Although he is not seen, the only reference to Mycroft Holmes in the Basil Rathbone / Nigel Bruce series is made in this film.
This marks the third Sherlock Holmes' movie starring Basil Rathbone where Holmes faces Prof. James Moriarty, after The Adventures of Sherlock Holmes (1939) and Sherlock Holmes and the Secret Weapon (1942). Curiously, Moriarty is portrayed by a different actor in each movie: George Zucco, Lionel Atwill, and Henry Daniell, respectively.
This was Henry Daniell's third of three different roles in the Rathbone Sherlock Holmes works, following Sherlock Holmes and the Voice of Terror (1942) as as an ally of Holmes, Anthony Lloyd, and Sherlock Holmes in Washington (1943) as William Easter.
Inspector Gregson (Matthew Boulton) assumes the role of the Scotland Yard contact normally filled by Inspector Lestrade. Most likely Dennis Hoey was working on another picture (probably A Thousand and One Nights (1945), to judge from production schedules) and was unavailable. Holmes does mention his absence at one point.
Only with the color version would the viewer know Lydia's final outfit was green; there is no mention of the wardrobe color in the movie. It was assumed in the original B&W version Lydia was wearing green sometime during the movie.