First report of vaginal infection caused by Enterococcus raffinosus

Case report

During the second month of hospitalization by the Department of Haematology, a 50-year-old immunocompromised patient with acute leukaemia developed severe vulvovaginal itching and burning. At speculum examination, erythema and discharge were observed. Vaginal pH was 5.2, the amine test was negative and microscopy (Gram stain) showed the presence of numerous Gram-positive cocci and leukocytes. We obtained swab specimens from the middle third of the vagina and placed them on Sabouraud dextrose agar, sheep blood agar, MacConkey agar and Mannitol salt agar (all from Biolife). The plates were incubated in air. A second blood agar plate was inoculated and incubated anaerobically, whilst a third blood agar plate and a Thayer-Martin plate were inoculated and incubated in an atmosphere of 5 % CO₂. All cultures were kept at 37 °C and examined after 24 and 48 h. After 24 h of incubation, smooth and α-haemolytic colonies (>150 c.f.u.), about 1 mm in diameter, were observed on all blood agar plates. The organism was found to be a Gram-positive coccus that formed pairs and short chains, and non-motile at 36 °C. None of Lancefield group A, B, C, D, F and G antisera produced a positive reaction (SLIDEX Strepto Plus; bioMérieux) (Facklam & Collins, 1989) and the catalase test was negative. No other organisms of known vaginal pathogenicity were isolated. The isolate was identified as Enterococcus raffinosus by using the VITEK 2 system (card GP; bioMérieux) with 99 % certainty, and the identification was confirmed by using the API system (Rapid ID 32 Strep; bioMérieux). Antibiotic susceptibility testing was performed with brain heart infusion agar (Biolife) using the disc diffusion method and was interpreted as per Clinical and Laboratory Standards Institute guidelines (Chirurgi et al., 1991; NCCLS, 2000; Prakash et al., 2005). The organism was found to be resistant to penicillin, ampicillin, amoxicillin/clavulanate, ampicillin/sulbactam, piperacillin/tazobactam, imipenem, meropenem, ciprofloxacin, clindamycin, erythromycin, cotrimoxazole, tetracycline, amikacin, gentamicin, netilmicin and tobramicin, and susceptible only to vancomycin, teicoplanin and rifampicin. Discs were provided by Oxoid.

Pending the results from cultures, the patient was treated empirically and unsuccessfully with oral amoxicillin/clavulanate (1000 mg, every 12 h). Instead, the vaginal symptoms disappeared suddenly within the second day of teicoplanin therapy that the patient had received empirically because of the outbreak of a Gram-positive cocci bacteraemia (data not shown). One week after glycopeptide treatment, vaginal examination and cultures were repeated but they did not show signs of inflammation or E. raffinosus colonies, respectively. Further, Gram stain showed the restoration of a physiological vaginal environment.

Discussion

Enterococcus species are one of the main causes of nosocomial infections. Although Enterococcus faecalis and Enterococcus faecium represent 90 % of clinical isolates, the incidence of enterococcal species other than E. faecalis and E. faecium is increasing (Prakash et al., 2005). In particular, uncommonly E. raffinosus has been isolated from hospitalized patients. E. raffinosus is a facultative, motile or non-motile, non-encapsulated, non-sporulating organism. The cells are Gram-positive and are arranged singly, in pairs or short chains. On blood agar with 5 % sheep blood, colonies are smooth, grey, α-haemolytic or non-haemolytic and ≤1 mm in diameter. This species is characterized by the presence of pyrrolidonylarylmidase and leucine aminopeptidase activities, hydrolysis of aesculin and utilization of mannitol, sorbitol, raffinose, sucrose and l-arabinose, and the absence of catalase activity. Further, E. raffinosus occasionally presents Lancefield Group D antigen (Facklam & Collins, 1989; Freyaldenhoven et al., 2005; Sandoe et al., 2001).

The species E. raffinosus was recognized in 1989, when the bacterium was distinguished from the phenotypically similar Enterococcus avium by the ability of the former to utilize raffinose. Misidentification is possible by using API 20 Strep and API 32 Strep (bioMérieux), as E. raffinosus is not included in the database for these products. Instead, VITEK 2, Rapid ID 32 Strep and the BD Phoenix system include this species in the Gram-positive identification database (Freyaldenhoven et al., 2005; Grayson et al., 1991; Sandoe et al., 2001).

E. raffinosus is associated with urinary tract infections, wounds, ulcers and plagues infections, intra-abdominal and inguinal abscesses, Bartolin gland abscesses, biliary and peritoneal infections, vertebral osteomyelitis, endocarditis and bacteraemia (Chirurgi et al., 1991; Sandoe et al., 2001). A case of haematoma infection has been reported recently (Freyaldenhoven et al., 2005). Isolation of E. raffinosus is usually related to long-term hospitalization, antibiotic prophylaxis and therapies, urinary catheterization and surgical procedures (Chirurgi et al., 1991; Prakash et al., 2005; Sandoe et al., 2001). It is difficult to provide a standard susceptibility pattern for E. raffinosus, and its emerging multidrug resistance is increasing in importance (Gordon et al., 1992), as resistance to penicillin, aminoglycosides, fluoroquinolones, carbapenems and glycopeptides has been described in the literature. In particular, resistance to penicillin is mediated mostly by mutations in penicillin-binding proteins. Further, gentamicin resistance is often associated with resistance to erythromycin, tetracycline and minocycline (Chirurgi et al., 1991; Grayson et al., 1991; Sapico et al., 1989; Straut et al., 1996; Tanimoto et al., 2006; Wilke et al., 1997).

To date, little has been published concerning E. raffinosus. In the case described here, the patient had received parenteral piperacillin–tazobactam, imipenem and vancomycin during the first month of admission to hospital, due to the onset of bacteraemia caused by Escherichia coli, and of a catheter-related Staphylococcus epidermidis bloodstream infection (data not shown). Other factors or underlying diseases able to increase the risk of acquiring such an enterococcal infection were not present. E. raffinosus is known to only inhabit the oropharynx of domestic cats, as part of the commensal flora; nothing else is known about its natural habitat (Sandoe et al., 2001). It is not known whether the patient in this case kept any pets.

To our knowledge, this report describes the first case of vaginitis caused by E. raffinosus, confirming the role of this organism as a cause of nosocomial infections. Further, it focuses on the emerging occurrence of antimicrobial resistance in this uncommon species. Hence, the authors re-emphasize the importance of the implementation of infection control measures, in order to limit the nosocomial spread of unusual and atypical bacteria, and the emergence of multidrug resistance among them.