Dystroglycan glycosylation and muscular dystrophy

Dystroglycan is an integral member of the skeletal muscle dystrophin glycoprotein complex, which links dystrophin to proteins in the extracellular matrix. Recently, a group of human muscular dystrophy disorders have been demonstrated to result from defective glycosylation of the α-dystroglycan subunit. Genetic studies of these diseases have identified six genes that encode proteins required for the synthesis of essential carbohydrate structures on dystroglycan. Here we highlight their known or postulated functions. This glycosylation pathway appears to be highly specific (dystroglycan is the only substrate identified thus far) and to be highly conserved during evolution.     

革兰阴性细菌耐药特性的双聚类分析

目的通过双聚类分析了解临床分离的革兰阴性菌的耐药特征。方法采用K-B法对临床分离的113株细菌进行药物敏感试验,用软件WHONET 5.6和MATLAB对药敏试验结果进行统计分析。结果传统耐药分析方法表明113株细菌总体耐药率较低,其中大肠埃希菌、铜绿假单胞菌和鲍曼不动杆菌的耐药率则较高,而肺炎克雷伯菌耐药率较低。通过双聚类分析,所有菌株被聚为三大类,I类菌株占23.0%,耐药率最高,几乎对18种药物都耐药,细菌种类以肺炎克雷伯菌、铜绿假单胞菌、鲍曼不动杆菌为主;Ⅱ类菌株占56.6%,耐药率普遍较低,以肺炎克雷伯菌为主;Ⅲ类菌株占20.4%,菌株的耐药率介于I和Ⅱ类之间,包括肺炎克雷伯菌、铜绿假单胞菌和大肠埃希菌。其中Ⅱ大类,根据所耐抗生素的不同又分为Ⅱ-A、Ⅱ-B、Ⅱ-C三个亚类,每一亚类都具有相似的耐药特点。结论本实验收集的革兰阴性菌株根据耐药特征被聚为Ⅰ、Ⅱ、Ⅲ类,耐药程度为IⅢⅡ,双聚类分析法有利于快速找到具有相同耐药特征的菌株以及不同菌株之间的耐药差别。     

对安贞医院革兰阴性杆菌的耐药趋势调查

目的分析安贞医院感染菌中,居前5位的革兰阴性杆菌的耐药趋势,为临床合理使用抗生素提供必要依据。方法对2000～2002年该院分离的医院感染菌株中,居前5位的革兰阴性杆菌的耐药性,进行回顾性分析。结果革兰阴性杆菌中前5位细菌,对3代头孢菌素的耐药性增高。大多数细菌对亚胺培南和美洛培南敏感。阿米卡星的耐药性有所下降。大肠埃希菌对喹诺酮类抗生素耐药率在85％以上。结论不规范使用抗生素,使细菌的耐药性越来越高,交替使用抗生素可能是降低细菌对抗生素耐药性的有效方法。医院应宏观控制使用抗生素。     

Characterization of reduced susceptibility to chlorhexidine among Gram-negative bacteria

Chlorhexidine gluconate (CHG) is one of the most commonly used antiseptic, acting against Gram-negative, Gram-positive bacteria, yeast and fungi. However, over use may lead to reduced susceptibility of different bacteria to CHG. This study aimed to characterize the CHG susceptibility among Gram-negative strains in Israel, to evaluate factors that may affect this susceptibility, and to compare CHG susceptibility between ESBLs bacteria to strains without these enzymes. Pseudomonas aeruginosa, Proteus mirabilis, Klebsiella spp, Escherichia coli, and Acinetobacter baumannii were isolated from clinical samples of 193 patients hospitalized at Padeh–Poriya Medical Center. Phenotypic CHG susceptibility was assessed by determining minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). The highest CHG MIC was found among P. mirabilis. The differences between the CHG MIC values among the different strains were statistically significant (p < 0.001). ESBL-positive strains had higher MIC values as compared to ESBL-negative strains (p = 0.030). A significant association was found between CHG susceptibility and sample source (p = 0.015). In conclusion, the information gathered here significantly improves our knowledge on the reduced susceptibility to CHG among Gram-negative bacteria in Israel. Moreover, ESBL-positive bacteria are less susceptible to CHG and finally, bacteria in sputum, wounds, and body fluids are less CHG-susceptible.     

Analysis of glycosylation motifs and glycosyltransferases in Bacteria and Archaea

The process of glycosylation has been studied extensively in prokaryotes but many questions still remain unanswered. Glycosyltransferase is the enzyme which mediates glycosylation and has its preference for the target glycosylation sites as well as for the type of glycosylation i.e. N-linked and O-linked glycosylation. In this study we carried out the bioinformatics analysis of one of the key enzymes of pgl locus from Campylobacter jejuni, known as PglB, which is distributed widely in bacteria and AglB from archaea. Relatively little sequence similarity was observed in the archaeal AglB(s) as compared to those of the bacterial PglB(s). In addition we tried to the answer the question of as to why not all the sequins Asp-X-Ser/Thr have an equal opportunity to be glycosylated by looking at the influence of the neighboring amino acids but no significant conserved pattern of the flanking sites could be identified. The software tool was developed to predict the potential glycosylation sites in autotransporter protein, the virulence factors of gram negative bacteria, and our results revealed that the frequency of glycosylation sites was higher in adhesins (a subclass of autotransporters) relative to the other classes of autotransporters.     

常见革兰阴性菌耐药性分析与合理用药研究

目的 分析芜湖市第一人民医院革兰阴性病原菌分布情况及对常见抗菌药物的耐药程度,为临床抗菌药物的选择及其合理使用提供参考。方法 收集2015年1月至2017年12月我院临床科室送检的细菌培养标本,采用法国生物梅里埃公司生产VITEK2 compact全自动微生物分析系统及其配套试剂盒进行细菌鉴定及药敏试验,数据采用WHONET 5.6软件进行统计分析。对年度耐药株数使用R语言3.4.0软件进行分析。结果 2015—2017年我院共检出病原菌3 728株,其中革兰阳性菌1 055株,占28.3%;革兰阴性菌2 673株,占71.7%。革兰阴性菌中主要致病菌为大肠埃希菌、肺炎克雷伯菌、铜绿假单胞杆菌、鲍曼不动杆菌,对青霉素类和第一代头孢菌素类抗菌药物高水平耐药。2017年这4种细菌对头孢唑林的耐药率均已达100.0%。鲍曼不动杆菌对头孢曲松的耐药率逐年上升,对头孢替坦连续3年耐药率均为100.0%,对氨苄西林/舒巴坦、氟喹诺酮类以及头孢吡肟耐药率未逐年上升,对亚胺培南耐药率变化显著,差异均有统计学意义（P<0.05）。铜绿假单胞杆菌对氨曲南耐药率逐年上升,对氨基糖苷类及头孢他啶耐药率逐年下降,差异均有统计学意义（P<0.05）。大肠埃希菌和肺炎克雷伯菌对头孢吡肟耐药率逐年下降,差异有统计学意义（P<0.05）。结论 我院住院患者病原菌以革兰阴性菌为主,不同病原菌对药物的敏感性各异,临床医生应结合患者情况和药敏结果综合分析,合理选用抗菌药物。     

The Streptomyces flora of Jordan and its' potential as a source of antibiotics active against antibiotic-resistant Gram-negative bacteria

Detection and identification of members of the genus Streptomyces are of great value because they provide a rich source of antibiotics. Toward the goal of identifying additional novel antibiotics, a total of 292 different Streptomyces isolates were recovered from 54 soil samples collected from 28 different locations in Jordan. These were then characterized by conventional methods and assessed for their activity against two antibiotic-resistant Gram-negative isolates of Escherichia coli and Klebsiella pneumoniae. Results revealed that grey, white and yellow series isolates were the most abundant, with 15% of the Streptomyces isolates active against at least one of the test pathogens. Most of the active isolates exhibited activity against E. coli (96%), while less activity was exhibited against K. pneumoniae (18%). Overall screening revealed the characterization of six Streptomyces isolates (I₇, AC₃₂, G₁₇, Z₁₁, Bb₃₆ and AQ₁₆) which inhibited the growth of both pathogens. All were obtained from a region characterized by low-nutrient soils and harsh conditions. The unusual antibiotic profile of these isolates stressed their potential as a source of novel antibiotics.     

重庆地区儿童2000年至2004年首位革兰阴性细菌和阳性细菌的耐药性监测

目的了解重庆地区儿童感染的分离至临床标本的首位革兰阴性细菌和阳性细菌对常用抗生素的耐药趋势,指导临床合理使用抗生素。方法常规方法分离、培养细菌,应用美国德灵公司WalkAway-40细菌鉴定仪对2000年至2004年我院细菌室分离至临床标本的首位革兰阴性细菌和阳性细菌共2854株进行细菌鉴定及药敏试验。结果2000年至2004年检出的首位革兰阴性细菌和阳性细菌分别为大肠埃希菌和金黄色葡萄球菌。2000年至2004年前5位革兰阴性菌5777株,革兰阳性菌1565株,其中大肠埃希菌2090株,金黄色葡萄球菌764株,分别占36.2%和48.8%;5年间大肠埃希菌对氨苄西林、头孢吡肟、头孢西丁、庆大霉素、亚胺培南、环丙沙星、头孢噻肟、头孢他啶的总耐药率分别为80.9%、37.5%、15.4%、54.0%、0.8%、34.0%、46.6%、46.2%;金黄色葡萄球菌对青霉素、红霉素、复方新诺明、万古霉素、阿莫西林/克拉维酸的总耐药率分别为95.6%、63.4%、5.8%、0%、11.0%。结论通过细菌耐药监测发现：大肠埃希菌对常用抗生素的总耐药率变化不大,金黄色葡萄球菌对常用抗生素的总耐药率有下降趋势,应引起临床医生重视。     

Mechanism of chlorpromazine binding by Gram-positive and Gram-negative bacteria

Chlorpromazine forms charge-transfer complexes with xanthene dyes in bacteria. These complexes permit the differentiation of Gram-positive and Gram-negative bacteria in both light and polarization microscopy. The birefringence induced by the charge-transfer complex might explain the molecular basis of bacterial staining.The charge-transfer complexes formed between chorpromazine and xanthene dyes accumulate in the bacterial cell, mainly inside the bacterial cell wall. The complexes give the cells a color, which depends on the chemical composition of the staining structure, and in particular the polysaccharides of the cell wall in bacteria.Metachromatic granules were seen inside Gram-positive bacteria after chlorpromazine and rose bengal staining. Although the nature of these granules remains unclear, this type of binding may have a role in the inhibition of biochemical processes in the bacterial cells.     

Micromorphology of Gram-negative hydrogen bacteria

The fine structure of the cell envelope, of membrane systems and of cytoplasmic inclusions of Gram-negative aerobic hydrogen bacteria has been studied. The results have been tabulated, and three main groups could be recognized: Group 1: Alcaligenes eutrophus, A. paradoxus, A. ruhlandii, Pseudomonas facilis, P. flava, P. pseudoflava, P. palleronii, and Aquaspirillum autotrophicum; Group 2: Corynebacterium autotrophicum and strains MA 2 and SA 35; Group 3: Paracoccus denitrificans. Special structures related to the chemoautotrophic way of life of the hydrogen bacteria were not observed.Abbreviations CM cytoplasmic membrane - OM outer membrane     

Discriminative features of type I and type III secreted proteins from Gram-negative bacteria

The amino acid composition of sequences and structural attributes (α-helices, β-sheets) of C-and N-terminal fragments (50 amino acids) were compared to annotated (SWISS-PROT/ TrEMBL) type I (20 sequences) and type III (22 sequences) secreted proteins of Gram-negative bacteria. The discriminant analysis together with the stepwise forward and backward selection of variables revealed the frequencies of the residues Arg, Glu, Gly, Ile, Met, Pro, Ser, Tyr, Val as a set of strong (1-P < 0.001) predictor variables to discriminate between the sequences of type I and type III secreted proteins with a cross-validated accuracy of 98.6–100 %. The internal and external validity of discriminant analysis was confirmed by multiple (15 repeats) test-retest procedures using a randomly split original set of proteins; this validation method demonstrated an accuracy of 100 % for 191 non-selected (retest) sequences. The discriminant analysis was also applied using selected variables from the propensities for β-sheets and polarity of C-terminal fragments. This approach produced the next highest and comparable cross-validated classification accuracy for randomly selected and retest proteins (85.4–86.0 % and 82.4–84.5 %, respectively). The proposed sets of predictor variables could be used to assess the compatibility between secretion substrates and secretion pathways of Gram-negative bacteria by means of discriminant analysis.     

Gram-negative bacteria associated with timber as a potential respiratory hazard for woodworkers

The concentration and species composition of viableGram-negative bacteria was determined in samples of coniferous wood (Scots pine) and deciduous wood (European beech), and in air samples taken during the processing of these woods in sawmills. The concentration of Gram-negative bacteria in the sapwood of pine logs ranged from 1.0 × 10¹ to 6.2 × 10⁴ CFU/g. Their concentration in the air of the pine processing sawmill was within a range of 1.0 × 10²–6.3 × 10² CFU/m³. Concentration of Gram-negative bacteria in the sapwood of beech logs was similar to that in the sapwood of pine logs, ranging from 1.0 × 10¹ to 4.6 × 10⁴ CFU/g. However, the concentration of these bacteria in the air of a beech processing sawmill was within a range of 7.8 × 10³–1.3 × 10⁴ CFU/m³, being significantly higher than in a pine processing sawmill. Enterobacteriaceae strains, mostly Enterobacter spp. and Rahnella spp., made up 70–75% of Gram-negative bacteria isolated from pine and beech wood and from the air pollutedwith sawdust from these woods. The aerial exposure to Gram-negative bacteria possessing endotoxic andallergenic properties poses potential risk ofoccupational respiratory disorders among sawmillworkers, in particular those processing beech wood.     

Buffering capacity and H+ membrane conductance of Gram-negative bacteria

Abstract Buffering capacity and membrane H⁺ conductance were examined in seven Gram-negative species: Aquaspirillum serpens, Pseudomonas aeruginosa, Alcaligenes faecalis, Escherichia coli, Salmonella typhimurium, Proteus mirabilis and Aeromonas hydrophila . All strains of Enterobacteriaceae studied here showed a decrease in both parameters as the external pH increased, over the pH range studied. The other four species presented an increase in buffering capacity and membrane conductance to protons as the external pH increased from 5.5 to 7.0.     

The importance of black-pigmented Gram-negative anaerobes in human infections

Abstract Black-pigmented Gram-negative anaerobic rods are found on mucosal surfaces as indigenous flora. With mucosal damage due to disease, trauma or surgery, these organisms may invade tissues and set up infection. Other important factors determining whether or not infection results include ‘inoculum’ size, synergy with other organisms and production of virulence factors that include capsules, lipopolysaccharide, attachment factors, proteases, collagenase, neuraminidase, and phospholipase A; also, they may have fibrinolytic and anti-phagocytic activity and may degrade complement and IgG and IgM. Pigmented anaerobes are found in all types of infections including such serious infections as bacteraemia, endocarditis, intracranial abscess, necrotizing pneumonia and necrotizing fasciitis, generally as part of a mixed infecting flora, and they play a key role in experimental mixed infections. They dominate or are prominent in infections involving organisms originating in the oropharynx, such as central nervous system, head and neck, dental and pleuropulmonary infections. Therapy of infections involving pigmented anaerobes includes surgery plus antimicrobial agents; a significant percentage of strains produce β-lactamase. Much remains to be done to determine the relative importance of the various taxa of black-pigmented Gram-negative anaerobes and of the different virulence factors produced by them.     

Nonspecific porins of the outer membrane of Gram-negative bacteria: Structure and functions

The structure and functional properties of nonspecific porins (β-structured integral proteins of the outer membrane of Gram-negative bacteria) are overviewed. The characteristic features of porin spatial structure related to the principles of β-barrel construction and pore geometry are considered. The data concerning nonspecific diffusion of low-molecular substances and dynamic behavior of porin channels dependent on the distribution of charged amino acid residues in different structural domains of the porin molecule are presented. The methods and approaches used in the study of functional activity of porins are surveyed. The data on modulation of pore-forming activity of these proteins by external factors and membrane components are considered separately.     

Design and synthesis of potent Gram-negative specific LpxC inhibitors

Antibiotic resistant hospital acquired infections are on the rise, creating an urgent need for novel bactericidal drugs. Enzymes involved in lipopolysaccharide (LPS) biosynthesis are attractive antibacterial targets since LPS is the major structural component of the outer membrane of Gram-negative bacteria. Lipid A is an essential hydrophobic anchor of LPS and the first committed step in lipid A biosynthesis is catalyzed by a unique zinc dependent metalloamidase, UDP-3-O-(R-3-hydroxymyristoyl)-N-acetylglucosamine deacetylase (LpxC). LpxC is an attractive Gram-negative only target that has been chemically validated by potent bactericidal hydroxamate inhibitors that work by coordination of the enzyme’s catalytic zinc ion. An exploratory chemistry effort focused on expanding the SAR around hydroxamic acid zinc-binding ‘warheads’ lead to the identification of novel compounds with enzyme potency and antibacterial activity similar to CHIR-090.     

Regulation and function of axon guidance and adhesion molecules during olfactory map formation

The olfactory system presents a practical model for investigating basic mechanisms involved in patterning connections between peripheral sensory neurons and central targets. Our understanding of olfactory map formation was advanced greatly by the discovery of cAMP signaling as an important determinant of glomerular positioning in the olfactory bulb. Additionally, several cell adhesion molecules have been identified recently that are proposed to regulate homotypic interactions among projecting axons. From these studies a model has emerged to partially explain the wiring of axons from widely dispersed neuron populations in the nasal cavity to relatively stereotyped glomerular positions. These advances have revitalized interest in axon guidance molecules in establishing olfactory topography, but also open new questions regarding how these patterns of guidance cues are established and function, and what other pathways, such as glycosylation, might be involved. This review summarizes the current state of this field and the important molecules that impact on cAMP-dependent mechanism in olfactory axon guidance.