Haemophilus influenzae HP1 Bacteriophage Encodes a Lytic Cassette with a Pinholin and a Signal-Arrest-Release Endolysin.

HP1 is a temperate bacteriophage, belonging to the Myoviridae family and infecting Haemophilus influenzae Rd. By in silico analysis and molecular cloning, we characterized lys and hol gene products, present in the previously proposed lytic module of HP1 phage. The amino acid sequence of the lys gene product revealed the presence of signal-arrest-release (SAR) and muraminidase domains, characteristic for some endolysins. HP1 endolysin was able to induce lysis on its own when cloned and expressed in Escherichia coli, but the new phage release from infected H. influenzae cells was suppressed by inhibition of the secretion (sec) pathway. Protein encoded by hol gene is a transmembrane protein, with unusual C-out and N-in topology, when overexpressed/activated. Its overexpression in E. coli did not allow the formation of large pores (lack of leakage of ß-galactosidase), but caused cell death (decrease in viable cell count) without lysis (turbidity remained constant). These data suggest that lys gene encodes a SAR-endolysin and that the hol gene product is a pinholin. HP1 SAR-endolysin is responsible for cell lysis and HP1 pinholin seems to regulate the cell lysis and the phage progeny release from H. influenzae cells, as new phage release from the natural host was inhibited by deletion of the hol gene.

Characterization and Analysis of 2-(2-Phenylethyl)-chromone Derivatives from Agarwood (Aquilaria crassna) by Artificial Holing for Different Times.

A total of fifty-six chromones, including seven 5,6,7,8-tetrahydro-2-(2-phenylethyl)-chromones (THPECs), five 5,6-epoxy-2-(2-phenylethyl)chromones (EPECs), seven 5,6:7,8-diepoxy-2-(2-phenylethyl)chromones (DEPECs) and thirty-seven 2-(2-phenylethyl)chromones of the flidersia type (FTPECs), were characterized by HPLC/DAD/ESI/MS/MS in three agarwood samples (from Aquilaria crassna) induced by artificial holing with different holing times. The characteristic fragmentation behavior of DEPECs and EPECs, and the methods to distinguish these four types of chromones by MS analysis were described for the first time. In addition, it was found that the relative contents of DEPECs and EPECs were down-regulated, while the relative contents of THPECs and FTPECs were up-regulated for the samples from two, four and five years of the agarwood formation time. However, the relative contents of six most widespread and abundant FTPECs presented roughly upward based on the formation time. These results could be referenced to distinguish different agarwood samples collected from different formation time.

Sesquiterpenoids from Chinese Agarwood Induced by Artificial Holing.

Two new sesquiterpenoids, 3-oxo-7-hydroxylholosericin A (1) and 1,5;8,12-diepoxy-guaia-12-one (2), together with seven known sesquiterpenoids 3-9, were isolated from Chinese agarwood induced by artificial holing originating from Aquilaria sinensis (Lour.) Gilg. Their structures were identified by spectroscopic techniques (UV, IR, 1D and 2D NMR) and MS analyses. The absolute configuration of compound 1 was determined by comparison of its measured CD curve with that of calculated data for 1 and ent-1. The NMR data of 3 were reported in this study for the first time. Compounds 1, 2, 4-6, together with the EtOAc extract showed moderate inhibitory activities against acetylcholinesterase, and compounds 4-6, 8 exhibited antibacterial activities against Staphylococcus aureus or Ralstonia solanacearum.

Quantitative evaluation method of smoke exhaust performance and application on exhaust volume optimization in tunnel fires under lateral centralized mode.

This study investigated the indicators in the quantitative method of evaluating the smoke exhaust performance, which provided a theoretical basis for the optimization of the smoke extraction system under the lateral centralized mode in tunnel fires. The criterion was proposed for plug-holing, and the theoretical models of smoke exhaust efficiency were established to distinguish whether the plug-holing occurs or not. The relationship between efficiency, effectiveness, and efficacy was analyzed from the perspective of smoke and heat exhaust. Meanwhile, this evaluation method was applied to the optimization of exhaust volume in a practical engineering through FDS numerical simulation. The results show that Ri is a vital basis for reflecting the movement form of smoke and the exhaust effect. The critical value of Ri is 1.09 when plug-holing occurs in a standard three-lane immersed tunnel, resulting in a significant reduction in the efficiency of smoke exhaust. The greater the exhaust volume of the exhaust fan, the greater the Ri value, the higher the total smoke and heat exhaust efficiency, and the better the exhaust effectiveness of smoke inlets without plug-holing. Under longitudinal ventilation, the optimal exhaust volume is 180 m3/s at 20 MW and 360 m3/s at 50 MW in the application case.

Characterization of a Novel Bacteriophage swi2 Harboring Two Lysins Can Naturally Lyse Escherichia coli.

The novel virulent Siphoviridae bacteriophage swi2 was isolated from a pig farm, and its biological characteristics, genome architecture, and infection-related properties were characterized. Phage swi2 has a high titer of 1.01 × 1012 PFU/mL with good tolerance to UV rays and remains stable in the pH range of 6-10 and at temperatures less than 50°C. One-step growth analysis revealed that phage swi2 had a 25 min latent period with a large burst size (1,000 PFU/cell). The biological characteristics indicated that swi2 had good host infectivity and effective lytic activities. The genome of phage swi2 is composed of 47,611 bp with a G + C content of 46.50%. Eighty-nine orfs were predicted, and only 18 of them have known functions. No virulence genes or drug resistance genes were found in the genome. Genome sequence comparison of phage swi2 showed that there were a total of 10 homologous phages in the database with low similarity (less than 92.51% nucleotide identity and 66% query coverage). The predicted host lysis-related genes of phage swi2 consist of one holin, two endolysins, and Rz/Rz1 equivalents. Antibacterial activity assays showed that both endolysins could naturally reduce the host Escherichia coli 51 titers by -1 log unit both in vitro and in vivo, EDTA showed no obvious synergistic action, and holin had no lytic effects on the host cell. These results provide necessary information for the development of antibiotic alternatives for the treatment of multidrug-resistant Escherichia coli infection.

Five new epoxy-5,6,7,8-tetrahydro-2-(2-phenylethyl)chromones from Chinese agarwood by artificial holing.

Five optically active epoxy-5,6,7,8-tetrahydro-2-(2-phenylethyl)chromone derivatives (1-5) with four chirality centers in the condensed cyclohexene ring were isolated from artificial holing agarwood originating from Aquilaria sinensis (Lour.) Gilg. Their structures were elucidated by spectroscopic techniques (UV, IR, 1D and 2D NMR) and MS analyses. The absolute configuration of 2 was elucidated by TDDFT-ECD calculations and ECD spectra of 1, 3-5 allowed their configurational assignment as well. All of the new compounds contained a condensed oxirane ring, and compound 1 was the only 6,7-epoxy-2-(2-phenylethyl)-5,6,7,8-tetrahydrochromone derivative that has ever been found in agarwood. AChE inhibitory activity was tested for the first time of these new compounds by using modified Ellman's colorimetric method. Compounds 3 and 5 exhibited inhibitory activity against AChE with IC50 value of 441.6, and 155.6 µM, respectively.

Five new eudesmane-type sesquiterpenoids from Chinese agarwood induced by artificial holing.

Five new eudesmane-type sesquiterpenoids (1-5), along with six known ones (6-11), were isolated from Chinese agarwood induced by artificial holing originating from Aquilaria sinensis (Lour.) Gilg (Thymelaeaceae). The structures of the new sesquiterpenoids were established by spectroscopic methods including UV, IR, MS, 1D, and 2D NMR. Compounds 1, 3, 6 and 7 exhibited antibacterial activities against both Staphylococcus aureus and Ralstonia solanacearum, and compound 5 only showed an inhibitory activity towards S. aureus. Compounds 1, 6, 7 and 10 showed weak acetylcholinesterase inhibitory activity.

2-(2-phenylethyl)chromone derivatives from Chinese agarwood induced by artificial holing.

Three new 2-(2-phenylethyl)chromone derivatives (1-3), together with thirteen known ones (4-16), were isolated from the EtOAc extract of Chinese agarwood induced by artificial holing, originating from Aquilaria sinensis (Lour.) Gilg (Thymelaeaceae). The chemical structures of the new compounds were identified by spectroscopic techniques (UV, IR, MS, 1D and 2D NMR). Compounds 1, 6, 15 and 16 exhibited inhibitory effects on Staphylococcus aureus, and compounds 15 and 16 showed inhibitory effects on Ralstonia solanacearum. Compounds 1-3, 7, 9, 11, 12, 15 and 16 exhibited acetylcholinesterase inhibitory activity. A possible biogenetic pathway of compounds 1-16 was proposed to show the relationships between diepoxy-tetrahydro-2-(2-phenylethyl)chromones, epoxy-tetrahydro-2-(2-phenylethyl) chromones, tetrahydro-2-(2-phenylethyl)chromones, and 2-(2-phenylethyl)chromones of the flidersia type, the four main types of 2-(2-phenylethyl)chromones found in agarwood, on the basis of their appearances in different stage of agarwood formation.

Perfusing chemotherapy by percutaneous lung puncture "holing" for pulmonary tuberculoma-a ten-year single center experience.

BACKGROUND: Pulmonary tuberculoma is a special form of secondary pulmonary tuberculosis, with a poor response to drug treatment. We used the method of drug administration via percutaneous lung puncture "holing" to treat pulmonary tuberculoma and observe its short- and long-term efficacy, summing up our 10-year clinical experience. METHODS: A total of 54 patients with pulmonary tuberculoma were included in this study. They themselves were taken as the control group. Three to six months of conventional anti-tuberculosis treatment was conducted firstly. Then those patients with no changes of sizes in tuberculoma were recommended to receive drug administration via percutaneous lung puncture. Isoniazid (INH, 0.1 g) and amikacin (AMK, 0.2 g) were injected into tuberculoma (once or twice per week, 10 times as a course of treatment). RESULTS: After two months of drug treatment by lung puncture, the sputum smear test showed the negative conversion rate of tubercle bacillus was 87% (13/15), and the positive conversion rate was 8% (3/39). The tuberculosis bacillus culture indicated that the negative conversion rate was 100% (7/7). The reexamination after one year showed the negative conversion rate of tubercle bacillus in the sputum smear test was 80% (4/5). About 58% (31/54) of tuberculoma disappeared or significantly reduced, in which, 40% (21/54) of tuberculoma disappeared. The tuberculoma diameter reduced from 3.6 cm × 2.8 cm to 1.7 cm × 1.1 cm on average. Side-effects included postoperative pneumothorax 9% (5/54), hemoptysis 7% (4/54) and fever 11% (6/54). A total of 34 patients were followed up for five years, and the disappearance rate of tuberculoma was up to 47% (16/34), with no recurrence. CONCLUSIONS: The drug administration via percutaneous lung puncture-"holing" in pulmonary tuberculoma takes a significant effect obviously, good short- and long-term effects and less side effects.