Original article / research
|
|||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Vancomycin Resistant Enterococcal Urinary Tract Infection: A Potential Threat |
|||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
|
Sabba Mussadiq, Rajesh Kumar Verma, Dharmendra Prasad Singh, Mohd Mubashir Ali Khan 1. Senior Resident, Department of Microbiology, Uttar Pradesh University of Medical Sciences, Etawah, Uttar Pradesh, India. 2. Professor, Department of Microbiology, Uttar Pradesh University of Medical Sciences, Etawah, Uttar Pradesh, India. 3. Professor, Department of Microbiology, Uttar Pradesh University of Medical Sciences, Etawah, Uttar Pradesh, India. 4. Associate Professor, Department of Urology, Uttar Pradesh University of Medical Sciences, Etawah, Uttar Pradesh, India. |
|||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Correspondence
Address : Mohd Mubashir Ali Khan, 106 B, Type 4, Old Doctors Campus, UPUMS, Saifai, Etawah, Uttar Pradesh, India. E-mail: mubashiirkhan@gmail.com |
|||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| ABSTRACT |  | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| : Enterococci are considered less virulent organisms, but have incurred recognition for being notorious for their acquisition and transfer of resistance. The greatest potential threat posed by enterococci is vancomycin resistance. The transfer of enterococcal vancomycin resistance to Staphylococcus aureus has been achieved making scientists apprehensive of its consequences. Aim: To find the prevalence of Vancomycin-Resistant Enterococci (VRE) and to determine the antimicrobial resistance pattern in enterococcal urinary isolates. Materials and Methods: A cross-sectional study was carried out on all urinary samples suspected of Urinary Tract Infection (UTI) received for duration of one year from April 2021 to March 2022 in the Department of Microbiology at Uttar Pradesh University of Medical Sciences, Saifai, Etawah, Uttar Pradesh. Enterococci were isolated and identified with a VITEK-2® COMPACT (bioMérieux) automated system. The antibiotic susceptibility pattern was determined by Kirby-Bauer disc diffusion method. Further, confirmation of VRE was done by Minimum Inhibitory Concentration (MIC) E-test. Results: A total of 128 urinary enterococcal isolates were identified with the male-to-female ratio 1.37:1 and mean age of patients was 37.18±22.64 years. Out of total, 71.87% were identified as Enterococcus faecalis followed by 24.21% Enterococcus faecium and the rare species (4%) including E. durans, E. hirae, E. raffinosus. The prevalence of VRE was found to be 8.6%. Maximum resistance by isolates has been shown against ampicillin, erythromycin, ciprofloxacin, and doxycycline. All isolates were sensitive to linezolid. Nitrofurantoin resistance was observed in 4.34% and 25.80% of E. faecalis and E. faecium isolates, respectively. Conclusion: In this study, it was revealed that the emergence of VRE in urinary isolates with antimicrobial resistance was higher among E. faecium. All this puts pressure on strict compliance with a multidimensional approach with collaboration of antibiotic stewardship, educational and surveillance programs. | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
 |
|||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Keywords : Bladder infection, Gram-positive cocci, Prevalence | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| INTRODUCTION | | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| Enterococcus is one of the pathogens forming the acronym ESKAPE (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) group, which accounts for the majority of nosocomial infections worldwide and has also been considered a pathogen of high priority by WHO in its global list of significant antibiotic-resistant bacteria (1). Enterococcus was previously classified as group D Streptococci but with the help of molecular analysis, they have been placed in a separate genus, Enterococcus (2). They are gram-positive cocci which are indigenous inhabitants of the gastrointestinal tract, oral cavity, and genital tract of humans (3). Genus Enterococcus, includes a variety of species, but only two species are considered to cause the majority of infections, these are Enterococcus faecalis (85-90%) and Enterococcus faecium (5-15%) (2). Enterococci are one of the dominant uropathogens. Besides infecting the urinary tract, they are the second prominent cause of hospital-acquired urinary infections and also cause other infections like bacteraemia, wound infections, intra-abdominal and pelvic infections, urinary catheter-associated infections, and infective endocarditis (4). They rarely cause meningitis or respiratory tract infections (5). The inception of urinary VRE has emerged as a dreadful infection in immunocompromised patients (6). Patients who have enterococcal UTIs, either exhibit minimal or no symptoms. Symptoms are mostly related to catheterisation and instrumentation (7). Complications and recurrences associated with UTIs are due to pre-existing urinary tract abnormalities (8). Enterococci are considered less virulent organisms but have incurred recognition for being notorious for their resistance traits (9). The world has seen an increase in drug-resistant bacteria at a remarkable rate and enterococcus is one amongst them. The greatest potential threat posed by enterococci is vancomycin resistance (6). The empirical use of antibiotics gives bacteria an edge of selective pressure to acquire resistance (10). Enterococci are intrinsically resistant to antimicrobials used frequently and also acquire resistance by mutation or in conjugation with the transfer of genetic material (11). The transfer of enterococcal vancomycin resistance to Staphylococcus aureus has been achieved and scientists are apprehensive that it may emerge in nature as a new pandemic with devastating consequences (12),(13). Past reports of VRE emerged in the late 1980s and since then it has become of paramount concern for researchers (10). The prevalence of VRE in Europe is demonstrated to vary between 1% to 30%, whereas in the United States it is estimated to be about 30%, which is considered to be hospital-acquired (2). Amongst the phenotypic vancomycin resistance, most common in the United States are Van A and Van B, about 70% and 25%, respectively (6). Van A enterococci are resistant to both vancomycin and teicoplanin, whereas Van B is resistant only to vancomycin and preserve susceptibility to teicoplanin (11). The assessment of resistance varies extensively in different geographical latitudes, depending on their antimicrobial stewardship programs, empirical practice patterns, and prevalence of habitant-resistant enterococcal species (5). Patients colonised with VRE remain a threat for hospital outbreaks as they serve as a reservoir of VRE and subsequently spread infection from patient to patient (10),(14). Animals are said to be reservoirs, from which VRE transmits to man via the food chain (11). Hospitalised patients, hospital equipment, environmental surfaces, and VRE patient areas, may also serve as a reservoir for VRE (14). UTIs caused by VRE bear serious health and socio-economic burden including the cost of hospitalisation (2). Categorising VRE associated urinary colonisation, asymptomatic bacteriuria, and UTIs to determine optimal treatment options, duration of therapy, and eventually decreasing mortality and healthcare costs (15). The global dissemination of recalcitrant VRE showing an increasing trend is a critical situation strikingly the most important is their ability to acquire resistance to the presently available antimicrobial agents, along with the dearth of new agents, with very few in development (6),(11),(16). Therefore, the present study was undertaken to know the prevalence and antimicrobial resistance pattern of enterococcal urinary isolates and to determine the phenotypes of VRE. | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| MATERIAL AND METHODS | | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| A prospective cross-sectional study was performed on urinary samples of patients suspected of UTI for routine culture and sensitivity. The samples were received from both outpatient and inpatient areas in the Department of Microbiology at Uttar Pradesh University of Medical Sciences, Saifai, a Tertiary care hospital Etawah, Uttar Pradesh, India for a period of one year from April 2021 to March 2022. Inclusion criteria: Urine samples from both outpatient and inpatients of our hospital, of all age groups and both sexes complaining of urinary symptoms were included in the study. Exclusion criteria: Body fluids, sputum, blood, and specimens other than urine were excluded from the study. Additionally, urine specimens with bacterial growth other than enterococci were also excluded. Study Procedure Using the semiquantitative method, urine samples were cultured on Cysteine Lactose Electrolyte Deficient agar (CLED), reading the plates after 24 hours of aerobic incubation at 37ºC. Colony counts yielding bacterial growth of >105 CFU/mL were considered significant bacteriuria. Presumptive identification of the genus Enterococcus was done based on using colony morphology, gram-staining, hydrolysis by bile esculin and catalase. Species identification was done separately by using the VITEK-2® COMPACT (bioMérieux, Marcy I’Etoile, France) automated system (17). Antibiotic susceptibility testing was done for all the enterococcal isolates by the Kirby-Bauer disc diffusion method on Mueller-Hinton agar and the results were interpreted as per Clinical and Laboratory Standards Institute (CLSI) guidelines (18). Muller-Hinton agar plates were surface seeded by preparing a uniform suspension of culture with turbidity adjusted to 0.5 McFarland standard. The plates were read with the naked eye using transmitted light for appreciating the presence or absence of a zone of inhibition around each antibiotic disc. Quality control was done by using the ATCC strain of Enterococcus faecalis, 29212 which was procured from HiMedia Laboratories Pvt., Ltd., Mumbai, Maharastra, India. The following commonly used antibiotic discs and their disc potency that was used for susceptibility testing of enterococcal isolates is as follows: Ampicillin 10 μg, Erythromycin 15 μg, Tetracycline 30 μg, Doxycycline 30μg, Ciprofloxacin 5 μg, Vancomycin 30 μg, Teicoplanin 30 μg, High Level Gentamycin (HLG) 120 μg, Linezolid 30 μg, Nitrofurantoin 300 μg. All enterococcal isolates with an inhibition zone diameter of ≤14 mm or any growth within the zone of inhibition were considered vancomycin-resistant and isolates with a zone diameter of ≤10 mm were considered resistant to teicoplanin by disc diffusion method (18). VRE isolates by disc diffusion method were further confirmed by E-test strips (MIC range: 0.016 to 256 μg/mL) and read according to the manufacturer’s recommendations. Enterococci that had MIC >32 μg/mL were noted as resistant to both vancomycin and teicoplanin according to CLSI guidelines (18). Phenotypic characterisation of VRE was classified as Van A and Van B, with Van A strains showing resistance to both vancomycin as well as teicoplanin, whereas Van B phenotypes remain susceptible 2to teicoplanin. Culture media, antiobiotic discs and E-test strips used in the study were secured from HiMedia Laboratories Pvt., Ltd., Mumbai, Maharastra, India. Statistical Analysis Data was segregated, coded, and recorded in the Microsoft® Excel spreadsheet program. Descriptive percentages and frequencies were represented by constructing tables. Mean, standard deviation, and ratios were summarised for continuous variables, whereas frequencies and proportions were for categorical variables. All statistical analysis was performed with statistical software using SPSS version 24.0 by IBM USA. | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| RESULTS | | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| In the present study, 1996 urine samples received in the department were screened for UTIs. Out of which 300 isolates were culture positive. A total of 128 (42.66%) isolates were identified as Enterococcal spp. and 172 other bacteria. Of 128 enterococcal isolates, 74 (57.81%) were males and 54 (42.18%) were females. The male-to-female ratio was 1.37:1. The mean age of patients was 37.18±22.64 years (range: 7 days to 92 years). The highest isolates 43 (33.6%) were from the age group 21-40 years, followed by 38 (29.68%) from age up to 20 years, 26 (20.31%) aged 41-60 years, 18 (14.06%) were aged 61-80 years and 3 (2.34%) were ≥80 years. Inpatients were 87.5% and outpatients were 12.5%. Among the inpatients, 37 (33%) isolates were obtained from the General Surgery ward followed by 28 (25%) from the Urology ward (Table/Fig 1). Five different species of enterococci were identified as Enterococcus faecalis 92 (71.87%), Enterococcus faecium 31 (24.21%), and rare species E. durans 3 (2.34%), E. hirae 1 (0.78%), E. raffinosus 1 (0.78%) (Table/Fig 2). The antimicrobial resistance pattern by the Kirby-Bauer disc diffusion method is depicted in (Table/Fig 3). The resistance pattern of E. faecalis to commonly used drugs was erythromycin (86.95%), ampicillin (84.78%), ciprofloxacin (82.60%), doxycycline (81.52%). E. faecium showed resistance mostly (90.32%) to ampicillin, ciprofloxacin, (88%) to erythromycin, and HLG, (83.87%) to doxycycline. All five isolates were 100% sensitive to linezolid. Resistance toward nitrofurantoin was found (4.34%) in E. faecalis and (25.80%) in E. faecium. E. raffinosus was resistant to none of the antibiotics tested (Table/Fig 3). Antibiotic resistance pattern of VREs compared to Vancomycin Sensitive Enterococcal (VSE) islates depicated in (Table/Fig 4). All the VRE strains were resistant (100%) to erythromycin, ciprofloxacin and HLG. Around 90% VRE were also resistant to ampicillin and doxycycline. Among VSE strains; 82 (70.08%) were HLG resistant. VSE strains showed highest resistance to erythromycin, ampicillin and ciprofloxacin. All VSE strains were sensitive to teicoplanin. Out of 128 enterococcal isolates by disc diffusion method, 11 were resistant to vancomycin as such the prevalence of VRE was found to be 8.6%, of which nine were E. faecium and two were E. faecalis. Vancomycin resistance was not found in other species. Further confirmation of vancomycin-resistant isolates by E-test showed no false susceptibility, as Minimium Inhibitory Concentrations (MICs) of all 11 isolates were >32 μg/mL. One of the two vancomycin-resistant E. faecalis and 8 vancomycin-resistant E. faecium isolates had MIC values >256 μg/mL for vancomycin (Table/Fig 5). Three out of seven E. faecium had MIC values >256 μg/mL for teicoplanin (Table/Fig 6). Seven isolates of E. faecium showed Van A phenotype. Four isolates showed Van B phenotype and were distributed among both species equally (Table/Fig 7). | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| DISCUSSION | | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| A survey done by the Centers for Disease Control (CDC) and prevention found a 20-fold increase in VRE cases in a short period (16). This shows the vulnerability of VRE acquisition and dissemination at a significant rate in healthcare centers, which is a matter of great concern. In our study, the prevalence of VRE was found to be 8.6%, which is similar to the study done from South India by Fernandes SC and Dhanashree B (3). Although there are studies from India and other countries that showed the prevalence of VRE as low as 0.43% and others as high as 42.9% (Table/Fig 8) (4),(19),(20),(21),(22),(23),(24),(25). Among the enterococcal isolates, E. faecalis and E. faecium were commonly isolated, with the predominance of the former species. Other rare species such as E. durans, E.hirae, and E. raffinosus were also isolated which corroborates with the findings of other studies done in India and China (21),(26). In a study done from Bangalore by Sreeja S et al., E. faecalis and E. faecium were found to be the only enterococcal species from various clinical specimens as identification by conventional methods by them may not be able to identify strains with atypical phenotypes (5),(20). This can also be related to different strains circulating in different geographical regions, even study settings, methodologies used, and presentation of cases (2),(11),(13). More isolates were from inpatients 112 (87.5%), which seemed to be evidently in excess as compared to outpatients 16 (12.5%), this being in concordance with a study done in Ethiopia, to show hospital-acquired enterococcal infection at a rise (5). As such, eminent importance should be given to the infection control measures in the hospitals, to recede the disseminating infection (6),(14). In the current study, young patients were found to suffer from enterococcal UTIs. The highest prevalence of 33.6% was seen in the age group 21-40 years, with similar findings recorded in a study done in Nigeria (19). Our study also showed a male predominance of UTIs which can be related to a major contribution from the urology ward, male-dominated and long-term use of indwelling catheters in these wards leads to the formation of biofilm resulting in persistent infection by drug-resistant microorganisms (2),(26). In the present study, E. faecium was found to be more drug-resistant than E. faecalis as was evidenced by other study (22). Among 11 VRE isolated in this study, E. faecium 9 (81.81%) was the dominant species, whereas our results were not in congruence with a study from Uttar Pradesh by Yadav G et al., who identified E. faecalis to be the only vancomycin-resistant isolate (23). Vancomycin resistance is less in other enterococcal species (27). Maximum resistance by enterococcal isolates has been shown against ampicillin, erythromycin, ciprofloxacin, and doxycycline. HLG resistance was seen more in E. faecium 27 (88%) than E. faecalis 66 (71.73%), with a similar resistance pattern reported from Mumbai by Karmarkar MG et al., (9). The resistance offered to HLG restricts the use of aminoglycosides for resistant enterococci, thus leaving the therapeutic realm with very few drug options. Both isolates E. faecalis and E. faecium displayed low resistance to nitrofurantoin (4.34%) and (25.80%) respectively. This makes nitrofurantoin to be a pertinent option for treating VRE UTI, as it attains good concentration levels in urine, which is in agreement with the recommendations by other researchers [23,28]. None of the Enterococcal spp. in our study, showed resistance to linezolid, which is incomparable with the study from North-East India reporting a resistance of 4.5% (29). Linezolid can be preserved for use in VRE UTI in intricate cases such as complicated renal diseases and urosepsis (15). While we compared the resistance profile of VRE with vancomycin-sensitive enterococci and it was noteworthy to find that all VRE (100%) were resistant to erythromycin, ciprofloxacin and HLG, which may be due to unusual ability of enterococci to acquire resistance (6). Other authors from Bihar have reported maximum resistance to pencillin (100%), Ampicillin (91.65) and piperacillin (75.0%) among VRE isolates (13). Incompatible antibiotic resistance patterns could be due to compliance with infection control measures and monitoring techniques for the detection of VRE, surveillance for colonisation, and hand-washing practice (10). This is also impacted by regional antimicrobial practice and screening patterns, and isolation policies of the healthcare facilities. Among the 11 VRE, 9 isolates in our study showed a high level of resistance to vancomycin (MIC >256 μg/mL) which can be compared to the study done in Kuwait (30). This study shows that 7 (63.63%) of VRE isolates had Van A phenotype, with similarly high rates of Van A phenotype reported from South India by Praharaj I et al., (4). The reliability of methodologies put into use in different laboratories as per their resources, ease, and testing frequency indicate a need for further testing before establishing results, as such, the findings of our study must be inspected with caution and therefore are not generalisable. Limitation(s) There are a few limitations to our study. Firstly, in this study, the molecular approach for definitive speciation was not done due to limited resources. Secondly, enterococcal isolates from samples other than urine might have provided more pertinent data. However, these presumptive results can help clinicians in the empirical choice of drugs for treating VRE. | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| CONCLUSION | | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| This study reveals an 8.6% prevalence of VRE UTI however, which can be a subtle signal for the emergence of VRE demanding VRE surveillance in healthcare facilities. Moreover, Enterococcal spp. with its apt ability to acquire and transmit antimicrobial resistance along with insufficient treatment options is a threat to the ecosystem. We reported increased drug resistance by Enterococcal species especially by E. faecium. All this puts pressure on strict compliance of multidimensional approach with collaboration of antibiotic stewardship, educational and surveillance programs. Aggressive and active infection control measures need to be proposed to contain the spread of VRE. Nitrofurantoin may be considered a choice for treating uncomplicated infection guided by the regional patterns of drug resistance. Nevertheless, further studies should be done using molecular diagnostics together with phenotypic modalities which can facilitate inaccurate and definite detection of VRE, also identifying highly heterogeneous strains. Adequate measures must be taken to contain, resistant enterococcal strains from dissemination, to prevent outbreak like situation in hospitals worldwide. | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
|
|||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
|
|||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
|
|||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
