From correlation to causation: dissecting the detection methodologies of oral bacterial-fungal crosstalk in health and disease

In the oral microecosystem, cross-kingdom interactions between bacteria and fungi are pivotal ecological events that determine the equilibrium between oral health and disease. Historically, technical limitations have confined research to describing correlations in microbial community composition, hindering the elucidation of underlying pathogenic mechanisms. This review aims to systematically outline the evolution of detection technologies for oral bacterial-fungal interactions and critically discuss the paradigm shift from identifying “associative correlations” to deciphering “causal mechanisms.” The literature search encompassed studies ranging from traditional culture and molecular identification to high-throughput sequencing for ecological network construction, as well as cutting-edge multi-omics integration and high-resolution imaging technologies. Beyond cataloguing technical advances, we critically evaluate the level of causal inference supported by each methodological class, with particular attention to temporality, confounding, ecological simplification, and the need for perturbational validation in disease-relevant models. Representative studies are used to illustrate how evidence can progress from co-occurrence and spatial proximity to functional dependency and mechanism.