The emergence of acquired high-level resistance among Enterococcus species has renewed interest in mechanisms of resistance to glycopeptide antibiotics in gram-positive bacteria. In Enterococcus faecalis and Enterococcus faecium, resistance is encoded by the van gene cluster and is due to the production of a peptidoglycan precursor terminating in D-alanyl-D-lactate, to which vancomycin does not bind. MostLeuconostoc and many Lactobacillus species are intrinsically resistant to high levels of glycopeptide antibiotics, but the mechanism of resistance has not been elucidated. To determine whether the mechanisms of resistance are similar in intrinsically resistant bacteria, cytoplasmic pepti4oglycan precursors were isolated from Leuconostoc mesenteroides and Lactobacilus casei and analyzed by mass spectrometry, revealing structuresGlycopeptide antibiotics such as vancomycin act by binding to the D-alanyl-D-alanine terminus of stem pentapeptides present in bacterial peptidoglycan (16,21). It had been widely assumed that this terminus was ubiquitous among eubacterial species producing peptidoglycan and that vancomycin resistance was therefore unlikely to emerge in the absence of a barrier to vancomycin binding such as the outer membrane of gram-negative bacteria. However, high-level transmissible resistance to glycopeptides has recently been recognized among several gram-positive species, particularly enterococci, in which the incidence of resistance has been rising among clinical isolates (2).Most Leuconostoc and Pediococcus and some Lactobacillus species have been described as being intrinsically resistant to glycopeptides (9,15,17,20), but the mechanism of resistance has not been systematically investigated, perhaps because these organisms were considered to be of little clinical significance. However, the widespread use of vancomycin has led to more frequent recognition of these species as opportunistic pathogens (9,20 depsipeptide (D-alanyl-D-hydroxy acid) which is added in place of D-alanyl-D-alanine to the UDP-muramyl-tripeptide precursor, resulting in a peptidoglycan precursor that is not bound by vancomycin (6). This hypothesis was supported by the observation that the MIC for resistant enterococci was diminished when tested with high concentrations of D-amino acids that can be incorporated into pentapeptide and that are bound by vancomycin to a greater extent than D-2-hydroxy acids (23). Subsequently, the cytoplasmic peptidoglycan precursor in resistant E. faecium and E. faecalis was identified as UDP-

With the aim to discuss the similarities and differences of phytochemicals in Moringa oleifera leaves collected from China (CML) and India (IML) in mind, comparative ultra-performance liquid chromatography coupled with quadrupole-time-of-flight mass spectrometry (UPLC-QTOF-MS) analysis was performed in this study. A screening analysis based on a UNIFI platform was first carried out to discuss the similarities. Next, untargeted metabolomic analysis based on multivariate statistical analysis was performed to discover the differences. As a result, a total of 122 components, containing 118 shared constituents, were characterized from CML and IML. The structure types included flavonoids, alkaloids, glyosides, organic acids and organic acid esters, iridoids, lignans, and steroids, etc. For CML, 121 compounds were characterized; among these, 18 potential biomarkers with higher contents enabled differentiation from IML. For IML, 119 compounds were characterized; among these, 12 potential biomarkers with higher contents enabled differentiation from CML. It could be concluded that both CML and IML are rich in phytochemicals and that CML is similar to IML in the kinds of the compounds it contains, except for the significant differences in the contents of some compounds. This comprehensive phytochemical profile study provides a basis for explaining the effect of different growth environments on secondary metabolites and exists as a reference for further research into or applications of CML in China.

Cardioplegic reperfusion during a long term ischemic period interrupts cardiac surgery and also increase cellular edema due to repeated administration. We reviewed the clinical experiences on myocardial protection of one single perfusion with histidine-ketoglutarate-tryptophan (HTK) for infants. This retrospective study included 118 infants who underwent open-heart surgery between January 2004 and December 2007. We divided the entire cohort into two groups: In group H (n = 63), myocardial protection was carried out with one single perfusion with HTK solution, and in group S (n = 55) with conventional St. Thomas crystalloid cardioplegia. The duration of cardiopulmonary bypass (CPB) did not differ between these two groups, but the duration of aortic cross-clamping time in group H was significantly shorter than that in group S (p < 0.05). During reperfusion, the spontaneous re-beating rate was higher in group H (p < 0.05). There were no differences in doses of inotropic agent and creatinekinase (CK) values on postoperative day 1 between these two groups, but the level of CK in group H was significantly less than that in group S on postoperative day 2 (p < 0.01). The mortality in group H was lower than in group S (p < 0.05). The HTK group had shorter cross-clamping time and more frequent spontaneous defibrillation than St. Thomas group. We propose that HTK is valid for some complicated cardiac surgeries with long term cross-clamping time.

BackgroundCassava (Manihot esculenta Crantz) is a major crop extensively cultivated in the tropics as both an important source of calories and a promising source for biofuel production. Although stable gene expression have been used for transgenic breeding and gene function study, a quick, easy and large-scale transformation platform has been in urgent need for gene functional characterization, especially after the cassava full genome was sequenced.MethodsFully expanded leaves from in vitro plantlets of Manihot esculenta were used to optimize the concentrations of cellulase R-10 and macerozyme R-10 for obtaining protoplasts with the highest yield and viability. Then, the optimum conditions (PEG4000 concentration and transfection time) were determined for cassava protoplast transient gene expression. In addition, the reliability of the established protocol was confirmed for subcellular protein localization.ResultsIn this work we optimized the main influencing factors and developed an efficient mesophyll protoplast isolation and PEG-mediated transient gene expression in cassava. The suitable enzyme digestion system was established with the combination of 1.6% cellulase R-10 and 0.8% macerozyme R-10 for 16 h of digestion in the dark at 25 °C, resulting in the high yield (4.4 × 107 protoplasts/g FW) and vitality (92.6%) of mesophyll protoplasts. The maximum transfection efficiency (70.8%) was obtained with the incubation of the protoplasts/vector DNA mixture with 25% PEG4000 for 10 min. We validated the applicability of the system for studying the subcellular localization of MeSTP7 (an H+/monosaccharide cotransporter) with our transient expression protocol and a heterologous Arabidopsis transient gene expression system.ConclusionWe optimized the main influencing factors and developed an efficient mesophyll protoplast isolation and transient gene expression in cassava, which will facilitate large-scale characterization of genes and pathways in cassava.