Genes conferring carbapenem resistance have spread worldwide among gram-negative bacteria. Subtyping of these genes has epidemiological value due to the global cross-border movement of people. Subtyping of bla_IMP genes that frequently detected in Japan appears to be important in public health settings; however, there are few useful tools for this purpose. We developed a subtyping screening tool based on PCR direct sequencing, which targets the internal sequences of almost all bla_IMP genes. The tool used bipartite multiplex primers with M13 universal sequences at the 5’-end. According to in silico analysis, among the 78 known IMP-type genes, except for bla_IMP-81, 77 detected genes were estimated to be differentiated. In vitro evaluation indicated that sequences of amplicons of IMP-1, IMP-6, IMP-7, and IMP-20 templates were identical to their respective subtypes. Even if the amplicons were small or undetectable through the first PCR, sufficient amplicons for DNA sequencing were obtained through a second PCR using the M13 universal primers. In conclusion, our tool can be possibly used for subtype screening of bla_IMP, which is useful for the surveillance of bacteria with bla_IMP in clinical and public health settings or environmental fields.

赋予碳青霉烯抗性的基因已在全球革兰氏阴性菌中传播。由于人们的全球跨境流动，这些基因的分型具有流行病学价值。在日本经常检测到的bla _IMP基因的亚型分型似乎在公共卫生环境中很重要；然而，很少有用于此目的的有用工具。我们开发了一种基于 PCR 直接测序的亚型筛选工具，该工具针对几乎所有bla _IMP基因的内部序列。该工具使用在 5' 端具有 M13 通用序列的二分多重引物。根据计算机分析，在已知的 78 个 IMP 型基因中，除了bla _IMP-81，估计有 77 个检测到的基因被区分。体外评估表明，IMP-1、IMP-6、IMP-7 和 IMP-20 模板的扩增子序列与其各自的亚型相同。即使扩增子很小或无法通过第一次 PCR 检测到，也可以通过使用 M13 通用引物的第二次 PCR 获得足够的 DNA 测序扩增子。总之，我们的工具可能用于bla _IMP的亚型筛选，这对于在临床和公共卫生环境或环境领域中监测bla _IMP细菌很有用。