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)     

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.     

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.     

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.     

Safety and Efficacy of Everolimus With Exemestane vs. Exemestane Alone in Elderly Patients With HER2-Negative,Hormone Receptor–Positive Breast Cancer in BOLERO-2

BackgroundPostmenopausal women with hormone receptor–positive (HR⁺) breast cancer in whom disease progresses or there is recurrence while taking a nonsteroidal aromatase inhibitor (NSAI) are usually treated with exemestane (EXE), but no single standard of care exists in this setting. The BOLERO-2 trial demonstrated that adding everolimus (EVE) to EXE improved progression-free survival (PFS) while maintaining quality of life when compared with EXE alone. Because many women with HR⁺ advanced breast cancer are elderly, the tolerability profile of EVE plus EXE in this population is of interest.Patients and MethodsBOLERO-2, a phase III randomized trial, compared EVE (10 mg/d) and placebo (PBO), both plus EXE (25 mg/d), in 724 postmenopausal women with HR⁺ advanced breast cancer recurring/progressing after treatment with NSAIs. Safety and efficacy data in elderly patients are reported at 18-month median follow-up.ResultsBaseline disease characteristics and treatment histories among the elderly subsets (≥ 65 years, n = 275; ≥ 70 years, n = 164) were generally comparable with younger patients. The addition of EVE to EXE improved PFS regardless of age (hazard ratio, 0.59 [≥ 65 years] and 0.45 [≥ 70 years]). Adverse events (AEs) of special interest (all grades) that occurred more frequently with EVE than with PBO included stomatitis, infections, rash, pneumonitis, and hyperglycemia. Elderly EVE-treated patients had similar incidences of these AEs as did younger patients but had more on-treatment deaths.ConclusionAdding EVE to EXE offers substantially improved PFS over EXE and was generally well tolerated in elderly patients with HR⁺ advanced breast cancer. Careful monitoring and appropriate dose reductions or interruptions for AE management are recommended during treatment with EVE in this patient population.     

Localization of anti-lipopolysaccharide factor (ALFPm3) in tissues of the black tiger shrimp, Penaeus monodon, and characterization of its binding properties

Anti-lipopolysaccharide factor (ALF) is an antimicrobial peptide originally identified from horseshoe crabs and recently found in several shrimp species. ALFPm3, the most abundant isoform in the black tiger shrimp, Penaeus monodon, has been shown to possess a broad spectrum of antimicrobial activity against Gram-negative and Gram-positive bacteria, and filamentous fungi. In this study, a potential role for ALFPm3 in the shrimp innate immunity was revealed by examining the distribution of the protein in shrimp tissues in response to Vibrio harveyi challenge. Immunohistochemistry using anti-ALFPm3 antibody showed that the ALFPm3 protein is primarily localized in hemocytes and the positive cells observed at the injection site and in the cephalothorax are infiltrating hemocytes that migrate into shrimp tissues after bacterial injection. A rapid increase in the number of hemocytes producing ALFPm3 observed in V. harveyi-injected shrimp suggests a likely important function of the protein in defense against invading pathogens. ALFPm3 was shown to be able to bind to Gram-negative and Gram-positive bacterial cells and their major cell wall components, lipopolysaccharide (LPS) and lipoteichoic acid (LTA), respectively. The results suggested that ALFPm3 performs its antibacterial activity by binding to component(s) of the bacterial cell wall.