The gut metabolite trimethylamine-N-oxide (TMAO) has gained recognition for potential involvement in cardiovascular disease (CVD), though whether this reflects a biomarker function or a causal role in disease pathogenesis awaits elucidation. Elevated TMAO is reported in chronic disorders including obesity, type 2 diabetes (T2D) and metabolic syndrome (MetS). Shifts in gut biome compositions may contribute to changes in formation across diseases. We explored relationships between dietary intake, gut biome profiles, circulating TMAO and cardiometabolic disease status. Biobanked plasma samples from well-defined (age and gender matched) healthy, overweight, obese, and MetS subjects (n=35-40 per group) were analysed. Patient health/medical and lifestyle history (including T2D risk) were assessed along with dietary composition (3-day food diary), blood biochemistry and gut microbial composition (16s rRNA sequencing). Circulating TMAO and its precursor substrates were assayed in stored plasma via LC-MS/MS. Across the data set, TMAO correlated significantly with insulin (p<0.0001), cholesterol (p<0.05) and seafood intake (p<0.0001). Conversely, markers of gut biome diversity were negatively correlated with MetS criteria (p<0.001), BMI (p<0.001), glucose (p<0.01) and diabetes risk (p<0.001), while correlating positively with HDL (p=0.0001) and select dietary macro- and micronutrients. These correlations grew stronger when data were limited to individuals from the obese or MetS groups. Microbiome profiles from the MetS group were statistically less diverse based on operational taxonomic unit count (p<0.005) and Shannon (p<0.05) indexes. Circulating carnitine was significantly higher in patients with MetS (p<0.05), though no other significant differences were present in precursors or TMAO itself. These results suggest that while biome shifts arise with disease status, this does not translate into changes in TMAO. Biome changes may be explained by diet, though additional investigation is needed to elucidate these relationships and influences of additional variables, including gut permeability.