Synthesis of benzoylthiourea derivatives and analysis of their antibacterial performance against planktonic Staphylococcus aureus and its biofilms

Following the appearance of several antimicrobial agents to control the spread of infections, two major challenges have emerged: (i) the occurrence and blowout of multiresistant bacteria and the increase of chronic diseases and (ii) difficult-to-eradicate infections. In this study, we tested five benzoylthiourea derivatives for their ability to inhibit and stop bacterial growth and evaluated the possible influence of 1,2,4-triazolyl-benzoylthiourea derivative 4 on the formation and eradication of Staphylococcus aureus biofilms. Benzoylthiourea derivatives 4 , 6 , 10 , 11 and 13 were obtained in one or two steps with low cost and subjected to tests to identify their minimum inhibitory concentration (MIC) and minimum bactericidal concentration. In vitro tests were also performed to assess their effects on biofilm formation and in preformed biofilms and scanning electron microscopy was used to visualize the effects on biofilm formation. The 1,2,4-triazolyl-benzoylthiourea derivative 4 showed bacteriostatic activity against the S. aureus HU25 clinical strain with an MIC of 16 µg ml⁻¹, which is below the toxic concentration (at 2500 µg ml⁻¹, 62·25% of the cells remained viable). Compound 4 also effectively prevented biofilm formation at the three subinhibitory concentrations tested (1/2 MIC, 1/4 MIC and 1/8 MIC) as confirmed by scanning electron microscopy. For breakdown of formed biofilms, the main influence was at a subinhibitory concentration (1/2 MIC). These findings make compound 4 a strong candidate for studies on the development of new antimicrobial and antibiofilm agents.     

Effects of water gradients on soil enzyme activity and active organic carbon composition under Carex lasiocarpa marsh

The responses of soil enzyme activity of freshwater marsh, microbial biomass carbon (MBC), dissolved organic carbon (DOC) and aboveground biomass to water gradients were studied with Carex lasiocarpa pot culture experiment. The relationships between soil enzyme activity and MBC, DOC and aboveground biomass were discussed. The water gradients were W1, 15 cm; W2, ?5 cm; W3, ?5–5 cm; W4, submerged. The results indicated that acid phosphatase, invertase and urease activities were decreased with the increase of water level, while catalase activity was increased with moisture content increasing. Drying-wetting alternation (W3) increased soil enzyme activities if compared with W1. MBC content followed the order of W3 > W1 > W2 > W4, and the activities of invertase, urease and catalase were significantly positively correlated with MBC (p < 0.05). DOC content presented the order of W4 > W1 > W3 > W2, and the activities of urease and acid phosphatase were most significantly negatively correlated with DOC (p < 0.01). In addition, drying-wetting alternation promoted the growth of Carex lasiocarpa. When water submerged plants, the growth of Carex lasiocarpa was significantly inhibited. The aboveground biomass was positively related to soil enzyme activities. There were close relationships between the activities of invertase, urease and catalase and the growth situation of Carex lasiocarpa.     

Enhanced litter input rather than changes in litter chemistry drive soil carbon and nitrogen cycles under elevated CO2: a microcosm study

Elevated CO₂ has been shown to stimulate plant productivity and change litter chemistry. These changes in substrate availability may then alter soil microbial processes and possibly lead to feedback effects on N availability. However, the strength of this feedback, and even its direction, remains unknown. Further, uncertainty remains whether sustained increases in net primary productivity will lead to increased long‐term C storage in soil. To examine how changes in litter chemistry and productivity under elevated CO₂ influence microbial activity and soil C formation, we conducted a 230‐day microcosm incubation with five levels of litter addition rate that represented 0, 0.5, 1.0, 1.4 and 1.8 × litterfall rates observed in the field for aspen stand growing under control treatments at the Aspen FACE experiment in Rhinelander, WI, USA. Litter and soil samples were collected from the corresponding field control and elevated CO₂ treatment after trees were exposed to elevated CO₂ (560 ppm) for 7 years. We found that small decreases in litter [N] under elevated CO₂ had minor effects on microbial biomass carbon, microbial biomass nitrogen and dissolved inorganic nitrogen. Increasing litter addition rates resulted in linear increase in total C and new C (C from added litter) that accumulated in whole soil as well as in the high density soil fraction (HDF), despite higher cumulative C loss by respiration. Total N retained in whole soil and in HDF also increased with litter addition rate as did accumulation of new C per unit of accumulated N. Based on our microcosm comparisons and regression models, we expected that enhanced C inputs rather than changes in litter chemistry would be the dominant factor controlling soil C levels and turnover at the current level of litter production rate (230 g C m⁻² yr⁻¹ under ambient CO₂). However, our analysis also suggests that the effects of changes in biochemistry caused by elevated CO₂ could become significant at a higher level of litter production rate, with a trend of decreasing total C in HDF, new C in whole soil, as well as total N in whole soil and HDF.     

四川宜宾七种中药提取物体外抗菌活性研究

用肉汤二倍稀释法和琼脂平板培养计数法,研究四川宜宾七种中药的提取物对大肠杆菌、沙门氏菌和巴氏杆菌的体外抑制作用.结果表明：黄柏提取物的抗菌活性最强,最小抑菌浓度(Minimum inhibitory concentration,MIC)为15．6~125 mg·mL-1,最小杀菌浓度(Minimum bactericidal concentration,MBC)为31．25~250 mg·mL-1;栀子提取物抗菌活性次之,MIC 为62．5~125 mg·mL-1,MBC 为125~＞250 mg·mL-1;佛手提取物有较稳定的抗菌活性,MIC均为125 mg·mL-1,MBC为125 mg·mL-1和250 mg·mL-1;姜黄、杜仲、何首乌、细毡毛忍冬的提取物抗菌活性相对较差,大部分 MIC≥250 mg·mL-1.说明四川宜宾的黄柏和栀子的提取物抗菌活性较强,佛手提取物抗菌活性较稳定,有进一步研究的价值.     

The Podosphaera fusca TUB2 gene,a molecular “Swiss Army knife” with multiple applications in powdery mildew research

The powdery mildew fungus Podosphaera fusca (synonym Podosphaera xanthii) is the main causal agent of cucurbit powdery mildew and one of the most important limiting factors for cucurbit production worldwide. Despite the fungus' economic importance, very little is known about the physiological and molecular processes involved in P. fusca biology and pathogenesis. In this study, we isolated and characterised the β-tubulin-encoding gene of P. fusca (PfTUB2) to develop molecular tools with different applications in powdery mildew research. PfTUB2 is predicted to encode a protein of 447 amino acid residues. The coding region is interrupted by six introns that occur at approximately the same positions as the introns present in other fungal TUB2-like genes. Once cloned, the PfTUB2 sequence information was used in different applications. Our results showed that the TUB2 gene is a good marker for molecular phylogenetics in powdery mildew fungi but it is unsuitable for the analysis of intraspecific diversity in P. fusca. The expression of PfTUB2 was proven to be stable in different temperature conditions, supporting its use as a reference gene in quantitative gene expression studies. Furthermore, an allele-specific PCR assay for the detection of resistance to methyl-2-benzimidazole carbamate (MBC) fungicides in P. fusca was developed based on the correlation between the single amino acid change E198A in β-tubulin and the MBC resistance phenotype. Lastly, PfTUB2 was used as a target gene in the development of a high-throughput method to quantify fungal growth in plant tissues.     

10种中草药提取物体外抗铜绿假单胞菌作用研究

通过对粗糠柴等10种中草药采用80%乙醇室温下浸渍制备的提取物进行体外抗铜绿假单胞菌及其耐药菌活性研究,并采取药敏纸片法测定临床分离菌株的耐药性。结果表明:这10种中草药80%乙醇提取物中,粗糠柴的乙酸乙酯层对铜绿假单胞菌标准菌及其耐药菌的抑菌效果最好,其抑菌圈直径范围在10~17 mm之间,MIC范围在0.125~0.5 mg·mL~(-1)之间,MBC范围在0.5~1 mg·mL~(-1)之间;正丁醇层、水层的抑菌活性较乙酸乙酯层弱,石油醚层对铜绿假单胞菌没有效果。而小叶藤黄、滇南红厚壳、续随子的乙酸乙酯层,巴豆、罗汉松、肉桂醇提物对铜绿假单胞菌及其耐药菌株有较弱抗菌活性;滇南红厚壳的正丁醇层、续随子乙酸乙酯层以及大八角和郁金的醇提物对铜绿假单胞菌及其耐药菌株均无活性。从这些数据中可以得出,粗糠柴的乙酸乙酯层、正丁醇层和水层对铜绿假单胞菌及其耐药菌有较好的抑菌活性,尤以乙酸乙酯层活性最好,而粗糠柴的石油醚层没有活性。     

Isolation of oxidative stress response mutants in Escherichia coli for their use as a genetic screen to identify new anti-mycobacterials as functional analogues of isoniazid

Summary Tuberculosis is a leading killer disease of the world with increasing mortality due to HIV-infected individuals becoming highly prone to this infection. An attempt has been made in the present work to identify novel plant-derived compounds active against Mycobacterium tuberculosis (MTB) through construction of a target based bio-screen to facilitate rapid screening of anti-TB plant compounds. To achieve this, construction of a genetically modified model system was attempted in fast growing, non-pathogenic, Escherichia coli in which experimental testing is relatively easier and rapid as compared to M. tuberculosis, which is pathogenic and slow growing in nature. The exquisitely high sensitivity of M. tuberculosis to isoniazid (INH) has been attributed to lesions in oxyR, a gene that positively regulates the expression of a set of hydrogen peroxide-inducible genes in E. coli and S. typhimurium. Moreover in the mechanism of emergence of INH resistance in M. tuberculosis, oxidative stress response has been implicated. In this study, mutants of E. coli defective in oxidative stress response function were derived and used to screen plant compounds, which might interfere with the oxidative stress response in MTB. Since MTB is inherently known to be oxyR defective and thus being highly sensitive to INH, mutants defective in oxidative stress response were isolated to construct a model system in E. coli, which is otherwise INH resistant, having functional oxyR. These mutants showed simultaneous sensitivity to oxidative stress-causing agents like hydrogen peroxide and cumene hydroperoxide. To further define the mutational lesions, complementation studies were carried out through mobilization of cloned wild type genes involved in the oxidative stress response and in this way a biological screen was constructed to identify plant compounds/essential oils/extracts/oil components which induce oxidative stress. The positives were finally tested for activity against M. tuberculosis strain H37Rv using the radiometric BACTEC 460 TB system. Interestingly, the bioactives were found to be active against the pathogen with marked potency, as the reduction in δGI values for the identified bioactives against M. tuberculosis were significant. The study demonstrates application of a specific target-based genetic model system in E. coli as a rapid high throughput screen in identifying anti-mycobacterials from plants.     

Potential impacts of climate warming on active soil organic carbon contents along natural altitudinal forest transect of Changbai Mountain

Understanding soil carbon fractions and their responses to the global warming is important for improving soil carbon management of natural altitudinal forest ecosystem. In this study, the contents of soil total organic carbon (SOC), soil labile organic carbon (LOC), and microbial biomass carbon (MBC) in soil upper layers (0–20 cm) were measured along a natural altitudinal transect in the north slope of Changbai Mountain. The results showed that under natural conditions the contents of SOC and LOC were largest in Betula ermanii forest (altitude 1996 m), moderate in spruce-fir forest (altitude 1350 m), and smallest in Korean pine mixed broad-leaf tree forest (altitude 740 m). MBC contents in different forest ecosystems decreased in the order of Betula ermanii forest, Korean pine mixed broad-leaf tree forest, and dark coniferous forest. In addition, the responses of SOC, LOC, and MBC to soil warming were conducted by relocating intact soil cores from high- to low-elevation forests for one year. As expected, the soil core relocation caused significant increase in soil temperature but made no significant effect on soil moisture. After one year incubation, soil relocation significantly decreased SOC contents, whereas the contents of LOC, MBC, and the ratios of LOC to SOC and MBC to SOC increased.     

Antimitotische Aktivität von Methylbenzimidazol-2-ylcarbamat (MBC)

Zusammenfassung Die unbeeinflußte Mitose vonAspergillus nidulans entspricht in zahlreichen ultrastrukturellen Details der Mitose anderer Asco- und Basidiomyceten. Unter dem Einfluß von 4 M MBC sind folgende Parameter der morphometrischen Analyse des Zellkern und des Kern-assoziierten Organells (NAO) verändert: 1. Es werden keine Mikrotubuli gebildet. 2. Das NAO ist extrem vergrößert und fragmentiert in zwei gleiche Teile, die sich auf der Kernhülle entlang bewegen und sich später teilen können. 3. Das Kernvolumen hat sich beträchtlich vergrößert. — Es wird diskutiert, ob MBC einen spezifischen Typ der NAO-Duplikation verursacht und sich die Zellkerne ohne Mikrotubuli teilen.

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Antimitotic activity of methylbenzimidazol-2-ylcarbamate (MBC)II. Electronmicroscopical investigations on duplication of the nucleus-associated organelle (centriolar plaque, MTOC, KCE) inAspergillus nidulans

Summary The unaffected mitosis ofAspergillus nidulans corresponds in many ultrastructural details with the mitosis of other asco- and basidiomycetes. Under the influence of 4 M MBC the following parameters are altered as shown by morphometric analysis of both nucleus and nucleus-associated organelle (NAO): 1. No microtubules are formed. 2. The NAO is extremely enlarged and fragments into two equal parts which move along the nuclear envelope and may divide another time. 3. The nuclear volume is considerably increased.—It is discussed whether MBC causes a specific type of NAO-duplication and nuclei can divide without microtubules.

Meinem Lehrer Prof. M. Girbardtzum 60. Geburtstag gewidmet     

Synthesis and in vitro antimicrobial activity of new 4-phenyl-5-methyl-4H-1,2,4-triazole-3-thione derivatives

This study presents the synthesis, spectral analysis and antimicrobial evaluation of a new series of substituted 1,2,4-triazole (5a–i) and 1,3,4-thiadiazole derivatives (9a, c, g, h). New compounds were obtained by cyclization reaction of acyl thiosemicarbazide derivatives in the presence of alkaline and acidic media. All synthesized compounds were screened for their in vitro antimicrobial activities. Nine of the compounds had potential activity against Gram-positive bacteria (MIC?=?3.91–500 µg/mL). Some compounds showed good activity especially against: Micrococcus luteus ATCC 10240 (MIC?=?3.91?31.25 µg/mL), Bacillus subtilis ATCC 6633 (MIC?=?15.63? 62.5 µg/mL), and Staphylococcus aureus ATCC 25923 (MIC?=?15.63?125 µg/mL).