Emergence of OXA-72-producing Acinetobacter baumannii Belonging to High-Risk Clones (CC15 and CC79) in Different Brazilian States

To the Editor—Carbapenem resistance limits treatment options and causes major therapeutic problems; it has been continuously reported worldwide among Acinetobacter baumannii isolates. Carbapenem resistance in A. baumannii is frequently associated with Ambler class D carbapenemase, mainly blaOXA-23. Until now, there have been only a few reports of other oxacilinases, such as blaOXA-72, in Brazil. ¹ Multilocus sequence typing (MLST) seems to be a reliable tool for investigating population structure and global A. baumannii epidemiology. In Brazil, most carbapenem-resistant blaOXA-23–producing A. baumannii have been associated with clonal complexes CC79 and CC15. ² To the best of our knowledge, our report here is the first report of the epidemic clonal complex CC15 associated with A. baumannii carrying blaOXA-72. Furthermore, we describe the spread of A. baumannii high-risk clones (CC79 and CC15) carrying blaOXA-72 gene in 3 Brazilian states.

As part of a surveillance study performed by our research group (Laboratório de Pesquisa em Resistência Bacteriana – LABRESIS), we evaluated 94 A. baumannii-calcoaceticus isolates from 4 Brazilian states (São Paulo, Rio de Janeiro, Rio Grande do Sul, and Paraná) identified between April and October 2013. The isolates were screened for oxacilinases genes (ie, OXA-23, OXA-24/40, OXA-51, OXA-58, and OXA-143) by polymerase chain reaction (PCR). Class 1 and 2 integrons were detected by PCR for the integrase gene. ³ Among these 94 isolates from 3 states (São Paulo, Paraná and Rio Grande do Sul), 11 isolates (11.7%) presented positive results for the blaOXA-24/40, which was identified by sequencing as the variant blaOXA-72 (ABI 3500 Genetic Analyzer). Species identification was performed using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF) and a gyrB multiplex PCR, and all blaOXA-72–producing isolates were identified as A. baumannii.

Clonal diversity of the A. baumannii OXA-72 producers was investigated using repetitive-sequence-based PCR (REP-PCR). ⁴ Results were interpreted according to a dendrogram constructed using BioNumerics v6.5 software (bioMèrieux, Marcy-l'Étoile, France). REP-PCR analysis showed the presence of 3 clonal groups (A, B, and C; Table 1).

TABLE 1 Molecular Typing Analysis of blaOXA-72–Producing A. baumannii in Different Brazilian States

NOTE. REP-PCR, repetitive-sequence–based polymerase chain reaction; CC, clonal complex; ST, sequence type; MLST, multilocus sequence typing; Int, integron.

^a Sequence type by Institute Pasteur MLST scheme.

^b MLST clonal complex defined by Institute Pasteur.

The OXA-72 A. baumannii producers were typed using the MLST scheme from the Pasteur Institute (http://www.pasteur.fr). Analyses of the allele sequences and sequence type (ST) were performed via the A. baumannii MLST Institute Pasteur website (http://pubmlst.org/abaumannii/). The relationships among the new and existing STs were evaluated using the eBURST program to determinate the clonal relation of A. baumannii carrying blaOXA-72 among different states. MLST demonstrated that the isolates belonged to 5 different sequence types: ST79, ST180, ST730, and the new ST890 and ST891 (Table 1). According to the MLST data analysis and the eBURST algorithm, the isolates belonged to the epidemic clonal complexes CC15 (ST180 and ST890) and CC79 (ST79 and ST730), and the ST891 was a singleton. CC15 was restricted to the state of Paraná, and CC79 was present in Rio Grande do Sul and São Paulo states. Class 1 integrons were detected in all isolates belonging to CC15, and all isolates of CC79 presented class 2 integrons (Table 1).

Acinetobacter baumannii is characterized by a remarkable capability to acquire antibiotic resistance determinants, intra- and interhospital outbreaks, and national and international clonal dissemination. Worldwide dissemination of multidrug-resistant A. baumannii OXA-23–producers has been associated with specific clones, such as international clone 1 (CC1^IP), international clone 2 (CC2^IP), and CC15^IP. ⁵ In Brazil, the extensive dissemination of CC79 and CC15 have been associated with blaOXA-23 production in different states. ^{1 , 2} Furthermore, Stietz et al ⁶ reported that ST79 is also frequently observed among OXA-23–producing A. baumannii isolates in Argentina, which demonstrates the dispersal of this CC79 not only in Brazil but also in South America countries. ⁶ The same study demonstrated the spread of blaOXA-72 related to the epidemic clone CC79. ⁶

Acinetobacter baumannii belonging to CC15 have had evolutionary success, and they usually exhibit multidrug-resistant phenotypes, which has facilitated their rapid clonal expansion in recent years. This clonal complex has been related to OXA-23 outbreaks in several European countries, including Italy, Spain, Greece, and Turkey, as well as an outbreak in South America. ⁵ To the best of our knowledge, this is the first description of CC15 associated with OXA-72 production reported in the literature.

The screening for integrase genes demonstrated the presence of class 1 and 2 integrons associated with CC15 and CC79, respectively (Table 1). Interestingly, Martins et al ⁷ showed the same distribution of class 1 and 2 integrons, related to specific CCs such as CC15 and CC79. ⁷ In a previous study, we reported a high prevalence of class 2 integrons in southern Brazil. Indeed, these results might be explained by the association of class 2 integrons to CC79, which are prevalent in South America. ⁸

The OXA-72 enzyme was first identified in A. baumannii from Thailand in 2004. Later, this enzyme was reported in Acinetobacter spp. clinical isolates from China, South Korea, Taiwan, Italy, Spain, and France. ⁹ Until now, there have only been a few reports of A. baumannii carrying the blaOXA-72 gene in Brazil. ¹ In the present study, we have described the association of A. baumannii carrying blaOXA-72 with the epidemic clones CC15, CC79, and a new ST (ST 891). Camargo et al ² described blaOXA-72 producers of CC79 in Brazil. In contrast, the data presented in our study have demonstrated not only the association of blaOXA-72 producers to CC79 but also to CC15. The ability of these particular clones to acquire antibiotic resistance should trigger efforts to prevent the spread of OXA-72–producing isolates as occurred with blaOXA-23. ¹⁰ These data indicate the potential for this gene to spread to different countries and distinct geographical regions.