Replacement of Culture with Molecular Testing for Diagnosis Infectious Diseases

Key points

Introduction

Despite significant advances in molecular testing platforms, in vitro cultivation of microbial pathogens on artificial media remains a hallmark of infectious disease diagnostics. Culture-based approaches allow for the recovery, identification, and antimicrobial susceptibility testing of a specific pathogen. However, there are many limitations to the use of culture. The growth of an organism can take days to weeks; definitive testing for identification can be performed only on pure culture or isolated colony, which may introduce additional delays; some organisms cannot be cultivated in vitro (eg, some viruses, parasites) or have specialized growth requirements that cannot be met by most routine clinical microbiology laboratories (eg, Mycoplasma species). Ultimately, culture is labor intensive and is associated with long turnaround times. Molecular testing has exquisite sensitivity and specificity and is associated with faster turnaround times. Limitations of molecular assays include high cost, the need for specific primer sequences based on an a priori knowledge of the pathogen target sequence, and laboratory personnel with specific expertise in molecular techniques. Nonetheless, there are many areas where laboratories have transitioned away from culture-based pathogen detection to molecular methods. This article discusses the considerations for adoption of molecular assays as a replacement (in whole or in part) for culture, and to highlight examples where this approach has been successfully implemented.

Discussion

The movement away from culture-based testing to molecular testing for infectious diseases largely began in academic medical centers and specialized reference laboratories, which implemented laboratory-developed molecular testing. Recently, there has been a shift in adoption of commercially available, sample-to-answer testing platforms for more routine pathogens. The nature of these tests allows use by smaller community hospitals and other laboratories located outside of large systems and academic centers. The decision to migrate culture-based diagnostic testing to molecular testing must include an assessment of several factors, including test volume, analytical performance, workflow and throughput, availability of trained personnel, and cost. Regulatory and accreditation factors must also be taken into consideration.

Early adoption of molecular testing as a replacement for culture occurred in the diagnostic virology laboratory. The ability to read cytopathic effect in cell culture and identify specific viruses based on the cytopathic effect observed in various cell lines is an expert skill that has waned in laboratories over time owing to staff attrition and a shortage of new medical laboratory scientists who are interested in microbiology. This situation has caused a shift in virology testing to shorter turnaround time and shell vial cultures, which require their own level of expertise to read immunofluorescent stains. The loss of skilled personnel and the inability to propagate some viruses in culture in vitro led to the replacement of culture-based virology testing with molecular testing. Viruses such as human immunodeficiency virus, hepatitis A, hepatitis B, and hepatitis C were among the first molecular tests used both for the diagnosis of infection and monitoring of viral load over time to determine the effectiveness of antiviral therapy. Testing for other clinically important viruses soon shifted to molecular testing. Respiratory virus testing, for example, has largely transitioned to multiplex molecular panels that allow for a diagnosis of influenza virus A, influenza virus B, and respiratory syncytial virus (as well as other respiratory viruses for which there are no specific antiviral therapies, such as human metapneumovirus) from a single specimen in a single test. The diagnosis of infectious diarrhea has also shifted from culture, specialized microscopy, and rapid antigen testing methods to multiplex molecular gastrointestinal panels. Etiologic causes of infectious gastroenteritis include bacteria, viruses, and parasites, and clinical presentation typically does not allow for a specific diagnosis because diarrhea is the predominant symptom. Therefore, a battery of tests must be ordered, yet often the causative agent remains unidentified. Gastrointestinal panels provide the laboratory with a single test for a single specimen and yield results for a breadth of infectious agents. Performance of these culture-independent diagnostic tests is reported to be excellent