The Newest Vital Sign (NVS) is a simple, quick and accurate screening test for health literacy (HL). It has been validated for different languages but, to date, not for the Croatian language. The aim of this study was to develop a linguistically validated Croatian version of the NVS and to use it at a later stage in a pilot study of health literacy assessment of hospital patients in Croatia. A full linguistic validation procedure was applied, including forward and backward translation, expert panel review, cognitive interview with 10 respondents from general population, and full involvement in the procedure of one of the screening test developers, the lead author of the NVS-UK version. HL testing on 100 hospital patients (55% women, median age 63.5 years) revealed 58% of patients had less than adequate HL level (scores less than 4), and mean NVS total score was 3.34. A positive significant association was observed between HL and educational level (p = 0.002). A high percentage of pati)