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 Table of Contents  
ORIGINAL ARTICLE
Year : 2020  |  Volume : 15  |  Issue : 3  |  Page : 421-425

Vancomycin-resistant enterococci causing bacteriuria in hospitalized patients from Northwest India


Department of Microbiology, JNUIMSRC, Jaipur, Rajasthan, India

Date of Submission20-Mar-2020
Date of Decision30-Apr-2020
Date of Acceptance25-May-2020
Date of Web Publication1-Feb-2021

Correspondence Address:
Dr. S N Tayade
Department of Microbiology, JNUIMSRC, Jaipur, Rajasthan
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/jdmimsu.jdmimsu_69_20

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  Abstract 


Background: Vancomycin-resistant enterococci (VRE) are an important opportunistic pathogen in hospitalized patients due to their resistance to commonly administered antimicrobial agents. Material and Methods: A pilot study consisting of 100 urinary enterococcal isolates with significant bacteriuria (>105 CFU/ml) was conducted to assess the resistance pattern in our hospital. Culture, identification, and antibiotic sensitivity testing were performed as per standard protocols. Resistance to vancomycin was confirmed by minimum inhibitory concentration method using commercially available E strips (Hi Media). Results: Fifty-four percent (54/100) of isolates were Enterococcus faecalis, 33% (33/100) Enterococcus faecium followed by 13% (13/100) nonfaecalis nonfaecium species. 4% (4/100) of enterococci were vancomycin resistant out of which 1 (25%) was resistant to linezolid. All VRE were sensitive to nitrofurantoin. Conclusion: Emerging resistance to vancomycin in cases of significant bacteriuria found in our study raises a concern for reserving higher antibiotics such as linezolid, daptomycin, and quinupristin-dalfopristin. Efforts are required to assess the clinical significance of VRE in terms of colonization and infection and switch to evidence-based treatment plan.

Keywords: Bacteriuria, colonization, multidrug resistance, vancomycin-resistant enterococci


How to cite this article:
Gupta S, Srivastava P, Yadav S, Tayade S N. Vancomycin-resistant enterococci causing bacteriuria in hospitalized patients from Northwest India. J Datta Meghe Inst Med Sci Univ 2020;15:421-5

How to cite this URL:
Gupta S, Srivastava P, Yadav S, Tayade S N. Vancomycin-resistant enterococci causing bacteriuria in hospitalized patients from Northwest India. J Datta Meghe Inst Med Sci Univ [serial online] 2020 [cited 2021 Mar 4];15:421-5. Available from: http://www.journaldmims.com/text.asp?2020/15/3/421/308573




  Introduction Top


Enterococci are isolated from cases of urinary tract infections (UTIs), septicemia, endocarditis, peritonitis, meningitis, and simple wound infections in most of the health-care settings. UTI by enterococci is particularly caused in catheterized, intensive care unit (ICU), and other immunocompromised patients. Many studies from India have reported Enterococci to be the common cause of nosocomial UTI after  Escherichia More Details coli and Klebsiella spps.[1],[2] Currently, more than 54 enterococcal species are known but Enterococcus faecalis and Enterococcus faecium have been predominantly isolated from clinical specimen.[3],[4],[5] Recent studies have reported an increased prevalence of rare species such as Enterococcus avium, Enterococcus durans, Enterococcus dispar, Enterococcus casseliflavus, Enterococcus gallinarum, Enterococcus raffinosus, and Enterococcus malodoratus.[5],[6],[7]

This organism has drawn attention due to its intrinsic resistance to antimicrobials such as beta lactams, low concentrations of aminoglycosides, clindamycin, and fluoroquinolones.[8] Studies have also reported acquired resistance to the high concentrations of beta lactams and aminoglycosides, glycopeptides, tetracyclines, erythromycin, fluoroquinolones, etc.[9] Enterococci have been able to thrive in hospital environment primarily due to its intrinsic resistance and second, ability to acquire resistance by mutation and receipt of foreign material by plasmids and transposons.[10] Thus, multidrug resistance gets introduced in the genome of hospitalized patients receiving multiple courses of antimicrobials for the treatment of UTI or other indications due to antibiotic selection pressure.

Available data from the United States and India suggest that E. faecium exhibit a higher degree of resistance to vancomycin (10% to >90%) followed by E. faecalis among all other enterococcal species.[3],[4],[11] CDC reported 75.6% E. faecium to be vancomycin resistant.[12] Studies have also demonstrated low level vancomycin resistance among other species such as E. gallinarum,[6] Enterococcus munditii, E. raffinosus, E. durans, E. avium, and Enterococcus hirae.[13]

Although Clinical and Laboratory Standards Institute (CLSI) has recommended a breakpoint of =4 mg/L (susceptible dose dependent) with dose 8–12 mg/kg Q24H of daptomycin for the treatment of vancomycin-resistant enterococci (VRE) infections, it has not been approved by the US Food and Drug Administration till date.[14] There is a dearth of clinical data for management and duration of therapy in UTI's harboring VRE with reference to hospital strains circulating in Indian subcontinent. Hence, differentiation of enterococci from Group D streptococci and identification up to the species level is essential. Species-wise laboratory data on vancomycin resistance in enterococci are needed in addition to clinical examination to decide duration of treatment in VRE in UTIs. We have undertaken this pilot study to assess the resistance pattern of various enterococcal species especially vancomycin from cases of significant bacteriuria (=105 CFU/ml) in hospitalized patients.


  Material and Methods Top


One hundred urine samples from patients attending various outpatient and inpatient departments of our hospital over 1 year from October 2017 to September 2018 were included in this prospective study. The study was approved by institutional ethics committee and informed consent was obtained from all patients. A colony count of =105 CFU/ml was significant and indicative of UTI. All patients with colony count <105 CFU/ml of urine specimen representing colonization and outpatients were excluded from the study.

The strains were identified and speciated according to standard laboratory procedures as per the scheme of Facklam and Collins.[15] Antimicrobial susceptibility was determined by Kirby–Bauer disc diffusion method using antibiotics such as ampicillin (10 μg), erythromycin (15 μg), doxycycline (30 μg), ciprofloxacin (5 μg), levofloxacin (5 μg), nitrofurantoin (300 μg), norfloxacin (10 μg), fosfomycin (200 μg), vancomycin (30 μg), teicoplanin (30 μg), and linezolid (15 μg) (HiMedia Laboratories, Mumbai, Maharashtra, India) as per recommendations of CLSI.[14] Resistance to vancomycin was confirmed by the determination of minimum inhibitory concentration (MIC) by E-strip test. (EM 060, Vancomycin Ezy MICTM Strip [0.016–256 mcg/ml] Hi media). Enterococci which had MIC >32 mcg/ml were considered resistant; 8–16 mcg/ml as intermediately resistant; and MIC of 4 mcg/ml as susceptible to vancomycin as per CLSI.[14]

Statistical analysis

Data were analyzed using? IBM SPSS version 26 (SPSS Inc., Chicago, Ill., USA). The analysis of variance was used to compare the resistance pattern between three or more species of enterococci. Pearson's coefficient (P value) <0.05 was considered as statistically significant.

Ethical clearance

The Institutional Ethics Committee of JNUIMSRC has approved the Research work proposed to be carried out at JNUIMSRC, Jaipur, Rajasthan Date: 2nd March, 2017 with Reference no JNUIMSRC/EC/2017/202.


  Results Top


Maximum number of patients (36% [36/100]) were between 41 and 60 years of age out of which 52.77% (19/36) were males. Fifty-four percent (54/100) of patients were from medicine, 22% (22/100) general surgery, 13% (13/100) obstetrics and gynecology, 6% (06/100) pediatrics, and 5% (5/100) chest and tuberculosis departments, respectively. The most common enterococcal isolate was E. faecalis (54/100 [54%]), followed by E. faecium (33/100 [33%]), E. avium (5/100 [5%]), E. hirae (4/100 [4%]), E. durans (2/100 [2%]), and E. raffinosus (2/100 [2%]) [Table 1].
Table 1: Age-wise distribution of various enterococcal isolates from urine

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E. faecium showed significantly higher resistance to vancomycin (P = 0.007; P < 0.01) and teicoplanin (P = 0.001; P < 0.01) as compared to E. faecalis. All isolates of E. avium and E. hirae were resistant ciprofloxacin, levofloxacin, and nitrofurantoin [Table 2]. Four isolates of E. faecium were resistant to vancomycin both by Kirby–Bauer disk diffusion and E-strip method (MIC > 256 mcg/ml) [Figure 1].
Table 2: Percentage distribution of antibiotic resistance pattern among various enterococcal isolates from urine

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Figure 1: Minimum inhibitory concentration by E strip test for strain of vancomycin-resistant enterococci

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All isolates of VRE were found to be resistant to ampicillin, erythromycin, ciprofloxacin, levofloxacin, teicoplanin, norfloxacin, fosfomycin 50% (2/4) resistant to doxycycline, and 25% (1/4) resistant to linezolid. Comparative antibiogram of vancomycin sensitive and vancomycin-resistant isolates shows that VRE were significantly more resistant to ampicillin and teicoplanin (P < 0.01). All strains showed higher resistance (92.7%–100%) to fluoroquinolones such as ciprofloxacin, levofloxacin, and norfloxacin [Table 3].
Table 3: Comparative resistance pattern between vancomycin resistant and vancomycin sensitive enterococci

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  Discussion Top


In the present study, elderly group of 41–60 years was most susceptible to enterococcal UTI. E. faecalis (54%) was found to be the most predominant isolate in the enterococcal UTIs followed by E. faecium. Same has been postulated by other Indian and foreign studies.[3],[13],[16] Resilient nature of E. faecalis helps its survival in acidic environments and responsible for its preponderance as the endogenous flora in the human body. Rare species such as E. avium, E. hirae, E. durans, and E. raffinosus were also isolated as previously reported by many recent studies.[7],[13],[14] This suggests that these species are changing their ecological habitat from animal, plant, soil, and water to humans and can add to the existing antibiotic resistance.

In the present study, E. faecium strains exhibited a significantly higher resistance to vancomycin and teicoplanin as compared to E. faecalis (P < 0.05). One strain was resistant to Linezolid. Higher level resistance of E. faecium to ampicillin and aminoglycosides have been highlighted by many studies which makes their synergistic combination ineffective.[17] A Canadian study has documented decreased sensitivity of fluoroquinolones and ampicillin against vancomycin-resistant E. faecium.[8] French literature has described the role of E. faecalis (V583) in disease severity correlated with antibiotic usage.[18] In the present study, 52% (52/100) of E. faecalis and 32% (32/100) of E. faecium were resistant to norfloxacin. In the present study, E. avium strains showed resistance to fluoroquinilones, ampicillin, erythromycin, and doxycycline sparing nitrofurantoin and fosfomycin for treatment enterococcal UTIs similar to previous reports.[17] It can be inferred that E. faecium shows higher degree of resistance to vancomycin and teicoplanin as compared to E. faecalis. Among non faecalis–non faecium species E. hirae and E. avium are important in terms of their occurrence and resistance from clinical samples. Thus, the identification of urinary isolates up to the species level from cases of significant enterococcal bacteriuria in the laboratory is utmost essential.

A VRE prevalence of 4% (4/100, E. faecium) was found among urinary isolates in our study. Concordant Indian studies have reported VRE prevalence between 2% and 35.3% from various geographical areas.[3],[4],[6],[19],[20] VRE prevalence varies as per geographical location and local epidemiology. A meta-analysis of published literature have documented VRE acquisition rate of 8.8% in ICUs across the United States, Australia, Taiwan, Holland, and Argentina. Approximately 10.6% get colonized on admission to ICUs which is a major determinant of VRE dynamics and the risk of VRE-related infections is closely related to colonization.[21]

VRE transmission from sources such as sewage, stools of farm animals, animals, and their products may result in an increased human reservoir of VRE leading to its colonization. Studies have documented risk factors such as hospital stay >48 h, ventilation, administration of the 3rd generation cephalosporins as independent risk factors for infection/colonization with VRE.[4] Lipopolysaccharide and flagellin of Gram-negative bacteria produce a C-type lectin Reg III γ which is capable of killing VRE. Hence, destruction of Gram-negative flora in the gut by the antibiotics play a role in its colonization with VRE.[22] Significant growth of VRE from urine in the microbiology laboratory without significant pyuria is colonization and should be reported with caution. Gut colonization by VRE should be determined by evaluating fecal samples of the patient and differentiated from clinically significant infections.

Various mechanisms have been suggested for emergence of vancomycin resistance like change of the D-Ala-D-Ala terminus of the protein precursors to D-Ala-D-lactate and D-Ala-D-Ser which is responsible for decreased affinity toward binding of vancomycin to peptidoglycan. The presence of nine phenotypic variants (VanA, VanB, Van C, VanD, VanE, VanG, VanL, VanM, and VanN) is responsible for acquired glycopeptide resistance. Van A gene cluster being present on transposon Tn1546 and Van B genotypes is more common and responsible for transferrable resistance.[23]

We compared the antimicrobial resistance pattern of vancomycin sensitive and VRE and found that all VRE (100%) were resistant to antimicrobials such as fluoroquinolones (ciprofloxacin, levofloxacin, and norfloxacin), ampicillin, erythromycin, teicoplanin, and fosfomycin. Twenty-five percent (1/4) of VRE strains were found to be resistant to linezolid and 50% (2/4) resistant to doxycycline. Our results are parallel with studies reporting decreased susceptibility/total resistance of VRE to routine antimicrobials such as doxycycline, ciprofloxacin, erythromycin, gentamycin, and teicoplanin.[24],[25] Purohit et al. from Delhi reported a resistance of 1.6% to linezolid and 3.2% to tigecycline among VRE.[5] Incidences and consequences of linezolid-resistant VRE have been discussed previously published literature.[26] Nitrofurantoin being sensitive to all VRE strains in the present study is a good therapeutic option in concordance to other results.[27],[28] Others have suggested aminopenicillins for treating vancomycin-resistant UTI regardless of the organism's ampicillin susceptibility.[28],[29]

Linezolid, daptomycin, quinupristin-dalfopristin, and tigecycline are last resorts to treat patients infected with VRE.[11] Latest inventions such as tedizolid, telavancin, dalbavancin, and oritavancin may prove to a milestone for the future treatment of VRE infectins. The evaluation should be done on a case-by-case basis due to concerns of toxicity, resistance, and insufficient supportive data. Multidrug resistance found among VRE isolates in our study raises a concern regarding its spread as a nosocomial pathogen and reserving last resort anti-VRE agents for complicated UTIs leading to septicemia.

Limitations

We did not perform genotypic studies including Van A, Van B, and Van C in VRE due to resource limitations. Second, the determination of high level resistance to gentamycin and streptomycin and third, inclusion of enterococcal isolates from samples other than urine would have yielded more relevant data. Research probing the sensitivity pattern of reserve antibiotics with larger number of samples is required to generate evidence for formulating policies for the management of VRE.


  Conclusion Top


VRE isolated from clinically significant bacteriuria cases pose a threat for its dissemination in the community. By results of this study, we recommend speciation of all clinically significant isolates, MIC for the detection of vancomycin resistance as per CLSI guidelines, differentiation of VRE colonization from infection and judicious use of reserve drugs in the era of emerging multidrug resistance among VRE isolates.

Acknowledgment

We are grateful to the Jaipur National University Institute of Medical Science and Research center (JNUIMSRC), Jaipur, Rajasthan, for providing infrastructure and resources for carrying out this research work.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
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