Detection and identification of microorganisms by gene amplification and sequencing.

Gene amplification and sequencing have led to the discovery of new pathogens as agents of disease and have enabled us to better classify microorganisms from culture. Sequence-based identification of bacteria and fungi using culture is more objective and accurate than conventional methods, especially for classifying unusual microorganisms that are emerging pathogens in immunocompromised hosts. Although a powerful tool, the interpretation of sequence-based classification can be challenging as microbial taxonomy grows more complex, without known clinical correlatives. Additionally, broad-range gene polymerase chain reaction and sequencing have emerged as alternative, culture-independent methods for detecting pathogens from clinical material. The promise of this technique has remained strong, limited mainly by contamination and inadequate sensitivity issues. This review explains sequence-based microbial classification, with emphasis on relating the complex world of microbial taxonomy to a clinical context. Additionally, this review discusses a rational approach to broad-range bacterial polymerase chain reaction and gene sequencing when applied directly to clinical samples.