Walckenaer Estelle, Leflon-Guibout Véronique, Nicolas-Chanoine Marie-Hélène
Service de Microbiologie, Faculté de Médecine D. Diderot, Paris, France.
J Microbiol Methods. 2008 Dec;75(3):405-10. doi: 10.1016/j.mimet.2008.07.011. Epub 2008 Jul 22.
Although Raoultella planticola and Raoultella ornithinolytica were described more than 20 years ago, identifying them remains difficult. The reliability of the chromosomal bla gene for this identification was evaluated in comparison with that of the 16S rDNA and rpoB genes in 35 Raoultella strains from different origins. Of the 26 strains previously identified as R. planticola by biochemical tests alone or in association with molecular methods, 21 harboured a bla gene with 99.8% identity with the bla gene of two reference R. ornithinolytica strains (bla(ORN) gene) and 5 harboured a bla gene with 99.2% identity with the bla gene of two reference R. planticola strains (bla(PLA) gene). The 9 isolates previously identified as R. ornithinolytica harboured a bla(ORN) gene. The bla gene-based identification was confirmed by 16S rDNA and rpoB sequencing. The 21 isolates newly identified as R. ornithinolytica had a test negative for ornithine decarboxylase (ODC). Molecular experiments suggested one copy of ODC-encoding gene in both ODC-negative R. ornithinolytica and R. planticola strains and two copies in ODC-positive R. orninthinolytica strains. Analysis of the 35 bla genes allowed us (i) to confirm an identity of only 94% between the bla genes of the two Raoultella species while this identity was > 98% for rpoB and > 99% for 16S rDNA genes and (ii) to develop and successfully apply a bla PCR RFLP assay for Raoultella spp. identification. Overall, this study allowed us to discover ODC-negative R. ornithinolytica and to provide a reliable Raoultella identification method widely available as not requiring sequencing equipment.
尽管植生拉乌尔菌和鸟氨酸拉乌尔菌在20多年前就已被描述，但对它们进行鉴定仍然困难。在来自不同来源的35株拉乌尔菌中，将染色体bla基因用于该鉴定的可靠性与16S rDNA和rpoB基因的可靠性进行了比较评估。在之前仅通过生化试验或结合分子方法鉴定为植生拉乌尔菌的26株菌株中，21株携带与两株鸟氨酸拉乌尔菌参考菌株的bla基因具有99.8%同一性的bla基因（bla(ORN)基因），5株携带与两株植生拉乌尔菌参考菌株的bla基因具有99.2%同一性的bla基因（bla(PLA)基因）。之前鉴定为鸟氨酸拉乌尔菌的9株分离株携带bla(ORN)基因。基于bla基因的鉴定通过16S rDNA和rpoB测序得到证实。新鉴定为鸟氨酸拉乌尔菌的21株分离株鸟氨酸脱羧酶（ODC）试验呈阴性。分子实验表明，ODC阴性的鸟氨酸拉乌尔菌和植生拉乌尔菌菌株中ODC编码基因有一个拷贝，而ODC阳性的鸟氨酸拉乌尔菌菌株中有两个拷贝。对35个bla基因的分析使我们能够（i）确认两种拉乌尔菌物种的bla基因之间仅有94%的同一性，而rpoB基因的同一性>98%，16S rDNA基因的同一性>99%，以及（ii）开发并成功应用一种用于拉乌尔菌属鉴定的bla PCR RFLP检测方法。总体而言，本研究使我们发现了ODC阴性的鸟氨酸拉乌尔菌，并提供了一种可靠的拉乌尔菌鉴定方法，该方法广泛可用，因为不需要测序设备。
