不動桿菌屬 (Acinetobacter) 和其他葡萄糖非發酵菌在臨床上是重要的病原菌。然而，臨床實驗室卻沒有一套簡單又正確的方法可以用來區分Acinetobacrer。Acinetobacter中的A. calcoaceticus (genospecies 1), A. baumannii (genospeices 2), genospeices 3 (未命名)，genospecies 13TU (未命名)，因生化特性十分相近，故統稱為Acinetobacter calcoaceticus-Acinetobacter baumannii complex (Acb complex), Acb complex為Acinetobacter中分離率最高的細菌。本研究評估核糖體核酸基因的內轉錄區 (internal transceribed spacer region, ITS) 序列作為鑑定臨床Acb complex之可行性。目前GenBank資料庫中所提供的ITS序列仍不足，因此需要架構完整的ITS序列資料庫，才能用來鑑定Acb complex或其他臨床常見非發酵菌。本研究以PCR擴增108株 (42種) 參考菌株的ITS序列，並結合GenBank資料庫的序列，並初步以此資料庫做為鑑定Acb complex之工具。以此資料庫測試82株Acb complex臨床分離株，這82株臨床分離株並進一步以API 20 NE測定，結果63株為A. baumannnii，19株為A. calcoaceticus。若以ITS序列分析則發現這63株A. baumannii中，有2株為genospecies 3，14株可能為未命名的Acinetobacter spp.，所以事實上臨床鑑定之A. baumannii可能有高達 25.4%為鑑定錯誤。在19株A. calcoaceticus中，有16株為genospecies 3，而3株可能為未命名的Acinetobacter spp.，所以事實上臨床鑑定之A. calcoaceticus可能皆為鑑定錯誤。這些未命名的Acinetobacter spp.中，可分為五個genotypes，其中絕大多數 (13株，76%) 為同一genotype。這些未命名的Acinetobacter spp.需進一步以DNA-DNA hybridization的方法確認其基因種(genospecies)。本研究中的ITS序列資料可做為未來發展DNA chip之基礎。

Members of the genus Acinetobacter and other nonfermentative gram-negative bacteria are clinically important pathogens. However, clinical laboratories still lack simple methods that allow accurate identification of these species. For example, species of the Acinetobacter calcoaceticus- Acinetobacter baumannii complex (Acb complex) that includes Acinetobacter calcoaceticus (genospecies 1), A. baumannii (genospecies 2), unnamed genospecies 3 and 13TU are difficult to be differentiated. In fact, a complete discrimination between these species is possible only by DNA-DNA hybridization that is a complex and time-consuming method and cannot be performed in most clinical laboratories. In this study, the feasibility of using the sequences of 16S-23S rDNA spacer region (ITS) for the identification of Acb complex was evaluated. The ITS data in GenBank were insufficient to identify clinically relevant nonfermenters. The ITS regions from 108 strains (42 species) were amplified by PCR and sequenced; the sequence data in combination with those available in GenBank were used to construct an ITS sequence database for the identification of Acb complex. The database was used to test 82 clinical isolates of Acb complex; these microorgranisms included 63 isolates of A. baumannii and 19 isolates of A. calcoaceticus, as verified by API 20 NE. It is interesting to find among the 63 isolates of A. baumannii, 2 isolates might be genospecies 3 and 14 isolates were ungroupable, as revealed by ITS sequence analysis. Therefore, it is possible that 25.4% of clinical isolates of A. baumannii may be misidentified. Furthermore, among the 19 clinical isolates of A. calcoaceticus, 16 isolates might be genospecies 3 and 3 isolates were ungroupable as evidenced by ITS sequence analysis, and it seems that the designation of A. calcoaceticus to clinical isolates is, under most conditions, not correct. The 17 ungroupable isolates could be divided into 5 genotypes. The delineation of these ungroupable isolates, with their ITS sequences close to genospecies 2 and 13TU, needs further investigation such as DNA-DNA hybridization experiment. The ITS sequence database of nonfermenters may be useful for developing a DNA chip for identification of these microorganisms.

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