Bacteremia caused by an Acinetobacter junii strain harboring class 1 integron and diverse DNA mobile elements

Introduction: Infections caused by Acinetobacter junii are rarely reported. However, some outbreaks of septicemia in neonates and pediatric oncology patients, as well as meningitis, peritonitis, and ocular infection have been described. Since it is highly infrequent to find the molecular characterization of A. junii strains in literature, in this study we described the molecular characterization of A. junii isolates recovered from blood samples of a renal transplant patient. Methodology: The case was defined as a catheter-related bacteremia caused by A. junii. The patient responded favorably after catheter removal and treatment with ciprofloxacin. Results and Conclusion: The complete molecular characterization of the isolate showed that it harbored a class 1 integron and diverse DNA mobile elements. This explains its genomic plasticity for acquiring antimicrobial resistance determinants and for adapting to a nosocomial niche.

In contrast to A. baumannii infections, the infections caused by A. junii are easy to treat because this bacterium is commonly susceptible to antimicrobial agents [5].Still, carbapenem-resistant A. junii producing OXA-type carbapenemases and IMP-4 have been described [8,9].The presence of plasmids containing antibiotic resistance genes, commonly found in A. baumannii, shows that horizontal genetic transfer may be possible between both species of Acinetobacter.By this genetic exchange the therapeutic options to treat A. junii infections might be limited.
Mobile elements and resistance determinants associated to antibiotic resistance have been well studied for A. baumannii [10][11][12], in contrast there are few reports of these elements in A. junii [13,14].
Genetically characterized A. junii strains is rarely described in literature.This study was undertaken to investigate whether A. junii can be a reservoir of mobile elements.In addition, we studied the occurrence of resistant determinants in A. junii which had previously been described in A. baumannii isolates from our lab, to investigate their possible intra-species transfer.

Bacterial Strain
Positive growth were obtained in two blood culture sets (taken at two different moments) and in the blood culture drawn through the catheter hub (BacT/Alert; bioMérieux, Marcy l'Etoile, France) from a 41 years old female patient.She was admitted to the hospital for pathological fracture of the left tibia secondary to brown tumor.She had a history of papillary thyroid carcinoma and renal transplant in 2006 treated with tacrolimus and corticosteroids since that date.
The isolates were analyzed using VITEK 2 Compact (bioMérieux, Marcy l'Etiole, France) system.Polymrase chain reaction (PCR) amplifications and sequence analysis of the 16S rRNA and rpoB genes were carried out to confirm the identification [15].Sequencing was performed on both DNA strands using ABIPrism 3100 BioAnalyzer equipment for sequencing (Macrogen Inc., Seoul, Republic of South Korea).

Antibiotic susceptibility
The antibiotic susceptibility test was performed using the VITEK 2 System (bioMerieux, Marcy, L'Etoile, France) employing the panel AST-082 (GNS susceptibility card).The minimum inhibitory concentration (MIC) results were interpreted using the Clinical and Laboratory Standards Institute (CLSI) categories.
All positive PCR amplification products were sequenced on both DNA strands, using an ABI Prism3100 BioAnalyzer equipment (Macrogen Inc., Seoul, Republic of South Korea) and nucleotide sequences were analyzed using the Blast v2.0 software (http://www.ncbi.nlm.nih.gov/BLAST/).
Due to the fact there is little information published on the molecular mechanisms and mobile elements associated to antimicrobial resistance in A. junii, we decided to study the resistance determinants and mobile elements based on their wide distribution in our country [22,19].
Among the PCR reactions carried out to detect resistance determinants, positive results were only obtained for sul1, sul2, strA, strB, aphA1 and aac(6')-Ib (Table 1), which explained the resistance found in the strains to gentamicin, amikacin and TMP-SMX.
The mobile elements found in the A. junii isolates were the insertion sequences IS26, ISAba1, ISAba125, IS1008 and ISCR2, which is in agreement with the high occurrence of insertion sequences in Acinetobacter spp.[12,13].Negative results were obtained to determine the presence not only of plasmids belonging to the incompatibility plasmids tested in this study, but also of transposons mostly present in our clinical isolates.
We observed the presence of one class 1 integron, whereas we obtained negative results for class 2 integron amplification, which is the most wide-spread class of integron in our A. baumannii strains in our region.The amplification of the variable region (vr-1) of the integron revealed the presence of the gene cassettes arr3-aac(6')-Ib, genes that codify for a rifampin ADP-ribosylating transferase and an aminoglycoside-(6')-N-acetyltransferase, respectively.
The association between the IS26 and the aphA1 gene, encoding for APH (3')-I aminoglycoside phosphotransferase enzyme and conferring kanamycin and neomycin resistance, is frequent in A. baumannii strains [23,24].Thus we decided to test the association of these two genes in our strain obtaining positive results for the amplification reactions carried out.This result showed that in A. junii, the aphA1 gene can also be linked with IS26 as it was described for A. baumannii.
Concerning the phenotype of these strains, the presence of the aac(6')-Ib gene, can explain the observed resistance to amikacin.However, no resistance to rifampicin was observed suggesting that the arr3 gene cassettes maybe weakly expressed or do not confer resistance to our strains.
The molecular characterization of the A. junii isolates showed the presence of different mobile genetic elements and determinants associated to horizontal gene transfer; these elements might also play an important role in the acquisition and development of antibiotic resistance in this species.
Infections caused by species of Acinetobacter other than A. baumannii have been reported in literature [25,15].The existence of molecular techniques that allow correct species identification give an important contribution to the epidemiology of non-baumannii Acinetobacter and to the knowledge on the real prevalence of these species and the type of infections associated with them.In addition, information regarding the antimicrobial resistance mechanisms and mobile genetics elements contained in these species can help to establish a more accurate treatment and stop their spreading.

Table 1 .
Antibiotic resistance determinants, mobile elements and integrons studied with the corresponding results obtained in the Acinetobacter junii strains.