Objectives Regular HIV testing in men who have sex with men (MSM) enables timely entry into care and reduces the likelihood of HIV transmission. We aimed to assess HIV-testing behaviour and associated factors in MSM by urbanisation of place of residence. Design Data were derived from online survey ('Men & Sexuality') in the Netherlands, which was mainly advertised on social media (Facebook and Instagram), dating websites, apps for MSM (Grindr and PlanetRomeo) and gay media. Primary and secondary outcome measures HIV testing was defined as recent (<1 year), not recent (>= 1 year) or never. Using multinominal regression analyses, factors associated with not recent testing and never testing, compared with recent testing, were assessed among MSM living in highly (>2500 residences/km(2)) or non-highly (<= 2500 residences/km(2)) urbanised areas. Participants The study sample included 3815 MSM, currently living in the Netherlands. The mean age was 36 years (SD 14.7), and 67.6% were highly educated. Results In highly urbanised areas, 11.8% was never and 19.8% was not recently HIV-tested. In non-highly urbanised areas, this was 25.2% and 19.6%. Among MSM living in highly urbanised areas, independently associated with never and not recent testing were younger age, self-identification as bisexual, fewer sex partners, never notified of HIV and no recent condomless anal intercourse. Among MSM living in non-highly urbanised areas, lower perceived HIV severity, higher perceived HIV risk and a lower proportion gay friends were associated with never and not recent testing. Among never tested MSM, those in non-highly urbanised areas preferred self-sampling/self-testing over facility-based testing; those in highly urbanised areas preferred testing at healthcare facilities. Conclusions The proportion of never tested MSM was high (25%) in non-highly urbanised areas in the Netherlands. MSM living in non-highly urbanised areas may possibly be reached with targeted approaches to increase HIV testing uptake such as ...