Background: This longitudinal study aimed to investigate (risk factors for) persistence of carriage and molecular characteristics of extended-spectrum and pAmpC β-lactamase-producing (ESBL/pAmpC) Escherichia coli and Klebsiella pneumoniae (ESBL-E/K) in adults in the Dutch community. Methods: Following a cross-sectional study (ESBL-E/K prevalence 4.5%), a subset of ESBL-E/K positive (n=76) and negative (n=249) individuals volunteered to provide five monthly faecal samples and questionnaires. ESBL-E/K was cultured using selective enrichment/culture and MLSTs were determined. ESBL/pAmpC-genes were analysed using PCR and sequencing. Plasmids were characterized and subtyped by plasmid MLST. Risk factors for persistent carriage were analysed using logistic regression. Results: Of the initially ESBL-E/K positive participants, 25/76 (32.9%) remained positive in all subsequent samples; 51/76 persons (67.1%) tested ESBL-E/K negative at some time point during follow-up of which 31 (40.8%) stayed negative throughout the longitudinal study. Carriers often carried the same ESBL-gene and plasmid, but sometimes in different ESBL-E/K strains, indicative for horizontal transfer of plasmids. Of the 249 initially ESBL-E/K negative participants, the majority (n=218, 87.6%) tested negative during eight months follow-up, whereas 31/249 (12.4%) participants acquired an ESBL-E/K. E. coli phylogenetic group B2 and D and travel to ESBL high prevalence countries were associated with prolonged carriage. Conclusion: ESBL-E/K carriage persisted for more than eight months in 32.9% of the initially ESBL-positive individuals, while 12.4% of initially negatives acquired ESBL-E/K during the study. A single positive test result provides no accurate prediction for prolonged carriage. Acquisition/loss of ESBL-E/K does not seem to be a random process, but differs between bacterial genotypes.