With heightened global concern of microbial drug resistance, advanced methods for early and accurate diagnosis of infection are urgently needed. Analysis of exhaled breath volatile organic compounds (VOCs) toward detecting microbial infection potentially allows a highly informative and noninvasive alternative to current genomics and culture-based methods. We performed a systematic review of research literature reporting human and animal exhaled breath VOCs related to microbial infections. In this Review, we find that a wide range of breath sampling and analysis methods are used by researchers, which significantly affects interstudy method comparability. Studies either perform targeted analysis of known VOCs relating to an infection, or non-targeted analysis to obtain a global profile of volatile metabolites. In general, the field of breath analysis is still relatively immature, and there is much to be understood about the metabolic production of breath VOCs, particularly in a host where both commensal microflora as well as pathogenic microorganisms may be manifested in the airways. We anticipate that measures to standardize high throughput sampling and analysis, together with an increase in large scale collaborative international trials, will bring routine breath VOC analysis to improve diagnosis of infection closer to reality.