The 52-year-old leaves Bootham Crescent with the Minstermen sitting 22nd in League Two.
York, who lost 3-1 at Newport County on Saturday in Worthington's final match in charge, have won just one of their 12 matches in the league this season.
"After Saturday's result and in the best interests of York City, I have made the difficult decision to leave," Worthington told the club's official website.
"In recent weeks, performances have not been up to my high standards and I take full responsibility for this."
The former Northern Ireland boss succeeded Gary Mills as York manager in March 2013.