Digital PCR in the Clinical Microbiology Laboratory: Another Tool on the Molecular Horizon

Digital PCR was first described in 1999. Improvements in instrumentation and reagents have increased its routine use in molecular pathology laboratories. For clinical microbiology laboratories, digital PCR applications could cover a broad range of purposes, from absolute pathogen quantification to the detection of rare mutations that confer resistance to antibiotics. In this review, we describe several potential applications of digital PCR in the clinical microbiology laboratory. As this method continues to evolve, digital PCR may become an important tool on the horizon for clinical and molecular microbiology laboratories.     

Clinical microbiologists facing an anthrax alert

Microbiological war and terrorist attacks are made to weaken populations by transmitting pathogenic and epidemic microorganisms. These bacteria or viruses are often difficult to diagnose. Anthrax alerts following September 2001 showed that most clinical microbiology laboratories were not adequately prepared, using obsolete diagnostic methods or being too slow to use accurate tools when facing a major threat. Following this period, most microbiology laboratories were prepared for bioterrorism alerts, in order to provide accurate and rapid results, although such events are rare and unexpected. In this review, we describe the organization and preparedness of our clinical microbiology laboratory regarding bioterrorism risk, although its main task is to perform routine diagnostic microbiology tests. To illustrate the difficulties, we briefly describe an anthrax alert.     

Biologie moléculaire et microbiologie clinique en 2007: Généralités - Partie 1

Molecular biology is omnipresent in medical biology especially in microbiology. Many papers report studies on its importance in diagnosis, prognosis, therapeutic field, epidemiology, natural (or not) biological risks alike. The important sum of reports on this subject does not always bring an obvious answer to the part that molecular biology could play in a microbiology laboratory either public or private. This article summarizes the contributions of this specialized field in microbiology and brings some questions on its contribution. Is molecular biology a real progress in microbiology? What are the main indications of molecular biology today in this field? According to the infectious particle, the answers are not always simple. They can be obvious in some cases (for instance, hepatitis C) or less obvious in other cases (tuberculosis for instance). In this article divided in two parts, the first one report an introduction to molecular biology in microbiology and the following second part will deal with the main applications in microbiology.     

Biologie moléculaire et microbiologie clinique en 2007: Les applications et leur avenir – Partie 2

Molecular biology is omnipresent in medical biology especially in microbiology. Many papers report studies on its importance in diagnosis, prognosis, therapeutic field, epidemiology, natural (or not) biological risks alike. The important sum of reports on this subject does not always bring an obvious answer to the part that molecular biology could play in a microbiology laboratory either public or private. This article summarizes the contributions of this specialized field in microbiology and brings some questions on its contribution. Is molecular biology a real progress in microbiology? What are the main indications of molecular biology today in this field? According to the infectious particle, the answers are not always simple. They can be obvious in some cases (for instance, hepatitis C) or less obvious in other cases (tuberculosis for instance). In the first part of this article, we reported general remarks in molecular biology related to microbiology. The second part report some important applications underlining the important extent of molecular biology in this field.     

Digital microbiology

BackgroundDigitalization and artificial intelligence have an important impact on the way microbiology laboratories will work in the near future. Opportunities and challenges lie ahead to digitalize the microbiological workflows. Making efficient use of big data, machine learning, and artificial intelligence in clinical microbiology requires a profound understanding of data handling aspects.ObjectiveThis review article summarizes the most important concepts of digital microbiology. The article gives microbiologists, clinicians and data scientists a viewpoint and practical examples along the diagnostic process.SourcesWe used peer-reviewed literature identified by a PubMed search for digitalization, machine learning, artificial intelligence and microbiology.ContentWe describe the opportunities and challenges of digitalization in microbiological diagnostic processes with various examples. We also provide in this context key aspects of data structure and interoperability, as well as legal aspects. Finally, we outline the way for applications in a modern microbiology laboratory.ImplicationsWe predict that digitalization and the usage of machine learning will have a profound impact on the daily routine of laboratory staff. Along the analytical process, the most important steps should be identified, where digital technologies can be applied and provide a benefit. The education of all staff involved should be adapted to prepare for the advances in digital microbiology.     

Onsite microbiology services and outsourcing microbiology and offsite laboratories--advantage and disadvantage, thinking of effective utilization

In recent years, budget restrictions have prompted hospital managers to consider outsourcing microbiology service. But there are many advantages onsite microbiology services. Onsite microbiology services have some advantages. 1) High recovery rate of microorganism. 2) Shorter turn around time. 3) Easy to communicate between physician and laboratory technician. 4) Effective utilization of blood culture. 5) Getting early information about microorganism. 6) Making antibiogram (microbiological local factor). 7) Getting information for infection control. The disadvantages are operating costs and labor cost. The important point of maximal utilization of onsite microbiology service is close communication between physicians to microbiology laboratory. It will be able to provide prompt and efficient report to physicians through discussion about Gram stain findings, agar plate media findings and epidemiological information. The rapid and accurate identification of pathogen affords directed therapy, thereby decreasing the use of broad-spectrum antibiotics and shortening the length of hospital stay and unnecessary ancillary procedures. When the physician use outsourcing microbiology services, should discuss with offsite laboratories about provided services. Infection control person has to arrange data of susceptibility about every isolate and monitoring multi-drug resistant organism. Not only onsite microbiology services but also outsourcing microbiology services, to communicate bedside and laboratory is most important point of effective utilization.     

Applications of Artificial Intelligence in Clinical Microbiology Diagnostic Testing

The use of artificial intelligence (AI) computer software to interpret data has become part of our everyday lives, and these AI algorithms are becoming part of our everyday laboratory practices. Many AI tools are beginning to demonstrate their real or potential utility in clinical microbiology laboratory practice. In this introduction to applications of AI in clinical microbiology diagnostic testing, the authors introduce AI and machine learning to those familiar with routine clinical microbiology practice. The discussion explores the role of AI for image analysis including Gram stains, ova and parasite exam, and digital plate reading of bacterial cultures. AI's role in advanced analysis of matrix-assisted laser desorption-ionization/time of flight mass spectrometry (MALDI-TOF) mass spectral data and whole genome sequence data of microbes is also discussed. In the future, computers and clinical laboratory scientists will work more closely together to provide optimal efficiency and quality in clinical microbiology laboratory practice, and this close collaboration between humans and machines is expected to improve patient care.     

Impact of Microbiology Practice on Cumulative Prevalence of Respiratory Tract Bacteria in Patients with Cystic Fibrosis

Investigators participating in the Epidemiologic Study of Cystic Fibrosis project began to collect microbiological, pulmonary, and nutritional data on cystic fibrosis (CF) patients at 180 North American sites in 1994. Part of this study was a survey undertaken in August 1995 to determine microbiology laboratory practices with regard to pulmonary specimens from CF patients. The survey included a section on test ordering, completed by a site clinician, and a section on test performance and reporting, completed by each site’s clinical microbiology laboratory staff. Seventy-nine percent of the surveys were returned. There was intersite consistency of microbiology laboratory practices in most cases. The majority of sites follow most of the CF Foundation consensus conference recommendations. There were differences in the frequency at which specimens for culture were obtained, in the use of selective media for Staphylococcus aureus and Haemophilus influenzae, and in the use of a prolonged incubation for Burkholderia cepacia. These variations in practice contribute to prevalence differences among sites and may result in differences in clinical care.     

Typing of Staphylococcus epidermidis clinical strains by a selected panel of Brazilian killer yeasts

Discrimination of multi-resistant microorganisms in small clinical microbiology laboratories is frequently based on the biologic profile (biotype) of phenotypic markers, such as antimicrobial susceptibility patterns (antibiograms) and serologic or enzymatic typing, but few use indicative microorganisms. The purpose of this study was to evaluate the power of a panel of selected killer yeasts for differentiating and discriminating clinical isolates of Staphylococcus epidermidis from two care hospitals and clinical microbiology laboratories from the South of Brazil. The short panel of eleven killer yeasts was capable of discriminating 100% of the sensitive strains of S. epidermidis using quantitative data matrix (QDM) and differentiating them from strains of coagulase-positive Staphylococcus. Therefore, this phenotypic methodology proved to be valid as a discriminatory tool when applied to these clinical bacteria strains, besides being simple and feasible for routine use even in microbiological laboratories with minimal resources.     

The impact of a clinic for adults with HIV infection on the microbiology laboratory

The Infectious Diseases Clinic (IDC) discussed serves adults who are seropositive for human immunodeficiency virus. The authors reviewed the outpatient and inpatient microbiology tests of a three-month period during 1989 for a systematic sample of IDC patients. The 249 patients in the sample had 682 microbiology tests performed during the period (mean 2.7 tests per patient). Tests most frequently requested were mycobacterial culture, routine blood culture, and cryptococcal antigen determination. Patients with acquired immunodeficiency syndrome (43% of IDC patients) accounted for 63% of the requested IDC tests. IDC patients comprised about 2.4% of patients served but accounted for 3.9% of the requested microbiology tests and 6.6% of the microbiology work load for reported tests. Using Centers for Disease Control case projections, the authors estimated that services to IDC patients in 1993 would comprise 6.6% of all microbiology tests and 10.6% of the microbiology work load. The implications of these data for microbiology probably also apply to other laboratory testing and emphasize the need for more efficient ways to use and perform diagnostic studies required by patients with HIV infection.     

How to: identify non-tuberculous Mycobacterium species using MALDI-TOF mass spectrometry

BackgroundThe implementation of MALDI-TOF MS for microorganism identification has changed the routine of the microbiology laboratories as we knew it. Most microorganisms can now be reliably identified within minutes using this inexpensive, user-friendly methodology. However, its application in the identification of mycobacteria isolates has been hampered by the structure of their cell wall. Improvements in the sample processing method and in the available database have proved key factors for the rapid and reliable identification of non-tuberculous mycobacteria isolates using MALDI-TOF MS.AimsThe main objective is to provide information about the proceedings for the identification of non-tuberculous isolates using MALDI-TOF MS and to review different sample processing methods, available databases, and the interpretation of the results.SourcesResults from relevant studies on the use of the available MALDI-TOF MS instruments, the implementation of innovative sample processing methods, or the implementation of improved databases are discussed.ContentInsight about the methodology required for reliable identification of non-tuberculous mycobacteria and its implementation in the microbiology laboratory routine is provided.ImplicationsMicrobiology laboratories where MALDI-TOF MS is available can benefit from its capacity to identify most clinically interesting non-tuberculous mycobacteria in a rapid, reliable, and inexpensive manner.     

Editorial – The Microbiology Laboratorian in a Post-technology World: Indispensable or Obsolete?

Technology continues to have a major impact on the way that laboratory medicine is practiced. Automation, culture-independent pathogen detection methods, and rapid antimicrobial susceptibility testing are all influencing practice in the clinical microbiology laboratory and treatment of patients in the clinical practice arena. The mining of big data and its ability to guide clinical decisions through artificial intelligence and machine learning will continue to alter the value assigned to laboratory medicine. This editorial explores these changes and the challenges they pose to investigate the skill set that will be required from this and the next generation of laboratory leaders to successfully leverage the role of the clinical microbiology laboratory in a post-technology world.     

Basic Concepts of Microarrays and Potential Applications in Clinical Microbiology

Summary: The introduction of in vitro nucleic acid amplification techniques, led by real-time PCR, into the clinical microbiology laboratory has transformed the laboratory detection of viruses and select bacterial pathogens. However, the progression of the molecular diagnostic revolution currently relies on the ability to efficiently and accurately offer multiplex detection and characterization for a variety of infectious disease pathogens. Microarray analysis has the capability to offer robust multiplex detection but has just started to enter the diagnostic microbiology laboratory. Multiple microarray platforms exist, including printed double-stranded DNA and oligonucleotide arrays, in situ-synthesized arrays, high-density bead arrays, electronic microarrays, and suspension bead arrays. One aim of this paper is to review microarray technology, highlighting technical differences between them and each platform''s advantages and disadvantages. Although the use of microarrays to generate gene expression data has become routine, applications pertinent to clinical microbiology continue to rapidly expand. This review highlights uses of microarray technology that impact diagnostic microbiology, including the detection and identification of pathogens, determination of antimicrobial resistance, epidemiological strain typing, and analysis of microbial infections using host genomic expression and polymorphism profiles.     

Biomedical mass spectrometry in today's and tomorrow's clinical microbiology laboratories

Clinical microbiology is a conservative laboratory exercise where base technologies introduced in the 19th century remained essentially unaltered. High-tech mass spectrometry (MS) has changed that. Within a few years following its adaptation to microbiological diagnostics, MS has been introduced, embraced, and broadly accepted by clinical microbiology laboratories throughout the world as an innovative tool for definitive bacterial species identification. Herein, we review the current state of the art with respect to this exciting new technology and discuss potential future applications.     

Photograms for microbiological assays.

The use of photograms provides a permanent record of microbiological assays in which diffusion of a substance in agar is measured. The accuracy of this procedure is comparable to direct measurement. This technique is inexpensive, does not require special photographic equipment, and is applicable to many tests commonly employed in clinical microbiology and immunology laboratories.     

Laboratory evaluation of leukocyte esterase and nitrite tests for the detection of bacteriuria.

We compared the sensitivity, specificity, and predictive values of the 1-min leukocyte esterase test and the test for urinary nitrite alone and in combination as screening tests for bacteriuria in over 5,000 clinical urine specimens. The leukocyte esterase-nitrite combination had a sensitivity of 79.2%, a specificity of 81%, and a negative predictive value of a negative test of 94.5% for specimens with greater than or equal to 10(5) CFU/ml. Although the sensitivity of this test was too low to allow its use as the only screening test for bacteriuria, it may serve as a useful adjunct to culturing and other urine-processing systems in the microbiology laboratory.     

Comparative performance of Fungichrom I, Candifast and API 20C Aux systems in the identification of clinically significant yeasts.

To compare the performance of current chromogenic yeast identification methods, three commercial systems (API 20C Aux, Fungichrom I and Candifast) were evaluated in parallel, along with conventional tests to identify yeasts commonly isolated in this clinical microbiology laboratory. In all, 116 clinical isolates, (68 Candida albicans, 12 C. parapsilosis, 12 C. glabrata and 24 other yeasts) were tested. Germ-tube production, microscopical morphology and other conventional methods were used as standards to definitively identify yeast isolates. The percentage of isolates identified correctly varied between 82.7% and 95.6%. Overall, the performance obtained with Fungichrom I was highest with 95.6% identification (111 of 116 isolates). The performance of API 20C Aux was higher with 87% (101 of 116 isolates) than that of Candifast with 82.7% (96 of 116). The Fungichrom I method was found to be rapid, as 90% of strains were identified after incubation for 24 h at 30 degrees C. Both of the chromogenic yeast identification systems provided a simple, accurate alternative to API 20C Aux and conventional assimilation methods for the rapid identification of most commonly encountered isolates of Candida spp. Fungichrom seemed to be the most appropriate system for use in a clinical microbiology laboratory, due to its good performance with regard to sensitivity, ease of use and reading, rapidity and the cost per test.     

Evaluation of an Online Program To Teach Microbiology to Internal Medicine Residents

Microbiology rounds are an integral part of infectious disease consultation service. During microbiology rounds, we highlight microbiology principles using vignettes. We created case-based, interactive, microbiology online modules similar to the vignettes presented during microbiology rounds. Since internal medicine residents rotating on our infectious disease elective have limited time to participate in rounds and learn microbiology, our objective was to evaluate the use of the microbiology online modules by internal medicine residents. We asked residents to complete 10 of 25 online modules during their infectious disease elective. We evaluated which modules they chose and the change in their knowledge level. Forty-six internal medicine residents completed assessments given before and after accessing the modules with an average of 11/20 (range, 6 to 19) and 16/20 (range, 9 to 20) correct questions, respectively (average improvement, 5 questions; P = 0.0001). The modules accessed by more than 30 residents included those related to Clostridium difficile, anaerobes, Candida spp., Streptococcus pneumoniae, influenza, Mycobacterium tuberculosis, and Neisseria meningitidis. We demonstrated improved microbiology knowledge after completion of the online modules. This improvement may not be solely attributed to completing the online modules, as fellows and faculty may have provided additional microbiology education during the rotation.     

Serum bactericidal test.

The serum bactericidal test represents one of the few in vitro tests performed in the clinical microbiology laboratory that combines the interaction of the pathogen, the antimicrobial agent, and the patient. Although the use of such a test antedates the antimicrobial era, its performance, results, and interpretation have been subject to question and controversy. Much of the confusion concerning the serum bactericidal test can be avoided by an understanding of the various factors which influence bactericidal testing. In addition, the methodologic aspects of the serum bactericidal test have recently been addressed and should place this test on firmer ground. New information on the clinical utility of this test is becoming available; additional data are needed to establish more clearly the usefulness of the serum bactericidal test in specific infections. Such clinical trials from multiple centers will enable firmer recommendations for the future use of the serum bactericidal test.