In vitro Characterization of Phenylacetate Decarboxylase, a Novel Enzyme Catalyzing Toluene Biosynthesis in an Anaerobic Microbial Community.

Anaerobic bacterial biosynthesis of toluene from phenylacetate was reported more than two decades ago, but the biochemistry underlying this novel metabolism has never been elucidated. Here we report results of in vitro characterization studies of a novel phenylacetate decarboxylase from an anaerobic, sewage-derived enrichment culture that quantitatively produces toluene from phenylacetate; complementary metagenomic and metaproteomic analyses are also presented. Among the noteworthy findings is that this enzyme is not the well-characterized clostridial p-hydroxyphenylacetate decarboxylase (CsdBC). However, the toluene synthase under study appears to be able to catalyze both phenylacetate and p-hydroxyphenylacetate decarboxylation. Observations suggesting that phenylacetate and p-hydroxyphenylacetate decarboxylation in complex cell-free extracts were catalyzed by the same enzyme include the following: (i) the specific activity for both substrates was comparable in cell-free extracts, (ii) the two activities displayed identical behavior during chromatographic separation of cell-free extracts, (iii) both activities were irreversibly inactivated upon exposure to O2, and (iv) both activities were similarly inhibited by an amide analog of p-hydroxyphenylacetate. Based upon these and other data, we hypothesize that the toluene synthase reaction involves a glycyl radical decarboxylase. This first-time study of the phenylacetate decarboxylase reaction constitutes an important step in understanding and ultimately harnessing it for making bio-based toluene.

Effects of imposed salinity gradients on dissimilatory arsenate reduction, sulfate reduction, and other microbial processes in sediments from two California soda lakes.

Salinity effects on microbial community structure and on potential rates of arsenate reduction, arsenite oxidation, sulfate reduction, denitrification, and methanogenesis were examined in sediment slurries from two California soda lakes. We conducted experiments with Mono Lake and Searles Lake sediments over a wide range of salt concentrations (25 to 346 g liter(-1)). With the exception of sulfate reduction, rates of all processes demonstrated an inverse relationship to total salinity. However, each of these processes persisted at low but detectable rates at salt saturation. Denaturing gradient gel electrophoresis analysis of partial 16S rRNA genes amplified from As(V) reduction slurries revealed that distinct microbial populations grew at low (25 to 50 g liter(-1)), intermediate (100 to 200 g liter(-1)), and high (>300 g liter(-1)) salinity. At intermediate and high salinities, a close relative of a cultivated As-respiring halophile was present. These results suggest that organisms adapted to more dilute conditions can remain viable at high salinity and rapidly repopulate the lake during periods of rising lake level. In contrast to As reduction, sulfate reduction in Mono Lake slurries was undetectable at salt saturation. Furthermore, sulfate reduction was excluded from Searles Lake sediments at any salinity despite the presence of abundant sulfate. Sulfate reduction occurred in Searles Lake sediment slurries only following inoculation with Mono Lake sediment, indicating the absence of sulfate-reducing flora. Experiments with borate-amended Mono Lake slurries suggest that the notably high (0.46 molal) concentration of borate in the Searles Lake brine was responsible for the exclusion of sulfate reducers from that ecosystem.

Spontaneous kidney allograft rupture.

Spontaneous renal allograft rupture is one of the most dangerous complications of kidney transplantation, which can result in graft loss. This condition needs immediate surgical intervention. Conservative management has dismal results. Its prevalence varies from 0.3% to 3%. Rupture occurs in first few weeks after transplantation. Predisposing factors for graft rupture are acute rejection, acute tubular necrosis, and renal vein thrombosis. There are growing reports about successful results of repairing these ruptured kidneys. In this study, we reviewed the medical records of 1682 patients who received kidney allografts from living donors from 1986 through 2003. There were six (0.35%) cases of renal allograft rupture. All were preceded by acute graft rejection. They were treated with antirejection medications. In first three cases, the kidney allografts were removed because the procedure of choice in this situation is graft nephrectomy; but in three next cases we repaired the ruptured grafts with good results in two of them. In conclusion, the procedure of choice for kidney allograft rupture is graft repair.

The results of grade IV cystocele repair using mesh.

PURPOSE: To evaluate the results of grade IV cystocele repair by 4-corner bladder and bladder neck suspension technique, using prolene mesh. MATERIAL AND METHODS: Thirty-one women with a median age of 61 years and severe anterior vaginal wall prolapse (grade IV cystocele) were treated by 4-corner bladder and bladder neck suspension technique, using prolene mesh. Of these, 3 had associated uterine prolapse, rectocele, and enterocele, one had rectocele and enterocele, and 18 had rectocele only. In these cases, pelvic floor defects were also repaired simultaneously and in 3, vaginal hysterectomy was done. Twelve patients had a previous failed cystocele repair. In a 32-month follow-up, the patients were evaluated with vaginal examination and upright cystography. Urinary continence during increased intra-abdominal pressure was also assessed, based on subjective symptoms. RESULTS: None of the patients had cystocele recurrence. Urinary continence during increased intra-abdominal pressure was seen in all of the patients. Intraoperative rectal or bladder injury did not occur. Transfusion was not required in any of the cases. Early complications (6 to 8 weeks postoperatively) included irritative urinary symptom in 17 patients, of whom, 8 had documented urinary tract infection that were treated successfully. Late complications were spotting in 3 cases (two were treated with topical estrogen and vaginal mucosal repair was done in one), dyspareunia in 4 sexually active patients, changes in urination pattern in 28 (improved significantly with behavioral therapy), long-term urge incontinence (>8 weeks) in 5 (medical treatment was successful in these patients), and prolonged intermittent catheterization in 1. Pelvic abscess and migration of mesh were not observed. CONCLUSION: According to our findings, using mesh in patients with grade IV cystocele, who had a previous failed surgery or weakness in supportive pelvic tissue, is an appropriate treatment modality.