Enterobacter radicincitans sp. nov., a plant growth promoting species of the family Enterobacteriaceae

A plant growth promoting bacterial isolate (D5/23T) from the phyllosphere of winter wheat, able to fix atmospheric nitrogen and to produce auxines and cytokinins was investigated in a polyphasic taxonomy approach. Phylogenetic analyses using the 16S rRNA gene sequence of the strain clearly indicated that the strain belonged to the family Enterobacteriaceae, most closely related to Enterobacter cloacae with 99.0% and Enterobacter dissolvens with 98.5% sequence similarity. Phylogenetic analysis derived from the sequence of the rpoB gene showed the highest sequence similarities to Enterobacter cowanii (93.0%) but supported the distinct position of strain D5/23T. The isolate produced a fatty acid pattern typical for members of the family Enterobacteriaceae. On the basis of the phylogenetic analyses, DNA-DNA hybridizations, and the unique physiological and biochemical characteristics, we propose that strain D5/23T represents a new species of the genus Enterobacter for which we propose the name Enterobacter radicincitans sp. nov.     

A simplified method for assay of hydrogenase activities of H<Subscript>2</Subscript> evolution and uptake in <Emphasis Type="Italic">Enterobacter aerogenes</Emphasis>

Assay of hydrogenase activity pertaining to H₂ production needs anaerobic conditions. To establish a simplified method for assay of hydrogenase activities by using intact cells of Enterobater aerogenes, different chemicals capable of enhancing the cell-wall permeability to electron mediators were examined. As a result, Triton X-100 and CTAB were found to be appropriate for H₂ uptake and evolution activities of the intact cells, respectively. This method enabled H₂ uptake and evolution activities of the intact cells to be easily detected. This is also the first report of the presence of H₂ uptake hydrogenase activity in E. aerogenes.Revisions requested 2 March 2005 and 21 April 2005; Revisions received 12 April 2005 and 17 May 2005     

Purification, characterization, and gene cloning of 4-hydroxybenzoate decarboxylase of Enterobacter cloacae P240

We found the occurrence of 4-hydroxybenzoate decarboxylase in Enterobacter cloacae P240, isolated from soils under anaerobic conditions, and purified the enzyme to homogeneity. The purified enzyme was a homohexamer of identical 60 kDa subunits. The purified decarboxylase catalyzed the nonoxidative decarboxylation of 4-hydroxybenzoate without requiring any cofactors. Its K _m value for 4-hydroxybenzoate was 596 μM. The enzyme also catalyzed decarboxylation of 3,4-dihydroxybenzoate, for which the K _m value was 6.80 mM. In the presence of 3 M KHCO₃ and 20 mM phenol, the decarboxylase catalyzed the reverse carboxylation reaction of phenol to form 4-hydroxybenzoate with a molar conversion yield of 19%. The K _m value for phenol was calculated to be 14.8 mM. The gene encoding the 4-hydroxybenzoate decarboxylase was isolated from E. cloacae P240. Nucleotide sequencing of recombinant plasmids revealed that the 4-hydroxybenzoate decarboxylase gene codes for a 475-amino-acid protein. The amino acid sequence of the enzyme is similar to those of 4-hydroxybenzoate decarboxylase of Clostridium hydroxybenzoicum (53% identity), VdcC protein (vanillate decarboxylase) of Streptomyces sp. strain D7 (72%) and 3-octaprenyl-4-hydroxybenzoate decarboxylase of Escherichia coli (28%). The hypothetical proteins, showing 96–97% identities to the primary structure of E. cloacae P240 4-hydroxybenzoate decarboxylase, were found in several bacterial strains.     

Nonrecovery of varying proportions of viable bacteria during spread plating governed by the extent of spreader usage and proposal for an alternate spotting-spreading approach to maximize the CFU

Aims: To elucidate the cause of high variations and inconsistencies in bacterial CFU observed within and between different experiments while assessing viable bacterial counts through spread plating (SP). Methods and Results: Following the inconsistent results, CFU estimations were undertaken through conventional SP using the spreader, or a modified approach that did not use spreader employing four organisms. The latter approach involving spotting‐and‐tilt‐spreading of inoculum on agar surface [spotting spreading (SS)] yielded higher CFU by 11–120% over the weighted average depending on the organism and diluent. The adverse effect owing to the spreader was the most obvious in Escherichia coli followed by Staphylococcus epidermidis, Enterobacter cloacae and Bacillus pumilus. Plate attributes that determined the surface moisture levels of agar medium and the spreading practice adopted by the personnel formed two other major influencing factors. Plating for shorter periods (<60 s) using fresh 15/20 ml plates caused loss of 3–12% CFU owing to inoculum adhesion to spreader irrespective of glass or polypropylene make. On the other hand, prolonging the plating brought down the CFU significantly. Spreader movement on agar surface subsequent to the exhaustion of free moisture, which was marked by the experiencing of some friction to smooth spreader movement, was detrimental to vegetative cells, while Bacillus spores were less affected. Conclusions: The study brings out that the way SP is carried out exerts significant effects on CFU influenced by plate conditions. Prolonged use of spreader on dry agar surface could be highly detrimental to bacterial cells. A mild use of spreader accounting for spreader‐adhering inoculum or the practice of SS not involving the spreader is recommended. Significance and Impact of the Study: This study unravels the effects owing to the spreader on bacterial cells and the CFU and recommends an alternate approach of SS to minimize CFU inconsistencies and to maximize the viable bacterial counts.     

229株阴沟肠杆菌的标本分布及耐药性分析

目的 了解阴沟肠杆菌在医院感染的标本分布和耐药情况,为临床合理选择和应用抗生素提供依据.方法 采用VITEK 60全自动微生物分析仪和配套的GNI、GNS-143、GNI-448,对229株阴沟肠杆菌进行分离鉴定和药敏试验,药敏结果使用WHONET 5.5软件进行分析.结果 从2007年1月至2011年9月共分离到阴沟肠杆菌229株,59.0％来自于呼吸道标本,其次是尿液占13.5％,再次为创面分泌物/脓液占12.7％.阴沟肠杆菌对美洛培南、亚胺培南和头孢哌酮/舒巴坦的耐药率分别为0.0％、0.4％和2.5％,对氨苄西林、头孢唑啉、头孢西丁的耐药率分别为98.7％、96.9％、97.5％.结论 阴沟肠杆菌耐药机制复杂,对抗生素具有多重耐药性,临床应合理使用抗菌药物,以减少阴沟肠杆菌耐药性的产生和在院内扩散.     

肠杆菌新解释标准指导临床用药的实用性

采用新旧肠杆菌的2种解释标准,分析肠杆菌的耐药性,比较2种解释标准的临床意义。收集本院2009至2010两年临床分离的912株大肠埃希菌、328株肺炎克雷伯菌,用Kirby-Bauer法作药敏试验,用WO-NET5.4软件先设置CLSI推荐的旧的肠杆菌解释标准,分析耐药性。再设置CLSI推荐的新的肠杆菌解释标准[1]修改头孢曲松、头孢他啶、头孢噻肟、氨曲南和亚胺培南的解释标准,分析其耐药性。用肠杆菌旧的解释标准分析常规检测分离出的大肠埃希菌236株产酶株;肺炎克雷伯66株产酶株;产酶株比非产酶株的耐药率高。用新的头孢菌素和氨曲南的解释标准:对于大肠埃希菌头孢曲松的耐药率提高5.2%;头孢他啶提高10.4%;头孢噻肟提高10.2%;氨曲南提高7.1%;对于肺炎克雷伯菌头孢曲松的耐药率提高6.7%;头孢他啶提高4.3%;头孢噻肟提高10.1%;氨曲南提高2.5%;未向临床报产酶株。采用碳青霉烯类抗生素新的解释标准大肠埃希菌和肺炎克雷伯菌对亚胺培南的耐药分别提高了0.4%和0.6%,对美罗培南的耐药率分别提高了0.2%和0.6%。新的肠杆菌解释标准更能客观分析肠杆菌的耐药性,对指导临床合理用药,控制耐药株的蔓延更具实用价值。     

Effect of environmental stresses on the sensitivity of Enterobacter sakazakii in powdered infant milk formula to gamma radiation

Aim:  To evaluate the effect of starvation, heat, cold, acid, alkaline, chlorine and ethanol stresses on the resistance of Enterobacter sakazakii in powdered infant milk formula (PIMF) towards gamma radiation.
Methods and Results:  Stressed cells of E. sakazakii ATCC 51329 and four other food isolate strains were mixed individually with PIMF, kept overnight at room temperature, and then exposed to gamma radiation up to 7·5 kGy. The D ₁₀-values were determined using linear regression and for the stressed E. sakazakii strains these values ranged from 0·82 to 1·95 kGy.
Conclusions:  Environmental stresses did not significantly change the sensitivity of most E. sakazakii strains to ionizing radiation.
Significance and Impact of the Study:  Data obtained established that most forms of environmental stress are unlikely to significantly enhance the resistance of E. sakazakii strains to lethal, low dose irradiation treatment.