Complete genetic characterization of carbapenem-resistant Acinetobacter johnsonii, co-producing NDM-1, OXA-58, and PER-1 in a patient source

Abstract

The emergence of carbapenemase-producing Acinetobacter spp. has been widely reported and become a global threat. However, carbapenem-resistant A. johnsonii strains are relatively rare and without comprehensive genetic structure analysis, especially for isolates collected from human specimen. Here, one A. johnsonii AYTCM strain, co-producing NDM-1, OXA-58, and PER-1 enzymes, was isolated from sputum in China in 2018. Antimicrobial susceptibility testing showed that it was resistant to meropenem, imipenem, ceftazidime, ciprofloxacin, and cefoperazone/sulbactam. Whole-genome sequencing and bioinformatic analysis revealed that it possessed 11 plasmids. bla _OXA-58 and bla _PER-1 genes were located in the pAYTCM-1 plasmid. Especially, a complex class 1 integron consisted of a 5' conserved segment (5' CS) and 3' CS, which was found to carry sul1, arr-3, qnrVC6, and bla _PER-1 cassettes. Moreover, the bla _NDM-1 gene was located in 41,087 conjugative plasmids and was quite stable even after 70 passages under antibiotics-free conditions. In addition, six prophage regions were identified. Tracking of closely related plasmids in the public database showed that pAYTCM-1 was similar to pXBB1-9, pOXA23_010062, pOXA58_010030, and pAcsw19-2 plasmids, which were collected from the strains of sewage in China. Concerning the pAYTCM-3 plasmids, results showed that strains were collected from different sources and their hosts were isolated from various countries, such as China, USA, Japan, Brazil, and Mexico, suggesting that a wide spread occurred all over the world. In conclusion, early surveillance is warranted to avoid the extensive spread of this high-risk clone in the healthcare setting.

Keywords: Acinetobacter johnsonii; NDM-1; OXA-58; PER-1; carbapenem resistance; integron.

Figure 1

RAST annotation of A. johnsonii AYTCM strain. The number of each subsystem category is shown on the right of column.

Figure 2

Circular map and genetic environment of the pAYTCM-1 plasmid. (A) Circular map of the pAYTCM-1 plasmid. Different filled boxes indicate various open reading frames (ORFs). The GC content and GC skew are shown in the inner rings. Resistance genes (red filled boxes), mobile genetic elements (yellow filled boxes), T4SS region (green filled boxes), and mercury resistance region (purple filled boxes). Light blue represents other ORFs. (B) Genetic environment of the bla _OXA-58 gene. The red filled arrow indicates the position of the bla _OXA-58 gene. Blue filled arrows indicate ISAjo2 and ISAjo2Δ. Purple filled arrow indicates ISAlw15. Arrows’ directions indicate the ORF directions. 9-bp target site duplications (TSD) are shown upstream and downstream of ISAjo2 and ISAjo2Δ using white or red filled circles, respectively. (C) Structure of the class 1 integron containing bla _PER-1. intl1 is shown as a black filled box. attl is shown as a white filled box. The 5′ conserved segment (5′ CS) and 3′ CS of class 1 integron are labeled. The various kinds of resistance genes were shown as different colors with the names labeled above with the orientation indicated by thin black arrows.

Figure 3

Plasmid comparison with pXBB1-9 using Circoletto. Ribbons represent the alignments produced by BLAST, their width the alignment length, and the colors the alignment bitscore in four quartiles: blue for the first 25% of the maximum bitscore, green for the next 25%, orange for the third, and finally red for the top bitscores of between 75% and 100%.

Figure 4

Circular map of pAYTCM-3. Different filled boxes indicate various open reading frames (ORFs). The GC content and GC skew are shown in the inner rings. Resistance genes are shown as a red filled box. Mobile genetic elements are shown as a yellow filled box. The T4SS region and T4CP are indicated as a green filled box. The relaxase-encoding gene is shown in the purple filled box. Light blue represents other ORFs. Moreover, the position of oriT was also labeled.

Figure 5

Predicted prophage regions within the A johnsonii AYTCM chromosome. (A) Six prophage regions positions in the chromosome. Green filled boxes mean the intact prophage regions (score > 90), filled boxes mean the questionable prophage regions (score 70–90), filled boxes mean the incomplete prophage regions (score < 70). (B) Structure of two intact and two questionable prophage regions. Genes are colored based on predicted functions.