bluuebunny/crossref_metadata_2025_split huggingface dataset:
Broth Microdilution Test
Authors: Gamage Dulini , Chandrasekaran Sukantha , Carpenter Darcie E. 
Published: January 2025

Abstract

The broth microdilution MIC method is used to measure the
in vitro
activity of an antimicrobial agent against a bacterial isolate. A sterile multiwell plastic tray containing various concentrations of antimicrobialagents is inoculated with a standardized number of test bacteria. MICs obtained are interpreted as susceptible, susceptible dose‐dependent, intermediate, resistant, or nonsusceptible based on the criteria listed in the CLSI MIC standard. Infections caused by both staphylococci and streptococci with inducible clindamycin resistance may fail to respond to clindamycin therapy. Thymidine can interfere with performance of sulfonamides and trimethoprim. For
Streptococcus pneumoniae
with trimethoprim and sulfonamides Antagonists in the medium may allow some slight growth.
Gradient Diffusion Tests
Authors: Dwivedi Hari P. , Brocco Fabio , Matuschek Erika 
Published: January 2025

Abstract
A standardized inoculum of bacteria is swabbed onto the surface of a low‐thymine‐thymidine‐concentration, cation‐adjusted Mueller‐Hinton agar plate, in accordance with the ISO 16782 standard. Gradient diffusion test strips, i.e., strips containing a continuous gradient of antimicrobial concentrations, are placed on the agar surface. After overnight incubation, an elliptical zone of inhibition forms as the antimicrobial agent inhibits growth. The MIC is read where growth intersects the gradient test strip. MICs can be interpreted in categories based on the breakpoints established by breakpoint‐establishing agencies such as the Food and Drug Administration (FDA), the CLSI, or the European Committee on Antimicrobial Susceptibility Testing (EUCAST). The gradient test strips are commercial antimicrobial gradient products and are not specifically addressed in CLSI or EUCAST standards. The CLSI does acknowledge FDA‐cleared antimicrobial gradient tests that perform comparably to CLSI reference methods as acceptable for use in clinical laboratories.
Detection of VRSA, VISA, and Vancomycin‐Heteroresistant
Staphylococcus aureus
(hVISA)
Authors: Morrisette Taylor , Rybak Michael J. 
Published: January 2025

Abstract

Vancomycin‐resistant
Staphylococcus aureus
(VRSA) and vancomycin‐intermediate
S. aureus
(VISA) can be detected using routine broth microdilution methods (BMD); the vancomycin agar screen test is another available method. Detection of VISA and VRSA isolates should preferentially be performed by BMD, agar dilution, or gradient diffusion methods. The detection of vancomycin heteroresistance in
S. aureus
(hVISA) is challenging and labor‐intensive, and it is complicated in the clinical realm by a lack of routine screening procedures. The presence of hVISA in clinical specimens has been associated with vancomycin treatment failures in some studies, but this is poorly validated clinically. The PAP‐AUC ratio is considered the gold standard method for detection of hVISA. The chapter covers proper quality control principles for analytical tests. With postanalytical considerations, it provides attention to common test limitations, interpretive challenges, and results reporting.
Agar Screen Test to Detect Vancomycin Resistance in
Enterococcus
spp.
Authors: Maris Alexander S. , Tao Lili 
Published: January 2025

Abstract
This chapter describes agar dilution screen test in detail. The vancomycin agar screen is used to detect vancomycin‐resistant enterococcal colonies that have been isolated from clinical or surveillance cultures. MIC and species identification tests are subsequently required to determine if the isolate is a true vancomycin‐resistant enterococcus (VRE). VRE isolates possess acquired vancomycin resistance determinants that may be horizontally transferred with infection prevention implications; such VRE isolates contrast with isolates possess chromosomally mediated intrinsic low‐level vancomycin resistance, which does not pose infection prevention concerns. The chapter covers proper quality control principles for analytical tests. With postanalytical considerations, it provides attention to common test limitations, interpretive challenges, and results reporting.
Synergy Testing: Broth Microdilution and Broth Macrodilution Checkerboard Methods
Authors: Doern Christopher D. , Van Belkum Alex 
Published: January 2025

Abstract

Although uncommon, microbiology laboratories may use a two‐dimensional, two‐agent broth microdilution checkerboard to evaluate combinations of antimicrobial agents against microorganisms. A broth macrodilution limited series, agar dilution method, gradient diffusion, or disk diffusion method can also be used, although filter‐based methods have been described as well. Test methods are based on Clinical and Laboratory Standards Institute and European Committee on Antimicrobial Susceptibility Testing broth dilution susceptibility methods for evaluating the inhibitory or bactericidal activity of specific concentrations in combination at a fixed time.
In vitro
interactions are calculated algebraically and interpreted as synergistic, indifferent, or antagonistic depending on whether the antibacterial activity of the combination is greater than, equivalent to, or less than the activities of the individual agents, respectively.
In vitro
interactions can also be represented geometrically with isobolograms. Synergism testing between more formal antibiotics and natural compounds is increasingly reported.
Evaluation and Verification of Antimicrobial Susceptibility Test Systems
Authors: Bobenchik April M. 
Published: January 2025

Abstract
A wide variety of antimicrobial susceptibility test systems are available to clinical microbiologists. This procedure provides guidance on evaluating and choosing a system or method appropriate for the laboratory. The primary focus should be on determination of the systems' capabilities, assessment of costs and workflow, and examination of performance data. This chapter describes a procedure for analyzing results generated from evaluating a new system in comparison with those generated from a reference system. Some of the suggestions in this protocol can be considered to fulfill the verification requirements. The chapter covers proper quality control principles for analytical tests. With postanalytical considerations, it provides attention to common test limitations, interpretive challenges, and results reporting.
Selecting Antimicrobial Agents for Testing and Reporting
Authors: Bobenchik April M. , Simner Trish 
Published: January 2025

Abstract
Developing protocols for antimicrobial susceptibility testing (AST) and reporting is best done by the clinical laboratory with input from the antimicrobial stewardship program, infectious disease service, infection control, and the pharmacy and therapeutics committee, as available at a given institution. The goals are to provide clinically relevant information that will support appropriate and cost‐effective utilization of antimicrobial agents to avoid reporting results that may adversely affect patient care or public health. Selective reporting is a reporting strategy that reports certain AST results based on defined criteria such as organism identification, body site, or patient demographics. Cascade reporting involves reporting results for specific agents based on the overall antimicrobial susceptibility profile of an isolate. These reporting techniques may withhold (i.e., suppress) some AST results from the final report provided to clinical end users, in part to support antimicrobial stewardship efforts.
Susceptibility Testing
Authors: Sussland Zdenek , Saubolle Michael A. 
Published: January 2025

Abstract

The Clinical and Laboratory Standards Institute has published guideline M24, and its supplement M62, which describe standard methods for appropriate
in vitro
antimicrobial susceptibility testing against aerobic actinomycetes. The broth microdilution method, providing MICs, is the method recommended for susceptibility testing of the aerobic actinomycetes, including the
Nocardia
spp.,
Rhodococcus
spp.,
Tsukamurella
spp.,
Gordonia
spp.,
Streptomyces
spp., and
Actinomadura
. These procedures have been primarily used for the most commonly isolated species of the
Nocardia
genus but may be cautiously used for other clinically relevant genera of aerobic actinomycetes. Commercially prepared microdilution plates with the appropriate antimicrobial agents are available but have not yet been approved by the Food and Drug Administration. A disk diffusion test with sulfisoxazole may be used only to verify sulfonamide MIC results and to confirm concentration of organisms in the inoculum.
Solid Media Used for Isolation
Authors: Wengenack Nancy L. 
Published: January 2025

Abstract

Both liquid and solid media are recommended for optimal recovery of mycobacteria in culture. Pure cultures of mycobacteria, particularly
Mycobacterium tuberculosis
, are still required, despite the availability of molecular technology, to obtain phenotypic antimicrobial susceptibility test results and for strain typing. The advantage of solid medium is that it enables the detection of mixed cultures and contaminants. Egg‐based or agar‐based media may be used. The main advantage of an egg‐based medium is that it best supports the growth of
M. tuberculosis
and permits niacin testing, if needed. The advantages of agar‐based media include less contamination and earlier and easier visibility of colonial morphology. Colonial morphology can aid in the identification of mycobacteria. Use of both nonselective and selective media is needed for optimal mycobacterial isolation, the latter containing one or more antimicrobial agents to prevent overgrowth by contaminating bacteria or fungi.
Susceptibility Testing by the Agar Proportion Method for
Mycobacterium tuberculosis
Complex
Authors: Escuyer Vincent E. , Salfinger Max 
Published: January 2025

Abstract

Antimicrobial susceptibility testing should be performed on each patient's initial
Mycobacterium tuberculosis
complex isolate, in relapse or retreatment cases and when acquired drug resistance is suspected. The modified agar proportion method is used to determine the susceptibility of
M. tuberculosis
complex, indirectly from a pure culture. Quadrant plates are used that contain either Middlebrook 7H10 agar without antimicrobial agents or 7H10 agar with antimicrobial agents at various concentrations. Agar proportion testing compares the growth of an isolate on solid agar medium with and without antimycobacterial drug. The concentrations of the drugs tested are based on the recommendations of the CLSI. The agar proportion method also allows the determination of the proportion or percentage of resistance among the tested isolates. This chapter covers proper quality control principles for analytical tests. With analytical considerations, it provides attention to common test limitations, interpretive challenges, and results reporting.
Fungal Taxonomy and Nomenclature
Authors: Lockhart Shawn R. , Wiederhold Nathan P. 
Published: January 2025

Abstract
Defining the rules for fungal taxonomy is an evolving process. Changes in nomenclature continue to have significant implications for understanding fungi in the environment, fungal disease, and the treatment of infections. Fungi differ from bacteria in that many of them are capable of undergoing a sexual cycle and multiplying through meiotic division. They can exist in two states: an asexual state that divides only through mitotic division, called the anamorph state, and a sexual state that divides by meiotic division, called the teleomorph state. As most fungal infections are caused by a single isolate, mating in the laboratory is rare and the anamorph state is most often identified. However, on rare occasions a homothallic fungus is isolated and the teleomorph state is identified. The Clinical and Laboratory Standards Institute is working on a standard for the incorporation of bacterial and fungal taxonomy in the clinical laboratory.
Specimen Examination
Authors: Abdolrasouli Alireza , Howell Susan 
Published: January 2025

Abstract

The microscopic examination of clinical specimens for the presence of fungi plays an important part in the laboratory diagnosis of most fungal infections. Microscopic examination may provide a rapid indication of the cause of an infection, allowing the prompt initiation of appropriate antifungal therapy. An initial macroscopic examination of the specimen is also important. When the specimen is too small to allow full examination, culture usually takes precedence over microscopic examination due to the higher sensitivity and specificity of culture. Pretreatment of clinical specimens is often required in order to concentrate fungal cells or to see the fungal material more clearly.
Pneumocystis jirovecii
can be detected microscopically in a variety of respiratory specimens including induced sputum, bronchoalveolar lavage fluid, tracheal aspirate, and bronchial brush. Both
P. jirovecii
‐positive and
P. jirovecii
‐negative but yeast‐positive material should be used for immunospecific staining.
Clinical Breakpoints and Epidemiological Cutoff Values
Authors: Dufresne Philippe J. , Fuller Jeff 
Published: January 2025

Abstract

Interpreting the value of
in vitro
susceptibility test results relies not only on the quality and standardization of test methods but also on the quality and standardization of the methods used to derive interpretive thresholds. Clinical breakpoints (CBPs) are developed from a collection of
in vitro
and
in vivo
data so that the interpretation of a susceptibility test result correlates with the likelihood of a good patient outcome. Epidemiological cutoff values, which categorize MIC values as wild‐type or non‐wildtype, are developed strictly from
in vitro
MIC data and correlate with the likelihood of a cultured isolate harboring an acquired mechanism of resistance to the tested antimicrobial agent. Clinical and Laboratory Standards Institute CBPs have been developed for the echinocandins, fluconazole, and voriconazole against the more common species of
Candida
as well as for voriconazole against
Aspergillus fumigatus
. EUCAST provides CBPs for yeasts and molds for susceptibility testing performed according to its broth microdilution protocol.
Additional Techniques for Stool Examination
Authors: Shawar Ribhi , Garcia Lynne S. , Johnson Lisa 
Published: January 2025

Abstract

Agar plate cultures are recommended for the recovery of
Strongyloides stercoralis
larvae and tend to be more sensitive than some of the other diagnostic methods.
Strongyloides stercoralis
larvae are usually the only larvae found in stool specimens. Occasionally, it is necessary to examine stool specimens for scolices and proglottids of cestodes and for adult nematodes and trematodes to confirm the diagnosis and/or to identify a species. AimTab may provide clinically useful information about carbohydrate metabolism by semiquantitatively determining the amount of reducing substance in stool. Harada‐Mori Technique employs a filter paper to which fecal material is added and a test tube into which the filter paper is inserted. The qualitative fecal fat test is used to screen for malabsorption (e.g., due to intestinal inflammation) and maldigestion (e.g., due to pancreatic insufficiency) of dietary fat.
Introduction: Traditional Methods for Laboratory Diagnosis of Viral and Chlamydial Infections
Authors: Hodinka Richard L. 
Published: January 2025

Abstract

This part introduction presents an overview of the key concepts discussed in the subsequent chapters. The part focuses on some of the more traditional culture and rapid immunologic methods for the detection and identification of viruses in a diagnostic laboratory. It includes
Chlamydia
culture given that this method is virtually always performed in a clinical virology laboratory or a distinct section of the clinical microbiology laboratory due to the need for a cell culture system and appropriate facilities. Traditional methods continue to have a place in regional and national reference laboratories to help identify and characterize new viruses or variants of well‐recognized viruses not detected by molecular methods, for active surveillance to detect and monitor disease outbreaks, and in support of research activities for antiviral drug and vaccine development.
Immunoassays for Detection of Infectious Diseases
Authors: Theel Elitza S. 
Published: January 2025

Abstract

Serologic testing for the detection of antibodies developed against, or antigens released from, human pathogens continues to play an important diagnostic role for numerous infectious agents, ranging from spirochetes to arboviruses, fungi, parasites, and mycobacteria, among others. Serologic testing, including both contemporary methods, such as chemiluminescence or multiplex flow immunoassays, and classic techniques, such as immunodiffusion and complement fixation assays, remains an integral part of the infectious disease diagnostics portfolio. While commercially available serologic assays are numerous, the clinical utility of many of these methods is rather limited. For example, while many manufacturers offer immunoassays for detection of antibodies against
Helicobacter pylori
,
Chlamydia trachomatis
, and
Candida
spp. and anti‐HSV IgM, among others, given the availability of direct detection methods for these pathogens, with high sensitivity and specificity, serologic testing no longer plays a relevant diagnostic role and should be avoided. This chapter focuses on serologic assays where result interpretation may be challenging.
Quality Assessment and Improvement (Quality Assurance)
Authors: Kehl Sue , Harriott Melphine , Samuel Linoj , Culbreath Karissa , Thompson Cecilia 
Published: January 2025

Abstract
A quality management system is an integrated system of laboratory activities to ensure quality and continued improvement throughout the testing process. Quality assessment employs internal and external monitors or audits to verify the quality of laboratory processes. Continual improvement is the process whereby opportunities are identified, either through assessments or other means, and strategies are developed to improve quality. Proficiency testing or external quality assessment is an important component of a laboratory's external assessments. A central component of nonconforming event management is root cause analysis. This is usually limited to high‐risk, high‐cost, high‐volume events or events associated with adverse patient outcome. Clinical microbiology laboratories can benefit from specific feedback on microbiology services and incorporate any documented quality deficiencies into the quality improvement program. A thorough self‐review of the accreditation requirements and examination of protocols can improve quality by identifying areas of nonconformance and can allow the laboratory to correct deficiencies prior to the external inspection.
Verification and Validation of Test System Performance Specifications
Authors: Martin Rebekah M. , Loeffelholz Michael 
Published: January 2025

Abstract
Clinical laboratory testing in the United States is regulated by the Centers for Medicare and Medicaid Services through the Clinical Laboratory Improvement Amendments (CLIA). CLIA provides quality standards for clinical laboratory testing in order to ensure patient safety and improve patient care. Two critical components of quality systems in the laboratory are verification and validation of test system performance. This procedure will provide guidance for clinical microbiology laboratories in the United States for designing and performing verification and validation procedures. CLIA regulations state that nonwaived test systems which are either Food and Drug Administration cleared or approved must undergo verification of performance specifications by the clinical laboratory prior to reporting patient results with that system. The verification process must assess precision, accuracy, reportable range, and reference intervals of the assay where applicable. The validation process must assess precision, accuracy, reportable range, reference intervals, analytical sensitivity, and analytical specificity of the assay where applicable.
Bacillus anthracis
and
Bacillus cereus
biovar anthracis—Anthrax
Authors: Lee Philip A. , Rowlinson Marie‐Claire 
Published: January 2025

Abstract

Bacillus anthracis
, the agent of anthrax, causes a zoonotic disease that is transmissible to humans, most often through consumption or handling of contaminated products. It is an aerobic, spore‐forming, nonmotile, large Gram‐positive bacterium. Phylogenetically related to
B. anthracis
,
Bacillus cereus
biovar anthracis was initially recognized in the early 2000s in Cameroon and Côte d'Ivoire. While rare in the United States,
B. anthracis
causes sporadic outbreaks in wildlife and livestock. This presents an environmental and occupational exposure risk to ranchers and veterinarians. Laboratory in‐house testing includes Gram staining and culturing from the following: lower respiratory tract specimens, wound specimens, and blood cultures. The chapter covers proper quality control principles for analytical tests. With postanalytical considerations, it provides attention to common test limitations, interpretive challenges, and results reporting.
Toxins: Botulinum Toxin (
Clostridium botulinum
) and Staphylococcal Enterotoxin B (
Staphylococcus aureus
)
Authors: Egan Christina , Perry Michael J. 
Published: January 2025

Abstract

Pathogenic microorganisms can contain genes that allow toxin production. Two toxin‐producing organisms are
Clostridium botulinum
and
Staphylococcus aureus
.
C. botulinum
is a group of anaerobic organisms, commonly found in soil and aquatic habitats throughout the world. Staphylococcal enterotoxin B (SEB) is one of several exotoxins that may be produced by
S. aureus
. Depending on the strain,
S. aureus
can produce a variety of extracellular proteins that act as virulence factors, including toxic shock syndrome toxin 1, exfoliative toxins A and B, and a family of structurally related proteins classified as enterotoxins. Testing for SEB may be performed by clinical laboratories with appropriate protocols and safety mitigation. The chapter covers safety considerations, specimen collection, and specimen processing, incubation and examination of cultures for preanalytical considerations. For analytical considerations, it provides quality control and identification. With postanalytical considerations, the chapter provides attention to summary/special considerations.
Procedures for the Storage of Microorganisms
Authors: She Rosemary C. , Butler‐Wu Susan M. 
Published: January 2025

Abstract

This chapter presents methods that can be used for the storage of bacteria, protozoa, fungi, and viruses. The simplest method for maintaining the short‐term viability of microorganisms, usually used for bacteria, is periodic subculture to fresh medium. Stability may be longer for non‐fastidious organisms such as
Pseudomonas aeruginosa
and storage at –80 degree Celsius may allow for long‐term preservation using liquid Amies. Archiving of patient specimens containing analytes of interest helps clinical laboratories prepare for future lot‐to‐lot comparisons, quality control needs, and new assay verification studies. For valuable or large collections of microorganisms in long‐term storage, the International Society for Biological and Environmental Repositories provides guidance on emergency preparedness. It is good practice to maintain backup copies of written or electronic data linked to the stored organisms. The field of microbiology continues to rapidly evolve in this era of genomics, transcriptomics, and proteomics.