Ze-Hua Cui1,2, Wei-Na Ni1,2, Tian Tang1,2, Bing He1,2, Zi-Xing Zhong1,2, Liang-Xing Fang1,2, Liang Chen3, Chong Chen1,2, Chao-Yue Cui1,2, Ya-Hong Liu1,2, Xiao-Ping Liao1,2, Jian Sun1,2. 1. National Risk Assessment Laboratory for Antimicrobial Resistance of Animal Original Bacteria, South China Agricultural University, Guangzhou, China. 2. Laboratory of Veterinary Pharmacology, College of Veterinary Medicine, South China Agricultural University, Guangzhou 510642, P. R. China. 3. Hackensack-Meridian Health Center for Discovery and Innovation, Nutley, NJ, USA.
Abstract
OBJECTIVES: The emergence and spread of plasmid-encoded tet(X3/X4) genes that confer high-level tigecycline and eravacycline resistance in Escherichia coli and Acinetobacter spp. pose serious threats to human and animal health. We developed a rapid and robust assay to detect Tet(X3/X4) in Gram-negative bacteria based on eravacycline degradation by the presence of the Tet(X) enzyme in the test strain. METHODS: This tetracycline inactivation method (TIM) is based on the degradation of eravacycline by the Tet(X3/X4)-producing strain, which results in reduced eravacycline activity against an acid-producing thermophile Bacillus stearothermophilus indicator strain. For Tet(X)-negative strains, eravacycline retains its antimicrobial activity. Coupled with a pH-sensitive dye (bromocresol purple), the reduced colorimetric inhibition zone can be measured to determine the production of Tet(X3/X4). One hundred and eighteen isolates, including 30 tet(X4)-positive E. coli, 30 tet(X3)-positive Acinetobacter spp. and 58 tet(X)-negative E. coli and Acinetobacter spp., were examined to evaluate the performance of this TIM. RESULTS: The sensitivity and specificity for E. coli carrying tet(X4) was 96.7% and 100%, respectively, and for Acinetobacter spp. carrying tet(X3) both were 100%. The TIM assay can be completed within 6.5 h. CONCLUSIONS: The TIM is a simple, rapid and cost-effective method for the detection of plasmid-mediated high-level tigecycline resistance in E. coli and Acinetobacter spp.
OBJECTIVES: The emergence and spread of plasmid-encoded tet(X3/X4) genes that confer high-level tigecycline and eravacycline resistance in Escherichia coli and Acinetobacter spp. pose serious threats to human and animal health. We developed a rapid and robust assay to detect Tet(X3/X4) in Gram-negative bacteria based on eravacycline degradation by the presence of the Tet(X) enzyme in the test strain. METHODS: This tetracycline inactivation method (TIM) is based on the degradation of eravacycline by the Tet(X3/X4)-producing strain, which results in reduced eravacycline activity against an acid-producing thermophile Bacillus stearothermophilus indicator strain. For Tet(X)-negative strains, eravacycline retains its antimicrobial activity. Coupled with a pH-sensitive dye (bromocresol purple), the reduced colorimetric inhibition zone can be measured to determine the production of Tet(X3/X4). One hundred and eighteen isolates, including 30 tet(X4)-positive E. coli, 30 tet(X3)-positive Acinetobacter spp. and 58 tet(X)-negative E. coli and Acinetobacter spp., were examined to evaluate the performance of this TIM. RESULTS: The sensitivity and specificity for E. coli carrying tet(X4) was 96.7% and 100%, respectively, and for Acinetobacter spp. carrying tet(X3) both were 100%. The TIM assay can be completed within 6.5 h. CONCLUSIONS: The TIM is a simple, rapid and cost-effective method for the detection of plasmid-mediated high-level tigecycline resistance in E. coli and Acinetobacter spp.