Beta-Lactamases and Carbapenemases
By Dr. Zubair Khalid, DVM, MS, PhD ·
Quick Answer
- Clinical laboratories choose between phenotypic carbapenemase detection tests like the modified carbapenem inactivation method (mCIM) and molecular assays based on their workflow capacity, cost, and need for resistance gene identification.
- The mCIM test offers a low-cost phenotypic approach that detects carbapenemase activity directly from cultured isolates, while molecular methods provide definitive gene-level identification of carbapenemase types.
- No single detection method identifies all carbapenemase variants, and laboratories must understand the performance limitations of each approach when interpreting results for patient management decisions.
Clinical Context for Beta-Lactamase Detection
Antimicrobial resistance represents one of the most pressing challenges in modern medicine and agriculture. The World Health Organization identifies antimicrobial resistance as a global health threat that requires coordinated surveillance and stewardship efforts across human health, animal health, and environmental sectors. Beta-lactamases are enzymes produced by bacteria that hydrolyze beta-lactam antibiotics, rendering them ineffective against infections. These enzymes represent the most common mechanism of resistance to penicillins, cephalosporins, and carbapenems among gram-negative bacteria.
Carbapenems are often considered agents of last resort for treating infections caused by multidrug-resistant gram-negative organisms. The emergence and spread of carbapenemase-producing bacteria have created urgent diagnostic challenges for clinical laboratories. Carbapenemases are beta-lactamases that can hydrolyze carbapenems and most other beta-lactam antibiotics, leaving clinicians with limited treatment options for infected patients.
The accurate detection and classification of beta-lactamases, especially carbapenemases, directly influences patient management decisions. When a laboratory identifies a carbapenemase-producing organism, clinicians can adjust therapy to use agents that remain active against the specific resistance mechanism. Conversely, failure to detect these enzymes can lead to treatment failure and poor patient outcomes.
Laboratories face a complex decision when selecting detection methods. Phenotypic tests detect enzyme activity through observable bacterial growth patterns, while molecular tests detect the genetic determinants that encode these enzymes. Each approach has distinct advantages and limitations that must be weighed in the context of laboratory resources, testing volume, and clinical need.
Beta-Lactamase Classification and Mechanisms
Beta-lactamases are grouped into classes based on their molecular structure and functional characteristics. The Ambler classification system divides these enzymes into four classes designated A, B, C, and D. Classes A, C, and D include serine beta-lactamases that use a serine residue at their active site to hydrolyze beta-lactam antibiotics. Class B enzymes are metallo-beta-lactamases that require zinc ions for their catalytic activity.
Class A beta-lactamases include the extended-spectrum beta-lactamases such as TEM and SHV variants that confer resistance to penicillins and cephalosporins. These enzymes are commonly found in Enterobacteriaceae and are often encoded on plasmids that can transfer between bacterial species. Class A also includes some carbapenemases such as KPC, which is a significant cause of carbapenem resistance in healthcare settings.
Class B metallo-beta-lactamases include the NDM, VIM, and IMP families. These enzymes require zinc for activity and can hydrolyze carbapenems and all other beta-lactams except monobactams. Metallo-beta-lactamases are not inhibited by the beta-lactamase inhibitors used in clinical practice, which complicates treatment approaches.
Class C beta-lactamases are AmpC enzymes that are typically encoded on chromosomes but can also be plasmid-mediated. These enzymes confer resistance to cephalosporins and are often inducible in certain bacterial species. Class D beta-lactamases include the OXA-type enzymes, some of which have carbapenemase activity, particularly in Acinetobacter species.
The clinical significance of carbapenemase-producing organisms has driven the development of multiple detection methods. Laboratories must understand the enzyme classes and their properties to select appropriate detection strategies and interpret results correctly.
Phenotypic Detection Methods
Phenotypic methods detect carbapenemase activity by observing the ability of an organism to inactivate carbapenem antibiotics. These methods rely on the growth characteristics of bacteria in the presence of carbapenem disks or strips and are widely used in clinical laboratories because they are relatively inexpensive and do not require specialized equipment.
Modified Carbapenem Inactivation Method
The modified carbapenem inactivation method is a phenotypic test that detects carbapenemase activity in Enterobacteriaceae and Pseudomonas aeruginosa. The test involves incubating a suspension of the test organism with a meropenem disk, then placing the disk on a lawn of a susceptible indicator organism. If the test organism produces a carbapenemase, it inactivates the meropenem, allowing the indicator organism to grow around the disk.
The mCIM procedure requires careful attention to timing and incubation conditions. A loopful of the test organism is suspended in tryptic soy broth, and a meropenem disk is added to the suspension. After a specific incubation period, the disk is removed and placed on a Mueller-Hinton agar plate that has been inoculated with a standardized suspension of the indicator organism. The plate is incubated overnight, and the zone of inhibition around the disk is measured.
Interpretation of the mCIM result depends on the zone diameter. A zone of inhibition that is smaller than the established threshold indicates carbapenemase production. The method has been validated for detection of carbapenemases in Enterobacteriaceae and Pseudomonas aeruginosa, and it is recommended by the Clinical and Laboratory Standards Institute for routine testing.
The mCIM method has several advantages for clinical laboratories. It is inexpensive, requires no specialized equipment, and can be performed in any laboratory that performs routine antimicrobial susceptibility testing. The method detects functional carbapenemase activity instead of the presence of specific genes, which means it can detect novel carbapenemases that may not be targeted by molecular assays.
EDTA-Modified Carbapenem Inactivation Method
The EDTA-modified carbapenem inactivation method (eCIM) is an extension of the mCIM that helps distinguish metallo-beta-lactamases from other carbapenemase types. EDTA is a chelating agent that binds zinc ions, which are required for the activity of metallo-beta-lactamases. When EDTA is added to the test system, metallo-beta-lactamases are inactivated, and the carbapenem antibiotic remains active against the indicator organism.
The eCIM is performed in parallel with the mCIM using the same test organism. A second meropenem disk is incubated with the test organism in a buffer containing EDTA. After incubation, the disk is placed on the indicator plate alongside the mCIM disk. The zone of inhibition around the eCIM disk is compared with the zone around the mCIM disk.
Interpretation of the eCIM requires comparison of the two zone diameters. If the eCIM zone is larger than the mCIM zone by a defined amount, the organism produces a metallo-beta-lactamase. If the zones are similar, the carbapenemase is likely a serine beta-lactamase. This information helps guide treatment decisions because metallo-beta-lactamases have different therapeutic implications than serine carbapenemases.
The eCIM method is most useful when performed in conjunction with the mCIM. It provides additional classification information without requiring molecular testing. However, the method has limitations and may not detect all carbapenemase types, particularly those that produce low levels of enzyme or that have unusual activity profiles.
Other Phenotypic Detection Approaches
Several other phenotypic approaches are available for carbapenemase detection. The modified Hodge test was historically used but has been replaced by more accurate methods in many laboratories. The Carba NP test detects carbapenemase activity by measuring pH changes that occur when carbapenem is hydrolyzed. This test is rapid and can be performed directly on bacterial colonies.
The Carba NP test uses a colorimetric reaction to detect carbapenemase activity. The test organism is mixed with a solution containing imipenem and a pH indicator. If the organism produces a carbapenemase, the antibiotic is hydrolyzed, producing an acidic product that changes the color of the indicator. The test can provide results within a few hours and does not require specialized equipment.
Combination disk tests use disks containing carbapenem alone and carbapenem with a beta-lactamase inhibitor. An increase in zone diameter around the disk with the inhibitor indicates the presence of a beta-lactamase that is inhibited by that compound. These tests can help distinguish between different carbapenemase types.
Each phenotypic method has specific performance characteristics and limitations. Laboratories should select methods that have been validated for the organisms they routinely test and should understand the interpretation criteria for each method.
Molecular Detection Methods
PCR-Based Assays
Molecular detection methods identify the genetic determinants that encode beta-lactamases and carbapenemases. Polymerase chain reaction (PCR) assays are the most widely used molecular methods in clinical laboratories. These assays amplify specific DNA sequences that are unique to particular carbapenemase genes, allowing detection of the gene even when the enzyme is not expressed.
Conventional PCR assays detect a single gene or a small panel of genes in a single reaction. These assays require gel electrophoresis to visualize the amplified products, which adds time and complexity to the workflow. Real-time PCR assays use fluorescent probes to detect amplification as it occurs, providing results more quickly and reducing the risk of contamination.
Multiplex PCR assays can detect multiple carbapenemase genes in a single reaction. These assays are designed to detect the most common carbapenemase genes, including KPC, NDM, VIM, IMP, and OXA-48-like genes. The ability to detect multiple targets simultaneously makes multiplex PCR efficient for laboratories that test large numbers of isolates.
PCR assays provide definitive identification of specific carbapenemase genes, which is valuable for epidemiological surveillance and infection control. Knowing the specific gene type can help track the spread of resistance within a facility or region. This information is also useful for selecting treatment options because some carbapenemase types have different treatment implications.
Whole Genome Sequencing
Whole genome sequencing (WGS) provides the most comprehensive approach to detecting beta-lactamase and carbapenemase genes. This method sequences the entire genome of the bacterial isolate and identifies all resistance genes present. WGS can detect known carbapenemase genes and can also identify novel variants that may not be detected by targeted PCR assays.
WGS has become more accessible in recent years as sequencing costs have decreased and bioinformatics tools have improved. However, the method requires significant technical expertise and computational resources. The turnaround time for WGS is longer than for PCR-based methods, which may limit its use for urgent clinical decisions.
The interpretation of WGS data requires careful analysis of the sequencing results. Resistance genes must be identified and their potential impact on antibiotic susceptibility assessed. The presence of a resistance gene does not always correlate with phenotypic resistance, and the expression of the gene can be influenced by regulatory mechanisms and other factors.
WGS is particularly valuable for surveillance and outbreak investigations. It can identify the specific strain of bacteria and determine whether isolates from different patients are related. This information helps infection control teams understand the transmission of resistant organisms within healthcare settings.
Isothermal Amplification Methods
Isothermal amplification methods amplify nucleic acids at a constant temperature, eliminating the need for thermal cycling equipment. These methods include loop-mediated isothermal amplification (LAMP) and recombinase polymerase amplification (RPA). These techniques can provide rapid results and are suitable for point-of-care testing or laboratories with limited infrastructure.
LAMP assays for carbapenemase detection have been developed for several gene targets. These assays use a set of primers that recognize multiple regions of the target gene, providing high specificity. The amplification can be detected by turbidity, fluorescence, or color change, making the method accessible to laboratories without specialized equipment.
Isothermal methods offer several advantages for resource-limited settings. They do not require expensive thermal cyclers and can be performed with simpler equipment. The results can be obtained in a shorter time than traditional PCR, which may be important for clinical decision-making.
The main limitation of isothermal amplification methods is the need for careful primer design and validation. The specificity of these assays depends on the primer sequences, and cross-reactivity with related genes can produce false-positive results. Laboratories must validate these methods for their specific testing needs.
Comparison of Detection Methods
Performance Characteristics
The performance of carbapenemase detection methods varies depending on the organism being tested and the specific carbapenemase gene present. Phenotypic methods detect functional enzyme activity, while molecular methods detect the presence of specific genes. Each approach has different strengths and limitations that must be considered when selecting a detection strategy.
Phenotypic methods such as the mCIM have high sensitivity for detecting carbapenemase activity in Enterobacteriaceae. These methods can detect carbapenemases that are expressed at sufficient levels to produce a positive result. However, they may miss carbapenemases that are expressed at low levels or that have reduced activity against the specific carbapenem used in the test.
Molecular methods have high specificity for the genes they are designed to detect. A positive PCR result indicates the presence of the target gene, but it does not confirm that the gene is expressed and producing functional enzyme. Some isolates may carry carbapenemase genes that are not expressed, leading to a discrepancy between molecular and phenotypic results.
The sensitivity of molecular methods depends on the panel of genes included in the assay. If an isolate produces a carbapenemase that is not included in the panel, the molecular test will be negative even though the organism is resistant. This is a significant limitation of targeted molecular assays.
Workflow and Turnaround Time
The workflow for carbapenemase detection varies significantly between phenotypic and molecular methods. Phenotypic methods such as the mCIM require an overnight incubation step, which means results are not available until the day after the test is set up. This turnaround time may be acceptable for routine testing but can delay treatment decisions for critically ill patients.
Molecular methods can provide results in a few hours, depending on the specific assay and the availability of equipment. Real-time PCR assays can be completed in approximately two to three hours after the DNA extraction step. This faster turnaround time can be important for patients with serious infections where early identification of carbapenemase production may influence treatment.
The workflow for molecular testing requires additional steps compared with phenotypic methods. DNA extraction must be performed before the amplification reaction, and the quality of the DNA can affect the results. Laboratories must also have the appropriate equipment and trained personnel to perform molecular testing.
The choice between phenotypic and molecular methods often depends on the laboratory resources and the clinical need. Laboratories with high testing volumes may benefit from the efficiency of molecular methods, while smaller laboratories may prefer the lower cost of phenotypic methods.
Cost Considerations
The cost of carbapenemase detection methods varies widely. Phenotypic methods such as the mCIM are inexpensive and use materials that are already available in most clinical laboratories. The cost of these methods is primarily the labor time required to perform the test and interpret the results.
Molecular methods require more expensive reagents and equipment. The cost of PCR reagents, DNA extraction kits, and consumables can be significant, especially for laboratories that test a large number of isolates. The initial investment in equipment such as thermal cyclers or real-time PCR instruments can also be substantial.
The cost-effectiveness of different methods depends on the testing volume and the clinical impact of the results. For laboratories that test a small number of isolates, the lower cost of phenotypic methods may be more appropriate. For laboratories with high testing volumes, the efficiency of molecular methods may justify the higher cost.
Laboratories must also consider the cost of false-positive and false-negative results. A false-negative result can lead to treatment failure and poor patient outcomes, while a false-positive result can lead to unnecessary use of alternative antibiotics. The cost of these outcomes should be considered when selecting a detection method.
Laboratory Implementation and Quality Control
Method Selection and Validation
The selection of a carbapenemase detection method should be based on the specific needs of the laboratory and the patient population it serves. Laboratories should consider the types of organisms they encounter, the testing volume, the available resources, and the clinical need for rapid results. The method should be validated for the organisms and specimen types that will be tested.
Validation of a new detection method requires testing against a panel of well-characterized isolates. The panel should include isolates with known carbapenemase genes, isolates without carbapenemase genes, and isolates with other resistance mechanisms. The performance of the new method should be compared with a reference method to determine its sensitivity and specificity.
The Clinical and Laboratory Standards Institute provides standards for antimicrobial susceptibility testing and related methods. These standards describe the procedures for performing and interpreting susceptibility tests, including carbapenemase detection methods. Laboratories should follow these standards to ensure the reliability of their results.
Quality control is essential for all carbapenemase detection methods. Laboratories should test control strains with known carbapenemase activity and known negative strains with each batch of tests. The results of the quality control tests should be documented and reviewed to ensure the method is performing correctly.
Quality Control and Assurance
Quality control for carbapenemase detection methods includes the use of reference strains with known results. The American Type Culture Collection provides authenticated biological materials that can be used for quality control. These strains have been characterized and can be used to verify that the detection method is working correctly.
The quality control strains should be tested in the same manner as the clinical isolates. The results should be compared with the expected results for each strain. If the quality control results do not match the expected results, the test should be repeated and the problem investigated.
Quality assurance also includes the documentation of all testing procedures and results. The laboratory should maintain records of the quality control results, the test results, and any problems that occur. These records are important for identifying trends and for demonstrating the reliability of the testing process.
The laboratory should also participate in external quality assessment programs to verify the accuracy of its testing. These programs provide samples with known results that are tested by the laboratory and compared to the results of other laboratories. Participation in these programs can identify problems with the testing process and help improve the quality of the results.
Training and Competency
The personnel who perform carbapenemase detection tests must be properly trained and demonstrate competency in the testing procedures. Training should include the principles of the method, the steps of the procedure, and the interpretation of the results. The training should also cover the safety precautions that are required when handling bacterial cultures.
Competency assessment should be performed on a regular basis to ensure that the personnel can perform the test correctly. The assessment should include the performance of the test on known samples and the interpretation of the results. The results of the competency assessment should be documented and reviewed.
The American Society for Microbiology provides educational resources and training opportunities for microbiology professionals. These resources can help laboratory personnel stay current with the latest methods and best practices for carbapenemase detection. The ASM also provides guidance on laboratory practice and workforce development.
The laboratory should also have procedures for the review of the results by a qualified professional. The results of carbapenemase detection tests should be reviewed by a clinical microbiologist or other qualified individual before they are reported. This review can help identify errors and ensure that the results are interpreted correctly.
Common Failure Patterns and Troubleshooting
False-Negative Results
False-negative results occur when a carbapenemase-producing organism is not detected by the test. This can happen for several reasons. The organism may produce a carbapenemase that is not detected by the method being used, or the enzyme may be expressed at a level that is below the detection threshold of the test.
For phenotypic methods, false-negative results can occur when the carbapenemase has low activity against the specific carbapenem used in the test. Some carbapenemases have different activity profiles against different carbapenems, and the test may not detect the enzyme if the activity is low. The test conditions, such as the incubation time and the inoculum size, can also affect the sensitivity of the method.
For molecular methods, false-negative results can occur when the isolate contains a carbapenemase gene that is not included in the assay panel. The molecular test can only detect the genes that are targeted by the primers or probes. If the isolate produces a different carbapenemase, the test will be negative.
The laboratory should be aware of the limitations of the method and should consider the possibility of false-negative results. If the clinical presentation suggests a carbapenemase-producing organism but the test is negative, the laboratory should consider using a different method or referring the isolate to a reference laboratory.
False-Positive Results
False-positive results occur when the test indicates the presence of a carbapenemase in an organism that does not produce the enzyme. This can occur when the test is not specific for carbapenemase activity or when the test detects a gene that is present but not expressed.
For phenotypic methods, false-positive results can occur when the organism has other resistance mechanisms that produce a similar result. For example, the combination of a beta-lactamase and a porin mutation can reduce the susceptibility to carbapenems, which may be misinterpreted as carbapenemase activity. The test conditions can also affect the specificity of the method.
For molecular methods, false-positive results can occur when the test detects a gene that is present but not expressed. The presence of a carbapenemase gene does not always mean that the enzyme is produced and functional. The gene may be present in a form that is not expressed, or the expression may be regulated by other factors.
The laboratory should be aware of the possibility of false-positive results and should confirm the results with a different method when necessary. The clinical significance of a positive result should be considered in the context of the patient's condition and the results of other tests.
Interpretation Challenges
The interpretation of carbapenemase detection results can be challenging, especially when the results are borderline or when the different methods provide conflicting results. The laboratory must have clear criteria for the interpretation of the results and should document the criteria in the standard operating procedures.
The interpretation of the mCIM and eCIM results requires careful measurement of the zone diameters. The zones should be measured using a consistent method, and the results should be compared to the established thresholds. The interpretation of the results should be performed by a trained individual who is familiar with the method.
The interpretation of molecular results requires consideration of the genes that are detected and the potential impact of the genes on the resistance phenotype. The presence of a carbapenemase gene should be considered in the context of the antimicrobial susceptibility testing results. The laboratory should also consider the possibility of multiple resistance mechanisms in the same organism.
The laboratory should have a process for the review of the results and the resolution of discrepancies. When the results of different methods are conflicting, the laboratory should investigate the cause of the discrepancy and determine the most reliable result. The results should be reported with the appropriate comments and limitations.
Laboratory Workflow and Decision Making
Step-by-Step Implementation
The implementation of a carbapenemase detection program requires a systematic approach. The laboratory should first assess its needs and resources, then select the appropriate method, and then validate the method for its specific use. The following steps provide a framework for the implementation.
The first step is to assess the laboratory's testing needs. This includes the types of organisms that are tested, the volume of testing, and the clinical impact of the results. The laboratory should also consider the available equipment and the expertise of the personnel.
The second step is to select the detection method. The laboratory should compare the available methods and select the one that best meets its needs. The selection should consider the performance of the method, the workflow, and the cost.
The third step is to validate the method. The laboratory should test the method against a panel of well-characterized isolates to ensure that it provides reliable results. The validation should include the assessment of the sensitivity and specificity of the method.
The fourth step is to implement the method in the routine workflow. The laboratory should develop standard operating procedures and train the personnel. The method should be integrated into the routine testing process.
The fifth step is to monitor the performance of the method. The laboratory should perform quality control testing and participate in the external quality assessment. The results should be reviewed to identify any problems and to improve the testing process.
Biosafety Considerations
The testing of carbapenemase-producing organisms requires appropriate biosafety precautions. The organisms that produce carbapenemases are often multidrug-resistant and can be difficult to treat. The laboratory should follow the biosafety guidelines for the handling of these organisms.
The Biosafety in Microbiological and Biomedical Laboratories (BMBL) provides guidelines for the safe handling of microorganisms in the laboratory. The guidelines describe the biosafety levels and the practices that are required for each level. The laboratory should determine the appropriate biosafety level for the organisms it handles and follow the required practices.
The laboratory should use appropriate personal protective equipment, including gloves, lab coats, and eye protection. The work should be performed in a biosafety cabinet when there is a risk of aerosolization. The laboratory should also have procedures for the decontamination of the work surfaces and the disposal of the waste.
The laboratory should also have procedures for the handling of the specimens and the cultures. The specimens should be transported and stored in a safe manner. The cultures should be handled in a way that minimizes the risk of exposure to the laboratory personnel.
Professional Escalation Criteria
The laboratory should have criteria for the escalation of the results to the appropriate professionals. When a carbapenemase-producing organism is detected, the laboratory should notify the clinical team and the infection control team. The notification should include the results of the testing and the implications for the patient management.
The laboratory should also have criteria for the referral of the isolates to a reference laboratory. When the laboratory cannot identify the carbapenemase or when the results are uncertain, the isolate should be referred to a reference laboratory for further testing. The reference laboratory can perform additional tests to identify the specific carbapenemase and to provide guidance for the treatment.
The escalation criteria should be defined in the standard operating procedures. The laboratory should have a clear process for the notification of the results and the referral of the isolates. The process should be reviewed regularly to ensure that it is effective.
At a Glance
| Detection Method | Principle | Time to Result | Key Advantage | Key Limitation |
|---|---|---|---|---|
| Modified Carbapenem Inactivation Method (mCIM) | Phenotypic detection of carbapenemase activity | Overnight | Low cost, no specialized equipment | Cannot identify specific carbapenemase type |
| EDTA-Modified Carbapenem Inactivation Method (eCIM) | Phenotypic detection with EDTA to distinguish metallo-beta-lactamases | Overnight | Distinguishes metallo-beta-lactamases from serine beta-lactamases | Requires parallel testing with mCIM |
| PCR-Based Assays | Molecular detection of specific carbapenemase genes | 2 to 4 hours | High sensitivity for targeted genes, rapid results | Detects only genes in the panel, requires specialized equipment |
| Whole Genome Sequencing | Molecular detection of all resistance genes | 1 to 3 days | Comprehensive detection of known and novel genes | High cost, requires bioinformatics expertise |
Practical Implementation Steps
The implementation of a carbapenemase detection program should follow a structured approach that includes the assessment of the laboratory needs, the selection of the method, the validation of the method, and the integration of the method into the routine workflow.
The laboratory should first assess the types of organisms it tests and the clinical impact of the results. This assessment should include the review of the local epidemiology of carbapenemase-producing organisms and the needs of the clinical team. The laboratory should also consider the available resources, including the equipment and the personnel.
The selection of the method should be based on the assessment of the laboratory needs. The laboratory should compare the available methods and select the one that provides the best balance of performance, cost, and workflow. The laboratory should also consider the need for the classification of the carbapenemase type, which may require the use of the eCIM or molecular methods.
The validation of the method should be performed using a panel of well-characterized isolates. The panel should include isolates with known carbapenemase genes and isolates without carbapenemase genes. The validation should assess the sensitivity and specificity of the method and should be documented in the laboratory records.
The integration of the method into the routine workflow should include the development of the standard operating procedures and the training of the personnel. The method should be incorporated into the routine testing process, and the results should be reported in a timely manner. The laboratory should also establish the process for the escalation of the results to the clinical team.
Common Failure Patterns
The most common failure patterns in carbapenemase detection include the use of the methods that are not validated for the organisms being tested, the misinterpretation of the results, and the lack of the appropriate quality control. These failures can lead to incorrect results and can have a significant impact on the patient management.
The use of the method that is not validated for the organisms being tested can lead to false-negative or false-positive results. The laboratory should validate the method for the types of organisms that it tests. The validation should include the assessment of the performance of the method for the specific organisms.
The misinterpretation of the results can occur when the laboratory does not follow the interpretation criteria. The laboratory should have clear criteria for the interpretation of the results and should train the personnel in the interpretation. The results should be reviewed by a qualified individual before they are reported.
The lack of the appropriate quality control can lead to the failure of the method. The laboratory should test the quality control strains with each batch of testing. The quality control results should be reviewed to ensure that the method is working correctly.
The laboratory should also be aware of the limitations of the methods and should consider the possibility of the false results. When the results are not consistent with the clinical presentation, the laboratory should investigate the cause and consider the use of the alternative methods.
Safety and Regulatory Context
The testing of carbapenemase-producing organisms is performed in the context of the broader antimicrobial resistance surveillance and stewardship efforts. The World Health Organization identifies antimicrobial resistance as a global health concern and recommends the surveillance and stewardship across the human health, veterinary, and environmental sectors. The laboratory testing for carbapenemase is an important component of the surveillance and the stewardship.
The laboratory should follow the biosafety guidelines for the handling of the organisms. The CDC Biosafety in Microbiological and Biomedical Laboratories provides the guidelines for the safe handling of the microorganisms. The laboratory should implement the appropriate biosafety practices for the level of the risk.
The laboratory should also follow the standards for the antimicrobial susceptibility testing. The Clinical and Laboratory Standards Institute provides the standards for the testing and the interpretation of the results. The laboratory should follow these standards to ensure the reliability of the results.
The laboratory should also participate in the quality assurance programs and the ongoing assessment. The participation in the external quality assessment programs can help the laboratory identify the weaknesses in the testing and improve the reliability of the results.
Frequently Asked Questions
What is the difference between the mCIM and the eCIM?
The mCIM detects the carbapenemase activity in the organism, while the eCIM is performed in parallel with the mCIM and uses EDTA to distinguish the metallo-beta-lactamases from the serine beta-lactamases. The eCIM result is interpreted by comparing the zone of inhibition with the mCIM result.
When should a laboratory use a molecular method instead of a phenotypic method?
A laboratory should use a molecular method when the rapid results are needed for the patient management or when the identification of the specific carbapenemase gene is important for the epidemiological surveillance. The molecular methods can also be used to confirm the results of the phenotypic methods.
What are the limitations of the phenotypic methods for carbapenemase detection?
The phenotypic methods detect the functional activity of the carbapenemase, but they do not identify the specific gene. The methods can also produce false-negative results when the enzyme is produced at a low level or when the enzyme has a low activity against the carbapenem used in the test.
What are the limitations of the molecular methods for carbapenemase detection?
The molecular methods detect the presence of the specific genes, but they do not confirm that the gene is expressed and produces the functional enzyme. The methods can also produce false-negative results when the isolate produces a carbapenemase that is not included in the assay panel.
How should a laboratory validate a carbapenemase detection method?
The laboratory should validate the method using a panel of well-characterized isolates that includes the isolates with known carbapenemase genes and the isolates without the carbapenemase genes. The validation should assess the sensitivity and the specificity of the method.
What quality control is required for carbapenemase detection?
The laboratory should test the quality control strains with each batch of the testing. The quality control strains should have the known results and should be tested in the same manner as the clinical isolates. The quality control results should be reviewed to ensure that the method is working correctly.
When should a laboratory refer an isolate to a reference laboratory?
The laboratory should refer the isolate to a reference laboratory when the testing results are not clear or when the isolate is suspected to produce a carbapenemase that cannot be identified by the laboratory. The reference laboratory can provide the additional testing and the guidance for the management.
What biosafety precautions are required for the testing of carbapenemase-producing organisms?
The laboratory should follow the biosafety guidelines for the handling of the organisms. The work should be performed in a biosafety cabinet when there is a risk of aerosolization. The laboratory should use the appropriate personal protective equipment and should have the procedures for the decontamination and the waste disposal.
Using the Evidence
| Source | Best use in this topic | Important limitation |
|---|---|---|
| Biosafety in Microbiological and Biomedical Laboratories | official guidance | Check the linked page for current local requirements |
| Microbiology Resources | official guidance | Check the linked page for current local requirements |
| Antimicrobial Resistance | official guidance | Check the linked page for current local requirements |
Related Bacteriology Guides
- Streptococcus agalactiae in Farmed Tilapia: Clinical Outbreaks and Molecular Detection
- Duck Viral Enteritis: Clinical Diagnosis and Management
- Duck Viral Enteritis: Clinical Signs and Management
- Duck Viral Enteritis: Clinical Disease and Management
- Avian Coryza in Chickens: Clinical Signs, Diagnosis, and Management
References and Further Reading
- Biosafety in Microbiological and Biomedical Laboratories. CDC and NIH.
- Microbiology Resources. American Society for Microbiology.
- Antimicrobial Resistance. World Health Organization.
- Bacteriological Analytical Manual. U.S. Food and Drug Administration.
- Antimicrobial Susceptibility Testing. Clinical and Laboratory Standards Institute.
- Culture Collections. American Type Culture Collection.
- NCBI Pathogen Detection. National Center for Biotechnology Information.
- Klebsiella pneumoniae carbapenemase variants: the new threat to global public health.. Clinical microbiology reviews, 2023.
- Beta-lactamase and carbapenemase detection methods: an overview of recent patents.. Recent patents on anti-infective drug discovery, 2014.
- Carbapenemase-producing Pseudomonas aeruginosa -an emerging challenge.. Emerging microbes & infections, 2022.
- Laboratory detection of carbapenemases among Gram-negative organisms.. Clinical microbiology reviews, 2024.
This article is educational and does not replace validated laboratory procedures, clinical diagnosis, institutional biosafety review, or public-health guidance.