The human mycobiome refers to the fungal communities residing across body sites and plays pivotal roles in human health and disease. This chapter summarizes technical advances and current knowledge on the human mycobiome and discusses its clinical implications. Although high-throughput sequencing-based approaches have greatly improved the resolution of profiling fungal populations compared to the traditional culture-based methods, researchers should be aware of the inherent limitations of each approach and choose the most appropriate one or combination based on specific context of their study. We highlight the research progress on the composition of mycobiome and its cross talk with the host in the gastrointestinal tract, respiratory tract, oral cavity, genital tract, tumor tissues, and skin surface. The complex cross-kingdom interactions with bacteria and the emergence of new fungal pathogens-potentially driven by environment factors, emphasize the need to integrate mycobiome studies into broader microbial networks and the One Health frameworks. Together, this chapter underscores the potential of the human mycobiome as a diagnostic and therapeutic target in various diseases and advocates for interdisciplinary efforts to address the impact of fungi on human health.