandular fistula-in-ano. It can be performed with minimal morbidity, no mortality, an acceptable recurrence rate, and little alteration in anorectal continence.

Complete obstruction of the sigmoid colon treated by primary resection and anastomosis--an improved technique (preliminary report). Report of three cases.

Three patients with complete colonic obstruction treated by primary resection and anastomosis with intraoperative colon tube decompression and bowel lumen sterilization without a protective colostomy are presented. An improved colonic decompressor was used. It is postulated that this procedure is an alternative safe technique in patients with colonic obstruction in whom an end-colostomy, mucous fistula, or Hartmann pouch would be necessary.

Vaginography: an easy and safe technique for diagnosis of colovaginal fistulas.

Colovaginal fistulas are often difficult to demonstrate. Vaginography is a simple, safe, and effective technique.

Rectocele repair. Four years' experience.

A retrospective review of 64 rectocele repairs done over a four-year period was performed. The most common indication for repair was constipation. Thirty-five patients were repaired transanally, and 29 were repaired transvaginally. The overall morbidity was 34 percent, and the overall mortality was 0 percent. The most common complication was urinary retention in 12.5 percent. There was no difference in complications between techniques. Of 46 patients contacted for follow-up, 25 (54 percent) still complained of constipation, 17 (34 percent) had partial incontinence, 8 (17 percent) noted persistent rectal pain, 15 (32 percent) mentioned occasional rectal bleeding, and 10 (22 percent) complained of vaginal tightness or sexual dysfunction. Thirty-seven (80 percent) patients stated that they had improved after surgery. Except for persistent rectal pain, there was no difference in results between transanal and transvaginal repairs. Those undergoing transvaginal repair had a much greater problem with pain. Our relatively poor results may be due to an unselective approach to rectocele repair. The presence of both constipation and a rectocele does not imply an association, and a complete anorectal physiologic examination should precede repair. There is no functional difference between transvaginal and transanal rectocele repair.

Prevention of colostomy in partial colonic obstruction by intraoperative rectal tube irrigation.

Four patients with partial obstruction of the colon, in whom adequate standard preparation for colonic resection was not possible, are presented. Clinically, these patients did not show sufficient signs of obstruction to require a preliminary colostomy. A technique of quarantining the area of tumor resection from the abdominal cavity is described, allowing for adequate preparation of the colon lumen at the site of anastomosis. After completion of the anastomosis, a large caliber (36 French) colonic tube is inserted, per rectum, through the anastomosis and the proximal colon irrigated with Betadine solution adequately emptying and sterilizing the lumen of the bowel to allow for primary anastomosis without a protective colostomy. The criteria for selection of patients for this technique are discussed. No complications, of significance, ensued. This method of intraoperative rectal tube irrigation, for prevention of colostomy in partial colonic obstruction, is safe when properly performed in the appropriate patient.

Colonoscopy-assisted colostomy--an alternative to laparotomy: report of two cases.

PURPOSE: The objective of this study was to evaluate the feasibility of performing fecal diversion with the help of a colonoscope without a concomitant laparotomy. METHODS: Colostomies were performed on two patients who needed fecal diversion and who would benefit from avoiding the morbidity of laparotomy. A colonoscope was used in each case to guide the surgeon in selecting the appropriate bowel segment. RESULTS: No complications related to the colostomy were noted in either patient. CONCLUSIONS: The technique of colonoscopy-assisted colostomy that we have described offers an acceptable method of creating a stoma without the need for laparotomy.

A Bacillus subtilis gene induced by cold shock encodes a membrane phospholipid desaturase.

Bacillus subtilis grown at 37 degrees C synthesizes saturated fatty acids with only traces of unsaturated fatty acids (UFAs). However, when cultures growing at 37 degrees C are transferred to 20 degrees C, UFA synthesis is induced. We report the identification and characterization of the gene encoding the fatty acid desaturase of B. subtilis. This gene, called des, was isolated by complementation of Escherichia coli strains with mutations in either of two different genes of UFA synthesis. The des gene encodes a polypeptide of 352 amino acid residues containing the three conserved histidine cluster motifs and two putative membrane-spanning domains characteristic of the membrane-bound desaturases of plants and cyanobacteria. Expression of the des gene in E. coli resulted in desaturation of palmitic acid moieties of the membrane phospholipids to give the novel mono-UFA cis-5-hexadecenoic acid, indicating that the B. subtilis des gene product is a delta5 acyl-lipid desaturase. The des gene was disrupted, and the resulting null mutant strains were unable to synthesize UFAs upon a shift to low growth temperatures. The des null mutant strain grew as well as its congenic parent at 20 or 37 degrees C but showed severely reduced survival during stationary phase. Analysis of operon fusions in which the des promoter directed the synthesis of a lacZ reporter gene showed that des expression is repressed at 37 degrees C, but a shift of cultures from 37 to 20 degrees C resulted in a 10- to 15-fold increase in transcription. This is the first report of a membrane phospholipid desaturase in a nonphotosynthetic organism and the first direct evidence for cold induction of a desaturase.

Transcriptional control of the low-temperature-inducible des gene, encoding the delta5 desaturase of Bacillus subtilis.

The Bacillus subtilis des gene encodes the cold-inducible Delta5 lipid desaturase involved in the formation of unsaturated fatty acids from saturated phospholipid precursors. Here, we describe the expression pattern of the des gene in response to a temperature downshift from 37 to 20 degrees C. We found that the synthesis of des mRNA is undetectable at 37 degrees C but dramatically induced upon the temperature downshift. Decay characteristics of the des transcript as well as the in vivo decay of B. subtilis bulk mRNA were investigated. The results showed that the stability of the des transcript as well as of bulk mRNA lasted substantially longer at 20 degrees C than at 37 degrees C. Functional expression of des at 37 degrees C was achieved by exchanging its promoter with the non-cold shock spac promoter. These data provide the first direct evidence that temperature-mediated control of transcription is the major mechanism regulating the mRNA levels of the B. subtilis desaturase. The present results also demonstrate that the only component of the desaturation system regulated by temperature is the desaturase enzyme.

Mucosal advancement in the treatment of anal fistula.

One hundred eighty-nine patients with anal fistula treated within an eight-month to seven-year period by anal fistulectomy and rectal mucosal advancement are presented. An 80 percent follow-up revealed a 90 percent asymptomatic group and a ten percent group who had minor symptoms. Eight percent of the symptomatic patients had minor soiling; 7 percent were incontinent for gas, and 6 percent were incontinent for loose stools. No patient was incontinent for solid feces. There was a 1.5 percent rate of recurrent anal fistula comparable to other techniques.

Reconstruction of rectovaginal fistula with sphincter disruption by combining rectal mucosal advancement flap and anal sphincteroplasty.

PURPOSE: This study evaluated the effectiveness of combining advancement flap with sphincteroplasty in patients symptomatic with rectovaginal fistula and anal sphincter disruption. METHODS: Twenty patients with rectovaginal fistulas and anal sphincter disruptions after vaginal deliveries underwent combined rectal mucosal advancement flap and anal sphincteroplasty between July 1986 and July 1993. The mean age of the patients was 30 (range, 18-40) years and the mean duration of symptoms was 54.8 weeks (range, 7 weeks to 6 years). In addition to mucosal advancement flap repair, 13 patients underwent two-layer repair of anal sphincters (with reapproximation of the puborectalis in 8 of the patients); 6 patients underwent one-layer overlap repair of anal sphincters (with reapproximation of the puborectalis in 2 of the patients); and 1 patient underwent reapproximation of internal anal sphincter alone because squeeze pressures were adequate, as determined by anal manometry. RESULTS: Postoperatively, vaginal discharge of stool and flatus was eliminated entirely in all 20 patients. Perfect anal continence of stool and flatus was restored in 14 patients (70 percent). Incontinence was improved but not eliminated in six patients (4 incontinent to liquid stool and 2 to flatus), and two patients required perineal pads. Subjectively, 19 patients (95 percent) reported the result as excellent or good. There were no complications. CONCLUSION: The combination of mucosal advancement flap and anal sphincteroplasty is a safe and highly effective procedure for correcting rectovaginal fistula with sphincter disruption after obstetrical injuries.

Survival after colonic perforation during barium-enema examination. Modified radical surgical debridement.

Four of five patients survived perforation of the colon incidental to barium-enema examination, while the other survived without sequela of infection until hemiplegia and pneumonia supervened two and 36 days, respectively, postoperatively. It is postulated that adequate intravenous fluids, early operation, with complete cleansing by antibiotic irrigation and mechanical debridement of the abdominal cavity, along with excision of the perforated segment of bowel with establishment of an end colostomy or ileostomy and distal mucous fistula, if possible, are keys to survival in these patients. Triple antibiotics, using ampicillin, gentamicin, and clindamycin, or metronidazole, should be administered in all patients as soon as the diagnosis is made, and continued postoperatively as indicated until there is no evidence of infection. An intraluminal intestinal tube, for stichless plication of the small bowel, may be helpful in some patients. A stomacone should be used for a barium enema through a colostomy stoma. Care should be exercised in performing barium contrast studies in patients with inflammatory bowel disease.

The efficacy of barium-enema examinations in patients with anorectal disease.

This retrospective study examines the value of an air contrast barium enema examination in detecting proximal neoplasia in the patient presenting with benign anorectal disease as determined by history, physical examination, rigid, and flexible sigmoidoscopy. In 428 of these patients, the roentgenographic studies showed proximal colonic cancer or polyps in less than 1 percent of patients reviewed. In addition, a review of 402 patients with known colon and rectal cancer were surveyed using the same criteria for diagnosis, and less than 1 percent were misinterpreted as having benign anorectal disease.

Urecholine prophylaxis for urinary retention in anorectal surgery.

A randomized prospective trial with 108 patients undergoing anorectal surgery was conducted comparing the use of Urecholine orally or subcutaneously to no treatment controls. There was no difference in postoperative urinary retention rates and caudal or general anesthesia, nor was there an earlier postoperative bowel movement with Urecholine. The volume of intravenous fluids significantly affected retention rates.

Surgical treatment of low rectovaginal fistulas.

Forty women with low rectovaginal fistulas were operated upon over a 9-year period. The etiology of the fistula in the majority was obstetric. Nine women had prior attempts to repair the fistula. All 40 women were managed with endorectal advancement flap with the addition of sphincteroplasty or perineal body reconstruction in 15 patients and rectocele repair in six patients. Postoperative complications included urinary difficulties (two patients) and wound complications (three patients). There were two recurrences. All women treated with sphincteroplasty or perineal body reconstruction were continent. Seven women complained of varying degrees of incontinence postoperatively; none had undergone sphincter or perineal body reconstruction. Endorectal advancement flap is a safe and effective operation for women with rectovaginal fistulas. Concomitant sphincteroplasty or perineal body reconstruction should be performed in women with historical, physical, or manometric evidence of incontinence.

Abdominal colon and rectal operations in the elderly.

Sixty-seven abdominal operations for colon and rectal disorders were performed on 56 patients 80 years of age or older from January 1, 1984 to June 30, 1989. Nine patients required multiple operations. Sixty-two procedures (92 percent) were performed on patients in their ninth decade; two operations were performed on patients 95 years of age or older. Forty-five patients (80 percent) were operated upon for carcinoma. Operations included segmental colectomy (33 patients), low anterior resection (12 patients), total abdominal colectomy (3 patients) and abdominoperineal resection (2 patients). Forty patients were classified as ASA Class III; the majority were monitored in the surgical intensive care unit for a mean of 2.84 days. Thirty patients were monitored with arterial catheters and 21 with central invasive monitoring. Operative mortality was 7 percent (4 patients). Two patients died from diffuse carcinomatosis; one patient had a fatal myocardial infarction. The final death occurred from multisystem organ failure following anastomotic dehiscence. Twenty-seven operations were performed without postoperative complications; 18 operations were followed by a single minor complication. The average hospital stay was 18.96 days. All patients were admitted from home. Thirty-three returned home postoperatively; 16 were discharged to an extended care facility. In conclusion, elderly patients with colon and rectal disorders can be operated upon with acceptable morbidity and mortality. Age alone should not interdict surgical therapy.

Delayed repair of obstetric injuries of the anorectum and vagina. A stratified surgical approach.

PURPOSE: We categorized the various types of post-obstetric injuries of the anorectum and vagina encountered in a five-year period. The operative procedures used to repair these injuries and the functional outcome after surgery were assessed. METHODS: Between 1986 and 1991, 52 patients were surgically treated for obstetric injuries of the anorectum and vagina; 48 patients were available for follow-up study. Four clinical injury types were identified: Type I, incontinent and sphincter (11 patients); Type II, rectovaginal fistula (16 patients); Type III, rectovaginal fistula and incontinent and sphincter (11 patients); and Type IV, cloaca-like defect (10 patients). The mean age of the patients was 30 years, the mean duration of symptoms before surgery was 13 months, and the mean follow-up period was 16 months. The major component of surgical repair for each injury type was: Type I, overlap repair of external anal sphincter; Type II, rectal mucosal advancement flap; Type III, overlap repair of external anal sphincter and rectal mucosal advancement flap; and Type IV, overlap repair of external anal sphincter, anterior levatorplasty, and anal and vaginal mucosal reconstruction. Fecal diversion was not performed in any patient. Specific questions were asked at the most recent follow-up assessment to determine results. RESULTS: Continence status postoperatively was classified as perfect, impaired, or poor; poor was defined as no improvement or worse. Postoperative continence (perfect impaired, or poor) was, respectively: Type I (11 patients), 64 percent, 36 percent, and 0 percent; Type II (16 patients), 56 percent, 0 percent, and 44 percent; Type III (11 patients), 64 percent, 36 percent, and 0 percent; and Type IV (10 patients), 90 percent, 10 percent, and 0 percent. Vaginal discharge of stool was eliminated in all patients with a rectovaginal fistula. Subjectively, 92 percent of the patients had excellent or good results. Complications included wound hematoma (n = 2), fecal impaction (n = 2), urinary retention (n = 1), and urinary tract infection (n = 1). CONCLUSION: Patients with Type II injuries had the worst results (P < 0.001). These patients should be evaluated for anal incontinence before surgery to assess the need for a concomitant sphincteroplasty.

Molecular basis of thermosensing: a two-component signal transduction thermometer in Bacillus subtilis.

Both prokaryotes and eukaryotes respond to a decrease in temperature with the expression of a specific subset of proteins. Although a large body of information concerning cold shock-induced genes has been gathered, studies on temperature regulation have not clearly identified the key regulatory factor(s) responsible for thermosensing and signal transduction at low temperatures. Here we identified a two-component signal transduction system composed of a sensor kinase, DesK, and a response regulator, DesR, responsible for cold induction of the des gene coding for the Delta5-lipid desaturase from Bacillus subtilis. We found that DesR binds to a DNA sequence extending from position -28 to -77 relative to the start site of the temperature-regulated des gene. We show further that unsaturated fatty acids (UFAs), the products of the Delta5-desaturase, act as negative signalling molecules of des transcription. Thus, a regulatory loop composed of the DesK-DesR two-component signal transduction system and UFAs provides a novel mechanism for the control of gene expression at low temperatures.

A new silicone-prosthesis in the modified Thiersch operation.

Experience with a new silicone prosthesis in the modified Thiersch operation for rectal procidentia in 16 extremely poor-risk patients is presented. The technique of implantation, structural details of the prosthesis, and the clinical results are described. The use of a new silicone prosthesis in the modified Thiersch procedure is a viable alternative in this group of patients. Surgical technique is a primary determining factor in preventing complications.