Central Wisconsin Christian joined the conference after leaving the East Central Flyway Conference in 2004, and two years later the Trailways Conference took three more former East Central Flyway schools into the fold: Lourdes Academy, Markesan and Oakfield. All three schools joined the North Division with Pardeeville moving over to the South Division to accommodate the expansion:[2]
North Division
South Division
Cambria-Friesland
Abundant Life Christian
Central Wisconsin Christian
Deerfield
Green Lake
Dodgeland
Lourdes Academy
Fall River
Markesan
Hustisford
Montello
Johnson Creek
Oakfield
Pardeeville
Princeton
Rio
Randolph
Williams Bay
Valley Christian
This alignment would only last for two years, as the Trailways split into Central, North and South Divisions in 2008:[3]
Central Division
North Division
South Division
Cambria-Friesland
Green Lake
Abundant Life Christian
Central Wisconsin Christian
Lourdes Academy
Deerfield
Markesan
Montello
Dodgeland
Pardeeville
Oakfield
Fall River
Randolph
Princeton
Hustisford
Valley Christian
Johnson Creek
Rio
Williams Bay
In 2012, Green Lake and Princeton High Schools entered into a cooperative agreement for their schools' athletics, and the newly created program remained in the North Division.[4] That same year, Horicon[5] and Madison Country Day School[6] in Waunakee joined the Trailways, and Rio moved to the North Division to keep membership at six schools. The Trailways Conference was realigned to a two-division format in 2013:
North Division
South Division
Cambria-Friesland
Abundant Life Christian
Central Wisconsin Christian
Deerfield
Green Lake/Princeton
Dodgeland
Lourdes Academy
Fall River
Markesan
Horicon
Montello
Hustisford
Oakfield
Johnson Creek
Pardeeville
Madison Country Day
Randolph
Rio
Valley Christian
Williams Bay
In 2014, Wayland Academy in Beaver Dam joined the Trailways South Division,[7] and St. Ambrose Academy in Madison joined with Abundant Life Christian School for most sponsored sports (they would move to their own stand-alone programs in 2023).[8] The North/South divisional alignment would last for two more years before the Trailways realigned to three divisions in 2016:
Central Division
North Division
South Division
Cambria-Friesland
Green Lake/Princeton
Abundant Life Christian
Central Wisconsin Christian
Lourdes Academy
Deerfield
Horicon
Markesan
Dodgeland
Pardeeville
Montello
Fall River
Randolph
Oakfield
Hustisford
Wayland Academy
Valley Christian
Johnson Creek
Madison Country Day
Rio
St. Ambrose Academy
Williams Bay
This alignment would only last a single season, as Palmyra-Eagle and Parkview joined the Trailways as all-sport members in 2017 after leaving the Rock Valley Conference,[9] bringing conference membership to the current twenty-four schools and the current East/South/West divisional alignment.