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One of the oldest clubs in Greek football, Anagennisi was formed in 1904 in the southwestern Thessalian city of Karditsa as the football team of the multisport club ASA (Greek: Αθλητικός Σύλλογος Αναγέννησης - Athlitikos Syllogos Anagennisis), the Athletic Association Anagennisi. The name of the club comes from the Greek word for Renaissance. The statute of the club was signed personally by Crown Prince Constantine I.

From its inception in 1904, the football team of ASA was not officially recognized as a distinct entity within the organization. As more of the club's athletes began to show interest in the sport, the football team began to train, and play locally organized games, in the area of central Karditsa (which later became the Pavsilipo Park). With the sport's expanding popularity in Greece in the following decades, in 1924 the football team was finally given its own department within ASA. Playing in the independent, regional Thessalian League from 1929 to 1962, it joined the newly established Greek second division in time for Season 1962-63. When the Greek football league became professional in 1979, the football department of ASA was reformed as a Football Public Limited Company, or PAE (Greek: ΠΑΕ - Ποδοσφαιρική Ανώνυμη Εταιρία / Podosferiki Anonymi Eteria) and continued to compete as PAE Anagennisi Karditsas (FC Anagennisi Karditsa).

One of the highlights in Anagennisi Karditsa's history is the victory over Olympiacos for the Greek Cup in 1993-94 as well as the win against Paniliakos for the Greek semi-professional Cup final in 1981. In 2008 Anagennisi Karditsa reach the Gamma Ethniki play-off and beat Ilioupoli 2-0 in Athens to win promotion to Beta Ethniki.

Anagennisi Karditsa is a well-supported club by regional football standards, and their passion is very well known. They hold the record for the most fans to ever attend an away game for Gamma Ethniki in 2008, with 2.000 supporters in the play-off match against Ilioupoli. In 2008-09 season Anagennisi Karditsa had the fourth best average in attendance despite having only avoided relegation in the last fixture against Veria.