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      Vancomycin Resistance due to vanA Gene Expression in an Aerococcus viridans Isolate: First Case in Korea

      brief-report
      , M.D. 1 , , M.S. 1 , , M.D. 1 , , , M.D. 1 , , M.D. 2
      Annals of Laboratory Medicine
      The Korean Society for Laboratory Medicine

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          Abstract

          Dear Editor, Aerococcus viridans is a catalase-negative gram-positive coccus that appears in clusters, tetrads, or irregular arrangements [1]. This organism is generally considered as a contaminant in clinical cultures, but is also infrequently reported as a clinically significant isolate that causes endocarditis, bacteremia, spondylodiscitis, and urinary tract infections [2 3 4 5]. Although most A. viridans strains were susceptible to penicillin and other commonly used antibiotics, Uh et al [6] described a case of bacteremia caused by an A. viridans strain showing high resistance to penicillin, erythromycin, clindamycin, and ceftriaxone. In 2014, Zhou et al [7] reported a peritoneal dialysis-related infection caused by vancomycin-resistant A. viridans harboring the vanA gene. We report a case of a vancomycin-resistant A. viridans isolate obtained from an excisional biopsy wound. As far as we know, no study regarding vancomycin-resistant A. viridans has yet been published in Korea. A 77-yr-old farmer visited the emergency room complaining of severe chills. Swelling in an external wound was observed on the inguinal areas where previous excisional biopsy was performed. He had recently been diagnosed as having colon adenocarcinoma and angioimmunoblastic T-cell lymphoma. On admission, his body temperature was 39.0℃. Hematological investigation revealed a hemoglobin level of 9.2 g/dL, white blood cell (WBC) count of 18.96×109/L (segmented neutrophils; 92.2%), and platelet count of 262×109/L. Serum C-reactive protein (CRP) level (14.36 mg/dL, reference range: <0.30 mg/dL) was elevated. Two aerobic and anaerobic blood culture sets were incubated in the BacT/Alert 3D system (bioMérieux, Durham, NC, USA). The mucus-like whitish aspirate from the wound was plated onto 5% sheep blood agar (BD Diagnostic Systems, Sparks, MD, USA), MacConkey agar (BD Diagnostic Systems), and fluid thioglycollate medium (BD Diagnostic Systems). Some colonies grew on the 5% sheep blood agar after 24 hr aerobic incubation at 35℃; these colonies were identified as oxacillin-resistant Staphylococcus hemolyticus by VITEK 2 (bioMérieux, Marcy l'Etoile, France). No growth was detected in the blood cultures after five days of incubation. Thirteen days after hospital admission, chemotherapy for lymphoma was initiated. Although WBC count and CRP level were within reference ranges at that time, mucus-like discharge from the wound remained, and CRP started to rise steadily. A. viridans and S. hemolyticus were repeatedly simultaneously isolated from wound cultures. The identification probability of A. viridans by VITEK 2 was 98%. Antimicrobial susceptibility test by a MicroScan MICroSTREP plus panel (Beckman Coulter, Brea, CA, USA) showed that A. viridans was susceptible to tetracycline, but resistant to vancomycin, penicillin, cefotaxime, ceftriaxone, sulfamethoxazole/trimethoprim, meropenem, and levofloxacin. To confirm the species identification and antimicrobial susceptibilities, 16S rRNA sequence analysis, minimal inhibitory concentration (MIC) determination for vancomycin (Daewoong Lilly, Seoul, Korea) and teicoplanin (Narion Merrell Dow, Seoul, Korea), and PCR to detect vanA and vanB genes were performed. MICs were performed by using the broth microdilution method according to CLSI guidelines [8]. PCR amplification of 16S rRNA was performed by using primers 16SF (5′-TAA YAC ATG CAA GTC GAR CG-3′) and 608R (5′-TAT TAC CGC GGC TGC TGG CA-3′), and sequencing was conducted by using the Big Dye Terminator Cycle Sequencing kit (Applied Biosystems, Foster City, CA, USA) and an ABI PRISM 3730 genetic analyzer (Applied Biosystems). All sequences were analyzed by using the basic local alignment search tool and ribosomal database project. The 450-bp 16S rRNA gene sequence from our isolate showed 100% similarity to and 99% query coverage with several A. viridans strains (GenBank accession no. KR140225.1, LN998006.1, EU169542.1). The primers and PCR procedure for amplification of vanA and vanB genes were previously described [9]. Each DNA product was tested by using gel electrophoresis, where vanA gene was conclusively detected. A. viridans showed a high degree of resistance to vancomycin (<128 µg/mL) and teicoplanin (64 µg/mL), indicating potential acquisition of the vanA gene. The physician added sulfamethoxazole/trimethoprim to teicoplanin and meropenem combination therapy. Consequently, exudate lessened, and WBC count and CRP level also decreased (Fig. 1). It is difficult to distinguish simple colonization from real infection when A. viridans is identified in a wound specimen. However, we paid attention to our A. viridans isolate because this patient struggled with chemotherapy before A. viridans was isolated and because the resistance may spread to other gram-positive cocci through transfer of vanA from A. viridans [10].

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          Most cited references10

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          VanA-type vancomycin-resistant Staphylococcus aureus.

          Since 2002, nine methicillin (meticillin)-resistant Staphylococcus aureus (MRSA) strains that are also resistant to vancomycin (VRSA) have been reported in the United States, including seven clinical isolates from Michigan. The strains harbor a plasmid-borne Tn1546 element following conjugation from a glycopeptide-resistant Enterococcus strain. In the second step, Tn1546 transposed to a resident plasmid in five strains; the acquired plasmid behaved as a suicide gene delivery vector, and the incoming DNA had been rescued by illegitimate recombination. Surprisingly, combination of a glycopeptide with a beta-lactam has a strong synergistic effect against VRSA, both in vitro and in an animal model, despite resistance of the strains to both drug classes when administered separately. This results from the fact that the late peptidoglycan precursors ending in D-alanine-D-lactate (D-Ala-D-Lac) that are mainly synthesized in the presence of glycopeptide inducers are not substrates for PBP2', which is the only transpeptidase that remains active in the presence of oxacillin. One VRSA strain is partially dependent on vancomycin for growth due to a mutation in the host D-Ala:D-Ala ligase, thus having to rely on the inducible resistance pathway for cell wall synthesis. Competition growth experiments in the absence of inducer between the MRSA recipient and isogenic VRSA transconjugant revealed a disadvantage for the transconjugant, accounting, in part, for the low level of dissemination of the VRSA clinical isolates. The association of multiple molecular and environmental factors has been implicated in the regional emergence of VRSA in Michigan.
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            Aerococci and aerococcal infections.

            Aerococcus is a genus that comprises seven species, of which Aerococcus urinae, and Aerococcus sanguinicola are emerging human pathogens. Aerococci are gram positive cocci that are easily misidentified as streptococci or staphylococci, and thus the incidence of aerococcal infections has been underestimated. With the introduction of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) clinical microbiologists now have access to a rapid and accurate method to identify aerococci. A. urinae and A. sanguinicola are isolated in a small proportion of urinary specimens in many laboratories and many patients with bacteriuria with aerococci have symptoms of urinary tract infection (UTI). A. urinae, and also A. sanguinicola, cause invasive infections including infective endocarditis (IE) with many reported fatalities. Especially older men with urinary tract abnormalities are at risk for bacteraemia with A. urinae but the prognosis of bacteraemia without IE is favourable. Penicillin is appropriate for treatment of invasive infections and in IE, addition of an aminoglycoside should be considered. Treatment of UTI with aerococci is complicated by uncertainty about the effect of trimethoprim-sulphametoxazole and fluoroquinolones on aerococci. This review will discuss identification of Aerococcus spp., antibiotic resistance, the clinical presentation and management of aerococcal infections as well as the virulence mechanisms of these bacteria.
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              Urinary tract infection caused by Aerococcus viridans, a case report.

              Aerococcus viridans organisms are gram-positive, usually airborne cocci that are widely distributed in hospital environments. These bacteria have infrequently been encountered as a human pathogen causing bacteremia, endocarditis and urinary tract infections. The clinical significance of these bacteria may be overlooked due to their fastidious growth and often confused with other strains of streptococci. We report a case of urinary tract infection with bacteremia caused by A. viridans in an 87 year-old male nursing home resident. The patient presented with a fever of 103 degrees F, dysuria, hematuria and weakness for three days. Urinalysis showed large amount of blood, more than one hundred white cells/HPF and 4+ bacteria. Laboratory tests revealed, white blood cell count of 2300/cu mm (neutrophils 80%, bands 7%, lymphocytes 11% and monocytes 2%), hemoglobin 15.4 gm/dL, blood urea nitrogen 23 mg/dL and creatinine 1.2 mg/dL. Urine culture yielded growth of 10(5) CFU of A. viridans. The patient was treated for ten days with levofloxacin (both IV + PO). To date, no clinical case report of this nature has been described implicating A. viridans in urinary tract infections. Increased awareness and more studies of this genus should lead to the identification of their potential role in human infections.
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                Author and article information

                Journal
                Ann Lab Med
                Ann Lab Med
                ALM
                Annals of Laboratory Medicine
                The Korean Society for Laboratory Medicine
                2234-3806
                2234-3814
                May 2017
                17 February 2017
                : 37
                : 3
                : 288-289
                Affiliations
                [1 ]Department of Laboratory Medicine, Yonsei University Wonju College of Medicine, Wonju, Korea.
                [2 ]Department of Internal Medicine, Yonsei University Wonju College of Medicine, Wonju, Korea.
                Author notes
                Corresponding author: Young Uh. Department of Laboratory Medicine, Yonsei University Wonju College of Medicine, 20 Ilsan-ro, Wonju 26426, Korea. Tel: +82-33-741-1592, Fax: +82-33-731-0506, u931018@ 123456yonsei.ac.kr
                Article
                10.3343/alm.2017.37.3.288
                5339105
                28224779
                657588a5-891a-4bdc-8865-daa8af39cace
                © The Korean Society for Laboratory Medicine

                This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License ( http://creativecommons.org/licenses/by-nc/4.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

                History
                : 04 October 2016
                : 11 October 2016
                : 29 December 2016
                Categories
                Letter to the Editor
                Clinical Microbiology

                Clinical chemistry
                Clinical chemistry

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