Obligate aerobic, gram-positive, weak acid-fast, nonmotile bacilli, Tsukamurella tyrosinosolvens: Minireview of a rare opportunistic pathogen

Tsukamurella species are obligate aerobic,gram-positive,weak acid-fast,nonmotile bacilli.They are found in various environments,such as soil,water,sludge,and petroleum reservoir wastewater,and belong to the order Actinomycetales.In 2016,there was a reclassification of species within the genus Tsukamurella,merging the species Tsukamurella tyrosinosolvens(T.tyrosinosolvens)and Tsukamurella carboxydivorans.Tsukamurella species are clinically considered to be a rare opportunistic pathogen,because most reported cases have been related to bacteremia and intravascular prosthetic devices and immunosuppression.To date,it has been isolated only from human specimens,and has always been associated with clinical disease;human infections are very rare.Reported infections have included pneumonia,brain abscesses,catheter-related bloodstream infections,ocular infections,bacteremia,and sepsis presenting with septic pulmonary emboli in patients who are immunocompromised.To date,there is no commercially available test for identification.On the other hand,sequence-based identification,including matrix-assisted laser desorption ionization time-of-flight mass spectrometry,is an alternative method for identifying clinical isolates that are either slow growers or difficult to identify through biochemical profiling.The golden standards for diagnosis and optimal management still remain to be determined.However,newer molecular biological techniques can provide accurate identification,and contribute to the appropriate selection of definitive therapy for infections caused by this organism.Combinations of several antimicrobial agents have been proposed for treatment,though the length of treatment for infections has yet to be determined,and should be individualized according to clinical response.Immunocompromised patients often experience severe cases due to infection,and life-threatening T.tyrosinosolvens events associated with dissemination and/or failure of source control have occurred.Favorable prognoses can be achieved through earlier identification of the cause of infection,as well as successful management,including appropriate antibiotic therapy together with source control.Further analyses of similar cases are required to establish the most adequate diagnostic methods and treatment regimens for infections.

First Report of Tsukamurella conjunctivitidis CAPD-related Peritonitis

Introduction Using the information obtained from analysis of 16S rRNA gene sequences,Tsukamurella was first proposed as a genus in 1988,1 despite the fact that the first strain of this group of bacteria was described in 1941,2 and the first human isolate was reported in 1971.3 Similar to related genera of the order Corynebacteriales,such as Nocardia,Rhodococcus,and Gordonia,members of Tsukamurella are Gram-positive,aerobic,catalase-positive,and partially acid-fast as a result of the presence of mycolic acid in the cell envelope.Due to their similar phenotypic properties,differentiation of these genera and speciation within these genera is difficult in most clinical microbiology laboratories.

冢村菌致导管相关血流感染1例并文献复习

目的探讨冢村菌所致导管相关血流感染的临床表现和诊治,提高临床对该病的认识。方法回顾1例冢村菌引起的导管相关血流感染患者的临床资料,并以英文检索词“Tsukamurella”检索PubMed数据库,以中文检索词“冢村菌”或“冢村氏菌”检索中国知网、万方和维普数据库,获取国内外报道的冢村菌致导管相关血流感染的病例资料并进行总结分析。结果患者系37岁男性,基础疾病为短肠综合征、脾功能亢进、腹腔硬纤维瘤,长期留置有经外周静脉穿刺中心静脉导管,主要起病症状为发热,经血培养和导管尖端培养明确诊断为冢村菌致导管相关血流感染,先后经头孢哌酮-舒巴坦、亚胺培南-西司他丁抗感染后治愈。文献检索获取到56个病例,多为免疫功能受损患者。多数病例采用化学分类和/或分子技术鉴定方法进行了病原菌鉴定。在治疗过程中,大多患者的血管内导管被拔除。除1例患者结局未知,55例患者中有54例生存,1例死亡,但不能明确其是否死于冢村菌感染。结论冢村菌是一种机会致病菌,引起的临床感染相当少见,其鉴定有赖于质谱技术和基因测序。除了选用合适的抗生素,拔除血管内导管对控制感染往往必不可少。

生物拆分制备(S)-α-乙基-2-氧-1-吡咯烷乙酸工艺研究

耐酪氨酸冢村氏菌(Tsukamurella tyrosinosolvens)E105可选择性催化(R,S)-α-乙基-2-氧-1-吡咯烷乙酸乙酯生物拆分制备左乙拉西坦关键手性中间体(S)-α-乙基-2-氧-1-吡咯烷乙酸.通过单因素实验和响应面法对菌株产酶培养条件进行优化,并考察含酶静息细胞催化制备(S)-α-乙基-2-氧-1-吡咯烷乙酸的反应过程.结果表明:最优产酶培养条件为装液量30%,接种量4%,发酵36h;培养基组成为葡萄糖17.70g/L,酵母膏16.53g/L,NH4Cl 9.5g/L,K2HPO42g/L,KH2PO41g/L,NaCl 0.6g/L,MgSO4·7H2O 0.5g/L,初始pH值7.32.在优化条件下,酶活力为19.33U/g,细胞干重达4.13g/L,分别较优化前提高了32.6%和60.1%.当细胞加量为30g/L,底物浓度60mmol/L,反应体系为300mL,拆分24h后,产率和ee值分别为48%和99%,(S)-α-乙基-2-氧-1-吡咯烷乙酸浓度达28.8mmol/L,较优化前提高了32.1%.

α-乙基-2-氧-1-吡咯烷乙酸乙酯水解酶产生菌发酵培养基的响应面优化

对耐酪氨酸冢村氏菌(Tsukamurella tyrosinosolvens)E105菌株生物拆分外消旋体α-乙基-2-氧-1-吡咯烷乙酸乙酯制备左乙拉西坦关键手性中间体(S)-α-乙基-2-氧-1-吡咯烷乙酸的发酵培养基进行优化,以提高其产酶活力.首先考察了碳源和氮源对菌种产酶活力的影响,并通过Plackett-Burman实验得出影响该菌株产水解酶的最主要因素为酵母粉质量浓度和初始pH值.继而采用中心组合实验并结合响应面分析优化发酵培养基组成,确定了最佳的酵母粉质量浓度为12.1g/L,最佳的初始pH值为7.06.在优化的条件下酶活力可达1 024.6U/mL,较优化前提高了67%,表明采用响应面法优化发酵培养基组成是提高该菌株产酶活力的有效途径之一.

利用纤维床生物反应器制备左乙拉西坦关键手性中间体

利用纤维床生物反应器固定耐酪氨酸冢村氏菌(Tsukamurella tyrosinosolvens)E105细胞不对称水解(R,S)-α-乙基-2-氧代-1-吡咯烷乙酸乙酯制备左乙拉西坦关键手性中间体(S)-α-乙基-2-氧代-1-吡咯烷乙酸.考察了载体的化学修饰、缓冲液pH和离子强度等对棉纤维载体吸附细胞的影响,确定了最优的菌体吸附条件;在此基础上研究了纤维床生物反应器细胞动态吸附动力学以及利用纤维床生物反应器固定化细胞不对称水解(R,S)-α-乙基-2-氧代-1-吡咯烷乙酸乙酯制备左乙拉西坦关键手性中间体(S)-α-乙基-2-氧代-1-吡咯烷乙酸反应的过程曲线、细胞负载量对产率和ee值的影响,继而,考察了纤维床反应器中固定化细胞的重复利用性能.结果表明:在1.115mol/L,pH7.0的磷酸钾缓冲液中,以聚乙烯亚胺修饰棉纤维为固定化载体,细胞最大吸附量可达185mg/g;细胞在纤维床生物反应器上的动态吸附符合一级动力学,动力学方程为Ln(C/C0)=-0.947t;以细胞负载浓度为50g/L的纤维床生物反应器进行不对称水解反应,反应36h后产率和ee值分别为43.2%和98.2%,且纤维床反应器固定化细胞具有一定的重复利用性能.研究结果为纤维床反应器与发酵罐偶联进行半连续化制备左乙拉西坦手性中间体奠定了一定的理论基础.

一株冢村菌的筛选及其对邻苯二甲酸二丁酯的降解特性研究

为了获得邻苯二甲酸酯(PAEs)生物降解菌,从农田土壤筛选出一株能够以PAEs为唯一碳源和能源生长的微生物,命名为SD2。采用聚合酶链式反应(PCR)扩增得到其16S rDNA并构建分子系统发育树,可鉴定出其为冢村菌(Tsukamurella)。研究该菌株在不同的温度、pH值和转速条件下对邻苯二甲酸二丁酯(DBP)的降解情况,同时采用高效液相色谱(HPLC)测定该菌株对DBP的降解能力。结果表明,SD2对DBP的最佳降解条件为:温度为30℃,pH值为9,转速为150 r/min。在该最佳条件下,SD2在72 h内可以完全降解500 mg/L的DBP。底物广谱试验表明,该菌能够降解长短链的PAEs化合物及常见的代谢产物。同时采用气相质谱联用仪(GC-MS)检测发现,DBP在降解的过程中产生了中间代谢产物邻苯二甲酸丁基甲酯和邻苯二甲酸(PA),预示着该菌具备完全降解DBP的能力。这是首次关于冢村菌降解PAEs污染物的报道,能够为PAEs污染环境的生物降解及修复提供新的微生物资源。

两株PGPR菌株的花生定殖及对根际细菌群落结构的影响

为探讨2株根际促生菌耐酪氨酸束村氏菌P9和吡咯伯克霍尔德氏菌P10对花生的促生机制。利用GFP及利福平对2个菌株进行标记、结合扫描电镜观察,追踪了2株PGPR菌株在花生组织中的定殖动态;并通过16S rRNA全长测序对菌株接种花生根际土壤的细菌多样性进行分析。结果表明,利福平标记的P9和P10菌株具有良好的遗传稳定性,其生长和促生特性与原始菌株基本一致。GFP标记菌株可在花生的根尖及其分生区定殖;利福平标记菌株可稳定定殖在土壤及花生的根、茎部,且接菌30 d后定殖数仍保持在10CFU/g数量级。与未接菌植株根际土壤相比,P9、P10及混合菌株接种组的细菌群落相似性更高;接菌组的Flavihumibacter、unidentified_Rhizobiaceae的相对丰度显著增加,芽孢杆菌属、链霉菌属等的丰度较CK有不同程度增加,溶杆菌属、无色杆菌属及假黄单胞菌属等的丰度降低。2株PGPR菌株均可通过直接定殖在植株组织中、间接影响土壤细菌群落结构而发挥对花生的促生作用,混合菌株接种效果更优。研究结果明析了2株促生菌的促生机制,并为菌株的应用提供了科学依据。