Bacteria on living or inert surfaces usually form biofilms which make them highly resistant to antibiotics and immune clearance. Herein, we develop a simple approach to overcome the above conundrum through lysozyme-associated liposomal gentamicin (LLG). The association of lysozyme to the surface of liposomes can effectively reduce the fusion of liposomes and undesirable payload release in regular storage or physiological environments. The LLG was more effective at damaging established biofilms and inhibiting biofilm formation of pathogens including Gram-positive and Gram-negative bacteria than gentamicin alone. This strategy may provide a novel approach to treat infections due to bacterial biofilm.
Environmental iron concentrations coordinately regulate transcription of genes involved in iron acquisition and virulence via the ferric uptake regulation (fur) system. We identified and sequenced the fur gene and flanking regions of three Bartonella species. The most notable difference between Bartonella Fur and other Fur proteins was a substantially higher predicted isoelectric point. No promoter activity or Fur autoregulation was detected using a gfp reporter gene fused to the 204 nucleotides immediately upstream of the Bartonella fur gene. Bartonella henselae fur gene expression complemented a Vibrio cholerae fur mutant.Bartonella, an extremely fastidious gram-negative bacillus, causes cat scratch disease, bacillary angiomatosis, and other syndromes (2,13,15,29). Few data exist regarding the pathogenic mechanisms of this hemophilic bacterium (5), which can occupy two alternate niches: the iron-rich gut of obligately hematophagous arthropods and the iron-restricted bloodstream of mammals (11,32). Acquisition of iron and expression of many virulence factors are under transcriptional regulation by the fur gene product, the ferric uptake regulation (Fur) protein, and its homodimeric complex (7). At sufficient intracellular iron levels, the corepressors Fur and Fe 2ϩ form an active Fur-Fe 2ϩ complex that binds a consensus sequence ("iron box," a 19-bp hyphenated dyad repeat [22] or three repeats of 6 bp of the sequence NAT[A,T]AT [7]) in the promoter region of genes regulated by Fur, down-regulating genes encoding iron-scavenging proteins (7,22). We hypothesized that Bartonella species possess a fur gene homolog with a gene regulatory system influenced by iron levels.Bacterial strains. Strains and plasmids used in this study are listed in Table 1. All Bartonella strains were used at low passage numbers (passes 1 through 3). B. henselae and B. quintana strains were grown on fresh chocolate agar (14), which provided a replete iron source. B. bacilliformis was grown on fresh heart infusion agar supplemented with 5% defibrinated rabbit blood (Hemostat Labs, Dixon, Calif.). Plates were incubated at 34°C (B. henselae and B. quintana) or 29°C (B. bacilliformis) in an enriched CO 2 environment for 5 to 7 days. Iron availability to B. henselae and B. quintana was restricted by adding the ferric-specific chelating agent EDDHA (ethylene diamine dihydroxy-o-phenylacetic acid) (The Complete Green Company, El Segundo, Calif.) (25) or by decreasing the hemoglobin (Hb) concentration in agar.Vibrio strains were grown overnight in Luria-Bertani medium with the appropriate selective antibiotic(s), with or without the iron chelator 2,2-dipyridyl (Sigma-Aldrich, Inc., St. Louis, Mo.), as previously described (8). Because of limited growth at 37°C by CML13(pSYP4), all strains were grown at 30°C and then incubated at 37°C for 60 to 90 min before assays. Selective antibiotics were added to growth media as required, at the following concentrations: ampicillin, 100 g/ml; chloramphenicol, 25 g/ml; kanamycin, 50 g/ml; streptomycin, 100 g/ml...
The mixture of glucose and β-disaccharide (MGD) synthesized by transglycosylation of glucose as a low-cost soluble carbon source can efficiently induce cellulase production in Trichoderma reesei, which holds potential for the biorefining of lignocellulosic biomass. However, it is not yet fully understood how MGD induces T. reesei cellulase. In this study, transcriptomic analyses were conducted to investigate the molecular basis of MGD for lignocellulose-degrading enzyme production of T. reesei Rut C30 compared with that on lactose. Particular attention was paid to CAZymes, transcription factors, transporters and other protein processing pathways related to lignocellulose degradation. As a result, MGD can elicit transcription of GH5-, GH6- and GH7-encoding cellulases that is up to 1.4-fold higher than that induced by lactose, but GH11- and GH74-encoding xylanases are downregulated by 1.7- and 4.4-fold, respectively. Gene expression profiles suggest that the transcription activators xyr1 and vib1 are significantly upregulated and that the mitogen-activated protein kinase pathway is strengthened compared to the case of lactose induction. In addition, hac1-encoding UPR-specific transcription factors are significantly upregulated by MGD, which may be enhanced due to proper folding and processing of nascent proteins. These findings provide a theoretical basis for further understanding the characterization of efficient cellulase production using MGD as an inducer in T. reesei and offer potential strategies for strain improvement.
Sucrose, as the main transporter of photosynthate, is transported by sucrose transporters. In this study, we cloned Ib-SUT3 gene from sweet potato (Ipomoea batatas Lam. GenBank accession number MN233361), by RACE technique. The full-length sequence of the gene is 1607 bp and the complete open reading frame is 1518 bp, encoding 505 amino acids. The predicted protein has the molecular weight of 53.82 kD and isoelectric point of 9.19, containing 12 transmembrane domains. Multiple sequence alignment analysis indicated the protein, belonging to Group I, was significantly different from Group IV in evolution and shared high similarities with SUTs from other plant species. IbSUT3 had the function of transporting sucrose verified by SUSY7/ura3 system. Subcellular localization showed that IbSUT3 protein was located in the tobacco protoplasts plasma membrane. The qRT-PCR results showed that IbSUT3 highly expressed in leaves and induced by drought, high salt, low temperature and exogenous abscisic acid, suggesting that IbSUT3 is involved in response to various abiotic stresses, including abscisic acid.
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