Historically, anti-vaccination sentiment has existed in many populations. Mass media plays a large role in disseminating and sensationalizing vaccine objections, especially via the medium of the Internet. Based on studies of processing fluency, we assumed that anti-influenza vaccination online messages to be more readable and more fluently processed than pro-influenza vaccination online messages, which may consequently sway the opinions of some audiences. The aim of this study was to compare readability of anti- and pro-influenza vaccination online messages in Japan using a measure of readability.Web searches were conducted at the end of August 2016 using two major Japanese search engines (Google.jp and Yahoo!.jp). The included websites were classified as “anti”, “pro”, or “both” depending on the claims, and “health professional” or “non-health professional” depending on the writers' expertise. Readability was determined using a validated measure of Japanese readability (the Japanese sentence difficulty discrimination system). Readability of “health professional” websites was compared with that of “non-health professional” websites, and readability of “anti” websites was compared with that of “pro” websites, using the t-test.From a total of 145 websites, the online messages written by non-health professionals were significantly easier to read than those written by health professionals (p = 0.002, Cohen's d = 0.54). Anti-influenza vaccination messages were significantly easier to read than pro-influenza vaccination messages (p < 0.001, Cohen's d = 0.74).When health professionals prepare pro-influenza vaccination materials for publication online, we recommend they check for readability using readability assessment tools and improve the text for easy reading if necessary.