|Year : 2023 | Volume
| Issue : 1 | Page : 43-48
Screening for carriers of carbapenemase producing Enterobacteriaceae in critical care units
Kevin Thevarmadam Raju, Thomas S Kuruvilla, Sharon Joseph
Department of Microbiology, Father Muller Medical College, Mangalore, Karnataka, India
|Date of Submission||24-Nov-2022|
|Date of Decision||26-Dec-2022|
|Date of Acceptance||31-Dec-2022|
|Date of Web Publication||15-Mar-2023|
Thomas S Kuruvilla
Professor, Department of Microbiology, Father Muller Medical College, Mangalore, Karnataka - 575 002
Source of Support: None, Conflict of Interest: None
Background: The increasing cases of carbapenemase resistant Enterobacteriaceae (CRE) across the world is a cause of concern. Asymptomatic carriage of CRE in critical care units is a menace to infection control. Aims: This study determines the carriage rate of CRE in patients admitted to the intensive care units (ICU's) and evaluates the potential risk factors, leading to colonization in patients with CRE. Materials and Methods: Sixty rectal swabs from patients in the ICU's were screened for carriage of CRE. The samples were inoculated onto ChromID CARBA SMART bi-plate. The organisms showing color appearances as per the manufacturer's instructions were considered as CRE. Routine disk diffusion technique was also employed and CRE was defined as an organism belonging to the Enterobacteriaceae family which was resistant to either imipenem or meropenem. Results: The organisms isolated were identified and the percentage of carriage of carbapenem-resistant organisms was 12 (20%), of which Klebsiella pneumoniae was 4 (33.3%), Escherichia coli 6 (50%), Citrobacter freundii 1 (8.3%), and Enterobacter spp. 1 (8.3%). Out of these, 2 (3.3%) showed OXA 48 type resistance seen with K. pneumoniae and E. coli. Prior hospitalization, the use of high-end antibiotics and patients who have undergone surgeries were the most common potential risk factors for colonization with CRE. Conclusion: The prompt detection of CRE by routine screening using cost-effective methods and reduction of potential risk factors for gut colonization reduce the transmission of drug resistance in any hospital setting and pave the way for better antibiotic stewardship and appropriate contact isolation precautions.
Keywords: Asymptomatic carriage, carbapenemase, Enterobacteriaceae, resistance, screening
|How to cite this article:|
Raju KT, Kuruvilla TS, Joseph S. Screening for carriers of carbapenemase producing Enterobacteriaceae in critical care units. Saudi J Health Sci 2023;12:43-8
| Introduction|| |
Since almost two decades, carbapenems were the final drug of choice in the treatment of Enterobacteriaceae that were resistant to multiple drugs, especially in the intensive care units. Surveillance of carbapenemase-producing Enterobacteriaceae also referred to as carbapenem-resistant Enterobacteriaceae (CRE) is an essential part of preventing outbreaks, strengthen hospital infection control protocols, and antibiotic stewardship. Screening of carriers in ICU's is one way to track these unknown CRE colonizers and can be done using chromogenic media and an assessment of potential risk factors can help prevent the spread of carbapenem resistance.
Chromogenic medium is a feasible and cost-effective method to promote the identification of CRE pathogens from any clinical sample or from culture isolates at a rapid pace. Molecular characterization by methods such as Xpert Carba is also available although expensive for routine screening it can be used if necessary for the confirmation of the carbapenamase class.
One of the challenges in recent times in health-care facilities are infections by carbapenemase-producing Gram-negative pathogens, especially in high-risk patients in ICU. Among all the components of management, treatment options and infection control are the most challenging and close monitoring of these respective mechanisms of infection is necessary to prevent outbreaks in a hospital setup. Antibiotic resistance always carries with it unfavorable outcomes, which include high mortality, prolonged hospital stay, and rising health-related expenditure.,
Infections due to CRE have recently been listed under the high-risk category to a community by the Centers for Disease Control and Prevention Atlanta, USA, stressing the need to prevent these infections and their transmission. According to Calfee and Jenkins, hospitalized patients have a tendency for colonization ranging from 3% to 7%, although in a critical care setup, it can be much higher. In an Indian study conducted by Wattal C et. al, the prevalence of CRE in a critical care setup ranges between 13% and 51%. Active surveillance and a robust compliance with existing hospital infection control policies and protocols along with a meticulous approach, in general, can help prevent the spread of CRE, especially in an ICU.
Enterobacteriaceae, such as Escherichia coli and Klebsiella pneumoniae are pathogens that are responsible for a variety of infections such as cystitis, pyelonephritis, septicemia, pneumonia, peritonitis, and meningitis. Apart from these two bacteria, there are other potential pathogenic species in the Enterobacteriaceae family that result in similar human infections caused by Citrobacter, Enterobacter, Proteus, Providencia, and Serratia marcescens which are transmitted within hospitals by colonizing the hands of health-care personnel and by contamination of the food or water that is consumed.
Carbapenems have a major role in the treatment of potentially grave hospital-acquired infections caused by these Enterobacteriaceae that may produce enzymes like extended-spectrum beta-lactamases. Detection of carbapenemase resistance in Enterobacteriaceae is very difficult by routine Kirby-Bauer disc diffusion methods in most microbiology laboratories, but phenotypic methods such as the modified Hodge test and modified carbapenem inactivation method (mCIM) are used for simple detection of Enterobacteriaceae that produce carbapenemase. There are a variety of carbapenemases like metallo-β-lactamases belonging to Ambler class B (IMP, VIM, and NDM) and K. pneumoniae carbapenemase of Ambler class A and OXA; Ambler class D that are prevalent in Enterobacteriaceae in various countries around the globe. Those who harbor these resistant strains in their gastrointestinal tract are considered reservoirs for cross-transmission in a hospital setting. Thus, a rectal swab specimen is a good sampling method for CRE surveillance.
There are many tested culture techniques for screening carbapenem-resistant Enterobacteriaceae, such as CHROM agar, mSuper CARBA (MSCA), and ChromID CARBA SMART (IDCA). The bioMérieux company of France has developed a ChromID CARBA SMART bi-plate, with OXA-48 screening media (ChromID OXA-48) on one half of the plate and a CRE screening media (ChromID CARBA) on the other. This ChromID CARBA is a type of selective agar for identifying CRE and has a good sensitivity and specificity of >90% for various non-OXA CRE.
There is however no established standard method for screening these CRE. Direct molecular methods can also be used since it is more sensitive and rapid. These molecular methods are expensive and can detect only a few known genes. An added drawback is that bacteria will not be available for routine biochemical identification or antibiotic sensitivity testing.
| Materials and Methods|| |
This observational, descriptive type of study was done from various ICUs in collaboration with the department of microbiology of a tertiary care hospital, for duration of 2 months. After voluntary consent, rectal swabs from admitted patients in the ICUs were obtained and transported to the microbiology laboratory for processing. A total of 60 patients all above 18 years, admitted to these critical care units were screened for carriage of CRE. All patients <18 years old and not admitted to the ICUs of the hospital were not taken for the study. The demographic details of the patients and associated probable risk factors like previous admission to any health-care facility or ICU, use of high-end antibiotics, previous history of any surgery over the past 90 days, or the use of any invasive devices or development of any systemic infection or chronic conditions such as chronic obstructive pulmonary disease (COPD) or chronic renal failure, autoimmune diseases, malignancies, and steroid use were recorded.
A rectal swab was collected from all patients within 12–48 h of being admitted to the ICUs. A sterile swab moistened with a little bit of sterile saline was inserted around 1 inch beyond the anal canal sphincter and the swab was gently rotated, so that a good sample from the rectal orifice could be obtained. On withdrawal, the swab was placed in its test tube and immediately sent to be processed in the microbiology laboratory.
This rectal swab was inoculated into peptone water and then the suspension was aseptically inoculated onto the ChromID CARBA SMART bi-plate. After plating, the culture petri dishes were then incubated at 37°C for around 18–24 h. The organisms showing color appearances as per the manufacturer's instructions (green–blue-to-browny green, pink-to-burgundy, or brown colonies) were considered CRE.
These colored colonies were also biochemically identified by manual methods and their antibiotic sensitivity test by the Kirby-Bauer disc diffusion method following the Clinical Laboratory Standards Institute guidelines were done to confirm resistance to carbapenems. The antibiotic panel tested included aztreonam (30 μg), piperacillin-tazobactam (100/10 μg), ciprofloxacin (5 μg), amikacin (30 μg), imipenem (10 μg), and meropenem (10 μg) (HiMedia Laboratories, India Pvt. Ltd.).
A supportive phenotypic test was done for isolates that were intermediate or susceptible for imipenem, i.e., 16 mm–21 mm. The mCIM test was done for carbapenemase production as recommended by the CLSI 2018 guideline before reporting the carbapenem susceptibility results. Here, 1 μL loopful of Enterobacteriaceae colonies grown on blood agar were mixed in 2 mL of trypticase soy broth (TSB). An antibiotic disk containing meropenem was put in the suspension and then incubated for 4 h at 35°C. A 0.5 McFarland suspension of E. coli ATCC 25922 standard strains was emulsified in saline by the direct colony suspension method. A lawn culture of this E. coli ATCC 25922 was done on a Mueller–Hinton agar (MHA) plate. The meropenem disk from the TSB was removed and placed on this MHA plate on which E. coli ATCC 25922 indicator strain was inoculated. The culture petri dishes were incubated at 35°C for 18–24 h. A zone size diameter of 6–15 mm or any colony growing within a 16–18 mm zone were considered positive and a inhibition zone size of ≥19 mm was taken as a negative result.
CRE was defined as an organism belonging to Enterobacteriaceae family which is resistant to either imipenem or meropenem. The data were analyzed using a software SPSS 17.0 (SPSS Inc., Chicago, IL, USA). The distribution of continuous variables was presented as a mean ± standard deviation. Other tests such as the Chi-square and Fisher's exact test were used to analyze categorical variables.
| Results|| |
A total number of 60 fecal samples were tested. A majority of the patients were male 37 (61.7%) and 29 (48.3%) were more than 60 years of age. The tally of samples that showed CRE, non-CRE, and negative (i.e., culture plates with no growth) is shown in [Table 1].
|Table 1: Total isolates of carbapenemase Enterobacteriaceae, noncarbapenemase Enterobacteriaceae and negative samples|
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The potential risk factor parameters analyzed, of the 60 cases reveals that 14 (23.3%) of the cases had a previous history of admission to other health-care facilities. The use of high-end reserve antibiotics/antifungals (carbapenems, colistin, tigecycline, vancomycin, linezolid, teicoplanin, and caspofungin) were seen in 8 (13.3%) of the cases and antibiotic escalations were done based on culture reports. Four (6.7%) cases underwent surgeries such as hysterectomy, laparotomy, intestinal obstruction, hip repair, and burr hole and were admitted to the ICU for critical care. Nine (15%) had systemic infections such as urinary tract infections, pulmonary tuberculosis, leptospirosis, pneumonia, and malaria. The elderly patients 37 (61.7%) had long standing diseases such as high blood pressure, ischemic changes of the heart, Type 2 diabetes mellitus, asthma, obesity, and chronic obstructive pulmonary disease (COPD). Nine (15%) patients were on steroids as part of their treatment protocol. Death was the outcome in 8 (13.3%) and some of the cases admitted and were due to septic shock (neutropenic)/acute respiratory distress syndrome, complications from tuberculosis, cerebrovascular accident, and cholecystitis. The potential risk parameters of the patients who were and were not colonized with CRE are shown in [Table 2].
|Table 2: Potential risk factors leading to carbapenemase Enterobacteriaceae colonization|
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The organisms that were isolated from the feces samples were also identified biochemically as K. pneumoniae, Klebsiella oxytoca, E. coli, Citrobacter freundii, and Enterobacter spp. The color codes of these isolates that showed carbapenem resistance seen on ChromID CARBA SMART bi-plate agar were E. coli pink to burgundy (For OXA-48), bluish green to bluish gray for Klebsiella, Enterobacter, and Citrobacter species in [Table 3] and [Figure 1].
|Figure 1: CRE Klebsiella (L), Escherichia coli (R) colonies on ChromID CARBA SMART respectively|
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|Table 3: Carbapenemase Enterobacteriaceae colonizers and their colony colors on ChromID CARBA SMART|
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The percentage of carriage of carbapenem-resistant organisms (CRE) were 12 (20%) of which K. pneumoniae was 4 (33.3%), E. coli 6 (50%), C. freundii 1 (8.3%) and Enterobacter spp 1 (8.3%) as depicted in [Table 3]. Out of these, 2 (3.3%) showed OXA-48 type resistance and was seen with K. pneumoniae and E. coli.
| Discussion|| |
The increasing cases of CRE across the world is a cause of concern and carbapenems being the reserve drug for multidrug resistant Enterobacteriaceae may get hydrolyzed by carbapenemases. The carriage of CRE for prolonged periods in the feces is a source for the spread of these pathogens by cross-contamination, leading to urinary tract infections, abdominal infections, and even sepsis.
In today's scenario, there are no known standard protocols for routine surveillance of fecal carriage in India. In our study CRE colonization was detected in 12 (20%) cases when compared to a study by Yamuna et. al where colonization with CRE by fecal swabs was only 7.8%. However our carriage rates did closely match with a study done by Papadimitriou-Olivgeris et al. who detected a colonization rate of 28%. Fourteen (23.3%) of the cases studied had a previous history of admission to other health care facilities. This was a significant risk factor (P = 0.038) in our study for colonization with CRE when compared to a study by Ramanathan et al., where CRE in previously hospitalized was 8.6%. He had also studied the CRE colonization of the patients from the community and found it to be little lower at 6.8%. Since our hospital is a tertiary care hospital with many referrals it could be a reason for high gut carriage of CRE's.
In our study 8 (13.3%) cases with CRE colonization were started on high-end reserve antibiotics such as meropenem, colistin, tigecycline, vancomycin, linezolid, and teicoplanin. There was no significant association of these cases with CRE colonization (P = 0.211) which was in variance with a study by Swaminathan et al. He further analyzed other risk factors such as central line insertion, endotracheal intubation and Foley's catheterization where there was no significant association with CRE colonization. In contrast, our study risk factors like patient on dialysis and Foley's catheterization had a significant association with increased CRE colonization (P = 0.014 and 0.028), respectively. Four (6.7%) cases that underwent surgeries in the past 90 days had CRE colonization which was significant (P = 0.048) and this finding was similar to studies by Yamuna et. al where a statistical association with CRE colonization (P = 0.0167) was seen.
Those cases colonized with CRE and who developed systemic infections were not however infected by the same CRE isolates in our study. OXA-48 class of CRE was seen in cases with underlying hypertension, ischemia, diabetes, and high-end antibiotic exposure, but only the latter factor had a significant correlation. A challenge in OXA-48 detection is that it exhibits sensitivity to broad-spectrum antibiotics like cephalosporins and eventually may remain nonresponsive to carbapenems.
Among the various members of Enterobacteriacae family, the presence of carbapenemases was observed mainly in E. coli 6 (50%) and K. pneumoniae 4 (33.3%) and was in consensus with a study by Xu et. al where K. pneumoniae (39.3%) and E. coli (21.97%) were reported to have a high resistance to carbapenems. The overall resistance to both imipenem and meropenem among the CRE isolated were the same in the present study at 29.4% despite literature that quotes a higher resistance to meropenem is usually seen due to added mechanisms of increased efflux pumps where the expulsion of the drug out of the bacterial cell is seen which is not seen for imipenem. The use of cephalosporins and fluoroquinolones were considered a potential risk for infections by CRE that had been noticed in earlier studies. However, Zhao et al. did not find any such correlation and neither did we see it in our study.
The culture-based ChromID CARBA SMART bi-plate, and an additional mCIM phenotypic test to confirm that the isolates were CRE or not were relatively user friendly and rapid screening tools in our study when compared to expensive molecular alternatives such as polymerase chain reaction, Xpert Carba and Matrix assisted laser desorption ionization time-of-flight mass spectrometry. However, every methodology has its own advantages and disadvantages. The media with color ingredients making it chromogenic can be a satisfactorily quick means of detecting CRE, but its greatest potential lies in the use of rectal swab sampling. Therefore, it would be imperative that rectal samples can be suited for a risk factor associated active surveillance. Chromogenic media, however, may give rise to false-positives results. Thus, it is useful in doubtful cases to confirm the positives with either a disc diffusion assay or by molecular methods.
| Conclusion|| |
The current study reveals a high carriage rate of CRE in critical care units. Prior hospitalization, the use of high-end antibiotics and patients who have undergone surgeries were the most common potential risk factors for being colonized with CRE. The utility of certain in situ devices for patients also can lead to prolonged hospital stay and thus invariably contribute to high carriage of CRE. Routine screening using cost-effective rapid methodologies will help curtail spread of CRE in critical care units and thereby control hospital-acquired infections and help better monitoring of the antibiotic stewardship protocols in tertiary health care facilities.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Amjad A, Mirza IA, Abbasi S, Farwa U, Malik N, Zia F. Modified Hodge test: A simple and effective test for detection of carbapenemase production. Iran J Microbiol 2011;3:189-93.
Bakthavatchalam YD, Anandan S, Veeraraghavan B. Laboratory detection and clinical implication of oxacillinase-48 like Carbapenemase: The hidden threat. J Glob Infect Dis 2016;8:41-50.
Ramanathan YV, Venkatasubramanian R, Nambi PS, Ramabathiran M, Venkataraman R, Thirunarayan MA, et al.
Carbapenem-resistant enterobacteriaceae screening: A core infection control measure for critical care unit in India? Indian J Med Microbiol 2018;36:572-6.
] [Full text]
Calfee D, Jenkins SG. Use of active surveillance cultures to detect asymptomatic colonization with carbapenem-resistant Klebsiella pneumoniae
in intensive care unit patients. Infect Control Hosp Epidemiol 2008;29:966-8.
Papadimitriou-Olivgeris M, Marangos M, Fligou F, Christofidou M, Bartzavali C, Anastassiou ED, et al.
Risk factors for KPC-producing Klebsiella pneumoniae
enteric colonization upon ICU admission. J Antimicrob Chemother 2012;67:2976-81.
Wattal C, Goel N, Oberoi JK, Raveendran R, Datta S, Prasad KJ. Surveillance of multidrug resistant organisms in tertiary care hospital in Delhi, India. J Assoc Physicians India 2010;58 Suppl: 32-6.
Mariappan S, Sekar U, Kamalanathan A. Carbapenemase-producing Enterobacteriaceae
: Risk factors for infection and impact of resistance on outcomes. Int J Appl Basic Med Res 2017;7:32-9.
Vrioni G, Daniil I, Voulgari E, Ranellou K, Koumaki V, Ghirardi S, et al
. A comparative evaluation of a prototype chromogenic medium (ChromID CARBA) for detecting carbapenemase-producing Enterobacteriaceae
in surveillance rectal swabs. J Clin Microbiol 2012;50:1841-6.
Jing X, Zhou H, Min X, Zhang X, Yang Q, Du S, et al.
The simplified carbapenem inactivation method (sCIM) for simple and accurate detection of carbapenemase-producing gram-negative Bacilli. Front Microbiol 2018;9:2391.
Creighton J, Wang H. Evaluation of CHROMagar™ mSuperCARBA™ for the detection of carbapenemase producing gram-negative organisms. N Z J Med Lab Sci 2016;70:101-5.
Lee SY, Octavia S, Chew KL. Detection of OXA-carbapenemase-producing Enterobacteriaceae
with chromID CARBA SMART screening plate. Pathology 2019;51:108-10.
Mohan B, Prasad A, Kaur H, Hallur V, Gautam N, Taneja N. Fecal carriage of carbapenem-resistant Enterobacteriaceae
and risk factor analysis in Hospitalised patients: A single Centre study from India. Indian J Med Microbiol 2017;35:555-62.
] [Full text]
Chu YW, Afzal-Shah M, Houang ET, Palepou MI, Lyon DJ, Woodford N, et al.
IMP-4, a novel metallo-beta-lactamase from nosocomial Acinetobacter
spp. collected in Hong Kong between 1994 and 1998. Antimicrob Agents Chemother 2001;45:710-4.
McGettigan SE, Andreacchio K, Edelstein PH. Specificity of ertapenem susceptibility screening for detection of Klebsiella pneumoniae
carbapenemases. J Clin Microbiol 2009;47:785-6.
Swaminathan M, Sharma S, Poliansky Blash S, Patel G, Banach DB, Phillips M, et al.
Prevalence and risk factors for acquisition of carbapenem-resistant Enterobacteriaceae
in the setting of endemicity. Infect Control Hosp Epidemiol 2013;34:809-17.
Mulvey MR, Grant JM, Plewes K, Roscoe D, Boyd DA. New Delhi metallo-β-lactamase in Klebsiella pneumoniae
and Escherichia coli
, Canada. Emerg Infect Dis 2011;17:103-6.
Schechner V, Kotlovsky T, Tarabeia J, Kazma M, Schwartz D, Navon-Venezia S, et al.
Predictors of rectal carriage of carbapenem-resistant Enterobacteriaceae
(CRE) among patients with known CRE carriage at their next hospital encounter. Infect Control Hosp Epidemiol 2011;32:497-503.
Zhao ZC, Xu XH, Liu MB, Wu J, Lin J, Li B. Fecal carriage of carbapenem-resistant Enterobacteriaceae
in a Chinese university hospital. Am J Infect Control 2014;42:e61-4.
[Table 1], [Table 2], [Table 3]