Real Betis became the first La Liga club to sack a coach since the season re-started after being halted by the COVID-19 pandemic.
Real Betis lost their first game back 2-0 to Sevilla and drew 2-2 with Granada in mid-week.
Defeat by Athletic Bilbao left Real Betis 14th in the standings on 34 points after 30 games.
They are eight points above Real Mallorca who occupy the final spot outside of the relegation zone.
Rubi was named Real Betis coach last summer after guiding Espanyol to seventh last season, succeeding Quique Setien who went on to become the FC Barcelona manager last January.
He won eight out of 30 league games with the team, including only one away victory.