Watanabe Mako, Nakano Ryuichi, Yamamoto Keizo, Nakano Akiyo, Suzuki Yuki, Saito Kai, Nakashima Satoko, Endo Kentaro, Narita Kazuya, Yano Hisakazu
Department of Microbiology and Infectious Diseases, Nara Medical University, 840 Shijo-cho, Kashihara 6348521, Nara, Japan.
Department of Chemistry, Nara Medical University, 88 Shijo-cho, Kashihara 6340813, Nara, Japan.
Antibiotics (Basel). 2025 Aug 12;14(8):823. doi: 10.3390/antibiotics14080823.
The genus harbors class C (AmpC) and class A β-lactamases. produces an inducible AmpC β-lactamase controlled by the LysR-type transcriptional regulator AmpR and cytosolic amidase AmpD. produces the class A β-lactamase Sed-1, whose expression is believed to be regulated by the transcriptional regulator SedR and AmpD. NR2807, isolated in Japan, is resistant to third-generation cephalosporins and displays extended-spectrum β-lactamase characteristics. Here, we sought to understand the mechanism for successful resistance to third-generation cephalosporins by investigating the regulators controlling Sed-1 production. Plasmids containing and (pCR2807) or truncated (pCR2807ΔSedR) were constructed and introduced into . Antibiotic-resistant mutants of NR2807 were obtained, and enzyme kinetics were assessed. The AmpD mutant (pCR2807/ML4953) showed an 8-fold increase in cefotaxime MIC and an 8.46-fold increase in Sed-1 activity compared to the wild-type (pCR2807/ML4947). However, induction of pCR2807/ML4947 also led to a 1.32-fold higher Sed-1 activity, indicating semi-inducibility. Deletion of (pCR2807ΔSedR/ML4947) led to a 4-fold decrease in cefotaxime MIC and 1.93-fold lower Sed-1 activity, confirming SedR as an activator. While wild-type ATCC51115 is susceptible to third-generation cephalosporins, the AmpD mutation in NR2807 led to Sed-1 overproduction and resistance to this class of antibiotics. Finally, mutagenesis revealed that amino acid substitution in Sed-1 conferred resistance to ceftazidime and extended-spectrum β-lactamase characteristics. Sed-1 producers, though usually susceptible to third-generation cephalosporins, may develop extended-spectrum β-lactamase traits due to AmpD or Sed-1 mutations, thereby requiring careful monitoring.
该菌属含有C类（AmpC）和A类β-内酰胺酶。产生一种由LysR型转录调节因子AmpR和胞质酰胺酶AmpD控制的可诱导型AmpCβ-内酰胺酶。产生A类β-内酰胺酶Sed-1，其表达据信受转录调节因子SedR和AmpD调控。在日本分离得到的NR2807对第三代头孢菌素耐药，并表现出超广谱β-内酰胺酶特性。在此，我们通过研究控制Sed-1产生的调节因子，试图了解其对第三代头孢菌素成功耐药的机制。构建了含有sedR和ampD（pCR2807）或截短的sedR（pCR2807ΔSedR）的质粒，并将其导入NR2807。获得了NR2807的抗生素耐药突变体，并评估了酶动力学。与野生型（pCR2807/ML4947）相比，AmpD突变体（pCR2807/ML4953）的头孢噻肟MIC增加了8倍，Sed-1活性增加了8.46倍。然而，pCR2807/ML4947的诱导也导致Sed-1活性高出1.32倍，表明具有半诱导性。sedR的缺失（pCR2807ΔSedR/ML4947）导致头孢噻肟MIC降低4倍，Sed-1活性降低1.93倍，证实SedR为激活剂。虽然野生型NR2807 ATCC51115对第三代头孢菌素敏感，但NR2807中的AmpD突变导致Sed-1过量产生并对这类抗生素耐药。最后，诱变显示Sed-1中的氨基酸取代赋予了对头孢他啶的耐药性和超广谱β-内酰胺酶特性。产生Sed-1的菌株虽然通常对第三代头孢菌素敏感，但可能由于AmpD或Sed-1突变而产生超广谱β-内酰胺酶特性，因此需要仔细监测。
