Psichogiou M, Tassios P T, Avlamis A, Stefanou I, Kosmidis C, Platsouka E, Paniara O, Xanthaki A, Toutouza M, Daikos G L, Tzouvelekis L S
First Department of Propaedeutic Medicine, Medical School, University of Athens, Athens, Greece.
J Antimicrob Chemother. 2008 Jan;61(1):59-63. doi: 10.1093/jac/dkm443. Epub 2007 Nov 13.
To determine the current frequency and study the characteristics of VIM-1-producing Klebsiella pneumoniae isolates from bloodstream infections in Greek hospitals.
All blood isolates of K. pneumoniae were prospectively collected during 2004-06 in three teaching hospitals located in Athens. MICs of antibiotics were determined by the Etest. Extended-spectrum- (ESBL) and metallo-beta-lactamase (MBL) production was examined by clavulanate- and EDTA-based techniques, respectively. Isolates were typed by PFGE of XbaI-digested genomic DNA. Detection of bla(VIM-1) and mapping of the VIM-1-encoding integrons were performed by PCR and sequencing. Beta-lactamase activities were analysed by IEF and imipenem hydrolysis was assessed by spectrophotometry. VIM-1-encoding plasmids were transferred to Escherichia coli by conjugation and transformation and characterized by Inc/rep typing and RFLP.
Sixty-seven (37.6%) of 178 K. pneumoniae blood isolates were bla(VIM-1)-positive (VPKP); 77.8% of these were from ICUs. All VPKP isolates were multidrug-resistant. The MICs of carbapenems for VPKP varied from the susceptible range to high-level resistance overlapping with those of MBL-negative isolates. The EDTA-imipenem synergy methods had reduced sensitivity in detecting VPKP isolates when the MICs were in the susceptible range. ESBL production was common among VPKP isolates (n = 45, 67.2%) as indicated by resistance to aztreonam and confirmed by a clavulanate-based double-disc synergy test. The responsible ESBL was always an SHV-5-type enzyme as indicated by IEF. PFGE identified eight clusters (A-H) of VPKP isolates with related (>80%) patterns, as well as four unique types. Both inter-hospital spread of several clones and genotypic similarities among susceptible, ESBL-positive and VPKP isolates were also observed. Location of bla(VIM-1) and expression of VIM-1 were studied in 12 isolates representing the eight PFGE clusters. In all isolates, bla(VIM-1) was part of a class 1 integron that also carried aacA4, dhfrI, aadA and sulI. In eight isolates (clusters C, D, G and H), the bla(VIM-1) integron was located in transferable IncN plasmids. A cluster F isolate carried a VIM-1-encoding, self-transferable plasmid that was not typeable by Inc/rep typing. VIM-1-encodingreplicons were not identified in three isolates (PFGE clusters A, B and E). VPKP isolates exhibited differences in imipenem-hydrolysing activities which, however, were not correlated with the respective carbapenem MICs.
A multiclonal epidemic of bla(VIM-1)-carrying K. pneumoniae is under way in the majorhospitals in Greece. Microorganisms producing both VIM-1 and SHV-5 constitute the prevalent multidrug-resistant population of K. pneumoniae in this setting.
确定希腊医院血流感染中产VIM-1肺炎克雷伯菌分离株的当前频率并研究其特征。
2004年至2006年期间，前瞻性收集雅典三家教学医院的所有肺炎克雷伯菌血液分离株。采用Etest法测定抗生素的最低抑菌浓度（MIC）。分别采用基于克拉维酸和乙二胺四乙酸（EDTA）的技术检测超广谱β-内酰胺酶（ESBL）和金属β-内酰胺酶（MBL）的产生情况。通过对经XbaI酶切的基因组DNA进行脉冲场凝胶电泳（PFGE）对分离株进行分型。通过聚合酶链反应（PCR）和测序检测bla(VIM-1)基因并对VIM-1编码整合子进行定位。通过等电聚焦（IEF）分析β-内酰胺酶活性，通过分光光度法评估亚胺培南水解情况。通过接合和转化将携带VIM-1编码的质粒转移至大肠埃希菌，并通过Inc/rep分型和限制性片段长度多态性（RFLP）进行鉴定。
178株肺炎克雷伯菌血液分离株中有67株（37.6%）bla(VIM-1)基因阳性（VIM-1阳性肺炎克雷伯菌，VPKP）；其中77.8%来自重症监护病房（ICU）。所有VPKP分离株均对多种药物耐药。VPKP对碳青霉烯类药物的MIC值从敏感范围到高水平耐药不等，与MBL阴性分离株的MIC值重叠。当MIC值处于敏感范围时，EDTA-亚胺培南协同试验检测VPKP分离株的敏感性降低。如对氨曲南耐药并经基于克拉维酸的双纸片协同试验证实，VPKP分离株中常见ESBL产生（n = 45，67.2%）。IEF结果显示，相关ESBL始终为SHV-5型酶。PFGE鉴定出VPKP分离株的八个簇（A - H），其图谱相关（>80%），以及四种独特类型。还观察到几个克隆在医院间传播以及敏感、ESBL阳性和VPKP分离株之间的基因型相似性。在代表八个PFGE簇的12株分离株中研究了bla(VIM-1)基因的位置和VIM-1的表达。在所有分离株中，bla(VIM-1)是1类整合子的一部分，该整合子还携带aacA4、dhfrI、aadA和sulI。在八株分离株（簇C、D、G和H）中，bla(VIM-1)整合子位于可转移的IncN质粒中。一株簇F分离株携带一个编码VIM-1的自我转移质粒，无法通过Inc/rep分型进行鉴定。在三株分离株（PFGE簇A、B和E）中未鉴定出VIM-1编码复制子。VPKP分离株在亚胺培南水解活性方面存在差异，然而，这与各自的碳青霉烯类药物MIC值无关。
希腊各大医院正在发生携带bla(VIM-1)肺炎克雷伯菌的多克隆流行。同时产生VIM-1和SHV-5的微生物构成了这种情况下肺炎克雷伯菌中普遍存在的多重耐药菌群。
