Development of new ethanologenic Escherichia coli strains for fermentation of lignocellulosic biomass.

Two new ethanologenic strains (FBR4 and FBR5) of Escherichia coli were constructed and used to ferment corn fiber hydrolysate. The strains carry the plasmid pLOI297, which contains the genes from Zymomonas mobilis necessary for efficiently converting pyruvate into ethanol. Both strains selectively maintained the plasmid when grown anaerobically. Each culture was serially transferred 10 times in anaerobic culture with sugar-limited medium containing xylose, but no selective antibiotic. An average of 93 and 95% of the FBR4 and FBR5 cells, respectively, maintained pLOI297 in anaerobic culture. The fermentation performances of the repeatedly transferred cultures were compared with those of cultures freshly revived from stock in pH-controlled batch fermentations with 10% (w/v) xylose. Fermentation results were similar for all the cultures. Fermentations were completed within 60 h and ethanol yields were 86-92% of theoretical. Maximal ethanol concentrations were 3.9-4.2% (w/v). The strains were also tested for their ability to ferment corn fiber hydrolysate, which contained 8.5% (w/v) total sugars (2.0% arabinose, 2.8% glucose, and 3.7% xylose). E. coli FBR5 produced more ethanol than FBR4 from the corn fiber hydrolysate. E. coli FBR5 fermented all but 0.4% (w/v) of the available sugar, whereas strain FBR4 left 1.6% unconsumed. The fermentation with FBR5 was completed within 55 h and yielded 0.46 g of ethanol/g of available sugar, 90% of the maximum obtainable.

Adaptive response to cold temperatures in Vibrio vulnificus.

The effectiveness of rapid chilling or freezing of oysters to reduce Vibrio vulnificus levels in shellfish may be compromised by product handling procedures that permit cold adaptation. When a V. vulnificus culture was shifted from 35 degrees C to 6 degrees C conditions, it underwent transition to a non-culturable state. Cells adapted to 15 degrees C prior to change to 6 degrees C condition, however, remain viable and culturable. In addition, cultures adapted to 15 degrees C were able to survive better upon freezing at -78 degrees C compared with cultures frozen directly from 35 degrees C. Inhibition of protein synthesis by addition of chloramphenicol in a V. vulnificus culture immediately prior to the exposure to the adaptive temperature eliminated inducible cold tolerance. These results suggest that cold-adaptive "protective" proteins may enhance survival and tolerance at cold temperatures. In addition, removal of iron from the growth medium by adding 2,2'-Dipyridyl prior to cold adaptation decreased the viability by approximately 2 logarithm levels. This suggests that iron plays an important role in adaptation at cold temperatures. Analysis of total cellular proteins on an SDS polyacrylamide gel electrophoresis, labeled with 35S-methionine during exposure at 15 degrees C, showed elevated expressions of a 6-kDa and a 40-kDa protein and decreased expression of an 80-kDa protein. These results suggest that, for V. vulnificus, survival and tolerance at cold temperatures could be due to the expression of cold-adaptive proteins other than previously documented major cold shock proteins such as CS7.4 and CsdA. In this study, for the first time we have shown that exposure to an intermediate cold temperature (15 degrees C) causes a cold adaptive response, helping this pathogen remain in culturable state when exposed to a much colder temperature (6 degrees C). This adaptive nature to cold temperatures could be important for shellfish industry efforts to reduce the risk of V. vulnificus infection from consuming raw oysters.

Homarine as a feeding deterrent in common shallow-water antarctic lamellarian gastropodMarseniopsis mollis: A rare example of chemical defense in a marine prosobranch.

The common bright yellow antarctic lamellarian gastropodMarseniopsis mollis was examined for the presence of defensive chemistry. Proton nuclear magnetic resonance (NMR) spectroscopy indicated that a major component of ethanolic extracts purified by reversed-phase column chromatography was homarine. Further high-performance liquid chromatography (HPLC) analysis of the mantle, foot, and viscera verified the presence of homarine in all body tissues at concentrations ranging from 6 to 24 mg/g dry tissue. A conspicuous macroinvertebrate predator of the shallow antarctic benthos, the sea starOdontaster validus, always rejected live individuals ofM. mollis, while readily feeding on pieces of fish tail muscle. Filter paper disks treated with shrimp elicited a broad range of feeding behaviors in the sea starO. validus (movement of disc to mouth, extrusion of cardiac stomach, humped feeding posture). Shrimp disks treated with homarine (0.4 and 4 mg/disk) were rejected byO. validus significantly more frequently than control disks treated with solvent carrier and shrimp or shrimp alone. The highest concentration of homarine tested not only caused feeding deterrence, but in several sea stars a flight response was noted. Homarine was not detected in the tunic of the antarctic ascidianCnemidocarpa verrucosa, a presumed primary prey ofM. mollis. Nonetheless, crude extracts of the epizooites that foul the tunic (primarily the bryozoans and hydroids) contain homarine, suggestingM. mollis may ingest and derive its chemistry from these organisms. This appears to be only the third example of chemical defense in a member of the Order Mesogastropoda. As the vestigial internalized shell ofM. mollis is considered a primitive condition, the findings of this study lend support to the hypothesis that chemical defense evolved prior to shell loss in shell-less gastropods.

Chemical defense of common antarctic shallow-water nudibranchTritoniella belli eliot (Mollusca: Tritonidae) and its prey,Clavularia frankliniana rouel (Cnidaria: Octocorallia).

Extracts of the dorid nudibranchTritoniella belli and stoloniferan coralClavularia frankliniana were chromatographed and analyzed by(1)H NMR and thin-layer chromatography. Three glycerol ethers were detected inT. belli, primarily 1-O-hexadecyl glycerol (chimyl alcohol). Chimyl alcohol was also detected after gradient flash chromatography and reverse-phase HPLC purification in the tissues ofC. frankliniana. The common omnivorous predatory Antarctic sea starOdontaster validus, a likely predator of benthic invertebrates, showed feeding deterrence to small cubes ofT. belli mantle tissue placed on the tube feet along the ambulacral feeding groove, while always extruding the cardiac stomach when presented with cubes of shrimp tissue of similar size. Filter-paper disks soaked in an aqueous shrimp solution and then dried were found to elicit a broad range of feeding behaviors inO. validus, including movement of the shrimp disk to the mouth, extrusion of the cardiac stomach, and the assumption of a humped feeding posture. Chimyl alcohol-treated shrimp disks caused significant feeding deterrence in sea stars when compared with control disks (solvent plus shrimp treated disks alone).T. belli andC. frankliniana appear to employ a defensive compound that has been found in a variety of temperate and tropical mollusks, where it has been demonstrated to deter fish predators. We provide evidence for further deterrent capabilities of chimyl alcohol and of its trophic relationship in the polar ecosystem of McMurdo Sound, Antarctica.

Purification and Characterization of an alpha-l-Arabinofuranosidase from Butyrivibrio fibrisolvens GS113.

An alpha-l-arabinofuranosidase (EC 3.2.1.55) was purified from the cytoplasm of Butyrivibrio fibrisolvens GS113. The native enzyme had an apparent molecular mass of 240 kDa and was composed of eight polypeptide subunits of 31 kDa. The enzyme displayed an isoelectric point of 6.0, a pH optimum of 6.0 to 6.5, a pH stability of 4.0 to 8.0, and a temperature optimum of 45 degrees C and was stable to 55 degrees C. The K(m) and V(max) for p-nitrophenyl-alpha-l-arabinofuranoside were 0.7 mM and 109 mumol/min/mg of protein, respectively. The enzyme was specific for the furanoside configuration and also readily cleaved methylumbelliferyl-alpha-l-arabinofuranoside but had no activity on a variety of other nitrophenyl- or methylumbelliferyl glycosides. When the enzyme was incubated with cellulose, carboxymethyl cellulose, or arabinogalactan, no release of sugars was found. Arabinose was found as the hydrolysis product of oatspelt xylan, corn endosperm xylan, or beet arabinan. No activity was detected when either coumaric or ferulic acid ester linked to arabinoxylobiose was used as substrates, but arabinoxylobiose was degraded to arabinose and xylobiose. Since B. fibrisolvens GS113 possesses essentially no extracellular arabinofuranosidase activity, the major role of the purified enzyme is apparently in the assimilation of arabinose-containing xylooligosaccharides generated from xylosidase, phenolic esterase, xylanase, and other enzymatic activities on xylans.

Chemotherapy-based control of schistosomiasis haematobia. II. Metrifonate vs. praziquantel in control of infection-associated morbidity.

To determine the relative efficacy of metrifonate and praziquantel in controlling urinary tract morbidity due to Schistosoma haematobium infection, a random allocation treatment trial was performed among 1,813 school age S. haematobium-infected children from the Msambweni area of Coast Province, Kenya. Following baseline examination for infection, hematuria, proteinuria, and ultrasonographic urinary tract abnormalities, oral treatment with either metrifonate (10 mg/kg, repeated at 4 month intervals) or praziquantel (1 dose of 40 mg/kg) was given to infected subjects. Prevalence of morbidity was reassessed 12 months later for each treatment group. Results indicated equivalent patient improvement in response to either regimen: prevalence of hematuria fell from 75% to 17% after either praziquantel or metrifonate therapy. Similarly, prevalence of proteinuria was significantly reduced from 73% to 29% (metrifonate) or 27% (praziquantel) after therapy. Metrifonate and praziquantel caused similar reductions in bladder granulomata and bladder thickening; however, no reduction in hydronephrosis was noted with either drug. Analysis of outcomes in population subgroups defined by age, sex, pretreatment intensity of infection, or severity of pretreatment morbidity showed no consistent advantage for either drug. In this endemic area, both agents provide effective control of morbidity due to urinary schistosomiasis.

MR imaging of uterine inversion.

Inversion of the uterus is a postpartum complication that is an obstetric emergency. This report describes a case of inversion of the uterus in which clinical evaluation was inconclusive, and magnetic resonance (MR) of the pelvis revealed findings virtually pathognomonic of incomplete inversion of the uterus with far greater conspicuity than on corresponding ultrasound. The MR findings directly resulted in earlier therapeutic intervention with a possible decrease in morbidity.

Neonatal aortic thrombosis treated with intra-arterial urokinase therapy.

We report a newborn infant with neonatal aortic thrombosis (a complication of umbilical artery catheterization) successfully treated by intra-arterial urokinase therapy.

Magnetic resonance imaging of rectal carcinoma.

Five patients with a diagnosis of rectal carcinoma were examined by magnetic resonance and computerized tomography (CT) to determine if magnetic resonance imaging (MRI) added any information to that obtained by computerized tomography. In each case, the imaging studies agreed with the surgical description of the tumor. The studies agreed on the presence of perirectal fat invasion and tumor position. Both modalities were positive for involved lymph nodes in the patient with metastatic disease to pelvic nodes, but both studies underestimated the number of nodes involved. These five cases were examined using two magnetic resonance scanners. The older one operated at 0.3 tesla (T) while the newer scanner operated at 1 T. Spatial resolution was improved with the newer scanner. In addition, the difference in signal intensity between tumor tissue and normal tissue was greater on the 1 T scanner. Tissue differentiation was useful in one case in which CT suggested extension of tumor into the vagina. The extension was confirmed on the MRI scan.

Ultrasound findings in the adreno-genital syndrome (congenital adrenal hyperplasia).

The adreno-genital syndrome (congenital adrenal hyperplasia) is caused by deficiency of an enzyme (usually C-21 hydroxylase) necessary for adrenal production of cortisol, which results in excessive accumulation of androgenic precursors. It is the most common cause of female pseudohermaphroditism. There have been a few sporadic reports of ultrasonic demonstration of enlarged adrenals in the adreno-genital syndrome. To determine whether ultrasonography could be used to establish or exclude the diagnosis, ultrasonic examinations were performed on eight newborn infants with ambiguous genitalia who were subsequently proven to have the adreno-genital syndrome. The adrenals were found to be enlarged in three patients, at the upper limit of normal in three patients, and in two infants the adrenals were well within normal limits in size. The uterus was identified in seven of the eight patients, but was not seen for technical reasons in one. It is concluded that ultrasound is useful in the evaluation of infants with congenital adrenal hyperplasia to establish the presence of a uterus and to demonstrate enlargement of the adrenals, but it should be cautioned that finding adrenals of normal size does not exclude the diagnosis. Key words ultrasound, adreno-genital syndrome, congenital adrenal hyperplasia.

Magnetic resonance imaging of ruptured Baker's cyst.

The clinical presentation of a ruptured Baker's cyst and of thrombophlebitis can be identical. Venography, arthrography, and more recently ultrasonography have been used to differentiate these two entities. We report a patient with a ruptured Baker's cyst who was also evaluated using magnetic resonance imaging (MRI). The soft-tissue abnormalities were better demonstrated by MRI than by the other imaging methods.

Chemotherapy-based control of schistosomiasis haematobia. I. Metrifonate versus praziquantel in control of intensity and prevalence of infection.

To determine the effect of targeted field administration of oral chemotherapeutic agents on the prevalence, intensity, and morbidity of Schistosoma haematobium infections, we initiated a long-term school-based program in the Msambweni area of Kwale District, Coast Province, Kenya. Prior to treatment, 69% of the children examined (ages 4-21, n = 2,628) were infected; 34% had moderate or heavy infections (greater than 100 eggs/10 ml urine). Infected individuals were randomized to receive, during one year, either metrifonate (10 mg/kg x 3 doses) or praziquantel, (40 mg/kg x 1 dose). At the end of the first year, prevalence of infection fell to 19%; only 2% of the pupils remained in the moderately and heavily infected groups. Corresponding decreases in the prevalence of hematuria (54% in 1984 vs. 16% in 1985) and proteinuria (56% in 1984 vs. 26% in 1985) were noted. These were associated with significant declines in bladder thickening and irregularities noted during ultrasound examinations, but not with decreases in hydronephrosis. There was no significant difference in the post-treatment prevalence or intensity of infection after treatment with metrifonate as compared with praziquantel. These results demonstrate that field-applied chemotherapy with either agent offers a practical strategy for the control of S. haematobium infection and its associated morbidity.

Juvenile rheumatoid arthritis: assessment with MR imaging.

Thirty-three joints of the appendicular skeleton in 15 children with juvenile rheumatoid arthritis were examined with magnetic resonance (MR) imaging to determine if it could demonstrate synovial hypertrophy and status of the articular cartilage. Presumed synovial hypertrophy was seen in 13 joints as masses of varying sizes of low to intermediate signal intensity on T1- and T2-weighted images; sometimes foci of increased signal intensity, most likely due to fluid or inflammation, were seen on T2-weighted images. Probable abnormal articular cartilage was detected in ten joints, and MR imaging also demonstrated epiphyseal overgrowth, bone erosions, joint effusions, and joint space narrowing. Because MR imaging appears to provide an objective method of evaluating both synovial hypertrophy and status of articular cartilage, it may prove to be useful in monitoring progression of juvenile rheumatoid arthritis and response to therapy.

Hemophilic arthropathy: assessment with MR imaging.

Fifteen patients with hemophilia, 14 of whom had hemophilic arthropathy, were examined with magnetic resonance (MR) imaging to determine if it could be used to assess hemophilic arthropathy, especially synovial hypertrophy and the status of the articular cartilage. Thirty-five joints of the appendicular skeleton were imaged. Four joints in two patients were clinically normal. Synovial hypertrophy was detected in 28 joints and appeared as areas of low to intermediate signal intensity on T1- and T2-weighted images, with foci of increased signal intensity on T2-weighted images (presumed to be due to areas of fluid or inflammation) in 16 joints. Abnormal articular cartilage was demonstrated in 26 joints; bone lesions, fluid collections, and joint space narrowing could also be seen. MR imaging appears to be useful in depicting the components of hemophilic arthropathy.

Chondromalacia patellae: assessment with MR imaging.

Magnetic resonance (MR) images of the posterior patellar hyaline articular cartilage were obtained in 23 subjects to determine if MR imaging could accurately demonstrate the patellar cartilage. Arthroscopy was used as the standard of reference. Three subjects were asymptomatic volunteers. In the remaining 20 who had patellofemoral pain, arthroscopy was performed before MR imaging in seven and afterward in 12; one did not undergo arthroscopy. MR imaging showed focal areas of swelling of the patellar cartilage, focal hypointensity, surface irregularity, areas of thinning, and areas of cartilage loss with exposure of subchondral bone. The surgical findings agreed with those from MR images in all seven patients who underwent arthroscopy before MR imaging and in ten of the 12 who underwent surgery afterward. MR imaging is an accurate means of examining the posterior patellar cartilage and should be considered as an alternative to diagnostic arthroscopy when chondromalacia patellae is suspected.

MR imaging of osteochondral lesions of talus.

Ten patients with osteochondral lesions of the talus had magnetic resonance (MR) imaging to determine if this modality could accurately characterize the bone lesion and overlying articular cartilage. Eight patients had arthroscopy, five of whom also had arthrotomy. Magnetic resonance detected abnormal cartilage in all eight patients and disruption of cartilage in four of five, proven at surgery. One patient had disrupted cartilage at surgery not diagnosed by MR. Four patients had excision of the osteochondral fragment and curettage of the talar defect and in these patients MR accurately characterized the bony lesion. Three patients had conservative surgical procedures (drilling) and the extent of the bony lesion could not be determined at surgery. Magnetic resonance can accurately characterize the osteocartilaginous components of talus lesions and even though it should not replace plain radiography for diagnosis, it is ideally suited for evaluation of the articular cartilage and has proven to be useful in planning surgical procedures.

CT and MR imaging in staging bladder neoplasms.

Thirteen patients with bladder carcinoma were examined by magnetic resonance (MR) imaging and CT to determine the relative accuracy of the modalities in staging bladder carcinoma. Magnetic resonance correctly identified the presence or absence of extravesical spread of carcinoma in 10 of 13 patients (77%). Two patients were understaged and one was overstaged. Computed tomography correctly staged nine of 13 patients (69%), with three patients understaged and one patient overstaged. Neither MR nor CT was able to determine depth of invasion of the bladder wall in the absence of extravesical spread.

Magnetic resonance imaging of the prostate.

Magnetic resonance imaging was utilized in 18 patients with prostatic cancer and compared with the findings in normal volunteers (Pontes et al., 1985), benign prostatic hyperplasia (Hricak et al., 1983), acute prostatitis (Walsh and Jewett, 1980) and chronic prostatitis (ACS, 1986). Sixteen of the 18 patients with carcinoma demonstrated inhomogeneous signal intensity, however, a similar appearance was also seen in 5 patients with benign prostatic hyperplasia. It does not appear that magnetic resonance imaging is able to reliably differentiate benign from malignant prostatic disease. Extra-prostatic tumor extension and pelvic adenopathy was demonstrated and the technique offers promise for the pre-operative staging of patients with known prostatic carcinomas.