Identification of an alkaline-tolerant cyclodextrin-metabolizing bacterium and characterization of its cyclodextrinase gene

Bacillus circulans A11, an alkaline-tolerant cyclodextrin-metabolizing bacterium isolated from South-East Asian soil, was reidentified as Paenibacillus sp. A11 based on 16S rRNA gene sequence comparison, G + C content and cellular fatty acid composition. Levels of similarity of the 16S rRNA gene between strain A11 and the Paenibacillus species were 90-99%, while similarity with Bacillus circulans was only 86%. The major cellular fatty acid was anteiso-C15:0 which accounted for 59.3% of the total cellular fatty acids and the G+C content was 50.3 mol%. The CDase gene coding for this enzyme was cloned into E. coli. The open reading frame of the CDase gene was 1,959 bp encoding a CDase of 653 amino acid residues. At maximum growth, the specific activity of the recombinant CDase from E. coli was higher than that of Paenibacillus sp. A11. By SDS-PAGE, the translation product of the recombinant gene showed the same mobility as the purified CDase from the original strain. CDase from both Paenibacillus sp. A11 and E. coli produced glucose and maltose as dominant end-products of beta-CD hydrolysis. The ratio of maltose to glucose was 1:2.     

Domain inhibitory and bacteriostatic activities of the five-domain Kazal-type serine proteinase inhibitor from black tiger shrimp Penaeus monodon

Serine proteinase inhibitors (SPIs) in multi-cellular organisms are important modulators of proteinase activities in various biological processes. A five-domain Kazal-type SPI SPIPm2 from the black tiger shrimp Penaeus monodon is presumably involved in innate immune response. The SPIPm2 with the domain P1 residues T, A, E, K and E was isolated from the hemocyte cDNA libraries and found to strongly inhibit subtilisin and elastase, and weakly inhibit trypsin. To unravel further the inhibitory activity of each domain, we subcloned, over-expressed and purified each individual SPI domain. Their inhibitory specificities against trypsin, subtilisin and elastase were determined. Domain 1 was found to be inactive. Domains 2, 3 and 5 inhibited subtilisin. Domain 2 inhibited also elastase. Domain 4 weakly inhibited subtilisin and trypsin. The intact SPIPm2 inhibitor was found to possess bacteriostatic activity against the Bacillus subtilis but not the Bacillus megaterium, Staphylococcus aureus, Vibrio harveyi 639 and Escherichia coli JM109. Domains 2, 4 and 5 contributed to this bacteriostatic activity.     

Genotypic and phenotypic characterization of Lactobacillus plantarum strains isolated from Thai fermented fruits and vegetables

Ten Lactobacillus strains originally isolated from Thai fruits and vegetables fermentation were characterized by various phenotypic and genotypic methods. The phenotypic analysis using the method of carbohydrate fermentation patterns (API50CHL) revealed that the isolates belonged to the L. plantarum species. This was further confirmed by 16S rRNA gene sequencing. Multilocus sequence typing (MLST) revealed a strongly clonal population structure and a low genotypic diversity in this collection. However, the analyzed L. plantarum population demonstrated a higher level of diversification after API50CHL that reflects the role of available carbohydrate sources in bacterial evolution. Our results support the postulate that a combination of conventional biochemical and genotyping methods allows a thorough characterization and identification of isolates. We propose that genotypic characterization could be complemented by biochemical characterization to discriminate L. plantarum strains. (© 2009 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Phase II trial of paclitaxel, carboplatin, and concurrent radiation therapy for locally advanced non-small-cell lung cancer

We conducted a phase II trial to investigate the efficacy of concurrent chemoradiation in patients with stage III non-small-cell lung cancer (NSCLC). Thirty patients with inoperable NSCLC were enrolled onto a multicenter phase II trial of concurrent chemoradiation therapy. Patients received six weekly cycles of paclitaxel 45 mg/m(2) over 1 h; carboplatin at (area under the curve) AUC 2; and radiation therapy of 60 Gy. Radiation was administered to the primary tumor and regional lymph nodes (40 Gy over 4 weeks) followed by a boost to the primary tumor (20 Gy in 2 weeks). After the initial phase of concurrent chemoradiation, patients received an additional four cycles of paclitaxel 175 mg/m(2) over 3 h and carboplatin at AUC 6 every 3 weeks. The overall objective response rate of 30 assessable patients was 76.7%. At the median follow-up time of 13.1 months, the median survival time was 14.5 months (95% CI, 10.59-18.48). The median progression-free survival was 10.5 months (95% CI, 7.72-13.28). The major toxicity was hematologic. The incidence of grade 3 esophagitis was 10%. In conclusion, this chemoradiation regimen is well tolerated and shows significant clinical results for locally advanced NSCLC. Locoregional failure rate remains an important issue with this newer chemotherapeutic regimen. A novel chemotherapy and radiation therapy is clearly needed.     

Flavones with free radical scavenging activity fromGoniothalamus tenuifolius

From the leaves of Goniothalamus tenuifolius, a new natural product namely 3'-hydroxy-3,5,7,4'-tetramethoxyflavone (1) was isolated, along with seven other known compounds (2-8). Each of these isolates was evaluated for free radical scavenging activity on the DPPH decoloration test. The data obtained in this study suggested that the ortho 3',4'-diphenolic structure was essential for the activity of these flavonol derivatives.     

A five-domain Kazal-type serine proteinase inhibitor from black tiger shrimp Penaeus monodon and its inhibitory activities

A novel five-domain Kazal-type serine proteinase inhibitor, SPIPm2, identified from the hemocyte cDNA library of black tiger shrimp Penaeus monodon was successfully expressed in the Escherichia coli expression system. The expressed recombinant SPIPm2 (rSPIPm2) as inclusion bodies was solubilized with a sodium carbonate buffer, pH10, and purified by gel filtration chromatography. The molecular mass of rSPIPm2 was determined using MALDI-TOF mass spectrometry to be 29.065 kDa. The inhibitory activities of rSPIPm2 were tested against trypsin, alpha-chymotrypsin, subtilisin and elastase. The inhibitor exhibited potent inhibitory activities against subtilisin and elastase, weak inhibitory activity against trypsin, and did not inhibit chymotrypsin. Tight-binding inhibition assay suggested that the molar ratios of SPIPm2 to subtilisin and elastase were 1:2 and 1:1, respectively. The inhibition against subtilisin and elastase was a competitive type with inhibition constants (Ki) of 0.52 and 3.27 nM, respectively. The inhibitory activity of SPIPm2 against subtilisin implies that, in shrimp, it may function as a defense component against proteinases from pathogenic bacteria but the elastase inhibitory function is not known.     

Case report of metastatic familial pheochromocytoma treated with cisplatin and 5-fluorouracil

Summary We report on a rare case of malignant pheochromocytoma in a patient with a family history of this disease. After three cycles of treatment with cisplatin and 5-fluorouracil, a decrease in the need for antihypertensive treatment occurred, which lasted almost 2 years despite the discontinuation of chemotherapy. The patient showed an objective response, which was technically a minor response, although in this slow-growing tumor it was of major clinical significance. This chemotherapy regimen may play a role in the management of malignant pheochromocytoma.